不同葡萄糖、胰岛素水平对人肝细胞性肝癌生长作用研究
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摘要
随着社会的发展,人们已逐渐关注到糖尿病、恶性肿瘤已成为威胁人类生命健康的全球性问题。近年来,越来越多的流行病学和临床资料显示,糖尿病患者易患恶性肿瘤。有研究发现,2型糖尿病发生恶性肿瘤时以消化道肿瘤占第一位。原发性肝癌是世界上最常见且恶性程度最高的消化道肿瘤之一,其发病率居世界恶性肿瘤第五位,死亡率居第三位。我国是肝癌的高发区之一,每年死于肝癌的人数占全世界肝癌死亡人数的50%以上。由于肝癌的高发病率和死亡率,关于其病因学与发病机制的研究已成为国内外学者关注的焦点。目前已知,乙型肝炎病毒、丙型肝炎病毒感染、嗜酒等与肝细胞性肝癌发生发展关系密切,但仍有部分肝癌的病因尚不明确。2型糖尿病与肝细胞性肝癌的发生发展是否存在相关性?其分子机制是什么?仍不十分清楚,而且目前关于两者关系的研究多局限于流行病学调查和临床病例对照研究方面,具体的实验研究还甚少,国内外相关研究报道资料也较少。
依据2型糖尿病病理生理的改变,可将2型糖尿病的自然病程分为4个阶段,每个阶段均有葡萄糖、胰岛素水平的不同变化,血葡萄糖和胰岛素水平的变化贯穿于2型糖尿病的全过程。若2型糖尿病与肝细胞性肝癌有关,其自然病程中不同葡萄糖、胰岛素水平的变化是否对肝细胞肝癌的生物学特征产生影响?目前国内外相关研究报道资料较少,需要进一步对其进行研究。肝癌的发生和发展是一个多基因、多因素、多阶段的复杂演变过程,涉及到多种癌基因的激活、抑癌基因的失活以及由此引起的细胞生物学行为的改变等。环氧合酶-2(COX-2)是一种诱生型酶,在大多数正常组织和细胞中不表达,而多参与炎症、肿瘤等病理过程,在80%的肿瘤标本中都存在有COX-2的高表达,COX-2可通过促癌基因的激活和／或抑癌基因失活,促细胞增殖和／或抑制细胞凋亡,促肿瘤血管生成,促肿瘤细胞侵袭、转移,使机体产生免疫耐受致监视能力降低等多方面作用促肿瘤发生发展。血管内皮细胞生长因子(VEGF)是一种内皮细胞的特异性有丝分裂原,能够调节血管及淋巴管的生成,是目前已知作用最强、特异性最高的促血管生成因子。VEGF表达调控受多种因素的影响,且其异常表达与肿瘤及某些疾病的发生发展密切相关。肝癌是一种富血供实体肿瘤,生长快,易复发转移,其中血管生成环节在肿瘤的生长、侵袭、转移过程中起重要作用。所以本实验模拟2型糖尿病病程中不同葡萄糖、胰岛素浓度的变化环境,检测其对人肝癌SMMC-7721细胞体外生长的影响及对COX-2、VEGF基因表达的影响,初步探讨2型糖尿病与肝细胞性肝癌之间的关系,并为肝细胞性肝癌的临床防治工作提供新思路和理论依据。
实验方法
1.肝细胞性肝癌危险因素的病例对照研究
选取于我院2006年1月～2011年12月期间住院的1350例肝细胞性肝癌患者(病例组)的病例资料和同期住院的1350例非肿瘤患者(对照组)的病例资料,对相关数据进行搜集、整理,采用x2检验比较两组一般人口学特征的差异和2型糖尿病与肝细胞性肝癌发病的相关性,Logistic回归分析评估肝细胞性肝癌危险因素的比值比(odds ratio, OR)及95%可信区间(95%confidence interval,95%CI)。
2.实验用人肝癌SMMC-7721细胞培养浓度和时间的选择
通过倒置显微镜下观察并结合MTT方法评价细胞生长状态和数量,以此确定实验用人肝癌SMMC-7721细胞最佳接种浓度和合适培养时间。
3.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞增殖的影响
应用MTT方法检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)对人肝癌SMMC-7721细胞作用48小时后的影响,通过检测细胞在492nm波长下的吸光度(A492值),分析细胞增殖能力。
4.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞凋亡的影响
使用流式细胞仪检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)对人肝癌SMMC-7721细胞作用48小时后细胞凋亡率的变化,分析细胞凋亡情况。
5.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞COX-2、VEGFmRNA表达的影响
应用半定量RT-PCR法检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml))及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)条件下,各组人肝癌SMMC-7721细胞中COX-2、VEGFmRNA的表达情况。
6.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞COX-2、VEGF蛋白表达的影响
应用VVestern Blot方法检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml))及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)条件下,各组人肝癌SMMC-7721细胞中COX-2、VEGF蛋白的表达情况。
统计学分析
采用SPSS16.0统计软件对所得数据进行统计学分析。计量资料数据以均数±标准差(x±s)表示,二分类变量采用x2检验,危险因素采用Logistic回归分析,多组间均数的比较采用方差分析。以a=0.05作为检验性水准。
结果
1.嗜酒、乙型肝炎病毒感染、丙型肝炎病毒感染、2型糖尿病和肝癌家族史为肝细胞性肝癌的发病危险因素(P<0.05),其中2型糖尿病可增加肝细胞性肝癌发病风险的比值比OR=1.881,95%CI:1.314-2.671;而嗜烟、肥胖与肝细胞性肝癌的发病无关(P>0.05)。
2.为避免实验过程中人肝癌SMMC-7721细胞本身生长因素对实验结果的影响,我们摸索了实验用细胞接种的最佳浓度为2×105个/ml,细胞培养的合适时间为48小时。
3.随着葡萄糖浓度(5mmol/L,10mmol/L,20mmol/L)和胰岛素浓度(l0μIU/ml,50μIU/ml,100μIU/ml)的提高,人肝癌SMMC-7721细胞增殖能力逐渐增强,其中100μIU/ml葡萄糖组的增殖能力最强(0.6564±0.054)；联合葡萄糖+胰岛素并未显示出对增殖能力的协同作用,其中20+100组细胞增殖能力较其他联合组(5+10,5+100,20+10)强(P<0.05)。
4.随着葡萄糖(5mmol/L,10mmol/L,20mmol/L)和胰岛素(10μIU/ml,50μIU/ml100IU/ml)浓度的提高,人肝癌SMMC-7721凋亡细胞的数量逐渐减少,其中以胰岛素100μIU/ml组的细胞凋亡率最低(7.54±0.9%)；联合组(5+10,5+100,20+100,20+10)细胞凋亡率并未显示协同作用,其中20+100组细胞凋亡率较其他联合组(5+10,5+100,20+10)低(P<0.05)。
5.随着葡萄糖(5mmol/L,10mmol/L,20mmol/L)浓度的升高,人肝癌SMMC-7721细胞中VEGF mRNA表达明显增高,其中20mmol/L组细胞中VEGF mRNA相对表达量为2.784±0.07,远高于其他组(P<0.05),而COX-2mRNA表达增高与VEGFmRNA增高并不一致,其中10mmol/L组COX-2mRNA表达最高,为0.57±0.05,高于其他组(P<0.05)。不同的是随着胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)水平的升高,人肝癌SMMC-7721细胞中VEGF和COX-2mRNA表达量均明显增高,100μIU/ml组细胞VEGF和COX-2mRNA相对表达量分别为1.71±0.03和1.744±0.05,明显高于其他组P<0.05)。同时,联合组(5+10,5+100,20+100,20+10)中,20+100组细胞中VEGF mRNA相对表达量较其他组明显增高(P<0.05),但COX-2mRNA表达无明显变化。
6.随着葡萄糖(5mmol/L,10mmol/L,20mmol/L)浓度的升高,VEGF蛋白表达明显增高,其中20mmol/L组细胞VEGF蛋白相对表达量为2.53±±0.11,远高于其他组(P<0.05),而COX-2蛋白表达增高与VEGF蛋白增高并不一致,其中10mmol/L组COX-2蛋白表达最高,为0.74±0.08,高于其他组(P<0.05)。随着胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)浓度的升高,VEGF和COX-2蛋白表达量均明显增高,l00μIU/ml组细胞VEGF和COX-2蛋白相对表达量分别为2.05±0.14和0.92±0.15,明显高于其他组(P<0.05)。同时,联合组(5+10,5+100,20+100,20+10)中,20+100组细胞VEGF蛋白相对表达量较其他组明显增高,但COX-2表达无明显变化。
结论
1.2型糖尿病可增加肝细胞性肝癌的发病风险,其是肝细胞性肝癌发病的独立危险因素之一；2型糖尿病与肝细胞性肝癌的发病相关,且与嗜酒、乙肝和丙肝病毒感染之间存在协同作用。
2.高浓度葡萄糖和胰岛素可以增加人肝癌SMMC-7721细胞的增殖能力和减少细胞凋亡,其中胰岛素的作用最为明显,且两者无明显协同作用。
3.高浓度葡萄糖可以提高人肝癌SMMC-7721细胞中VEGF基因的表达,但不能提高COX-2基因表达；高浓度胰岛素不仅可以提高VEGF基因的表达,还可以提高COX-2基因表达；葡萄糖与胰岛素联合对两基因的表达无明显协同作用。
With the development of society, diabetes mellitus(DM) and malignant disease have been the global issueses that threaten human life and health. In recent years, more and more epidemic and clinical dataes have shown that diabetic patients were liable to malignant disease. Some researches have found that digestive cancer was at the most particular risk when the malignant disease was simultaneous with DM. Hepatocullular carcinoma(HCC) was one of the most popular digestive cancer in the world with a high-grade malignancy, and incidence and mortality rate of HCC were respectively fifth and third high-grade of all these cancer. There were the most HCC patients in China and about50%patients died of HCC were in China, so HCC have been making a threat to peoples'health and life, and also be focused by reseachers at home and abroad. At present, the infection of hepatitis B virus,hepatitis c virus and alcohol have close relations with HCC, but the pathogenesis of some kinds of HCC are still undefined. What is the relationship of type2diabetes mellitus and HCC? What is the molecular mechanism? We need to explore. At the same time, the documents of relative research were infrequent and limited to epidemiological investigation and case control study.
According to the physiopathologic change of type2DM, we divide course of this disease into four stages with the different change of glucose and insulin. The change of levels of blood glucose and insulin run through the overall process of type2DM. The relationship between HCC and type2DM implied that the change of levels of blood glucose and insulin might influence the biological characteristics of HCC. As these relative researches are relatively rare, further and deeper studies have been in desired. The generation and development of HCC underwent multi-gene, multi-factor and muti-stage process, which involved activation of oncogenes and inactivation of tumor suppressor genes, and induced to change of biological behaviour. Cyclooxygenase-2(COX-2) frequently participate in the pathological process of inflammation and tumor, and was expressed in80%turner samples, but did not express in normal tissues and cells. COX-2could promote the activation of oncogenes and inactivation of tumor suppressor genes, followed by cell proliferation and/or cell apoptosis, tumor angiogenesis, invasiveness and metastasis of cancer cells, immune tolerance,and so on. So to speak that COX-2could promote the generation and development of cancer. Vascular endothelial cell growth factor(VEGF) was a kind of specific mitogen in endothelial cells. VEGF could regulate the generation ofblood and lymphatic vessel with the most effective function in angiogenic factor. The expression of VEGF was regulated by many elements and had a close relation with cancer and other disease. HCC was a kind of solid tumor with abundant blood supply, and the generation of blood vessel played an important role in the tumor angiogenesis, invasiveness and metastasis of cancer cells. In this study, we imitated the change of different levels of glucose and insulin in type2DM, detected the growth influence of HCC cell line SMMC-7721, and the expression regulation of COX-2and VEGF genes. The results of this study could preliminary expound the relationship between HCC and type2DM, and provide new thinking and theory of clinial study for HCC.
Methods
1. Case control study of HCC with risk factors
1350cases of HCC patients and non-cancer patients (control) from the first affiliated hospital of Zhengzhou university were selected, and the relative data was then collect and classified. The difference of demography characteristic and the correlation of type2DM with HCC were evaluated by using chi-square test, and the odds ratio(OR) of risk factors and95%confidence interval(95%CI) for HCC were evaluated by Logistic regression analysis.
