RESUMO
BACKGROUND: The objective of the present study was to assess the hard and soft tissue differences of skeletal Class III malocclusion patients treated with orthodontic-orthognathic surgery treatment between two decompensation approaches including extraction of maxillary premolars in preoperative orthodontics and clockwise rotation of the maxilla in orthognathic surgery. METHODS: 22 skeletal Class III patients with the crowding of maxillary dental arch less than 3mm were included in this study. These patients were divided into two groups: extraction group and non-extraction group. Lateral cephalograms taken before preoperative orthodontic treatment and after postoperative orthodontic treatment were used to analyze the differences of hard and soft tissues between two groups. Independent t test was used to evaluate the differences of variables between extraction group and non-extraction group. RESULTS: After treatment, there was significant difference of Wits between extraction group and non-extraction group (- 4.34 mm vs - 2.82 mm, respectively, P <0.05). Co-Gn was significantly greater in non-extraction group than in extraction group (77.18 mm vs 71.58 mm, P <0.05). U1-SN and L1-MP in extraction group were significantly closer to the normal values than non-extraction group (P <0.05). Regarding the change of variables before and after orthodontic-orthognathic treatment, NLA (7.25° vs 1.46°, P <0.01) and G-Sn-Pog' (8.06° vs 4.62°, P <0.05) were significantly greater in extraction group than in non-extraction group. CONCLUSION: For patients with skeletal Class III malocclusion, extraction of maxillary premolars in preoperative orthodontic treatment can more effectively eliminate the dental compensation and achieve a more harmonious facial profile compared to clockwise rotation of the maxilla in orthognathic surgery. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .
Assuntos
Má Oclusão Classe III de Angle , Procedimentos Cirúrgicos Ortognáticos , Humanos , Mandíbula/cirurgia , Má Oclusão Classe III de Angle/cirurgia , Maxila/cirurgia , CefalometriaRESUMO
DNA nanomaterials have attracted ever-increasing attention over the past decades due to their incomparable programmability and multifunctionality. In particular, DNA dendrimer nanostructures, as a major research focus, have been applied in the fields of biosensing, therapeutics, and protein engineering, benefiting from their highly branched configuration. With the aid of specific recognition probes and inherent signal amplification, DNA dendrimers can achieve ultrasensitive detection of nucleic acids, proteins, cells, and other substances, such as lipopolysaccharides (LPS), adenosine triphosphate (ATP), and exosomes. By virtue of their void-containing structures and biocompatibility, DNA dendrimers can deliver drugs or functional nucleic acids into target cells in chemotherapy, immunotherapy, and gene therapy. Furthermore, DNA dendrimers are being applied in protein engineering for efficient directed evolution of proteins. This review summarizes the main research progress of DNA dendrimers, concerning their assembly methods and biomedical applications as well as the emerging challenges and perspectives for future research.
Assuntos
Pesquisa Biomédica , DNA/química , Dendrímeros/química , Nanoestruturas/química , Humanos , Tamanho da PartículaRESUMO
How the nervous system regulates bone remodeling is an exciting area of emerging research in bone biology. Accumulating evidence suggest that neurotransmitter-mediated inputs from neurons may act directly on osteoclasts. Dopamine is a neurotransmitter that can be released by hypothalamic neurons to regulate bone metabolism through the hypothalamic-pituitary-gonadal axis. Dopamine is also present in sympathetic nerves that penetrate skeletal structures throughout the body. It has been shown that dopamine suppresses osteoclast differentiation via a D2-like receptors (D2R)-dependent manner, but the intracellular secondary signaling pathway has not been elucidated. In this study, we found that cAMP-response element binding protein (CREB) activity responds to dopamine treatment during osteoclastogenesis. Considering the critical role of CREB in osteoclastogenesis, we hypothesize that CREB may be a critical target in dopamine's regulation of osteoclast differentiation. We confirmed that D2R is also present in RAW cells and activated by dopamine. Binding of dopamine to D2R inhibits the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) signaling pathway which ultimately decreases CREB phosphorylation during osteoclastogenesis. This was also associated with diminished expression of osteoclast markers that are downstream of CREB. Pharmacological activation of adenylate cyclase (to increase cAMP production) and PKA reverses the effect of dopamine on CREB activity and osteoclastogenesis. Therefore, we have identified D2R/cAMP/PKA/CREB as a candidate pathway that mediates dopamine's inhibition of osteoclast differentiation. These findings will contribute to our understanding of how the nervous and skeletal systems interact to regulate bone remodeling. This will enable future work toward elucidating the role of the nervous system in bone development, repair, aging, and degenerative disease.
