Strange, C. | bioRxiv Science

bioRxiv2025

Alpha-1 Antitrypsin Overexpressing Mesenchymal Stem/Stromal Cells Reverses Type 1 Diabetes via Promoting Treg Function and CD8+ T cell exhaustion

Mesenchymal stem/stromal cell (MSC) therapy holds great promise as both a therapeutic option and as a biofactory, as cells produce therapeutic proteins to augment their efficacy in disease treatment. This study investigates the therapeutic effects and the mechanistic insights of alpha-1 antitrypsin overexpressing MSCs (AAT-MSCs) in diabetes prevention and treatment. A single infusion of AAT-MSCs not only delayed diabetes onset but reversed new-onset type 1 diabetes (T1D) in the nonobese diabetic (NOD) mice. Using single-cell RNA sequencing, flow cytometry, and functional analyses, we characterized the impact of AAT-MSCs on immune cells, particularly CD4+ and CD8+ T cells, in pancreatic lymph nodes (PLNs) and islets of NOD mice. AAT-MSCs enhanced the immunosuppressive function and the communication of regulatory T cells (Tregs) with other immune cells while reducing the numbers of T helper 1 (Th1) cells and CD8+ cytotoxic T cells. In vitro experiments further confirmed the capacity of AAT-MSCs to promote the proliferation of Tregs, which consequently fostered an exhausted phenotype in CD8+ T cells, thereby facilitating {beta} cell survival and potentially aiding in diabetes remission. Thus, our findings underscore the significant protective effects of AAT-MSCs, delineate their novel mechanistic insight on recipient immune cells, and provide evidence for the clinical application of AAT-MSCs in treating T1D.

cell biology↗

bioRxiv2023

Overexpression of Alpha-1 Antitrypsin Increases the Proliferation of Mesenchymal Stem Cells by Upregulation of Cyclin D1 and is Independent of the Wnt Signaling Pathway

Alaph-1 antitrypsin overexpressing mesenchymal stromal/stem cells (AAT-MSCs) showed improved innate properties with a faster proliferation rate when studied for their protective effects in mouse models of diseases. Here, we investigated the potential mechanism(s) by which AAT gene insertion increases MSC proliferation. Human bone marrow-derived primary or immortalized MSCs (iMSCs) or AAT-MSCs (iAAT-MSCs) were used in the study. Cell proliferation was measured by cell counting and cell cycle analysis. Possible pathways involved in the pro-proliferation effect of AAT were investigated by measuring mRNA and protein expression of key cell cycle genes. Interval cell counting showed increased proliferation in AAT-MSCs or iAAT-MSCs compared to their corresponding MSC controls. Cell cycle analysis revealed more cells progressing into the S and G2/M phases in iAAT-MSCs, with a notable increase in the cell cycle protein, Cyclin D1. Moreover, treatment with Cyclin D1 inhibitors showed that the increase in proliferation is due to Cyclin D1 and that the AAT protein is upstream and a positive regulator of Cyclin D1. Furthermore, AATs effect on Cyclin D1 is independent of the Wnt signaling pathway as there were no differences in the expression of regulatory proteins, including GSK3{beta} and {beta}-Catenin in iMSC and iAAT-MSCs. In summary, our results indicate that AAT gene insertion in an immortalized MSC cell line increases cell proliferation and growth by increasing Cyclin D1 expression and consequently causing cells to progress through the cell cycle at a significantly faster rate.

cell biology↗