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1.
Nat Commun ; 15(1): 2529, 2024 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-38514612

RESUMO

Transcortical vessels (TCVs) provide effective communication between bone marrow vascular system and external circulation. Although osteocytes are in close contact with them, it is not clear whether osteocytes regulate the homeostasis of TCVs. Here, we show that osteocytes maintain the normal network of TCVs by transferring mitochondria to the endothelial cells of TCV. Partial ablation of osteocytes causes TCV regression. Inhibition of mitochondrial transfer by conditional knockout of Rhot1 in osteocytes also leads to regression of the TCV network. By contrast, acquisition of osteocyte mitochondria by endothelial cells efficiently restores endothelial dysfunction. Administration of osteocyte mitochondria resultes in acceleration of the angiogenesis and healing of the cortical bone defect. Our results provide new insights into osteocyte-TCV interactions and inspire the potential application of mitochondrial therapy for bone-related diseases.


Assuntos
Osteócitos , Osteócitos/metabolismo , Células Endoteliais , Osso e Ossos , Mitocôndrias
2.
Bone Res ; 12(1): 13, 2024 02 26.
Artigo em Inglês | MEDLINE | ID: mdl-38409111

RESUMO

Poor bone quality is a major factor in skeletal fragility in elderly individuals. The molecular mechanisms that establish and maintain bone quality, independent of bone mass, are unknown but are thought to be primarily determined by osteocytes. We hypothesize that the age-related decline in bone quality results from the suppression of osteocyte perilacunar/canalicular remodeling (PLR), which maintains bone material properties. We examined bones from young and aged mice with osteocyte-intrinsic repression of TGFß signaling (TßRIIocy-/-) that suppresses PLR. The control aged bone displayed decreased TGFß signaling and PLR, but aging did not worsen the existing PLR suppression in male TßRIIocy-/- bone. This relationship impacted the behavior of collagen material at the nanoscale and tissue scale in macromechanical tests. The effects of age on bone mass, density, and mineral material behavior were independent of osteocytic TGFß. We determined that the decline in bone quality with age arises from the loss of osteocyte function and the loss of TGFß-dependent maintenance of collagen integrity.


Assuntos
Remodelação Óssea , Osteócitos , Humanos , Idoso , Masculino , Animais , Camundongos , Remodelação Óssea/fisiologia , Colágeno/farmacologia , Envelhecimento , Fator de Crescimento Transformador beta/farmacologia
3.
Eur J Pharmacol ; 967: 176364, 2024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38316249

RESUMO

Osteocytes, as mechanosensitive cells residing within bone tissue, hold a pivotal role in averting the occurrence and progression of osteoporosis. The apoptosis of osteocytes induced by unloading is one of the contributing factors to osteoporosis, although the underlying molecular mechanisms have not been fully elucidated. PTH 1-34 is known to promote bone formation and inhibit bone loss by targeting osteoblasts and osteocytes. However, it is not known whether PTH 1-34 can inhibit osteocyte apoptosis under unloading conditions and the molecular mechanisms involved. In this study, we employed a Random Positioning Machine (RPM) to emulate unloading conditions and cultured MLO-Y4 osteocyte-like cells, in order to unravel the mechanisms through which PTH 1-34 constrains osteocyte apoptosis amidst unloading circumstances. Our findings revealed that PTH 1-34 activated autophagy while suppressing endoplasmic reticulum stress by curtailing the generation of reactive oxygen species (ROS) in MLO-Y4 osteocyte-like cells during unloading conditions. By shedding light on the osteoporosis triggered by skeletal unloading, this study contributes vital insights that may pave the way for the development of pharmacological interventions.


