RESUMO
Parathyroid hormone-related protein (PTHrP) and its receptor are abundantly expressed throughout the renal parenchyma, where PTHrP exerts a modulatory action on renal function. PTHrP upregulation is a common event associated with the mechanism of renal injury and repair. However, no study has yet explored the putative excretion of PTHrP in urine, including its potential relationship with renal function. In the present study, we tested this hypothesis by studying the well-known rat model of acute renal injury induced by the chemotherapeutic agent cisplatin. Using Western blot analysis, we could detect a single protein band, corresponding to intact PTHrP, in the urine of both control and cisplatin-injected rats, whose levels were significantly higher in the latter group. PTHrP was detected in rat urine by dot blot, and its quantification with two specific ELISA kits showed that, compared with control rats, those treated with cisplatin displayed a significant increase in urinary PTHrP (expressed as the PTHrP-to-creatinine ratio or 24-h excretion). In addition, a positive correlation between urinary PTHrP excretion and serum creatinine was found in these animals. In conclusion, our data demonstrate that PTHrP is excreted in rat urine and that this excretion is higher with the decrease of renal function. This suggests that urinary PTHrP levels might be a renal function marker.
Assuntos
Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/urina , Antineoplásicos/toxicidade , Cisplatino/toxicidade , Proteína Relacionada ao Hormônio Paratireóideo/urina , Injúria Renal Aguda/patologia , Animais , Biomarcadores/urina , Creatinina/urina , Rim/patologia , Testes de Função Renal , Masculino , Ratos , Ratos WistarRESUMO
The potential involvement of type 2 diabetes mellitus (T2DM) as a risk factor for colon cancer (CC) has been previously reported. While several clinical studies show a higher incidence of CC and a lower survival rate in diabetics, others report no association. Our own experience indicates that diabetes does not seem to worsen the prognosis once the tumor is present. Despite this controversy, there are no wide-spectrum molecular studies that delve into the impact of T2DM-related mechanisms in colon carcinogenesis. Here, we present a transcriptomic and proteomic profiling of paired tumor and normal colon mucosa samples in a cohort of 42 CC patients, 23 of which have T2DM. We used gene set enrichment and network approaches to extract relevant pathways in diabetics, referenced them to current knowledge, and tested them using in vitro techniques. Through our transcriptomics approach, we identified an unexpected overlap of pathways overrepresented in diabetics compared to nondiabetics, in both tumor and normal mucosa, including diabetes-related metabolic and signaling processes. Proteomic approaches highlighted several cancer-related signaling routes in diabetics found only in normal mucosa, not in tumors. An integration of the transcriptome and proteome analyses suggested the deregulation of key pathways related to colon carcinogenesis which converged on tumor initiation axis TEAD/YAP-TAZ as a potential initiator of the process. In vitro studies confirmed upregulation of this pathway in nontumor colon cells under high-glucose conditions. In conclusion, T2DM associates with deregulation of cancer-related processes in normal colon mucosa adjacent to tissue which has undergone a malignant transformation. These data support that in diabetic patients, the local microenvironment in normal colon mucosa may be a factor driving field cancerization promoting carcinogenesis. Our results set a new framework to study links between diabetes and colon cancer, including a new role of the TEAD/YAP-TAZ complex as a potential driver.
Assuntos
Neoplasias do Colo/etiologia , Neoplasias do Colo/genética , Diabetes Mellitus Tipo 2/complicações , Animais , Carcinogênese/genética , Carcinogênese/patologia , Estudos de Coortes , Feminino , Regulação Neoplásica da Expressão Gênica , Genômica , Glucose/metabolismo , Humanos , Hiperglicemia/complicações , Mucosa Intestinal/patologia , Masculino , Camundongos Nus , Transdução de Sinais/genética , Transcriptoma/genética , Regulação para Cima/genéticaRESUMO
Denosumab (Dmab) is a humanized monoclonal antibody that blocks RANKL (receptor activator for nuclear factor κB ligand), thereby exerting a potent bone antiresorptive action. Dmab treatment leads to a dramatic and sustained increase in bone mass through mechanisms that are currently under debate. It is also a matter of controversy whether this potent action of Dmab could lead to intrabone dystrophic mineralization. Recent research has uncovered a possible anabolic role of Dmab involving RANKL-dependent reverse signaling in osteoblasts, and that bone marrow adipocytes can modulate osteoclastogenesis through the production of RANKL. We comment here on potential pathways which might account for the anabolic action of Dmab. The impact of this proposed mechanism needs to be addressed in further research.
