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1.
Eur J Endocrinol ; 184(5): 687-697, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33683213

RESUMO

Objective: Activation of brown adipose tissue is a promising strategy to treat and prevent obesity and obesity-related disorders. Activation of uncoupling protein 1 (UCP1) leads to uncoupled respiration and dissipation of stored energy as heat. Induction of UCP1-rich adipocytes in white adipose tissue, a process known as 'browning', serves as an alternative strategy to increase whole body uncoupling capacity. Here, we aim to assess the association between parathyroid hormone (PTH) receptor expression and UCP1 expression in human adipose tissues and to study PTH effects on human white and brown adipocyte lipolysis and UCP1 expression. Design: A descriptive study of human neck adipose tissue biopsies substantiated by an interventional study on human neck-derived adipose tissue cell models. Methods: Thermogenic markers and PTH receptor gene expression are assessed in human neck adipose tissue biopsies and are related to individual health records. PTH-initiated lipolysis and thermogenic gene induction are assessed in cultured human white and brown adipocyte cell models. PTH receptor involvement is investigated by PTH receptor silencing. Results: PTH receptor gene expression correlates with UCP1 gene expression in the deep-neck adipose tissue in humans. In cell models, PTH receptor stimulation increases lipolysis and stimulates gene transcription of multiple thermogenic markers. Silencing of the PTH receptor attenuates the effects of PTH indicating a direct PTH effect via this receptor. Conclusion: PTH 1 receptor stimulation by PTH may play a role in human adipose tissue metabolism by affecting lipolysis and thermogenic capacity.


Assuntos
Adipócitos/efeitos dos fármacos , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo/efeitos dos fármacos , Lipólise/efeitos dos fármacos , Hormônio Paratireóideo/farmacologia , Receptor Tipo 1 de Hormônio Paratireóideo/metabolismo , Termogênese/efeitos dos fármacos , Adipócitos/metabolismo , Tecido Adiposo/metabolismo , Adulto , Idoso , Estudos Transversais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Proteína Desacopladora 1/metabolismo , Adulto Jovem
2.
Am J Physiol Renal Physiol ; 319(3): F541-F551, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32744087

RESUMO

Plasma phosphate (Pi) levels are tightly controlled, and elevated plasma Pi levels are associated with an increased risk of cardiovascular complications and death. Two renal transport proteins mediate the majority of Pi reabsorption: Na+-phosphate cotransporters Npt2a and Npt2c, with Npt2a accounting for 70-80% of Pi reabsorption. The aim of the present study was to determine the in vitro effects of a novel Npt2a inhibitor (PF-06869206) in opossum kidney (OK) cells as well as determine its selectivity in vivo in Npt2a knockout (Npt2a-/-) mice. In OK cells, Npt2a inhibitor caused dose-dependent reductions of Na+-dependent Pi uptake (IC50: ~1.4 µmol/L), whereas the unselective Npt2 inhibitor phosphonoformic acid (PFA) resulted in an ~20% stronger inhibition of Pi uptake. The dose-dependent inhibitory effects were present after 24 h of incubation with both low- and high-Pi media. Michaelis-Menten kinetics in OK cells identified an ~2.4-fold higher Km for Pi in response to Npt2a inhibition with no significant change in apparent Vmax. Higher parathyroid hormone concentrations decreased Pi uptake equivalent to the maximal inhibitory effect of Npt2a inhibitor. In vivo, the Npt2a inhibitor induced a dose-dependent increase in urinary Pi excretion in wild-type mice (ED50: ~23 mg/kg), which was completely absent in Npt2a-/- mice, alongside a lack of decrease in plasma Pi. Of note, the Npt2a inhibitor-induced dose-dependent increase in urinary Na+ excretion was still present in Npt2a-/- mice, a response possibly mediated by an off-target acute inhibitory effect of the Npt2a inhibitor on open probability of the epithelial Na+ channel in the cortical collecting duct.


Assuntos
Fosfatos/metabolismo , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIa/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Linhagem Celular , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Gambás , Hormônio Paratireóideo/farmacologia , Técnicas de Patch-Clamp , Distribuição Aleatória , Proteínas Cotransportadoras de Sódio-Fosfato Tipo IIa/genética
4.
J Appl Oral Sci ; 28: e20190690, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32348445

RESUMO

Objective To investigate the effects of intro-oral injection of parathyroid hormone (PTH) on tooth extraction wound healing in hyperglycemic rats. Methodology 60 male Sprague-Dawley rats were randomly divided into the normal group (n=30) and DM group (n=30). Type 1 diabetes mellitus (DM) was induced by streptozotocin. After extracting the left first molar of all rats, each group was further divided into 3 subgroups (n=10 per subgroup), receiving the administration of intermittent PTH, continuous PTH and saline (control), respectively. The intermittent-PTH group received intra-oral injection of PTH three times per week for two weeks. A thermosensitive controlled-release hydrogel was synthesized for continuous-PTH administration. The serum chemistry was determined to evaluate the systemic condition. All animals were sacrificed after 14 days. Micro-computed tomography (Micro-CT) and histological analyses were used to evaluate the healing of extraction sockets. Results The level of serum glucose in the DM groups was significantly higher than that in the non-DM groups (p<0.05); the level of serum calcium was similar in all groups (p>0.05). Micro-CT analysis showed that the DM group had a significantly lower alveolar bone trabecular number (Tb.N) and higher trabecular separation (Tb.Sp) than the normal group (p<0.05). The histological analyses showed that no significant difference in the amount of new bone (hard tissue) formation was found between the PTH and non-PTH groups (p>0.05). Conclusions Bone formation in the extraction socket of the type 1 diabetic rats was reduced. PTH did not improve the healing of hard and soft tissues. The different PTH administration regimes (continuous vs. intermittent) had similar effect on tissue healing. These results demonstrated that the metabolic characteristics of the hyperglycemic rats produced a condition that was unable to respond to PTH treatment.


