【摘要】：目的研究多種惡性膠質(zhì)瘤相關(guān)抗原表位肽激活的樹突狀細(xì)胞(GDC)致敏的細(xì)胞毒性T淋巴細(xì)胞(GDC-CTL)對人腦膠質(zhì)瘤細(xì)胞系U87的體外細(xì)胞毒活性,以及其對BALB/c nude裸小鼠神經(jīng)膠質(zhì)瘤模型的體內(nèi)抗腫瘤作用。方法制備多種惡性膠質(zhì)瘤相關(guān)抗原致敏的GDC和GDC-CTL細(xì)胞,用流式細(xì)胞儀對細(xì)胞表型進(jìn)行分析。設(shè)立不同的效靶比(5∶1,10∶1,20∶1),用CCK8法測定GDC-CTL細(xì)胞體外對U87細(xì)胞的殺傷率,用酶聯(lián)免疫吸附實(shí)驗(yàn)(ELISA)法檢測GDC-CTL與U87細(xì)胞共培養(yǎng)48 h后培養(yǎng)上清液中γ干擾素(IFN-γ)水平,設(shè)立未經(jīng)GDC激活的T淋巴細(xì)胞為對照組。用BALB/c Nude裸小鼠U87細(xì)胞皮下腫瘤模型觀察GDC-CTL細(xì)胞體內(nèi)抑瘤作用,實(shí)驗(yàn)分為空白對照組、模型組、靜脈治療組和局部治療組,空白對照組于背部皮下注射0.9%Na Cl,其他3組于背部皮下注射1×107U 87細(xì)胞,在腫瘤長徑達(dá)3 mm后模型組于腫瘤局部注射0.9%Na Cl,靜脈治療組和局部治療組分別于尾靜脈和腫瘤局部注射2×107GDC-CTL,每周注射3次,共2周,注射體積均為0.2 m L。觀察4組小鼠的腫瘤體積生長情況,取腫瘤組織進(jìn)行病理學(xué)檢測。結(jié)果 GDC高表達(dá)CD83、CD1a和HLA-DR,表明成功誘導(dǎo)GDC成熟。CD3+T淋巴細(xì)胞比例為93.00%,CD3+CD8+T淋巴細(xì)胞為69.00%,表明GDC-CTL成功激活。效靶比為5∶1,10∶1,20∶1時:GDC-CTL組和對照組對U87細(xì)胞的殺傷率分別為(24.35±1.12)%vs(15.21±0.91)%,(38.57±2.10)%vs(23.35±1.30)%,(59.44±3.79)%vs(35.23±2.33)%;GDC-CTL組和對照組與U87細(xì)胞共培養(yǎng)48 h后培養(yǎng)上清液中IFN-γ水平分別為(405.36±27.65)vs(371.11±23.23)pg·m L~(-1),(1509.22±97.16)vs(913.54±48.35)pg·m L~(-1),(2429.57±183.18)vs(1814.97±123.24)pg·m L~(-1),在效靶比為10∶1和20∶1時,2組間差異有統(tǒng)計學(xué)意義(P0.05)。靜脈治療組和局部治療組對BALB/c nude裸小鼠皮下神經(jīng)膠質(zhì)瘤的生長抑制率分別為34.83%和45.37%,與模型組的100.00%比較,差異均有統(tǒng)計學(xué)意義(P0.05),與模型組相比,局部治療組和靜脈治療組均可見腫瘤細(xì)胞顯著減少。結(jié)論 GDC-CTL是一種高效的免疫細(xì)胞,具有較強(qiáng)的體內(nèi)外抑制膠質(zhì)瘤生長的作用。
[Abstract]:Objective to study the cytotoxic activity of cytotoxic T lymphocytes (GDC-CTL) sensitized by dendritic cells (GDC) activated by epitope peptides associated with malignant glioma on human glioma cell line U87 in vitro. And its anti-tumor effect on BALB/c nude mouse glioma model in vivo. Methods GDC and GDC-CTL cells sensitized with various malignant glioma associated antigens were prepared and their phenotypes were analyzed by flow cytometry. Different effective / target ratios (5: 1 10: 1 20: 1) were established. The cytotoxicity of GDC-CTL cells to U87 cells was measured by CCK8 assay. The levels of IFN- 緯 in the supernatant of GDC-CTL and U87 cells co-cultured for 48 h were detected by enzyme-linked immunosorbent assay (ELISA). T lymphocytes without GDC activation were established as control group. BALB/c Nude nude mice U87 cell tumor model was used to observe the anti-tumor effect of GDC-CTL cells in vivo. The experiment was divided into three groups: blank control group, model group, intravenous treatment group and local treatment group. 0.9%Na Cl1 was injected subcutaneously into the back of the control group, and 1 脳 107 U87 cells were injected into the back in the other three groups. After the tumor reached 3 mm in length, the model group was injected with 0.9%Na Cl locally, and the intravenous treatment group and the local treatment group were injected with 2 脳 107 GDC-CTL in the tail vein and tumor respectively, three times a week for 2 weeks. The injection volume was 0.2mL. Tumor volume growth was observed in 4 groups of mice and tumor tissues were taken for pathological examination. Results the overexpression of CD83a CD1a and HLA-DR1 by GDC indicated that the ratio of T lymphocytes induced to GDC maturation. CD3 T lymphocytes was 93.00 and CD3 CD8 T lymphocytes were 69.00, indicating that GDC-CTL was successfully activated. 鏁堥澏姣斾負(fù)5鈭，

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