Assuntos
Denosumab/farmacologia , Osteogênese/efeitos dos fármacos , Ligante RANK/antagonistas & inibidores , Animais , Densidade Óssea/efeitos dos fármacos , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/metabolismo , Humanos , Achados Incidentais , Osteoblastos/efeitos dos fármacos , Osteoblastos/fisiologia , Osteoclastos/efeitos dos fármacos , Osteoclastos/fisiologia , Ligante RANK/imunologiaRESUMO
BACKGROUND: Multiple observational studies suggest an increased risk of colon cancer in patients with diabetes mellitus (DM). This can theoretically be the result of an influence of the diabetic environment on carcinogenesis or the tumor biologic behavior. AIM: To gain insight into the influence of a diabetic environment on colon cancer characteristics and outcomes. MATERIAL AND METHODS: Retrospective analysis of clinical records in an academic tertiary care hospital with detailed analysis of 81 diabetic patients diagnosed of colon cancer matched with 79 non-diabetic colon cancer patients. The impact of streptozotocin-induced diabetes on the growth of colon cancer xenografts was studied in mice. RESULTS: The incidence of DM in 1,137 patients with colorectal cancer was 16%. The diabetic colon cancer cases and non-diabetic colon cancer controls were well matched for demographic and clinical variables. The ECOG Scale Performance Status was higher (worse) in diabetics (ECOG ≥1, 29.1% of controls vs 46.9% of diabetics, p = 0.02), but no significant differences were observed in tumor grade, adjuvant therapy, tumor site, lymphovascular invasion, stage, recurrence, death or cancer-related death. Moreover, no differences in tumor variables were observed between patients treated or not with metformin. In the xenograft model, tumor growth and histopathological characteristics did not differ between diabetic and nondiabetic animals. CONCLUSION: Our findings point towards a mild or negligible effect of the diabetes environment on colon cancer behavior, once cancer has already developed.
Assuntos
Neoplasias do Colo/patologia , Complicações do Diabetes/patologia , Idoso , Idoso de 80 Anos ou mais , Animais , Carcinogênese , Proliferação de Células , Transformação Celular Neoplásica , Neoplasias do Colo/complicações , Neoplasias do Colo/epidemiologia , Complicações do Diabetes/epidemiologia , Progressão da Doença , Humanos , Hiperglicemia/complicações , Masculino , Camundongos , Pessoa de Meia-Idade , Estudos Retrospectivos , Atenção Terciária à SaúdeRESUMO
Diabetes mellitus (DM) induces bone deterioration, while mechanical stimulation promotes osteocyte-driven bone formation. We aimed to evaluate the interaction of acute exposure (24 h) to high glucose (HG) with both the pro-survival effect conferred to osteocytic MLO-Y4 cells and osteoblastic MC3T3-E1 cells by mechanical stimulation and the interaction of these cells with osteoclast precursor RAW264.7 cells. We found that 24 h of HG (25 mM) pre-exposure prevented both cell survival and ERK and ß-catenin nuclear translocation upon mechanical stimulation by fluid flow (FF) (10 min) in both MLO-Y4 and MC3T3-E1 cells. However, migration of RAW 264.7 cells was inhibited by MLO-Y4 cell-conditioned medium (CM), but not by MC3T3-E1 cell-CM, with HG or FF. This inhibitory effect was associated with consistent changes in VEGF, RANTES, MIP-1α, MIP-1ß MCP-1, and GM-CSF in MLO-Y4 cell-CM. RAW264.7 proliferation was inhibited by MLO-Y4 CM under static or HG conditions, but it increased by FF-CM with or without HG. In addition, both FF and HG abrogated the capacity of RAW 264.7 cells to differentiate into osteoclasts, but in a different manner. Thus, HG-CM in static condition allowed formation of osteoclast-like cells, which were unable to resorb hydroxyapatite. In contrast, FF-CM prevented osteoclastogenesis even in HG condition. Moreover, HG did not affect basal RANKL or IL-6 secretion or their inhibition induced by FF in MLO-Y4 cells. In conclusion, this in vitro study demonstrates that HG exerts disparate effects on osteocyte mechanotransduction, and provides a novel mechanism by which DM disturbs skeletal metabolism through altered osteocyte-osteoclast communication.
Assuntos
Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Glucose/farmacologia , Mecanotransdução Celular/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Osteócitos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Células-Tronco/efeitos dos fármacos , Células 3T3 , Animais , Comunicação Celular/efeitos dos fármacos , Citocinas/metabolismo , Camundongos , Osteoclastos/metabolismo , Osteócitos/metabolismo , Estimulação Física , Células RAW 264.7 , Células-Tronco/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Worldwide deaths from diabetes mellitus (DM) and colorectal cancer increased by 90% and 57%, respectively, over the past 20 years. The risk of colorectal cancer was estimated to be 27% higher in patients with type 2 DM than in non-diabetic controls. However, there are potential confounders, information from lower income countries is scarce, across the globe there is no correlation between DM prevalence and colorectal cancer incidence and the association has evolved over time, suggesting the impact of additional environmental factors. The clinical relevance of these associations depends on understanding the mechanism involved. Although evidence is limited, insulin use has been associated with increased and metformin with decreased incidence of colorectal cancer. In addition, colorectal cancer shares some cellular and molecular pathways with diabetes target organ damage, exemplified by diabetic kidney disease. These include epithelial cell injury, activation of inflammation and Wnt/ß-catenin pathways and iron homeostasis defects, among others. Indeed, some drugs have undergone clinical trials for both cancer and diabetic kidney disease. Genome-wide association studies have identified diabetes-associated genes (e.g. TCF7L2) that may also contribute to colorectal cancer. We review the epidemiological evidence, potential pathophysiological mechanisms and therapeutic implications of the association between DM and colorectal cancer. Further studies should clarify the worldwide association between DM and colorectal cancer, strengthen the biological plausibility of a cause-and-effect relationship through characterization of the molecular pathways involved, search for specific molecular signatures of colorectal cancer under diabetic conditions, and eventually explore DM-specific strategies to prevent or treat colorectal cancer.