Assuntos
Hormônios e Agentes Reguladores de Cálcio/farmacologia , Diabetes Mellitus Experimental/fisiopatologia , Hormônio Paratireóideo/farmacologia , Extração Dentária/métodos , Alvéolo Dental/efeitos dos fármacos , Cicatrização/efeitos dos fármacos , Animais , Glicemia/análise , Cálcio/sangue , Hidrogéis , Masculino , Osteogênese/efeitos dos fármacos , Osteogênese/fisiologia , Distribuição Aleatória , Ratos Sprague-Dawley , Reprodutibilidade dos Testes , Ferida Cirúrgica/tratamento farmacológico , Fatores de Tempo , Alvéolo Dental/diagnóstico por imagem , Resultado do Tratamento , Microtomografia por Raio-X
5.
Toxicology ; 436: 152429, 2020 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-32156525

RESUMO

Excessive systemic uptake of inorganic fluorides causes disturbances of bone homeostasis. The mechanism of skeletal fluorosis is still uncertain. This study aimed to study the effect of fluoride on osteocyte-driven osteoclastogenesis and probe into the role of PTH in this process. IDG-SW3 cells seeded in collagen-coated constructs were developed into osteocyte-like cells through induction of mineral agents. Then, osteocyte-like cells were exposed to fluoride in the presence or absence of parathyroid hormone (PTH). Cell viability and their capacity to produce receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG) and sclerostin (SOST) were detected by MTT and Western blot assays, respectively. Finally, a transwell coculture system using osteocyte-like cells seeded in the low compartment, and osteoclast precursors added in the inserts was developed to observe the osteocyte-driven osteoclasogenesis response to fluoride with or without PTH, and the expression of molecules involved in this mechanism were measure by real time RT-PCR. Results showed that osteocytes withstood a toxic dose of fluoride, and yet PTH administration significantly reduced osteocytes viability. PTH amplified the effect of fluoride on the expression of osteoclastogenesis-related molecules in osteocyte, but did not enlarged the stimulating effect of fluoride on osteoclastogenesis drove by osteocyte coculture. Gene expression levels of TRAP, RANK, JNK and NFAtc1 significantly increased in fluoride affected osteoclast precursor cocultured with osteocyte-like cells. The impact of fluoride on osteocyte-driven osteoclast differentiation was stronger than that of PTH. In conclusion, osteocyte played a pivotal role on the mechanism underlying fluoride-affected osteoclastogenesis in which RANK-JNK-NFATc1 signaling pathway was involved, and PTH had a significant impact in this process.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Osteócitos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Fluoreto de Sódio/toxicidade , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Cocultura , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Fatores de Transcrição NFATC/genética , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/metabolismo , Osteoclastos/patologia , Osteócitos/metabolismo , Osteócitos/patologia , Osteoprotegerina/genética , Osteoprotegerina/metabolismo , Hormônio Paratireóideo/farmacologia , Ligante RANK/genética , Ligante RANK/metabolismo , Células RAW 264.7 , Receptor Ativador de Fator Nuclear kappa-B/genética , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Transdução de Sinais , Fosfatase Ácida Resistente a Tartarato/genética , Fosfatase Ácida Resistente a Tartarato/metabolismo
6.
Z Gerontol Geriatr ; 53(2): 163-170, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31950363

RESUMO

Osteoporotic bones heal more slowly and ineffectively than normal bones. A combination of antibodies against sclerosing protein (Scl-Ab), and parathyroid hormone 1-34 (PTH 1-34) may improve healing. A standard osteoporotic rat model was established 12 weeks after bilateral ovarian resection (OVX). Bone defects were created in the right femora of 80 rats, which were randomly divided into 4 groups: control, Scl-Ab (25 mg/kg twice weekly), PTH (60 µg/kg of PTH 1-34 three times a week) and PTH plus Scl-Ab. After 12 weeks of treatment the rats were sacrificed and blood and the distal femora were harvested for biochemical evaluation, histology, microcomputed tomography and biomechanical testing. Compared to the control group, monotherapy and combination therapy with PTH and/or Scl-Ab promoted the formation of new bone, enhanced maximum femoral loading and increased the levels of procollagen type I N­terminal propeptide (PINP) and osteocalcin. The administration of PTH + Scl-Ab maximally enhanced bone defect healing. Combination treatment was better than either treatment alone, indicating a synergistic effect.


