miR-200c-3p regulates α4 integrin-mediated T cell adhesion and migration.

A microRNA miR-200c-3p is a regulator of epithelial-mesenchymal transition to control adhesion and migration of epithelial and mesenchymal cells. However, little is known about whether miR-200c-3p affects lymphocyte adhesion and migration mediated by integrins. Using TK-1 (a T lymphoblast cell) as a model of T cell, here we show that repressed expression of miR-200c-3p upregulated α4 integrin-mediated adhesion to and migration across mucosal addressin cell adhesion molecule-1 (MAdCAM-1). Conversely, overexpression of miR-200c-3p downregulated α4 integrin-mediated adhesion and migration. Unlike in epithelial cells, miR-200c-3p did not target talin, a conformation activator of integrin, but, targeted E26-transformation-specific sequence 1 (ETS1), a transcriptional activator of α4 integrin, in T cells. Treatment of the miR-200c-3p-low-expressing TK-1 cells that possessed elevated α4 integrin with ETS1 small interfering RNA (siRNA) resulted in the reversion of the α4 integrin expression, supporting that ETS1 is a target of miR-200c-3p. A potential proinflammatory immune-modulator retinoic acid (RA) treatment of TK-1 cells elicited a significant reduction of miR-200c-3p and simultaneously a marked increase in ETS1 and α4 integrin expression. An anti-inflammatory cytokine TGF-ß1 treatment elevated miR-200c-3p, thereby downregulating ETS1 and α4 integrin expression. These results suggest that miR-200c-3p is an important regulator of α4 integrin expression and functions and may be controlled by RA and TGF-ß1 in an opposite way. Overexpression of miR-200c-3p could be a novel therapeutic option for treatment of gut inflammation through suppressing α4 integrin-mediated T cell migration.

MicroRNA Profiles in Intestinal Epithelial Cells in a Mouse Model of Sepsis.

Sepsis is a systemic inflammatory disorder that leads to the dysfunction of multiple organs. In the intestine, the deregulation of the epithelial barrier contributes to the development of sepsis by triggering continuous exposure to harmful factors. However, sepsis-induced epigenetic changes in gene-regulation networks within intestinal epithelial cells (IECs) remain unexplored. In this study, we analyzed the expression profile of microRNAs (miRNAs) in IECs isolated from a mouse model of sepsis generated via cecal slurry injection. Among 239 miRNAs, 14 miRNAs were upregulated, and 9 miRNAs were downregulated in the IECs by sepsis. Upregulated miRNAs in IECs from septic mice, particularly miR-149-5p, miR-466q, miR-495, and miR-511-3p, were seen to exhibit complex and global effects on gene regulation networks. Interestingly, miR-511-3p has emerged as a diagnostic marker in this sepsis model due to its increase in blood in addition to IECs. As expected, mRNAs in the IECs were remarkably altered by sepsis; specifically, 2248 mRNAs were decreased, while 612 mRNAs were increased. This quantitative bias may be possibly derived, at least partly, from the direct effects of the sepsis-increased miRNAs on the comprehensive expression of mRNAs. Thus, current in silico data indicate that there are dynamic regulatory responses of miRNAs to sepsis in IECs. In addition, the miRNAs that were increased with sepsis had enriched downstream pathways including Wnt signaling, which is associated with wound healing, and FGF/FGFR signaling, which has been linked to chronic inflammation and fibrosis. These modifications in miRNA networks in IECs may lead to both pro- and anti-inflammatory effects in sepsis. The four miRNAs discovered above were shown to putatively target LOX, PTCH1, COL22A1, FOXO1, or HMGA2, via in silico analysis, which were associated with Wnt or inflammatory pathways and selected for further study. The expressions of these target genes were downregulated in sepsis IECs, possibly through posttranscriptional modifications of these miRNAs. Taken together, our study suggests that IECs display a distinctive miRNA profile which is capable of comprehensively and functionally reshaping the IEC-specific mRNA landscape in a sepsis model.

The Spike Glycoprotein of SARS-CoV-2 Binds to ß1 Integrins Expressed on the Surface of Lung Epithelial Cells.

The spike glycoprotein attached to the envelope of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) binds to and exploits angiotensin-converting enzyme 2 (ACE2) as an entry receptor to infect pulmonary epithelial cells. A subset of integrins that recognize the arginyl-glycyl-aspartic acid (RGD) sequence in the cognate ligands has been predicted in silico to bind the spike glycoprotein and, thereby, to be exploited for viral infection. Here, we show experimental evidence that the ß1 integrins predominantly expressed on human pulmonary epithelial cell lines and primary mouse alveolar epithelial cells bind to this spike protein. The cellular ß1 integrins support adhesive interactions with the spike protein independently of ACE2, suggesting the possibility that the ß1 integrins may function as an alternative receptor for SARS-CoV-2, which could be targeted for the prevention of viral infections.

The Lectin-Like Domain of Thrombomodulin Inhibits ß1 Integrin-Dependent Binding of Human Breast Cancer-Derived Cell Lines to Fibronectin.

Thrombomodulin is a molecule with anti-coagulant and anti-inflammatory properties. Recently, thrombomodulin was reported to be able to bind extracellular matrix proteins, such as fibronectin and collagen; however, whether thrombomodulin regulates the binding of human breast cancer-derived cell lines to the extracellular matrix remains unknown. To investigate this, we created an extracellular domain of thrombomodulin, TMD123-Fc, or domain deletion TM-Fc proteins (TM domain 12-Fc, TM domain 23-Fc) and examined their bindings to fibronectin in vitro by ELISA. The lectin-like domain of thrombomodulin was found to be essential for the binding of the extracellular domain of thrombomodulin to fibronectin. Using a V-well cell adhesion assay or flow cytometry analysis with fluorescent beads, we found that both TMD123-Fc and TMD12-Fc inhibited the binding between ß1 integrin of human breast cancer-derived cell lines and fibronectin. Furthermore, TMD123-Fc and TMD12-Fc inhibited the binding of activated integrins to fibronectin under shear stress in the presence of Ca2+ and Mg2+ but not under strong integrin-activation conditions in the presence of Mg2+ without Ca2+. This suggests that thrombomodulin Fc fusion protein administered exogenously at a relatively early stage of inflammation may be applied to the development of new therapies that inhibit the binding of ß1 integrin of breast cancer cell lines to fibronectin.