title: Activated protein C ameliorates diabetic nephropathy by epigenetically inhibiting the redox enzyme p66Shc.
abstract: The coagulation protease activated protein C (aPC) confers cytoprotective effects in various in vitro and in vivo disease models, including diabetic nephropathy. The nephroprotective effect may be related to antioxidant effects of aPC. However, the mechanism through which aPC may convey these antioxidant effects and the functional relevance of these properties remain unknown. Here, we show that endogenous and exogenous aPC prevents glomerular accumulation of oxidative stress markers and of the redox-regulating protein p66(Shc) in experimental diabetic nephropathy. These effects were predominately observed in podocytes. In vitro, aPC inhibited glucose-induced expression of p66(Shc) mRNA and protein in podocytes (via PAR-1 and PAR-3) and various endothelial cell lines, but not in glomerular endothelial cells. Treatment with aPC reversed glucose-induced hypomethylation and hyperacetylation of the p66(Shc) promoter in podocytes. The hyperacetylating agent sodium butyrate abolished the suppressive effect of aPC on p66(Shc) expression both in vitro and in vivo. Moreover, sodium butyrate abolished the beneficial effects of aPC in experimental diabetic nephropathy. Inhibition of p66(Shc) expression and mitochondrial translocation by aPC normalized mitochondrial ROS production and the mitochondrial membrane potential in glucose-treated podocytes. Genetic ablation of p66(Shc) compensated for the loss of protein C activation in vivo, normalizing markers of diabetic nephropathy and oxidative stress. These studies identify a unique mechanism underlying the cytoprotective effect of aPC. Activated PC epigenetically controls expression of the redox-regulating protein p66(Shc), thus linking the extracellular protease aPC to mitochondrial function in diabetic nephropathy.
authors.affiliation: Department of Clinical Chemistry and Pathobiochemistry,  Otto-von-Guericke-University, 39120 Magdeburg,  Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Bock
authors.forename: Khurrum
authors.initials: K
authors.lastname: Shahzad
authors.forename: Hongjie
authors.initials: H
authors.lastname: Wang
authors.forename: Stoyan
authors.initials: S
authors.lastname: Stoyanov
authors.forename: Juliane
authors.initials: J
authors.lastname: Wolter
authors.forename: Wei
authors.initials: W
authors.lastname: Dong
authors.forename: Pier Giuseppe
authors.initials: PG
authors.lastname: Pelicci
authors.forename: Muhammed
authors.initials: M
authors.lastname: Kashif
authors.forename: Satish
authors.initials: S
authors.lastname: Ranjan
authors.forename: Simone
authors.initials: S
authors.lastname: Schmidt
authors.forename: Robert
authors.initials: R
authors.lastname: Ritzel
authors.forename: Vedat
authors.initials: V
authors.lastname: Schwenger
authors.forename: Klaus G
authors.initials: KG
authors.lastname: Reymann
authors.forename: Charles T
authors.initials: CT
authors.lastname: Esmon
authors.forename: Thati
authors.initials: T
authors.lastname: Madhusudhan
authors.forename: Peter P
authors.initials: PP
authors.lastname: Nawroth
authors.forename: Berend
authors.initials: B
authors.lastname: Isermann
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D002087:Butyrates / Q000494:pharmacology; D047369:Chromatin Immunoprecipitation; D019175:DNA Methylation / Q000187:drug effects; D017931:DNA Primers / Q000235:genetics; D003921:Diabetes Mellitus, Experimental / Q000150:complications*; D003928:Diabetic Nephropathies / Q000188:drug therapy* / Q000209:etiology; D063185:Epigenetic Repression / Q000187:drug effects*; D055785:Gene Knockdown Techniques; D015151:Immunoblotting; D007150:Immunohistochemistry; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D050199:Podocytes / Q000378:metabolism; D011486:Protein C / Q000494:pharmacology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055765:Shc Signaling Adaptor Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D018709:Statistics, Nonparametric; D013347:Subcellular Fractions
chemical_list: D002087:Butyrates; D017931:DNA Primers; D011486:Protein C; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1