Assuntos
Anticorpos/administração & dosagem , Proteínas Morfogenéticas Ósseas/imunologia , Remodelação Óssea/fisiologia , Consolidação da Fratura/efeitos dos fármacos , Hormônio Paratireóideo/uso terapêutico , Animais , Densidade Óssea/efeitos dos fármacos , Calo Ósseo/efeitos dos fármacos , Modelos Animais de Doenças , Quimioterapia Combinada , Feminino , Humanos , Ovariectomia , Hormônio Paratireóideo/farmacologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Microtomografia por Raio-X/métodos
7.
J Steroid Biochem Mol Biol ; 196: 105500, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31629064

RESUMO

Cyp27b1 and Cyp24a1 are reciprocally regulated in the kidney by the key hormones PTH, FGF23, and 1,25(OH)2D3. Our recent genomic studies in mice identified a complex kidney-specific enhancer module located within the introns of adjacent Mettl1 (M1) and Mettl21b (M21) genes that mediate basal and PTH induction of Cyp27b1 as well as suppression by FGF23 and 1,25(OH)2D3. Gross deletion of these segments in mice has severe consequences on skeletal health, and directly affects Cyp27b1 expression in the kidney. Deletion of both M1 and M21 submodules together fully eliminates basal Cyp27b1 expression in the kidney, leading to a systemic and skeletal phenotype similar to that of the Cyp27b1-KO mouse due to depletion of 1,25(OH)2D3 and high PTH. Cyp24a1 levels in the double KO mouse were low due to compensatory regulation by elevated PTH and reduced FGF23. However, expression of Cyp27b1 and retention of its regulation by inflammation (LPS) in the NRTCs remained unperturbed. Dietary normalization of calcium, phosphate, PTH, and FGF23 rescues this aberrant phenotype and normalizes the skeletal issues. Cyp24a1 is controlled by its own unique enhancers for 1,25(OH)2D3, FGF23, and PTH. We were also able to eliminate these activities in mice. Collectively, the hormone-mediated enhancer regulation of both Cyp27b1 and Cyp24a1 in the kidney is responsible for the circulating levels of 1,25(OH)2D3 in the blood which in turn primarily affects calcium and phosphate regulation. Importantly, we can now manipulate this system with our enhancer deletion animal models to study 1,25(OH)2D3 production in non-renal target cells and tissues not only in disease, where it is known to affect the immune system, but also in healthy individuals. Here we will review our studies that have defined a finely balanced homeostatic control mechanism employed by PTH and FGF23 with catastrophic toxicity protection from 1,25(OH)2D3 in the genomic regulation of vitamin D metabolism and its accompanied control of mineral maintenance.


Assuntos
25-Hidroxivitamina D3 1-alfa-Hidroxilase/genética , Homeostase/fisiologia , Rim/metabolismo , Vitamina D3 24-Hidroxilase/genética , Vitamina D/metabolismo , 25-Hidroxivitamina D3 1-alfa-Hidroxilase/metabolismo , Animais , Calcitriol/farmacologia , Cálcio/metabolismo , Cálcio/farmacologia , Fatores de Crescimento de Fibroblastos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Humanos , Rim/efeitos dos fármacos , Hormônio Paratireóideo/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Vitamina D3 24-Hidroxilase/metabolismo
8.
J Cell Physiol ; 235(2): 1209-1221, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31276209

RESUMO

OBJECTIVE: Parathyroid hormone (PTH) is a main systemic mediator of calcium and phosphate homeostasis in the bone. Dental pulp stem cells (DPSCs) have been extensively studied in the regeneration of bone and tooth tissues. This paper aims to uncover the influences of PTH on the proliferative ability and osteo/odontogenic differentiation of DPSCs, as well as the underlying mechanisms. MATERIALS AND METHODS: The optimal concentration of PTH on DPSCs was determined by alkaline phosphatase (ALP) activity assay, ALP staining and western blot analysis. Proliferative ability and cell cycle distribution of DPSCs were analyzed by Cell counting kit-8, 5-ethynyl-20-deoxyuridine assay, and flow cytometry. Osteo/odontogenic capacity of DPSCs was evaluated and finally, the involvement of mitogen-activated protein kinase (MAPK) pathway was assessed. RESULTS: Purified DPSCs were obtained by enzymatic digestion, which presented a typical fibroblast-like morphology. 10-9 mol/L PTH was concerned as the optimal concentration for DPSCs induction. 10-9 mol/L PTH treatment did not change the proliferative rate of DPSCs (p > .05). Relative expressions of DSPP/DSPP, RUNX2/RUNX2, OSX/OSX, and ALP/ALP were upregulated in PTH-treated DPSCs relative to control group. Particularly, their mRNA/protein levels at Day 7 were markedly higher relative to those at Day 3 (p < .05 or p < .01). Mineralized nodules were formed after PTH induction, and calcium content increased by cetylpyridinium chloride quantitative analysis. Mechanistically, the protein levels of p-ERK and p-P38 significantly increased after PTH treatment, and the inhibitors targeting MAPK were identified that weakened the effects of PTH on the committed differentiation of DPSCs. CONCLUSIONS: PTH enhances the osteo/odontogenic differentiation capacity of DPSCs via ERK and P38 signaling pathways.


