Objectives: The differentiation of CD4+ T cells is regulated by a complex and fine signaling pathway composed of many molecules during immune response, and the molecular mechanism for regulating T-bet expression is unclear. Mediator complex subunit 1 (Med1) can combine with a variety of co-factors to regulate gene transcription, promote cell proliferation and survival, and affect invariant natural killer T cell (iNKT) development. This study aims to investigate the effect of Med1 on T cell development and CD4+ T cell differentiation in immune response.
Methods: Mice with T cell-specific knockout of Med1 gene (Med1F/FCD4cre+, KO) were constructed and verified. The percentage and number of CD4+ and CD8+ T cells in thymus, spleen, and lymph nodes of KO mice and control (Con) mice (Med1F/FCD4cre-) were detected by flow cytometry. After 8 days of infection with lymphocytic choriomeningitis virus (LCMV), the percentage and number of CD4+ T cells or antigen-specific (GP66+) CD4+ T cells, the percentage and number of Th1 cells (Ly6c+PSGL1+) in CD4+ T cells or antigen-specific CD4+ T cells were examined in the spleen of mice. Moreover, the fluorescence intensity of T-bet in CD4+ T cells or antigen-specific CD4+ T cells was analyzed.
Results: Compared with the Con group, the percentage and number of CD4+ T cells and CD8+ T cells in the thymus, CD4+ T cells in the spleen and lymph nodes of the KO group showed no significant differences (all P>0.05), but the percentage and number of CD8+ T cells in the spleen and lymph nodes of the KO group were diminished significantly (all P<0.05). After 8 days of infection with LCMV, there was no significant difference in the percentage and number of CD4+ T cells or antigen-specific CD4+ T cells in the spleen between the KO group and the Con group (all P>0.05), while in comparison with the Con group, the percentage and number of Th1 cells in CD4+ T cells or antigen-specific CD4+ T cells, and the expression of T-bet in CD4+ T cells or antigen-specific CD4+ T cells were significantly reduced in the spleen of the KO group (all P<0.05).
Conclusions: Specific knockout of Med1 in T cells does not affect the development of CD4+ and CD8+ T cells in the thymus, but does affect the maintenance of peripheral CD8+ T cells. In the immune response, Med1 gene deletion affects the expression of transcription factor T-bet, which in turn to reduce Th1 cell differentiation.
目的: CD4+ T细胞在免疫应答过程中的分化受到由众多分子组成的复杂而精细的信号通路的调控，且调控T-bet表达的分子机制并未阐明。中介体复合物亚基1(mediator complex subunit 1，Med1)与多种辅助因子结合调节基因转录，促进细胞存活和增殖，影响恒定自然杀伤T细胞(invariant natural killer T cell，iNKT)的发育。本研究旨在探讨Med1对T细胞发育及CD4+ T细胞在免疫应答中分化的影响。方法: 构建T细胞特异性敲除Med1基因(KO)小鼠(Med1F/FCD4cre+)并对其进行验证。采用流式细胞术检测KO组和对照(Con)组(Med1F/FCD4cre-)小鼠胸腺、脾和淋巴结中CD4+和CD8+ T细胞的百分比及数目；用淋巴细胞脉络丛脑膜炎病毒(lymphocytic choriomeningitis virus，LCMV)感染小鼠8 d后，检测小鼠脾中CD4+ T细胞及抗原特异性(GP66+)CD4+ T细胞的百分比及数目、Th1细胞(Ly6c+PSGL1+)占CD4+ T细胞及抗原特异性CD4+ T细胞的百分比及数目、CD4+ T细胞及抗原特异性CD4+ T细胞中T-bet的荧光强度。结果: KO组与Con组小鼠胸腺中CD4+和CD8+ T细胞的百分比及数目、脾和淋巴结中CD4+ T细胞的百分比及数目差异均无统计学意义(均P>0.05)，但KO组脾和淋巴结中CD8+ T细胞的百分比及数目较Con组均显著降低(均P<0.05)。感染LCMV后，KO组与Con组小鼠脾中CD4+ T细胞及抗原特异性CD4+ T细胞的百分比及数目差异均无统计学意义(均P>0.05)，但KO组Th1细胞占CD4+ T细胞及抗原特异性CD4+ T细胞的百分比及数目、CD4+ T细胞及抗原特异性CD4+ T细胞中T-bet的表达较Con组均显著降低(均P<0.05)。结论: T细胞中特异性敲除Med1基因不影响胸腺中CD4+和CD8+ T细胞的发育，但是影响外周CD8+ T细胞的维持。在免疫应答中Med1基因缺失影响转录因子T-bet的表达，最终导致CD4+ T细胞向Th1细胞分化减少。.
Keywords: T cell development; T-bet; Th1 cell; immune response; mediator complex subunit 1.