Exosomes Derived from GDNF-Modified Human Adipose Mesenchymal Stem Cells Ameliorate Peritubular Capillary Loss in Tubulointerstitial Fibrosis by Activating the SIRT1/eNOS Signaling Pathway

Abstract

Background: Glial-derived neurotrophic factor (GDNF) has been demonstrated to promote the therapeutic effect of mesenchymal stem cells on ameliorating renal injury. The mechanism may involve the transfer of endogenous molecules, but these factors remain unknown. Methods: GDNF was transfected into human adipose mesenchymal stem cells via a lentiviral transfection system and exosomes were isolated (GDNF-AMSC-exos). Using unilateral ureteral obstruction (UUO) mouse models and human umbilical vein endothelial cells (HUVECs) against hypoxia/serum deprivation (H/SD) injury models we investigated whether GDNF-AMSC-exos ameliorate peritubular capillary (PTC) loss in tubulointerstitial fibrosis, and whether this effect is meditated by SIRT1 signaling pathway. Additionally, by using SIRT1 activators or siRNA, the roles of the candidate mRNA and its downstream gene in GDNF-AMSC-exo-induced regulation of endothelial cell function were assessed. Findings: GDNF-AMSC-exos resulted in significantly decreased PTC rarefaction and renal fibrosis scores in mice with UUO. In vitro studies revealed that GDNF-AMSC-exos exert cytoprotective effects on HUVECs against H/SD injury by stimulating migration, angiogenesis and conferring apoptosis resistance. Mechanistically, GDNF-AMSC-exos enhanced SIRT1 signaling, which accompanied by increased protein levels of phosphorylated eNOS (p-eNOS). In addition, we have confirmed the SIRT1-eNOS interaction in HUVECs by immunoprecipitation. Interpretation: Our study unveils a mechanism by which exosomes ameliorate renal fibrosis: GDNF-AMSC-exos may activate an angiogenesis program in surviving PTCs after injury by activating the SIRT1/eNOS signaling pathway. This finding suggests that inhibition of chronic kidney disease (CKD) progression requires long-term suppression of fibrosis via renal microvascular repair and angiogenesis and that GDNF-AMSC-exos therapy is a promising strategy for achieving this goal. Funding Statement: This work were supported by projects of the National Natural Science Foundation of China (81270769 and 81803473); a project of the Jiangsu Provincial Natural Science Foundation (BK20161172); a project of the Jiangsu Provincial Commission of Health and Family Planning (H201628); projects of the Jiangsu Provincial Post Graduate Innovation Plan (KYCX18-2178, KYCX17-1708, SJCX17-0560, SJCX18-0715); the Municipal Key Research and Development Project of Xuzhou (KC18212); the “Liu ge yi Gong Cheng” project of Jiangsu High-Level Personnel (LGY2016043); a school class project of Xuzhou Medical University (2017KJ13); and the Xuzhou Science and Technology Bureau Applied Basic Research Program( KC18041 and KC 19069). Declaration of Interests: All the authors declared no competing interests. Ethics Approval Statement: Approval of all research involving human participants was obtained from the Institutional Review Board of the Affiliated Hospital of Xuzhou Medical University (permit number: xyfylw2013032). All experiments involving animals followed the animal use protocol enacted by the Institutional Animal Care and Use Committee of Xuzhou Medical University.