Department of General Thoracic Surgery, Osaka City University, 1-4-3 Asahi-machi, Abeno-ku, Osaka, 545-8585, Japan.
Department of Food Science and Nutrition Health, Kyoto Prefectural University, Kyoto, Japan.
Inflammopharmacology. 2018 Apr;26(2):435-445. doi: 10.1007/s10787-017-0371-y. Epub 2017 Jul 3.
Alveolar macrophages are key contributors to both the promotion and resolution of inflammation in the lung and are categorized into pro-inflammatory (M1) and anti-inflammatory (M2) phenotypes. The change in M1/M2 balance has been reported in various pulmonary diseases and is a target for therapeutic intervention. The aim of this study was to assess the modulation of M1/M2 phenotype in alveolar macrophages by water-soluble carbon monoxide-releasing molecule-3 (CORM-3). Rat alveolar macrophages (AM) (NR8383) in culture were stimulated with LPS (5 ng/ml)/IFN-γ (10 U/ml) or IL-4 (10 ng/ml)/IL-13 (10 ng/ml) to induce M1 and M2 phenotypes, respectively. Expression of M1 phenotype markers, iNOS and TNF-α, and M2 phenotype markers, CD206 and Ym-1, was assessed by western blotting after 1, 3, 6, or 24 h in the absence or presence of CORM-3 (0.15 mM) treatment. Inactive CORM-3 (iCORM-3) was used as a control. Treatment of naïve (unstimulated) AM with CORM-3 promoted progression of the M2 phenotype as evidenced by the increased expression of CD206 (at 1 h; 1.8-fold) and Ym-1 (at 3 h; 1.9-fold), respectively. Surprisingly, CORM-3 treatment also upregulated the expression of iNOS protein as assessed 6 h following stimulation of AM with CORM-3 (2.6-fold). On the contrary, CORM-3 effectively reduced LPS/IFN-γ-induced expression of iNOS protein (0.6-fold); however, it had no effect on TNF-α expression. Finally, CORM-3 acutely (1-3 h) upregulated CD206 (1.4-fold) and Ym-1 (1.6-fold) levels in IL-4-/IL-13-treated (M2-stimulus) macrophages. These findings indicate that CORM-3 modulates macrophage M1 and M2 phenotypes in vitro with respect to continuous suppression of iNOS expression in M1-polarized macrophages and transient (early-phase) upregulation of CD206 and Ym-1 proteins in M2-polarized macrophages.
肺泡巨噬细胞是肺部炎症发生和消退的关键贡献者，可分为促炎（M1）和抗炎（M2）表型。在各种肺部疾病中已经报道了 M1/M2 平衡的改变，这是治疗干预的靶点。本研究旨在评估水溶性一氧化碳释放分子-3（CORM-3）对肺泡巨噬细胞 M1/M2 表型的调节作用。在不存在或存在 CORM-3（0.15mM）处理的情况下，分别用 LPS（5ng/ml）/IFN-γ（10U/ml）或 IL-4（10ng/ml）/IL-13（10ng/ml）刺激培养的大鼠肺泡巨噬细胞（NR8383），以诱导 M1 和 M2 表型。在没有或存在 CORM-3 处理的情况下，分别在 1、3、6 或 24 小时后通过 Western blot 评估 M1 表型标志物诱导型一氧化氮合酶（iNOS）和 TNF-α以及 M2 表型标志物 CD206 和 Ym-1 的表达。用无活性 CORM-3（iCORM-3）作为对照。用 CORM-3 处理未刺激的（未刺激）AM 可促进 M2 表型的进展，这表现为 CD206（在 1 小时；增加 1.8 倍）和 Ym-1（在 3 小时；增加 1.9 倍）的表达增加。令人惊讶的是，在用 CORM-3 刺激 AM 6 小时后，CORM-3 处理还上调了 iNOS 蛋白的表达（增加 2.6 倍）。相反，CORM-3 有效地降低了 LPS/IFN-γ 诱导的 iNOS 蛋白表达（降低 0.6 倍）；然而，它对 TNF-α的表达没有影响。最后，CORM-3 在急性（1-3 小时）中上调了 IL-4-/IL-13 处理（M2 刺激）巨噬细胞中 CD206（增加 1.4 倍）和 Ym-1（增加 1.6 倍）的水平。这些发现表明，CORM-3 可以调节体外巨噬细胞的 M1 和 M2 表型，持续抑制 M1 极化巨噬细胞中的 iNOS 表达，并在 M2 极化巨噬细胞中短暂（早期）上调 CD206 和 Ym-1 蛋白。