Assuntos
Osteócitos , Osteoporose , Apoptose , Autofagia , Osteoblastos , Hormônio Paratireóideo , Animais , Camundongos
4.
Bone ; 181: 117028, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38309412

RESUMO

INTRODUCTION: Osteocytes modulate bone adaptation in response to mechanical stimuli imparted by the deforming bone tissue in which they are encased by communicating with osteoclasts and osteoblasts as well as other osteocytes in the lacuna-canalicular network through secreted cytokines and chemokines. Understanding the transcriptional response of osteocytes to mechanical stimulation in situ could identify new targets to inhibit bone loss or enhance bone formation in the presence of diseases like osteoporosis or metastatic cancer. We compared the mechanically regulated transcriptional response of osteocytes in trabecular bone following one or three days of controlled mechanical loading. METHODS: Porcine trabecular bone explants were cultured in a bioreactor for 48 h and subsequently loaded twice a day for one day or 3 days. RNA was isolated and sequenced, and the Tuxedo suite was used to identify differentially expressed genes and pathway analysis was conducted using Ingenuity Pathway Analysis (IPA). RESULTS: There were about 4000 differentially expressed genes following in situ culture relative to fresh bone. One hundred six genes were differentially expressed between the loaded and non-loaded groups following one day of loading compared to 913 genes after 3 d of loading. Only 45 of these were coincident between the two time points, indicating an evolving transcriptome. Clustering and principal component analysis indicated differences between the loaded and non-loaded groups after 3 d of loading. DISCUSSION: With sustained loading, there was a nine-fold increase in the number of differentially expressed genes, suggesting that osteocytes respond to loading through sequential activation of downstream genes in the same pathways. The differentially expressed genes were related to osteoarthritis, osteocyte, and chondrocyte signaling pathways. We noted that NFkB and TNF signaling are affected by early loading and this may drive downstream effects on the mechanobiological response. Moreover, these genes may regulate catabolic effects of mechanical disuse through their actions on pre-osteoclasts in the bone marrow niche.


Assuntos
Osso Esponjoso , Osteócitos , Animais , Suínos , Osteócitos/metabolismo , Transcriptoma/genética , Osso e Ossos , Osteoblastos , Estresse Mecânico
5.
Orthop Surg ; 16(3): 733-744, 2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38384174

RESUMO

OBJECTIVE: Glucocorticoid (GC) overuse is strongly associated with steroid-induced osteonecrosis of the femoral head (SINFH). However, the underlying mechanism of SINFH remains unclear. This study aims to investigate the effect of dexamethasone (Dex)-induced oxidative stress on osteocyte apoptosis and the underlying mechanisms. METHODS: Ten patients with SINFH and 10 patients with developmental dysplasia of the hips (DDH) were enrolled in our study. Sixty rats were randomly assigned to the Control, Dex, Dex + N-Acetyl-L-cysteine (NAC), Dex + Dibenziodolium chloride (DPI), NAC, and DPI groups. Magnetic resonance imaging (MRI) was used to examine edema in the femoral head of rats. Histopathological staining was performed to assess osteonecrosis. Immunofluorescence staining with TUNEL and 8-OHdG was conducted to evaluate osteocyte apoptosis and oxidative damage. Immunohistochemical staining was carried out to detect the expression of NOX1, NOX2, and NOX4. Viability and apoptosis of MLO-Y4 cells were measured using the CCK-8 assay and TUNEL staining. 8-OHdG staining was conducted to detect oxidative stress. 2',7'-Dichlorodihydrofluorescein diacetate (DCFH-DA) staining was performed to measure reactive oxygen species (ROS). The expression of NOX1, NOX2, and NOX4 in MLO-Y4 cells was analyzed by Western blotting. Multiple comparisons were performed using one-way analysis of variance (ANOVA). RESULTS: In patients and the rat model, hematoxylin-eosin (HE) staining revealed a significantly higher rate of empty lacunae in the SINFH group than in the DDH group. Immunofluorescence staining indicated a significant increase in TUNEL-positive cells and 8-OHdG-positive cells in the SINFH group compared to the DDH group. Immunohistochemical staining demonstrated a significant increase in the expression of NOX1, NOX2, and NOX4 proteins in SINFH patients compared to DDH patients. Moreover, immunohistochemical staining showed a significant increase in the proportion of NOX2-positive cells compared to the Control group in the femoral head of rats. In vitro, Dex significantly inhibited the viability of osteocyte cells and induced apoptosis. After Dex treatment, the intracellular ROS level increased. However, Dex treatment did not alter the expression of NOX proteins in vitro. Additionally, NAC and DPI inhibited the generation of intracellular ROS and partially alleviated osteocyte apoptosis in vivo and in vitro. CONCLUSION: This study demonstrates that GC promotes apoptosis of osteocyte cells through ROS-induced oxidative stress. Furthermore, we found that the increased expression of NOXs induced by GC serves as an important source of ROS generation.