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Diferenciação Celular/efeitos dos fármacos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Dopamina/farmacologia , Osteoclastos/metabolismo , Sistemas do Segundo Mensageiro/efeitos dos fármacos , Animais , Masculino , Camundongos , Células RAW 264.7RESUMO
OBJECTIVE: To improve the production yield of N-glycosylated anti-VEGFR2 (vascular endothelial growth factor receptor 2) monobody (FN3VEGFR2-Gly) in lpp knockout Escherichia coli cells harboring Campylobacter jejuni N-glycosylation pathway. RESULTS: The leaky CLM37-Δlpp strain efficiently secreted FN3VEGFR2-Gly into culture medium. The extracellular levels of glycosylated FN3VEGFR2-Gly in CLM37-Δlpp culture medium were approximately 11 and 15 times higher compared to those in CLM37 cells via IPTG and auto-induction, respectively. In addition, the highest level of total glycosylated FN3VEGFR2-Gly (70 ± 3.4 mg/L) was found in culture medium via auto-induction. Furthermore, glycosylated FN3VEGFR2-Gly was more stable than unglycosylated FN3VEGFR2-Gly in this expression system, but their bioactivities were relatively similar. CONCLUSIONS: Lpp knockout leaky E. coli strain combined with auto-induction method can enhance the extracellular production of homogenous N-glycosylated FN3VEGFR2-Gly, and facilitate the downstream protein purification. The findings of this study may provide practical implications for the large-scale production and cost-effective harvesting of N-glycosylation proteins.
Assuntos
Anticorpos , Escherichia coli/genética , Espaço Extracelular/metabolismo , Domínio de Fibronectina Tipo III/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular , Anticorpos/química , Anticorpos/genética , Anticorpos/imunologia , Anticorpos/metabolismo , Glicosilação , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/imunologiaRESUMO
Escherichia coli has been considered as a promising host for the production of N-glycosylated therapeutic proteins and glycoconjugate vaccines. In this study, we developed a simple and efficient strategy for improving the production of N-glycosylated recombinant proteins by combining auto-induction with the use of a leaky E. coli strain. A leaky E. coli strain, designated as CLM37-Δlpp, was engineered by deleting the Braun's lipoprotein (lpp) gene of E. coli strain CLM37. Three distinct acceptor model N-glycosylated proteins, glyco-tagged human tenth fibronectin type III domain (FN3-Gly), enhanced green fluorescent protein (eGFP-Gly), and scFv of vascular endothelial growth factor receptor 3 (scFv-VEGFR3-Gly) were then expressed in CLM37-Δlpp, which carried an N-glycosylation machinery from Campylobacter jejuni for the investigation of glycoprotein production. As much as 75%, 65%, and 60% of the glycosylated FN3-Gly, eGFP-Gly, and scFv-VEGFR3-Gly, respectively, were found in the culture medium. The yields of glycosylated FN3-Gly, eGFP-Gly, and scFv-VEGFR3-Gly were 106 ± 7.4 mg/L, 65 ± 2.5 mg/L, and 62 ± 4.3 mg/L, respectively, which were more than three folds the corresponding yields obtained when these proteins were expressed in CLM37, the unmodified strain. The results suggested that this simplified approach could improve the production of N-glycosylated proteins with E. coli to facilitate large-scale production.