Assuntos
Neoplasias Colorretais/epidemiologia , Diabetes Mellitus Tipo 2/epidemiologia , Antineoplásicos/uso terapêutico , Neoplasias Colorretais/tratamento farmacológico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Humanos , Hipoglicemiantes/uso terapêuticoRESUMO
There is an unmet need to understand the mechanisms underlying skeletal deterioration in diabetes mellitus (DM) and to develop therapeutic approaches to treat bone fragility in diabetic patients. We demonstrate herein that mice with type 1 DM induced by streptozotocin exhibited low bone mass, inferior mechanical and material properties, increased bone resorption, decreased bone formation, increased apoptosis of osteocytes, and increased expression of the osteocyte-derived bone formation inhibitor Sost/sclerostin. Further, short treatment of diabetic mice with parathyroid hormone related protein (PTHrP)-derived peptides corrected these changes to levels undistinguishable from non-diabetic mice. In addition, diabetic mice exhibited reduced bone formation in response to mechanical stimulation, which was corrected by treatment with the PTHrP peptides, and higher prevalence of apoptotic osteocytes, which was reduced by loading or by the PTHrP peptides alone and reversed by a combination of loading and PTHrP peptide treatment. In vitro experiments demonstrated that the PTHrP peptides or mechanical stimulation by fluid flow activated the survival kinases ERKs and induced nuclear translocation of the canonical Wnt signaling mediator ß-catenin, and prevented the increase in osteocytic cell apoptosis induced by high glucose. Thus, PTHrP-derived peptides cross-talk with mechanical signaling pathways to reverse skeletal deterioration induced by DM in mice. These findings suggest a crucial role of osteocytes in the harmful effects of diabetes on bone and raise the possibility of targeting these cells as a novel approach to treat skeletal deterioration in diabetes. Moreover, our study suggests the potential therapeutic efficacy of combined pharmacological and mechanical stimuli to promote bone accrual and maintenance in diabetic subjects. © 2016 American Society for Bone and Mineral Research.
Assuntos
Osso e Ossos/anatomia & histologia , Osso e Ossos/fisiologia , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Experimental/fisiopatologia , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Proteínas Adaptadoras de Transdução de Sinal , Adiposidade/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Fenômenos Biomecânicos , Densidade Óssea/efeitos dos fármacos , Reabsorção Óssea/genética , Reabsorção Óssea/patologia , Osso e Ossos/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 1/fisiopatologia , Regulação da Expressão Gênica/efeitos dos fármacos , Glicoproteínas/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular , Masculino , Camundongos Endogâmicos C57BL , Modelos Biológicos , Tamanho do Órgão/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Osteócitos/efeitos dos fármacos , Osteócitos/metabolismo , Osteogênese/efeitos dos fármacos , Suporte de CargaRESUMO
Osteoarthritis (OA) is characterized by degenerative changes in the whole joint leading to physical disability in the elderly population. This condition is associated with altered bone metabolism in subchondral areas suggesting that therapeutic strategies aimed at modifying bone cell metabolism may be of interest. We have investigated the effects of several parathyroid hormone-related protein (PTHrP)-derived peptides (1-37): (N-terminal), (107-111) and (107-139) (C-terminal) on senescence features induced by inflammatory stress in human OA osteoblasts. Incubation of these primary cells with interleukin(IL)-1ß led to an increased expression of senescence markers senescence-associated-ß-galactosidase activity, γH2AX foci, p16, p21, p53, and caveolin-1. PTHrP (107-111) and PTHrP (107-139) significantly reduced all these parameters. Both peptides decreased the production of IL-6 and prostaglandin E2 which was the consequence of cyclo-oxygenase-2 downregulation. PTHrP (107-139) also reduced tumor necrosis factor-α release. These anti-inflammatory effects would be related to the reduction of nuclear factor-κB activation by both peptides and activator protein-1 by PTHrP (107-139). The three PTHrP peptides favored osteoblastic function although the C-terminal domain of PTHrP was more efficient than its N-terminal domain. Our data support an anti-senescence and anti-inflammatory role for the C-terminal moiety of PTHrP with potential applications in chronic inflammatory conditions such as OA.
