中华肿瘤杂志
中華腫瘤雜誌
중화종류잡지
CHINESE JOURNAL OF ONCOLOGY
2014年
3期
177-182
,共6页
石立莹%陈骏%钟启平%耿鹏%何建民
石立瑩%陳駿%鐘啟平%耿鵬%何建民
석립형%진준%종계평%경붕%하건민
仙台病毒Tianjin株%缺损干扰颗粒%树突状细胞%T淋巴细胞%免疫%小鼠
仙檯病毒Tianjin株%缺損榦擾顆粒%樹突狀細胞%T淋巴細胞%免疫%小鼠
선태병독Tianjin주%결손간우과립%수돌상세포%T림파세포%면역%소서
Sendai virus Tianjin strain%Defective interfering particles%Dendritic cells%T-lymphocytes%Immunity%Mice
目的 通过小鼠结肠癌动物模型探讨仙台病毒Tianjin株缺损干扰颗粒(DIP)的抗肿瘤作用及其机制.方法 将CT26细胞悬液0.1 ml皮下注射Bal B/c小鼠背部,建立小鼠结肠癌模型,将小鼠随机分为Tianjin株DIP组(注射CT26后第4、7、10和13天,每天瘤内注射1次Tianjin株DIP 0.1 ml)和生理盐水对照组(瘤内注射生理盐水0.1ml),每组10只.通过测量肿瘤大小和观察小鼠存活率确定DIP的抑瘤作用.采用流式细胞仪和酶联免疫吸附(ELISA)法检测DIP在体外对小鼠树突状细胞(DCs)成熟及分泌白细胞介素6(IL-6)、干扰素α(IFN-α)和肿瘤坏死因子α(TNF-α)的促进作用.采用real time RT-PCR和免疫组化法检测DIP对小鼠肿瘤组织中CD4+、CD8+T细胞及CD1 1c+ DCs表达的影响.结果 至注射CT26后第22天,Tianjin株DIP组小鼠的肿瘤体积为(33.2 ±2.0)mm3,与生理盐水对照组[(2 376.0±130.8)mm3]比较,差异有统计学意义(P＜0.01).至注射CT26后第50天,Tianjin株DIP组小鼠的存活率为90.0％,明显高于生理盐水对照组(30.0％,P＜0.01).流式细胞仪检测显示,小鼠DCs表面CD40、CD80和CD86标志分子的表达率随病毒剂量的升高而升高,且Tianjin株DIP组和完整病毒组之间的差异无统计学意义(均P＞0.05).ELISA法检测显示,DIP能够促进小鼠DCs分泌IL-6、IFN-α和TNF-α,且呈剂量依赖性.real time RT-PCR检测结果显示,停止给药后24、48和120 h取得的小鼠肿瘤组织中CD4、CD8和CD11c mRNA的表达水平均有所升高,且在120h表达达高峰.免疫组化检测结果显示,Tianjin株DIP组肿瘤组织中CD4+T细胞、CD8+T细胞和CD1 1c+ DCs占总细胞的百分比分别为(21.60±1.49)％、(22.12±2.84)％和(23.05±2.91)％,而生理盐水对照组分别为(2.62±0.60)％、(4.05 ±0.12)％和(3.10±0.09)％,差异均有统计学意义(均P＜0.05).结论 Tianjin株DIP可以在小鼠体内发挥抗肿瘤作用,其作用机制与激活DCs和T细胞诱导的抗肿瘤免疫有关.
目的 通過小鼠結腸癌動物模型探討仙檯病毒Tianjin株缺損榦擾顆粒(DIP)的抗腫瘤作用及其機製.方法 將CT26細胞懸液0.1 ml皮下註射Bal B/c小鼠揹部,建立小鼠結腸癌模型,將小鼠隨機分為Tianjin株DIP組(註射CT26後第4、7、10和13天,每天瘤內註射1次Tianjin株DIP 0.1 ml)和生理鹽水對照組(瘤內註射生理鹽水0.1ml),每組10隻.通過測量腫瘤大小和觀察小鼠存活率確定DIP的抑瘤作用.採用流式細胞儀和酶聯免疫吸附(ELISA)法檢測DIP在體外對小鼠樹突狀細胞(DCs)成熟及分泌白細胞介素6(IL-6)、榦擾素α(IFN-α)和腫瘤壞死因子α(TNF-α)的促進作用.採用real time RT-PCR和免疫組化法檢測DIP對小鼠腫瘤組織中CD4+、CD8+T細胞及CD1 1c+ DCs錶達的影響.結果 至註射CT26後第22天,Tianjin株DIP組小鼠的腫瘤體積為(33.2 ±2.0)mm3,與生理鹽水對照組[(2 376.0±130.8)mm3]比較,差異有統計學意義(P＜0.01).至註射CT26後第50天,Tianjin株DIP組小鼠的存活率為90.0％,明顯高于生理鹽水對照組(30.0％,P＜0.01).流式細胞儀檢測顯示,小鼠DCs錶麵CD40、CD80和CD86標誌分子的錶達率隨病毒劑量的升高而升高,且Tianjin株DIP組和完整病毒組之間的差異無統計學意義(均P＞0.05).ELISA法檢測顯示,DIP能夠促進小鼠DCs分泌IL-6、IFN-α和TNF-α,且呈劑量依賴性.real time RT-PCR檢測結果顯示,停止給藥後24、48和120 h取得的小鼠腫瘤組織中CD4、CD8和CD11c mRNA的錶達水平均有所升高,且在120h錶達達高峰.免疫組化檢測結果顯示,Tianjin株DIP組腫瘤組織中CD4+T細胞、CD8+T細胞和CD1 1c+ DCs佔總細胞的百分比分彆為(21.60±1.49)％、(22.12±2.84)％和(23.05±2.91)％,而生理鹽水對照組分彆為(2.62±0.60)％、(4.05 ±0.12)％和(3.10±0.09)％,差異均有統計學意義(均P＜0.05).結論 Tianjin株DIP可以在小鼠體內髮揮抗腫瘤作用,其作用機製與激活DCs和T細胞誘導的抗腫瘤免疫有關.