Assuntos
Polpa Dentária/citologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hormônio Paratireóideo/farmacologia , Células-Tronco/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Diferenciação Celular , MAP Quinases Reguladas por Sinal Extracelular/genética , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Células-Tronco/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/genética
9.
J Cell Physiol ; 235(2): 1480-1493, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31301073

RESUMO

Type 1 diabetes (T1D) is correlated with osteopenia primarily due to low bone formation. Parathyroid hormone (PTH) is a known anabolic agent for bone, the anabolic effects of which are partially mediated through the Wnt/ß-catenin signaling pathway. In the present study, we first determined the utility of intermittent PTH treatment in a streptozotocin-induced T1D mouse model. It was shown that the PTH-induced anabolic effects on bone mass and bone formation were attenuated in T1D mice compared with nondiabetic mice. Further, PTH treatment failed to activate ß-catenin signaling in osteoblasts of T1D mice and was unable to improve osteoblast proliferation and differentiation. Next, the Col1-3.2 kb-CreERTM; ß-cateninfx(ex3) mice were used to conditionally activate ß-catenin in osteoblasts by injecting tamoxifen, and we addressed whether or not preactivation of ß-catenin boosted the anabolic action of PTH on T1D-related bone loss. The results demonstrated that pretreatment with activation of osteoblastic ß-catenin followed by PTH treatment outperformed PTH or ß-catenin activation monotherapy and led to greatly improved bone structure, bone mass, and bone strength in this preclinical model of T1DM. Further analysis demonstrated that osteoblast proliferation and differentiation, as well as osteoprogenitors in the marrow, were all improved in the combination treatment group. These findings indicated a clear advantage of developing ß-catenin as a target to improve the efficacy of PTH in the treatment of T1D-related osteopenia.


Assuntos
Anabolizantes/farmacologia , Osso e Ossos/efeitos dos fármacos , Diabetes Mellitus Tipo 1/metabolismo , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Hormônio Paratireóideo/farmacologia , beta Catenina/metabolismo , Animais , Densidade Óssea/efeitos dos fármacos , Osso e Ossos/metabolismo , Complicações do Diabetes/metabolismo , Complicações do Diabetes/patologia , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 1/patologia , Camundongos
10.
J. appl. oral sci ; 28: e20190690, 2020. graf
Artigo em Inglês | LILACS, BBO - Odontologia | ID: biblio-1101255

RESUMO

Abstract Objective To investigate the effects of intro-oral injection of parathyroid hormone (PTH) on tooth extraction wound healing in hyperglycemic rats. Methodology 60 male Sprague-Dawley rats were randomly divided into the normal group (n=30) and DM group (n=30). Type 1 diabetes mellitus (DM) was induced by streptozotocin. After extracting the left first molar of all rats, each group was further divided into 3 subgroups (n=10 per subgroup), receiving the administration of intermittent PTH, continuous PTH and saline (control), respectively. The intermittent-PTH group received intra-oral injection of PTH three times per week for two weeks. A thermosensitive controlled-release hydrogel was synthesized for continuous-PTH administration. The serum chemistry was determined to evaluate the systemic condition. All animals were sacrificed after 14 days. Micro-computed tomography (Micro-CT) and histological analyses were used to evaluate the healing of extraction sockets. Results The level of serum glucose in the DM groups was significantly higher than that in the non-DM groups (p<0.05); the level of serum calcium was similar in all groups (p>0.05). Micro-CT analysis showed that the DM group had a significantly lower alveolar bone trabecular number (Tb.N) and higher trabecular separation (Tb.Sp) than the normal group (p<0.05). The histological analyses showed that no significant difference in the amount of new bone (hard tissue) formation was found between the PTH and non-PTH groups (p>0.05). Conclusions Bone formation in the extraction socket of the type 1 diabetic rats was reduced. PTH did not improve the healing of hard and soft tissues. The different PTH administration regimes (continuous vs. intermittent) had similar effect on tissue healing. These results demonstrated that the metabolic characteristics of the hyperglycemic rats produced a condition that was unable to respond to PTH treatment.


Assuntos
Animais , Masculino , Ratos , Hormônio Paratireóideo/farmacologia , Extração Dentária/métodos , Cicatrização/efeitos dos fármacos , Alvéolo Dental/efeitos dos fármacos , Diabetes Mellitus Experimental/fisiopatologia , Osteogênese/efeitos da radiação , Osteogênese/fisiologia , Glicemia/análise , Distribuição Aleatória , Cálcio/sangue , Ratos Sprague-Dawley , Hidrogéis , Ferida Cirúrgica/tratamento farmacológico
11.
PLoS One ; 14(12): e0226163, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31821371

RESUMO

OBJECTIVE: PTH1-34 (parathyroid hormone 1-34) is the only clinical drug to promote osteogenesis. MSCs (mesenchymal stem cells) have multidirectional differentiation potential and are closely related to fracture healing. This study was to explore the effects of PTH1-34 on proliferation and differentiation of endothelial cells and MSCs in vitro, and on angiogenesis, and MSCs migration during fracture healing in vivo. METHODS: Mice with stabilized fracture were assigned to 4 groups: CON, PTH (PTH1-34 40 µg/kg/day), MSC (transplanted with 105 MSCs), PTH+MSCs. Mice were sacrificed 14 days after fracture, and callus tissues were harvested for microCT scan and immunohistochemistry analysis. The effects of PTH1-34 on angiogenesis, and MSCs differentiation and migration were assessed by wound healing, tube formation and immunofluorescence staining. RESULTS: Treatment with either PTH1-34, or MSCs promoted bone healing and vascular formation in fracture callus. The callus bone mass, bone volume, and bone mineral density were all greater in PTH and/or MSC groups than they were in CON (p<0.05). PTH1-34 increased small vessels formation (diameter ≤50µm), whereas MSCs increased the large ones (diameter >50µm). Expression of CD31 within calluses and trabecular bones were significantly higher in PTH1-34 treated group than that of not (p<0.05). Expression of CD31, VEGFR, VEGFR2, and vWF was upregulated, and wound healing and tube formation were increased in MSCs treated with PTH1-34 compared to that of control. CONCLUSIONS: PTH1-34 improved the proliferation and differentiation of endothelial cells and MSCs, enhancing migration of MSCs to bone callus to promote angiogenesis and osteogenesis, and facilitating fracture healing.