2. Optimizations of concentration and time for HCC SMMC-7721cell culture
Optimizations of concentration and time for HCC SMMC-7721cell culture were confirmed by observation from inverted microscope and using MTT assay to evaluate cell quantity and condition.
3. Proliferation of HCC SMMC-7721cells treated with different levels of glucose and insulin
HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L, lOmmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours, and MTT assay was used to detect the proliferation capacity.
4. Apoptosis of HCC SMMC-7721cells treated with different levels of glucose and insulin
HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L,10mmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours, and flow cytometry (FCM) was used to assay the apoptosis rate and analysis the apoptosis condition of cells.
5. Expression of COX-2and VEGF mRNA
Reverse transcript PCR(RT-PCR) was applied to detect the expression of COX-2and VEGF mRNA in HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L, lOmmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours.
6. Expression of COX-2and VEGF protein
Westernblot was used to evaluated the expression of COX-2and VEGF mRNA in HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L,10mmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours.
Statistical Analysis
Statistical analysis was performed with chi-square test, Logistic regression analysis and ANOVA analysis of variance. Statistical significance was set at α=0.05.
Results
1. Results showed that infection of hepatitis B virus and hepatitis C virus, alcoholomania, type2DM and family history of HCC were morbidity risk factors of HCC(P<0.05), and type2DM could increase the onset risk of HCC(odds ratio(OR)=1.881,95%CI:1.314～2.671), while smoking and fat were independent to HCC(P>0.05)
2. Optimizations of concentration and time for HCC SMMC-7721cell culture was proceeded to avoid the influence by the cells'growth factors, results showed that the optimum concentration was2×105cells/ml, and cells were appropriately cultured for48h.
3. In this part, we found that the proliferation capacity of SMMC-7721cells was enhanced gradually following by the concentration increasing of glucose(5mmol/L,10mmol/L,20mmol/L) and insulin(10μIU/ml,50μIU/ml,100μlU/ml). The proliferation of cells in100μIU/ml insulin group was the highest of all groups, but the combination groups did not showed the synergistic effect to proliferation capacity. The group(20+100) cells had a better proliferation compared with other combination groups (P<0.05)
4.The apoptosis cells of SMMC-7721decreased gradually following by the concentration increasing of glucose(5mmol/L,10mmol/L,20mmol/L) and insulin(10μIU/ml,50μIU/ml,100μIU/ml). The apoptosis rate of cells in100μIU/ml insulin group was the lowest of all groups, but the combination groups did not showed the synergistic effect to cells apoptosis.
5. With the increasing of glucose concentration(5mmol/L, lOmmol/L,20mmol/L), expression of VEGF mRNA in SMMC-7721cells was increased obviously:the relative VEGF mRNA expression of HCC cells in20mmol/L glucose group was2.78±0.07, which was higher than other groups (P<0.05), while the relative COX-2mRNA expression change did not keep track of VEGF mRNA, that is the highest expression of COX-2was in10mmol/L group(0.57±0.05)(P<0.05). Unlike to glucose group, With the increasing of insulin concentration (10μIU/ml,50μIU/ml,100μIU/ml), expression of VEGF and COX-2in SMMC-7721cells was increased obviously, relatively expression of VEGF and COX-2mRNA were respectively1.71±0.03and1.74±0.05, which were higher than other groups (P<0.05) Meanwhile, relatively expression of VEGF of cells in the combination groups was increased obvious (P<0.05), but COX-2did not changed evidently.
6. With the increasing of glucose concentration(5mmol/L,10mmol/L,20mmol/L), expression of VEGF protein in SMMC-7721cells was increased obviously:the relative VEGF protein expression of HCC cells in20mmol/L glucose group was2.53±0.11, which was higher than other groups (P<0.05), while the relative COX-2mRNA expression change did not keep track of VEGF protein, that is the highest expression of COX-2was in10mmol/L group(0.74±0.08)(P<0.05). With the increasing of insulin concentration (10μIU/ml,50μIU/ml,100μIU/ml), expression of VEGF and COX-2protein in SMMC-7721cells was increased obviously, relatively expression of VEGF and COX-2protein were respectively2.05±0.14and0.92±0.15, which were higher than other groups (P<0.05). Meanwhile, relatively expression of VEGF protein of cells in the combination groups was increased obvious (P<0.05), but COX-2did not changed evidently.
Conclusions
1. As a risk factor, type2DM could increase the onset risk of HCC. The positive correlation was type2DM and HCC. Type2DM was a independent risk factor and exerted synergistic effects with other risk factors such as HBV/HCV, alcoholic consumption on the development of HCC。
2. High concentration of glucose and insulin could increase proliferation ability and decrease the apoptosis of HCC cells, and the effect of insulin was stronger than glucose, but combination of glucose and insulin did not show any synergistic effect.
3. High concentration of glucose could increase expression of VEGF, but had no effect to COX-2expression; while high concentration of insulin could increase both VEGF and COX-2gene expression, but anyhow combination of glucose and insulin had little synergistic effect to these two genes expression.
依据2型糖尿病病理生理的改变,可将2型糖尿病的自然病程分为4个阶段,每个阶段均有葡萄糖、胰岛素水平的不同变化,血葡萄糖和胰岛素水平的变化贯穿于2型糖尿病的全过程。若2型糖尿病与肝细胞性肝癌有关,其自然病程中不同葡萄糖、胰岛素水平的变化是否对肝细胞肝癌的生物学特征产生影响?目前国内外相关研究报道资料较少,需要进一步对其进行研究。肝癌的发生和发展是一个多基因、多因素、多阶段的复杂演变过程,涉及到多种癌基因的激活、抑癌基因的失活以及由此引起的细胞生物学行为的改变等。环氧合酶-2(COX-2)是一种诱生型酶,在大多数正常组织和细胞中不表达,而多参与炎症、肿瘤等病理过程,在80%的肿瘤标本中都存在有COX-2的高表达,COX-2可通过促癌基因的激活和／或抑癌基因失活,促细胞增殖和／或抑制细胞凋亡,促肿瘤血管生成,促肿瘤细胞侵袭、转移,使机体产生免疫耐受致监视能力降低等多方面作用促肿瘤发生发展。血管内皮细胞生长因子(VEGF)是一种内皮细胞的特异性有丝分裂原,能够调节血管及淋巴管的生成,是目前已知作用最强、特异性最高的促血管生成因子。VEGF表达调控受多种因素的影响,且其异常表达与肿瘤及某些疾病的发生发展密切相关。肝癌是一种富血供实体肿瘤,生长快,易复发转移,其中血管生成环节在肿瘤的生长、侵袭、转移过程中起重要作用。所以本实验模拟2型糖尿病病程中不同葡萄糖、胰岛素浓度的变化环境,检测其对人肝癌SMMC-7721细胞体外生长的影响及对COX-2、VEGF基因表达的影响,初步探讨2型糖尿病与肝细胞性肝癌之间的关系,并为肝细胞性肝癌的临床防治工作提供新思路和理论依据。
实验方法
1.肝细胞性肝癌危险因素的病例对照研究
选取于我院2006年1月～2011年12月期间住院的1350例肝细胞性肝癌患者(病例组)的病例资料和同期住院的1350例非肿瘤患者(对照组)的病例资料,对相关数据进行搜集、整理,采用x2检验比较两组一般人口学特征的差异和2型糖尿病与肝细胞性肝癌发病的相关性,Logistic回归分析评估肝细胞性肝癌危险因素的比值比(odds ratio, OR)及95%可信区间(95%confidence interval,95%CI)。
2.实验用人肝癌SMMC-7721细胞培养浓度和时间的选择
通过倒置显微镜下观察并结合MTT方法评价细胞生长状态和数量,以此确定实验用人肝癌SMMC-7721细胞最佳接种浓度和合适培养时间。
3.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞增殖的影响
应用MTT方法检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)对人肝癌SMMC-7721细胞作用48小时后的影响,通过检测细胞在492nm波长下的吸光度(A492值),分析细胞增殖能力。
4.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞凋亡的影响
使用流式细胞仪检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)对人肝癌SMMC-7721细胞作用48小时后细胞凋亡率的变化,分析细胞凋亡情况。
5.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞COX-2、VEGFmRNA表达的影响
应用半定量RT-PCR法检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml))及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)条件下,各组人肝癌SMMC-7721细胞中COX-2、VEGFmRNA的表达情况。
6.不同浓度葡萄糖、胰岛素对人肝癌SMMC-7721细胞COX-2、VEGF蛋白表达的影响
应用VVestern Blot方法检测不同浓度葡萄糖(5mmol/L,10mmol/L,20mmol/L)、胰岛素(10μIU/ml,50μIU/ml,100μIU/ml))及葡萄糖+胰岛素(5+10,5+100,20+100,20+10)条件下,各组人肝癌SMMC-7721细胞中COX-2、VEGF蛋白的表达情况。
统计学分析
采用SPSS16.0统计软件对所得数据进行统计学分析。计量资料数据以均数±标准差(x±s)表示,二分类变量采用x2检验,危险因素采用Logistic回归分析,多组间均数的比较采用方差分析。以a=0.05作为检验性水准。
结果
1.嗜酒、乙型肝炎病毒感染、丙型肝炎病毒感染、2型糖尿病和肝癌家族史为肝细胞性肝癌的发病危险因素(P<0.05),其中2型糖尿病可增加肝细胞性肝癌发病风险的比值比OR=1.881,95%CI:1.314-2.671;而嗜烟、肥胖与肝细胞性肝癌的发病无关(P>0.05)。
2.为避免实验过程中人肝癌SMMC-7721细胞本身生长因素对实验结果的影响,我们摸索了实验用细胞接种的最佳浓度为2×105个/ml,细胞培养的合适时间为48小时。
3.随着葡萄糖浓度(5mmol/L,10mmol/L,20mmol/L)和胰岛素浓度(l0μIU/ml,50μIU/ml,100μIU/ml)的提高,人肝癌SMMC-7721细胞增殖能力逐渐增强,其中100μIU/ml葡萄糖组的增殖能力最强(0.6564±0.054)；联合葡萄糖+胰岛素并未显示出对增殖能力的协同作用,其中20+100组细胞增殖能力较其他联合组(5+10,5+100,20+10)强(P<0.05)。
4.随着葡萄糖(5mmol/L,10mmol/L,20mmol/L)和胰岛素(10μIU/ml,50μIU/ml100IU/ml)浓度的提高,人肝癌SMMC-7721凋亡细胞的数量逐渐减少,其中以胰岛素100μIU/ml组的细胞凋亡率最低(7.54±0.9%)；联合组(5+10,5+100,20+100,20+10)细胞凋亡率并未显示协同作用,其中20+100组细胞凋亡率较其他联合组(5+10,5+100,20+10)低(P<0.05)。
5.随着葡萄糖(5mmol/L,10mmol/L,20mmol/L)浓度的升高,人肝癌SMMC-7721细胞中VEGF mRNA表达明显增高,其中20mmol/L组细胞中VEGF mRNA相对表达量为2.784±0.07,远高于其他组(P<0.05),而COX-2mRNA表达增高与VEGFmRNA增高并不一致,其中10mmol/L组COX-2mRNA表达最高,为0.57±0.05,高于其他组(P<0.05)。不同的是随着胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)水平的升高,人肝癌SMMC-7721细胞中VEGF和COX-2mRNA表达量均明显增高,100μIU/ml组细胞VEGF和COX-2mRNA相对表达量分别为1.71±0.03和1.744±0.05,明显高于其他组P<0.05)。同时,联合组(5+10,5+100,20+100,20+10)中,20+100组细胞中VEGF mRNA相对表达量较其他组明显增高(P<0.05),但COX-2mRNA表达无明显变化。
6.随着葡萄糖(5mmol/L,10mmol/L,20mmol/L)浓度的升高,VEGF蛋白表达明显增高,其中20mmol/L组细胞VEGF蛋白相对表达量为2.53±±0.11,远高于其他组(P<0.05),而COX-2蛋白表达增高与VEGF蛋白增高并不一致,其中10mmol/L组COX-2蛋白表达最高,为0.74±0.08,高于其他组(P<0.05)。随着胰岛素(10μIU/ml,50μIU/ml,100μIU/ml)浓度的升高,VEGF和COX-2蛋白表达量均明显增高,l00μIU/ml组细胞VEGF和COX-2蛋白相对表达量分别为2.05±0.14和0.92±0.15,明显高于其他组(P<0.05)。同时,联合组(5+10,5+100,20+100,20+10)中,20+100组细胞VEGF蛋白相对表达量较其他组明显增高,但COX-2表达无明显变化。
结论
1.2型糖尿病可增加肝细胞性肝癌的发病风险,其是肝细胞性肝癌发病的独立危险因素之一；2型糖尿病与肝细胞性肝癌的发病相关,且与嗜酒、乙肝和丙肝病毒感染之间存在协同作用。
2.高浓度葡萄糖和胰岛素可以增加人肝癌SMMC-7721细胞的增殖能力和减少细胞凋亡,其中胰岛素的作用最为明显,且两者无明显协同作用。
3.高浓度葡萄糖可以提高人肝癌SMMC-7721细胞中VEGF基因的表达,但不能提高COX-2基因表达；高浓度胰岛素不仅可以提高VEGF基因的表达,还可以提高COX-2基因表达；葡萄糖与胰岛素联合对两基因的表达无明显协同作用。
With the development of society, diabetes mellitus(DM) and malignant disease have been the global issueses that threaten human life and health. In recent years, more and more epidemic and clinical dataes have shown that diabetic patients were liable to malignant disease. Some researches have found that digestive cancer was at the most particular risk when the malignant disease was simultaneous with DM. Hepatocullular carcinoma(HCC) was one of the most popular digestive cancer in the world with a high-grade malignancy, and incidence and mortality rate of HCC were respectively fifth and third high-grade of all these cancer. There were the most HCC patients in China and about50%patients died of HCC were in China, so HCC have been making a threat to peoples'health and life, and also be focused by reseachers at home and abroad. At present, the infection of hepatitis B virus,hepatitis c virus and alcohol have close relations with HCC, but the pathogenesis of some kinds of HCC are still undefined. What is the relationship of type2diabetes mellitus and HCC? What is the molecular mechanism? We need to explore. At the same time, the documents of relative research were infrequent and limited to epidemiological investigation and case control study.
