中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2015年
6期
1387-1390
,共4页
恶性黑色素瘤%Toll样受体4%叉头样转录因子%免疫逃逸
噁性黑色素瘤%Toll樣受體4%扠頭樣轉錄因子%免疫逃逸
악성흑색소류%Toll양수체4%차두양전록인자%면역도일
Malignant melanoma%Toll-like receptor 4%Forkhead/winged helix transcription factor P3%Immune evasion
目的 观察Toll样受体4(TLR4)在皮肤恶性黑色素瘤(CMM)中的表达及对叉状头/翅膀状螺旋转录因子(Foxp3)表达的影响,探讨其在肿瘤免疫逃逸中的作用.方法 采用免疫组织化学链霉菌抗生物素蛋白-过氧化物酶(SP)法检测TLR4和Foxp3在42例CMM、36例色素痣及30例正常皮肤的表达.采用反转录-聚合酶链反应(RT-PCR)法和Western blot法检测脂多糖(LPS)活化黑色素瘤A375细胞TLR4后Foxp3 mRNA和蛋白水平的表达.设计并构建TLR4小干扰RNA(siRNA)慢病毒表达载体转染A375细胞,检测TLR4基因沉默对Foxp3 mRNA和蛋白表达的影响.细胞计数试剂盒(CCK-8)实验检测CD4+ CD25-T淋巴细胞增殖能力.结果 在CMM、色素痣及正常皮肤组织,TLR4、Foxp3的阳性表达率分别为66.7％(28/42)、41.7％ (15/36)、16.7％(5/30)和95.2％ (40/42)、11.1％ (4/36)、0％ (0/30),TLR4、Foxp3在CMM组织的阳性表达率显著高于色素痣及正常皮肤组织,差异有统计学意义(P＜0.05),且两者表达呈显著正相关(r =0.883,P＜0.05).LPS活化A375细胞TLR4可显著上调Foxp3 mRNA和蛋白水平的表达,TLR4基因沉默后再用LPS刺激A375细胞,Foxp3表达较对照组明显降低,且肿瘤细胞对CD4+ CD25-T淋巴细胞增殖的抑制作用明显减弱(P＜0.05).结论 TLR4信号通路激活可能通过上调Foxp3表达参与恶性黑色素瘤的免疫逃逸.
目的 觀察Toll樣受體4(TLR4)在皮膚噁性黑色素瘤(CMM)中的錶達及對扠狀頭/翅膀狀螺鏇轉錄因子(Foxp3)錶達的影響,探討其在腫瘤免疫逃逸中的作用.方法 採用免疫組織化學鏈黴菌抗生物素蛋白-過氧化物酶(SP)法檢測TLR4和Foxp3在42例CMM、36例色素痣及30例正常皮膚的錶達.採用反轉錄-聚閤酶鏈反應(RT-PCR)法和Western blot法檢測脂多糖(LPS)活化黑色素瘤A375細胞TLR4後Foxp3 mRNA和蛋白水平的錶達.設計併構建TLR4小榦擾RNA(siRNA)慢病毒錶達載體轉染A375細胞,檢測TLR4基因沉默對Foxp3 mRNA和蛋白錶達的影響.細胞計數試劑盒(CCK-8)實驗檢測CD4+ CD25-T淋巴細胞增殖能力.結果 在CMM、色素痣及正常皮膚組織,TLR4、Foxp3的暘性錶達率分彆為66.7％(28/42)、41.7％ (15/36)、16.7％(5/30)和95.2％ (40/42)、11.1％ (4/36)、0％ (0/30),TLR4、Foxp3在CMM組織的暘性錶達率顯著高于色素痣及正常皮膚組織,差異有統計學意義(P＜0.05),且兩者錶達呈顯著正相關(r =0.883,P＜0.05).LPS活化A375細胞TLR4可顯著上調Foxp3 mRNA和蛋白水平的錶達,TLR4基因沉默後再用LPS刺激A375細胞,Foxp3錶達較對照組明顯降低,且腫瘤細胞對CD4+ CD25-T淋巴細胞增殖的抑製作用明顯減弱(P＜0.05).結論 TLR4信號通路激活可能通過上調Foxp3錶達參與噁性黑色素瘤的免疫逃逸.
목적 관찰Toll양수체4(TLR4)재피부악성흑색소류(CMM)중적표체급대차상두/시방상라선전록인자(Foxp3)표체적영향,탐토기재종류면역도일중적작용.방법 채용면역조직화학련매균항생물소단백-과양화물매(SP)법검측TLR4화Foxp3재42례CMM、36례색소지급30례정상피부적표체.채용반전록-취합매련반응(RT-PCR)법화Western blot법검측지다당(LPS)활화흑색소류A375세포TLR4후Foxp3 mRNA화단백수평적표체.설계병구건TLR4소간우RNA(siRNA)만병독표체재체전염A375세포,검측TLR4기인침묵대Foxp3 mRNA화단백표체적영향.세포계수시제합(CCK-8)실험검측CD4+ CD25-T림파세포증식능력.결과 재CMM、색소지급정상피부조직,TLR4、Foxp3적양성표체솔분별위66.7％(28/42)、41.7％ (15/36)、16.7％(5/30)화95.2％ (40/42)、11.1％ (4/36)、0％ (0/30),TLR4、Foxp3재CMM조직적양성표체솔현저고우색소지급정상피부조직,차이유통계학의의(P＜0.05),차량자표체정현저정상관(r =0.883,P＜0.05).LPS활화A375세포TLR4가현저상조Foxp3 mRNA화단백수평적표체,TLR4기인침묵후재용LPS자격A375세포,Foxp3표체교대조조명현강저,차종류세포대CD4+ CD25-T림파세포증식적억제작용명현감약(P＜0.05).결론 TLR4신호통로격활가능통과상조Foxp3표체삼여악성흑색소류적면역도일.
Objective To explore the relationship between the expression of Toll-like receptor 4 (TLR4) and forkhead/winged helix transcription factor P3 (Foxp3) in cutaneous malignant melanoma (CMM) and the role of TLR4 in immune evasion.Methods Immunohistochemistry was used to detect the expression of TLR4 and Foxp3 in 42 cases of CMM,36 cases of pigmented nevus and 30 cases of normal skin specimens.Lipopolysaccharide (LPS) was employed as exogenous ligands to activate TLR4 in A375 cells.The mRNA and protein levels of Foxp3 were detected by reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting.Specific TLR4 small interfering RNA (siRNA) was synthesized and transfected into A375 cells by lentivirus.The expression of Foxp3 and the proliferation of CD4 + CD25-T lymphocytes were examined.Results The positive expression rate of TLR4 and Foxp3 in CMM (66.7％ and 95.2％) was significantly higher than in the pigmented nevus (41.7％ and 11.1％) and normal skin (16.7％ and 0％) (P ＜0.05),and they were positively correlated (r =0.883,P＜0.05).The expression of Foxp3 at mRNA and protein levels was significantly increased by LPS stimulation.siRNA could suppress the expression level of Foxp3 and down-regulate the inhibitory ability of tumor cells on the proliferation of CD4 + CD25-T lymphocytes (P ＜ 0.05).Conclusion TLR4 signaling pathway may be involved in CMM immune evasion process by upregulating Foxp3 expression.
