HLA-G、HLA-E在卵巢恶性肿瘤免疫逃逸中的作用及药物干预的实验研究
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摘要
卵巢癌是妇科恶性肿瘤中死亡率居第一的恶性疾患,就诊时绝大多数患者已处于晚期。近30年来,虽经世界各国妇科肿瘤学家的不懈努力,成功探索出在肿瘤细胞减灭术的基础上辅助化疗的治疗策略,但晚期卵巢癌的5年生存率仍徘徊在20%左右。因此,研究卵巢恶性肿瘤的发生发展机制,寻求最根本有效的治疗方案,提高卵巢癌症患者生存质量一直是妇科肿瘤学者面临的重大挑战。人类白细胞抗原- G(human leukocyte antigen G,HLA-G)和人类白细胞抗原-E(human leukocyte antigen E,HLA-E)是近年来发现的非经典HLA-I类分子,HLA-G能与NK细胞、T细胞亚类、抗原递呈细胞、B细胞和单核细胞系的抑制性受体结合,从而抑制机体的免疫功能;HLA-G还可促进肿瘤细胞表达HLA-E,而后者也可通过与NK细胞和T细胞表面的抑制性受体相互作用而抑制两者的细胞溶解作用。HLA-G和HLA-E最早在胎盘绒毛膜外滋养层细胞表面发现有大量表达,二者在胎儿免疫耐受方面的作用机制已得到国内外学者的公认,并且对于两种抗原与肿瘤的关系也进行了初探。1998年,HLA-G抗原mRNA表达首先在皮肤黑色素瘤细胞系被报道,随后其在肾肿瘤、乳腺肿瘤、结直肠癌、肺癌、淋巴瘤、膀胱癌、胶质母细胞瘤中相继报道,大多数研究认为恶性肿瘤表达HLA-G抗原,且其抗原表达与肿瘤的免疫逃逸有关。但迄今为止二者在卵巢恶性肿瘤中的表达情况报道甚少且尚无定论。2006年国外研究报道了孕激素、绒毛膜促性腺激素可通过上调HLA-G的水平以达到保胎的目的,由此推测临床的引产药物如天花粉蛋白、米非司酮等引起流产的机理可能与其能下调HLA-G、HLA-E的水平有关,进而推测这些药物在免疫逃逸方面的抗肿瘤机制,为恶性肿瘤临床免疫治疗、生物治疗开辟出新的途径,该方面的研究目前国内外未见报道。本研究试图通过检测卵巢恶性肿瘤实体组织和体外培养细胞中HLA-G、HLA-E的mRNA和蛋白水平,分析其在卵巢肿瘤免疫逃逸中的作用。观察顺铂、天花粉蛋白和米非司酮对卵巢癌细胞生长的抑制作用及药物作用前后细胞中HLA-G、HLA-E mRNA和蛋白水平的变化,明确药物的抗肿瘤作用及其机制。并用卵巢癌细胞株进行裸鼠动物模型试验,观察药物对肿瘤生长及对肿瘤组织中HLA-G、HLA-E表达情况的影响,在此基础上寻找出有效的抗癌药物,为卵巢恶性肿瘤发生机制及生物治疗研究提供新的思路
和理论依据。第一部分卵巢恶性肿瘤HLA-G、HLA-E mRNA及蛋白表达水平与其临床生物学行为的关系
目的:检测HLA-G、HLA-E在良、恶性卵巢肿瘤和正常卵巢组织中的表达及与卵巢恶性肿瘤生物学行为的关系。
方法:采用半定量逆转录-聚合酶链反应(Reversed Transcript-Polymerase Chain Reaction, RT-PCR)和流式细胞分析技术检测60例卵巢恶性肿瘤、30例良性肿瘤和10例正常卵巢组织中HLA-G、HLA-E的mRNA和蛋白水平,并结合肿瘤组织类型、临床分期、病理分级等临床病理特征进行综合分析。
结果:①恶性组、良性组及正常组卵巢组织HLA-G mRNA阳性表达率分别为93.33%、73.33%、60%,蛋白阳性表达率分别为61.67%、16.67%、10%。经χ2检验,恶性组HLA-G mRNA和蛋白阳性表达率均显著高于良性组与正常组;而良性组同正常组相比,统计学差异不显著。②恶性组、良性组及正常组卵巢组织HLA-E mRNA阳性率分别为96.67%、83.33%、70%,蛋白阳性表达率分别为75%、36.67%、30%。经χ2检验,恶性组HLA- E mRNA阳性表达率显著高于正常组,也有高于良性组的趋势(χ2=3.272,P=0.07);恶性组HLA-E蛋白阳性表达率显著高于良性组及正常组。而良性组同正常组相比,HLA-E表达无统计学差异。③不同组织类型之间HLA-G mRNA和蛋白表达均无统计学差异。不同临床分期之间HLA-G mRNA表达无统计学差异,但Ⅰ/Ⅱ期有低于Ⅲ/Ⅳ期的趋势(χ2=4.133,P=0.077);HLA-G蛋白阳性表达率随临床分期进展而增高。不同分化程度之间HLA-G mRNA表达有统计学差异,高分化组HLA-G mRNA和蛋白表达显著低于低分化组。④不同组织类型之间HLA-E mRNA和蛋白表达均无统计学差异。不同临床分期之间HLA- E mRNA表达没有统计学差异,但Ⅰ/Ⅱ期有低于Ⅲ/Ⅳ期的趋势(χ2=4.828,P=0.086);HLA- E蛋白表达率随临床分期进展而增高。不同分化程度之间HLA-E mRNA和蛋白阳性表达率有统计学差异,高分化组显著低于低分化组。
结论:①卵巢恶性肿瘤组HLA-G、HLA-E表达均显著高于良性肿瘤组和正常组;②HLA-G、HLA-E表达与卵巢恶性肿瘤组织类型无关;与临床分期和病理分级显著相关,临床分期越晚、组织分化程度越差,抗原表达水平越高。③HLA-G、HLA-E可能参与了卵巢恶性肿瘤的发生、发展过程,从而使癌细胞产生免疫耐受,逃逸宿主的免疫监视。
第二部分卵巢癌细胞中HLA-G、HLA-E mRNA及蛋白表达水平检测
目的:检测HLA-G、HLA-E在卵巢癌细胞中mRNA和蛋白水平的表达情况,为进一步开展卵巢癌的临床免疫治疗提供细胞基础实验依据。
方法:培养卵巢癌细胞株SKOV3、3AO、OVCAR3,采用RT-PCR和流式细胞分析技术检测细胞中mRNA和蛋白水平的表达情况,并以高表达HLA-G的绒癌JEG-3细胞做为阳性对照。
结果:卵巢癌SKOV3细胞HLA-G、HLA-E mRNA和蛋白表达均为阴性;卵巢癌3AO细胞HLA-G mRNA和蛋白表达为阴性,HLA-E mRNA和蛋白表达为阳性;卵巢癌OVCAR3细胞HLA-G mRNA表达为阳性、蛋白表达为阴性,HLA-E mRNA和蛋白表达均为阳性;绒癌JEG-3细胞HLA-G、HLA-E mRNA和蛋白表达均为强阳性。
结论:虽然不同卵巢癌细胞株表达HLA-G、HLA-E水平有所不同,但还是从细胞水平验证了HLA-G、HLA-E在卵巢癌免疫逃逸方面的作用。并为下部分药物干预研究提供了同时表达HLA-G和HLA-E的卵巢癌OVCAR3细胞系。
第三部分天花粉蛋白、米非司酮、顺铂对卵巢癌细胞生长的影响
目的:检测天花粉蛋白、米非司酮及顺铂对卵巢癌细胞系生长的影响,明确药物的抗肿瘤作用。
方法:培养卵巢癌细胞株SKOV3、3AO、OVCAR3,MTT技术观察不同浓度天花粉蛋白、米非司酮、顺铂对三种细胞株生长增殖能力的影响。
结果:①10μg/ml、50μg/ml、100μg/ml、500μg/ml、1000μg/ml天花粉蛋白对SKOV3细胞的生长抑制率分别为0、3.67%、18.60%、34.46%、46.96%;对3AO细胞分别为0、2.97%、9.17%、35.01%、48.98%;对OVCAR3细胞分别为1.06%、9.03%、14.95%、39.66%、58.65%。药物浓度越高,对细胞生长抑制率越强。②2.5μg/ml、5μg/ml、10μg/ml、20μg/ml、40μg/ml米非司酮对SKOV3细胞的生长抑制率分别为2.45%、4.52%、5.98%、21.67%、30.03%;对3AO细胞分别为11.38%、17.93%、20.82%、45.75%、58.63%;对OVCAR3细胞分别为8%、24.88%、47.6%、61.58%、76.94%。③1.5625μg/ ml、3.125μg/ ml、6.25μg/ ml、12.5μg/ ml、25μg/ ml顺铂对SKOV3细胞的生长抑制率分别为5.68%、17.27%、21.75%、45.32%、54.98%;对3AO细胞分别为25.55%、39.81%、63.07%、86.93%、95.28%;对OVCAR3细胞分别为17.82%、29.25%、60.15%、70.57%、85.67%。顺铂对细胞生长抑制作用呈浓度依赖性。
结论:天花粉蛋白、米非司酮和顺铂对卵巢癌细胞SKOV3、3AO、OVCAR3生长具有明显抑制作用,并呈浓度依赖性。从体外实验明确了天花粉蛋白、米非司酮、顺铂具有抗卵巢癌作用,为进一步药物机制的研究及动物体内实验的开展提供了理论依据。
第四部分天花粉蛋白、米非司酮、顺铂对卵巢癌细胞中HLA-G、HLA-E mRNA及蛋白表达水平的影响
目的:探讨天花粉蛋白、米非司酮、顺铂是否能够通过下调HLA-G、HLA-E的表达,达到抑制肿瘤生长的目的。
方法:继续培养表达HLA-G、HLA-E的卵巢癌OVCAR3细胞株和高表达HLA-G的绒毛膜癌JEG-3细胞株,采用RT-PCR和流式细胞分析技术检测天花粉蛋白、米非司酮、顺铂对细胞中HLA-G、HLA-E mRNA和蛋白表达水平的影响。参照第三部分结果,选取药物浓度:天花粉蛋白注射液浓度为500μg/ml、1000μg/ml,米非司酮终浓度为20μg/ml、40μg/ml,顺铂浓度为6.25μg/ml、12.5μg/ml。
结果:①天花粉蛋白能明显下调OVCAR3细胞中HLA-G mRNA水平,对照组、500μg/ml组、1000μg/ml组光密度值分别为1.00±0.12、0.64±0.24、0.36±0.08,其中1000μg/ml组中HLA-G mRNA光密度值显著低于500μg/ml组(P﹤0.01);天花粉蛋白能明显下调OVCAR3细胞中HLA-E mRNA和蛋白表达水平,对照组、500μg/ml组、1000μg/ml组光密度值分别为1.14±0.09、0.82±0.05、0.65±0.06,荧光指数分别为2.29±0.12、1.77±0.09、1.37±0.11,而且,1000μg/ml组中HLA- E mRNA和蛋白表达水平显著低于500μg/ml组(均为P﹤0.01)。②米非司酮能明显下调OVCAR3细胞中HLA-G mRNA水平,对照组、20μg/ml组、40μg/ml组OVCAR3细胞中光密度值分别为1.02±0.14、0.67±0.14、0.35±0.12;20μg/ml组和40μg/ml组天花粉蛋白能明显下调OVCAR3细胞HLA-E mRNA和蛋白表达水平,均为P﹤0.01。③顺铂对OVCAR3细胞中HLA-G mRNA表达无明显影响,对照组、6.25μg/ml组、12.5μg/ml组HLA-G mRNA光密度值分别为1.02±0.09、0.98±0.11、0.94±0.07,组间方差分析F=1.138,P>0.05;对照组、6.25μg/ml组、12.5μg/ml组OVCAR3细胞中HLA- E mRNA光密度值分别为0.96±0.16、0.94±0.08、0.81±0.11,组间方差分析F=2.601,P>0.05。12.5μg/ml浓度顺铂作用于OVCAR3后,细胞中HLA-E mRNA表达有下降趋势,但无统计学意义(P=0.052)。6.25μg/ml和12.5μg/ml顺铂均能明显下调OVCAR3细胞HLA-E蛋白表达水平,分别为P<0.05、P<0.01。④同样浓度的天花粉蛋白、米非司酮都能明显下调绒癌JEG-3细胞中HLA-G、HLA-E mRNA和蛋白表达水平,均为P<0.01。但顺铂对JEG-3细胞中HLA-G、HLA-E表达的影响无统计学差异。
结论:三种药物可不同程度的拮抗卵巢癌免疫逃逸,天花粉蛋白和米非司酮治疗卵巢癌的作用机制同顺铂有所不同,与顺铂联合应用,可能会取得更好的治疗效果,为进一步动物移植瘤模型观察药物疗效提供了可行性依据。
第五部分天花粉蛋白、米非司酮、顺铂对荷人卵巢癌裸鼠肿瘤生长及移植瘤组织HLA-G、HLA-E mRNA及蛋白表达水平的影响
目的:模拟肿瘤生成的体内环境,进一步获得在体内药物干预后HLA-G、HLA-E水平变化的详实材料,深入探讨天花粉蛋白、米非司酮、顺铂在卵巢恶性肿瘤治疗中的作用机制,为临床药物的合理应用提供更充分有力的动物实验依据。
方法:①培养卵巢癌OVCAR3细胞株。②饲养BALB/c裸鼠,将细胞悬液注入裸鼠右侧肩胛背部,每只0.2ml,含细胞数1.2×106,约7天后明显成瘤,接种成功率为100%。③待裸鼠生长至接种后第2周,肿瘤界限清楚后,将裸鼠随机分为5组,每组7只,包括:对照组、天花粉蛋白组、米非司酮组、顺铂组、顺铂和米非司酮联合用药组。④用药4周后,处死裸鼠,观察各组皮下移植瘤体积,采用RT-PCR方法和流式细胞技术检测不同分组瘤组织中HLA-G、HLA-E mRNA和蛋白表达情况。
结果:①对照组、天花粉蛋白组、米非司酮组、顺铂组及联合组移植瘤体积分别为( 6100.83±1000.69 ) mm3、( 3500.46±562.04 ) mm3、(3689.30±1017.66)mm3、(3491.40±820.13)mm3、(2532.55±762.56)mm3。各用药组抑瘤率均明显小于对照组,米非司酮组和顺铂组对肿瘤的抑制率都明显低于联合组。②对照组、天花粉蛋白组、米非司酮组、顺铂组及联合组HLA-G mRNA表达光密度值分别为1.01±0.17、0.71±0.18、0.62±0.10、0.89±0.11、0.49±0.10,蛋白表达荧光指数分别为2.38±0.43、1.45±0.35、1.26±0.31、2.29±0.35、1.08±0.25。HLA-G mRNA和蛋白表达在顺铂组无明显改变,在其余各组都显著降低。米非司酮和联合组HLA-G mRNA和蛋白表达显著低于顺铂组,前两组相比,差别无统计学差异。HLA-G mRNA和蛋白表达之间有相关性(r=0.692,P﹤0.01)。③对照组、天花粉蛋白组、米非司酮组、顺铂组及联合组HLA-E mRNA表达光密度值分别为0.99±0.19、0.67±0.18、0.76±0.17、0.96±0.16、0.69±0.21,蛋白表达荧光指数分别为2.27±0.19、2.02±0.17、1.59±0.15、2.17±0.19、1.36±0.25。HLA-E mRNA和蛋白表达在顺铂组无明显改变,在其余各组都显著降低,米非司酮和联合组HLA-E mRNA和蛋白表达显著低于顺铂组。HLA-E mRNA和蛋白表达之间有相关性(r=0.472, P﹤0.01)。④HLA-G与HLA- E mRNA表达相关性比较,r=0.388, P﹤0.05;蛋白表达相关性比较,r=0.668, P﹤0.01。
结论:①天花粉蛋白、米非司酮和顺铂都能明显抑制裸鼠皮下抑制瘤生长,米非司酮和顺铂联合应用对肿瘤的抑制作用明显强于单一用药组,表明米非司酮和顺铂对卵巢癌的治疗有协同作用。②与顺铂不同,天花粉蛋白和米非司酮的抗肿瘤作用机制之一是能够明显下调肿瘤组织中HLA-G和HLA-E的表达水平,打破机体对肿瘤的免疫抑制,恢复效应细胞的杀瘤功能,同顺铂联合使用,有望取得更好的治疗效果。③HLA-G mRNA和蛋白表达之间、HLA-E mRNA和蛋白表达之间及HLA-G和HLA-E两种抗原表达之间统计学分析存在相关性,提示临床可以采用比较简单实用的检测手段测定抗原水平,将其作为判断肿瘤恶性程度、监测病情进展的标志物。
The death rate of human ovarian carcinoma is the first in gynecological malignant tumor and most of the patients at advanced stage when they are found. The experts of gynecology all over the world successfully strived and explored the way of cytoreductive surgery and union chemotherapy which based on the operation in the last 30 years. But the survival rate of five years of human ovarian carcinoma patients at advanced stage still rambled about 20%. So to investigate the mechanism of how the human ovarian carcinoma happened and developed and to explore the most valid treatment prescription and elevating the quality of patients who suffered from cancer are great challenge faced by expert of gynecology constantly. Human leukocyte antigen-G (HLA-G) and human leukocyte antigen-E (HLA-E) are both non-classical HLA-I molecules, which discovered in recent years. HLA-G could inhibit human immune function by combining with the rejection capability receptors of NK cells, T cells subtype, antigen presenting cells, B cells and monocaryon cells system. HLA-G could stimulate tumor cells to over-express HLA-E, which could inhibit cytolysis of NK cells and T cells by interacting with the rejection capability receptors of them. Initially HLA-G and HLA-E were observed on the extravillous cytotrophoblast. And the effect on the fetal-maternal immune tolerance has generally accepted by the public. Moreover, the scholar carried out rudiment investigation in the relationship between the antigens and tumor. After the report of being expressed in the melanoma cells in 1998, HLA-G was reported in kidney tumor, breast tumor, colorectal cancer, lung cancer, lymphoma, carcinoma of bladder, spongioblastoma. Most of studies indicated that malignant tumor expressed HLA-G antigen and the expression concerned with immune escape of tumor. But the reports were very few about the expression of the antigens in malignant tumor of ovary and there was no final conclusion to date. It was reported aboard in 2006 that progestin, choriogonadotrophin could achieve the goal of tocolysis by up-regulating the HLA-G level. So we presumed that the mechanism of the inducing abortion drug, such as trichosanthin and mifepristone might concerned with down-regulating the expression of HLA-G and HLA-E. We could also presume the anti-tumor mechanism of these drugs in immune escape aspect, which opened up a new pathway in the immunotherapy and biotherapy of malignant tumor and there was no report about this at present. We tried to analyze the effect of HLA-G and HLA-E in immune escape of tumor by detecting their expression in the tissues of ovarian malignant tumor and cultured cells in vitro, observe the inhibiting effect of trichosanthin, mifepristone and DDP on the ovarian cancer cells and compared the changes from mRNA and protein level of HLA-G and HLA-E. Then we could identify the anti-tumor effect and mechanism of the drugs. We established the models of xenografted tumor by transplanting human ovarian carcinoma OVCAR3 cell into the nude mice and observed the influence of the drugs on the tumor’s bulk and the expression of HLA-G, HLA-E. We expected to search out more valid anticancer drugs at this fundament and provide creative thinking and grounds for the generating mechanism of ovarian cancer and its biotherapy.