According to the physiopathologic change of type2DM, we divide course of this disease into four stages with the different change of glucose and insulin. The change of levels of blood glucose and insulin run through the overall process of type2DM. The relationship between HCC and type2DM implied that the change of levels of blood glucose and insulin might influence the biological characteristics of HCC. As these relative researches are relatively rare, further and deeper studies have been in desired. The generation and development of HCC underwent multi-gene, multi-factor and muti-stage process, which involved activation of oncogenes and inactivation of tumor suppressor genes, and induced to change of biological behaviour. Cyclooxygenase-2(COX-2) frequently participate in the pathological process of inflammation and tumor, and was expressed in80%turner samples, but did not express in normal tissues and cells. COX-2could promote the activation of oncogenes and inactivation of tumor suppressor genes, followed by cell proliferation and/or cell apoptosis, tumor angiogenesis, invasiveness and metastasis of cancer cells, immune tolerance,and so on. So to speak that COX-2could promote the generation and development of cancer. Vascular endothelial cell growth factor(VEGF) was a kind of specific mitogen in endothelial cells. VEGF could regulate the generation ofblood and lymphatic vessel with the most effective function in angiogenic factor. The expression of VEGF was regulated by many elements and had a close relation with cancer and other disease. HCC was a kind of solid tumor with abundant blood supply, and the generation of blood vessel played an important role in the tumor angiogenesis, invasiveness and metastasis of cancer cells. In this study, we imitated the change of different levels of glucose and insulin in type2DM, detected the growth influence of HCC cell line SMMC-7721, and the expression regulation of COX-2and VEGF genes. The results of this study could preliminary expound the relationship between HCC and type2DM, and provide new thinking and theory of clinial study for HCC.
Methods
1. Case control study of HCC with risk factors
1350cases of HCC patients and non-cancer patients (control) from the first affiliated hospital of Zhengzhou university were selected, and the relative data was then collect and classified. The difference of demography characteristic and the correlation of type2DM with HCC were evaluated by using chi-square test, and the odds ratio(OR) of risk factors and95%confidence interval(95%CI) for HCC were evaluated by Logistic regression analysis.
2. Optimizations of concentration and time for HCC SMMC-7721cell culture
Optimizations of concentration and time for HCC SMMC-7721cell culture were confirmed by observation from inverted microscope and using MTT assay to evaluate cell quantity and condition.
3. Proliferation of HCC SMMC-7721cells treated with different levels of glucose and insulin
HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L, lOmmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours, and MTT assay was used to detect the proliferation capacity.
4. Apoptosis of HCC SMMC-7721cells treated with different levels of glucose and insulin
HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L,10mmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours, and flow cytometry (FCM) was used to assay the apoptosis rate and analysis the apoptosis condition of cells.
5. Expression of COX-2and VEGF mRNA
Reverse transcript PCR(RT-PCR) was applied to detect the expression of COX-2and VEGF mRNA in HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L, lOmmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours.
6. Expression of COX-2and VEGF protein
Westernblot was used to evaluated the expression of COX-2and VEGF mRNA in HCC SMMC-7721cells were treated with different levels of glucose(5mmol/L,10mmol/L,20mmol/L), insulin(10μIU/ml,50μIU/ml,100μIU/ml) and combination of glucose and insulin (5+10,5+100,20+100,20+10) for48hours.
Statistical Analysis
Statistical analysis was performed with chi-square test, Logistic regression analysis and ANOVA analysis of variance. Statistical significance was set at α=0.05.
Results
1. Results showed that infection of hepatitis B virus and hepatitis C virus, alcoholomania, type2DM and family history of HCC were morbidity risk factors of HCC(P<0.05), and type2DM could increase the onset risk of HCC(odds ratio(OR)=1.881,95%CI:1.314～2.671), while smoking and fat were independent to HCC(P>0.05)
2. Optimizations of concentration and time for HCC SMMC-7721cell culture was proceeded to avoid the influence by the cells'growth factors, results showed that the optimum concentration was2×105cells/ml, and cells were appropriately cultured for48h.
3. In this part, we found that the proliferation capacity of SMMC-7721cells was enhanced gradually following by the concentration increasing of glucose(5mmol/L,10mmol/L,20mmol/L) and insulin(10μIU/ml,50μIU/ml,100μlU/ml). The proliferation of cells in100μIU/ml insulin group was the highest of all groups, but the combination groups did not showed the synergistic effect to proliferation capacity. The group(20+100) cells had a better proliferation compared with other combination groups (P<0.05)
4.The apoptosis cells of SMMC-7721decreased gradually following by the concentration increasing of glucose(5mmol/L,10mmol/L,20mmol/L) and insulin(10μIU/ml,50μIU/ml,100μIU/ml). The apoptosis rate of cells in100μIU/ml insulin group was the lowest of all groups, but the combination groups did not showed the synergistic effect to cells apoptosis.
5. With the increasing of glucose concentration(5mmol/L, lOmmol/L,20mmol/L), expression of VEGF mRNA in SMMC-7721cells was increased obviously:the relative VEGF mRNA expression of HCC cells in20mmol/L glucose group was2.78±0.07, which was higher than other groups (P<0.05), while the relative COX-2mRNA expression change did not keep track of VEGF mRNA, that is the highest expression of COX-2was in10mmol/L group(0.57±0.05)(P<0.05). Unlike to glucose group, With the increasing of insulin concentration (10μIU/ml,50μIU/ml,100μIU/ml), expression of VEGF and COX-2in SMMC-7721cells was increased obviously, relatively expression of VEGF and COX-2mRNA were respectively1.71±0.03and1.74±0.05, which were higher than other groups (P<0.05) Meanwhile, relatively expression of VEGF of cells in the combination groups was increased obvious (P<0.05), but COX-2did not changed evidently.
6. With the increasing of glucose concentration(5mmol/L,10mmol/L,20mmol/L), expression of VEGF protein in SMMC-7721cells was increased obviously:the relative VEGF protein expression of HCC cells in20mmol/L glucose group was2.53±0.11, which was higher than other groups (P<0.05), while the relative COX-2mRNA expression change did not keep track of VEGF protein, that is the highest expression of COX-2was in10mmol/L group(0.74±0.08)(P<0.05). With the increasing of insulin concentration (10μIU/ml,50μIU/ml,100μIU/ml), expression of VEGF and COX-2protein in SMMC-7721cells was increased obviously, relatively expression of VEGF and COX-2protein were respectively2.05±0.14and0.92±0.15, which were higher than other groups (P<0.05). Meanwhile, relatively expression of VEGF protein of cells in the combination groups was increased obvious (P<0.05), but COX-2did not changed evidently.
Conclusions
1. As a risk factor, type2DM could increase the onset risk of HCC. The positive correlation was type2DM and HCC. Type2DM was a independent risk factor and exerted synergistic effects with other risk factors such as HBV/HCV, alcoholic consumption on the development of HCC。
2. High concentration of glucose and insulin could increase proliferation ability and decrease the apoptosis of HCC cells, and the effect of insulin was stronger than glucose, but combination of glucose and insulin did not show any synergistic effect.
3. High concentration of glucose could increase expression of VEGF, but had no effect to COX-2expression; while high concentration of insulin could increase both VEGF and COX-2gene expression, but anyhow combination of glucose and insulin had little synergistic effect to these two genes expression.
引文
1. Parkin D M. Global cancer statistics in the year 2000[J]. Lancet Oncol,2001, 2(9):533-543.
2.赵金艳,高春芳.代谢综合征与肝癌流行病学研究进展[J].第二军医大学学报,2010,31(4)：425-428.
3.王爱萍,刘超,庞春梅等.2型糖尿病与恶性肿瘤临床相关性研究[J].南京医科大学学报,2007,27(1)：55-58.
4. Davila JA, Morgan RO, Shaib Y, et al. Diabetes increases the risk of hepatocellular carcinoma in the United States:a population based case control study[J]. Gut,2005,54(3):533-539.
5.邵清,陈国凤,李梵.慢性丙型肝炎病毒感染者2型糖尿病流行病学调查[J].肝脏,2007,12(5)：366-367.
6. Serag H, Tran T, Everhart JE. Diabetes increases the risk of chronic liver disease and hepatocellular carcinoma[J]. Gastroenterology,2004,126(2):460-468.
7. Staattin P, Ove Bjor, Ferrari P, et al. Prospective study of Hyperglycemia and cancer risk[J]. Diabetes Care,2007,30(3):561-567.
8. Serag H, Richardson P, Everhart J. The role of diabetes in hepatocellular carcinoma:a case-control study among United States Veterans[J]. Am J Gastroenterol,2001,96(8):2462-2467.
9.季尚玮,王江滨.2型糖尿病对原发性肝癌发病的危险性分析[J]。肝脏,2007,12(3)：168-170.
10.王艳丽.糖尿病诱发原发性肝癌的危险性评价及与其他相关危险因素协同致病作用分析[D].吉林大学,2005(04).
11. Wang F, Gupta S, Holly FA. Diabetes mellitus and pancreatic cancer in a population-based case-control study in the San Franxisco Bay Area, Califomia[J]. Cancer Epidemiol Biomarkers Prev,2006,15(8):1458-1463.
12. Chun Gao, Shu-Kun Yao. Diabetes mellitus:a "true" independent risk factor for hepatocellular carcinoma?[J]. Hepatobiliary Pancreat Dis Int.2009,8(2):465-473.
13.向红丁,张化冰.ADA：糖尿病医学诊治标准-2010(摘要)[J].中国糖尿病 杂志,2010,18(3)：164-171.
14.陈小箐,周凯.原发性肝癌患病的糖尿病相关因素研究[J]. Acta Academiae Medicine Jiangxi,2009,49(5):80-82.
15.刘成,马翔,马依彤等.新疆地区汉族、维吾尔族、哈萨克族成年人群超重和肥胖流行病学调查[J].中华流行病学杂志,2011,31(10)：1139-1143.
16.杨秉辉,张博恒.重视肝癌的病因和预防[J].临床肝胆病杂志,2012,28(4)：253-255.
17. Wilson EB, Maher HC. Cancer and tuberculosis with some comments on cancer and other diseases[J]. Am J Cancer,1932,16(1):227-250.
18. Lawson DH, Gray JM, Mckillop C, et al. Diabetes mellitus and primary hepatocellular carcinoma[J]. QJ Med,1986,61(2):945-955.
19. Hemminki K, Li X, Sundquist J, et al. Risk of cancer following hospitalization for type 2 diabetes[J]. Oncologist,2010,15(6):548-555.