Assuntos
Senescência Celular/fisiologia , Osteoartrite/metabolismo , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Idoso , Células Cultivadas , Dinoprostona/metabolismo , Feminino , Imunofluorescência , Humanos , Mediadores da Inflamação/farmacologia , Interleucina-1beta/farmacologia , Interleucina-6/metabolismo , Masculino , Osteoartrite/prevenção & controle , Osteoblastos/citologia , Fragmentos de Peptídeos/farmacologia , Reação em Cadeia da Polimerase em Tempo Real , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Oxidative damage is an important contributor to the morphological and functional changes in osteoporotic bone. Aging increases the levels of reactive oxygen species (ROS) that cause oxidative stress and induce osteoblast apoptosis. ROS modify several signaling responses, including mitogen-activated protein kinase (MAPK) activation, related to cell survival. Both parathyroid hormone (PTH) and its bone counterpart, PTH-related protein (PTHrP), can regulate MAPK activation by modulating MAPK phosphatase-1 (MKP1). Thus, we hypothesized that PTHrP might protect osteoblasts from ROS-induced apoptosis by targeting MKP1. In osteoblastic MC3T3-E1 and MG-63 cells, H2 O2 triggered p38, JNK, ERK and p66Shc phosphorylation, and cell apoptosis. Meanwhile, PTHrP (1-37) rapidly but transiently increased ERK and Akt phosphorylation without affecting p38, JNK, or p66Shc activation. H2 O2 -induced p38 and ERK phosphorylation and apoptosis were both decreased by pre-treatment with specific kinase inhibitors or PTHrP (1-37) in both osteoblastic cell types. These dephosphorylating and prosurvival actions of PTHrP (1-37) were prevented by a phosphatase inhibitor cocktail, the phosphatase MKP1 inhibitor sanguinarine or a MKP1 siRNA. PTHrP (1-37) promptly enhanced MKP1 protein and gene expression and MKP1-dependent catalase activity in osteoblastic cells. Furthermore, exposure to PTHrP (1-37) adsorbed in an implanted hydroxyapatite-based ceramic into a tibial defect in aging rats increased MKP1 and catalase gene expression in the healing bone area. Our findings demonstrate that PTHrP counteracts the pro-apoptotic actions of ROS by a mechanism dependent on MKP1-induced dephosphorylation of MAPKs in osteoblasts. J. Cell. Physiol. 232: 785-796, 2017. © 2016 Wiley Periodicals, Inc.
Assuntos
Citoproteção/efeitos dos fármacos , Fosfatase 1 de Especificidade Dupla/metabolismo , Osteoblastos/enzimologia , Osteoblastos/patologia , Estresse Oxidativo/efeitos dos fármacos , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Animais , Apoptose/efeitos dos fármacos , Osso e Ossos/efeitos dos fármacos , Catalase/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Peróxido de Hidrogênio/toxicidade , Masculino , Camundongos , Osteoblastos/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Ratos Wistar , Fatores de Tempo , Regulação para Cima/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Bone is one of the most adaptable tissues in the body as it is continuously subjected to load bearing. In fact, mechanical loading is an important regulator of bone mass. The skeleton adjusts to load by changing its mass, shape and microarchitecture, depending on the magnitude of the strain. Mechanical stimulation is necessary for the development of the skeleton, whereas in adults physiological levels of strain help maintain bone mass by reducing bone resorption. On the other hand, an excessive level of strain or bone disuse induces bone loss. Osteocytes are long-lived cells comprising more than 90% of bone cellularity, which are embedded in the bone matrix forming a functional syncytium extending to the bone surface. These cells are considered to be the main bone cells responsible for translating mechanical strain into regulatory signals for osteoblasts and osteoclasts, leading to adapting bone responses to environmental changes. In this review, we discuss the complexity and well-orchestrated events that occur in bone mechanotransduction, focusing on osteocyte viability as an important biological response in this respect. Elucidation of the molecular mechanisms of bone mechanotransduction and the key role of osteocytes is opening new avenues for the treatment of bone loss-related diseases.