Part One The expression and significance of HLA-G and HLA-E in ovarian malignant tumor tissue.
Objective: To investigate the expression of HLA-G and HLA-E in ovarian tumor and normal tissue and its corelation with clinical pathological character.
Methods: Reversed Transcript-Polymerase Chain Reaction (RT-PCR) method and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein levels in 60 malignant tumor and 30 benign tumor of ovary and 10 normal ovarian tissues.
Results:①The positive rate of HLA-G mRNA expression in malignant group,benign group and normal group was 93.33%,73.33%,60.00% respectively, while the positive rate of HLA-G protein expression was 61.67%,16.67%,10.00% respectively. Result ofχ2 analysis showed that the positive rate of HLA-G mRNA and protein expression in malignant group were higher than that in benign group and normal group, while the statistical difference between the benign group and the normal group was not significant.②The positive rate of HLA-E mRNA expression in malignant group,benign group and normal group was 96.67%,83.33%,70.00% respectively, while the positive rate of HLA-E protein expression was 75.00%,36.67%,30.00% respectively. Result ofχ2 analysis showed that the positive rate of HLA-E mRNA in malignant group was higher than that in normal group, and also had higher tendency than in benign group(χ2=3.272, P=0.077); The positive rate of HLA-E protein expression in malignant group was higher than that in benign group and normal group. There was no statistical difference between benign group and normal group .③There was no statistical difference of the HLA-G mRNA and protein expression among different histological typies. Expression of HLA-G mRNA among different clinical stages also showed no statistical difference, but the expression in clinical I/II stage had lower tendency than III/IV stage (χ2=4.133,P=0.077); The expression of HLA-G protein was increasing according with clinical stage. There was statistical difference of the HLA-G mRNA and protein expression among differentation levels, the positive rate in well-differentiated group was lower than that in poorly differentiated group.④There was no statistical difference of the HLA-E mRNA and protein expression among different histological typies. Expression of HLA-E mRNA among different clinical stages also showed no statistical difference, but the expression in clinical I/II stage had lower tendency than III/IV stage (χ2=4.828,P=0.086); The expression of HLA-E protein was increasing according with clinical stage. There was statistical difference of the HLA-E mRNA and protein expression among differentiation levels, the positive rate in well-differentiated group was lower than that in poorly differentiated group.
Conclusion:①The expression of HLA-G and HLA-E were all higher in malignant group than in benign group and normal group.②The expression of HLA-G and HLA-E were not corelating with histological typy of ovarian malignant tumor, but had corelation with clinical stage and pathological differenitation.③HLA-G and HLA-E might participate in the development of ovarian malignant tumor.
Part Two The expression and significance of HLA-G and HLA-E in ovarian cancer cells.
Objective: To investigate the expression of HLA-G and HLA-E in ovarian cancer cells and provide experimental evidence for immunotherapy of ovarian cancer.
Methods: Human ovarian cancer SKOV3, 3AO and OVCAR3 cells were cultured in vitro. RT-PCR and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein levels, and using chorioepithelioma JEG-3 cells as positive control.
Results: The expression of HLA-G, HLA-E mRNA and protein in SKOV3 cells was negative. The expression of HLA-G mRNA and protein in 3AO cells was negative, while the expression of HLA-E mRNA and protein was positive. The expression of HLA-G mRNA in OVCAR3 cells was positive, protein expression was negative; The expression of HLA-E mRNA and protein in OVCAR3 cells was positive. The expression of HLA-G, HLA-E mRNA and protein in JEG-3 cells showed intense positive. Conclusion: Although different ovarian cancer cells showed different levels of HLA-G,HLA-E, the significance of HLA-G,HLA-E in the immune escape mechanism for ovarian cancer was confirmed.
Part Three The effect of trichosanthin,mifepristone, DDP on the proliferation of ovarian cancer cells.
Objective: To investigate the influence of trichosanthin,mifepristone, DDP on the proliferation of ovarian cancer cells and identify theirs anti-tumor role.
Methods: Human ovarian cancer SKOV3, 3AO and OVCAR3 cells were cultured in vitro. MTT method was performed to observe the influence of trichosanthin,mifepristone, DDP on the proliferation of ovarian cancer cells .
Results:①The inhibition ratio of trichosanthin about the concentration of 10μg/ml,50μg/ml,100μg/ml,500μg/ml,1000μg/ml for SKOV3 cell was 0, 3.67%, 18.60%, 34.46%, 46.96%, respectively; For 3AO cell was 0, 2.97%, 9.17%, 35.01%, 48.98%, respectively; For OVCAR3 cell was 1.06%, 9.03%, 14.95%, 39.66%, 58.65%. The higher the concentration, the higher the inhibition ratio.②The inhibition ratio of mifepristone about the concentration of 2.5μg/ml, 5μg/ml, 10μg/ml, 20μg/ml, 40μg/ml for SKOV3 cell was 2.45%, 4.52%, 5.98%, 21.67%, 30.03%, respectively; For 3AO cell was 11.38%, 17.93%, 20.82%, 45.75%, 58.63%, respectively; For OVCAR3 cell was 8%, 24.88%, 47.6%, 61.58%, 76.94%.③The inhibition ratio of DDP about the concentration of 1.5625μg/ ml, 3.125μg/ ml, 6.25μg/ ml, 12.5μg/ ml, 25μg/ ml for SKOV3 cell was 5.68%, 17.27%, 21.75%, 45.32%, 54.98%, respectively;For 3AO cell was 25.55%, 39.81%, 63.07%, 86.93%, 95.28%, respectively; For OVCAR3 cell was 17.82%, 29.25%, 60.15%, 70.57%, 85.67%. Conclusion: Trichosanthin, mifepristone, DDP inhibited the proliferation of ovarian cancer cells obviously , and the inhibition ratio according with the drug concentration, which confirmed the anti-cancer effect of the three drugs for ovarian cancer cells.
Part Four The influence of trichosanthin, mifepristone, DDP on the expression of HLA-G and HLA-E in ovarian cancer cells.
Objective: To investigate whether the expression of HLA-G and HLA-E can be downregulated by trichosanthin, mifepristone, DDP.
Methods: Human ovarian cancer OVCAR3 cells were cultured in vitro, which express both HLA-G and HLA-E. RT-PCR and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein levels when being treated with trichosanthin, mifepristone, DDP respectively.
Results:①The optical density value of HLA-G mRNA expressed by OVCAR3 cells in control group, 500μg/ml group and 1000μg/ml group was 1.00±0.12, 0.64±0.24, 0.36±0.08, respectively. Trichosanthin could down-regulate HLA-G mRNA level significantly and the optical density value of HLA-G mRNA in 1000μg/ml group was lower than that in 500μg/ml group. Trichosanthin could down-regulate HLA-E mRNA and protein levels significantly, the optical density value of HLA-E mRNA expressed in control group, 500μg/ml group and 1000μg/ml group was 1.14±0.09, 0.82±0.05, 0.65±0.06, fluorescence index was 2.29±0.12, 1.77±0.09, 1.37±0.11, respectively.②Mifepristone could down-regulate HLA-G mRNA level significantly, the optical density value in control group, 20μg/ml group and 40μg/ml group was 1.02±0.14, 0.67±0.14, 0.35±0.12, respectively. Mifepristone could down-regulate HLA-E mRNA and protein levels significantly.③DDP had no distinct influence on the HLA-G mRNA level, the optical density value in control group, 6.25μg/ml group and 12.5μg/ml group was 1.02±0.09, 0.98±0.11, 0.94±0.07, respectively. The optical density value of HLA-E mRNA expressed by OVCAR3 cells in control group, 6.25μg/ml group, 12.5μg/ml group was 0.96±0.16, 0.94±0.08, 0.81±0.11, respectively, and the difference was not significant. DDP could down-regulate HLA-E protein expressed significantly.④At the same concentration, trichosanthin and mifepristone could down-regulate the mRNA and protein levels of both HLA-G and HLA-E, while the difference for DDP was not significantly.
Conclusion: The three drugs could rivalry immune escape of ovarian cancer in different degree, combining trichosanthin or mifepristone to DDP might gain the better result.
Part Five The influence of trichosanthin, mifepristone, DDP on the growth of xenografted tumor and the expression of HLA-G and HLA-E
Objective: To investigate the therapy mechanism of trichosanthin, mifepristone, DDP on ovarian cancer .
Methods:①Human ovarian cancer OVCAR3 cells were cultured in vitro.②Raised BALB/c nude mice and infused cell suspension into the right shoulder of nude mice. Everyone of them received 0.2ml, which including 1.2×106 cells. The tumor formed after 7days and the inoculation successful rate was 100%.③When the xenografted tumor growed two weeks and the circumscription of tumor was distinct, the nude mice were randomly divided in five groups, which were control, TCS, MIF ,DDP, and DDP+ MIF group.④The nude mice were killed after 4 weeks’medication, and the volume of tumor were measured. RT-PCR method and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein in the xenografted tumor of nude mice in different groups.
Results:①The xenografted tumor of nude mice in control, TCS, MIF, DDP and DDP+MIF group was (6100.83±1000.69) mm3, (3500.46±562.04) mm3, (3689.30±1017.66) mm3, (3491.40±820.13) mm3 and (2532.55±762.56) mm3 , respectively. The tumor growth inhibiting rates of treated groups was obviously lower than the control group.②The optical density value of HLA-G mRNA in control, TCS, MIF, DDP and DDP+MIF group was 1.01±0.17, 0.71±0.18, 0.62±0.10, 0.89±0.11 and 0.49±0.10, and the fluorescence index number of the HLA-G protein was 2.38±0.43, 1.45±0.35, 1.26±0.31, 2.29±0.35, 1.08±0.25, respectively. Except DDP group, the expression of HLA-G mRNA and protein in treated group were notably lower than the control group., which of MIF and DDP+MIF groups was lower than DDP group.③The optical density value of HLA-E mRNA in control, TCS, MIF, DDP and DDP+MIF group was 0.99±0.19, 0.67±0.18, 0.76±0.17, 0.96±0.16 and 0.69±0.21, and the fluorescence index number of the HLA-E protein was 2.27±0.19, 2.02±0.17, 1.59±0.15, 2.17±0.19 and 1.36±0.25, respectively. Except DDP group, the expression of HLA-E mRNA and protein in treated group were notably lower than the control group.④There was consistency between HLA-G and HLA-E mRNA level(r=0.388, P<0.05) and protein level(r=0.668, P<0.01).
Conclusion: Trichosanthes, mifepristone, DDP could obviously inhibit the growing of the xenograft of nude mice. The depressant effect of DDP+MIF on the tumor was obviously stronger than single drug group. The anti-tumor mechanism of trichosanthin and mifepristone connected with the down-regulating of HLA-G, HLA-E, but DDP had no such effect, which suggested that the therapeutic efficacy would be much better if the DDP combined with trichosanthin or mifepristone.