Assuntos
Osteócitos , Osteonecrose , Humanos , Ratos , Animais , Dexametasona/efeitos adversos , Espécies Reativas de Oxigênio/metabolismo , Cabeça do Fêmur , Glucocorticoides/efeitos adversos , Apoptose , Esteroides/efeitos adversos , Esteroides/metabolismo , Estresse Oxidativo
6.
Bone ; 181: 117026, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38325651

RESUMO

Disuse osteoporosis is a prevalent complication among patients afflicted with rheumatoid arthritis (RA). Although reports have shown that the antirheumatic drug iguratimod (IGU) ameliorates osteoporosis in RA patients, details regarding its effects on osteocytes remain unclear. The current study examined the effects of IGU on osteocytes using a mouse model of disuse-induced osteoporosis, the pathology of which crucially involves osteocytes. A reduction in distal femur bone mass was achieved after 3 weeks of hindlimb unloading in mice, which was subsequently reversed by intraperitoneal IGU treatment (30 mg/kg; five times per week). Histology revealed that hindlimb-unloaded (HLU) mice had significantly increased osteoclast number and sclerostin-positive osteocyte rates, which were suppressed by IGU treatment. Moreover, HLU mice exhibited a significant decrease in osteocalcin-positive cells, which was attenuated by IGU treatment. In vitro, IGU suppressed the gene expression of receptor activator of NF-κB ligand (RANKL) and sclerostin in MLO-Y4 and Saos-2 cells, which inhibited osteoclast differentiation of mouse bone marrow cells in cocultures. Although IGU did not affect the nuclear translocation or transcriptional activity of NF-κB, RNA sequencing revealed that IGU downregulated the expression of early growth response protein 1 (EGR1) in osteocytes. HLU mice showed significantly increased EGR1- and tumor necrosis factor alpha (TNFα)-positive osteocyte rates, which were decreased by IGU treatment. EGR1 overexpression enhanced the gene expression of TNFα, RANKL, and sclerostin in osteocytes, which was suppressed by IGU. Contrarily, small interfering RNA-mediated suppression of EGR1 downregulated RANKL and sclerostin gene expression. These findings indicate that IGU inhibits the expression of EGR1, which may downregulate TNFα and consequently RANKL and sclerostin in osteocytes. These mechanisms suggest that IGU could potentially be used as a treatment option for disuse osteoporosis by targeting osteocytes.


Assuntos
Cromonas , Osteoporose , Sulfonamidas , Fator de Necrose Tumoral alfa , Animais , Humanos , Fator de Necrose Tumoral alfa/metabolismo , Osteócitos/metabolismo , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Linhagem Celular , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , Proteína 1 de Resposta de Crescimento Precoce/farmacologia , Ligantes , Osteoclastos/metabolismo , NF-kappa B/metabolismo , Osteoporose/tratamento farmacológico , Osteoporose/metabolismo , Ligante RANK/metabolismo
7.
Bone ; 180: 116998, 2024 03.
Artigo em Inglês | MEDLINE | ID: mdl-38184100

RESUMO

Osteon morphology provides valuable information about the interplay between different processes involved in bone remodelling. The correct quantitative interpretation of these morphological features is challenging due to the complexity of interactions between osteoblast behaviour, and the evolving geometry of cortical pores during pore closing. We present a combined experimental and mathematical modelling study to provide insights into bone formation mechanisms during cortical bone remodelling based on histological cross-sections of quiescent human osteons and hypothesis-testing analyses. We introduce wall thickness asymmetry as a measure of the local asymmetry of bone formation within an osteon and examine the frequency distribution of wall thickness asymmetry in cortical osteons from human iliac crest bone samples from women 16-78 years old. Our measurements show that most osteons possess some degree of asymmetry, and that the average degree of osteon asymmetry in cortical bone evolves with age. We then propose a comprehensive mathematical model of cortical pore filling that includes osteoblast secretory activity, osteoblast elimination, osteoblast embedment as osteocytes, and osteoblast crowding and redistribution along the bone surface. The mathematical model is first calibrated to symmetric osteon data, and then used to test three mechanisms of asymmetric wall formation against osteon data: (i) delays in the onset of infilling around the cement line; (ii) heterogeneous osteoblastogenesis around the bone perimeter; and (iii) heterogeneous osteoblast secretory rate around the bone perimeter. Our results suggest that wall thickness asymmetry due to off-centred Haversian pores within osteons, and that nonuniform lamellar thicknesses within osteons are important morphological features that can indicate the prevalence of specific asymmetry-generating mechanisms. This has significant implications for the study of disruptions of bone formation as it could indicate what biological bone formation processes may become disrupted with age or disease.


