Relaxin inhibits renal myofibroblast differentiation via RXFP1, the nitric oxide pathway, and Smad2.
The hormone relaxin inhibits renal myofibroblast differentiation by interfering with TGF-beta1/Smad2 signaling. However, the pathways involved in the relaxin-TGF-beta1/Smad2 interaction remain unknown. This study investigated the signaling mechanisms by which human gene-2 (H2) relaxin regulates myofibroblast differentiation in vitro by examining its effects on mixed populations of fibroblasts and myofibroblasts propagated from injured rat kidneys. Cultures containing approximately 60-70% myofibroblasts were used to determine which relaxin receptors, G-proteins, and signaling pathways were involved in the H2 relaxin-mediated regulation of alpha-smooth muscle actin (alpha-SMA; a marker of myofibroblast differentiation). H2 relaxin only inhibited alpha-SMA immunostaining and collagen concentration in the presence of relaxin family peptide receptor 1 (RXFP1). H2 relaxin also induced a transient rise in cAMP in the presence of G(i/o) inhibition, and a sustained increase in extracellular signal-regulated kinase (ERK)-1/2 phosphorylation. Furthermore, inhibition of neuronal nitric oxide synthase (nNOS), NO, and cGMP significantly blocked the inhibitory effects of relaxin on alpha-SMA and Smad2 phosphorylation, while the NO inhibitor, L-nitroarginine methyl ester (hydrochloride) (L-NAME) significantly blocked the inhibitory actions of relaxin on collagen concentration in vivo. These findings suggest that relaxin signals through RXFP1, and a nNOS-NO-cGMP-dependent pathway to inhibit Smad2 phosphorylation and interfere with TGF-beta1-mediated renal myofibroblast differentiation and collagen production.
Mookerjee
Ishanee
I
Howard Florey Institute, University of Melbourne, Victoria, Australia.
Hewitson
Tim D
TD
Halls
Michelle L
ML
Summers
Roger J
RJ
Mathai
Michael L
ML
Bathgate
Ross A D
RA
Tregear
Geoffrey W
GW
Samuel
Chrishan S
CS
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism*; D018000:Receptors, Peptide / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology; D012065:Relaxin / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects*; D051899:Smad2 Protein / Q000235:genetics / Q000378:metabolism*
D012333:RNA, Messenger; D043562:Receptors, G-Protein-Coupled; D018000:Receptors, Peptide; D011994:Recombinant Proteins; C504789:Rxfp1 protein, rat; D051899:Smad2 Protein; C497784:Smad2 protein, rat; D009569:Nitric Oxide; D012065:Relaxin