紫草素联合肿瘤细胞裂解物刺激DC疫苗的抗肿瘤效果

Antitumor effects of DC vaccine loaded with shikonin combined with tumor cell lysate

目的:探讨紫草素(SK)联合肿瘤细胞裂解物(TCL)刺激的DC疫苗的抗肿瘤效果。方法:体外制备SK和TCL刺激的正常小鼠源的DC疫苗,流式细胞术检测DC表面CD80、CD86的荧光强度。流式细胞术检测与SK+TCL刺激的DC共培养后的正常小鼠脾T细胞中T-bet和RORγt的表达,ELISA检测共培养上清液中IFN-γ、IL-12P70和TNF-α的含量。建立Lewis肺癌3LL细胞荷瘤小鼠模型,分为PBS(1 mL)+TCL(5×10^(5)个细胞/100μL)、SK-L(1.25 mg/kg)+TCL、SK-M(2.5 mg/kg)+TCL、SK-H(5 mg/kg)+TCL、紫杉醇(PTX 2 mg/kg)+TCL疫苗组。疫苗接种结束后10 d,检测移植瘤体积和小鼠生存率,LDH法检测脾CTL的杀伤能力。结果:SK+TCL刺激的DC疫苗表达出高水平CD80、CD86(均P<0.01);DC与T细胞共培养液上清中IL-12P70、IFN-γ、TNF-α含量均显著上升(均P<0.01),T细胞中T-bet和RORγt的表达显著性上调(均P<0.01)。成功构建Lewis肺癌荷瘤小鼠模型,SK-H+TCL刺激的DC疫苗治疗显著延缓了的移植瘤生长并提高了小鼠的存活率,且可强烈诱导脾CTL的杀伤能力(均P<0.01)。结论:SK+TCL刺激的DC疫苗可激活DC至成熟状态,上调T细胞中T-bet和RORγt的表达,启动Th1效应细胞,SK体内治疗Lewis肺癌荷瘤小鼠具有良好的抗肿瘤效果。

Objective:To investigate the anti-tumor effect of dendritic cells(DCs)vaccines loaded with shikonin(SK)and tumor cell lysate(TCL).Methods:DC vaccines of normal mouse origin loaded with SK and TCL were prepared in vitro.Fluorescence intensity of CD80 and CD86 on the surface of DCs was detected by flow cytometry.The expression of T-bet and RORγt in normal mouse splenic T cells co-cultured DCs that stimulated by SK+TCL was determined by flow cytometry,and the contents of IFN-γ,IL-12P70,and TNF-αin the co-culture supernatant were detected using ELISA.A Lewis lung cancer 3LL cell-bearing mouse model was established,and the mice were randomized into PBS+TCL group(PBS[1 mL]+TCL[5×10^(5)cell/100μL]),SK-L+TCL group(low SK concentration[1.25 mg/kg]+TCL),SK-M+TCL group(medium SK concentration[2.5 mg/kg]+TCL),and SK-H+TCL group(high SK concentration[5 mg/kg]+TCL),and paclitaxel(PTX)+TCL vaccine group(PTX[2 mg/kg]+TCL).Ten days after the end of vaccination,the solid tumor volume and survival rate of the mice were observed,and the killing capacity of splenic cytotoxic T lymphocytes(CTLs)was assessed by LDH assay.Results:DC vaccines loaded with SK+TCL showed high levels of CD80 and CD86 expression(both P<0.01).The levels of IL-12P70,IFN-γ,and TNF-αin the DC-T cell co-culture supernatants were significantly increased(all P<0.01),and the expression of T-bet and RORγt(both P<0.01)in T cells were significantly elevated.A successful Lewis lung carcinoma mouse model was established,and the DC vaccines loaded with SK-H+TCL significantly delayed the growth of transplanted tumors and increased the mouse survival rate,while strongly inducing the cytotoxic activity of splenic CTLs(all P<0.01).Conclusion:The SK+TCL-loaded DC vaccine can activate DCs to a mature state,up-regulate the expression of T-bet and RORγt in T cells,and initiate Th1 effector cells.SK shows promising antitumor effects in the treatment of Lewis lung carcinoma in mice.