和理论依据。第一部分卵巢恶性肿瘤HLA-G、HLA-E mRNA及蛋白表达水平与其临床生物学行为的关系
目的:检测HLA-G、HLA-E在良、恶性卵巢肿瘤和正常卵巢组织中的表达及与卵巢恶性肿瘤生物学行为的关系。
方法:采用半定量逆转录-聚合酶链反应(Reversed Transcript-Polymerase Chain Reaction, RT-PCR)和流式细胞分析技术检测60例卵巢恶性肿瘤、30例良性肿瘤和10例正常卵巢组织中HLA-G、HLA-E的mRNA和蛋白水平,并结合肿瘤组织类型、临床分期、病理分级等临床病理特征进行综合分析。
结果:①恶性组、良性组及正常组卵巢组织HLA-G mRNA阳性表达率分别为93.33%、73.33%、60%,蛋白阳性表达率分别为61.67%、16.67%、10%。经χ2检验,恶性组HLA-G mRNA和蛋白阳性表达率均显著高于良性组与正常组;而良性组同正常组相比,统计学差异不显著。②恶性组、良性组及正常组卵巢组织HLA-E mRNA阳性率分别为96.67%、83.33%、70%,蛋白阳性表达率分别为75%、36.67%、30%。经χ2检验,恶性组HLA- E mRNA阳性表达率显著高于正常组,也有高于良性组的趋势(χ2=3.272,P=0.07);恶性组HLA-E蛋白阳性表达率显著高于良性组及正常组。而良性组同正常组相比,HLA-E表达无统计学差异。③不同组织类型之间HLA-G mRNA和蛋白表达均无统计学差异。不同临床分期之间HLA-G mRNA表达无统计学差异,但Ⅰ/Ⅱ期有低于Ⅲ/Ⅳ期的趋势(χ2=4.133,P=0.077);HLA-G蛋白阳性表达率随临床分期进展而增高。不同分化程度之间HLA-G mRNA表达有统计学差异,高分化组HLA-G mRNA和蛋白表达显著低于低分化组。④不同组织类型之间HLA-E mRNA和蛋白表达均无统计学差异。不同临床分期之间HLA- E mRNA表达没有统计学差异,但Ⅰ/Ⅱ期有低于Ⅲ/Ⅳ期的趋势(χ2=4.828,P=0.086);HLA- E蛋白表达率随临床分期进展而增高。不同分化程度之间HLA-E mRNA和蛋白阳性表达率有统计学差异,高分化组显著低于低分化组。
结论:①卵巢恶性肿瘤组HLA-G、HLA-E表达均显著高于良性肿瘤组和正常组;②HLA-G、HLA-E表达与卵巢恶性肿瘤组织类型无关;与临床分期和病理分级显著相关,临床分期越晚、组织分化程度越差,抗原表达水平越高。③HLA-G、HLA-E可能参与了卵巢恶性肿瘤的发生、发展过程,从而使癌细胞产生免疫耐受,逃逸宿主的免疫监视。
第二部分卵巢癌细胞中HLA-G、HLA-E mRNA及蛋白表达水平检测
目的:检测HLA-G、HLA-E在卵巢癌细胞中mRNA和蛋白水平的表达情况,为进一步开展卵巢癌的临床免疫治疗提供细胞基础实验依据。
方法:培养卵巢癌细胞株SKOV3、3AO、OVCAR3,采用RT-PCR和流式细胞分析技术检测细胞中mRNA和蛋白水平的表达情况,并以高表达HLA-G的绒癌JEG-3细胞做为阳性对照。
结果:卵巢癌SKOV3细胞HLA-G、HLA-E mRNA和蛋白表达均为阴性;卵巢癌3AO细胞HLA-G mRNA和蛋白表达为阴性,HLA-E mRNA和蛋白表达为阳性;卵巢癌OVCAR3细胞HLA-G mRNA表达为阳性、蛋白表达为阴性,HLA-E mRNA和蛋白表达均为阳性;绒癌JEG-3细胞HLA-G、HLA-E mRNA和蛋白表达均为强阳性。
结论:虽然不同卵巢癌细胞株表达HLA-G、HLA-E水平有所不同,但还是从细胞水平验证了HLA-G、HLA-E在卵巢癌免疫逃逸方面的作用。并为下部分药物干预研究提供了同时表达HLA-G和HLA-E的卵巢癌OVCAR3细胞系。
第三部分天花粉蛋白、米非司酮、顺铂对卵巢癌细胞生长的影响
目的:检测天花粉蛋白、米非司酮及顺铂对卵巢癌细胞系生长的影响,明确药物的抗肿瘤作用。
方法:培养卵巢癌细胞株SKOV3、3AO、OVCAR3,MTT技术观察不同浓度天花粉蛋白、米非司酮、顺铂对三种细胞株生长增殖能力的影响。
结果:①10μg/ml、50μg/ml、100μg/ml、500μg/ml、1000μg/ml天花粉蛋白对SKOV3细胞的生长抑制率分别为0、3.67%、18.60%、34.46%、46.96%;对3AO细胞分别为0、2.97%、9.17%、35.01%、48.98%;对OVCAR3细胞分别为1.06%、9.03%、14.95%、39.66%、58.65%。药物浓度越高,对细胞生长抑制率越强。②2.5μg/ml、5μg/ml、10μg/ml、20μg/ml、40μg/ml米非司酮对SKOV3细胞的生长抑制率分别为2.45%、4.52%、5.98%、21.67%、30.03%;对3AO细胞分别为11.38%、17.93%、20.82%、45.75%、58.63%;对OVCAR3细胞分别为8%、24.88%、47.6%、61.58%、76.94%。③1.5625μg/ ml、3.125μg/ ml、6.25μg/ ml、12.5μg/ ml、25μg/ ml顺铂对SKOV3细胞的生长抑制率分别为5.68%、17.27%、21.75%、45.32%、54.98%;对3AO细胞分别为25.55%、39.81%、63.07%、86.93%、95.28%;对OVCAR3细胞分别为17.82%、29.25%、60.15%、70.57%、85.67%。顺铂对细胞生长抑制作用呈浓度依赖性。
结论:天花粉蛋白、米非司酮和顺铂对卵巢癌细胞SKOV3、3AO、OVCAR3生长具有明显抑制作用,并呈浓度依赖性。从体外实验明确了天花粉蛋白、米非司酮、顺铂具有抗卵巢癌作用,为进一步药物机制的研究及动物体内实验的开展提供了理论依据。
第四部分天花粉蛋白、米非司酮、顺铂对卵巢癌细胞中HLA-G、HLA-E mRNA及蛋白表达水平的影响
目的:探讨天花粉蛋白、米非司酮、顺铂是否能够通过下调HLA-G、HLA-E的表达,达到抑制肿瘤生长的目的。
方法:继续培养表达HLA-G、HLA-E的卵巢癌OVCAR3细胞株和高表达HLA-G的绒毛膜癌JEG-3细胞株,采用RT-PCR和流式细胞分析技术检测天花粉蛋白、米非司酮、顺铂对细胞中HLA-G、HLA-E mRNA和蛋白表达水平的影响。参照第三部分结果,选取药物浓度:天花粉蛋白注射液浓度为500μg/ml、1000μg/ml,米非司酮终浓度为20μg/ml、40μg/ml,顺铂浓度为6.25μg/ml、12.5μg/ml。
结果:①天花粉蛋白能明显下调OVCAR3细胞中HLA-G mRNA水平,对照组、500μg/ml组、1000μg/ml组光密度值分别为1.00±0.12、0.64±0.24、0.36±0.08,其中1000μg/ml组中HLA-G mRNA光密度值显著低于500μg/ml组(P﹤0.01);天花粉蛋白能明显下调OVCAR3细胞中HLA-E mRNA和蛋白表达水平,对照组、500μg/ml组、1000μg/ml组光密度值分别为1.14±0.09、0.82±0.05、0.65±0.06,荧光指数分别为2.29±0.12、1.77±0.09、1.37±0.11,而且,1000μg/ml组中HLA- E mRNA和蛋白表达水平显著低于500μg/ml组(均为P﹤0.01)。②米非司酮能明显下调OVCAR3细胞中HLA-G mRNA水平,对照组、20μg/ml组、40μg/ml组OVCAR3细胞中光密度值分别为1.02±0.14、0.67±0.14、0.35±0.12;20μg/ml组和40μg/ml组天花粉蛋白能明显下调OVCAR3细胞HLA-E mRNA和蛋白表达水平,均为P﹤0.01。③顺铂对OVCAR3细胞中HLA-G mRNA表达无明显影响,对照组、6.25μg/ml组、12.5μg/ml组HLA-G mRNA光密度值分别为1.02±0.09、0.98±0.11、0.94±0.07,组间方差分析F=1.138,P>0.05;对照组、6.25μg/ml组、12.5μg/ml组OVCAR3细胞中HLA- E mRNA光密度值分别为0.96±0.16、0.94±0.08、0.81±0.11,组间方差分析F=2.601,P>0.05。12.5μg/ml浓度顺铂作用于OVCAR3后,细胞中HLA-E mRNA表达有下降趋势,但无统计学意义(P=0.052)。6.25μg/ml和12.5μg/ml顺铂均能明显下调OVCAR3细胞HLA-E蛋白表达水平,分别为P<0.05、P<0.01。④同样浓度的天花粉蛋白、米非司酮都能明显下调绒癌JEG-3细胞中HLA-G、HLA-E mRNA和蛋白表达水平,均为P<0.01。但顺铂对JEG-3细胞中HLA-G、HLA-E表达的影响无统计学差异。
结论:三种药物可不同程度的拮抗卵巢癌免疫逃逸,天花粉蛋白和米非司酮治疗卵巢癌的作用机制同顺铂有所不同,与顺铂联合应用,可能会取得更好的治疗效果,为进一步动物移植瘤模型观察药物疗效提供了可行性依据。
第五部分天花粉蛋白、米非司酮、顺铂对荷人卵巢癌裸鼠肿瘤生长及移植瘤组织HLA-G、HLA-E mRNA及蛋白表达水平的影响
目的:模拟肿瘤生成的体内环境,进一步获得在体内药物干预后HLA-G、HLA-E水平变化的详实材料,深入探讨天花粉蛋白、米非司酮、顺铂在卵巢恶性肿瘤治疗中的作用机制,为临床药物的合理应用提供更充分有力的动物实验依据。
方法:①培养卵巢癌OVCAR3细胞株。②饲养BALB/c裸鼠,将细胞悬液注入裸鼠右侧肩胛背部,每只0.2ml,含细胞数1.2×106,约7天后明显成瘤,接种成功率为100%。③待裸鼠生长至接种后第2周,肿瘤界限清楚后,将裸鼠随机分为5组,每组7只,包括:对照组、天花粉蛋白组、米非司酮组、顺铂组、顺铂和米非司酮联合用药组。④用药4周后,处死裸鼠,观察各组皮下移植瘤体积,采用RT-PCR方法和流式细胞技术检测不同分组瘤组织中HLA-G、HLA-E mRNA和蛋白表达情况。
结果:①对照组、天花粉蛋白组、米非司酮组、顺铂组及联合组移植瘤体积分别为( 6100.83±1000.69 ) mm3、( 3500.46±562.04 ) mm3、(3689.30±1017.66)mm3、(3491.40±820.13)mm3、(2532.55±762.56)mm3。各用药组抑瘤率均明显小于对照组,米非司酮组和顺铂组对肿瘤的抑制率都明显低于联合组。②对照组、天花粉蛋白组、米非司酮组、顺铂组及联合组HLA-G mRNA表达光密度值分别为1.01±0.17、0.71±0.18、0.62±0.10、0.89±0.11、0.49±0.10,蛋白表达荧光指数分别为2.38±0.43、1.45±0.35、1.26±0.31、2.29±0.35、1.08±0.25。HLA-G mRNA和蛋白表达在顺铂组无明显改变,在其余各组都显著降低。米非司酮和联合组HLA-G mRNA和蛋白表达显著低于顺铂组,前两组相比,差别无统计学差异。HLA-G mRNA和蛋白表达之间有相关性(r=0.692,P﹤0.01)。③对照组、天花粉蛋白组、米非司酮组、顺铂组及联合组HLA-E mRNA表达光密度值分别为0.99±0.19、0.67±0.18、0.76±0.17、0.96±0.16、0.69±0.21,蛋白表达荧光指数分别为2.27±0.19、2.02±0.17、1.59±0.15、2.17±0.19、1.36±0.25。HLA-E mRNA和蛋白表达在顺铂组无明显改变,在其余各组都显著降低,米非司酮和联合组HLA-E mRNA和蛋白表达显著低于顺铂组。HLA-E mRNA和蛋白表达之间有相关性(r=0.472, P﹤0.01)。④HLA-G与HLA- E mRNA表达相关性比较,r=0.388, P﹤0.05;蛋白表达相关性比较,r=0.668, P﹤0.01。
结论:①天花粉蛋白、米非司酮和顺铂都能明显抑制裸鼠皮下抑制瘤生长,米非司酮和顺铂联合应用对肿瘤的抑制作用明显强于单一用药组,表明米非司酮和顺铂对卵巢癌的治疗有协同作用。②与顺铂不同,天花粉蛋白和米非司酮的抗肿瘤作用机制之一是能够明显下调肿瘤组织中HLA-G和HLA-E的表达水平,打破机体对肿瘤的免疫抑制,恢复效应细胞的杀瘤功能,同顺铂联合使用,有望取得更好的治疗效果。③HLA-G mRNA和蛋白表达之间、HLA-E mRNA和蛋白表达之间及HLA-G和HLA-E两种抗原表达之间统计学分析存在相关性,提示临床可以采用比较简单实用的检测手段测定抗原水平,将其作为判断肿瘤恶性程度、监测病情进展的标志物。
The death rate of human ovarian carcinoma is the first in gynecological malignant tumor and most of the patients at advanced stage when they are found. The experts of gynecology all over the world successfully strived and explored the way of cytoreductive surgery and union chemotherapy which based on the operation in the last 30 years. But the survival rate of five years of human ovarian carcinoma patients at advanced stage still rambled about 20%. So to investigate the mechanism of how the human ovarian carcinoma happened and developed and to explore the most valid treatment prescription and elevating the quality of patients who suffered from cancer are great challenge faced by expert of gynecology constantly. Human leukocyte antigen-G (HLA-G) and human leukocyte antigen-E (HLA-E) are both non-classical HLA-I molecules, which discovered in recent years. HLA-G could inhibit human immune function by combining with the rejection capability receptors of NK cells, T cells subtype, antigen presenting cells, B cells and monocaryon cells system. HLA-G could stimulate tumor cells to over-express HLA-E, which could inhibit cytolysis of NK cells and T cells by interacting with the rejection capability receptors of them. Initially HLA-G and HLA-E were observed on the extravillous cytotrophoblast. And the effect on the fetal-maternal immune tolerance has generally accepted by the public. Moreover, the scholar carried out rudiment investigation in the relationship between the antigens and tumor. After the report of being expressed in the melanoma cells in 1998, HLA-G was reported in kidney tumor, breast tumor, colorectal cancer, lung cancer, lymphoma, carcinoma of bladder, spongioblastoma. Most of studies indicated that malignant tumor expressed HLA-G antigen and the expression concerned with immune escape of tumor. But the reports were very few about the expression of the antigens in malignant tumor of ovary and there was no final conclusion to date. It was reported aboard in 2006 that progestin, choriogonadotrophin could achieve the goal of tocolysis by up-regulating the HLA-G level. So we presumed that the mechanism of the inducing abortion drug, such as trichosanthin and mifepristone might concerned with down-regulating the expression of HLA-G and HLA-E. We could also presume the anti-tumor mechanism of these drugs in immune escape aspect, which opened up a new pathway in the immunotherapy and biotherapy of malignant tumor and there was no report about this at present. We tried to analyze the effect of HLA-G and HLA-E in immune escape of tumor by detecting their expression in the tissues of ovarian malignant tumor and cultured cells in vitro, observe the inhibiting effect of trichosanthin, mifepristone and DDP on the ovarian cancer cells and compared the changes from mRNA and protein level of HLA-G and HLA-E. Then we could identify the anti-tumor effect and mechanism of the drugs. We established the models of xenografted tumor by transplanting human ovarian carcinoma OVCAR3 cell into the nude mice and observed the influence of the drugs on the tumor’s bulk and the expression of HLA-G, HLA-E. We expected to search out more valid anticancer drugs at this fundament and provide creative thinking and grounds for the generating mechanism of ovarian cancer and its biotherapy.
Part One The expression and significance of HLA-G and HLA-E in ovarian malignant tumor tissue.
Objective: To investigate the expression of HLA-G and HLA-E in ovarian tumor and normal tissue and its corelation with clinical pathological character.
Methods: Reversed Transcript-Polymerase Chain Reaction (RT-PCR) method and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein levels in 60 malignant tumor and 30 benign tumor of ovary and 10 normal ovarian tissues.
Results:①The positive rate of HLA-G mRNA expression in malignant group,benign group and normal group was 93.33%,73.33%,60.00% respectively, while the positive rate of HLA-G protein expression was 61.67%,16.67%,10.00% respectively. Result ofχ2 analysis showed that the positive rate of HLA-G mRNA and protein expression in malignant group were higher than that in benign group and normal group, while the statistical difference between the benign group and the normal group was not significant.②The positive rate of HLA-E mRNA expression in malignant group,benign group and normal group was 96.67%,83.33%,70.00% respectively, while the positive rate of HLA-E protein expression was 75.00%,36.67%,30.00% respectively. Result ofχ2 analysis showed that the positive rate of HLA-E mRNA in malignant group was higher than that in normal group, and also had higher tendency than in benign group(χ2=3.272, P=0.077); The positive rate of HLA-E protein expression in malignant group was higher than that in benign group and normal group. There was no statistical difference between benign group and normal group .③There was no statistical difference of the HLA-G mRNA and protein expression among different histological typies. Expression of HLA-G mRNA among different clinical stages also showed no statistical difference, but the expression in clinical I/II stage had lower tendency than III/IV stage (χ2=4.133,P=0.077); The expression of HLA-G protein was increasing according with clinical stage. There was statistical difference of the HLA-G mRNA and protein expression among differentation levels, the positive rate in well-differentiated group was lower than that in poorly differentiated group.④There was no statistical difference of the HLA-E mRNA and protein expression among different histological typies. Expression of HLA-E mRNA among different clinical stages also showed no statistical difference, but the expression in clinical I/II stage had lower tendency than III/IV stage (χ2=4.828,P=0.086); The expression of HLA-E protein was increasing according with clinical stage. There was statistical difference of the HLA-E mRNA and protein expression among differentiation levels, the positive rate in well-differentiated group was lower than that in poorly differentiated group.
Conclusion:①The expression of HLA-G and HLA-E were all higher in malignant group than in benign group and normal group.②The expression of HLA-G and HLA-E were not corelating with histological typy of ovarian malignant tumor, but had corelation with clinical stage and pathological differenitation.③HLA-G and HLA-E might participate in the development of ovarian malignant tumor.
Part Two The expression and significance of HLA-G and HLA-E in ovarian cancer cells.
Objective: To investigate the expression of HLA-G and HLA-E in ovarian cancer cells and provide experimental evidence for immunotherapy of ovarian cancer.
Methods: Human ovarian cancer SKOV3, 3AO and OVCAR3 cells were cultured in vitro. RT-PCR and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein levels, and using chorioepithelioma JEG-3 cells as positive control.
Results: The expression of HLA-G, HLA-E mRNA and protein in SKOV3 cells was negative. The expression of HLA-G mRNA and protein in 3AO cells was negative, while the expression of HLA-E mRNA and protein was positive. The expression of HLA-G mRNA in OVCAR3 cells was positive, protein expression was negative; The expression of HLA-E mRNA and protein in OVCAR3 cells was positive. The expression of HLA-G, HLA-E mRNA and protein in JEG-3 cells showed intense positive. Conclusion: Although different ovarian cancer cells showed different levels of HLA-G,HLA-E, the significance of HLA-G,HLA-E in the immune escape mechanism for ovarian cancer was confirmed.