Assuntos
Osso e Ossos/fisiologia , Mecanotransdução Celular , Osteócitos/citologia , Trifosfato de Adenosina/química , Animais , Densidade Óssea , Matriz Óssea , Reabsorção Óssea , Adesão Celular , Sobrevivência Celular , Citoesqueleto/metabolismo , Humanos , Óxido Nítrico/metabolismo , Osteoblastos/citologia , Osteoclastos/citologia , Osteócitos/metabolismo , Estresse MecânicoRESUMO
Reactive Oxygen Species (ROS) increase during aging, potentially affecting many tissues including brain, heart, and bone. ROS alter signaling pathways and constitute potential therapeutic targets to limit oxidative damaging effects in aging-associated diseases. Parathyroid hormone receptors (PTHR) are widely expressed and PTH is the only anabolic therapy for osteoporosis. The effects of oxidative stress on PTHR signaling and trafficking have not been elucidated. Here, we used Fluorescence Resonance Energy Transfer (FRET)-based cAMP, ERK, and calcium fluorescent biosensors to analyze the effects of ROS on PTHR signaling and trafficking by live-cell imaging. PTHR internalization and recycling were measured in HEK-293 cells stably transfected with HA-PTHR. PTH increased cAMP production, ERK phosphorylation, and elevated intracellular calcium. Pre-incubation with H2O2 reduced all PTH-dependent signaling pathways. These inhibitory effects were not a result of PTH oxidation since PTH incubated with H2O2 triggered similar responses. PTH promoted internalization and recycling of the PTHR. Both events were significantly reduced by H2O2 pre-incubation. These findings highlight the role of oxidation on PTHR signaling and trafficking, and suggest the relevance of ROS as a putative target in diseases associated with oxidative stress such as age-related osteoporosis.
Assuntos
Estresse Oxidativo , Receptores de Hormônios Paratireóideos/metabolismo , Transdução de Sinais , AMP Cíclico/metabolismo , Transferência Ressonante de Energia de Fluorescência , Células HEK293 , Humanos , Peróxido de Hidrogênio/metabolismo , Transporte ProteicoRESUMO
Diabetes mellitus (DM) and aging are associated with bone fragility and increased fracture risk. Both (1-37) N- and (107-111) C-terminal parathyroid hormone-related protein (PTHrP) exhibit osteogenic properties. We here aimed to evaluate and compare the efficacy of either PTHrP (1-37) or PTHrP (107-111) loaded into gelatin-glutaraldehyde-coated hydroxyapatite (HA-Gel) foams to improve bone repair of a transcortical tibial defect in aging rats with or without DM, induced by streptozotocin injection at birth. Diabetic old rats showed bone structural deterioration compared to their age-matched controls. Histological and µ-computerized tomography studies showed incomplete bone repair at 4 weeks after implantation of unloaded Ha-Gel foams in the transcortical tibial defects, mainly in old rats with DM. However, enhanced defect healing, as shown by an increase of bone volume/tissue volume and trabecular and cortical thickness and decreased trabecular separation, occurred in the presence of either PTHrP peptide in the implants in old rats with or without DM. This was accompanied by newly formed bone tissue around the osteointegrated HA-Gel implant and increased gene expression of osteocalcin and vascular endothelial growth factor (bone formation and angiogenic markers, respectively), and decreased expression of Sost gene, a negative regulator of bone formation, in the healing bone area. Our findings suggest that local delivery of PTHrP (1-37) or PTHrP (107-111) from a degradable implant is an attractive strategy to improve bone regeneration in aged and diabetic subjects. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2060-2070, 2016.
Assuntos
Regeneração Óssea/efeitos dos fármacos , Materiais Revestidos Biocompatíveis/farmacologia , Diabetes Mellitus Experimental/patologia , Sistemas de Liberação de Medicamentos , Durapatita/química , Implantes Experimentais , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Animais , Peso Corporal/efeitos dos fármacos , Gelatina/química , Regulação da Expressão Gênica/efeitos dos fármacos , Imageamento Tridimensional , Masculino , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Tíbia/diagnóstico por imagem , Tíbia/efeitos dos fármacos , Microtomografia por Raio-XRESUMO
The only bone anabolic agent currently available for osteoporosis treatment is parathyroid hormone (PTH)-either its N-terminal 1-34 fragment or the whole molecule of 1-84 aminoacids-whose intermittent administration stimulates new bone formation by targeting osteoblastogenesis and osteoblast survival. PTH-related protein (PTHrP) is an abundant factor in bone which shows N-terminal homology with PTH and thus exhibits high affinity for the same PTH type 1 receptor in osteoblasts. Therefore, it is not surprising that intermittently administered N-terminal PTHrP peptides induce bone anabolism in animals and humans. Furthermore, the C-terminal region of PTHrP also elicits osteogenic features in vitro in osteoblastic cells and in various animal models of osteoporosis. In this review, we discuss the current concepts about the cellular and molecular mechanisms whereby PTHrP may induce anabolic actions in bone. Pre-clinical studies and clinical data using N-terminal PTHrP analogs are also summarized, pointing to PTHrP as a promising alternative to current bone anabolic therapies.
