ABSTRACT
Overexpression or pharmacological activation of SIRT1 has been shown to extend the lifespan of mice and protect against aging-related diseases. Here we show that pharmacological activation of SIRT1 protects in two models of osteoporosis. Ovariectomized female mice and aged male mice, models for post-menopausal and aging-related osteoporosis, respectively, show significant improvements in bone mass upon treatment with SIRT1 agonist, SRT1720. Further, we find that calorie restriction (CR) results in a two-fold upregulation of sirt1 mRNA expression in bone tissue that is associated with increased bone mass in CR mice. Reciprocally, SIRT1 whole-body knockout (KO) mice, as well as osteoblast and osteoclast specific KOs, show a low bone mass phenotype; though double knockout mice (containing SIRT1 deleted in both osteoblasts and osteoclasts) do not show a more severe phenotype. Altogether, these findings provide strong evidence that SIRT1 is a positive regulator of bone mass and a promising target for the development of novel therapeutics for osteoporosis.
Subject(s)
Bone Density Conservation Agents/pharmacology , Bone Density/drug effects , Bone and Bones/drug effects , Heterocyclic Compounds, 4 or More Rings/pharmacology , Osteoporosis/drug therapy , Sirtuin 1/metabolism , Aging/drug effects , Aging/metabolism , Animals , Bone Density/physiology , Bone and Bones/diagnostic imaging , Bone and Bones/metabolism , Caloric Restriction , Disease Models, Animal , Female , Male , Mice, 129 Strain , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Transgenic , Osteoblasts/drug effects , Osteoblasts/metabolism , Osteoclasts/drug effects , Osteoclasts/metabolism , Osteoporosis/diagnostic imaging , Osteoporosis/metabolism , Ovariectomy , Phenotype , RNA, Messenger/metabolism , Sirtuin 1/geneticsABSTRACT
Activation of SIRT1 has previously been shown to protect mice against osteoporosis through yet ill-defined mechanisms. In this study, we outline a role for SIRT1 as a positive regulator of the master osteoblast transcription factor, RUNX2. We find that ex vivo deletion of sirt1 leads to decreased expression of runx2 downstream targets, but not runx2 itself, along with reduced osteoblast differentiation. Reciprocally, treatment with a SIRT1 agonist promotes osteoblast differentiation, as well as the expression of runx2 downstream targets, in a SIRT1-dependent manner. Biochemical and luciferase reporter assays demonstrate that SIRT1 interacts with and promotes the transactivation potential of RUNX2. Intriguingly, mice treated with the SIRT1 agonist, resveratrol, show similar increases in the expression of RUNX2 targets in their calvaria (bone tissue), validating SIRT1 as a physiologically relevant regulator of RUNX2.
Subject(s)
Core Binding Factor Alpha 1 Subunit/metabolism , Osteoblasts/metabolism , Transcription Factors/metabolism , Animals , Cell Differentiation/drug effects , Cell Differentiation/physiology , Male , Mice , Mice, Inbred C57BL , Osteoblasts/drug effects , Osteogenesis/drug effects , Osteogenesis/physiology , Resveratrol , Sirtuin 1/antagonists & inhibitors , Sirtuin 1/metabolism , Stilbenes/pharmacologyABSTRACT
Hepatocyte growth factor (HGF) has been shown to have anti-fibrotic, pro-angiogenic, and cardioprotective effects; however, it is highly unstable and expensive to manufacture, hindering its clinical translation. Recently, a HGF fragment (HGF-f), an alternative c-MET agonist, was engineered to possess increased stability and recombinant expression yields. In this study, we assessed the potential of HGF-f, delivered in an extracellular matrix (ECM)-derived hydrogel, as a potential treatment for myocardial infarction (MI). HGF-f protected cardiomyocytes from serum-starvation and induced down-regulation of fibrotic markers in whole cardiac cell isolate compared to the untreated control. The ECM hydrogel prolonged release of HGF-f compared to collagen gels, and in vivo delivery of HGF-f from ECM hydrogels mitigated negative left ventricular (LV) remodeling, improved fractional area change (FAC), and increased arteriole density in a rat myocardial infarction model. These results indicate that HGF-f may be a viable alternative to using recombinant HGF, and that an ECM hydrogel can be employed to increase growth factor retention and efficacy.
Subject(s)
Drug Delivery Systems , Hepatocyte Growth Factor/therapeutic use , Hydrogel, Polyethylene Glycol Dimethacrylate/chemistry , Myocardial Infarction/drug therapy , Myocardial Infarction/physiopathology , Protein Engineering , Ventricular Remodeling , Animals , Blood Vessels/drug effects , Blood Vessels/pathology , Cell Size/drug effects , Disease Models, Animal , Extracellular Matrix/drug effects , Extracellular Matrix/metabolism , Female , Fibrosis/pathology , Heart Function Tests , Humans , Myocardial Infarction/diagnostic imaging , Myocardial Infarction/pathology , Myocytes, Cardiac/pathology , Myocytes, Smooth Muscle/metabolism , Neovascularization, Physiologic/drug effects , Peptide Fragments/pharmacology , Peptide Fragments/therapeutic use , Proto-Oncogene Proteins c-met/metabolism , Rats, Sprague-Dawley , Sus scrofa , Ultrasonography , Ventricular Remodeling/drug effectsABSTRACT
Hepatocyte growth factor (HGF), through activation of the c-MET receptor, mediates biological processes critical for tissue regeneration; however, its clinical application is limited by protein instability and poor recombinant expression. We previously engineered an HGF fragment (eNK1) that possesses increased stability and expression yield and developed a c-MET agonist by coupling eNK1 through an introduced cysteine residue. Here, we further characterize this eNK1 dimer and show it elicits significantly greater c-MET activation, cell migration, and proliferation than the eNK1 monomer. The efficacy of the eNK1 dimer was similar to HGF, suggesting its promise as a c-MET agonist.
