RESUMEN
Parathyroid hormone (PTH) plays a central role in the regulation of serum calcium and phosphorus homeostasis, while parathyroid hormone-related protein (PTHrP) has important developmental roles. Both peptides signal through the same G protein-coupled receptor, the PTH/PTHrP or PTH type 1 receptor (PTH1R). PTHrP, normally a secreted protein, also contains a nuclear localization signal (NLS) that in vitro imparts functionality to the protein at the level of the nucleus. We investigated this functionality in vivo by introducing a premature termination codon in Pthrp in ES cells and generating mice that express PTHrP (1-84), a truncated form of the protein that is missing the NLS and the C-terminal region of the protein but can still signal through its cell surface receptor. Mice homozygous for the knock-in mutation (Pthrp KI) displayed retarded growth, early senescence, and malnutrition leading postnatally to their rapid demise. Decreased cellular proliferative capacity and increased apoptosis in multiple tissues including bone and bone marrow cells were associated with altered expression and subcellular distribution of the senescence-associated tumor suppressor proteins p16(INK4a) and p21 and the oncogenes Cyclin D, pRb, and Bmi-1. These findings provide in vivo experimental proof that substantiates the biologic relevance of the NLS and C-terminal portion of PTHrP, a polypeptide ligand that signals mainly via a cell surface G protein-coupled receptor.
Asunto(s)
Trastornos del Crecimiento/etiología , Señales de Localización Nuclear/deficiencia , Proteína Relacionada con la Hormona Paratiroidea/genética , Proteína Relacionada con la Hormona Paratiroidea/fisiología , Animales , Apoptosis , Células de la Médula Ósea/patología , Huesos/patología , Proliferación Celular , Técnicas de Sustitución del Gen , Trastornos del Crecimiento/genética , Desnutrición/etiología , Desnutrición/genética , Ratones , Proteínas Mutantes , Oncogenes , Tasa de Supervivencia , Proteínas Supresoras de TumorRESUMEN
Mice heterozygous for targeted disruption of Pthrp exhibit, by 3 months of age, diminished bone volume and skeletal microarchitectural changes indicative of advanced osteoporosis. Impaired bone formation arising from decreased BM precursor cell recruitment and increased apoptotic death of osteoblastic cells was identified as the underlying mechanism for low bone mass. The osteoporotic phenotype was recapitulated in mice with osteoblast-specific targeted disruption of Pthrp, generated using Cre-LoxP technology, and defective bone formation was reaffirmed as the underlying etiology. Daily administration of the 1-34 amino-terminal fragment of parathyroid hormone (PTH 1-34) to Pthrp+/- mice resulted in profound improvement in all parameters of skeletal microarchitecture, surpassing the improvement observed in treated WT littermates. These findings establish a pivotal role for osteoblast-derived PTH-related protein (PTHrP) as a potent endogenous bone anabolic factor that potentiates bone formation by altering osteoblast recruitment and survival and whose level of expression in the bone microenvironment influences the therapeutic efficacy of exogenous PTH 1-34.
Asunto(s)
Huesos , Osteoblastos/metabolismo , Osteogénesis/fisiología , Proteína Relacionada con la Hormona Paratiroidea/metabolismo , Hormona Paratiroidea/metabolismo , Fragmentos de Péptidos/metabolismo , Animales , Apoptosis/fisiología , Densidad Ósea , Enfermedades Óseas Metabólicas/genética , Enfermedades Óseas Metabólicas/patología , Huesos/anatomía & histología , Huesos/citología , Huesos/metabolismo , Marcación de Gen , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Osteoblastos/citología , Osteoporosis/patología , Osteoporosis/fisiopatología , Hormona Paratiroidea/administración & dosificación , Hormona Paratiroidea/genética , Hormona Paratiroidea/uso terapéutico , Proteína Relacionada con la Hormona Paratiroidea/genética , Fragmentos de Péptidos/administración & dosificación , Fragmentos de Péptidos/genética , Fragmentos de Péptidos/uso terapéuticoRESUMEN
We investigated the relative contributions of PTH and PTHrP to the skeletal phenotype of mice deficient in PTH (PTH(-/-)). PTH(-/-) mice and PTH(-/-) mice lacking one allele encoding PTHrP (PTH(-/-)PTHrP(+/-)) were compared. Both mutants displayed similar biochemical abnormalities of hypoparathyroidism, but skeletal PTHrP mRNA and protein were decreased in PTH(-/-)PTHrP(+/ -) mice. PTH(-/-) mice had increased trabecular bone volume with diminished bone turnover. PTHrP haploinsufficiency reduced trabecular bone of the PTH(-/-) mice to levels below those in wild-type animals by decreasing osteoprogenitor cell recruitment, enhancing osteoblast apoptosis, and diminishing bone formation. The results show that the increased trabecular bone volume in PTH-deficient mice is due to diminished PTH-induced osteoclastic bone resorption and persistent PTHrP-stimulated osteoblastic bone formation. They also illustrate the changing role of PTHrP during bone development, demonstrate its bone- forming function in the postnatal state, and support its pharmacological potential as an anabolic agent.
Asunto(s)
Densidad Ósea , Proteína Relacionada con la Hormona Paratiroidea/fisiología , Hormona Paratiroidea/fisiología , Animales , Resorción Ósea , Glicoproteínas/metabolismo , Ratones , Ratones Endogámicos C57BL , Osteoclastos/fisiología , Osteogénesis , Osteoprotegerina , Receptores Citoplasmáticos y Nucleares/metabolismo , Receptores del Factor de Necrosis TumoralRESUMEN
Thiazolidinediones (TZDs) increase peripheral tissue insulin sensitivity in patients with type 2 diabetes mellitus by activating the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma). In bone marrow stromal cell cultures and in vivo, activation of PPARgamma by high doses (20 mg/kg/day) of TZDs has been reported to alter stem cell differentiation by promoting commitment of progenitor cells to the adipocytic lineage while inhibiting osteoblastogenesis. Here, we have examined the in vivo effects of low-dose rosiglitazone (3 mg/kg/day) on bone, administered to mice by gavage for 90 days. Rosiglitazone-treated mice had increased weight when compared with controls, with no significant alterations in serum levels of glucose, calcium or parathyroid hormone (PTH). Bone mineral density (BMD) at the lumbar vertebrae (L1-L4), ilium/sacrum, and total body was diminished by rosiglitazone treatment. Histologically, bone was characterized by decreased trabecular bone volume and increased marrow space with no significant change in bone marrow adipocity. Decreased osteoblast number and activity due to increased apoptotic death of osteoblasts and osteocytes was apparent while osteoclast parameters and serum levels of osteocalcin, alkaline phosphatase activity, and leptin were unaltered by rosiglitazone treatment. Therefore, the imbalance in bone remodeling that follows rosiglitazone administration arises from increased apoptotic death of osteogenic cells and diminished bone formation leading to the observed decrease in trabecular bone volume and BMD. These novel in vivo effects of TZDs on bone are of clinical relevance as patients with type 2 diabetes mellitus and other insulin resistant states treated with these agents may potentially be at increased risk of osteoporosis.