Assuntos
Conservadores da Densidade Óssea/farmacologia , Osteoporose/tratamento farmacológico , Proteína Relacionada ao Hormônio Paratireóideo , Animais , Humanos , Osteogênese/efeitos dos fármacosRESUMO
HYPOTHESIS: Dental bleaching with H2O2 is a common daily practice in dentistry to correct discoloration of anterior teeth. The aim of this study has been to determine whether this treatment of human teeth affects growth, differentiation and activity of osteoclast-like cells, as well as the putative modulatory action of osteostatin and fibroblast growth factor 2 (FGF-2). EXPERIMENTS: Previously to the in vitro assays, structural, physical-chemical and morphological features of teeth after bleaching were studied. Osteoclast-like cells were cultured on human dentin disks, pre-treated or not with 38% H2O2 bleaching gel, in the presence or absence of osteostatin (100 nM) or FGF-2 (1 ng/ml). Cell proliferation and viability, intracellular content of reactive oxygen species (ROS), pro-inflammatory cytokine (IL-6 and TNFα) secretion and resorption activity were evaluated. FINDINGS: Bleaching treatment failed to affect either the structural or the chemical features of both enamel and dentin, except for slight morphological changes, increased porosity in the most superficial parts (enamel), and a moderate increase in the wettability degree. In this scenario, bleaching produced an increased osteoclast-like cell proliferation but decreased cell viability and cytokine secretion, while it augmented resorption activity on dentin. The presence of either osteostatin or FGF-2 reduced the osteoclast-like cell proliferation induced by bleaching. FGF-2 enhanced ROS content, whereas osteostatin decreased ROS but increased TNFα secretion. The bleaching effect on resorption activity was increased by osteostatin, but this effect was less evident with FGF-2. CONCLUSIONS: These findings further confirm the deleterious effects of tooth bleaching by affecting osteoclast growth and function as well as different modulatory actions of osteostatin and FGF-2.
Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Peróxido de Hidrogênio/farmacologia , Osteoclastos/efeitos dos fármacos , Proteína Relacionada ao Hormônio Paratireóideo/metabolismo , Fragmentos de Peptídeos/metabolismo , Clareamento Dental/efeitos adversos , Adolescente , Adsorção , Adulto , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Dentina/citologia , Dentina/efeitos dos fármacos , Dentina/metabolismo , Citometria de Fluxo , Humanos , Macrófagos/efeitos dos fármacos , Camundongos , Osteoclastos/citologia , Osteoclastos/metabolismo , Tamanho da Partícula , Espécies Reativas de Oxigênio/metabolismo , Propriedades de Superfície , Molhabilidade , Adulto JovemRESUMO
A promising bone graft substitute is porous titanium. Porous titanium, produced by selective laser melting (SLM), can be made as a completely open porous and load-bearing scaffold that facilitates bone regeneration through osteoconduction. In this study, the bone regenerative capacity of porous titanium is improved with a coating of osteostatin, an osteoinductive peptide that consists of the 107-111 domain of the parathyroid hormone (PTH)-related protein (PTHrP), and the effects of this osteostatin coating on bone regeneration were evaluated in vitro and in vivo. SLM-produced porous titanium received an alkali-acid-heat treatment and was coated with osteostatin through soaking in a 100 nM solution for 24 h or left uncoated. Osteostatin-coated scaffolds contained â¼0.1 µg peptide/g titanium, and in vitro 81% was released within 24 h. Human periosteum-derived osteoprogenitor cells cultured on osteostatin-coated scaffolds did not induce significant changes in osteogenic (alkaline phosphatase [ALP], collagen type 1 [Col1], osteocalcin [OCN], runt-related transcription factor 2 [Runx2]), or angiogenic (vascular endothelial growth factor [VEGF]) gene expression; however, it resulted in an upregulation of osteoprotegerin (OPG) gene expression after 24 h and a lower receptor activator of nuclear factor kappa-B ligand (RankL):OPG mRNA ratio. In vivo, osteostatin-coated, porous titanium implants increased bone regeneration in critical-sized cortical bone defects (p=0.005). Bone regeneration proceeded until 12 weeks, and femurs grafted with osteostatin-coated implants and uncoated implants recovered, respectively, 66% and 53% of the original femur torque strength (97±31 and 77±53 N·mm, not significant). In conclusion, the osteostatin coating improved bone regeneration of porous titanium. This effect was initiated after a short burst release and might be related to the observed in vitro upregulation of OPG gene expression by osteostatin in osteoprogenitor cells. Long-term beneficial effects of osteostatin-coated, porous titanium implants on bone regeneration or mechanical strength were not established here and may require optimization of the pace and dose of osteostatin release.