20. Noto H, Osameic K, Sasazuki T, et al. Substantially increased risk of cancer in patients with diabetes mellitus:a systematic review and meta-analysis of epidemiologic evidence in Japan[J]. J Diabetes Complications,2010, 24(5):345-353.
21. Adami HO, Chow WH, Nyren O. Excess risk of primary liver cancer in patients with diabetes mellitus[J]. Natl Cancer Inst.1996,88(2):1472-1477.
22. El Serag HB, Richardson PA, Everhart JE, et al. Diabetes increases the risk of chronic liver disease and hepatocellular carcinoma[J].2004,126(7):460-468.
23. Davila JA, Morgan RO, Shaib Y, et al. Diabetes increases the risk of hepatocellular carcinoma in the United States:a population based case control study[J]. Gut,2005,54(3):533-539.
24. Giovannucci E, Harlan DM, Archer MC, et al. Diabetes and cancer:a conaensus report[J]. Diabetes Care,2010,33(7):1674-1685.
25.周慧敏.不同浓度的葡萄糖和胰岛素对Colon肿瘤细胞分泌免疫抑制物的影响及其分子机制的研究[D].河北医科大学,2009(4).
26.赵晔.S-腺苷蛋氨酸对胃癌增殖和C-myc、uPA基因甲基化的影响[D].郑州大 学,2010(5).
27.于志清,潘海林,黎英荣等.中国亚热带地区糖尿病流行与分型研究[J].广西医科大学学报,1999,16(6)：728-731.
28. Chang CK, Ulrich CM. Hyperinsulinaemia and Hyperglycaemia:Possible Risk Factors of Colorectal Cancer Among Diabetic Patients[J]. Diabetologia,2003, 46(5):595-607.
29. Jiao L, Weinstein ST, Albanes D, et al. Evidence the serum levels of the soluble receptor for advanced glycation end products are inversely associated wiyh pancreatic cancer risk:a prospective study[J]. Cancer Res,2011,71(10): 3582-3589.
30. Krone CA, Ely JT. Controlling hyperglycemia as an adjunct to cancer therapy[J]. Integrative Cancer Therapies,2005,4(1):25-31.
31.苏苹,郭庆畲.糖尿病、降糖治疗和癌症[J].中国老年保健医学杂志,2010,8(5)：5-7.
32. Currie CJ, Poole CD, Gale EA. The influence of glucose-lowering therapies on cancer risk in type 2 diabetes[J]. Diabetologia,2009,52(9):1766-1777.
33. Sandulache VC, Ow TJ, Pickering CR, et al. Glucose, not glutamine, is the dominant energy source required for proliferation and survival of head and neck squamous carcinoma cells[J]. Cancer,2011,117(13):2926-2938.
34.孙丽.糖尿病与恶性肿瘤相关性的作用机制研究进展[J].中国药师,2009,12(12)：1730-1732.
35.田爱平.胰岛素信号途径—肿瘤治疗的新靶点?[J].癌症进展杂志,2009,7(1)：66-72.
36. Reinmuth N, Fan F, LiuW, et al. Impact of insulin-like growth factor receptor function angio genesis, growth,and metastasis of colon cancer[J]. Lab Invest, 2002,82(10):1377-1389.
37. Guan Z, Buckman SY, Baier LD, et al. IGF-I and insulin amplify IL-1 beta-induced nitric oxide and prostaglandin biosynthesis[J]. Am J Physiol.1998, 274(4Pt2):F673-679.
38. Gupta K, Krishnsswaray G, Karnad A, et al. Insulin:a novel factor in carcinogenesis[J]. Am J Med Sci,2002,323(3):140-145.
39. Healy LA, Ruan AM, Carroll P, et al. Metabolic syndrome, central obesity and insulin resistance are associated with adverse pathological features in postmenopausal breast cancer[J]. Clin Oncol,2010,22(4):281-288.
40. Goodwin PJ, Ennis M, Bahl M, et al.High insulin levels in newly diagnosed breast cancer patients reflect underlying insulin resistance and are associated with components of the insulin resistance syndrome[J]. Breast Cancer Res Treat,2008, 4(25):235-243.
41.田春桃,焦顺昌,魏秀芳.胰岛素对人乳腺癌细胞影响的体外实验研究[J].现代肿瘤医学,2006,14(11)：1338-1340.
42.赵敬,薛凤霞,华绍芳,等.胰岛素对子宫内膜癌细胞增殖和凋亡的影响[J].中华妇产科杂志,2007,42(10)：696-700.
43.龚勇,赵秋,杨芳,等.胰腺导管腺癌组织中WT1、IGF-IR表达与细胞凋亡的关系[J].世界华人消化杂志,2006,14(28)：2810-2814.
44. Kurtaran A, Li SR, Raderer M, et al. Technetium-99m-galactosyl-neogolycoalbumin combined with iodine-123-Tyr-(A14)-insulin visualizes human hepatocellular carcinomas. J Nucl Med,1995,36(10):1875-1881.
45. Mosmann T. Rapid colorimetric assay of cellular growth and survival:application to proliferation and cytotoxiciyt assays[J]. J Immunol Methods,1983, 65(1):55-63.
46. Hino N, Higashi T, Nouso K, et al. Apoptosis and proliferation of human hepatocellular carcinoma[J]. Liver,1996,16(4):123-129.
47.潘銮凤.分子生物学技术[M].上海：复旦大学出版,2008,158-160.
48. Uefuji K, Ichikura T, Mochizuki H, et al. Expression of cyclooxygenase in human gastric adenomas and adenocarcinomas[J]. J Surg Oncol,2001,76(1):26-30.
49.张琛,夏海滨.血管内皮细胞生长因子及其表达调控研究进展[J].现代肿瘤学杂志,2012,20(4)：841-845.
50.陈建国.中国肝癌发病趋势和一级预防.临床肝胆病杂志[J].2012,28(4)：256-260.
51. Joo YE, Oh WT, Rew JS, et al. Cyclooxygenase-2 expression is associated with well-differentiated and intestinal-type pathways in gastric carcinogenesis[J]. Digestion,2002,66(4):222-229.
52. Nashida T, Yabe Y, Fu HY, et al. Geranylgeranylacetone induces cyclooxygenase-2 expression in cultured rat gastric epithelial cells through NF-kappaB[J]. Dig Dis Sci,2007,52(8):1890-1896.
53. Chang CK, Ulrich CM. Hyperinsulinaemia and Hyperglycaemia:Possible Risk Factors of Colorectal Cancer Among Diabetic Patients[J]. Diabetologia,2003, 46(5):595-607.
54.赵锋,曾春,周伟雄,等.初发2型糖尿病患者血清COX-2活性与IL-18含量改变的意[J].热带医学杂志,2006,6(10)：108-1083.
55. Persaud SJ, Muller D, Belin VD, et al. The role of arachidonic acid and its metabolites in insulin secretion from human islets of langerhans[J]. Diabetes, 2007,56(1):197-203.
56. Persaud SJ, Bums CJ, Belin VD, et al. Glucose-induced regulation of COX-2 expression in human islets of langerhans[J]. Diabetes,2004,53(Suppl 1):S190-S192.
57. Yuan L, Ziegle RR, Haman NA. Metformin modulates insulin post-receptor signaling transduction in chronically insulin-treated HepG2 cells [J]. Acta Pharmacal Sin,2003,24(1):55-60.
58. Joan Fortuny, Yolanda B, Ramon Bosch, et al. Type 2 diabetes mellitus, its treatment and risk for lymphoma[J]. European Journal of cancer,2005, 41(12):1782-1787.
59.阮丹杰,王伟.炎性因子与胰岛素抵抗的研究进展[J].医学综述,2008,15(4)：508-511.
60. Pollak M. Insulin, insulin-like growth factors and neoplasia[J]. Best Pract. Res.Clin. Endocrinol,2008,22(4):625-628.
61. Lee HC, Tian B, Sedivy JM. Loss of Rafkinase inhibitor protein promotes cell proliferation and migration of human hepatoma cells[J]. Gastroenterology,2006, 131(7):1208-1217.
62. Valter Donadon, Massimiliano Balbi, Giorgio Zanette, et al. Hyperinsulinemia and risk for hepatocellullar carcinoma in patients with chronic liver disease and type 2 diabetes mellitus[J]. Gastroenterol,2009,3(5):465-467.
63. Kunlin J, Yonjhua Z, Yuniuan S, et al. Vascular endothelial growth factor(VEGF) stimulates neorogenesis in vitor and in vivo. Proc Natl Acad Sei USA,2000, 97(18):1024-1027.
64. Ferrara N, Henzel WJ. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells[J]. Biochem Biophys Res Commun,1989,161(2):851-858.
65. Ho QT, Kuo CJ. Vascular endothelial growth factor:biology and therapeutic applications[J]. Int J Biochem Cell Biol,2007,39(78):1349-1357.
66. Maharaj AS, D'Amore PA. Role for VEGF in adult [J]. Microvasc Res, 2007,74(2-3):100-113.
67. Tung Ping Poon R, Fan ST, Wong J, et al. Risk factors prevention and managent of postoperative recurrence after resection of hepatocellular carcinoma[J]. Ann Surg,2000,232(8):10-24.
68.王越晖,王心蕊,朱向红等。高糖、高胰岛素对兔视网膜Muller细胞血管内皮生长因子表达的影响。中华糖尿病杂志,2004,12(4)：277-280.
69. Girnita L, Girnita A, Brodin B, et al. Increased Expression of Insulin-like Growth Factor I Receptor in Malignant Cells Expressing Aberrant p53 Functional Impact [J]. Cancer Research,2000,60(18):5278-5283.
70. Kim KS, Seu YB, Raek SH, et al. Induction of cellular senescence by insulin-like growth factor binding protein-5 through a p53-dependent mechanism[J]. Molecular Biology of the Cell,2007,18(11):4543-4552.
71.彭军,章军建,谢军等。高浓度葡萄糖诱导血管内皮细胞凋亡及对VEGF表达的影响。Stroke and Nervous Diseases,2007,14(4):238-241.
1. Wilson EB, Maher HC. Cancer and tuberculosis with some comments on cancer and other diseases[J]. Am J Cancer,1932,16(1):227-250.
2. Coughlin SS, Calle EE, Teras LR, et al. Diabetes mellitus as a predictor of cancer mortality in a large cohort of US adults[J]. Am J Epidemiol,2004, 159(12):1160-1167.
3. Giovannucci E, Harlan DM, Archer MC, et al. Diabetes and cancer:a conaensus report[J]. Diabetes Care,2010,33(7):1674-1685.
4.王爱萍,刘超,庞春梅,等.2型糖尿病与恶性肿瘤临床相关性研究[J].南京医科大学学报,2007,27(1)：55-58.
5. Hemminki K, Li X, Sundquist J, et al. Risk of cancer following hospitalization for type 2 diabetes[J]. Oncologist,2010,15(6):548-555.
6. Noto H, Osameic K, Sasazuki T, et al. Substantially increased risk of cancer in patients with diabetes mellitus:a systematic review and meta-analysis of epidemiologic evidence in Japan[J]. J Diabetes Complications,2010, 24(5):345-353.
7. Parkin D M. Global cancer statistics in the year 2000[J]. Lancet Oncol,2001, 2(9):533-543.
8.陈建国,宋新明.中国肝癌发病水平的估算及分析[J].中国肿瘤,2005,14(1)：28-31.
9.蒋俊俊,钟秋安,梁浩,等.糖尿病与原发性肝癌关系的病例对照研究Meta分析[J].中国慢性病预防与控制,2010,18(2)：132-135.
10. Staattin P, Ove Bjor, Ferrari P, et al. Prospective study of Hyperglycemia and cancer risk[J]. Diabetes Care,2007,30(3):561-567.
11. Polesel J, Zucchetto A, Montecca M, et al. The impact of obesity and diabetes mellitus on the risk of hepatocellular carcinoma[J]. Ann Oncol,2009, 20(2):353-359.
12. Bonovas S, Filioussi K, Tsantes A, et al. Diabetes mellitus and risk of prostate cancer:A meta-analysis[J]. Diabetologia,2004,47(6):1071-1078.
13. Michels KB, Solomon CG, Hu FB, et al. Type 2 diabetes and subsequent incidence of breast cancer in the Nurses'Health Study[J]. Diabetes Care,2003, 26(6):1752-1758.