Assuntos
Ósteon , Osteoblastos , Humanos , Feminino , Adolescente , Adulto Jovem , Adulto , Pessoa de Meia-Idade , Idoso , Ósteon/anatomia & histologia , Osso e Ossos , Osteócitos , Osso Cortical
8.
Curr Osteoporos Rep ; 22(1): 105-114, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-38198034

RESUMO

PURPOSE OF REVIEW: The formation of a pre-metastatic niche (PMN), in which primary cancer cells prime the distant site to be favorable to their engraftment and survival, may help explain the strong osteotropism observed in multiple cancers, such as breast and prostate. PMN formation, which includes extracellular matrix remodeling, increased angiogenesis and vascular permeability, enhanced bone marrow-derived cell recruitment and immune suppression, has mostly been described in soft tissues. In this review, we summarize current literature of PMN formation in bone. We also present evidence of a potential role for osteocytes to be the primary mediators of PMN development. RECENT FINDINGS: Osteocytes regulate the bone microenvironment in myriad ways beyond canonical bone tissue remodeling, including changes that contribute to PMN formation. Perilacunar tissue remodeling, which has been observed in both bone and non-bone metastatic cancers, is a potential mechanism by which osteocyte-cancer cell signaling stimulates changes to the bone microenvironment. Osteocytes also protect against endothelial permeability, including that induced by cancer cells, in a loading-mediated process. Finally, osteocytes are potent regulators of cells within the bone marrow, including progenitors and immune cells, and might be involved in this aspect of PMN formation. Osteocytes should be examined for their role in PMN formation.


Assuntos
Neoplasias , Osteócitos , Masculino , Humanos , Osteócitos/patologia , Remodelação Óssea , Neoplasias/patologia , Osso e Ossos , Transdução de Sinais , Microambiente Tumoral
9.
Biomater Sci ; 12(4): 919-932, 2024 Feb 13.
Artigo em Inglês | MEDLINE | ID: mdl-38231154

RESUMO

During bone formation, osteoblasts are embedded in a collagen-rich osteoid tissue and differentiate into an extensive 3D osteocyte network throughout the mineralizing matrix. However, how these cells dynamically remodel the matrix and undergo 3D morphogenesis remains poorly understood. Although previous reports investigated the impact of matrix stiffness in osteocyte morphogenesis, the role of matrix viscoelasticity is often overlooked. Here, we report a viscoelastic alginate-collagen interpenetrating network (IPN) hydrogel for 3D culture of murine osteocyte-like IDG-SW3 cells. The IPN hydrogels consist of an ionically crosslinked alginate network to tune stress relaxation as well as a permissive collagen network to promote cell adhesion and matrix remodeling. Two IPN hydrogels were developed with comparable stiffnesses (4.4-4.7 kPa) but varying stress relaxation times (t1/2, 1.5 s and 14.4 s). IDG-SW3 cells were pre-differentiated in 2D under osteogenic conditions for 14 days to drive osteoblast-to-osteocyte transition. Cellular mechanosensitivity to fluid shear stress (2 Pa) was confirmed by live-cell calcium imaging. After embedding in the IPN hydrogels, cells remained highly viable following 7 days of 3D culture. After 24 h, osteocytes in the fast-relaxing hydrogels showed the largest cell area and long dendritic processes. However, a significantly larger increase of some osteogenic markers (ALP, Dmp1, hydroxyapatite) as well as intercellular connections via gap junctions were observed in slow-relaxing hydrogels on day 14. Our results imply that fast-relaxing IPN hydrogels promote early cell spreading, whereas slow relaxation favors osteogenic differentiation. These findings may advance the development of 3D in vivo-like osteocyte models to better understand bone mechanobiology.