Part Three The effect of trichosanthin,mifepristone, DDP on the proliferation of ovarian cancer cells.
Objective: To investigate the influence of trichosanthin,mifepristone, DDP on the proliferation of ovarian cancer cells and identify theirs anti-tumor role.
Methods: Human ovarian cancer SKOV3, 3AO and OVCAR3 cells were cultured in vitro. MTT method was performed to observe the influence of trichosanthin,mifepristone, DDP on the proliferation of ovarian cancer cells .
Results:①The inhibition ratio of trichosanthin about the concentration of 10μg/ml,50μg/ml,100μg/ml,500μg/ml,1000μg/ml for SKOV3 cell was 0, 3.67%, 18.60%, 34.46%, 46.96%, respectively; For 3AO cell was 0, 2.97%, 9.17%, 35.01%, 48.98%, respectively; For OVCAR3 cell was 1.06%, 9.03%, 14.95%, 39.66%, 58.65%. The higher the concentration, the higher the inhibition ratio.②The inhibition ratio of mifepristone about the concentration of 2.5μg/ml, 5μg/ml, 10μg/ml, 20μg/ml, 40μg/ml for SKOV3 cell was 2.45%, 4.52%, 5.98%, 21.67%, 30.03%, respectively; For 3AO cell was 11.38%, 17.93%, 20.82%, 45.75%, 58.63%, respectively; For OVCAR3 cell was 8%, 24.88%, 47.6%, 61.58%, 76.94%.③The inhibition ratio of DDP about the concentration of 1.5625μg/ ml, 3.125μg/ ml, 6.25μg/ ml, 12.5μg/ ml, 25μg/ ml for SKOV3 cell was 5.68%, 17.27%, 21.75%, 45.32%, 54.98%, respectively;For 3AO cell was 25.55%, 39.81%, 63.07%, 86.93%, 95.28%, respectively; For OVCAR3 cell was 17.82%, 29.25%, 60.15%, 70.57%, 85.67%. Conclusion: Trichosanthin, mifepristone, DDP inhibited the proliferation of ovarian cancer cells obviously , and the inhibition ratio according with the drug concentration, which confirmed the anti-cancer effect of the three drugs for ovarian cancer cells.
Part Four The influence of trichosanthin, mifepristone, DDP on the expression of HLA-G and HLA-E in ovarian cancer cells.
Objective: To investigate whether the expression of HLA-G and HLA-E can be downregulated by trichosanthin, mifepristone, DDP.
Methods: Human ovarian cancer OVCAR3 cells were cultured in vitro, which express both HLA-G and HLA-E. RT-PCR and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein levels when being treated with trichosanthin, mifepristone, DDP respectively.
Results:①The optical density value of HLA-G mRNA expressed by OVCAR3 cells in control group, 500μg/ml group and 1000μg/ml group was 1.00±0.12, 0.64±0.24, 0.36±0.08, respectively. Trichosanthin could down-regulate HLA-G mRNA level significantly and the optical density value of HLA-G mRNA in 1000μg/ml group was lower than that in 500μg/ml group. Trichosanthin could down-regulate HLA-E mRNA and protein levels significantly, the optical density value of HLA-E mRNA expressed in control group, 500μg/ml group and 1000μg/ml group was 1.14±0.09, 0.82±0.05, 0.65±0.06, fluorescence index was 2.29±0.12, 1.77±0.09, 1.37±0.11, respectively.②Mifepristone could down-regulate HLA-G mRNA level significantly, the optical density value in control group, 20μg/ml group and 40μg/ml group was 1.02±0.14, 0.67±0.14, 0.35±0.12, respectively. Mifepristone could down-regulate HLA-E mRNA and protein levels significantly.③DDP had no distinct influence on the HLA-G mRNA level, the optical density value in control group, 6.25μg/ml group and 12.5μg/ml group was 1.02±0.09, 0.98±0.11, 0.94±0.07, respectively. The optical density value of HLA-E mRNA expressed by OVCAR3 cells in control group, 6.25μg/ml group, 12.5μg/ml group was 0.96±0.16, 0.94±0.08, 0.81±0.11, respectively, and the difference was not significant. DDP could down-regulate HLA-E protein expressed significantly.④At the same concentration, trichosanthin and mifepristone could down-regulate the mRNA and protein levels of both HLA-G and HLA-E, while the difference for DDP was not significantly.
Conclusion: The three drugs could rivalry immune escape of ovarian cancer in different degree, combining trichosanthin or mifepristone to DDP might gain the better result.
Part Five The influence of trichosanthin, mifepristone, DDP on the growth of xenografted tumor and the expression of HLA-G and HLA-E
Objective: To investigate the therapy mechanism of trichosanthin, mifepristone, DDP on ovarian cancer .
Methods:①Human ovarian cancer OVCAR3 cells were cultured in vitro.②Raised BALB/c nude mice and infused cell suspension into the right shoulder of nude mice. Everyone of them received 0.2ml, which including 1.2×106 cells. The tumor formed after 7days and the inoculation successful rate was 100%.③When the xenografted tumor growed two weeks and the circumscription of tumor was distinct, the nude mice were randomly divided in five groups, which were control, TCS, MIF ,DDP, and DDP+ MIF group.④The nude mice were killed after 4 weeks’medication, and the volume of tumor were measured. RT-PCR method and flow cytometry were performed to examine the expression of HLA-G, HLA-E mRNA and protein in the xenografted tumor of nude mice in different groups.
Results:①The xenografted tumor of nude mice in control, TCS, MIF, DDP and DDP+MIF group was (6100.83±1000.69) mm3, (3500.46±562.04) mm3, (3689.30±1017.66) mm3, (3491.40±820.13) mm3 and (2532.55±762.56) mm3 , respectively. The tumor growth inhibiting rates of treated groups was obviously lower than the control group.②The optical density value of HLA-G mRNA in control, TCS, MIF, DDP and DDP+MIF group was 1.01±0.17, 0.71±0.18, 0.62±0.10, 0.89±0.11 and 0.49±0.10, and the fluorescence index number of the HLA-G protein was 2.38±0.43, 1.45±0.35, 1.26±0.31, 2.29±0.35, 1.08±0.25, respectively. Except DDP group, the expression of HLA-G mRNA and protein in treated group were notably lower than the control group., which of MIF and DDP+MIF groups was lower than DDP group.③The optical density value of HLA-E mRNA in control, TCS, MIF, DDP and DDP+MIF group was 0.99±0.19, 0.67±0.18, 0.76±0.17, 0.96±0.16 and 0.69±0.21, and the fluorescence index number of the HLA-E protein was 2.27±0.19, 2.02±0.17, 1.59±0.15, 2.17±0.19 and 1.36±0.25, respectively. Except DDP group, the expression of HLA-E mRNA and protein in treated group were notably lower than the control group.④There was consistency between HLA-G and HLA-E mRNA level(r=0.388, P<0.05) and protein level(r=0.668, P<0.01).
Conclusion: Trichosanthes, mifepristone, DDP could obviously inhibit the growing of the xenograft of nude mice. The depressant effect of DDP+MIF on the tumor was obviously stronger than single drug group. The anti-tumor mechanism of trichosanthin and mifepristone connected with the down-regulating of HLA-G, HLA-E, but DDP had no such effect, which suggested that the therapeutic efficacy would be much better if the DDP combined with trichosanthin or mifepristone.
引文
1. Favier B, LeMaoult J, Rouas-Freiss N, et al. Research on HLA-G: an update.Tissue Antigens, 2007, 69(3): 207-211
2. El Chérif Ibrahim, Nadia Guerra, Marie-José Terrier Lacombe. Tumor-specific Up-Regulation of the Nonclassical Class I HLA-G Antigen Expression in Renal Carcinoma. Cancer Research , 2001, 9(15): 6838-6845
3. Hansel DE, Rahman A, Wilentz RE, et al. HLA-G upregulation in pre-malignant and malignant lesions of the gastrointestinal tract. Int J Gastrointest Cancer, 2005, 35(1): 15-23
4. El-Chennawi FA, Auf FA, El-Diasty AM, et al. Expression of HLA-G in cancer bladder. Egypt J Immunol, 2005, 12(1): 57-64
5. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cells express HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
6. Monique H. Hurks, Markus M, Valter,et al. Uveal Melanoma: No Expression of HLA-G. Investigative Ophthalmology and Visual Science, 2001, 42: 3081-3084
7. Salvadori S, Martinelli G, Zier K. Resection of solid tumors reverses T cell defects and restores protective immunity. J Immunol, 2000, 164(4): 2214-20
8. Carosella ED, Paul P, Moreau P, et al. HLA-G and HLA-E: fundamental and pathophysiological aspects. Immunol Today, 2000, 21(11): 532-534
9. Riteau B, Rouas-Freiss N, Menier C, et al. HLA-G2, -G3, and -G4 isoforms expressed as nonmature cell surface glycoproteins inhibit NK and antigen-specific CTL cytolysis. J Immunol, 2001, 166(8): 5018-5026
10. Paul P, Rouas-Freiss N, Khalil-Daher I, et al. HLA-G expression in melanoma: a way for tumor cells to escape from immunosurveillance. Proc Natl Acad Sci U S A, 1998, 95(8): 4510-4515
11. Paul P, Cabestré FA, Le Gal FA,et al. Heterogeneity of HLA-G gene transcription and protein expression in malignant melanoma biopsies. Cancer Res, 1999, 59(8): 1954-1960
12. Ibrahim el C, Aractingi S, Allory Y,et al. Analysis of HLA antigen expression in benign and malignant melanocytic lesions reveals that upregulation of HLA-G expression correlates with malignant transformation, high inflammatory infiltration and HLA-A1 genotype. Int J Cancer, 2004, 108(2): 243-250
13. Yie SM, Yang H, Ye SR, et al. Expression of HLA-G is associated with prognosis in esophageal squamous cell carcinoma. Am J Clin Pathol, 2007, 128(6):1002~1009
14. Yie SM, Yang H, Ye SR, et al. Expression of human leukocyte antigen G (HLA-G) correlates with poor prognosis in gastric carcinoma. Ann Surg Oncol, 2007, 14(10): 2721-2729
15. Barrier BF, Kendall BS, Sharpe-Timms KL, et al. Characterization of human leukocyte antigen-G (HLA-G) expression in endometrial adenocarcinoma. Gynecol Oncol, 2006, 103(1): 25-30
16. Bukur J, Seliger B. The role of HLA-G for protection of human renal cell-carcinoma cells from immune-mediated lysis: implications for immunotherapies. Semin Cancer Biol, 2003, 13(5): 353-359
17. Middleton D, Curran M, Maxwell L. Natural killer cells and their receptors. Transpl Immunol, 2002, 10(2-3): 147-64
18. Moscoso J, Serrano-Vela JI, Pacheco R, et al. HLA-G, -E and -F: allelism, function and evolution. Transpl Immunol, 2006, 17(1): 61-64
19. Moretta L, Romagnani C, Pietra G, et al. NK-CTLs, a novel HLA-E-restricted T-cell subset. Trends Immunol, 2003, 24(3): 136-143
20. 张彩, 田志刚, 魏海明, 等. 非经典HLA I类分子（HLA-G和HLA-E）在人肿瘤细胞系的表达及的调节作用. 中国肿瘤临床, 2001, 28(4): 247-253
21. Derré L, Corvaisier M, Charreau B, et al. Expression and release of HLA-E by melanoma cells and melanocytes: potential impact on the response of cytotoxic effector cells. J Immunol, 2006, 177(5): 3100-3107
22. Wischhusen J, Friese MA, Mittelbronn M, et al. HLA-E protects glioma cells from NKG2D-mediated immune responses in vitro: implications for immune escape in vivo.J Neuropathol Exp Neurol, 2005, 64(6): 523-528
1. Favier B, LeMaoult J, Rouas-Freiss N, et al. Research on HLA-G: an update.Tissue Antigens, 2007, 69(3): 207-211
2. Smyth MJ, Hayakawa Y, Takeda K, et al. New aspects of natural killer-cell surveillance and therapy of cancer. Nat Rev Cancer, 2002, 2(11): 856-861
3. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3593
4. Middleton D, Curran M, Maxwell L. Natural killer cells and their receptors. Transplant Immunol, 2002, 10: 147-164
5. Biaddoni R, Cantoni C, Pende D, et al. Human natural killer cell receptors and co-receptors. Immunol Rev, 2001, 181: 203-214
6. Moscoso J, Serrano-Vela JI, Pacheco R, et al. HLA-G, -E and -F: allelism, function and evolution. Transpl Immunol, 2006, 17(1): 61-64
7. Blaschitz A, Hutter H, Leitner V, et al. Reaction patterns of monoclonal antibodies to HLA-G in human tissues and on cell lines: a comparative study. Hum, Immunol, 2000, 61(11): 1074-1085
8. Riteau B, Rouas-Freiss N, Menier C, et al. HLA-G2, -G3, and -G4 isoforms expressed as nonmature cell surface glycoproteins inhibit NK and antigen-specific CTL cytolysis. J Immunol, 2001, 166(8): 5018-5026
9. Bukur J, Malenica B, Huber C, et al. Altered expression of nonclassical HLA class Ib antigens in human renal cell carcinoma and its association with impaired immune response. Hum Immunol, 2003, 64(11): 1081-1092
10. Chang CC, Murphy SP, Ferrone S. Differential in vivo and in vitro HLA-G expression in melanoma cells: potential mechanisms. Hum Immunol, 2003, 64(11): 1057-1063