14. Sellers TA, Jensen LE, Vierkant RA, et al. Assocation of diabetes with mammographic breast density and breast cancer in the Minnesota breast cancer family study[J]. Cancer Causes Control,2007,18(5):505-515.
15. Wu AH, Yu MC, Tseng CC, et al. Diabetes and risk of breast cancer in Asian-American women[J]. Carcinogenesis,2007,28(7):1561-1566.
16. Ewertz M, Holmberg L, Tretli S, et al. Risk factors for male breast cancer-a case-control study from Scandinavia[J]. Acta Oncol,2001,40(4):467-471.
17. Talamini R, Franceschi S, Favero A, et al. Selected medical conditions and risk of breast cancer[J]. Br J Cancer,1997,75 (11):1699-1703.
18. Wolf I, Sadetzki S, Gluck I, et al. Association between diabetes mellitus and adverse characteristics of breast cancer at presentation[J]. Eur J Cancer,2006, 42(8):1077-1082.
19. Zhang Y, Liu Z, Yu X, et al. The association between metabolic abnormality and endometrial cancer:a large case-control study in China[J]. Gynecol Oncol,2010, 117(1):41-46.
20.薛莹,刘超.2型糖尿病与恶性肿瘤[J].国际内分泌代谢杂志,2007,27(1)：56-58.
21.宋力.糖尿病作为子宫内膜癌危险因素的Meat分析[J].内蒙古医学杂志,2009,41(11)：1291-1295.
22. Fisher WE, Boros LG, Schirmer WJ. Reversal of enhanced pancreatic cancer growth in diabetes by insulin[J]. Surgery,1995,118(2):453-457.
23. Shibata A, Mack TM, Paganini-Hill A, et al. A prospective study of pancreatic cancer in the elderly[J]. Int J Cancer,1994,58(1):46-49.
24. DeMeo MT. Pancreatic cancer and sugar diabetes[J]. Nutr Rev,2001, 59(4):112-115.
25. Silverman DT, Schiffman M, Everhart J, et al. Diabetes mellitus, other medical conditions and familial history of cancer as risk factors for pancreatic cancer[J]. Br J Cancer,1999,80(11):1830-1837.
26. Jee SH, Ohrr H, Sull JW, et al. Fasting serum glucose level and cancer risk in Korean men and women [J]. JAMA,2005,293(2):194-202.
27. Kuriki K, Hirose K, Tajima K. Diabetes and cancer risk for all and specific sites among Japanese men and women[J]. Eur J Cancer Prev,2007,16(1):83-89.
28. Wang F, Gupta S, Holly FA. Diabetes mellitus and pancreatic cancer in a population-based case-control study in the San Franxisco Bay Area, Califomia[J]. Cancer Epidemiol Biomarkers Prev,2006,15(8):1458-1463.
29. Kellerer M. Von Eye Corleta H, Muhlhofer A, et al. Insulin and Insulin-like growth factor-1 receptor tyrosine-kinase activities in human reanl carcinoma[J]. Int J Cancer,1995,62(5):501-507.
30. McKeown-Eyssen G. Epidemiology of colorectal cancer revisited:Are serum triglycerides and /or plasma glucose associated with risk [J]. Cancer Epidemiol Biomarkers Prev,1994,3(8):687-695.
31.胡水清,汤哲,张玫.糖尿病与结直肠癌发病及转移关系[J].中国肿瘤,2007,16(9)：689-698.
32. Larsson SC, Giovannucci E, Wolk A. Diabetes and colorectal cancer incidence in the cohort of Swedish men[J]. Diabetes Care,2005,28(7):1805-1807.
33. Seow A, Yuan JM, Koh WP, et al. Diabetea mellitus and risk of colorectal cancer in the Singapore Chinese Health Study[J]. J Natl Cancer Inst,2006, 98(2):135-138.
34. Yang YX, Hennessy S, Lewis JD, et al. Type 2 diabetes mellitus and the risk of colorectal cancer[J]. Clin Gastroenterol Hepatol,2005,3(6):587-594.
35. La Vecchia C, Negri E, Decarli A, et al. Diabetes mellitus and colorectal cancer risk [J]. Cancer Epidemiol Biomarkers Prev,1997,6(12):1007-1010.
36. Chung YW, Han DS, Park KH, et al. Insulin therapy and colorectal adenoma risk among patients with Type 2 diabetes mellitus:a case-control study in Korea [J]. Dis Colon Rectum,2008,51(5):593-597.
37. Yang YX, Hennessy S, Lewis JD.Insulin therapy and colorectal cancer risk among type 2 diabetes mellitus patients[J].Gastroenterology,2004,127(4):1044-1050.
38. Davila JA, Morgan RO, Shaib Y, et al. Diabetes increases the risk of hepatocellular carcinoma in the United States:a population based case control study[J]. Gut,2005,54(4):533-539.
39.季尚玮,王江滨.2型糖尿病对原发性肝癌发病的危险性分析[J]。肝脏,2007,12(3)：168-170.
40. Adami HO, Chow WH, Nyren O, et al. Excess risk of primary liver cancer in patients with diabetes mellitus[J]. J Natl Cancer Inst,1996,88(20):1472-1477.
41. Serag H, Tran T, Everhart JE. Diabetes increases the risk of chronic liver disease and hepatocellular carcinoma[J]. Gastroenterology,2004,126(2):460-468.
42. Serag H, Richardson P, Everhart J. The role of diabetes in hepatocellular carcinoma:a case-control study among United States Veterans[J]. Am J Gastroenterol,2001,96(8):2462-2467.
43.王艳丽.糖尿病诱发原发性肝癌的危险性评价及与其他相关危险因素协同致病作用分析[D].吉林大学,2005(04).
44. Kawamura Y, Ikeda K, Arase Y, et al. Diabetes mellitus worsens the recurrence rate after potentially curative therapy in patients with hepatocellular carcinoma associated with nonviral hepatitis [J]. J Gastroenterol Hepatol,2008,23 (11):1739-1746.
45. Lindblad P, Chow WH, Chan J, et al. The role of diabetes mellitus in the aetiology of reanl cell cancer[J]. Diabetologia,1999,42(1):107-112.
46. Inoue M, Iwasaki M, Otani T, et al. Diabetes mellitus and the risk of cancer result from a large scale population based cohort study in Japan[J]. Arch Intern Med, 2006,166(17):1871-1877.
47. Washio M, Mori M, Khan M, et al. Diabetes mellitus and kidney cancer risk:the result of Japan Collaborative Cohort Study for Evaluation of cancer risk[J]. Int J Urol,2007,14(5):393-397.
48. Atchison EA, Gridley G, Carreon JD, et al. Risk of cancer in a large cohort of U.S. veterans with diabetes[J]. Int J Cancer,2011,128 (3):635-643.
49. Pollak MN, Schernhammer ES, Hankinson SE. Insulin-like growth factors and neoplasia[J]. Nat Rev Cancer,2004,4(7):505-518.
50.龚勇,赵秋,杨芳,等.胰腺导管腺癌组织中WT1、IGF-IR表达与细胞凋亡的关系[J].世界华人消化杂志,2006,14(28)：2810-2814.
51. Leroith D, Helman L. The new kid on the block(ade) of the IGF-1 receptor[J]. Cancer Cell,2004,5(3):201-202.
52. Misawa A, Hosoi H, Arimoto A, et al. N-Myc Induction Stimulated by Insulin-like Growth Factor I through Mitoge-nactivated Protein Kinase Signaling Pathway in Human Neuroblastoma Cells [J]. Cancer Research,2000,60(1):64-69.
53. Girnita L, Girnita A, Brodin B, et al. Increased Expression of Insulin-like Growth Factor I Receptor in Malignant Cells Expressing Aberrant p53: Functional Impact [J]. Cancer Research,2000,60(18):5278-5283.
54. Stearns M, Tran J, Francis MK, et al. Activated ras enhances insulin-like growth factor 1 induction of vascular endothelial growth factor in prostate epithelial cells[J]. Cancer Res,2005,65(6):2085-2088.
55.耿蕾,崔竹梅,李玉军,等.HIF-1α、VEGF及IGF-I蛋白在子宫内膜癌中的表达及其意义[J].齐鲁医学杂志,2006,21(3)：207-209.
56. Kim KS, Seu YB, Raek SH, et al. Induction of cellular senescence by insulin-like growth factor binding protein-5 through a p53-dependent mechanism[J]. Molecular Biology of the Cell,2007,18(11):4543-4552.
57. Renehan AG, Zwahlen M, Minder C, et al. Insulin-like growth factor binding protein-3 and cancer risk:systematic review and meta-regression analysis[J]. Lancet,2004,363(9418):1346-1353.
58.谢锡驹,王水.IGF.-Ⅱ在乳腺癌中的现状及展望[J].江苏医药,2007,33(4)：383-385.
59. Gupta K, Krishnaswamy G, Karnad A, et al. Insulin:a novel factor in carcinogenesis[J]. Am J Med Sci,2002,323(3):140-145.
60. FisherWE, Muscrella P, Boros LG, et al. Vaeiableeffect of streptozotocin-diabetes on the growth of hamster pancreatic cancer(H2T) in the Syrian hamster and nude mouse [J]. Surgery,1998,123 (3):315-320.
61. Chang CK, Ulrich CM. Hyperinsulinaemia and Hyperglycaemia:Possible Risk Factors of Colorectal Cancer Among Diabetic Patients[J]. Diabetologia,2003, 46(5):595-607.
62. Gerstein HC. Dose insulin therapy promote, reduce, or have a neutral effect on cancer[J]. JAMA,2010,303(5):446-447.
63. Currie CJ, Poole CD, Gale EA. The influence of glucose-lowering therapies on cancer risk in type 2 diabetes[J]. Diabetologia,2009,52(9):1766-1777.
64. Sandulache VC, Ow TJ, Pickering CR, et al. Glucose, not glutamine, is the dominant energy source required for proliferation and survival of head and neck squamous carcinoma cells[J]. Cancer,2011,117(13):2926-2938.
65. Jiao L, Weinstein ST, Albanes D, et al. Evidence the serum levels of the soluble receptor for advanced glycation end products are inversely associated wiyh pancreatic cancer risk:a prospective study[J]. Cancer Res,2011,71(10): 3582-3589.
66. Krone CA, Ely JT. Controlling hyperglycemia as an adjunct to cancer therapy[J]. Integrative Cancer Therapies,2005,4(1):25-31.
67. Guastam acchia E, Resta F, Triggiani V, et al. Evidence for a putative relationship between type 2 diabetes and neoplasia with particular reference to breast cancer: role of homones, growth factor and specific repectors[J]. Curr Drug Targets Immune Endocr Metabol Disord,2004,4(1):59-66.
68. Giovannucci E, Harlan DM, Archer MC, et al. Diabetes and cancer:a consensus report[J]. Diabetes Care,2010,33(7):1674-1685.
69. Healy LA, Ruan AM, Carroll P, et al. Metabolic syndrome, central obesity and insulin resistance are associated with adverse pathological features in postmenopausal breast cancer[J]. Clin Oncol,2010,22(4):281-288.
70. Goodwin PJ, Ennis M, Bahl M, et al.High insulin levels in newly diagnosed breast cancer patients reflect underlying insulin resistance and are associated with components of the insulin resistance syndrome[J]. Breast Cancer Res Treat,2008, 4(25):235-243.
71.田春桃,焦顺昌,魏秀芳.胰岛素对人乳腺癌细胞影响的体外实验研究[J].现 代肿瘤医学,2006,14(11)：1338-1340.
72.赵敬,薛凤霞,华绍芳,等.胰岛素对子宫内膜癌细胞增殖和凋亡的影响[J].中华妇产科杂志,2007,42(10)：696-700.
73. Komninou D, Ayonote A, Richie JP, et al. Insulin resistance and its contribution to colon carcinogenesis[J]. Exp Biol Med,2003,228(4):396-405.
74. Chang CK, Ulrich CM. Hyperinsulinaemia and hyperglycaemia:possible risk factors of colorectal cancer among diabetic patients[J]. Diabetologia,2003, 46(5):595-607.
75. Samantha L, Sumit R, Paul V, et al. Increased cancer-related mortality for patients with type 2 diabetes who use sulfonylureas or insulin[J]. Diabetes Care,2006, 29(8):254-258.
76. Mannucci E, Monami M, Balzi D, et al. Doses of insulin and its analogues and cancer occurrence in insulin-treated type diabetic patients[J]. Diabetes Care,2010, 33(9):1997-2003.
