SHH adenovirus

Interleukin 4 (IL4) plays a decisive role in M2 macrophage polarization. Sonic Hedgehog (Shh) signaling can regulate fibrosis in central and peripheral tissues. Here, researchers found that cerebral ischemia in rats induced persistent fibrosis and massive macrophage infiltration in the ischemic core. In addition, the number of infiltrating macrophages was significantly positively correlated with the expression of fibronectin (FN). Elimination of circulating monocyte-derived macrophages by clodronate liposomes alleviated fibrotic scars. In addition, IL4 was significantly enhanced in rat ischemic brain tissue, and IL4-dependent M2 macrophage polarization was susceptible to fibrotic scar formation in the ischemic core. Meanwhile, macrophage-conditioned medium directly promoted fibroblast proliferation and production of extracellular matrix (ECM) proteins in vitro. Using pharmacological and genetic methods, it was confirmed that Shh protein secreted by M2 macrophages can promote fibrosis formation both in vitro and in vivo, and this promotion is mediated by key fibrosis-related regulatory proteins TGFβ1 and MMP9. At the same time, the repair effect of IL4 on angiogenesis, apoptosis and infarct volume in the ischemic core area of ​​MCAO/R rats was significantly weakened under the intervention of Shh signal inhibitors, and was positively correlated with the degree of fibrosis.

The researchers investigated whether the Shh signaling pathway affects the production of FN and Col1 in fibroblasts by co-culture with MCM in vitro and in vivo MCAO/R model. Immunoblotting analysis showed that the expression of FN and Col1 proteins in fibroblasts in the MCM/IL4 group was significantly higher than that in the MCM/DMEM group, Con group or DMEM/IL4/Cyc group, while this expression was inhibited in the MCM/IL4/Cyc group (Figure 1A). In vivo experiments, knocking down Shh by adenoviral infection reduced the levels of FN and Col1 proteins in the ischemic core area 7 days after MCAO/R (Figure 1B, C), and eliminated the effect of IL4 on FN and Col1 protein expression (Figure 1C). In addition, overexpression of Shh by adenoviral infection had opposite effects on FN and Col1 expression (Figure 1D). According to the results obtained, Shh signaling can regulate the expression of FN and Col1 in vitro and in vivo.

Figure 1. Shh signaling regulates the production of ECM proteins. (Huang J, et al. 2022)