11. Lefebvre S, Antoine M, Uzan S, et al. Specific activation of the non-classical class I histocompatibility HLA-G antigen and expression of the ILT2 inhibitory receptor in human breast cancer. J Pathol, 2002, 196(3): 266-274
12. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cellsexpress HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
13. Paul P, Cabestré FA, Le Gal FA, et al. Heterogeneity of HLA-G gene transcription and protein expression in malignant melanoma biopsies. Cancer Res, 1999, 59(8): 1954-1960
14. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3596
1. 天花粉蛋白对胃癌细胞株SGC-7901凋亡影响的体外研究. 中国医药导报, 2007, 16(2): 231-234
2. 王媛媛, 欧阳东云, 郑永唐. 天花粉蛋白体外抗人白血病和淋巴瘤细胞的作用机制. 中国试验血液学杂志, 2007, 15(4): 729-732
3. 周欣阳, 张天一, 施海燕, 等. 天花粉蛋白诱导H22肝癌细胞凋亡的研究. 南通医学院学报, 2003, 23(4): 371-374
4. 黄益玲, 黄利鸣, 石新兰, 等. 天花粉蛋白对宫颈癌HeLa细胞增殖和细胞凋亡的影响. 中国药理学通报, 2005, 21: 253-254
5. 孙璟, 涂水平, 吴裕炘, 等. 天花粉蛋白诱导胃癌细胞MKN-45凋亡中P53、Bcl-2、C-myc蛋白表达变化. 上海医学, 2004, 24(5): 292-294
6. 田维毅, 马春玲, 白惠卿. 天花粉及其组份对小鼠NK细胞杀伤活性的影响. 贵州中医, 2001, 25(11): 982-984
7. 邬伟秀, 郭峰. 天花粉对血液肿瘤患者红细胞免疫功能影响的试验研究. 上海免疫学杂志, 1995, 15(3): 156
8. Li A, Felix JC, Minoo P, et al. Effect of mifepristone on proliferation and apoptosis of Ishikawa endometrial adenocarcinoma cells. Fertil Steril, 2005, 84(1): 202-211
9. Poole AJ, Li Y, Kim Y, et al. Prevention of Brca1-mediated mammary tumorigenesis in mice by a progesterone antagonist. Science, 2006, 314(5804): 1467-1470
10. Goyeneche AA, Carón RW, Telleria CM. Mifepristone inhibits ovarian cancer cell growth in vitro and in vivo. Clin Cancer Res, 2007, 13(11): 3370-3379
11. Grunberg SM, Weiss MH, Russell CA, et al. Long-term administration of mifepristone (RU486): clinical tolerance during extended treatment of meningioma. Cancer Invest, 2006, 24(8): 727-733
12. Li DQ, Wang ZB, Bai J, et al. Effects of mifepristone on proliferation of human gastric adenocarcinoma cell line SGC-7901 in vitro. World J Gastroenterol, 2004, 10(18): 2628-2631
13. Labriola L, Salatino M, Proietti CJ, et al. Heregulin induces transcriptional activation of the progesterone receptor by a mechanism that requires functional ErbB-2 and mitogen-activated protein kinase activation in breast cancer cells. Mol Cell Biol, 2003, 23(3): 1095-1111
14. Sridhar S, Ali AA, Liang Y,et al. Differential expression of members of the tumor necrosis factor alpha-related apoptosis-inducing ligand pathway in prostate cancer cells. Cancer Res, 2001, 61(19): 7179-7183
15. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
16. Fukuda H, He PJ, Yokota K, et al. Progesterone-dependent and -independent expression of the multidrug resistance type I gene in porcine granulosa cells. Mol Cell Biochem, 2007, 298(1-2): 179-186
17. Kacinski BM, Flick MB, Sapi E. RU-486 can abolish glucocorticoid-induced increases in CSF-1 receptor expression in primary human breast carcinoma specimens. J Soc Gynecol Investig, 2001, 8(2): 114-116
18. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
19. Hyder SM, Chiappetta C, Stancel GM. Pharmacological and endogenous progestins induce vascular endothelial growth factor expression in human breast cancer cells. Int J Cancer, 2001, 92(4): 469-473
20. Schoenlein PV, Hou M, Samaddar JS, et al. Downregulation of retinoblastoma protein is involved in the enhanced cytotoxicity of 4-hydroxytamoxifen plus mifepristone combination therapy versus antiestrogen monotherapy of human breast cancer. Int J Oncol, 2007, 31(3): 643-655
1 Malmberg KJ, Levitsky V, Norell H, et al. IFN-gamma protects short-term ovarian carcinoma cell lines from CTL lysis via a CD94/NKG2A-dependent mechanism. J Clin Invest, 2002, 110(10): 1515-1523
2. Yie SM, Li LH, Li GM, et al. Progesterone enhances HLA-G gene expression in JEG-3 choriocarcinoma cells and human cytotrophoblasts in vitro. Hum Reprod, 2006, 21(1): 46-51
3. 陈春玲 廖秦平. hCG 促进绒癌细胞系中HLA - G的表达. 中国妇产科临床杂志, 2004, 5(2): 113-115
4. 黄益玲, 黄利鸣, 石新兰, 等. 天花粉蛋白对宫颈癌HeLa细胞增殖和细胞凋亡的影响. 中国药理学通报, 2005, 21: 253-254
5. 孙璟, 涂水平, 吴裕炘, 等. 天花粉蛋白诱导胃癌细胞MKN-45凋亡中P53、Bcl-2、C-myc蛋白表达变化. 上海医学, 2004, 24(5): 292-294
6. 田维毅, 马春玲, 白惠卿. 天花粉及其组份对小鼠NK细胞杀伤活性的影响. 贵州中医, 2001, 25(11): 982-984
7. 邬伟秀, 郭峰. 天花粉对血液肿瘤患者红细胞免疫功能影响的试验研究. 上海免疫学杂志, 1995, 15(3): 156
8. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
9. Sridhar S, Ali AA, Liang Y,et al. Differential expression of members of the tumor necrosis factor alpha-related apoptosis-inducing ligand pathway in prostate cancer cells. Cancer Res, 2001, 61(19): 7179-7183
10. Fukuda H, He PJ, Yokota K, et al. Progesterone-dependent and -independent expression of the multidrug resistance type I gene in porcine granulosa cells. Mol Cell Biochem, 2007, 298(1-2): 179-186
11. Kacinski BM, Flick MB, Sapi E. RU-486 can abolish glucocorticoid-induced increases in CSF-1 receptor expression in primary human breast carcinoma specimens. J Soc Gynecol Investig, 2001, 8(2): 114-116
12. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
13. Hyder SM, Chiappetta C, Stancel GM. Pharmacological and endogenous progestins induce vascular endothelial growth factor expression in human breast cancer cells. Int J Cancer, 2001, 92(4): 469-473
14. Schoenlein PV, Hou M, Samaddar JS, et al. Downregulation of retinoblastoma protein is involved in the enhanced cytotoxicity of 4-hydroxytamoxifen plus mifepristone combination therapy versus antiestrogen monotherapy of human breast cancer. Int J Oncol, 2007, 31(3): 643-655
15. 连利娟. 林巧稚妇科肿瘤学. 第4版. 北京: 人民卫生出版社, 2006: 70
16. Seo J, Park JS, Nam JH, et al. Association of CD94/NKG2A, CD94/NKG2C, and its ligand HLA-E polymorphisms with Behcet's disease. Tissue Antigens, 2007, 70(4): 307-313
17. Ulbrecht M, Maier S, Hofmeister V, et al. Truncated HLA-G isoforms are retained in the endoplasmic reticulum and insufficiently provide HLA-E ligands. Hum Immunol, 2004, 65(3): 200-208
1. 孙健, 吴志全, 薛琼. 天花粉蛋白抑制肝细胞癌肺转移的研究. 中华肝胆外科杂志, 2005, 11(4): 253-256
2. 王茂钦, 李莉, 王洁, 等. 天花粉蛋白致豚鼠全身过敏反应的实验研究. 安徽中医学院学报, 1999, 18: 47-49
3. 杨海文, 李咏梅, 罗仁. 重组免疫毒素对荷瘤小鼠抗肿瘤效果的实验研究. 南方医科大学学报, 2007, 10: 78-81
4. Yuan HD, Li MY, Shao XX, et al. Determination of the Binding Epitope for Anti-trichosanthin Monoclonal Antibody T(8)C(12). Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 1998, 30(2): 137-141
5. 刘颖，王丽莉，邓燕，等. 米非司酮体内增强顺铂对人卵巢癌细胞的抗癌作用. 第一军医大学学报，2003，23（3）：242-244
6. 王刚，郑闻亭，肖平，等. 米非司酮对耐药卵巢癌细胞增殖、凋亡及其对顺铂敏感性的影响. 癌症，2004，23（4）：406-411
7. Rocereto TF, Saul HM, Aikins JA Jr. et al. Phase II study of mifepristone (RU486) in refractory ovarian cancer. Gynecol Oncol, 2000, 77(3): 429-432
8. Blaschitz A, Juch H, Volz A, et al. Soluble HLA-G, the discussion is going on! Mol Hum Reprod, 2005, 11(10): 723-727
9. Chang CC, Ferrone S. HLA-G in melanoma: can the current controversies be solved? Semin Cancer Biol. 2003, 13(5): 361-369
10. Seo J, Park JS, Nam JH, et al. Association of CD94/NKG2A, CD94/NKG2C, and its ligand HLA-E polymorphisms with Behcet's disease. Tissue Antigens, 2007, 70(4): 307-313
11. Ulbrecht M, Maier S, Hofmeister V, et al. Truncated HLA-G isoforms are retained in the endoplasmic reticulum and insufficiently provide HLA-E ligands. Hum Immunol, 2004, 65(3): 200-208
1. Favier B, LeMaoult J, Rouas-Freiss N, et al. Research on HLA-G: an update.Tissue Antigens, 2007, 69(3): 207-211
2. Smyth MJ, Hayakawa Y, Takeda K, et al. New aspects of natural killer-cell surveillance and therapy of cancer. Nat Rev Cancer, 2002, 2(11): 856-861
3. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3593
4. Middleton D, Curran M, Maxwell L. Natural killer cells and their receptors. Transplant Immunol, 2002, 10: 147-164
5. Biaddoni R, Cantoni C, Pende D, et al. Human natural killer cell receptors and co-receptors. Immunol Rev, 2001, 181: 203-214
6. Alvarez-Arias DA, Campbell KS. Protein kinase C regulates expression and function of inhibitory killer cell Ig-like receptors in NK cells. J Immunol, 2007, 179(8): 5281-5290
7. Barber DF, Faure M, Long EO. LFA-1 contributes an early signal for NK cell cytotoxicity. J Immunol, 2004, 173(6): 3653-3659
8. Van der Merwe PA. Formation function of the immunological synapse. Curr Opin Immunol, 2002, 14(3): 293-298
9. Ferrone S, Whiteside TL. Tumor microenvironment and immune escape. Surg Oncol Clin N Am, 2007, 16(4):755-774
10. Rouas-Freiss N, Moreau P, Menier C, et al. HLA-G in cancer: a way to turn off the immune system. Semin Cancer Biol, 2003, 13(5): 325-336
11. Sullivan LC, Clements CS, Beddoe T, et al. The Heterodimeric Assembly of the CD94-NKG2 Receptor Family and Implications for Human Leukocyte Antigen-E Recognition. Immunity, 2007, 27(6): 900-911
12. Ebert EC. IL-15 converts human intestinal intraepithelial lymphocytes to CD94 producers of IFN-gamma and IL-10, the latter promoting Fas ligand-mediated cytotoxicity. Immunology, 2005, 115(1): 118-126
13. Zhang C, Tian ZG, Zhang J, The negative regulatory effect of IFN-gamma on cognitive function of human natural killer cells. Zhonghua Zhong Liu Za Zhi, 2004, 26(6): 324-327
14. Malmberg KJ, Levitsky V, Norell H, et al. IFN-gamma protects short-term ovarian carcinoma cell lines from CTL lysis via a CD94/NKG2A-dependent mechanism. J Clin Invest, 2002, 110(10): 1515-1523
15. Wilkinson GW, Tomasec P, Stanton RJ, et al. Modulation of natural killer cells by human cytomegalovirus. J Clin Virol, 2007, 7: 567-571
16. Paul P, Cabestre FA, Ibrahim EC, et al. Identification of HLA-G7 as a new splice variant of the HLA-G mRNAand expression of soluble HLA-G5,-G6,and -G7 transcripts in human transfected cells. Hum Immunol, 2000, 61(11): 1138-1149
17. Blaschitz A, Hutter H, Leitner V, et al. Reaction patterns of monoclonal antibodies to HLA-G in human tissues and on cell lines: a comparative study. Hum, Immunol, 2000, 61(11): 1074-1085
18. Riteau B, Rouas-Freiss N, Menier C, et al. HLA-G2, -G3, and -G4 isoforms expressed as nonmature cell surface glycoproteins inhibit NK and antigen-specific CTL cytolysis. J Immunol, 2001, 166(8): 5018-5026
19. Moscoso J, Serrano-Vela JI, Pacheco R, et al. HLA-G, -E and -F: allelism, function and evolution. Transpl Immunol, 2006, 17(1): 61-64
20. Contini P, Ghio M, Poggi A, et al. Soluble HLA-A,-B,-C and -G molecules induce apoptosis in T and NK CD8+ cells and inhibit cytotoxic T cell activity through CD8 ligation. Eur J Immunol. 2003, 33(1): 125-134
21. Dorling A, Monk NJ, Lechler RI. HLA-G inhibits the transendothelial migration of human NK cells. Eur J Immunol, 2000, 30(2): 586-593
22. Onno M, Pangault C, Le Friec G, et al. Modulation of HLA-G antigens expression by human cytomegalovirus: specific induction in activated macrophages harboring human cytomegalovirus infection. J Immunol, 2000, 164(12): 6426-6434
23. LeMaoult J, Krawice-Radanne I, Dausset J, et al. HLA-G1-expressing antigen-presenting cells induce immunosuppressive CD4+ T cells. Proc Natl Acad Sci U S A, 2004, 101(18): 7064-7069
24. Blaschitz A, Hutter H, Leitner V, et al. Reaction patterns of monoclonal antibodies to HLA-G in human tissues and on cell lines: a comparative study. Hum Immunol, 2000, 61(11): 1074-1085
25. Le Discorde M, Moreau P, Sabatier P, et al. Expression of HLA-G in human cornea, an immune-privileged tissue. Hum Immunol, 2003, 64(11): 1039-1044
26. Blaschitz A, Juch H, Volz A, et al. Soluble HLA-G, the discussion is going on! Mol Hum Reprod, 2005, 11(10): 723-727
27. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cells express HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
28. Noci I, Fuzzi B, Rizzo R, et al. Embryonic soluble HLA-G as a marker of developmental potential in embryos. Hum Reprod, 2005, 20(1): 138-146
29. Paul P, Rouas-Freiss N, Khalil-Daher I, et al. HLA-G expression in melanoma: a way for tumor cells to escape from immunosurveillance. Proc Natl Acad Sci U S A, 1998, 95(8): 4510-4515.
