龙须菜藻红蛋白抗肿瘤作用及其机制的研究
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摘要
本文研究了汕头南澳海域龙须菜(Gracilaria Lemaneiformis)藻红蛋白(Phycoerythrin,PE)的抗肿瘤活性。通过建立肿瘤小鼠模型,研究藻红蛋白对肝癌细胞H_(22)的抗肿瘤作用,并探讨藻红蛋白对H_(22)荷瘤小鼠免疫作用的影响和对肿瘤组织p53等基因表达的影响,对体外培养的肿瘤细胞H_(22)生长的抑制作用以及对肿瘤抑制基因p53表达的影响,以揭示其抗肿瘤机制。
小鼠体内抗肿瘤实验结果显示:经口灌胃给予PE,当给药剂量是300 mg/kg·d时,抑瘤率达到30.23%;对荷瘤小鼠红细胞中超氧化物岐化酶(SOD)活性和肝匀浆中过氧化脂质(LPO)含量的测定结果表明,PE具有提高机体抗氧化能力的作用;PE对胸腺、脾脏等免疫器官有很好的保护作用,PE能提高荷瘤小鼠脾淋巴细胞对ConA的增殖反应,增强巨噬细胞吞噬活性和NK细胞杀伤活性以及红细胞对肿瘤细胞的黏附性,并能促进TNF-α的分泌。
利用半定量RT-PCR法探讨了PE对H_(22)荷瘤肿瘤小鼠肿瘤组织中p53和Caspese-3基因转录水平的影响。结果表明,PE能促进肿瘤组织中肿瘤抑制基因p53和与凋亡相关的蛋白水解酶Caspese-3基因的表达,说明在体内PE具有促进肿瘤抑制基因表达,从而抑制肿瘤生长,甚至导致肿瘤细胞凋亡。
采用细胞生长曲线法检测PE对体外培养的H_(22)肿瘤细胞的影响,结果显示PE对H_(22)肿瘤细胞没有抑制作用。试验结果提示,PE具有的抗肿瘤活性不是直接对H_(22)细胞的作用,可能是通过促进肿瘤组织中肿瘤抑制基因p53和与凋亡相关的蛋白水解酶Caspese-3基因的表达、提高机体免疫力和抗氧化作用得以实现的。
本文还初步探讨了龙须菜多糖(PGL)的免疫活性,利用环磷酰胺建立免疫低下小鼠模型(IDMM),研究PGL对IDMM免疫的调节作用。结果表明,PGL具有提高免疫低下小鼠免疫力的作用,但这种作用无量效关系,与多糖所含硫酸基也无直接联系。
In our study,the antitumor effect of Phycoerythrin(PE) from Gracilaria Lemaneiformis, collected from the sea area around Nan'ao Island,Shantou city,Guangdong province was studied.Throuth the methods of immune response of mice beating H_(22) hepatocellular carcinoma and effection to transcription of p53 and Caspese-3 gene to discuss the effection and mechanism of the anti-tumor activity of PE.
The antitumor experiments indicated that the Phycoerythrin could significantly inhibit the growth of H_(22) hepatocellular carcinoma.At the dose of 300mg·kg~(-1) when given irrigatively(ig),the inhibition rate was 30.23%.The tests of superoxide dismutase(SOD) activity in red blood cell and lipoperoxide(LPO) level in liver homogenate suggested that the anticancer effect of phycobiliprotein might be related to its antioxidation.The results showed that PE can stop the decrease of body weight caused by tumor-beating and protect spleen and thymus gland and improved their activities.The results also showed that crude Phycoerythrin from Gracilaria lemaneiformis can improve lymphocyte responses induced by ConA of tumor beating mice,increased spleen cell turn to lymphocyte.PE can increase the activity of macrophage and natural killer cell,the adhesive ability of erythrocyte immunity to tumor cells and the production of TNF-α.the result showed that maybe increase the immune response of mice beating H_(22) hepatocellular carcinoma to inhibit the growth of the tumor,even cause the tumor to apoptosis.
The effect of PE to the transcription of p53 and Caspese-3 gene in the tumor of mice beating H_(22) hepatocellular carcinoma was studied by semi-quantitaty RT-PCR. The result showed that in vivo mRNA of p53 and Caspese-3 gene in the tumor of mice beating H_(22) hepatocellular carcinoma given irrigatively(ig) PE is over-expressed.But inhibitory effect and over-expression is not found,in vitro.
