title: Essential roles of the nitric oxide (no)/cGMP/protein kinase G type-Iα (PKG-Iα) signaling pathway and the atrial natriuretic peptide (ANP)/cGMP/PKG-Iα autocrine loop in promoting proliferation and cell survival of OP9 bone marrow stromal cells.
abstract: Inappropriate signaling conditions within bone marrow stromal cells (BMSCs) can lead to loss of BMSC survival, contributing to the loss of a proper micro-environmental niche for hematopoietic stem cells (HSCs), ultimately causing bone marrow failure. In the present study, we investigated the novel role of endogenous atrial natriuretic peptide (ANP) and the nitric oxide (NO)/cGMP/protein kinase G type-Iα (PKG-Iα) signaling pathway in regulating BMSC survival and proliferation, using the OP9 BMSC cell line commonly used for facilitating the differentiation of HSCs. Using an ANP-receptor blocker, endogenously produced ANP was found to promote cell proliferation and prevent apoptosis. NO donor SNAP (S-nitroso-N-acetylpenicillamine) at low concentrations (10 and 50 µM), which would moderately stimulate PKG activity, protected these BMSCs against spontaneous apoptosis. YC-1, a soluble guanylyl cyclase (sGC) activator, decreased the levels of apoptosis, similar to the cytoprotective effects of low-level NO. ODQ (1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one), which blocks endogenous NO-induced activation of sGC and thus lowers endogenous cGMP/PKG activity, significantly elevated apoptotic levels by 2.5- and three-fold. Pre-incubation with 8-Bromo-cGMP or ANP, which bypass the ODQ block, almost completely prevented the ODQ-induced apoptosis. A highly-specific PKG inhibitor, DT-3, at 20, and 30 µM, caused 1.5- and two-fold increases in apoptosis, respectively. ODQ and DT-3 also decreased BMSCs proliferation and colony formation. Small Interfering RNA gene knockdown of PKG-Iα increased apoptosis and decreased proliferation in BMSCs. The data suggest that basal NO/cGMP/PKG-Iα activity and autocrine ANP/cGMP/PKG-Iα are necessary for preserving OP9 cell survival and promoting cell proliferation and migration.
authors.affiliation: Cancer Molecular Biology Section, Nevada Cancer Institute, Las Vegas, Nevada 89135, USA.
authors.forename: Janica C
authors.initials: JC
authors.lastname: Wong
authors.forename: Ronald R
authors.initials: RR
authors.lastname: Fiscus
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D019898:Autocrine Communication; D001854:Bone Marrow Cells / Q000166:cytology / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D006152:Cyclic GMP / Q000378:metabolism*; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases / Q000235:genetics* / Q000378:metabolism; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000378:metabolism; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D039902:Multipotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011810:Quinoxalines / Q000494:pharmacology; D034622:RNA Interference; D015398:Signal Transduction; D000098662:Vasodilator-Stimulated Phosphoprotein
chemical_list: D015815:Cell Adhesion Molecules; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D008840:Microfilament Proteins; D010750:Phosphoproteins; D011810:Quinoxalines; D000098662:Vasodilator-Stimulated Phosphoprotein; D009569:Nitric Oxide; D009320:Atrial Natriuretic Factor; D052250:Nitric Oxide Synthase Type III; C496315:Nos3 protein, mouse; C499321:Parp1 protein, mouse; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases; C569720:Prkg1 protein, mouse; D006152:Cyclic GMP