77.华绍芳,薛凤霞,赵敬.磷脂酰基醇3激酶通路在胰岛素调节子宫内膜癌细胞增殖凋亡中的作用[J].中华实用妇科与产科杂志,2007,23(11)：843-845.
78. Joan Fortuny, Yolanda B, Ramon Bosch, et al. Type 2 diabetes mellitus, its treatment and risk for lymphoma[J]. European Journal of cancer,2005, 41(12):1782-1787.
79. Yang YX, Hennessy S, Lewis JD. Insulin therapy and colorectal cancer risk among type 2 diabetes mellitus patients[J]. Gastroenterology,2004, 127(4):1044-1050.
80. Evans JM, Donnelly LA, Emslie-Smith Am, et al. Metformin and reduced risk of cancer in diabetic patients[J]. BMJ,2005,330(7503):1304-1305.
81.Micic D, Cvijovic Q Trajkovic V, et al. Metformin:its emerging role in oncology[J]. Hormones,2011,10(1):5-15.
82. Poggi A, Zocchi MR. Mechanisms of tumor escape:role of tumor microenvironment in inducing apoptosis of cytolytic effector cells [J]. Arch Immunol Ther Exp,2006,54(5):323-333.
83.孙明谨,曾玉琴,李雪锋,等.糖尿病患者T,B细胞亚群的研究[J].中国现代医学杂志,2005,15(22)：3455-3457.
84. Schott S, Bierhaus A, Schuetz F, et al. Therapeutic effects of metformin in breast cancer:involvement of the immune system[J]. Cancer Immunol Immunother, 2011,Jue17.
85. Huang B, Pan PY, Li Q, et al. Gr-1+CD115+immature myeloid suppressor cells mediate the development of tumor-induced T regulatory cells and T-cell anergy in tumor-bearing host[J]. Cancer Res,2006,66(2):1123-1131.
86. Zea AH, Rodriguez PC, Atkins MB, et al. Arginase-producing myeloid suppressor cells in renal cell carcinoma patients a mechanism of tumor evasion[J]. Cancer Res,2005,65(8):3044-3048.
87. Rodriguez PC, Quiceno DG, Zabaleta J, et al. Arginase I production in the tumor microenvironment by mature myeloid cells inhibits T-cell receptor expression and antigen-specific T-cell responses[J]. Cancer Res,2004,64(16):5839-5849.
88. Balkwill F, Charles KA, Mantovani A. Smoldering and polarized inflammation in the initiation and promotion of malignant disease [J]. Cancer Cell,2005, 7(3):211-217.
89. Hussain SP, Harris CC. Inflammation and cancer:an ancient link with novel potentials[J]. Int J Cancer 2007,121(11):2373-2380.
90. Weinberg R, Mihich E. Eighteenth annual pezcoller symposium:tumor microenvironment and heterotypic interactions [J]. Cancer Res,2006,66 (24):11550-11553.
91.Coussens LM, Werb Z. Inflammation and cancer [J]. Nature,2002, 420(6917):860-867.
92. Karin M, Greten FR. NFkappaB:linking inflammation and immunity to cancer development and progression[J]. Nat Rev Immunol,2005,5(10):749-759.
93. Rius J, Guma M, Schachtrup C, et al. NFkappaB links innate immunity to the hypoxic response through transcriptional regulation of HIF-1alpha[J]. Nature, 2008,453(7196):807-811.
94. Bromberg J, Wang TC. Inflammation and Cancer:IL-6 and STAT3 Complete the Link[J]. Cancer Cell,2009,15(2):79-80.
95. Whiteside TL. The tumor microenvironment and its role in promoting tumor growth[J]. Oncogene,2008,27(45):5904-5912.
96. Zhang B. Targeting the stroma by T cells to limit tumor growth[J]. Cancer Res, 2008,68(23):9570-9573.
97. Feinberg AP, Tycko B. The history of cancer epigenetics[J]. Nat Rev Cancer, 2004,4(2):143-153.
98. Hodge DR, Cho E, Copeland TD, et al. IL-6 enhances the nuclear translocation of DNA cytosine-5-methyl-transferasel (DNMT1) via phosphorylation of the nuclear localization sequence by the AKT kinase[J], Cancer Genomics Proteomics, 2007,4(6):387-398.
99. Kanai Y. Alterations of DNA methylation and clinicopathological diversity of human cancers[J]. Pathol Int,2008,58(8):544-558.
100.赵金艳,高春芳.代谢综合征与肝癌流行病学研究进展[J].第二军医大学学报,2010,31(4)：425-428.
101. Svacina S. Colorectal cancer and diabetes[J]. Vnitr Lek,2011,57(4):378-380.
2.赵金艳,高春芳.代谢综合征与肝癌流行病学研究进展[J].第二军医大学学报,2010,31(4)：425-428.
3.王爱萍,刘超,庞春梅等.2型糖尿病与恶性肿瘤临床相关性研究[J].南京医科大学学报,2007,27(1)：55-58.
4. Davila JA, Morgan RO, Shaib Y, et al. Diabetes increases the risk of hepatocellular carcinoma in the United States:a population based case control study[J]. Gut,2005,54(3):533-539.
5.邵清,陈国凤,李梵.慢性丙型肝炎病毒感染者2型糖尿病流行病学调查[J].肝脏,2007,12(5)：366-367.
6. Serag H, Tran T, Everhart JE. Diabetes increases the risk of chronic liver disease and hepatocellular carcinoma[J]. Gastroenterology,2004,126(2):460-468.
7. Staattin P, Ove Bjor, Ferrari P, et al. Prospective study of Hyperglycemia and cancer risk[J]. Diabetes Care,2007,30(3):561-567.
8. Serag H, Richardson P, Everhart J. The role of diabetes in hepatocellular carcinoma:a case-control study among United States Veterans[J]. Am J Gastroenterol,2001,96(8):2462-2467.
9.季尚玮,王江滨.2型糖尿病对原发性肝癌发病的危险性分析[J]。肝脏,2007,12(3)：168-170.
10.王艳丽.糖尿病诱发原发性肝癌的危险性评价及与其他相关危险因素协同致病作用分析[D].吉林大学,2005(04).
11. Wang F, Gupta S, Holly FA. Diabetes mellitus and pancreatic cancer in a population-based case-control study in the San Franxisco Bay Area, Califomia[J]. Cancer Epidemiol Biomarkers Prev,2006,15(8):1458-1463.
12. Chun Gao, Shu-Kun Yao. Diabetes mellitus:a "true" independent risk factor for hepatocellular carcinoma?[J]. Hepatobiliary Pancreat Dis Int.2009,8(2):465-473.
13.向红丁,张化冰.ADA：糖尿病医学诊治标准-2010(摘要)[J].中国糖尿病 杂志,2010,18(3)：164-171.
14.陈小箐,周凯.原发性肝癌患病的糖尿病相关因素研究[J]. Acta Academiae Medicine Jiangxi,2009,49(5):80-82.
15.刘成,马翔,马依彤等.新疆地区汉族、维吾尔族、哈萨克族成年人群超重和肥胖流行病学调查[J].中华流行病学杂志,2011,31(10)：1139-1143.
16.杨秉辉,张博恒.重视肝癌的病因和预防[J].临床肝胆病杂志,2012,28(4)：253-255.
17. Wilson EB, Maher HC. Cancer and tuberculosis with some comments on cancer and other diseases[J]. Am J Cancer,1932,16(1):227-250.
18. Lawson DH, Gray JM, Mckillop C, et al. Diabetes mellitus and primary hepatocellular carcinoma[J]. QJ Med,1986,61(2):945-955.
19. Hemminki K, Li X, Sundquist J, et al. Risk of cancer following hospitalization for type 2 diabetes[J]. Oncologist,2010,15(6):548-555.
20. Noto H, Osameic K, Sasazuki T, et al. Substantially increased risk of cancer in patients with diabetes mellitus:a systematic review and meta-analysis of epidemiologic evidence in Japan[J]. J Diabetes Complications,2010, 24(5):345-353.
21. Adami HO, Chow WH, Nyren O. Excess risk of primary liver cancer in patients with diabetes mellitus[J]. Natl Cancer Inst.1996,88(2):1472-1477.
22. El Serag HB, Richardson PA, Everhart JE, et al. Diabetes increases the risk of chronic liver disease and hepatocellular carcinoma[J].2004,126(7):460-468.
23. Davila JA, Morgan RO, Shaib Y, et al. Diabetes increases the risk of hepatocellular carcinoma in the United States:a population based case control study[J]. Gut,2005,54(3):533-539.
24. Giovannucci E, Harlan DM, Archer MC, et al. Diabetes and cancer:a conaensus report[J]. Diabetes Care,2010,33(7):1674-1685.
25.周慧敏.不同浓度的葡萄糖和胰岛素对Colon肿瘤细胞分泌免疫抑制物的影响及其分子机制的研究[D].河北医科大学,2009(4).
26.赵晔.S-腺苷蛋氨酸对胃癌增殖和C-myc、uPA基因甲基化的影响[D].郑州大 学,2010(5).
27.于志清,潘海林,黎英荣等.中国亚热带地区糖尿病流行与分型研究[J].广西医科大学学报,1999,16(6)：728-731.
28. Chang CK, Ulrich CM. Hyperinsulinaemia and Hyperglycaemia:Possible Risk Factors of Colorectal Cancer Among Diabetic Patients[J]. Diabetologia,2003, 46(5):595-607.
29. Jiao L, Weinstein ST, Albanes D, et al. Evidence the serum levels of the soluble receptor for advanced glycation end products are inversely associated wiyh pancreatic cancer risk:a prospective study[J]. Cancer Res,2011,71(10): 3582-3589.
30. Krone CA, Ely JT. Controlling hyperglycemia as an adjunct to cancer therapy[J]. Integrative Cancer Therapies,2005,4(1):25-31.
31.苏苹,郭庆畲.糖尿病、降糖治疗和癌症[J].中国老年保健医学杂志,2010,8(5)：5-7.
32. Currie CJ, Poole CD, Gale EA. The influence of glucose-lowering therapies on cancer risk in type 2 diabetes[J]. Diabetologia,2009,52(9):1766-1777.
33. Sandulache VC, Ow TJ, Pickering CR, et al. Glucose, not glutamine, is the dominant energy source required for proliferation and survival of head and neck squamous carcinoma cells[J]. Cancer,2011,117(13):2926-2938.
34.孙丽.糖尿病与恶性肿瘤相关性的作用机制研究进展[J].中国药师,2009,12(12)：1730-1732.
35.田爱平.胰岛素信号途径—肿瘤治疗的新靶点?[J].癌症进展杂志,2009,7(1)：66-72.
36. Reinmuth N, Fan F, LiuW, et al. Impact of insulin-like growth factor receptor function angio genesis, growth,and metastasis of colon cancer[J]. Lab Invest, 2002,82(10):1377-1389.
37. Guan Z, Buckman SY, Baier LD, et al. IGF-I and insulin amplify IL-1 beta-induced nitric oxide and prostaglandin biosynthesis[J]. Am J Physiol.1998, 274(4Pt2):F673-679.
38. Gupta K, Krishnsswaray G, Karnad A, et al. Insulin:a novel factor in carcinogenesis[J]. Am J Med Sci,2002,323(3):140-145.
39. Healy LA, Ruan AM, Carroll P, et al. Metabolic syndrome, central obesity and insulin resistance are associated with adverse pathological features in postmenopausal breast cancer[J]. Clin Oncol,2010,22(4):281-288.
40. Goodwin PJ, Ennis M, Bahl M, et al.High insulin levels in newly diagnosed breast cancer patients reflect underlying insulin resistance and are associated with components of the insulin resistance syndrome[J]. Breast Cancer Res Treat,2008, 4(25):235-243.
41.田春桃,焦顺昌,魏秀芳.胰岛素对人乳腺癌细胞影响的体外实验研究[J].现代肿瘤医学,2006,14(11)：1338-1340.
42.赵敬,薛凤霞,华绍芳,等.胰岛素对子宫内膜癌细胞增殖和凋亡的影响[J].中华妇产科杂志,2007,42(10)：696-700.
43.龚勇,赵秋,杨芳,等.胰腺导管腺癌组织中WT1、IGF-IR表达与细胞凋亡的关系[J].世界华人消化杂志,2006,14(28)：2810-2814.