RESUMO
Metabolic bone diseases are global public health concerns and are primarily caused by uncontrolled osteoclast (OC) formation and activation. During OC differentiation, intracellular reactive oxygen species (ROS) stimulated by receptor activator of nuclear factor kappa-B ligand (RANKL) can serve as the signaling molecules to promote osteoclastic genes expression. Nuclear factor erythroid-2 related factor 2 (NRF2), a master mediator of cellular antioxidant response, also plays a critical role in OC differentiation through the regulation of redox homeostasis. In this study, we investigated the effects of three NRF2 inducers on osteoclastogenesis, including Bardoxolone methyl (CDDO-Me), Sulforaphane (SFN), and tert-butylhydroquinone (tBHQ). By treating RAW cells with three compounds, we found that NRF2 was activated and its downstream antioxidant genes were upregulated, and the RANKL-induced intracellular ROS production and osteoclastogenesis were impaired. Additionally, the expression of nuclear factor of activated T cells c1 (NFATC1), C-FOS and tumor necrosis factor alpha (TNFα) were inhibited after acute exposures (6â¯h) to the three compounds. Furthermore, suppressed the expression of osteoclast differentiation-associated genes, tartrate-resistant acid phosphatase (TRAP), cathepsin K (CTSK), matrix metalloproteinase-9 (MMP-9) and dendritic cell-specific transmembrane protein (DC-STAMP) were observed after prolonged exposures (5 days) to the compounds. Taken together, these results suggest that CDDO-Me, SFN and tBHQ attenuate RANKL-induced osteoclastogenesis via activation of NRF2-mediated antioxidant response. Among these compounds, relatively low concentrations of CDDO-Me showed stronger active and inhibitory effects on antioxidant response and osteoclastogenesis, respectively.
Assuntos
Antioxidantes/farmacologia , Hidroquinonas/farmacologia , Isotiocianatos/farmacologia , Ácido Oleanólico/análogos & derivados , Osteogênese/efeitos dos fármacos , Ligante RANK/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Células Cultivadas , Masculino , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2/metabolismo , Ácido Oleanólico/farmacologia , Osteoclastos/citologia , Osteoclastos/efeitos dos fármacos , Osteoclastos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , SulfóxidosRESUMO
Escherichia coli cells have been considered as promising hosts for producing N-glycosylated proteins since the successful production of N-glycosylated protein in E. coli with the pgl (N-linked protein glycosylation) locus from Campylobacter jejuni. However, one hurdle in producing N-glycosylated proteins in large scale using E. coli is inefficient glycan glycosylation. In this study, we developed a strategy for the production of N-glycosylated proteins with high efficiency via an optimized auto-induction method. The 10th human fibronectin type III domain (FN3) was engineered with native glycosylation sequon DFNRSK and optimized DQNAT sequon in C-terminus with flexible linker as acceptor protein models. The resulting glycosylation efficiencies were confirmed by Western blots with anti-FLAG M1 antibody. Increased efficiency of glycosylation was obtained by changing the conventional IPTG induction to auto-induction method, which increased the glycosylation efficiencies from 60% and 75% up to 90% and 100% respectively. Moreover, in the condition of inserting the glycosylation sequon in the loop of FN3 (the acceptor sequon with local structural conformation), the glycosylation efficiency was increased from 35% to 80% by our optimized auto-induction procedures. To justify the potential for general application of the optimized auto-induction method, the reconstituted lsg locus from Haemophilus influenzae and PglB from C. jejuni were utilized, and this led to 100% glycosylation efficiency. Our studies provided quantitative evidence that the optimized auto-induction method will facilitate the large-scale production of pure exogenous N-glycosylation proteins in E. coli cells.
