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目的 :观察表达 m IL- 18的重组腺病毒基因修饰的胎肝细胞 (Adm IL- 18/ BNL.CL2 )经脾移植对正常小鼠免疫功能的影响。方法 :实验组小鼠经脾移植 Adm IL- 18/ BNL.CL2 ,同时设 Lac Z病毒对照组 (Ad- Lac Z/ BNL.CL2 ) ,BNL.CL2 细胞对照组及空白对照组。2周后处死 ,留取血清 ,制备腹腔巨噬细胞、脾淋巴细胞、肝组织匀浆液 ,提取肝组织总 RNA。采用 ELISA法检测各组小鼠血清、腹腔 Mφ和脾细胞培养上清、肝匀浆中细胞因子的含量 ;采用半定量 RT- PCR法 ,检测肝组织细胞因子 m RNA相对表达量 ;以 LDH释放法测定腹腔 Mφ杀伤活性和脾 NK细胞活性 ,用 MTT还原比色法测定脾淋巴细胞的增殖活性。结果 :实验组小鼠血清、细胞培养上清及肝匀浆中 ,IL- 18、IL - 2、IFN-γ、TNF-α含量均高于其它对照组 ,而 IL - 4、IL - 10水平则低于对照组 ;半定量 RT- PCR结果与EL ISA检测结果一致 ;同时 ,实验组腹腔 Mφ的杀伤活性和脾 NK细胞活性 ,及脾淋巴细胞增殖活性也明显高于对照组。结论 :Adm IL - 18能有效转染至胎肝细胞并稳定表达 m IL- 18;Adm IL - 18基因修饰的胎肝细胞经脾移植后 ,可显著提高肝脏、脾脏免疫细胞活性 ,活化腹腔 Mφ,促进 Th1类细胞因子表达 ,抑制 Th2类细胞因子的分泌。
OBJECTIVE: To observe the effect of recombinant adenovirus gene-modified fetal hepatocyte (Adm IL-18 / BNL-CL2) expressing m IL-18 on immune function in normal mice after spleen transplantation. Methods: The mice in experimental group were transplanted with Adm IL-18 / BNL.CL2 by spleen transplantation, LacZ virus control group (Ad-Lac Z / BNL.CL2), BNL-CL2 cell control group and blank control group. Two weeks later sacrificed, serum was collected, peritoneal macrophages, splenic lymphocytes and liver homogenates were prepared and the total RNA of liver tissue was extracted. The serum, intraperitoneal Mφ and cytokines contents in liver homogenates of the mice in each group were detected by ELISA. Relative expression of cytokines in liver tissues was detected by semi-quantitative RT-PCR. The activity of Mφ and the activity of splenic NK cells in peritoneal cavity were determined by MTT assay and the proliferation activity of splenic lymphocytes by MTT assay. Results: The levels of IL - 18, IL - 2, IFN - γ and TNF - α in the serum, cell culture supernatant and liver homogenate of experimental mice were higher than those of other control groups, while the levels of IL - 4 and IL - 10 The results of semi-quantitative RT-PCR were consistent with those of ELISA. Meanwhile, the activity of Mφ and activity of splenic NK cells and splenic lymphocyte proliferation of experimental group were also significantly higher than that of control group. CONCLUSION: Adm IL - 18 can be effectively transfected into fetal hepatocytes and stably express m IL - 18. After transplanted with Adm IL - 18, fetal liver cells modified with Adm IL - 18 can significantly improve immune activity of liver and spleen, activate intraperitoneal Mφ , Promote the expression of Th1 cytokines and inhibit the secretion of Th2 cytokines.