Assuntos
Consolidação da Fratura/efeitos dos fármacos , Fraturas Ósseas/tratamento farmacológico , Células-Tronco Mesenquimais/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Hormônio Paratireóideo/farmacologia , Animais , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Feminino , Fraturas Ósseas/fisiopatologia , Células-Tronco Mesenquimais/fisiologia , Camundongos , Osteogênese/efeitos dos fármacos , Hormônio Paratireóideo/uso terapêutico
12.
FASEB J ; 33(12): 14394-14409, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31675485

RESUMO

Glucocorticoid (GC) therapy decreases bone mass and increases the risk of fractures. We investigated interactions between the GC dexamethasone (DEX) and the bone resorptive agents 1,25(OH)2-vitamin D3 (D3) and parathyroid hormone (PTH) on osteoclastogenesis. We observed a synergistic potentiation of osteoclast progenitor cell differentiation and formation of osteoclasts when DEX was added to either D3- or PTH-treated mouse bone marrow cell (BMC) cultures. Cotreatment of DEX with D3 or PTH increased gene encoding calcitonin receptor (Calcr), acid phosphatase 5, tartrate resistant (Acp5), cathepsin K (Ctsk), and TNF superfamily member 11 (Tnfsf11) mRNA, receptor activator of NF-κB ligand protein (RANKL), numbers of osteoclasts on plastic, and pit formation and release of C-terminal fragment of type I collagen from cells cultured on bone slices. Enhanced RANKL protein expression caused by D3 and DEX was absent in BMC from mice in which the GC receptor (GR) was deleted in stromal cells/osteoblasts. Synergistic interactions between DEX and D3 on RANKL and osteoclast formation were present in BMC from mice with attenuated GR dimerization. These data demonstrate that the GR cooperates with D3 and PTH signaling, causing massive osteoclastogenesis, which may explain the rapid bone loss observed with high dosages of GC treatment.-Conaway, H. H., Henning, P., Lie, A., Tuckermann, J., Lerner, U. H. Glucocorticoids employ the monomeric glucocorticoid receptor to potentiate vitamin D3 and parathyroid hormone-induced osteoclastogenesis.


Assuntos
Colecalciferol/farmacologia , Dexametasona/farmacologia , Osteogênese/efeitos dos fármacos , Hormônio Paratireóideo/farmacologia , Receptores de Glucocorticoides/metabolismo , Animais , Sinergismo Farmacológico , Deleção de Genes , Regulação da Expressão Gênica/efeitos dos fármacos , Camundongos , Ligante RANK/genética , Ligante RANK/metabolismo
13.
Biochem Pharmacol ; 169: 113627, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31476292

RESUMO

Parathyroid hormone (PTH)-related protein (PTHrP) (gene name Pthlh) was discovered as the factor responsible for the humoral hypercalcemia of malignancy. It shares such sequence similarity with PTH in the amino-terminal region that the two are equally able to act through a single G protein-coupled receptor, PTH1R. A number of biological activities are ascribed to domains of PTHrP beyond the amino-terminal domain. PTH functions as a circulating hormone, but PTHrP is generated locally in many tissues including bone, where it acts as a paracrine factor on osteoblasts and osteocytes. The present study compares how PTH and PTHrP influence cyclic AMP (cAMP) formation through adenylyl cyclase, the first event in cell activation through PTH1R. Brief exposure to full length PTHrP(1-141) in several osteoblastic cell culture systems was followed by sustained adenylyl cyclase activity for more than an hour after ligand washout. This effect was dose-dependent and was not found with shorter PTHrP or PTH peptides even though they were fully able to activate adenylyl cyclase with acute treatment. The persistent activation response to PTHrP(1-141) was seen also with later events in the cAMP/PKA pathway, including persistent activation of CRE-luciferase and sustained regulation of several CREB-responsive mRNAs, up to 24 h after the initial exposure. Pharmacologic blockade of endocytosis prevented the persistent activation of cAMP and gene responses. We conclude that full length PTHrP, the likely local physiological effector in bone, differs in intracellular action to PTH by undergoing endosomal translocation to induce a prolonged adenylyl cyclase activation in its target cells.


