Oxidative stress in the pathogenesis of renal injury: the effect of the antioxidant danshensu on renal fibrosis
von Hannah Maria Plöger geb. Neuenhofer
Datum der mündl. Prüfung:2022-12-07
Betreuer:Prof. Dr. Hassan Dihazi
Gutachter:Prof. Dr. Michael Müller
Gutachter:Prof. Dr. Margarete Schön
EnglischChronic kidney disease (CKD), often manifested in its end-stage as renal fibrosis, is a widespread illness, for example in children following congenital urinary obstruction and in adults due to an increasing number of lifestyle diseases. Currently only few therapeutic options exist, calling for the development of other treatment opportunities. Therefore, this study aimed to examine a promising treatment candidate, danshensu, an extract of Salvia miltiorrhizae (red sage), by analyzing its effect on oxidative stress, as pathogenetic contributor to fibrosis, and on fibrosis development itself. An in-vitro model of TK173 kidney fibroblasts stressed by reactive oxygen species (ROS) and an in-vivo mouse kidney model of unilateral ureteral obstruction (UUO)-induced fibrosis were applied. In the in-vitro model, danshensu treatment increased TK173 cell viability and improved cell viability in the MTT assay after H2O2-induced oxidative stress. Correspondingly, FACS quantification of DCF-marked ROS showed a reduction of intracellular oxidative stress. In Western blot, SOD1 expression, as an important protein in the cellular first line of defense against oxidative stress, and the expression of extracellular matrix (ECM) proteins fibronectin (FN1) and vimentin (VIM) were analyzed, revealing upregulation after danshensu treatment. In the in-vivo UUO mouse model, danshensu treatment resulted in a downregulation of the TGF-β pathway, a central inductor of fibrosis, determined indirectly through the reduction of nuclear pSmad2 in immunohistochemical staining. Additionally, expression of α-SMA, a marker of epithelial mesenchymal transition, and collagen 1 (Col1), a further ECM protein typically upregulated in fibrosis, were significantly downregulated in the renal tissue as a sign of fibrosis attenuation, suiting the reduction of tubular damage analyzed in PAS tissue stains. These observations are supported through data confirming similar effects of danshensu in other models and organs. Interestingly, upregulation of VIM supports its cytoprotective role in renal cells shown in past studies. This study shows that danshensu rescues renal cell lines from oxidative stress through upregulation of its antioxidant potential, reduces the expression of ECM proteins α-SMA and Col1 through inhibition of the pro-fibrotic TGF-β pathway, and thus has a protective effect on fibrosis progression. These positive effects allow the hope of establishing danshensu as a treatment opportunity for CKD in children and in adults.
Keywords: kidney fibrosis; oxidative stress; TGF-beta; chronic kidney disease; urinary obstruction