Inhibition of Sonic Hedgehog signaling inhibits fibrous scar formation and adversely affects functional outcome after ischemic brain injury in rats / 南方医科大学学报

ABSTRACT

OBJECTIVE@#To investigate the effects of inhibiting Sonic Hedgehog (Shh) signaling on fibrous scar formation and functional outcome after ischemic brain injury.@*METHODS@#Adult SD rats were randomized into sham-operated group, middle cerebral artery occlusion (MCAO) and reperfusion (I/R) group, I/R with intraventricular empty adenoviral vector (rAd-NC) injection group, and I/R with adenovirus-mediated Shh knockdown (rAd-ShShh) group. After the treatments, the neurological deficits of the rats were assessed, and the protein and mRNA expressions of fibronectin (Fn), α-SMA, and Shh in the ischemic hemisphere were detected with immunofluorescence assay and qPCR; TUNEL staining was used for detecting neural cell apoptosis. In the cell experiment, primary meningeal fibroblasts isolated from neonatal SD rats were pretreated for 24 h with TGF-β1 or TGF-β1 plus cyclopamine (CYC) before oxygen-glucose deprivation for 150 min followed by reoxygenation for 72 h (OGD/R). CCK-8 assay and scratch test were performed to examine the changes in cell proliferation and migration, and immunofluorescence assay, qPCR and Western blotting were used for detecting cell transformation and the expressions of Shh, α-SMA, and Fn.@*RESULTS@#Cerebral I/R injury significantly increased the protein and mRNA expressions of Shh, α-SMA, and Fn in the ischemic hemisphere of the rats, but their expression levels were significantly lowered by intraventricular injection of rAd-Shshh (P < 0.05), which obviously increased cell apoptosis in the ischemic hemisphere (P < 0.05) and improved modified mNSS and modified Bederson scores of the rats (P < 0.05). In the cell experiment, pretreatment with TGF-β1 and TGF-β1+CYC both increased the viability of the primary meningeal fibroblasts after OGD/R. TGF-β1 significantly enhanced the migration ability and induced obvious transformation of the exposed cells (P < 0.05), but these effects were significantly attenuated by co-treatment with CYC (P < 0.05). The expressions of Shh, α-SMA and Fn in the TGF-β1 group were all significantly higher in TGF-β1-treated cells (P < 0.05) and were obviously lowered by co-treatment with CYC (P < 0.05).@*CONCLUSION@#Inhibition of Shh signaling may inhibit fibrous scar formation and functional recovery in rats after ischemic brain injury.