Assuntos
AMP Cíclico/biossíntese , Endocitose/fisiologia , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Fragmentos de Peptídeos/farmacologia , Adenilil Ciclases/metabolismo , Animais , Células Cultivadas , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Proteínas Quinases Dependentes de AMP Cíclico/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Hormônio Paratireóideo/farmacologia , Receptor Tipo 1 de Hormônio Paratireóideo/fisiologia
14.
Braz Oral Res ; 33: e086, 2019 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-31483052

RESUMO

Treatment of patients with bisphosphonate usage is a significant concern for oral surgeons because it interferes with jaw bone turnover and regeneration. In case of adverse effects manifesting related to bisphosphonate use, oral surgeons are usually treating and keep the patient's symptoms under control. In this study, we aimed to investigate a new treatment protocol for medication-related osteonecrosis of the jaw (MRONJ). This treatment protocol consisted of administering human parathyroid hormone (hPTH) loaded chitosan microspheres which were prepared by ionotropic gelation method or/and the prepared microspheres were suspended in a poloxamer gel. After in-vitro optimization studies, the efficacy of the chosen formulations was evaluated in-vivo studies. Zoledronic acid was administered daily to forty-eight adult female Sprague-Dawley rats, divided into four experimental groups, at a daily concentration of 0.11 mg/kg over three weeks to induce the MRONJ model. At the end of this period, maxillary left molar teeth were extracted. In the first group, the subjects received no treatment. In the negative control group, poloxamer hydrogel containing empty microspheres were immediately applied to the soft tissues surrounding the extraction socket. The treatment group-1 was treated with local injections of poloxamer hydrogel containing hPTH. The treatment group-2 was treated with a single local injection of poloxamer hydrogel containing hPTH-loaded chitosan microspheres. Both treatment groups received a total of 7 µg of hPTH at the end of the treatment protocol. Our study demonstrates successful attenuation of MRONJ through a local drug delivery system combined with hPTH, as opposed to previously attempted treatment strategies.


Assuntos
Conservadores da Densidade Óssea/farmacologia , Quitosana/farmacologia , Maxila/efeitos dos fármacos , Hormônio Paratireóideo/farmacologia , Animais , Osteonecrose da Arcada Osseodentária Associada a Difosfonatos/tratamento farmacológico , Osteonecrose da Arcada Osseodentária Associada a Difosfonatos/etiologia , Conservadores da Densidade Óssea/efeitos adversos , Conservadores da Densidade Óssea/uso terapêutico , Quitosana/uso terapêutico , Preparações de Ação Retardada , Feminino , Humanos , Maxila/patologia , Microesferas , Modelos Animais , Hormônio Paratireóideo/uso terapêutico , Poloxâmero/administração & dosagem , Poloxâmero/química , Ratos Sprague-Dawley , Ácido Zoledrônico/efeitos adversos
15.
J Am Chem Soc ; 141(36): 14210-14219, 2019 09 11.
Artigo em Inglês | MEDLINE | ID: mdl-31418572

RESUMO

Peptide agonists of GPCRs and other receptors are powerful signaling molecules with high potential as biological tools and therapeutics, but they are typically plagued by instability and short half-lives in vivo. Nature uses protein glycosylation to increase the serum stability of secreted proteins. However, these extracellular modifications are complex and heterogeneous in structure, making them an impractical solution. In contrast, intracellular proteins are subjected to a simple version of glycosylation termed O-GlcNAc modification. In our studies of this modification, we found that O-GlcNAcylation inhibits proteolysis, and strikingly, this stabilization occurs despite large distances in primary sequence (10-15 amino acids) between the O-GlcNAc and the site of cleavage. We therefore hypothesized that this "remote stabilization" concept could be useful to engineer the stability and potentially additional properties of peptide or protein therapeutics. Here, we describe the application of O-GlcNAcylation to two clinically important peptides: glucagon-like peptide-1 (GLP-1) and the parathyroid hormone (PTH), which respectively help control glucose and calcium levels in the blood. For both peptides, we found O-GlcNAcylated analogs that are equipotent to unmodified peptide in cell-based activation assays, while several GLP-1 analogs were biased agonists relative to GLP-1. As we predicted, O-GlcNAcylation can improve the stability of both GLP-1 and PTH in serum despite the fact that the O-GlcNAc can be quite remote from characterized sites of peptide cleavage. The O-GlcNAcylated GLP-1 and PTH also displayed significantly improved in vivo activity. Finally, we employed structure-based molecular modeling and receptor mutagenesis to predict how O-GlcNAcylation can be accommodated by the receptors and the potential interactions that contribute to peptide activity. This approach demonstrates the potential of O-GlcNAcylation for generating analogs of therapeutic peptides with enhanced proteolytic stability.


Assuntos
Peptídeo 1 Semelhante ao Glucagon/farmacologia , Hormônio Paratireóideo/farmacologia , Engenharia de Proteínas , Receptores Acoplados a Proteínas G/agonistas , Peptídeo 1 Semelhante ao Glucagon/sangue , Peptídeo 1 Semelhante ao Glucagon/química , Glicosilação , Humanos , Hormônio Paratireóideo/sangue , Hormônio Paratireóideo/química , Conformação Proteica , Receptores Acoplados a Proteínas G/metabolismo
16.
PLoS One ; 14(7): e0219575, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31291372