44. Kurtaran A, Li SR, Raderer M, et al. Technetium-99m-galactosyl-neogolycoalbumin combined with iodine-123-Tyr-(A14)-insulin visualizes human hepatocellular carcinomas. J Nucl Med,1995,36(10):1875-1881.
45. Mosmann T. Rapid colorimetric assay of cellular growth and survival:application to proliferation and cytotoxiciyt assays[J]. J Immunol Methods,1983, 65(1):55-63.
46. Hino N, Higashi T, Nouso K, et al. Apoptosis and proliferation of human hepatocellular carcinoma[J]. Liver,1996,16(4):123-129.
47.潘銮凤.分子生物学技术[M].上海：复旦大学出版,2008,158-160.
48. Uefuji K, Ichikura T, Mochizuki H, et al. Expression of cyclooxygenase in human gastric adenomas and adenocarcinomas[J]. J Surg Oncol,2001,76(1):26-30.
49.张琛,夏海滨.血管内皮细胞生长因子及其表达调控研究进展[J].现代肿瘤学杂志,2012,20(4)：841-845.
50.陈建国.中国肝癌发病趋势和一级预防.临床肝胆病杂志[J].2012,28(4)：256-260.
51. Joo YE, Oh WT, Rew JS, et al. Cyclooxygenase-2 expression is associated with well-differentiated and intestinal-type pathways in gastric carcinogenesis[J]. Digestion,2002,66(4):222-229.
52. Nashida T, Yabe Y, Fu HY, et al. Geranylgeranylacetone induces cyclooxygenase-2 expression in cultured rat gastric epithelial cells through NF-kappaB[J]. Dig Dis Sci,2007,52(8):1890-1896.
53. Chang CK, Ulrich CM. Hyperinsulinaemia and Hyperglycaemia:Possible Risk Factors of Colorectal Cancer Among Diabetic Patients[J]. Diabetologia,2003, 46(5):595-607.
54.赵锋,曾春,周伟雄,等.初发2型糖尿病患者血清COX-2活性与IL-18含量改变的意[J].热带医学杂志,2006,6(10)：108-1083.
55. Persaud SJ, Muller D, Belin VD, et al. The role of arachidonic acid and its metabolites in insulin secretion from human islets of langerhans[J]. Diabetes, 2007,56(1):197-203.
56. Persaud SJ, Bums CJ, Belin VD, et al. Glucose-induced regulation of COX-2 expression in human islets of langerhans[J]. Diabetes,2004,53(Suppl 1):S190-S192.
57. Yuan L, Ziegle RR, Haman NA. Metformin modulates insulin post-receptor signaling transduction in chronically insulin-treated HepG2 cells [J]. Acta Pharmacal Sin,2003,24(1):55-60.
58. Joan Fortuny, Yolanda B, Ramon Bosch, et al. Type 2 diabetes mellitus, its treatment and risk for lymphoma[J]. European Journal of cancer,2005, 41(12):1782-1787.
59.阮丹杰,王伟.炎性因子与胰岛素抵抗的研究进展[J].医学综述,2008,15(4)：508-511.
60. Pollak M. Insulin, insulin-like growth factors and neoplasia[J]. Best Pract. Res.Clin. Endocrinol,2008,22(4):625-628.
61. Lee HC, Tian B, Sedivy JM. Loss of Rafkinase inhibitor protein promotes cell proliferation and migration of human hepatoma cells[J]. Gastroenterology,2006, 131(7):1208-1217.
62. Valter Donadon, Massimiliano Balbi, Giorgio Zanette, et al. Hyperinsulinemia and risk for hepatocellullar carcinoma in patients with chronic liver disease and type 2 diabetes mellitus[J]. Gastroenterol,2009,3(5):465-467.
63. Kunlin J, Yonjhua Z, Yuniuan S, et al. Vascular endothelial growth factor(VEGF) stimulates neorogenesis in vitor and in vivo. Proc Natl Acad Sei USA,2000, 97(18):1024-1027.
64. Ferrara N, Henzel WJ. Pituitary follicular cells secrete a novel heparin-binding growth factor specific for vascular endothelial cells[J]. Biochem Biophys Res Commun,1989,161(2):851-858.
65. Ho QT, Kuo CJ. Vascular endothelial growth factor:biology and therapeutic applications[J]. Int J Biochem Cell Biol,2007,39(78):1349-1357.
66. Maharaj AS, D'Amore PA. Role for VEGF in adult [J]. Microvasc Res, 2007,74(2-3):100-113.
67. Tung Ping Poon R, Fan ST, Wong J, et al. Risk factors prevention and managent of postoperative recurrence after resection of hepatocellular carcinoma[J]. Ann Surg,2000,232(8):10-24.
68.王越晖,王心蕊,朱向红等。高糖、高胰岛素对兔视网膜Muller细胞血管内皮生长因子表达的影响。中华糖尿病杂志,2004,12(4)：277-280.
69. Girnita L, Girnita A, Brodin B, et al. Increased Expression of Insulin-like Growth Factor I Receptor in Malignant Cells Expressing Aberrant p53 Functional Impact [J]. Cancer Research,2000,60(18):5278-5283.
70. Kim KS, Seu YB, Raek SH, et al. Induction of cellular senescence by insulin-like growth factor binding protein-5 through a p53-dependent mechanism[J]. Molecular Biology of the Cell,2007,18(11):4543-4552.
71.彭军,章军建,谢军等。高浓度葡萄糖诱导血管内皮细胞凋亡及对VEGF表达的影响。Stroke and Nervous Diseases,2007,14(4):238-241.
1. Wilson EB, Maher HC. Cancer and tuberculosis with some comments on cancer and other diseases[J]. Am J Cancer,1932,16(1):227-250.
2. Coughlin SS, Calle EE, Teras LR, et al. Diabetes mellitus as a predictor of cancer mortality in a large cohort of US adults[J]. Am J Epidemiol,2004, 159(12):1160-1167.
3. Giovannucci E, Harlan DM, Archer MC, et al. Diabetes and cancer:a conaensus report[J]. Diabetes Care,2010,33(7):1674-1685.
4.王爱萍,刘超,庞春梅,等.2型糖尿病与恶性肿瘤临床相关性研究[J].南京医科大学学报,2007,27(1)：55-58.
5. Hemminki K, Li X, Sundquist J, et al. Risk of cancer following hospitalization for type 2 diabetes[J]. Oncologist,2010,15(6):548-555.
6. Noto H, Osameic K, Sasazuki T, et al. Substantially increased risk of cancer in patients with diabetes mellitus:a systematic review and meta-analysis of epidemiologic evidence in Japan[J]. J Diabetes Complications,2010, 24(5):345-353.
7. Parkin D M. Global cancer statistics in the year 2000[J]. Lancet Oncol,2001, 2(9):533-543.
8.陈建国,宋新明.中国肝癌发病水平的估算及分析[J].中国肿瘤,2005,14(1)：28-31.
9.蒋俊俊,钟秋安,梁浩,等.糖尿病与原发性肝癌关系的病例对照研究Meta分析[J].中国慢性病预防与控制,2010,18(2)：132-135.
10. Staattin P, Ove Bjor, Ferrari P, et al. Prospective study of Hyperglycemia and cancer risk[J]. Diabetes Care,2007,30(3):561-567.
11. Polesel J, Zucchetto A, Montecca M, et al. The impact of obesity and diabetes mellitus on the risk of hepatocellular carcinoma[J]. Ann Oncol,2009, 20(2):353-359.
12. Bonovas S, Filioussi K, Tsantes A, et al. Diabetes mellitus and risk of prostate cancer:A meta-analysis[J]. Diabetologia,2004,47(6):1071-1078.
13. Michels KB, Solomon CG, Hu FB, et al. Type 2 diabetes and subsequent incidence of breast cancer in the Nurses'Health Study[J]. Diabetes Care,2003, 26(6):1752-1758.
14. Sellers TA, Jensen LE, Vierkant RA, et al. Assocation of diabetes with mammographic breast density and breast cancer in the Minnesota breast cancer family study[J]. Cancer Causes Control,2007,18(5):505-515.
15. Wu AH, Yu MC, Tseng CC, et al. Diabetes and risk of breast cancer in Asian-American women[J]. Carcinogenesis,2007,28(7):1561-1566.
16. Ewertz M, Holmberg L, Tretli S, et al. Risk factors for male breast cancer-a case-control study from Scandinavia[J]. Acta Oncol,2001,40(4):467-471.
17. Talamini R, Franceschi S, Favero A, et al. Selected medical conditions and risk of breast cancer[J]. Br J Cancer,1997,75 (11):1699-1703.
18. Wolf I, Sadetzki S, Gluck I, et al. Association between diabetes mellitus and adverse characteristics of breast cancer at presentation[J]. Eur J Cancer,2006, 42(8):1077-1082.
19. Zhang Y, Liu Z, Yu X, et al. The association between metabolic abnormality and endometrial cancer:a large case-control study in China[J]. Gynecol Oncol,2010, 117(1):41-46.
20.薛莹,刘超.2型糖尿病与恶性肿瘤[J].国际内分泌代谢杂志,2007,27(1)：56-58.
21.宋力.糖尿病作为子宫内膜癌危险因素的Meat分析[J].内蒙古医学杂志,2009,41(11)：1291-1295.
22. Fisher WE, Boros LG, Schirmer WJ. Reversal of enhanced pancreatic cancer growth in diabetes by insulin[J]. Surgery,1995,118(2):453-457.
23. Shibata A, Mack TM, Paganini-Hill A, et al. A prospective study of pancreatic cancer in the elderly[J]. Int J Cancer,1994,58(1):46-49.
24. DeMeo MT. Pancreatic cancer and sugar diabetes[J]. Nutr Rev,2001, 59(4):112-115.
25. Silverman DT, Schiffman M, Everhart J, et al. Diabetes mellitus, other medical conditions and familial history of cancer as risk factors for pancreatic cancer[J]. Br J Cancer,1999,80(11):1830-1837.
26. Jee SH, Ohrr H, Sull JW, et al. Fasting serum glucose level and cancer risk in Korean men and women [J]. JAMA,2005,293(2):194-202.
27. Kuriki K, Hirose K, Tajima K. Diabetes and cancer risk for all and specific sites among Japanese men and women[J]. Eur J Cancer Prev,2007,16(1):83-89.
28. Wang F, Gupta S, Holly FA. Diabetes mellitus and pancreatic cancer in a population-based case-control study in the San Franxisco Bay Area, Califomia[J]. Cancer Epidemiol Biomarkers Prev,2006,15(8):1458-1463.
29. Kellerer M. Von Eye Corleta H, Muhlhofer A, et al. Insulin and Insulin-like growth factor-1 receptor tyrosine-kinase activities in human reanl carcinoma[J]. Int J Cancer,1995,62(5):501-507.
30. McKeown-Eyssen G. Epidemiology of colorectal cancer revisited:Are serum triglycerides and /or plasma glucose associated with risk [J]. Cancer Epidemiol Biomarkers Prev,1994,3(8):687-695.
31.胡水清,汤哲,张玫.糖尿病与结直肠癌发病及转移关系[J].中国肿瘤,2007,16(9)：689-698.
32. Larsson SC, Giovannucci E, Wolk A. Diabetes and colorectal cancer incidence in the cohort of Swedish men[J]. Diabetes Care,2005,28(7):1805-1807.
33. Seow A, Yuan JM, Koh WP, et al. Diabetea mellitus and risk of colorectal cancer in the Singapore Chinese Health Study[J]. J Natl Cancer Inst,2006, 98(2):135-138.
34. Yang YX, Hennessy S, Lewis JD, et al. Type 2 diabetes mellitus and the risk of colorectal cancer[J]. Clin Gastroenterol Hepatol,2005,3(6):587-594.
35. La Vecchia C, Negri E, Decarli A, et al. Diabetes mellitus and colorectal cancer risk [J]. Cancer Epidemiol Biomarkers Prev,1997,6(12):1007-1010.
36. Chung YW, Han DS, Park KH, et al. Insulin therapy and colorectal adenoma risk among patients with Type 2 diabetes mellitus:a case-control study in Korea [J]. Dis Colon Rectum,2008,51(5):593-597.
37. Yang YX, Hennessy S, Lewis JD.Insulin therapy and colorectal cancer risk among type 2 diabetes mellitus patients[J].Gastroenterology,2004,127(4):1044-1050.
38. Davila JA, Morgan RO, Shaib Y, et al. Diabetes increases the risk of hepatocellular carcinoma in the United States:a population based case control study[J]. Gut,2005,54(4):533-539.