Assuntos
Hidrogéis , Osteócitos , Camundongos , Animais , Hidrogéis/metabolismo , Osteócitos/metabolismo , Osteogênese , Colágeno/metabolismo , Alginatos
10.
Sci Rep ; 14(1): 161, 2024 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-38168144

RESUMO

In the current study, the creation of a chitosan/alginate scaffold hydrogel with and without FeO-NPs or CuO-NPs was studied. From fetal ovine bone marrow mesenchymal stem cells (BM-MSCs) were isolated and cultivated. Their differentiation into osteocyte and adipose cells was investigated. Also, on the scaffolds, cytotoxicity and apoptosis were studied. To investigate the differentiation, treatment groups include: (1) BM-MSCs were plated in DMEM culture medium with high glucose containing 10% FBS and antibiotics (negative control); (2) BM-MSCs were plated in osteogenic differentiation medium (positive control); (3) positive control group + FeO-NPs, (4) positive control group + CuO-NPs; (5) BM-MSCs were plated in osteogenic differentiation medium on chitosan/alginate scaffold; (6) BM-MSCs were plated in osteogenic differentiation medium on chitosan/alginate/FeO-NPs scaffold; and (7) BM-MSCs were plated in osteogenic differentiation medium on chitosan/alginate/CuO-NPs scaffold. Alkaline phosphatase enzyme concentrations, mineralization rate using a calcium kit, and mineralization measurement by alizarin staining quantification were evaluated after 21 days of culture. In addition, qRT-PCR was used to assess the expression of the ALP, ColA, and Runx2 genes. When compared to other treatment groups, the addition of CuO-NPs in the chitosan/alginate hydrogel significantly increased the expression of the ColA and Runx2 genes (p < 0.05). However, there was no significant difference between the chitosan/alginate hydrogel groups containing FeO-NPs and CuO-NPs in the expression of the ALP gene. It appears that the addition of nanoparticles, in particular CuO-NPs, has made the chitosan/alginate scaffold more effective in supporting osteocyte differentiation.


Assuntos
Quitosana , Células-Tronco Mesenquimais , Ovinos , Animais , Quitosana/metabolismo , Tecidos Suporte , Osteogênese , Osteócitos , Alginatos/metabolismo , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Medula Óssea , Diferenciação Celular , Hidrogéis/metabolismo , Carneiro Doméstico , Células Cultivadas
11.
Sci Adv ; 10(3): eadi4298, 2024 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-38232158

RESUMO

Bone is one of the most common sites of tumor metastases. During the last step of bone metastasis, cancer cells colonize and disrupt the bone matrix, which is maintained mainly by osteocytes, the most abundant cells in the bone microenvironment. However, the role of osteocytes in bone metastasis is still unclear. Here, we demonstrated that osteocytes transfer mitochondria to metastatic cancer cells and trigger the cGAS/STING-mediated antitumor response. Blocking the transfer of mitochondria by specifically knocking out mitochondrial Rho GTPase 1 (Rhot1) or mitochondrial mitofusin 2 (Mfn2) in osteocytes impaired tumor immunogenicity and consequently resulted in the progression of metastatic cancer toward the bone matrix. These findings reveal the protective role of osteocytes against cancer metastasis by transferring mitochondria to cancer cells and potentially offer a valuable therapeutic strategy for preventing bone metastasis.


Assuntos
Neoplasias Ósseas , Osteócitos , Humanos , Osteócitos/metabolismo , Osso e Ossos , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Neoplasias Ósseas/secundário , GTP Fosfo-Hidrolases/genética , GTP Fosfo-Hidrolases/metabolismo , Mitocôndrias , Microambiente Tumoral
12.
Lasers Med Sci ; 39(1): 34, 2024 Jan 17.
Artigo em Inglês | MEDLINE | ID: mdl-38231300