RESUMO

Parathyroid hormone (PTH) is an anabolic bone drug approved by the US Food and Drug Administration (FDA) to treat osteoporosis. However, previous studies using cross-sectional designs have reported variable and sometimes contradictory results. The aim of the present study was to quantify the localized effect of PTH on the structural and densitometric behaviors of mouse tibia and their links with the global mechanical behavior of bone using a novel spatiotemporal image analysis approach and a finite element analysis technique. Twelve female C57BL/6J mice were divided into two groups: the control and PTH treated groups. The entire right tibiae were imaged using an in vivo micro-computed tomography (µCT) system eight consecutive times. Next, the in vivo longitudinal tibial µCT images were rigidly registered and divided into 10 compartments across the entire tibial space. The bone volume (BV), bone mineral content (BMC), bone tissue mineral density (TMD), and tibial endosteal and periosteal areas (TEA and TPA) were quantified in each compartment. Additionally, finite element models of all the tibiae were generated to analyze the failure behavior of the tibia. It was found that both the BMC and BV started to increase in the proximal tibial region, and then the increases extended to the entire tibial region after two weeks of treatment (p < 0.05). PTH intervention significantly reduced the TEA in most tibial compartments after two weeks of treatment, and the TPA increased in most tibial regions after four weeks of treatment (p < 0.05). Tibial failure loads significantly increased after three weeks of PTH treatment (p < 0.01). The present study provided the first evidence of the localized effect of PTH on bone structural and densitometric properties, as well as their links with the global mechanical behaviors of bone, which are important pieces of information for unveiling the mechanism of PTH intervention.


Assuntos
Densidade Óssea/efeitos dos fármacos , Osteoporose/tratamento farmacológico , Fraturas por Osteoporose/prevenção & controle , Hormônio Paratireóideo/farmacologia , Tíbia/efeitos dos fármacos , Animais , Densitometria/métodos , Modelos Animais de Doenças , Feminino , Análise de Elementos Finitos , Camundongos , Camundongos Endogâmicos C57BL , Osteoporose/complicações , Fraturas por Osteoporose/etiologia , Hormônio Paratireóideo/uso terapêutico , Análise Espaço-Temporal , Tíbia/diagnóstico por imagem , Tíbia/fisiologia , Suporte de Carga , Microtomografia por Raio-X
17.
Biochem Biophys Res Commun ; 516(2): 551-557, 2019 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-31235257

RESUMO

Developing methods for regenerating bones and uncovering the molecular mechanism underlying bone formation have great significance to human health. In the last decade, people have been using adipose-derived stem cells (ADSCs), that are capable of multilineage differentiation, to reconstruct defected bones. Uncovering the molecular mechanisms of the osteoblast differentiation of ADSCs will provide more understanding of ADSCs in the application of bone regeneration and perhaps new methods for osteoporosis treatment. Here we studied how parathyroid hormone (PTH1-34) acts on osteoinduced ADSCs to orchestrate bone formation and how Wnt4 signaling is involved in PTH-promoted bone formation from ADSCs. We found that PTH1-34 can phosphorylate SIK2, upregulate RANKL and downregulate SOST, thereby upregulating Wnt4 to promote the osteogenesis process of ADSCs. Though the knockdown of Wnt4 with shRNA interference barely affects the expression of upstream proteins (i.e., RANKL, SOST), it affects the expression of other downstream osteogenic proteins (i.e., Runx2, Osterix, and Osteocalcin), and then inhibit the osteogenesis process of ADSCs. Overall, PTH can affect the osteogenesis process of ADSCs by regulating SIK2 and Wnt4. We anticipate that this work will provide researchers with new insights into the bone regeneration with ADSCs.


Assuntos
Tecido Adiposo/citologia , Osteogênese , Hormônio Paratireóideo/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Células-Tronco/metabolismo , Proteína Wnt4/metabolismo , Animais , Masculino , Osteogênese/efeitos dos fármacos , Compostos de Fenilureia/farmacologia , Fosforilação/efeitos dos fármacos , Pirimidinas/farmacologia , Ratos Endogâmicos Lew , Transdução de Sinais/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos
18.
J Bone Miner Res ; 34(10): 1952-1963, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31173642

RESUMO

Intermittent parathyroid hormone (iPTH) treatment induces bone anabolic effects that result in the recovery of osteoporotic bone loss. Human PTH is usually given to osteoporotic patients because it induces osteoblastogenesis. However, the mechanism by which PTH stimulates the expansion of stromal cell populations and their maturation toward the osteoblastic cell lineage has not be elucidated. Mouse genetic lineage tracing revealed that iPTH treatment induced osteoblastic differentiation of bone marrow (BM) mesenchymal stem and progenitor cells (MSPCs), which carried the leptin receptor (LepR)-Cre. Although these findings suggested that part of the PTH-induced bone anabolic action is exerted because of osteoblastic commitment of MSPCs, little is known about the in vivo mechanistic details of these processes. Here, we showed that LepR+ MSPCs differentiated into type I collagen (Col1)+ mature osteoblasts in response to iPTH treatment. Along with osteoblastogenesis, the number of Col1+ mature osteoblasts increased around the bone surface, although most of them were characterized as quiescent cells. However, the number of LepR-Cre-marked lineage cells in a proliferative state also increased in the vicinity of bone tissue after iPTH treatment. The expression levels of SP7/osterix (Osx) and Col1, which are markers for osteoblasts, were also increased in the LepR+ MSPCs population in response to iPTH treatment. In contrast, the expression levels of Cebpb, Pparg, and Zfp467, which are adipocyte markers, decreased in this population. Consistent with these results, iPTH treatment inhibited 5-fluorouracil- or ovariectomy (OVX)-induced LepR+ MSPC-derived adipogenesis in BM and increased LepR+ MSPC-derived osteoblasts, even under the adipocyte-induced conditions. Treatment of OVX rats with iPTH significantly affected the osteoporotic bone tissue and expansion of the BM adipose tissue. These results indicated that iPTH treatment induced transient proliferation of the LepR+ MSPCs and skewed their lineage differentiation from adipocytes toward osteoblasts, resulting in an expanded, quiescent, and mature osteoblast population. © 2019 American Society for Bone and Mineral Research.


