Department of Respiratory and Critical Care Medicine, Shanxi Academy of Medical Sciences, Shanxi Bethune Hospital, Taiyuan, Shanxi 030032, P.R. China.
Department of Respiratory and Critical Care Medicine, The Second Clinical Medical College, Shanxi Medical University, Taiyuan, Shanxi 030009, P.R. China.
Mol Med Rep. 2020 May;21(5):2267-2275. doi: 10.3892/mmr.2020.11027. Epub 2020 Mar 13.
Adoptive transfer of bone marrow‑derived mesenchymal stem cells (BMSCs) significantly alleviates smoking‑induced chronic obstructive pulmonary disease (COPD) in rats. Considering the critical roles of T cells during COPD development, the present study aimed to further identify the molecular mechanisms underlying the antiproliferative effect of BMSCs on splenic T cells isolated from rats following chronic exposure to nicotine. Splenic T cells were co‑cultured with rat BMSCs at various ratios and subsequently, T‑cell proliferation was measured using the Cell Counting Kit‑8 assay. The effects of the inducible nitric oxide synthase (iNOS) inhibitor N‑nitro‑L‑arginine methylester (L‑NAME) and short hairpin (sh)RNA‑lentivirus‑mediated knockdown of iNOS in BMSCs on T‑cell proliferation were evaluated. The expression levels of iNOS and STAT5 phosphorylation in BMSCs and T cells, respectively, were assessed by reverse transcription‑quantitative PCR and western blotting. A higher ratio of BMSCs to T cells resulted in increased inhibition of T‑cell proliferation; therefore, the ratio of 1:20 was selected for further in vitro experiments. At a dose of 5 µM, L‑NAME displayed the strongest ability to reverse the antiproliferative effects of BMSCs in the co‑culture system. Both L‑NAME treatment and shRNA‑mediated knockdown of iNOS expression significantly decreased the suppressive effects of BMSCs, downregulated iNOS expression at the mRNA and protein levels in BMSCs, and enhanced STAT5 phosphorylation in T cells. BMSCs inhibited the proliferation of nicotine‑exposed T cells, which was associated with iNOS expression in BMSCs and decreased STAT5 phosphorylation in T cells. The present study indicated the potential mechanisms underlying the beneficial effects of BMSC infusion in patients with chronic smoking‑induced COPD and emphysema.
骨髓间充质干细胞（BMSCs）的过继转移可显著减轻大鼠吸烟诱导的慢性阻塞性肺疾病（COPD）。鉴于 T 细胞在 COPD 发展过程中的关键作用，本研究旨在进一步鉴定 BMSCs 对慢性尼古丁暴露大鼠脾 T 细胞增殖的抑制作用的分子机制。将脾 T 细胞与大鼠 BMSCs 以不同比例共培养，然后使用细胞计数试剂盒-8 检测 T 细胞增殖。评估诱导型一氧化氮合酶（iNOS）抑制剂 N-硝基-L-精氨酸甲酯（L-NAME）和 BMSCs 短发夹（sh）RNA-慢病毒介导的 iNOS 敲低对 T 细胞增殖的影响。通过逆转录-定量 PCR 和 Western blot 分别评估 BMSCs 和 T 细胞中 iNOS 和 STAT5 磷酸化的表达水平。BMSCs 与 T 细胞的更高比例导致 T 细胞增殖抑制增加；因此，选择 1:20 的比例用于进一步的体外实验。在 5 μM 的剂量下，L-NAME 显示出在共培养系统中逆转 BMSCs 增殖抑制作用的最强能力。L-NAME 处理和 shRNA 介导的 iNOS 表达下调均显著降低了 BMSCs 的抑制作用，下调了 BMSCs 中 iNOS 的 mRNA 和蛋白表达水平，并增强了 T 细胞中 STAT5 的磷酸化。BMSCs 抑制了尼古丁暴露的 T 细胞的增殖，这与 BMSCs 中的 iNOS 表达和 T 细胞中 STAT5 磷酸化减少有关。本研究表明了 BMSC 输注在慢性吸烟诱导的 COPD 和肺气肿患者中的有益作用的潜在机制。
