Np73α基因转染树突状细胞抗乳腺癌细胞的免疫效应

摘要

目的：研究ΔNp73α基因转染树突状细胞（ DC）诱导的特异性抗乳腺癌免疫效应。方法：人脐带血细胞经GM-CSF、IL-4、TNF-α等细胞因子诱导培养DC，流式细胞仪（FCM）检测DC成熟前后CD1a、CD83的表达变化情况。脂质体法将pcDNA-HA／ΔNp73α转染至DC，经Western blot检测转染情况。转染DC与自体T细胞共培养诱导特异性CTL。 MTT法测定T细胞增殖能力；ELISA法检测IFN-γ的分泌水平；LDH释放法检测T细胞对乳腺癌细胞MDA-MB-231的杀伤作用。结果：DC诱导成熟后，CD1a表达约占56％，CD83约占74％，与未成熟DC（ CD1a 19％，CD8313％）比较，差异有显著统计学意义（ P＜0.01）。 Western blot检测到DC-ΔNp73α组有一特异条带表达。 DC-ΔNp73α组诱导的特异性CTL对MDA-MB-231杀伤作用高于DC组（ P＜0.05），而且刺激T细胞增殖能力增强，分泌IFN-γ的水平升高，与空载体DC-pcDNA组及DC组比较有显著统计学意义（P＜0.01）。结论：以ΔNp73α转染DC制备的DC疫苗，具有显著诱导CTL杀伤乳腺癌细胞的作用。%Objective:To research the imumunological effect of dendritic cell transfected by recombinant plasmid of ΔNp73αon breast cancer.Methods:Cultivate dendritic cells by cytokines( GM-CSF,IL-4,TNF-α) from human umbilical cord blood in vitro, CD1a,CD83 were detected by flow cytometry.Recombinant plasmid pcDNA-HA/ΔNp73αwas used to transfected DCs,the transfection was detected by Western blot.Transfected DCs cocultivate with T cell induce specific cytotoxic lymphocytes( CTL).Proliferation ability of T cell was detected by MTT methods.IFN-γin the culture supernatant of the transfected DCs were detected by ELISA.The killing effect on breast cancer MDA-MB-231 was detected by LDH method.Results:The expression of CD1a(56%),CD83(74%)in mature DC was higher than immature DC(CD1a 19%,CD83 13%)(P<0.01).ΔNp73αtransfected group had a special band.The killing effect of specific CTL induced byΔNp73α-DC and T cells cocultivation on breast cancer cell MDA-MB-231 was statistic stronger than DC group(P<0.05),the proliferation of T cell and secretion level of IFN-γcompared with DC and pcDNA group were statistically significant(P<0.01).Conclusion:The CTL induced byΔNp73αtransfected DC vaccine has significant effect on killing breast cancer.