30. Paul P, Cabestré FA, Le Gal FA,et al. Heterogeneity of HLA-G gene transcription and protein expression in malignant melanoma biopsies. Cancer Res, 1999, 59(8): 1954-1960
31. Ibrahim el C, Aractingi S, Allory Y,et al. Analysis of HLA antigen expression in benign and malignant melanocytic lesions reveals that upregulation of HLA-G expression correlates with malignant transformation, high inflammatory infiltration and HLA-A1 genotype. Int J Cancer, 2004, 108(2): 243-250
32. Ibrahim el C, Allory Y, Commo F, et al. Altered pattern of major histocompatibility complex expression in renal carcinoma: tumor-specific expression of the nonclassical human leukocyte antigen-G molecule is restricted to clear cell carcinoma while up-regulation of other major histocompatibility complex antigens is primarily distributed in all subtypes of renal carcinoma. Am J Pathol, 2003, 162(2): 501-508
33. El-Chennawi FA, Auf FA, El-Diasty AM, et al. Expression of HLA-G in cancer bladder. Egypt J Immunol, 2005, 12(1): 57-64
34. Yie SM, Yang H, Ye SR, et al. Expression of human leucocyte antigen G (HLA-G) is associated with prognosis in non-small cell lung cancer. Lung Cancer, 2007, 58(2): 267-274
35. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cells express HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
36. Yie SM, Yang H, Ye SR, et al. Expression of HLA-G is associated with prognosis in esophageal squamous cell carcinoma. Am J Clin Pathol, 2007, 128(6): 1002-1009
37. Yie SM, Yang H, Ye SR, et al. Expression of human leukocyte antigen G (HLA-G) correlates with poor prognosis in gastric carcinoma. Ann Surg Oncol, 14(10): 2721-2729
38. Hansel DE, Rahman A, Wilentz RE, et al. HLA-G upregulation in pre-malignant and malignant lesions of the gastrointestinal tract. Int J Gastrointest Cancer, 2005, 35(1): 15-23
39. Leelawat K, Engprasert S, Pongchai-rerk U, et al. No expression of human leukocyte antigen G (HLA-G) in colorectal cancer cells. J Med Assoc Thai, 2004, 87(7): 816-818
40. Lefebvre S, Antoine M, Uzan S, et al. Specific activation of the non-classical class I histocompatibility HLA-G antigen and expression of the ILT2 inhibitory receptor in human breast cancer. J Pathol, 2002, 196(3): 266-274
41. Palmisano GL, Pistillo MP, Fardin P, et al. Analysis of HLA-G expression in breast cancer tissues. Hum Immunol, 2002, 63(11): 969-976
42. Yoon BS, Kim YT, Kim JW, et al. Expression of human leukocyte antigen-G and its correlation with interleukin-10 expression in cervical carcinoma. Int J Gynaecol Obstet, 2007, 98(1): 48-53
43. Barrier BF, Kendall BS, Sharpe-Timms KL, et al. Expression of human leukocyte antigen-G and its correlation with interleukin-10 expression in cervical carcinoma. Gynecol Oncol, 2006, 103(1): 25-30
44. Rezvany MR, Kazemi A, Hajifathali A, et al. Analysis of HLA-G gene expression in B-lymphocytes from chronic lymphocytic leukemia patients.Iran Biomed J, 2007, 11(2): 125-129
45. Morandi F, Levreri I, Bocca P, et al. Human neuroblastoma cells trigger an immunosuppressive program in monocytes by stimulating soluble HLA-G release. Cancer Res, 2007, 67(13): 6433-6441
46. Adithi M, Kandalam M, Ramkumar HL, et al. Retinoblastoma: expression of HLA-G. Ocul Immunol Inflamm, 2006, 14(4): 207-213
47. Chang CC, Ferrone S. HLA-G in melanoma: can the current controversies be solved? Semin Cancer Biol, 2003, 13(5): 361-369
48. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3596
49. Koller BH, Geraghty DE, Shimizu Y, et al. HLA-E. A novel HLA class I gene expressed in resting T lymphocytes. J Immunol, 1988, 141(3): 897-904
50. Grimsley C, Kawasaki A, Gassner C, et al. Definitive high resolution typing of HLA-E allelic polymorphisms: Identifying potential errors in existing allele data. Tissue Antigens, 2002, 60(3): 206-212
51. 赵丽丽, 曲迅, 梁路, 等. 妊娠子宫微环境中uNK细胞NKG2A和NKG2D及其配体表达的研究. 中国病理生理杂志, 2006, 22 (8): 1636-1639
52. Bai A, Broen J, Forman J. The pathway for processing leader-derived peptides that regulate the maturation and expression of Qa-1b. Immunity, 1998, 9(3): 413-421
53. Brooks CR, Elliott T, Parham P, et al. The inhibitory receptor NKG2A determines lysis of vaccinia virus-infected autologous targets by NK cells. J Immunol, 2006, 176(2): 1141-1147
54. 张彩, 田志刚, 魏海明, 等. 非经典HLA I类分子（HLA-G和HLA-E）在人肿瘤细胞系的表达及IFN-γ 的调节作用! 中国肿瘤临床, 2001， 28（4）: 247-250
55. Derré L, Corvaisier M, Charreau B, et al. Expression and release of HLA-E by melanoma cells and melanocytes: potential impact on the response of cytotoxic effector cells. J Immunol, 2006, 177(5): 3100-3107
56. Hirankarn N, Kimkong I, Mutirangura A. HLA-E polymorphism in patients with nasopharyngeal carcinoma. Tissue Antigens, 2004, 64(5): 588-592
57. Mittelbronn M, Simon P, L?ffler C, et al. Elevated HLA-E levels in human glioblastomas but not in grade I to III astrocytomas correlate with infiltrating CD8+ cells. J Neuroimmunol, 2007, 189(1-2): 50-58
58. Hodgkinson AD, Millward BA, Demaine AG. The HLA-E locus is associated with age at onset and susceptibility to type 1 diabetes mellitus. Hum Immunol, 2000, 61(3): 290-295
59. Matsunami K, Miyagawa S, Nakai R, et al. Protection against natural killer-mediated swine endothelial cell lysis by HLA-G and HLA-E. Transplant Proc, 2000, 32(5): 939-940
1. 姜国勇, 李思经, 翁曼丽, 等. 天花粉蛋白结构、核苷酸序列及基因表达研究进展. 山东农业科学, 1999, 2: 52-54
2. 姜国勇, 李思经, 翁曼丽, 等. 天花粉蛋白基因转化番茄的研究. 植物学报, 1999, 41: 334-336
3. 李建国.核糖体失活蛋白的研究进展. 分子植物育种, 2005, 3: 566-570
4. 鲁继斌, 于宏伟, 李亚明, 等. 抗人肺腺癌单克隆抗体-天花粉蛋白免疫毒素对荷人肺腺癌裸鼠模型的导向治疗. 中国医科大学学报, 1995, 24(3): 243-246
5. Yuan HD, Li MY, Shao XX, et al. Determination of the Binding Epitope for Anti-trichosanthin Monoclonal Antibody T(8)C(12). Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 1998, 30(2): 137-141
6. 杨海文, 李咏梅, 罗仁. 重组免疫毒素对荷瘤小鼠抗肿瘤效果的实验研究. 南方医科大学学报, 2007, 10(3): 54-57
7. 胡梅洁, 张曙, 章永平, 等. 天花粉蛋白和重组干扰素α-2b联合应用对消化道癌细胞的协同杀伤作用. 肿瘤, 1997, 17(6): 465-467
8. 孙健, 吴志全, 薛琼. 天花粉蛋白抑制肝细胞癌肺转移的研究. 中华肝胆外科杂志, 2005, 11(4): 253-256
9. 黄益玲, 黄利鸣, 石新兰, 等. 天花粉蛋白对宫颈癌HeLa细胞增殖和细胞凋亡的影响. 中国药理学通报, 2005, 21: 253-254
10. 王艳华, 黄利鸣. TSLC1基因甲基化与宫颈癌HeLa细胞凋亡相关性的研究. 中国现代医学杂志, 2007, 21(4): 46-49
11. 孙璟, 涂水平, 吴裕炘, 等. 天花粉蛋白诱导胃癌细胞MKN-45凋亡中P53、Bcl-2、C-myc蛋白表达变化. 上海医学, 2004, 24(5): 292-294
12. Xu L, Wang Y, Wang L, et al. TYchi, a novel chitinase with RNA N-glycosidase and anti-tumor activities. Front Biosci, 2008, 13: 3127-3135
13. 田维毅, 马春玲, 白惠卿. 天花粉及其组份对小鼠NK细胞杀伤活性的影响. 贵州中医, 2001, 25(11): 982-984
14. Li F, Mei Y, Wang Y, et al. Trichosanthin inhibits antigen-specific T cell expansion through nitric oxide-mediated apoptosis pathway. Cell Immunol, 2005, 234(1): 23-30
15. 郭峰, 钱保华, 花美仙, 等. 天花粉对卵巢癌患者红细胞天然免疫活性促进作用的实验研究. 中南药学, 2004, 2(6): 329-331
16. An Q, Wei S, Mu S, et al. Mapping the antigenic determinants and reducing the immunogenicity of trichosanthin by site-directed mutagenesis. J Biomed Sci, 2006, 13(5):,637-643
17. Zhou X, Yang N, Lu L, Ding Q, et al. Up-regulation of IL-10 expression in dendritic cells is involved in Trichosanthin-induced immunosuppression. Immunol Lett, 2007, 110(1): 74-81
18. Zhao S, Wang Y, Wei H. Trichosanthin induced Th2 polarization status. Cell Mol Immunol, 2006, 3(4): 297-301
19. Shaw PC, Lee KM, Wong KB. Recent advances in trichosanthin, a ribosome-inactivating protein with multiple pharmacological properties. Toxicon, 2005 45(6): 683-689
20. Ouyang DY, Chan H, Wang YY, et al. An inhibitor of c-Jun N-terminal kinases (CEP-11004) counteracts the anti-HIV-1 action of trichosanthin. Biochem Biophys Res Commun, 2006, 339(1): 25-29
21. 王茂钦, 李莉, 王洁, 等. 天花粉蛋白致豚鼠全身过敏反应的实验研究. 安徽中医学院学报, 1999, 18: 47-49
22. He XH, Shaw PC, Xu LH, et al. Site-directed polyethylene glycol modification of trichosanthin: effects on its biological activities, pharmacokinetics, and antigenicity. Life Sci, 1999, 64(14): 1163-1175
1. Swahn ML, Ugocsai G, Bygdeman M, et al. Effect of oral prostaglandin E2 on uterine contractility and outcome of treatment in women receiving RU 486 (mifepristone) for termination of early pregnancy. Hum Reprod, 1989, 4(1): 21-28
2. 山海直, 还藤克, 金山喜以. 抗孕激素制剂不同给药途径投药剂量投药时期对大鼠妊娠的影响. 不妊会志, 1992, 37(1): 118-124
3. WHO task force on post-ovulatory methods of fertility regulations. Comparison of two doses of mifepristone in combination with misoprostol for early medical abortion: a randomised trial. World Health Organisation Task Force on Post-ovulatory Methods of Fertility Regulation. BJOG, 2000, 107(4): 524-530.
4. 程利南. 米非司酮配伍米索前列醇终止10-16周妊娠的临床多中心随机比较研究. 中华妇产科杂志, 1999, 34: 268-271
5. Frydman R, Lelaidier C, Baton-Saint-Mleux C, et al. Labor induction in women at term with mifepristone (RU 486): a double-blind, randomized, placebo-controlled study. Obstet Gynecol, 1992, 80(6): 972-975
6. 王自能, 朱颖嫄, 卢凤昕. 米非司酮对人胎脑组织超微结构的影响. 实用医学杂志, 2000, 16: 274
7. 程利南, 童传良, 肖志琴. 不同剂量米非司酮用于急性避孕的临床多中心研究. 中华妇产科研究, 1999, 34: 335-338
8. WHO task force on post-ovulatory methods of fertility regulations. Use of mifepristone for treatment of ectopic pregnancy. Am Fam Physician， 2006， 73(10): 1703-1714
9. 杨幼林, 郑淑蓉, 张志文. 米非司酮对子宫肌瘤组织中表皮生长因子基因表达的影响. 中华妇产科杂志, 1998, 1: 56-58
10. Kettel LM, Murphy AA, Morales AJ, et al. Preliminary report on the treatment of endometriosis with low-dose mifepristone (RU 486). Am J Obstet Gynecol, 1998, 178(6): 1151-1156
11. Butts CL, Shukair SA, Duncan KM, et al. Progesterone inhibits mature rat dendritic cells in a receptor-mediated fashion.Int Immunol, 2007, 19(3): 287-296
12. El Etreby MF, Liang Y, et al. Antitumor activity of mifepristone in the human LNCaP, LNCaP-C4, and LNCaP-C4-2 prostate cancer models in nude mice. Prostate, 2000, 42(2): 99-106
13. Sridhar S, Ali AA, Liang Y, et al. Differential expression of members of the tumor necrosis factor alpha-related apoptosis-inducing ligand pathway in prostate cancer cells. Cancer Res, 2001, 61(19): 7179-7183
14. Li DQ, Wang ZB, Bai J, et al. Reversal of multidrug resistance in drug-resistant human gastric cancer cell line SGC7901/VCR by antiprogestin drug mifepristone. World J Gastroenterol, 2004, 10(12): 1722-1725
15. Kacinski BM, Flick MB, Sapi E. RU-486 can abolish glucocorticoid-induced increases in CSF-1 receptor expression in primary human breast carcinoma specimens. J Soc Gynecol Investig, 2001, 8(2): 114-116
16. Hyder SM, Chiappetta C, Stancel GM. Pharmacological and endogenous progestins induce vascular endothelial growth factor expression in human breast cancer cells. Int J Cancer, 2001, 92(4): 469-473
17. Bamberger AM, Bamberger CM, Gellersen B, et al. Modulation of AP-1 activity by the human progesterone receptor in endometrial adenocarcinoma cells. Proc Natl Acad Sci U S A, 1996, 93(12): 6169-6174
18. 张秋实, 李继俊, 林仲秋. 米非司酮对子宫内膜癌裸鼠移植瘤细胞凋亡调控的研究. 实用癌症杂志, 2004, 19(6): 582-584
19. 温泽清, 兰守敏, 赵兴波, 等. 口服大剂量米非司酮对子宫内膜癌细胞形态学和免疫组织化学的影响. 现代妇产科进展, 2001, 10(6): 422-424
20. Rose FV, Barnea ER. Response of human ovarian carcinoma cell lines to antiprogestin in mifepristone. Oncogene, 1996, 12(5): 999-1003
21. 孙丽君, 刘慧范, 郭海洲, 等. 米非司酮对人卵巢癌细胞系HO-8910的生长及细胞周期的影响. 中国肿瘤临床, 2000, 27(5): 366-369
22. Rocereto TF, Saul HM, Aikins JA Jr, et al. Phase II study of mifepristone (RU486) in refractory ovarian cancer. Gynecol Oncol, 2000, 77(3): 429-432
23. Clark K, Ji H, Feltovich H, Janowski J, et al. Mifepristone-induced cervical ripening: structural, biomechanical, and molecular events. Am J Obstet Gynecol, 2006, 194(5): 1391-1398
24. Kamradt MC, Mohideen N, Vaughan AT. RU486 increases radiosensitivity and restores apoptosis through modulation of HPV E6/E7 in dexamethasone-treated cervical carcinoma cells. Gynecol Oncol, 2000, 77(1): 177-182
25. Webster JI, Tonelli LH, Moayeri M, et al. Anthrax lethal factor represses glucocorticoid and progesterone receptor activity. Proc Natl Acad Sci U S A, 2003, 100(10): 5706-5711
26. Mark PJ, Waddell BJ. P-glycoprotein restricts access of cortisol and dexamethasone to the glucocorticoid receptor in placental BeWo cells. Endocrinology, 2006, 147(11): 5147-5152
27. Wang H, Lee EW, Zhou L, Leung PC, et al. Progesterone receptor (PR) isoforms PRA and PRB differentially regulate expression of the breast cancer resistance protein in human placental choriocarcinoma BeWo cells. Mol Pharmacol, 2008, 73(3): 845-854
28. Romieu G, Maudelonde T, Ulmann A, et al. The antiprogestin RU486 in advanced breast cancer: preliminary clinical trial. Bull Cancer, 1987, 74(4): 455-461
29. Liang Y, Besch-Williford C, Brekken RA, et al. Progestin-dependent progression of human breast tumor xenografts: a novel model for evaluating antitumor therapeutics. Cancer Res, 2007, 67(20): 9929-9936
30. Newfield RS, Spitz IM, Isacson C, et al. Long-term mifepristone (RU486) therapy resulting in massive benign endometrial hyperplasia. Clin Endocrinol (Oxf), 2001, 54(3): 399-404
2. El Chérif Ibrahim, Nadia Guerra, Marie-José Terrier Lacombe. Tumor-specific Up-Regulation of the Nonclassical Class I HLA-G Antigen Expression in Renal Carcinoma. Cancer Research , 2001, 9(15): 6838-6845
3. Hansel DE, Rahman A, Wilentz RE, et al. HLA-G upregulation in pre-malignant and malignant lesions of the gastrointestinal tract. Int J Gastrointest Cancer, 2005, 35(1): 15-23
4. El-Chennawi FA, Auf FA, El-Diasty AM, et al. Expression of HLA-G in cancer bladder. Egypt J Immunol, 2005, 12(1): 57-64
5. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cells express HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
6. Monique H. Hurks, Markus M, Valter,et al. Uveal Melanoma: No Expression of HLA-G. Investigative Ophthalmology and Visual Science, 2001, 42: 3081-3084
7. Salvadori S, Martinelli G, Zier K. Resection of solid tumors reverses T cell defects and restores protective immunity. J Immunol, 2000, 164(4): 2214-20
8. Carosella ED, Paul P, Moreau P, et al. HLA-G and HLA-E: fundamental and pathophysiological aspects. Immunol Today, 2000, 21(11): 532-534
9. Riteau B, Rouas-Freiss N, Menier C, et al. HLA-G2, -G3, and -G4 isoforms expressed as nonmature cell surface glycoproteins inhibit NK and antigen-specific CTL cytolysis. J Immunol, 2001, 166(8): 5018-5026
10. Paul P, Rouas-Freiss N, Khalil-Daher I, et al. HLA-G expression in melanoma: a way for tumor cells to escape from immunosurveillance. Proc Natl Acad Sci U S A, 1998, 95(8): 4510-4515
11. Paul P, Cabestré FA, Le Gal FA,et al. Heterogeneity of HLA-G gene transcription and protein expression in malignant melanoma biopsies. Cancer Res, 1999, 59(8): 1954-1960
12. Ibrahim el C, Aractingi S, Allory Y,et al. Analysis of HLA antigen expression in benign and malignant melanocytic lesions reveals that upregulation of HLA-G expression correlates with malignant transformation, high inflammatory infiltration and HLA-A1 genotype. Int J Cancer, 2004, 108(2): 243-250
13. Yie SM, Yang H, Ye SR, et al. Expression of HLA-G is associated with prognosis in esophageal squamous cell carcinoma. Am J Clin Pathol, 2007, 128(6):1002~1009
