Delayed fracture healing is a clinical problem in diabetic patients. However, the mechanisms of diabetic delayed bone repair remain unknown. Here, we investigate the role of macrophages in diabetic delayed bone repair after femoral bone injury in streptozotocin (STZ)-treated and plasminogen activator inhibitor-1 (PAI-1)-deficient female mice. STZ treatment significantly decreased the numbers of F4/80-positive cells (macrophages) but not granulocyte-differentiation antigen-1-positive cells (neutrophils) at the damaged site on day 2 after femoral bone injury in mice. It significantly decreased the messenger RNA (mRNA) levels of macrophage colony-stimulating factor, inducible nitric oxide synthase (iNOS), interleukin (IL)-6, and CD206 at the damaged site on day 2 after bone injury. Moreover, STZ treatment attenuated a decrease in the number of hematopoietic stem cells in bone marrow induced by bone injury. On the other hand, PAI-1 deficiency significantly attenuated a decrease in the number of F4/80-positive cells induced by STZ treatment at the damaged site on day 2 after bone injury in mice. PAI-1 deficiency did not affect the mRNA levels of iNOS and IL-6 in F4/80- and CD11b-double-positive cells from the bone marrow of the damaged femurs decreased by diabetes in mice. PAI-1 deficiency significantly attenuated the phagocytosis of macrophages at the damaged site suppressed by diabetes. In conclusion, we demonstrated that type 1 diabetes decreases accumulation and phagocytosis of macrophages at the damaged site during early bone repair after femoral bone injury through PAI-1 in female mice.

We recently revealed that plasminogen activator inhibitor-1 (PAI-1), a serine protease inhibitor, is involved in diabetes, osteoporosis and muscle wasting induced by glucocorticoid (GC) treatment in mice. In the present study, we investigated the detailed mechanisms by which GC induces muscle wasting through PAI-1 in vivo and in vitro. PAI-1 deficiency suppressed the mRNA levels of atrogin1 and muscle RING-Finger Protein 1 (MuRF1), ubiquitin ligases leading to muscle degradation, elevated by GC treatment in the gastrocnemius muscle of mice. In vitro study revealed that active PAI-1 treatment augmented the increase in atrogin1 mRNA levels enhanced by dexamethasone (Dex) in mouse myoblastic C2C12 cells. Moreover, a reduction in endogenous PAI-1 level by siRNA suppressed the mRNA levels of atrogin1 and MuRF1 enhanced by Dex in C2C12 cells. In contrast, a reduction in endogenous PAI-1 levels and active PAI-1 did not affect the phosphorylations of Akt and p70S6 kinase nor myogenic differentiation with or without Dex in C2C12 cells. In addition, PAI-1 deficiency blunted IGF-1 mRNA levels decreased by GC treatment in the gastrocnemius muscle of mice, although neither active PAI-1 nor a reduction in endogenous PAI-1 levels affected the levels of IGF-1 mRNA in C2C12 cells in the presence of Dex. In conclusion, our data suggest that paracrine PAI-1 is involved in GC-induced muscle wasting through the enhancement of muscle degradation in mice.

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