39.季尚玮,王江滨.2型糖尿病对原发性肝癌发病的危险性分析[J]。肝脏,2007,12(3)：168-170.
40. Adami HO, Chow WH, Nyren O, et al. Excess risk of primary liver cancer in patients with diabetes mellitus[J]. J Natl Cancer Inst,1996,88(20):1472-1477.
41. Serag H, Tran T, Everhart JE. Diabetes increases the risk of chronic liver disease and hepatocellular carcinoma[J]. Gastroenterology,2004,126(2):460-468.
42. Serag H, Richardson P, Everhart J. The role of diabetes in hepatocellular carcinoma:a case-control study among United States Veterans[J]. Am J Gastroenterol,2001,96(8):2462-2467.
43.王艳丽.糖尿病诱发原发性肝癌的危险性评价及与其他相关危险因素协同致病作用分析[D].吉林大学,2005(04).
44. Kawamura Y, Ikeda K, Arase Y, et al. Diabetes mellitus worsens the recurrence rate after potentially curative therapy in patients with hepatocellular carcinoma associated with nonviral hepatitis [J]. J Gastroenterol Hepatol,2008,23 (11):1739-1746.
45. Lindblad P, Chow WH, Chan J, et al. The role of diabetes mellitus in the aetiology of reanl cell cancer[J]. Diabetologia,1999,42(1):107-112.
46. Inoue M, Iwasaki M, Otani T, et al. Diabetes mellitus and the risk of cancer result from a large scale population based cohort study in Japan[J]. Arch Intern Med, 2006,166(17):1871-1877.
47. Washio M, Mori M, Khan M, et al. Diabetes mellitus and kidney cancer risk:the result of Japan Collaborative Cohort Study for Evaluation of cancer risk[J]. Int J Urol,2007,14(5):393-397.
48. Atchison EA, Gridley G, Carreon JD, et al. Risk of cancer in a large cohort of U.S. veterans with diabetes[J]. Int J Cancer,2011,128 (3):635-643.
49. Pollak MN, Schernhammer ES, Hankinson SE. Insulin-like growth factors and neoplasia[J]. Nat Rev Cancer,2004,4(7):505-518.
50.龚勇,赵秋,杨芳,等.胰腺导管腺癌组织中WT1、IGF-IR表达与细胞凋亡的关系[J].世界华人消化杂志,2006,14(28)：2810-2814.
51. Leroith D, Helman L. The new kid on the block(ade) of the IGF-1 receptor[J]. Cancer Cell,2004,5(3):201-202.
52. Misawa A, Hosoi H, Arimoto A, et al. N-Myc Induction Stimulated by Insulin-like Growth Factor I through Mitoge-nactivated Protein Kinase Signaling Pathway in Human Neuroblastoma Cells [J]. Cancer Research,2000,60(1):64-69.
53. Girnita L, Girnita A, Brodin B, et al. Increased Expression of Insulin-like Growth Factor I Receptor in Malignant Cells Expressing Aberrant p53: Functional Impact [J]. Cancer Research,2000,60(18):5278-5283.
54. Stearns M, Tran J, Francis MK, et al. Activated ras enhances insulin-like growth factor 1 induction of vascular endothelial growth factor in prostate epithelial cells[J]. Cancer Res,2005,65(6):2085-2088.
55.耿蕾,崔竹梅,李玉军,等.HIF-1α、VEGF及IGF-I蛋白在子宫内膜癌中的表达及其意义[J].齐鲁医学杂志,2006,21(3)：207-209.
56. Kim KS, Seu YB, Raek SH, et al. Induction of cellular senescence by insulin-like growth factor binding protein-5 through a p53-dependent mechanism[J]. Molecular Biology of the Cell,2007,18(11):4543-4552.
57. Renehan AG, Zwahlen M, Minder C, et al. Insulin-like growth factor binding protein-3 and cancer risk:systematic review and meta-regression analysis[J]. Lancet,2004,363(9418):1346-1353.
58.谢锡驹,王水.IGF.-Ⅱ在乳腺癌中的现状及展望[J].江苏医药,2007,33(4)：383-385.
59. Gupta K, Krishnaswamy G, Karnad A, et al. Insulin:a novel factor in carcinogenesis[J]. Am J Med Sci,2002,323(3):140-145.
60. FisherWE, Muscrella P, Boros LG, et al. Vaeiableeffect of streptozotocin-diabetes on the growth of hamster pancreatic cancer(H2T) in the Syrian hamster and nude mouse [J]. Surgery,1998,123 (3):315-320.
61. Chang CK, Ulrich CM. Hyperinsulinaemia and Hyperglycaemia:Possible Risk Factors of Colorectal Cancer Among Diabetic Patients[J]. Diabetologia,2003, 46(5):595-607.
62. Gerstein HC. Dose insulin therapy promote, reduce, or have a neutral effect on cancer[J]. JAMA,2010,303(5):446-447.
63. Currie CJ, Poole CD, Gale EA. The influence of glucose-lowering therapies on cancer risk in type 2 diabetes[J]. Diabetologia,2009,52(9):1766-1777.
64. Sandulache VC, Ow TJ, Pickering CR, et al. Glucose, not glutamine, is the dominant energy source required for proliferation and survival of head and neck squamous carcinoma cells[J]. Cancer,2011,117(13):2926-2938.
65. Jiao L, Weinstein ST, Albanes D, et al. Evidence the serum levels of the soluble receptor for advanced glycation end products are inversely associated wiyh pancreatic cancer risk:a prospective study[J]. Cancer Res,2011,71(10): 3582-3589.
66. Krone CA, Ely JT. Controlling hyperglycemia as an adjunct to cancer therapy[J]. Integrative Cancer Therapies,2005,4(1):25-31.
67. Guastam acchia E, Resta F, Triggiani V, et al. Evidence for a putative relationship between type 2 diabetes and neoplasia with particular reference to breast cancer: role of homones, growth factor and specific repectors[J]. Curr Drug Targets Immune Endocr Metabol Disord,2004,4(1):59-66.
68. Giovannucci E, Harlan DM, Archer MC, et al. Diabetes and cancer:a consensus report[J]. Diabetes Care,2010,33(7):1674-1685.
69. Healy LA, Ruan AM, Carroll P, et al. Metabolic syndrome, central obesity and insulin resistance are associated with adverse pathological features in postmenopausal breast cancer[J]. Clin Oncol,2010,22(4):281-288.
70. Goodwin PJ, Ennis M, Bahl M, et al.High insulin levels in newly diagnosed breast cancer patients reflect underlying insulin resistance and are associated with components of the insulin resistance syndrome[J]. Breast Cancer Res Treat,2008, 4(25):235-243.
71.田春桃,焦顺昌,魏秀芳.胰岛素对人乳腺癌细胞影响的体外实验研究[J].现 代肿瘤医学,2006,14(11)：1338-1340.
72.赵敬,薛凤霞,华绍芳,等.胰岛素对子宫内膜癌细胞增殖和凋亡的影响[J].中华妇产科杂志,2007,42(10)：696-700.
73. Komninou D, Ayonote A, Richie JP, et al. Insulin resistance and its contribution to colon carcinogenesis[J]. Exp Biol Med,2003,228(4):396-405.
74. Chang CK, Ulrich CM. Hyperinsulinaemia and hyperglycaemia:possible risk factors of colorectal cancer among diabetic patients[J]. Diabetologia,2003, 46(5):595-607.
75. Samantha L, Sumit R, Paul V, et al. Increased cancer-related mortality for patients with type 2 diabetes who use sulfonylureas or insulin[J]. Diabetes Care,2006, 29(8):254-258.
76. Mannucci E, Monami M, Balzi D, et al. Doses of insulin and its analogues and cancer occurrence in insulin-treated type diabetic patients[J]. Diabetes Care,2010, 33(9):1997-2003.
77.华绍芳,薛凤霞,赵敬.磷脂酰基醇3激酶通路在胰岛素调节子宫内膜癌细胞增殖凋亡中的作用[J].中华实用妇科与产科杂志,2007,23(11)：843-845.
78. Joan Fortuny, Yolanda B, Ramon Bosch, et al. Type 2 diabetes mellitus, its treatment and risk for lymphoma[J]. European Journal of cancer,2005, 41(12):1782-1787.
79. Yang YX, Hennessy S, Lewis JD. Insulin therapy and colorectal cancer risk among type 2 diabetes mellitus patients[J]. Gastroenterology,2004, 127(4):1044-1050.
80. Evans JM, Donnelly LA, Emslie-Smith Am, et al. Metformin and reduced risk of cancer in diabetic patients[J]. BMJ,2005,330(7503):1304-1305.
81.Micic D, Cvijovic Q Trajkovic V, et al. Metformin:its emerging role in oncology[J]. Hormones,2011,10(1):5-15.
82. Poggi A, Zocchi MR. Mechanisms of tumor escape:role of tumor microenvironment in inducing apoptosis of cytolytic effector cells [J]. Arch Immunol Ther Exp,2006,54(5):323-333.
83.孙明谨,曾玉琴,李雪锋,等.糖尿病患者T,B细胞亚群的研究[J].中国现代医学杂志,2005,15(22)：3455-3457.
84. Schott S, Bierhaus A, Schuetz F, et al. Therapeutic effects of metformin in breast cancer:involvement of the immune system[J]. Cancer Immunol Immunother, 2011,Jue17.
85. Huang B, Pan PY, Li Q, et al. Gr-1+CD115+immature myeloid suppressor cells mediate the development of tumor-induced T regulatory cells and T-cell anergy in tumor-bearing host[J]. Cancer Res,2006,66(2):1123-1131.
86. Zea AH, Rodriguez PC, Atkins MB, et al. Arginase-producing myeloid suppressor cells in renal cell carcinoma patients a mechanism of tumor evasion[J]. Cancer Res,2005,65(8):3044-3048.
87. Rodriguez PC, Quiceno DG, Zabaleta J, et al. Arginase I production in the tumor microenvironment by mature myeloid cells inhibits T-cell receptor expression and antigen-specific T-cell responses[J]. Cancer Res,2004,64(16):5839-5849.
88. Balkwill F, Charles KA, Mantovani A. Smoldering and polarized inflammation in the initiation and promotion of malignant disease [J]. Cancer Cell,2005, 7(3):211-217.
89. Hussain SP, Harris CC. Inflammation and cancer:an ancient link with novel potentials[J]. Int J Cancer 2007,121(11):2373-2380.
90. Weinberg R, Mihich E. Eighteenth annual pezcoller symposium:tumor microenvironment and heterotypic interactions [J]. Cancer Res,2006,66 (24):11550-11553.
91.Coussens LM, Werb Z. Inflammation and cancer [J]. Nature,2002, 420(6917):860-867.
92. Karin M, Greten FR. NFkappaB:linking inflammation and immunity to cancer development and progression[J]. Nat Rev Immunol,2005,5(10):749-759.
93. Rius J, Guma M, Schachtrup C, et al. NFkappaB links innate immunity to the hypoxic response through transcriptional regulation of HIF-1alpha[J]. Nature, 2008,453(7196):807-811.
94. Bromberg J, Wang TC. Inflammation and Cancer:IL-6 and STAT3 Complete the Link[J]. Cancer Cell,2009,15(2):79-80.
95. Whiteside TL. The tumor microenvironment and its role in promoting tumor growth[J]. Oncogene,2008,27(45):5904-5912.
96. Zhang B. Targeting the stroma by T cells to limit tumor growth[J]. Cancer Res, 2008,68(23):9570-9573.
97. Feinberg AP, Tycko B. The history of cancer epigenetics[J]. Nat Rev Cancer, 2004,4(2):143-153.
98. Hodge DR, Cho E, Copeland TD, et al. IL-6 enhances the nuclear translocation of DNA cytosine-5-methyl-transferasel (DNMT1) via phosphorylation of the nuclear localization sequence by the AKT kinase[J], Cancer Genomics Proteomics, 2007,4(6):387-398.
99. Kanai Y. Alterations of DNA methylation and clinicopathological diversity of human cancers[J]. Pathol Int,2008,58(8):544-558.
100.赵金艳,高春芳.代谢综合征与肝癌流行病学研究进展[J].第二军医大学学报,2010,31(4)：425-428.
101. Svacina S. Colorectal cancer and diabetes[J]. Vnitr Lek,2011,57(4):378-380.