RESUMO

PURPOSE: Long-term human immunodeficiency virus (HIV)-infected patients are considered at higher risk for osteoporosis. Among the various causes that lead these patients to lower bone health, there is the use of antiretroviral drugs (ARVs), especially protease inhibitors (PI), such as ritonavir (RTV). In this context, emerge the potential benefits of LED therapy, whose effects on bone cells are currently being extensively studied, showing a modulation in cell differentiation. However, it remains unclear if photobiostimulation might interfere with RTV effects on osteoblast differentiation. METHODS: In the present study, we investigated the effects of red LED (625 nm) irradiation (15 mW/cm2, 0.2 J/cm2, and 8 mW/cm2, 0.12 J/cm2) on osteoblast cell line MC3T3-E1 treated with RTV (2.5, 5, and 10 µg/mL). RESULTS: Our results indicated that red LED irradiation was able to reverse, or at least minimize, the deleterious effects of RTV on the osteoblasts. Neither the ARV treatments 5 and 10 µg/mL (104.4% and 95.01%) nor the LED protocols (100.3% and 105.7%) statistically altered cell viability, assessed by the MTT assay. Also, the alkaline phosphatase activity and mineralization showed a decrease in osteoblast activity followed by ARV exposure (39.3-73%), which was attenuated by LED in more than 70% with statistical significance (p < 0.05). CONCLUSION: In conclusion, photobiostimulation with red LED at 625 nm was associated with improved beneficial biological effects as a potential inducer of osteogenic activity on RTV-affected cells. This is the first study that investigated the benefits of red LED irradiation over ARV-treated in vitro osteoblasts.


Assuntos
Infecções por HIV , Ritonavir , Humanos , Ritonavir/efeitos adversos , Osteoblastos , Osteócitos , Infecções por HIV/tratamento farmacológico
13.
Dev Cell ; 59(2): 173-174, 2024 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-38262346

RESUMO

Although mechanical cues are known to influence the postnatal skeleton, the impact of bone cell mechano-transduction on early skeletal development remains less clear. In this issue of Developmental Cell, Collins et al. (2023) report that YAP/TAZ deletion in osteoblast precursors reduces Cxcl12 expression, leading to defects in bone vascularization.


Assuntos
Sinais (Psicologia) , Via de Sinalização Hippo , Humanos , Neovascularização Patológica , Osteoblastos , Osteócitos
14.
Cell Biochem Funct ; 42(1): e3924, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38269507

RESUMO

Designing biocompatible polymers using plant derivatives can be extremely useful in tissue engineering, nanomedicine, and many other fields of medicine. In this study, it was first looked into how chitosan/alginate scaffolds were made and characterized in the presence of berberine and barberry fruit extract. Second, the process of proliferation and differentiation of ovine fetal BM-MSCs (bone marrow-mesenchymal stem cells) was assessed on these scaffolds after BM-MSCs were extracted and confirmed by developing into osteocyte and adipose cells. To investigate the differentiation, treatment groups include (1) ovine fetal BM-MSCs were plated in Dulbecco's modified eagle medium culture medium with high glucose containing 10% fetal bovine serum and antibiotics (negative control), (2) ovine fetal BM-MSCs were plated in osteogenic differentiation medium (positive control group), (3) positive control group + barberry fruit extract, (4) positive control group + berberine, (5) ovine fetal BM-MSCs were plated in osteogenic differentiation medium on chitosan/alginate scaffold (hydrogel group), (6) ovine fetal BM-MSCs were plated in osteogenic differentiation medium on chitosan/alginate/barberry fruit extract scaffold (hydrogel group containing barberry fruit extract), and (7) ovine fetal BM-MSCs were plated in osteogenic differentiation medium on chitosan/alginate/berberine scaffold (hydrogel group containing berberine). Alkaline phosphatase (ALP) enzyme concentrations, mineralization rate using a calcium kit, and mineralization measurement by alizarin staining quantification were all found after 21 days of culture. In addition, real-time quantitative reverse transcription polymerase chain reaction was used to assess the expression of the ALP, COL1A2, and Runx2 genes. Days 5 and 7 had the lowest water absorption by the hydrogel scaffold containing barberry extract, which was significant in comparison to other groups (p < .05). Among the hydrogel scaffolds under study, the one containing barberry extract exhibited the lowest tensile strength, and this difference was statistically significant (p < .05). The chitosan/alginate hydrogel has the highest tensile strength of all of them. In comparison to the control and other treatment groups, the inclusion of berberine in the chitosan/alginate hydrogel significantly increased the expression of the ALP, Runx2, and COL1A2 genes (p < .05). The osteocyte differentiation of mesenchymal stem cells in in vitro settings appears to have been enhanced by the inclusion of berberine in the chitosan/alginate scaffold.


