Hilt Zachary T, Maurya Preeti, Tesoro Laura, Pariser Daphne N, Ture Sara K, Cleary Simon J, Looney Mark R, McGrath Kathleen E, Morrell Craig N
Aab Cardiovascular Research Institute (Z.T.H., P.M., L.T., D.N.P., S.K.T., C.N.M.), University of Rochester School of Medicine, NY.
Department of Microbiology and Immunology, Cornell University, Ithaca, NY (Z.T.H.).
Circ Res. 2021 Mar 5;128(5):655-669. doi: 10.1161/CIRCRESAHA.120.317119. Epub 2021 Jan 29.
Circulating monocytes can have proinflammatory or proreparative phenotypes. The endogenous signaling molecules and pathways that regulate monocyte polarization in vivo are poorly understood. We have shown that platelet-derived β2M (β-2 microglobulin) and TGF-β (transforming growth factor β) have opposing effects on monocytes by inducing inflammatory and reparative phenotypes, respectively, but each bind and signal through the same receptor. We now define the signaling pathways involved.
To determine the molecular mechanisms and signal transduction pathways by which β2M and TGF-β regulate monocyte responses both in vitro and in vivo.
Wild-type- (WT) and platelet-specific β2M knockout mice were treated intravenously with either β2M or TGF-β to increase plasma concentrations to those in cardiovascular diseases. Elevated plasma β2M increased proinflammatory monocytes, while increased plasma TGFβ increased proreparative monocytes. TGF-βR (TGF-β receptor) inhibition blunted monocyte responses to both β2M and TGF-β in vivo. Using imaging flow cytometry, we found that β2M decreased monocyte SMAD2/3 nuclear localization, while TGF-β promoted SMAD nuclear translocation but decreased noncanonical/inflammatory (JNK [jun kinase] and NF-κB [nuclear factor-κB] nuclear localization). This was confirmed in vitro using both imaging flow cytometry and immunoblots. β2M, but not TGF-β, promoted ubiquitination of SMAD3 and SMAD4, that inhibited their nuclear trafficking. Inhibition of ubiquitin ligase activity blocked noncanonical SMAD-independent monocyte signaling and skewed monocytes towards a proreparative monocyte response.
Our findings indicate that elevated plasma β2M and TGF-β dichotomously polarize monocytes. Furthermore, these immune molecules share a common receptor but induce SMAD-dependent canonical signaling (TGF-β) versus noncanonical SMAD-independent signaling (β2M) in a ubiquitin ligase dependent manner. This work has broad implications as β2M is increased in several inflammatory conditions, while TGF-β is increased in fibrotic diseases. Graphic Abstract: A graphic abstract is available for this article.
循环单核细胞可具有促炎或促修复表型。体内调节单核细胞极化的内源性信号分子和途径尚不清楚。我们已经表明，血小板衍生的β2M（β-2微球蛋白）和TGF-β（转化生长因子β）分别通过诱导炎症和修复表型对单核细胞产生相反的作用，但它们都通过相同的受体结合并发出信号。我们现在确定其中涉及的信号通路。
确定β2M和TGF-β在体外和体内调节单核细胞反应的分子机制和信号转导通路。
野生型（WT）和血小板特异性β2M基因敲除小鼠通过静脉注射β2M或TGF-β进行处理，以使血浆浓度升高至心血管疾病中的水平。血浆β2M升高会增加促炎单核细胞，而血浆TGF-β升高会增加促修复单核细胞。TGF-βR（TGF-β受体）抑制减弱了体内单核细胞对β2M和TGF-β的反应。使用成像流式细胞术，我们发现β2M降低了单核细胞SMAD2/3的核定位，而TGF-β促进了SMAD的核转位，但降低了非经典/炎症性（JNK [应激活化蛋白激酶]和NF-κB [核因子κB]）的核定位。这在体外使用成像流式细胞术和免疫印迹得到了证实。β2M而非TGF-β促进了SMAD3和SMAD4的泛素化，从而抑制了它们的核转运。泛素连接酶活性的抑制阻断了非经典的不依赖SMAD的单核细胞信号传导，并使单核细胞偏向促修复单核细胞反应。
我们的研究结果表明，血浆β2M和TGF-β升高会使单核细胞产生二分极化。此外，这些免疫分子共享一个共同的受体，但以泛素连接酶依赖的方式诱导依赖SMAD的经典信号传导（TGF-β）与不依赖SMAD的非经典信号传导（β2M）。这项工作具有广泛的意义，因为β2M在几种炎症状态下会升高，而TGF-β在纤维化疾病中会升高。图形摘要：本文提供了图形摘要。