Assuntos
Adipogenia/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Hormônio Paratireóideo/farmacologia , Receptores para Leptina/metabolismo , Animais , Feminino , Camundongos , Camundongos Transgênicos , Hormônio Paratireóideo/genética , Receptores para Leptina/genética
19.
J Biol Chem ; 294(25): 9722-9733, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31068415

RESUMO

Bone is a highly metabolic organ that undergoes continuous remodeling to maintain its structural integrity. During development, bones, in particular osteoblasts, rely on glucose uptake. However, the role of glucose metabolism in osteocytes is unknown. Osteocytes are terminally differentiated osteoblasts orchestrating bone modeling and remodeling. In these cells, parathyroid hormone (PTH) suppresses Sost/sclerostin expression (a potent inhibitor of bone formation) by promoting nuclear translocation of class IIa histone deacetylase (HDAC) 4 and 5 and the repression of myocyte enhancer factor 2 (MEF2) type C. Recently, Scriptaid, an HDAC complex co-repressor inhibitor, has been shown to induce MEF2 activation and exercise-like adaptation in mice. In muscles, Scriptaid disrupts the HDAC4/5 co-repressor complex, increases MEF2C function, and promotes cell respiration. We hypothesized that Scriptaid, by affecting HDAC4/5 localization and MEF2C activation, might affect osteocyte functions. Treatment of the osteocytic Ocy454-12H cells with Scriptaid increased metabolic gene expression, cell respiration, and glucose uptake. Similar effects were also seen upon treatment with PTH, suggesting that both Scriptaid and PTH can promote osteocyte metabolism. Similar to PTH, Scriptaid potently suppressed Sost expression. Silencing of HDAC5 in Ocy454-12H cells abolished Sost suppression but not glucose transporter type 4 (Glut4) up-regulation induced by Scriptaid. These results demonstrate that Scriptaid increases osteocyte respiration and glucose uptake by mechanisms independent of HDAC complex inhibition. In osteocytes, Scriptaid, similar to PTH, increases binding of HDAC5 to Mef2c with suppression of Sost but only partially increases receptor activator of NF-κB ligand (Rankl) expression, suggesting a potential bone anabolic effect.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Transportador de Glucose Tipo 4/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Hidroxilaminas/farmacologia , Osteócitos/metabolismo , Hormônio Paratireóideo/farmacologia , Quinolinas/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Hormônios e Agentes Reguladores de Cálcio/farmacologia , Células Cultivadas , Feminino , Transportador de Glucose Tipo 4/genética , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Fatores de Transcrição MEF2/genética , Fatores de Transcrição MEF2/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Osteócitos/citologia , Osteócitos/efeitos dos fármacos
20.
Am J Orthod Dentofacial Orthop ; 155(5): 670-680, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31053283

RESUMO

INTRODUCTION: This study investigated the effects of different doses of parathyroid hormone (PTH) on orthodontic tooth movement after mandibular ramus osteotomy and the associated dose-response relationship. METHODS: One-hundred twenty rabbits were divided into 2 experimental groups (A and B) and 2 control groups (control group and negative control group). An experimental model of mandibular ramus osteotomy with installation of an orthodontic tooth movement device was established in groups A and B and the control group. After surgery, groups A and B received intermittent subcutaneous injections of PTH, 20 and 40 µg/kg, respectively, and the control group received injections of normal saline solution. The negative control group underwent installation of the orthodontic tooth movement device without mandibular ramus osteotomy and received normal saline solution after surgery. Changes in expression of RANKL and RUNX2 in the periodontal tissues of the first molars were evaluated by means of immunohistochemical analysis and quantitative fluorescence polymerase chain reaction. RESULTS: Movement of the first molars was more rapid in group B than in group A in the 21 days after surgery. Significantly higher RANKL mRNA levels and lower RUNX2 mRNA levels were detected on the compression side of the periodontal tissues in groups A and B than in the control groups. There was a significant difference in RANKL and RUNX2 expression levels between group B and the control groups at all time points. CONCLUSIONS: Mandibular ramus osteotomy combined with high-dose PTH can increase catabolism on the compressed periodontal tissues, thereby accelerating remodeling of periodontal bone and promoting orthodontic tooth movement after surgery.


Assuntos
Osteotomia Mandibular , Hormônio Paratireóideo/farmacologia , Técnicas de Movimentação Dentária , Animais , Remodelação Óssea/fisiologia , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Relação Dose-Resposta a Droga , Humanos , Imuno-Histoquímica , Hormônio Paratireóideo/administração & dosagem , Reação em Cadeia da Polimerase , Ligante RANK/metabolismo , Coelhos
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