Assuntos
Berberina , Berberis , Quitosana , Células-Tronco Fetais , Ovinos , Animais , Quitosana/farmacologia , Subunidade alfa 1 de Fator de Ligação ao Core , Berberina/farmacologia , Osteócitos , Osteogênese , Alginatos/farmacologia , Hidrogéis
15.
Int J Mol Med ; 53(3)2024 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-38214344

RESUMO

Osteocyte function is critical for metabolism, remodelling and regeneration of bone tissue. In the present study, the roles of regulator of G protein signalling 18 (RGS18) were assessed in the regulation of osteocyte proliferation and bone formation. Target genes and signalling pathways were screened using the Gene Expression Omnibus (GEO) database and Gene Set Enrichment Analysis (GSEA). The function of RGS18 and the associated mechanisms were analysed by Cell Counting Kit 8 assay, 5­ethynyl­2'­deoxyuridine assay, flow cytometry, reverse transcription­quantitative PCR, western blotting and immunostaining. Overlap analysis of acutely injured subjects (AIS) and healthy volunteers (HVs) from the GSE93138 and GSE93215 datasets of the GEO database identified four genes: KIAA0825, ANXA3, RGS18 and LIPN. Notably, RGS18 was more highly expressed in peripheral blood samples from AIS than in those from HVs. Furthermore, RGS18 overexpression promoted MLO­Y4 and MC3T3­E1 cell viability, proliferation and S­phase arrest, but inhibited apoptosis by suppressing caspase­3/9 cleavage. Silencing RGS18 exerted the opposite effects. GSEA of GSE93138 revealed that RGS18 has the ability to regulate MAPK signalling. Treatment with the MEK1/2 inhibitor PD98059 reversed the RGS18 overexpression­induced osteocyte proliferation, and treatment with the ERK1/2 activator 12­O­tetradecanoylphorbol­13­acetate reversed the effects of RGS18 silencing on osteocyte proliferation. In conclusion, RGS18 may contribute to osteocyte proliferation and bone fracture healing via activation of ERK signalling.


Assuntos
MAP Quinases Reguladas por Sinal Extracelular , Osteócitos , Proteínas RGS , Humanos , Apoptose/genética , Proliferação de Células/genética , Proteínas de Ligação ao GTP , Transdução de Sinais , Animais , Camundongos , Células 3T3 , Proteínas RGS/genética
16.
Nat Commun ; 15(1): 890, 2024 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-38291059

RESUMO

Type 2 diabetes (T2D)-related fragility fractures represent an increasingly tough medical challenge, and the current treatment options are limited. Mechanical loading is essential for maintaining bone integrity, although bone mechano-responsiveness in T2D remains poorly characterized. Herein, we report that exogenous cyclic loading-induced improvements in bone architecture and strength are compromised in both genetically spontaneous and experimentally-induced T2D mice. T2D-induced reduction in bone mechano-responsiveness is directly associated with the weakened Ca2+ oscillatory dynamics of osteocytes, although not those of osteoblasts, which is dependent on PPARα-mediated specific reduction in osteocytic SERCA2 pump expression. Treatment with the SERCA2 agonist istaroxime was demonstrated to improve T2D bone mechano-responsiveness by rescuing osteocyte Ca2+ dynamics and the associated regulation of osteoblasts and osteoclasts. Moreover, T2D-induced deterioration of bone mechano-responsiveness is blunted in mice with osteocytic SERCA2 overexpression. Collectively, our study provides mechanistic insights into T2D-mediated deterioration of bone mechano-responsiveness and identifies a promising countermeasure against T2D-associated fragility fractures.


Assuntos
Diabetes Mellitus Tipo 2 , Osteócitos , Animais , Camundongos , Osso e Ossos , Cálcio/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Osteoblastos/metabolismo , Osteócitos/metabolismo
18.
Biomech Model Mechanobiol ; 23(1): 129-143, 2024 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-37642807