Assuntos
Regeneração Óssea/efeitos dos fármacos , Materiais Revestidos Biocompatíveis/farmacologia , Fêmur/patologia , Fêmur/fisiopatologia , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Fragmentos de Peptídeos/farmacologia , Titânio/farmacologia , Adolescente , Animais , Fenômenos Biomecânicos/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fêmur/diagnóstico por imagem , Fêmur/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Neovascularização Fisiológica/efeitos dos fármacos , Neovascularização Fisiológica/genética , Osteoblastos/efeitos dos fármacos , Osteoblastos/patologia , Osteoclastos/efeitos dos fármacos , Osteoclastos/patologia , Porosidade , Ratos Wistar , Microtomografia por Raio-XRESUMO
Osteocytes have a major role in the control of bone remodeling. Mechanical stimulation decreases osteocyte apoptosis and promotes bone accrual, whereas skeletal unloading is deleterious in both respects. PTH1R ablation or overexpression in osteocytes in mice produces trabecular bone loss or increases bone mass, respectively. The latter effect was related to a decreased osteocyte apoptosis. Here, the putative role of PTH1R activation in osteocyte protection conferred by mechanical stimulation was assessed. Osteocytic MLO-Y4 cells were subjected to mechanical stimuli represented by hypotonic shock (216 mOsm/kg) or pulsatile fluid flow (8 Hz, 10 dynes/cm(2)) for a short pulse (10 min), with or without PTH1R antagonists or after transfection with specific PTHrP or PTH1R siRNA. These mechanical stimuli prevented cell death induced within 6 hours by etoposide (50 µM), related to PTHrP overexpression; and this effect was abolished by the calcium antagonist verapamil (1 µM), a phospholipase C (PLC) inhibitor (U73122; 10 µM), and a PKA activation inhibitor, Rp-cAMPS (25 µM), in these cells. Each mechanical stimulus also rapidly induced ß-catenin stabilization and nuclear ERK translocation, which were inhibited by the PTH1R antagonist PTHrP(7-34) (1 µM), or PTH1R siRNA, and mimicked by PTHrP(1-36) (100 nM). Mechanical stretching by hypotonic shock did not affect cAMP production but rapidly (<1 min) stimulated Ca(i)(2+) transients in PTH1R-overexpressing HEK-293 cells and in MLO-Y4 cells, in which calcium signaling was unaffected by the presence of a PTHrP antiserum or PTHrP siRNA but inhibited by knocking down PTH1R. These novel findings indicate that PTH1R is an important component of mechanical signal transduction in osteocytic MLO-Y4 cells, and that PTH1R activation by PTHrP-independent and dependent mechanisms has a relevant role in the prosurvival action of mechanical stimulus in these cells.
Assuntos
Mecanotransdução Celular , Osteócitos/citologia , Osteócitos/metabolismo , Receptor Tipo 1 de Hormônio Paratireóideo/metabolismo , Animais , Cálcio/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , AMP Cíclico/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Soluções Hipotônicas/farmacologia , Mecanotransdução Celular/efeitos dos fármacos , Camundongos , Modelos Biológicos , Osteócitos/efeitos dos fármacos , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , beta Catenina/metabolismoRESUMO
Biopolymer-coated nanocrystalline hydroxyapatite (HA) made as macroporous foams which are degradable and flexible are promising candidates as orthopaedic implants. The C-terminal (107-111) epitope of parathyroid hormone-related protein (PTHrP) exhibits osteogenic properties. The main aim of this study was to evaluate whether PTHrP (107-111) loading into gelatin-glutaraldehyde biopolymer-coated HA (HAGlu) scaffolds would produce an optimal biomaterial for tissue engineering applications. HAGlu scaffolds with and without PTHrP (107-111) were implanted into a cavitary defect performed in both distal tibial metaphysis of adult rats. Animals were sacrificed after 4 weeks for histological, microcomputerized tomography and gene expression analysis of the callus. At this time, bone healing occurred only in the presence of PTHrP (107-111)-containing HAGlu implant, related to an increase in bone volume/tissue volume and trabecular thickness, cortical thickness and gene expression of osteocalcin and vascular cell adhesion molecule 1, but a decreased gene expression of Wnt inhibitors, SOST and dickkopf homolog 1. The autonomous osteogenic effect of the PTHrP (107-111)-loaded HAGlu scaffolds was confirmed in mouse and human osteoblastic cell cultures. Our findings demonstrate the advantage of loading PTHrP (107-111) into degradable HAGlu scaffolds for achieving an optimal biomaterial that is promising for low load bearing clinical applications.