목적 통과소서결장암동물모형탐토선태병독Tianjin주결손간우과립(DIP)적항종류작용급기궤제.방법 장CT26세포현액0.1 ml피하주사Bal B/c소서배부,건립소서결장암모형,장소서수궤분위Tianjin주DIP조(주사CT26후제4、7、10화13천,매천류내주사1차Tianjin주DIP 0.1 ml)화생리염수대조조(류내주사생리염수0.1ml),매조10지.통과측량종류대소화관찰소서존활솔학정DIP적억류작용.채용류식세포의화매련면역흡부(ELISA)법검측DIP재체외대소서수돌상세포(DCs)성숙급분비백세포개소6(IL-6)、간우소α(IFN-α)화종류배사인자α(TNF-α)적촉진작용.채용real time RT-PCR화면역조화법검측DIP대소서종류조직중CD4+、CD8+T세포급CD1 1c+ DCs표체적영향.결과 지주사CT26후제22천,Tianjin주DIP조소서적종류체적위(33.2 ±2.0)mm3,여생리염수대조조[(2 376.0±130.8)mm3]비교,차이유통계학의의(P＜0.01).지주사CT26후제50천,Tianjin주DIP조소서적존활솔위90.0％,명현고우생리염수대조조(30.0％,P＜0.01).류식세포의검측현시,소서DCs표면CD40、CD80화CD86표지분자적표체솔수병독제량적승고이승고,차Tianjin주DIP조화완정병독조지간적차이무통계학의의(균P＞0.05).ELISA법검측현시,DIP능구촉진소서DCs분비IL-6、IFN-α화TNF-α,차정제량의뢰성.real time RT-PCR검측결과현시,정지급약후24、48화120 h취득적소서종류조직중CD4、CD8화CD11c mRNA적표체수평균유소승고,차재120h표체체고봉.면역조화검측결과현시,Tianjin주DIP조종류조직중CD4+T세포、CD8+T세포화CD1 1c+ DCs점총세포적백분비분별위(21.60±1.49)％、(22.12±2.84)％화(23.05±2.91)％,이생리염수대조조분별위(2.62±0.60)％、(4.05 ±0.12)％화(3.10±0.09)％,차이균유통계학의의(균P＜0.05).결론 Tianjin주DIP가이재소서체내발휘항종류작용,기작용궤제여격활DCs화T세포유도적항종류면역유관.
Objective To explore the anti-tumor effect and its mechanism of Sendai virus Tianjin strain defective interfering particles (DIP) on mouse models of colon carcinoma.Methods CT26 cells (5 × 106/0.1 ml) were subcutaneously injected into the back of Bal B/c mice to establish murine colon carcinoma model.After the tumors reached 5 mm in diameter,the mice were randomly divided into Tianjin strain DIP group and saline control group.The former was intratumorally injected with Tianjin strain DIP (0.1 ml) once a day on day 4,7,10 and 13 after CT26 cell inoculation.The latter was intratumorally injected with the same volume of saline.Tumor volume and survival rate of the mice were calculated to confirm the anti-tumor effect of DIP.Flow cytometry and ELISA were used to examine the maturation and release of cytokines IL-6,IFN-α and TNF-α from murine myeloid dendritic cells (DCs) induced by Tianjin strain DIP.Moreover,real-time RT-PCR and immunohistochemistry were performed to identify whether the Tianjin strain DIP could induce infiltration of CD11c + DCs,CD4 + and CD8 + T cells in the tumors.Results On day 22 after CT26 cell inoculation,the average tumor volume of the Tianjin strain DIP group was (33.2 ± 2.0) mm3,significantly smaller than that of the control group [(2 376.0 ± 130.8) mm3,P ＜ 0.01].On day 50 after CT26 cell inoculation,the survival rate of mice was 90.0％ in the Tianjin strain DIP group,much higher than that of the control group (30.0％,P ＜ 0.01).Flow cytometry analysis showed that the expression of markers of DCs maturation,including CD40,CD80 and CD86,was dose-dependently increased by DIP or intact virus.No statistically significant difference was found betweent the DIP and intact virus groups.ELISA results showed that DIP could stimulate the secretion of IL-6,IFN-α and TNF-α from mouse DCs.The secretion of all of the cytokines was dose-dependently increased by DIP or intact virus.Real-time RT-PCR revealed that the expression of CD4,CD8 and CD11 c mRNAs was increased in tumors treated with DIP compared with that of the saline group at all time points.Moreover,the expression level of all of them remained maximal at 120 h after the last treatment.Immunohistochemical staining revealed that the ratios of CD4+,CD8+ T cells or CD11c+ DCs to total cells were (21.60±1.49)％,(22.12 ± 2.84) ％ and (23.05 ± 2.91)％,respectively,in the DIP-treated tumors.In the tumors treated by saline,the ratios were (2.62 ± 0.60) ％,(4.05 ± 0.12) ％ and (3.10 ± 0.09) ％,respectively.The difference between experimental group and control group had statistical significance.Conclusions Tianjin strain DIP may exert anti-tumor effect on tumor-bearing mice.The mechanism is related with the antitumor immunity induced by DCs and T cells.
