Abstract Body: Background and rationale. Diabetes mellitus (DM), particularly in the context of aging (>65 years), drives chronic inflammation and disrupts regenerative responses, resulting in impaired angiogenesis and poor wound healing. Clinical therapies and attempts at therapeutic angiogenesis fail to restore complete vascular networks. A key obstacle to advancement is the incomplete understanding of how DM alters expression of angiogenic mediators such as the VEGF-A, including the balance between its proangiogenic (VEGFA₁₆₅a) and antiangiogenic (VEGFA₁₆₅b) splice variants.
Results. Chronic DM people are most at risk for PAD-critical limb ischemia. We model this using aged experimental type 2 diabetes, leptin receptor (T2DM Lepr^db/db) mice. Our data demonstrated 52 weeks control C57BL/6J mice showed decreased blood flow recovery (~60%) compared to 12 weeks control after 21 days using femoral artery ligation model of PAD and macrophage-mediated vascular repair. T2DM Lepr^db/db mice at 12 weeks also showed delayed blood flow recovery (~50%). When aging was combined with diabetes, there was a further decrease in blood flow recovery (~30%) due to impaired angio/arteriogenesis. Single nuclei RNA-Seq from ischemic muscle harvested on day 3, the peak of early infiltrates inflammatory macrophages, and RNA-seq from polarized M1 macrophages showed excessive activation of inflammatory pathways including NLRP3, CXCL2 and IL-1β while expression of regenerative mediators (TGFβ-SMAD, ANGPT1, VEGF-A, VEGFR1/2) were reduced in aged old diabetic mice compare to control. Moreover, the RNA binding protein MSI2 expression, is reduced in both day 3 ischemic muscle and macrophages in diabetic mice and emerges as a central regulator of macrophage-mediated angiogenic signaling. RNA binding assay revealed decreased MSI2 binding to proangiogenic VEGFA mRNA in diabetic mice. Serial concentrations of Ro 08-2750, a selective inhibitor of MSI2, showed decreased proangiogenic VEGFA in C57BL/6J control. Lastly, transfection of old diabetic macrophages with a constitutively active MSI2 plasmid DNA restores VEGFA₁₆₅a expression.
Conclusion. Long-term diabetes disrupts angiogenesis by driving harmful inflammation and weakening angiogenic responses. MSI2 emerges as a potent therapeutic target; by enhancing regenerative pathways such as VEGF-A post-transcription and translation. It may be possible to reprogram diabetic macrophages by restoring neo/revascularization capacity in the context of diabetes.