Assuntos
Biopolímeros/química , Regeneração Óssea/efeitos dos fármacos , Materiais Revestidos Biocompatíveis , Durapatita/química , Gelatina/química , Glutaral/química , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Fragmentos de Peptídeos/farmacologia , Células 3T3 , Animais , Sequência de Bases , Primers do DNA , Camundongos , Microscopia Eletrônica de Varredura , Ratos , Reação em Cadeia da Polimerase em Tempo Real , Alicerces TeciduaisRESUMO
Insulin-like growth factor-I (IGF-I) deficiency causes growth delay, and IGF-I has been shown to partially mediate bone anabolism by parathyroid hormone (PTH). PTH-related protein (PTHrP) is abundant in bone, and has osteogenic features by poorly defined mechanisms. We here examined the capacity of PTHrP (1-36) and PTHrP (107-111) (osteostatin) to reverse the skeletal alterations associated with IGF-I deficiency. Igf1-null mice and their wild type littermates were treated with each PTHrP peptide (80 µg/Kg/every other day/2 weeks; 2 males and 4 females for each genotype) or saline vehicle (3 males and 3 females for each genotype). We found that treatment with either PTHrP peptide ameliorated trabecular structure in the femur in both genotypes. However, these peptides were ineffective in normalizing the altered cortical structure at this bone site in Igf1-null mice. An aberrant gene expression of factors associated with osteoblast differentiation and function, namely runx2, osteoprotegerin/receptor activator of NF-κB ligand ratio, Wnt3a , cyclin D1, connexin 43, catalase and Gadd45, as well as in osteocyte sclerostin, was found in the long bones of Igf1-null mice. These mice also displayed a lower amount of trabecular osteoblasts and osteoclasts in the tibial metaphysis than those in wild type mice. These alterations in Igf1-null mice were only partially corrected by each PTHrP peptide treatment. The skeletal expression of Igf2, Igf1 receptor and Irs2 was increased in Igf1-null mice, and this compensatory profile was further improved by treatment with each PTHrP peptide related to ERK1/2 and FoxM1 activation. In vitro, PTHrP (1-36) and osteostatin were effective in promoting bone marrow stromal cell mineralization in normal mice but not in IGF-I-deficient mice. Collectively, these findings indicate that PTHrP (1-36) and osteostatin can exert several osteogenic actions even in the absence of IGF-I in the mouse bone.
Assuntos
Fêmur/anormalidades , Transtornos do Crescimento/tratamento farmacológico , Perda Auditiva Neurossensorial/tratamento farmacológico , Fator de Crescimento Insulin-Like I/deficiência , Proteína Relacionada ao Hormônio Paratireóideo/uso terapêutico , Fragmentos de Peptídeos/uso terapêutico , Animais , Feminino , Fêmur/diagnóstico por imagem , Fêmur/efeitos dos fármacos , Fêmur/patologia , Regulação da Expressão Gênica/efeitos dos fármacos , Transtornos do Crescimento/patologia , Perda Auditiva Neurossensorial/patologia , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Camundongos , Osteoblastos/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoblastos/patologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Proteína Relacionada ao Hormônio Paratireóideo/química , Proteína Relacionada ao Hormônio Paratireóideo/metabolismo , Proteína Relacionada ao Hormônio Paratireóideo/farmacologia , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Fragmentos de Peptídeos/farmacologia , Fenótipo , Radiografia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
Parathyroid hormone-related protein (PTHrP) and its receptor type 1 (PTH1R) are extensively expressed in the kidney, where they are able to modulate renal function. Renal PTHrP is known to be overexpressed in acute renal injury. Recently, we hypothesized that PTHrP involvement in the mechanisms of renal injury might not be limited to conditions with predominant damage of the renal tubulointerstitium and might be extended to glomerular diseases, such as diabetic nephropathy (DN). In experimental DN, the overexpression of both PTHrP and the PTH1R contributes to the development of renal hypertrophy as well as proteinuria. More recent data have shown, for the first time, that PTHrP is upregulated in the kidney from patients with DN. Collectively, animal and human studies have shown that PTHrP acts as an important mediator of diabetic renal cell hypertrophy by a mechanism which involves the modulation of cell cycle regulatory proteins and TGF- ß 1. Furthermore, angiotensin II (Ang II), a critical factor in the progression of renal injury, appears to be responsible for PTHrP upregulation in these conditions. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Nefropatias Diabéticas/etiologia , Proteína Relacionada ao Hormônio Paratireóideo/metabolismo , Animais , Nefropatias Diabéticas/metabolismo , Humanos , Rim/metabolismo , Receptor Tipo 1 de Hormônio Paratireóideo/metabolismoRESUMO
Osteoporosis is characterized by low bone mineral density and/or poor bone microarchitecture leading to an increased risk of fractures. The skeletal alterations in osteoporosis are a consequence of a relative deficit of bone formation compared to bone resorption. Osteoporosis therapies have mostly relied on antiresorptive drugs. An alternative therapeutic approach for osteoporosis is currently available, based on the intermittent administration of parathyroid hormone (PTH). Bone anabolism caused by PTH therapy is mainly accounted for by the ability of PTH to increase osteoblastogenesis and osteoblast survival. PTH and PTH-related protein (PTHrP)-an abundant local factor in bone- interact with the common PTH type 1 receptor with similar affinities in osteoblasts. Studies mainly in osteoporosis rodent models and limited data in postmenopausal women suggest that N-terminal PTHrP peptides might be considered a promising bone anabolic therapy. In addition, putative osteogenic actions of PTHrP might be ascribed not only to its N-terminal domain but also to its PTH-unrelated C-terminal region. In this review, we discuss the underlying cellular and molecular mechanisms of the anabolic actions of PTH and the similar potential of PTH-related protein (PTHrP) to increase bone mass and improve bone regeneration.