RESUMEN
Regenerative therapy based on mesenchymal stem cells (MSCs) has great promise to achieve functional recovery in cerebral infarction patients. However, the survival rate of transplanted MSCs is extremely low because of destructive autophagy caused by the harsh ischemic microenvironment in cerebral infarct tissue. The mechanism by which fibronectin type III domain protein 5 (FNDC5) regulates autophagy of transplanted bone marrow-MSCs (BMSCs) following ischemic injury needs to be elucidated. In this study, we confirmed that FNDC5 promotes the survival of transplanted BMSCs in a rat cerebral infarction model. Furthermore, bioinformatic analysis and verification experiments revealed the transcription factor, Sp1, to be a key mediator of autophagy regulation by FNDC5. FNDC5 significantly inhibited BMSC autophagy by down-regulating Sp1 and the autophagy-related Sp1-target gene, ULK2. Transplanted BMSCs overexpressing FNDC5 (BMSCs-OE-FNDC5) promoted neurovascular proliferation and alleviated ischemic brain injury in cerebral infarct model rats. However, the increased survival and enhanced neuroprotective effect of transplanted BMSCs-OE-FNDC5 were reversed by simultaneous overexpression of Sp1. Our data indicate a role for FNDC5 in BMSC survival and reveal a novel mechanism of transcription regulation through Sp1 for the autophagy-related gene ULK2. Modulation of FNDC5 may promote survival capacity and improve the therapeutic effect of BMSCs in various tissues following ischemia.
RESUMEN
Background: Most bone marrow mesenchymal stem cell (BMSC) death is caused by the harsh ischemia and hypoxic microenvironment, which impacts the therapeutic effects of transplanted BMSCs. Fibronectin type III domain-containing protein 5 (FNDC5) and its cleaved product, irisin, are reportedly involved in cerebral protective effect. Research into whether FNDC5 plays a key role in the survival rate of BMSCs and cerebral infarction (CI) remains inadequate. The present study aimed to clarify the protective role of FNDC5 on the low viability of transplanted BMSCs and improve CI treatment outcomes. Methods: A lentivirus vector, which drives the expression of FNDC5, was constructed and used to transfect BMSCs. Cell Counting Kit-8 (CCK8), flow cytometry, immunofluorescence, and western blot were performed to evaluate the function of FNDC5-overexpressing BMSCs (BMSCs-OE-FNDC5) exposed to hypoxic and serum deprivation (H/SD) stress. Transmission electron microscopy (TEM) was used to monitor autophagy. In addition, BMSCs were engrafted into a middle cerebral artery occlusion (MCAO) rat model with or without FNDC5-overexpression (OE-FNDC5). The survival rate of transplanted BMSCs was evaluated by 5-ethynyl-2'-deoxyuridine (EdU) labeling. The CI volume was assessed by 2,3,5-triphenyl tetrazolium chloride (TTC) staining. Results: H/SD stress caused increased cell autophagy, apoptosis, and decreased cell viability of BMSCs, while OE-FNDC5 alleviated these injuries. The in vivo results showed that transplantation of BMSCs-OE-FNDC5 reduced the infarct volume in the rat MCAO model. Furthermore, OE-FNDC5 decreased neuronal apoptosis. The improved therapeutic efficacy of BMSCs-OE-FNDC5 may be attributable to the obviously increased cell survival number after transplantation. Conclusions: These results indicated that FNDC5 overexpression promotes BMSC survival in a CI model, which might provide a potential therapeutic target.