白介素-21基因转导人宫颈癌细胞产生T细胞和NK细胞依赖的抗肿瘤作用

Tumorigenesis of IL-21-transfected HeLa cells in SCID or hu-PBL-SCID mice

目的探讨白介素-21(interleukin-21,IL-21)基因抗肿瘤作用的机制。方法采用包含有人IL-21的质粒电转染至宫颈癌细胞(HeLa细胞),移植到用人外周淋巴细胞免疫重建和非免疫重建的重症联合免疫缺陷(severe combinedimmunodeficiency,SCID)小鼠体内,并用抗唾液酸神经节苷酯(anti-asialo GM1)删除小鼠的NK细胞,以及乳酸脱氢酶(lactate dehydrogenase,LDH)法检测荷瘤鼠的脾细胞毒性和酶联免疫吸附(enzyme linked immunosorbent assay,ELISA)双夹心法测定对靶细胞产生的IFN-γ。结果含有IL-21基因的宫颈癌细胞(HeLa-IL-21细胞)在免疫重建和非免疫重建小鼠中的生长完全被抑制,HeLa细胞在免疫重建鼠中的生长较非免疫重建小鼠滞后,肿瘤体积也明显小。删除NK细胞后暂时消除了HeLa-IL-21细胞对肿瘤的生长抑制,同时非免疫重建组消除肿瘤生长抑制较免疫重建组明显;免疫重建组荷HeLa-IL-21细胞小鼠脾细胞毒性为(16.55±4.53),高于非免疫重建组荷HeLa-IL-21细胞(8.32±2.12)(P<0.05)和荷HeLa细胞(3.42±1.56)(P<0.01),对靶细胞产生IFN-γ的量也大于免疫重建组荷HeLa细胞(5.12±2.34)(P<0.05)。结论在NK细胞缺乏的情况下,表达IL-21的HeLa细胞能产生T淋巴细胞依赖和不依赖的抗肿瘤作用。

Objective To explore the antitumor effect of interleukin 21（IL-21） after transfected into in HeLa cells in severe combined immunodeficiency（SCID） mice or human peripheral blood lymphocytes-SCID（hu-PBL-SCID） mice.Methods pcDNA4/HisMax-IL-21 was transfected into human cervical cancer cell line HeLa to produce HeLa-IL-21 cells.Totally 48 SCID mice were randomly and equally divided into SCID group（0.2 ml PBS,i.p.） and hu-PBL-SCID group（0.2 ml 2×10^7/ml hu-PBL,i.p.）.Then every group was further divided into 3 subgroups（n=6） to receive a subcutemeous implantation of HeLa,HeLa-vector or HeLa-IL-21 cells,and the rest 6 mice were intraperitoneally injected with anti-asialo GM1 50 mg/animal 1 d before and 4 d after HeLa-IL-21 cells transplantation for NK cell depletion.The growth of the tumor mass was observed.Flow cytometry was used to detect CD8-and DX5-positive cells in hu-PBL from hu-PBL-SCID mice,and spleen NK cells from SCID mice or hu-PBL-SCID mice.The secretion of IFN-γ and cytotoxicity of splenic cells from HeLa/HeLa-IL-21-bearing SCID mice or hu-PBL-SCID mice were detected with double sandwich ELISA assay and LDH assay respectively.Results Growth of HeLa-IL-21 tumors was significantly suppressed compared with that of HeLa cells,with a slow development and smaller volume.But this growth suppression was not observed in NK cell depletion groups,with SCID group more severe than hu-PBL-SCID group.hu-PBL-SCID mice bearing HeLa-IL-21 cells（16.55±4.53）had a higher toxicity in spleen cells than those SCID mice bearing HeLa-IL-21 cells（8.32±2.12） or HeLa cells（3.42±1.56）（P〈0.01）,and had larger production of IFN-γ than ofhu-PBL-SCID mice bearing splenocyts from hu-PBL-SCID bearing HeLa cells（5.12±2.34）（P〈0.05）.Conclusion Expression of IL-21 in HeLa cells can produce T cell dependent and-independent antitumor effects in an NK cell-defective condition.