The immunodeficiency mice mode(IDMM) was estabulished with Cyclophosphamide injected.The immunoregulatory activity of polysaccharides from Gracilaria lemaneiformis(PGL) was studied.The results showed that PGL could enhance immune- funcation of immnodepressive mice.But the results showed that there's no does-effect relationgship between activity of PGL enhancing immunofuncation and PGL.does-effect relationgship between activity of enhancing immunofuncation and contents of sulphate in PGL was not found.
小鼠体内抗肿瘤实验结果显示:经口灌胃给予PE,当给药剂量是300 mg/kg·d时,抑瘤率达到30.23%;对荷瘤小鼠红细胞中超氧化物岐化酶(SOD)活性和肝匀浆中过氧化脂质(LPO)含量的测定结果表明,PE具有提高机体抗氧化能力的作用;PE对胸腺、脾脏等免疫器官有很好的保护作用,PE能提高荷瘤小鼠脾淋巴细胞对ConA的增殖反应,增强巨噬细胞吞噬活性和NK细胞杀伤活性以及红细胞对肿瘤细胞的黏附性,并能促进TNF-α的分泌。
利用半定量RT-PCR法探讨了PE对H_(22)荷瘤肿瘤小鼠肿瘤组织中p53和Caspese-3基因转录水平的影响。结果表明,PE能促进肿瘤组织中肿瘤抑制基因p53和与凋亡相关的蛋白水解酶Caspese-3基因的表达,说明在体内PE具有促进肿瘤抑制基因表达,从而抑制肿瘤生长,甚至导致肿瘤细胞凋亡。
采用细胞生长曲线法检测PE对体外培养的H_(22)肿瘤细胞的影响,结果显示PE对H_(22)肿瘤细胞没有抑制作用。试验结果提示,PE具有的抗肿瘤活性不是直接对H_(22)细胞的作用,可能是通过促进肿瘤组织中肿瘤抑制基因p53和与凋亡相关的蛋白水解酶Caspese-3基因的表达、提高机体免疫力和抗氧化作用得以实现的。
本文还初步探讨了龙须菜多糖(PGL)的免疫活性,利用环磷酰胺建立免疫低下小鼠模型(IDMM),研究PGL对IDMM免疫的调节作用。结果表明,PGL具有提高免疫低下小鼠免疫力的作用,但这种作用无量效关系,与多糖所含硫酸基也无直接联系。
In our study,the antitumor effect of Phycoerythrin(PE) from Gracilaria Lemaneiformis, collected from the sea area around Nan'ao Island,Shantou city,Guangdong province was studied.Throuth the methods of immune response of mice beating H_(22) hepatocellular carcinoma and effection to transcription of p53 and Caspese-3 gene to discuss the effection and mechanism of the anti-tumor activity of PE.
The antitumor experiments indicated that the Phycoerythrin could significantly inhibit the growth of H_(22) hepatocellular carcinoma.At the dose of 300mg·kg~(-1) when given irrigatively(ig),the inhibition rate was 30.23%.The tests of superoxide dismutase(SOD) activity in red blood cell and lipoperoxide(LPO) level in liver homogenate suggested that the anticancer effect of phycobiliprotein might be related to its antioxidation.The results showed that PE can stop the decrease of body weight caused by tumor-beating and protect spleen and thymus gland and improved their activities.The results also showed that crude Phycoerythrin from Gracilaria lemaneiformis can improve lymphocyte responses induced by ConA of tumor beating mice,increased spleen cell turn to lymphocyte.PE can increase the activity of macrophage and natural killer cell,the adhesive ability of erythrocyte immunity to tumor cells and the production of TNF-α.the result showed that maybe increase the immune response of mice beating H_(22) hepatocellular carcinoma to inhibit the growth of the tumor,even cause the tumor to apoptosis.
The effect of PE to the transcription of p53 and Caspese-3 gene in the tumor of mice beating H_(22) hepatocellular carcinoma was studied by semi-quantitaty RT-PCR. The result showed that in vivo mRNA of p53 and Caspese-3 gene in the tumor of mice beating H_(22) hepatocellular carcinoma given irrigatively(ig) PE is over-expressed.But inhibitory effect and over-expression is not found,in vitro.
The immunodeficiency mice mode(IDMM) was estabulished with Cyclophosphamide injected.The immunoregulatory activity of polysaccharides from Gracilaria lemaneiformis(PGL) was studied.The results showed that PGL could enhance immune- funcation of immnodepressive mice.But the results showed that there's no does-effect relationgship between activity of PGL enhancing immunofuncation and PGL.does-effect relationgship between activity of enhancing immunofuncation and contents of sulphate in PGL was not found.
引文
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