Gu Guangli, Lv Xiaodan, Liu Gengfeng, Zeng Ruizhi, Li Shiquan, Chen Lan, Liang Zhaoliang, Wang Huiqin, Lu Fei, Zhan Lingling, Lv Xiaoping
Department of Gastroenterology, The First Affiliated Hospital of Guangxi Medical University, Nanning, China.
Department of Clinical Experimental Medicine, The First Affiliated Hospital of Guangxi Medical University, Nanning, China.
Front Pharmacol. 2021 Oct 11;12:734040. doi: 10.3389/fphar.2021.734040. eCollection 2021.
To investigate the immunological mechanism of bone marrow-derived mesenchymal stem cells (BM-MSCs) in inflammatory bowel disease (IBD). Mice with 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis were intraperitoneally injected with phosphate-buffered saline, BM-MSCs, BM-MSCs with tumor necrosis factor-induced protein 6 (Tnfaip6) knockdown mediated by RNA interference recombinant adenovirus, and BM-MSCs-infected with control adenovirus or recombinant mouse Tnfaip6. The disease activity index, weight loss, and histological scores were recorded. Serum levels of Tnfaip6 and pro- and anti-inflammatory cytokines, including interleukin (IL)-21, tumor necrosis factor-alpha (TNF-α), IL-10 were measured by enzyme-linked immunosorbent assay. The relative expression levels of these cytokines, B-cell lymphoma 6 (BCL-6) and fork-like transcription factor p3 (Foxp3) in the colon were determined by real-time quantitative PCR (RT-qPCR). BCL-6 and Foxp3 are the master regulators of follicular helper T cells (Tfh) and follicular regulatory T cells (Tfr), respectively. The infiltration of Tfh and Tfr in mesenteric lymph nodes (MLNs) and spleens was analyzed by flow cytometry. Compared to the normal control group, the expression levels of BCL-6 and IL-21 in the colon, Tfh infiltration, and ratios of Tfh/Tfr in the MLNs and spleen, and the serum concentrations of IL-21 and TNF-α increased significantly in the colitis model group ( < 0.05). Intraperitoneal injection of BM-MSCs or Tnfaip6 ameliorated weight loss and clinical and histological severity of colitis, downregulated the expression of BCL-6, IL-21, and TNF-α, upregulated the expression of Foxp3, IL-10, and Tnfaip6 ( < 0.05), increased Tfr and reduced the infiltration of Tfh in the MLNs and spleen, and downregulated the Tfh/Tfr ratio ( < 0.05). On the other hand, BM-MSCs lost the therapeutic effect and immune regulatory functions on Tfh and Tfr after Tnfaip6 knockdown. Tfh increase in the inflamed colon, Tfh decrease and Tfr increase during the colitis remission phase, and the imbalance of the Tfh/Tfr ratio is closely related to the progression of IBD. Tnfaip6 secreted by BM-MSCs alleviates IBD by inhibiting Tfh differentiation, promoting Tfr differentiation, and improving the imbalance of Tfh/Tfr in mice.
探讨骨髓间充质干细胞（BM-MSCs）在炎症性肠病（IBD）中的免疫机制。将2,4,6-三硝基苯磺酸（TNBS）诱导的结肠炎小鼠腹腔注射磷酸盐缓冲盐水、BM-MSCs、经RNA干扰重组腺病毒介导的肿瘤坏死因子诱导蛋白6（Tnfaip6）敲低的BM-MSCs，以及感染对照腺病毒或重组小鼠Tnfaip6的BM-MSCs。记录疾病活动指数、体重减轻和组织学评分。通过酶联免疫吸附测定法测量血清中Tnfaip6以及促炎和抗炎细胞因子的水平，包括白细胞介素（IL）-21、肿瘤坏死因子-α（TNF-α）、IL-10。通过实时定量PCR（RT-qPCR）测定结肠中这些细胞因子、B细胞淋巴瘤6（BCL-6）和叉状转录因子p3（Foxp3）的相对表达水平。BCL-6和Foxp3分别是滤泡辅助性T细胞（Tfh）和滤泡调节性T细胞（Tfr）的主要调节因子。通过流式细胞术分析肠系膜淋巴结（MLNs）和脾脏中Tfh和Tfr的浸润情况。与正常对照组相比，结肠炎模型组结肠中BCL-6和IL-21的表达水平、MLNs和脾脏中Tfh浸润以及Tfh/Tfr比值以及血清中IL-21和TNF-α浓度显著升高（<0.05）。腹腔注射BM-MSCs或Tnfaip6可改善体重减轻以及结肠炎的临床和组织学严重程度，下调BCL-6、IL-21和TNF-α的表达，上调Foxp3、IL-10和Tnfaip6的表达（<0.05），增加Tfr并减少MLNs和脾脏中Tfh的浸润，下调Tfh/Tfr比值（<0.05）。另一方面，Tnfaip6敲低后BM-MSCs失去了对Tfh和Tfr的治疗作用和免疫调节功能。在炎症性结肠中Tfh增加，在结肠炎缓解期Tfh减少而Tfr增加，并且Tfh/Tfr比值失衡与IBD的进展密切相关。BM-MSCs分泌的Tnfaip6通过抑制小鼠Tfh分化、促进Tfr分化以及改善Tfh/Tfr失衡来减轻IBD。