14. Yie SM, Yang H, Ye SR, et al. Expression of human leukocyte antigen G (HLA-G) correlates with poor prognosis in gastric carcinoma. Ann Surg Oncol, 2007, 14(10): 2721-2729
15. Barrier BF, Kendall BS, Sharpe-Timms KL, et al. Characterization of human leukocyte antigen-G (HLA-G) expression in endometrial adenocarcinoma. Gynecol Oncol, 2006, 103(1): 25-30
16. Bukur J, Seliger B. The role of HLA-G for protection of human renal cell-carcinoma cells from immune-mediated lysis: implications for immunotherapies. Semin Cancer Biol, 2003, 13(5): 353-359
17. Middleton D, Curran M, Maxwell L. Natural killer cells and their receptors. Transpl Immunol, 2002, 10(2-3): 147-64
18. Moscoso J, Serrano-Vela JI, Pacheco R, et al. HLA-G, -E and -F: allelism, function and evolution. Transpl Immunol, 2006, 17(1): 61-64
19. Moretta L, Romagnani C, Pietra G, et al. NK-CTLs, a novel HLA-E-restricted T-cell subset. Trends Immunol, 2003, 24(3): 136-143
20. 张彩, 田志刚, 魏海明, 等. 非经典HLA I类分子（HLA-G和HLA-E）在人肿瘤细胞系的表达及的调节作用. 中国肿瘤临床, 2001, 28(4): 247-253
21. Derré L, Corvaisier M, Charreau B, et al. Expression and release of HLA-E by melanoma cells and melanocytes: potential impact on the response of cytotoxic effector cells. J Immunol, 2006, 177(5): 3100-3107
22. Wischhusen J, Friese MA, Mittelbronn M, et al. HLA-E protects glioma cells from NKG2D-mediated immune responses in vitro: implications for immune escape in vivo.J Neuropathol Exp Neurol, 2005, 64(6): 523-528
1. Favier B, LeMaoult J, Rouas-Freiss N, et al. Research on HLA-G: an update.Tissue Antigens, 2007, 69(3): 207-211
2. Smyth MJ, Hayakawa Y, Takeda K, et al. New aspects of natural killer-cell surveillance and therapy of cancer. Nat Rev Cancer, 2002, 2(11): 856-861
3. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3593
4. Middleton D, Curran M, Maxwell L. Natural killer cells and their receptors. Transplant Immunol, 2002, 10: 147-164
5. Biaddoni R, Cantoni C, Pende D, et al. Human natural killer cell receptors and co-receptors. Immunol Rev, 2001, 181: 203-214
6. Moscoso J, Serrano-Vela JI, Pacheco R, et al. HLA-G, -E and -F: allelism, function and evolution. Transpl Immunol, 2006, 17(1): 61-64
7. Blaschitz A, Hutter H, Leitner V, et al. Reaction patterns of monoclonal antibodies to HLA-G in human tissues and on cell lines: a comparative study. Hum, Immunol, 2000, 61(11): 1074-1085
8. Riteau B, Rouas-Freiss N, Menier C, et al. HLA-G2, -G3, and -G4 isoforms expressed as nonmature cell surface glycoproteins inhibit NK and antigen-specific CTL cytolysis. J Immunol, 2001, 166(8): 5018-5026
9. Bukur J, Malenica B, Huber C, et al. Altered expression of nonclassical HLA class Ib antigens in human renal cell carcinoma and its association with impaired immune response. Hum Immunol, 2003, 64(11): 1081-1092
10. Chang CC, Murphy SP, Ferrone S. Differential in vivo and in vitro HLA-G expression in melanoma cells: potential mechanisms. Hum Immunol, 2003, 64(11): 1057-1063
11. Lefebvre S, Antoine M, Uzan S, et al. Specific activation of the non-classical class I histocompatibility HLA-G antigen and expression of the ILT2 inhibitory receptor in human breast cancer. J Pathol, 2002, 196(3): 266-274
12. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cellsexpress HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
13. Paul P, Cabestré FA, Le Gal FA, et al. Heterogeneity of HLA-G gene transcription and protein expression in malignant melanoma biopsies. Cancer Res, 1999, 59(8): 1954-1960
14. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3596
1. 天花粉蛋白对胃癌细胞株SGC-7901凋亡影响的体外研究. 中国医药导报, 2007, 16(2): 231-234
2. 王媛媛, 欧阳东云, 郑永唐. 天花粉蛋白体外抗人白血病和淋巴瘤细胞的作用机制. 中国试验血液学杂志, 2007, 15(4): 729-732
3. 周欣阳, 张天一, 施海燕, 等. 天花粉蛋白诱导H22肝癌细胞凋亡的研究. 南通医学院学报, 2003, 23(4): 371-374
4. 黄益玲, 黄利鸣, 石新兰, 等. 天花粉蛋白对宫颈癌HeLa细胞增殖和细胞凋亡的影响. 中国药理学通报, 2005, 21: 253-254
5. 孙璟, 涂水平, 吴裕炘, 等. 天花粉蛋白诱导胃癌细胞MKN-45凋亡中P53、Bcl-2、C-myc蛋白表达变化. 上海医学, 2004, 24(5): 292-294
6. 田维毅, 马春玲, 白惠卿. 天花粉及其组份对小鼠NK细胞杀伤活性的影响. 贵州中医, 2001, 25(11): 982-984
7. 邬伟秀, 郭峰. 天花粉对血液肿瘤患者红细胞免疫功能影响的试验研究. 上海免疫学杂志, 1995, 15(3): 156
8. Li A, Felix JC, Minoo P, et al. Effect of mifepristone on proliferation and apoptosis of Ishikawa endometrial adenocarcinoma cells. Fertil Steril, 2005, 84(1): 202-211
9. Poole AJ, Li Y, Kim Y, et al. Prevention of Brca1-mediated mammary tumorigenesis in mice by a progesterone antagonist. Science, 2006, 314(5804): 1467-1470
10. Goyeneche AA, Carón RW, Telleria CM. Mifepristone inhibits ovarian cancer cell growth in vitro and in vivo. Clin Cancer Res, 2007, 13(11): 3370-3379
11. Grunberg SM, Weiss MH, Russell CA, et al. Long-term administration of mifepristone (RU486): clinical tolerance during extended treatment of meningioma. Cancer Invest, 2006, 24(8): 727-733
12. Li DQ, Wang ZB, Bai J, et al. Effects of mifepristone on proliferation of human gastric adenocarcinoma cell line SGC-7901 in vitro. World J Gastroenterol, 2004, 10(18): 2628-2631
13. Labriola L, Salatino M, Proietti CJ, et al. Heregulin induces transcriptional activation of the progesterone receptor by a mechanism that requires functional ErbB-2 and mitogen-activated protein kinase activation in breast cancer cells. Mol Cell Biol, 2003, 23(3): 1095-1111
14. Sridhar S, Ali AA, Liang Y,et al. Differential expression of members of the tumor necrosis factor alpha-related apoptosis-inducing ligand pathway in prostate cancer cells. Cancer Res, 2001, 61(19): 7179-7183
15. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
16. Fukuda H, He PJ, Yokota K, et al. Progesterone-dependent and -independent expression of the multidrug resistance type I gene in porcine granulosa cells. Mol Cell Biochem, 2007, 298(1-2): 179-186
17. Kacinski BM, Flick MB, Sapi E. RU-486 can abolish glucocorticoid-induced increases in CSF-1 receptor expression in primary human breast carcinoma specimens. J Soc Gynecol Investig, 2001, 8(2): 114-116
18. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
19. Hyder SM, Chiappetta C, Stancel GM. Pharmacological and endogenous progestins induce vascular endothelial growth factor expression in human breast cancer cells. Int J Cancer, 2001, 92(4): 469-473
20. Schoenlein PV, Hou M, Samaddar JS, et al. Downregulation of retinoblastoma protein is involved in the enhanced cytotoxicity of 4-hydroxytamoxifen plus mifepristone combination therapy versus antiestrogen monotherapy of human breast cancer. Int J Oncol, 2007, 31(3): 643-655
1 Malmberg KJ, Levitsky V, Norell H, et al. IFN-gamma protects short-term ovarian carcinoma cell lines from CTL lysis via a CD94/NKG2A-dependent mechanism. J Clin Invest, 2002, 110(10): 1515-1523
2. Yie SM, Li LH, Li GM, et al. Progesterone enhances HLA-G gene expression in JEG-3 choriocarcinoma cells and human cytotrophoblasts in vitro. Hum Reprod, 2006, 21(1): 46-51
3. 陈春玲 廖秦平. hCG 促进绒癌细胞系中HLA - G的表达. 中国妇产科临床杂志, 2004, 5(2): 113-115
4. 黄益玲, 黄利鸣, 石新兰, 等. 天花粉蛋白对宫颈癌HeLa细胞增殖和细胞凋亡的影响. 中国药理学通报, 2005, 21: 253-254
5. 孙璟, 涂水平, 吴裕炘, 等. 天花粉蛋白诱导胃癌细胞MKN-45凋亡中P53、Bcl-2、C-myc蛋白表达变化. 上海医学, 2004, 24(5): 292-294
6. 田维毅, 马春玲, 白惠卿. 天花粉及其组份对小鼠NK细胞杀伤活性的影响. 贵州中医, 2001, 25(11): 982-984
7. 邬伟秀, 郭峰. 天花粉对血液肿瘤患者红细胞免疫功能影响的试验研究. 上海免疫学杂志, 1995, 15(3): 156
8. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
9. Sridhar S, Ali AA, Liang Y,et al. Differential expression of members of the tumor necrosis factor alpha-related apoptosis-inducing ligand pathway in prostate cancer cells. Cancer Res, 2001, 61(19): 7179-7183
10. Fukuda H, He PJ, Yokota K, et al. Progesterone-dependent and -independent expression of the multidrug resistance type I gene in porcine granulosa cells. Mol Cell Biochem, 2007, 298(1-2): 179-186
11. Kacinski BM, Flick MB, Sapi E. RU-486 can abolish glucocorticoid-induced increases in CSF-1 receptor expression in primary human breast carcinoma specimens. J Soc Gynecol Investig, 2001, 8(2): 114-116
12. El Etreby MF, Liang Y, Lewis RW. Induction of apoptosis by mifepristone and tamoxifen in human LNCaP prostate cancer cells in culture. Prostate, 2000, 43(1): 31-42
13. Hyder SM, Chiappetta C, Stancel GM. Pharmacological and endogenous progestins induce vascular endothelial growth factor expression in human breast cancer cells. Int J Cancer, 2001, 92(4): 469-473
14. Schoenlein PV, Hou M, Samaddar JS, et al. Downregulation of retinoblastoma protein is involved in the enhanced cytotoxicity of 4-hydroxytamoxifen plus mifepristone combination therapy versus antiestrogen monotherapy of human breast cancer. Int J Oncol, 2007, 31(3): 643-655
15. 连利娟. 林巧稚妇科肿瘤学. 第4版. 北京: 人民卫生出版社, 2006: 70
16. Seo J, Park JS, Nam JH, et al. Association of CD94/NKG2A, CD94/NKG2C, and its ligand HLA-E polymorphisms with Behcet's disease. Tissue Antigens, 2007, 70(4): 307-313
17. Ulbrecht M, Maier S, Hofmeister V, et al. Truncated HLA-G isoforms are retained in the endoplasmic reticulum and insufficiently provide HLA-E ligands. Hum Immunol, 2004, 65(3): 200-208
1. 孙健, 吴志全, 薛琼. 天花粉蛋白抑制肝细胞癌肺转移的研究. 中华肝胆外科杂志, 2005, 11(4): 253-256
2. 王茂钦, 李莉, 王洁, 等. 天花粉蛋白致豚鼠全身过敏反应的实验研究. 安徽中医学院学报, 1999, 18: 47-49
3. 杨海文, 李咏梅, 罗仁. 重组免疫毒素对荷瘤小鼠抗肿瘤效果的实验研究. 南方医科大学学报, 2007, 10: 78-81
4. Yuan HD, Li MY, Shao XX, et al. Determination of the Binding Epitope for Anti-trichosanthin Monoclonal Antibody T(8)C(12). Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 1998, 30(2): 137-141
5. 刘颖，王丽莉，邓燕，等. 米非司酮体内增强顺铂对人卵巢癌细胞的抗癌作用. 第一军医大学学报，2003，23（3）：242-244
6. 王刚，郑闻亭，肖平，等. 米非司酮对耐药卵巢癌细胞增殖、凋亡及其对顺铂敏感性的影响. 癌症，2004，23（4）：406-411
7. Rocereto TF, Saul HM, Aikins JA Jr. et al. Phase II study of mifepristone (RU486) in refractory ovarian cancer. Gynecol Oncol, 2000, 77(3): 429-432
8. Blaschitz A, Juch H, Volz A, et al. Soluble HLA-G, the discussion is going on! Mol Hum Reprod, 2005, 11(10): 723-727
9. Chang CC, Ferrone S. HLA-G in melanoma: can the current controversies be solved? Semin Cancer Biol. 2003, 13(5): 361-369
10. Seo J, Park JS, Nam JH, et al. Association of CD94/NKG2A, CD94/NKG2C, and its ligand HLA-E polymorphisms with Behcet's disease. Tissue Antigens, 2007, 70(4): 307-313
11. Ulbrecht M, Maier S, Hofmeister V, et al. Truncated HLA-G isoforms are retained in the endoplasmic reticulum and insufficiently provide HLA-E ligands. Hum Immunol, 2004, 65(3): 200-208
1. Favier B, LeMaoult J, Rouas-Freiss N, et al. Research on HLA-G: an update.Tissue Antigens, 2007, 69(3): 207-211
2. Smyth MJ, Hayakawa Y, Takeda K, et al. New aspects of natural killer-cell surveillance and therapy of cancer. Nat Rev Cancer, 2002, 2(11): 856-861
3. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3593
4. Middleton D, Curran M, Maxwell L. Natural killer cells and their receptors. Transplant Immunol, 2002, 10: 147-164
5. Biaddoni R, Cantoni C, Pende D, et al. Human natural killer cell receptors and co-receptors. Immunol Rev, 2001, 181: 203-214
6. Alvarez-Arias DA, Campbell KS. Protein kinase C regulates expression and function of inhibitory killer cell Ig-like receptors in NK cells. J Immunol, 2007, 179(8): 5281-5290
7. Barber DF, Faure M, Long EO. LFA-1 contributes an early signal for NK cell cytotoxicity. J Immunol, 2004, 173(6): 3653-3659
8. Van der Merwe PA. Formation function of the immunological synapse. Curr Opin Immunol, 2002, 14(3): 293-298
9. Ferrone S, Whiteside TL. Tumor microenvironment and immune escape. Surg Oncol Clin N Am, 2007, 16(4):755-774
10. Rouas-Freiss N, Moreau P, Menier C, et al. HLA-G in cancer: a way to turn off the immune system. Semin Cancer Biol, 2003, 13(5): 325-336
11. Sullivan LC, Clements CS, Beddoe T, et al. The Heterodimeric Assembly of the CD94-NKG2 Receptor Family and Implications for Human Leukocyte Antigen-E Recognition. Immunity, 2007, 27(6): 900-911
12. Ebert EC. IL-15 converts human intestinal intraepithelial lymphocytes to CD94 producers of IFN-gamma and IL-10, the latter promoting Fas ligand-mediated cytotoxicity. Immunology, 2005, 115(1): 118-126
13. Zhang C, Tian ZG, Zhang J, The negative regulatory effect of IFN-gamma on cognitive function of human natural killer cells. Zhonghua Zhong Liu Za Zhi, 2004, 26(6): 324-327
14. Malmberg KJ, Levitsky V, Norell H, et al. IFN-gamma protects short-term ovarian carcinoma cell lines from CTL lysis via a CD94/NKG2A-dependent mechanism. J Clin Invest, 2002, 110(10): 1515-1523
15. Wilkinson GW, Tomasec P, Stanton RJ, et al. Modulation of natural killer cells by human cytomegalovirus. J Clin Virol, 2007, 7: 567-571
16. Paul P, Cabestre FA, Ibrahim EC, et al. Identification of HLA-G7 as a new splice variant of the HLA-G mRNAand expression of soluble HLA-G5,-G6,and -G7 transcripts in human transfected cells. Hum Immunol, 2000, 61(11): 1138-1149
17. Blaschitz A, Hutter H, Leitner V, et al. Reaction patterns of monoclonal antibodies to HLA-G in human tissues and on cell lines: a comparative study. Hum, Immunol, 2000, 61(11): 1074-1085
18. Riteau B, Rouas-Freiss N, Menier C, et al. HLA-G2, -G3, and -G4 isoforms expressed as nonmature cell surface glycoproteins inhibit NK and antigen-specific CTL cytolysis. J Immunol, 2001, 166(8): 5018-5026
19. Moscoso J, Serrano-Vela JI, Pacheco R, et al. HLA-G, -E and -F: allelism, function and evolution. Transpl Immunol, 2006, 17(1): 61-64
20. Contini P, Ghio M, Poggi A, et al. Soluble HLA-A,-B,-C and -G molecules induce apoptosis in T and NK CD8+ cells and inhibit cytotoxic T cell activity through CD8 ligation. Eur J Immunol. 2003, 33(1): 125-134
21. Dorling A, Monk NJ, Lechler RI. HLA-G inhibits the transendothelial migration of human NK cells. Eur J Immunol, 2000, 30(2): 586-593
22. Onno M, Pangault C, Le Friec G, et al. Modulation of HLA-G antigens expression by human cytomegalovirus: specific induction in activated macrophages harboring human cytomegalovirus infection. J Immunol, 2000, 164(12): 6426-6434
23. LeMaoult J, Krawice-Radanne I, Dausset J, et al. HLA-G1-expressing antigen-presenting cells induce immunosuppressive CD4+ T cells. Proc Natl Acad Sci U S A, 2004, 101(18): 7064-7069
24. Blaschitz A, Hutter H, Leitner V, et al. Reaction patterns of monoclonal antibodies to HLA-G in human tissues and on cell lines: a comparative study. Hum Immunol, 2000, 61(11): 1074-1085
25. Le Discorde M, Moreau P, Sabatier P, et al. Expression of HLA-G in human cornea, an immune-privileged tissue. Hum Immunol, 2003, 64(11): 1039-1044
26. Blaschitz A, Juch H, Volz A, et al. Soluble HLA-G, the discussion is going on! Mol Hum Reprod, 2005, 11(10): 723-727
27. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cells express HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
28. Noci I, Fuzzi B, Rizzo R, et al. Embryonic soluble HLA-G as a marker of developmental potential in embryos. Hum Reprod, 2005, 20(1): 138-146
29. Paul P, Rouas-Freiss N, Khalil-Daher I, et al. HLA-G expression in melanoma: a way for tumor cells to escape from immunosurveillance. Proc Natl Acad Sci U S A, 1998, 95(8): 4510-4515.