RESUMO

Exercise and physical activity exert mechanical loading on the bones which induces bone formation. However, the relationship between the osteocyte lacunar-canalicular morphology and mechanical stress experienced locally by osteocytes transducing signals for bone formation is not fully understood. In this study, we used computational modeling to predict the effect of canalicular density, the number of fluid inlets, and load direction on fluid flow shear stress (FFSS) and bone strains and how these might change following the microstructural deterioration of the lacunar-canalicular network that occurs with aging. Four distinct computational models were initially generated of osteocytes with either ten or eighteen dendrites using a fluid-structure interaction method with idealized geometries. Next, a young and a simulated aged osteocyte were developed from confocal images after FITC staining of the femur of a 4-month-old C57BL/6 mouse to estimate FFSS using a computational fluid dynamics approach. The models predicted higher fluid velocities in the canaliculi versus the lacunae. Comparison of idealized models with five versus one fluid inlet indicated that with four more inlets, one-half of the dendrites experienced FFSS greater than 0.8 Pa, which has been associated with osteogenic responses. Confocal image-based models of real osteocytes indicated a six times higher ratio of canalicular to lacunar surface area in the young osteocyte model than the simulated aged model and the average FFSS in the young model (FFSS = 0.46 Pa) was three times greater than the aged model (FFSS = 0.15 Pa). Interestingly, the surface area with FFSS values above 0.8 Pa was 23 times greater in the young versus the simulated aged model. These findings may explain the impaired mechano-responsiveness of osteocytes with aging.


Assuntos
Envelhecimento , Osteócitos , Camundongos , Animais , Osteócitos/fisiologia , Estresse Mecânico , Camundongos Endogâmicos C57BL , Simulação por Computador , Dendritos
19.
Adv Sci (Weinh) ; 11(2): e2305842, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37967351

RESUMO

Bone metastases are a common cause of suffering in breast and prostate cancer patients, however, the interaction between bone cells and cancer cells is poorly understood. Using a series of co-culture, conditioned media, human cancer spheroid, and organ-on-a-chip experiments, this study reveals that osteocytes suppress cancer cell proliferation and increase migration via tumor necrosis factor alpha (TNF-α) secretion. This action is regulated by osteocyte primary cilia and associated intraflagellar transport protein 88 (IFT88). Furthermore, it shows that cancer cells block this mechanism by secreting transforming growth factor beta (TGF-ß), which disrupts osteocyte cilia and IFT88 gene expression. This bi-directional crosstalk signaling between osteocytes and cancer cells is common to both breast and prostate cancer. This study also proposes that osteocyte inhibition of cancer cell proliferation decreases as cancer cells increase, producing more TGF-ß. Hence, a positive feedback loop develops accelerating metastatic tumor growth. These findings demonstrate the importance of cancer cell-osteocyte signaling in regulating breast and prostate bone metastases and support the development of therapies targeting this pathway.


Assuntos
Neoplasias Ósseas , Neoplasias da Próstata , Masculino , Humanos , Osteócitos/metabolismo , Cílios , Próstata , Neoplasias Ósseas/metabolismo , Fator de Crescimento Transformador beta/metabolismo
20.
Ultrastruct Pathol ; 48(2): 128-136, 2024 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-38115187

RESUMO

Radiation exposure is a major health concern due to bone involvement including mandible, causing deleterious effects on bone metabolism, and healing with an increasing risk of infection and osteoradionecrosis. This study aims to investigate the radiotherapy-induced microstructural changes in the human mandible by scanning electron microscopy (SEM). Mandibular cortical bone biopsies were obtained from control, irradiated, and patients with osteoradionecrosis (ORN). Bone samples were prepared for light microscopy and SEM. The SEM images were analyzed for the number of osteons, number of Haversian canal (HC), diameter of osteon (D.O), the diameter of HC (D.HC), osteonal wall thickness (O.W.Th), number of osteocytes, and number of osteocytic dendrites. The number of osteons, D.O, D.HC, O.W.Th, the number of osteocytes, and osteocytic dendrites were significantly decreased in both irradiated and ORN compared to controls (p < .05). The number of HCs decreased in irradiated and ORN bone compared to the control group. However, this was statistically not significant. The deleterious effect of radiation continues gradually altering the bone quality, structure, cellularity, and vascularity in the long term (>5 years mean radiation biopsy interval). The underlying microscopic damage in bone increases its susceptibility and contributes further to radiation-induced bone changes or even ORN.


Assuntos
Osteorradionecrose , Humanos , Microscopia Eletrônica de Varredura , Osteorradionecrose/etiologia , Osteorradionecrose/patologia , Osteócitos/patologia , Ósteon , Mandíbula/patologia
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