30. Paul P, Cabestré FA, Le Gal FA,et al. Heterogeneity of HLA-G gene transcription and protein expression in malignant melanoma biopsies. Cancer Res, 1999, 59(8): 1954-1960
31. Ibrahim el C, Aractingi S, Allory Y,et al. Analysis of HLA antigen expression in benign and malignant melanocytic lesions reveals that upregulation of HLA-G expression correlates with malignant transformation, high inflammatory infiltration and HLA-A1 genotype. Int J Cancer, 2004, 108(2): 243-250
32. Ibrahim el C, Allory Y, Commo F, et al. Altered pattern of major histocompatibility complex expression in renal carcinoma: tumor-specific expression of the nonclassical human leukocyte antigen-G molecule is restricted to clear cell carcinoma while up-regulation of other major histocompatibility complex antigens is primarily distributed in all subtypes of renal carcinoma. Am J Pathol, 2003, 162(2): 501-508
33. El-Chennawi FA, Auf FA, El-Diasty AM, et al. Expression of HLA-G in cancer bladder. Egypt J Immunol, 2005, 12(1): 57-64
34. Yie SM, Yang H, Ye SR, et al. Expression of human leucocyte antigen G (HLA-G) is associated with prognosis in non-small cell lung cancer. Lung Cancer, 2007, 58(2): 267-274
35. Pangault C, Le Friec G, Caulet-Maugendre S, et al. Lung macrophages and dendritic cells express HLA-G molecules in pulmonary diseases. Hum Immunol, 2002, 63(2): 83-90
36. Yie SM, Yang H, Ye SR, et al. Expression of HLA-G is associated with prognosis in esophageal squamous cell carcinoma. Am J Clin Pathol, 2007, 128(6): 1002-1009
37. Yie SM, Yang H, Ye SR, et al. Expression of human leukocyte antigen G (HLA-G) correlates with poor prognosis in gastric carcinoma. Ann Surg Oncol, 14(10): 2721-2729
38. Hansel DE, Rahman A, Wilentz RE, et al. HLA-G upregulation in pre-malignant and malignant lesions of the gastrointestinal tract. Int J Gastrointest Cancer, 2005, 35(1): 15-23
39. Leelawat K, Engprasert S, Pongchai-rerk U, et al. No expression of human leukocyte antigen G (HLA-G) in colorectal cancer cells. J Med Assoc Thai, 2004, 87(7): 816-818
40. Lefebvre S, Antoine M, Uzan S, et al. Specific activation of the non-classical class I histocompatibility HLA-G antigen and expression of the ILT2 inhibitory receptor in human breast cancer. J Pathol, 2002, 196(3): 266-274
41. Palmisano GL, Pistillo MP, Fardin P, et al. Analysis of HLA-G expression in breast cancer tissues. Hum Immunol, 2002, 63(11): 969-976
42. Yoon BS, Kim YT, Kim JW, et al. Expression of human leukocyte antigen-G and its correlation with interleukin-10 expression in cervical carcinoma. Int J Gynaecol Obstet, 2007, 98(1): 48-53
43. Barrier BF, Kendall BS, Sharpe-Timms KL, et al. Expression of human leukocyte antigen-G and its correlation with interleukin-10 expression in cervical carcinoma. Gynecol Oncol, 2006, 103(1): 25-30
44. Rezvany MR, Kazemi A, Hajifathali A, et al. Analysis of HLA-G gene expression in B-lymphocytes from chronic lymphocytic leukemia patients.Iran Biomed J, 2007, 11(2): 125-129
45. Morandi F, Levreri I, Bocca P, et al. Human neuroblastoma cells trigger an immunosuppressive program in monocytes by stimulating soluble HLA-G release. Cancer Res, 2007, 67(13): 6433-6441
46. Adithi M, Kandalam M, Ramkumar HL, et al. Retinoblastoma: expression of HLA-G. Ocul Immunol Inflamm, 2006, 14(4): 207-213
47. Chang CC, Ferrone S. HLA-G in melanoma: can the current controversies be solved? Semin Cancer Biol, 2003, 13(5): 361-369
48. Masilamani M, Nguyen C, Kabat J, et al. CD94/NKG2A inhibits NK cell activation by disrupting the actin network at the immunological synapse. J Immunol, 2006, 177(6): 3590-3596
49. Koller BH, Geraghty DE, Shimizu Y, et al. HLA-E. A novel HLA class I gene expressed in resting T lymphocytes. J Immunol, 1988, 141(3): 897-904
50. Grimsley C, Kawasaki A, Gassner C, et al. Definitive high resolution typing of HLA-E allelic polymorphisms: Identifying potential errors in existing allele data. Tissue Antigens, 2002, 60(3): 206-212
51. 赵丽丽, 曲迅, 梁路, 等. 妊娠子宫微环境中uNK细胞NKG2A和NKG2D及其配体表达的研究. 中国病理生理杂志, 2006, 22 (8): 1636-1639
52. Bai A, Broen J, Forman J. The pathway for processing leader-derived peptides that regulate the maturation and expression of Qa-1b. Immunity, 1998, 9(3): 413-421
53. Brooks CR, Elliott T, Parham P, et al. The inhibitory receptor NKG2A determines lysis of vaccinia virus-infected autologous targets by NK cells. J Immunol, 2006, 176(2): 1141-1147
54. 张彩, 田志刚, 魏海明, 等. 非经典HLA I类分子（HLA-G和HLA-E）在人肿瘤细胞系的表达及IFN-γ 的调节作用! 中国肿瘤临床, 2001， 28（4）: 247-250
55. Derré L, Corvaisier M, Charreau B, et al. Expression and release of HLA-E by melanoma cells and melanocytes: potential impact on the response of cytotoxic effector cells. J Immunol, 2006, 177(5): 3100-3107
56. Hirankarn N, Kimkong I, Mutirangura A. HLA-E polymorphism in patients with nasopharyngeal carcinoma. Tissue Antigens, 2004, 64(5): 588-592
57. Mittelbronn M, Simon P, L?ffler C, et al. Elevated HLA-E levels in human glioblastomas but not in grade I to III astrocytomas correlate with infiltrating CD8+ cells. J Neuroimmunol, 2007, 189(1-2): 50-58
58. Hodgkinson AD, Millward BA, Demaine AG. The HLA-E locus is associated with age at onset and susceptibility to type 1 diabetes mellitus. Hum Immunol, 2000, 61(3): 290-295
59. Matsunami K, Miyagawa S, Nakai R, et al. Protection against natural killer-mediated swine endothelial cell lysis by HLA-G and HLA-E. Transplant Proc, 2000, 32(5): 939-940
1. 姜国勇, 李思经, 翁曼丽, 等. 天花粉蛋白结构、核苷酸序列及基因表达研究进展. 山东农业科学, 1999, 2: 52-54
2. 姜国勇, 李思经, 翁曼丽, 等. 天花粉蛋白基因转化番茄的研究. 植物学报, 1999, 41: 334-336
3. 李建国.核糖体失活蛋白的研究进展. 分子植物育种, 2005, 3: 566-570
4. 鲁继斌, 于宏伟, 李亚明, 等. 抗人肺腺癌单克隆抗体-天花粉蛋白免疫毒素对荷人肺腺癌裸鼠模型的导向治疗. 中国医科大学学报, 1995, 24(3): 243-246
5. Yuan HD, Li MY, Shao XX, et al. Determination of the Binding Epitope for Anti-trichosanthin Monoclonal Antibody T(8)C(12). Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai), 1998, 30(2): 137-141
6. 杨海文, 李咏梅, 罗仁. 重组免疫毒素对荷瘤小鼠抗肿瘤效果的实验研究. 南方医科大学学报, 2007, 10(3): 54-57
7. 胡梅洁, 张曙, 章永平, 等. 天花粉蛋白和重组干扰素α-2b联合应用对消化道癌细胞的协同杀伤作用. 肿瘤, 1997, 17(6): 465-467
8. 孙健, 吴志全, 薛琼. 天花粉蛋白抑制肝细胞癌肺转移的研究. 中华肝胆外科杂志, 2005, 11(4): 253-256
9. 黄益玲, 黄利鸣, 石新兰, 等. 天花粉蛋白对宫颈癌HeLa细胞增殖和细胞凋亡的影响. 中国药理学通报, 2005, 21: 253-254
10. 王艳华, 黄利鸣. TSLC1基因甲基化与宫颈癌HeLa细胞凋亡相关性的研究. 中国现代医学杂志, 2007, 21(4): 46-49
11. 孙璟, 涂水平, 吴裕炘, 等. 天花粉蛋白诱导胃癌细胞MKN-45凋亡中P53、Bcl-2、C-myc蛋白表达变化. 上海医学, 2004, 24(5): 292-294
12. Xu L, Wang Y, Wang L, et al. TYchi, a novel chitinase with RNA N-glycosidase and anti-tumor activities. Front Biosci, 2008, 13: 3127-3135
13. 田维毅, 马春玲, 白惠卿. 天花粉及其组份对小鼠NK细胞杀伤活性的影响. 贵州中医, 2001, 25(11): 982-984
14. Li F, Mei Y, Wang Y, et al. Trichosanthin inhibits antigen-specific T cell expansion through nitric oxide-mediated apoptosis pathway. Cell Immunol, 2005, 234(1): 23-30
15. 郭峰, 钱保华, 花美仙, 等. 天花粉对卵巢癌患者红细胞天然免疫活性促进作用的实验研究. 中南药学, 2004, 2(6): 329-331
16. An Q, Wei S, Mu S, et al. Mapping the antigenic determinants and reducing the immunogenicity of trichosanthin by site-directed mutagenesis. J Biomed Sci, 2006, 13(5):,637-643
17. Zhou X, Yang N, Lu L, Ding Q, et al. Up-regulation of IL-10 expression in dendritic cells is involved in Trichosanthin-induced immunosuppression. Immunol Lett, 2007, 110(1): 74-81
18. Zhao S, Wang Y, Wei H. Trichosanthin induced Th2 polarization status. Cell Mol Immunol, 2006, 3(4): 297-301
19. Shaw PC, Lee KM, Wong KB. Recent advances in trichosanthin, a ribosome-inactivating protein with multiple pharmacological properties. Toxicon, 2005 45(6): 683-689
20. Ouyang DY, Chan H, Wang YY, et al. An inhibitor of c-Jun N-terminal kinases (CEP-11004) counteracts the anti-HIV-1 action of trichosanthin. Biochem Biophys Res Commun, 2006, 339(1): 25-29
21. 王茂钦, 李莉, 王洁, 等. 天花粉蛋白致豚鼠全身过敏反应的实验研究. 安徽中医学院学报, 1999, 18: 47-49
22. He XH, Shaw PC, Xu LH, et al. Site-directed polyethylene glycol modification of trichosanthin: effects on its biological activities, pharmacokinetics, and antigenicity. Life Sci, 1999, 64(14): 1163-1175
1. Swahn ML, Ugocsai G, Bygdeman M, et al. Effect of oral prostaglandin E2 on uterine contractility and outcome of treatment in women receiving RU 486 (mifepristone) for termination of early pregnancy. Hum Reprod, 1989, 4(1): 21-28
2. 山海直, 还藤克, 金山喜以. 抗孕激素制剂不同给药途径投药剂量投药时期对大鼠妊娠的影响. 不妊会志, 1992, 37(1): 118-124
3. WHO task force on post-ovulatory methods of fertility regulations. Comparison of two doses of mifepristone in combination with misoprostol for early medical abortion: a randomised trial. World Health Organisation Task Force on Post-ovulatory Methods of Fertility Regulation. BJOG, 2000, 107(4): 524-530.
4. 程利南. 米非司酮配伍米索前列醇终止10-16周妊娠的临床多中心随机比较研究. 中华妇产科杂志, 1999, 34: 268-271
5. Frydman R, Lelaidier C, Baton-Saint-Mleux C, et al. Labor induction in women at term with mifepristone (RU 486): a double-blind, randomized, placebo-controlled study. Obstet Gynecol, 1992, 80(6): 972-975
6. 王自能, 朱颖嫄, 卢凤昕. 米非司酮对人胎脑组织超微结构的影响. 实用医学杂志, 2000, 16: 274
7. 程利南, 童传良, 肖志琴. 不同剂量米非司酮用于急性避孕的临床多中心研究. 中华妇产科研究, 1999, 34: 335-338
8. WHO task force on post-ovulatory methods of fertility regulations. Use of mifepristone for treatment of ectopic pregnancy. Am Fam Physician， 2006， 73(10): 1703-1714
9. 杨幼林, 郑淑蓉, 张志文. 米非司酮对子宫肌瘤组织中表皮生长因子基因表达的影响. 中华妇产科杂志, 1998, 1: 56-58
10. Kettel LM, Murphy AA, Morales AJ, et al. Preliminary report on the treatment of endometriosis with low-dose mifepristone (RU 486). Am J Obstet Gynecol, 1998, 178(6): 1151-1156
11. Butts CL, Shukair SA, Duncan KM, et al. Progesterone inhibits mature rat dendritic cells in a receptor-mediated fashion.Int Immunol, 2007, 19(3): 287-296
12. El Etreby MF, Liang Y, et al. Antitumor activity of mifepristone in the human LNCaP, LNCaP-C4, and LNCaP-C4-2 prostate cancer models in nude mice. Prostate, 2000, 42(2): 99-106
13. Sridhar S, Ali AA, Liang Y, et al. Differential expression of members of the tumor necrosis factor alpha-related apoptosis-inducing ligand pathway in prostate cancer cells. Cancer Res, 2001, 61(19): 7179-7183
14. Li DQ, Wang ZB, Bai J, et al. Reversal of multidrug resistance in drug-resistant human gastric cancer cell line SGC7901/VCR by antiprogestin drug mifepristone. World J Gastroenterol, 2004, 10(12): 1722-1725
15. Kacinski BM, Flick MB, Sapi E. RU-486 can abolish glucocorticoid-induced increases in CSF-1 receptor expression in primary human breast carcinoma specimens. J Soc Gynecol Investig, 2001, 8(2): 114-116
16. Hyder SM, Chiappetta C, Stancel GM. Pharmacological and endogenous progestins induce vascular endothelial growth factor expression in human breast cancer cells. Int J Cancer, 2001, 92(4): 469-473
17. Bamberger AM, Bamberger CM, Gellersen B, et al. Modulation of AP-1 activity by the human progesterone receptor in endometrial adenocarcinoma cells. Proc Natl Acad Sci U S A, 1996, 93(12): 6169-6174
18. 张秋实, 李继俊, 林仲秋. 米非司酮对子宫内膜癌裸鼠移植瘤细胞凋亡调控的研究. 实用癌症杂志, 2004, 19(6): 582-584
19. 温泽清, 兰守敏, 赵兴波, 等. 口服大剂量米非司酮对子宫内膜癌细胞形态学和免疫组织化学的影响. 现代妇产科进展, 2001, 10(6): 422-424
20. Rose FV, Barnea ER. Response of human ovarian carcinoma cell lines to antiprogestin in mifepristone. Oncogene, 1996, 12(5): 999-1003
21. 孙丽君, 刘慧范, 郭海洲, 等. 米非司酮对人卵巢癌细胞系HO-8910的生长及细胞周期的影响. 中国肿瘤临床, 2000, 27(5): 366-369
22. Rocereto TF, Saul HM, Aikins JA Jr, et al. Phase II study of mifepristone (RU486) in refractory ovarian cancer. Gynecol Oncol, 2000, 77(3): 429-432
23. Clark K, Ji H, Feltovich H, Janowski J, et al. Mifepristone-induced cervical ripening: structural, biomechanical, and molecular events. Am J Obstet Gynecol, 2006, 194(5): 1391-1398
24. Kamradt MC, Mohideen N, Vaughan AT. RU486 increases radiosensitivity and restores apoptosis through modulation of HPV E6/E7 in dexamethasone-treated cervical carcinoma cells. Gynecol Oncol, 2000, 77(1): 177-182
25. Webster JI, Tonelli LH, Moayeri M, et al. Anthrax lethal factor represses glucocorticoid and progesterone receptor activity. Proc Natl Acad Sci U S A, 2003, 100(10): 5706-5711
26. Mark PJ, Waddell BJ. P-glycoprotein restricts access of cortisol and dexamethasone to the glucocorticoid receptor in placental BeWo cells. Endocrinology, 2006, 147(11): 5147-5152
27. Wang H, Lee EW, Zhou L, Leung PC, et al. Progesterone receptor (PR) isoforms PRA and PRB differentially regulate expression of the breast cancer resistance protein in human placental choriocarcinoma BeWo cells. Mol Pharmacol, 2008, 73(3): 845-854
28. Romieu G, Maudelonde T, Ulmann A, et al. The antiprogestin RU486 in advanced breast cancer: preliminary clinical trial. Bull Cancer, 1987, 74(4): 455-461
29. Liang Y, Besch-Williford C, Brekken RA, et al. Progestin-dependent progression of human breast tumor xenografts: a novel model for evaluating antitumor therapeutics. Cancer Res, 2007, 67(20): 9929-9936
30. Newfield RS, Spitz IM, Isacson C, et al. Long-term mifepristone (RU486) therapy resulting in massive benign endometrial hyperplasia. Clin Endocrinol (Oxf), 2001, 54(3): 399-404
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