Assuntos
Escherichia coli/genética , Fibronectinas/genética , Sequência de Aminoácidos , Campylobacter jejuni/genética , Escherichia coli/metabolismo , Fibronectinas/química , Fibronectinas/metabolismo , Loci Gênicos , Glicoproteínas/química , Glicoproteínas/genética , Glicoproteínas/metabolismo , Glicosilação , Humanos , Microbiologia Industrial , Modelos Moleculares , Polissacarídeos/genética , Polissacarídeos/metabolismo , Conformação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Transdução de SinaisRESUMO
PURPOSE: Radiation therapy for head and neck cancer commonly leads to radiation sialadenitis. Emerging evidence has indicated that phenylephrine pretreatment reduces radiosensitivity in the salivary gland; however, the underlying cytoprotective mechanism remains unclear. Nicotinamide phosphoribosyltransferase (NAMPT) is not only a key enzyme for the nicotinamide adenine dinucleotide salvage pathway, but also a cytokine participating in cell survival, metabolism, and longevity, with a broad effect on cellular functions in physiology and pathology. However, the regulatory events of NAMPT in response to the irradiated salivary gland are unknown. METHODS AND MATERIALS: The cell viability of primary cultured submandibular gland cells was determined using the PrestoBlue assay. NAMPT expression was measured using reverse transcriptase polymerase chain reaction and Western blotting in vitro and in vivo. Silent information regulator 1 (SIRT1) and phosphorylated Akt protein levels were examined by Western blotting. The cellular locations of NAMPT and SIRT1 were detected by immunohistochemistry. NAMPT promoter activity was assessed using the luciferase reporter gene assay. RESULTS: NAMPT was mainly distributed in the cytoplasm of granular convoluted tubule cells and ductal cells in normal submandibular glands. mRNA and protein expression of NAMPT was downregulated after radiation but upregulated with phenylephrine pretreatment both in vivo and in vitro. Moreover, the protein expression of phosphorylated Akt and SIRT1 was decreased in irradiated glands, and phenylephrine pretreatment restored the expression of both. SIRT1 was mainly located in the cell nucleus and cytoplasm in the normal submandibular gland. Phenylephrine dramatically enhanced the expression of SIRT1, which was significantly reduced by radiation. Furthermore, phenylephrine induced a marked increase of NAMPT promoter activity. CONCLUSIONS: These findings reveal the regulatory mechanisms of NAMPT expression, which help to understand the mechanism of the cytoprotective role of phenylephrine on irradiated tissues.
Assuntos
Sobrevivência Celular/efeitos da radiação , Nicotinamida Fosforribosiltransferase/biossíntese , Fenilefrina/administração & dosagem , Protetores contra Radiação/administração & dosagem , Glândula Submandibular/fisiopatologia , Glândula Submandibular/efeitos da radiação , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Feminino , Masculino , Doses de Radiação , Ratos , Ratos Wistar , Glândula Submandibular/efeitos dos fármacos , Glândula Submandibular/enzimologia , Resultado do Tratamento , Regulação para Cima/efeitos dos fármacosRESUMO
Radiotherapy for malignant tumors of the head and neck commonly leads to radiation-induced sialadenitis as a result of radiation-induced salivary gland dysfunction. We demonstrated previously that phenylephrine could protect the irradiated submandibular gland against apoptosis, although the mechanism is unclear. In this study, we investigated the influence of phenylephrine pretreatment on the expressions of aquaporin 5 (AQP5) and c-Jun N-terminal kinase (JNK) that were presumed to have a role in radiation-induced salivary gland dysfunction. Rats pretreated with phenylephrine (5 mg/kg) were locally irradiated (20 Gy) in the head and neck region. The submandibular glands were removed on day 7 after irradiation. The expression of AQP5 and activation of JNK were measured by immunohistochemistry and Western blot. The localization of AQP5 at the apical and lateral plasma membrane of acinar cells was significantly reduced by irradiation, but markedly enhanced with phenylephrine pretreatment. The protein expression of AQP5 was decreased by 84.91% in irradiated glands, whereas it was fully recovered to the control level in phenylephrine-pretreated glands. Moreover, many acinar, ductal and granular convoluted tubular cells in the irradiated glands exhibited intense immunoreactivity for p-JNK, while in the phenylephrine-pretreated irradiated glands, only a few acinar cells exhibited very faint immunoreactivity for p-JNK. The protein expression level of p-JNK was increased by 41.65% in the irradiated alone glands, but was significantly decreased in the phenylephrine-pretreated irradiated glands. These results suggest that the protective mechanism of phenylephrine might be related to the improved expression of AQP5 and decreased activation of JNK. Pretreatment with phenylephrine in patients undergoing radiotherapy may provide a helpful strategy for suppression of radiation-induced sialadenitis.
Assuntos
Aquaporina 5/metabolismo , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Fenilefrina/farmacologia , Protetores contra Radiação/farmacologia , Glândula Submandibular/efeitos dos fármacos , Glândula Submandibular/efeitos da radiação , Animais , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Membrana Celular/efeitos da radiação , Polaridade Celular/efeitos dos fármacos , Polaridade Celular/efeitos da radiação , Masculino , Tamanho do Órgão/efeitos dos fármacos , Tamanho do Órgão/efeitos da radiação , Fosfoproteínas/metabolismo , Transporte Proteico/efeitos dos fármacos , Transporte Proteico/efeitos da radiação , Ratos , Ratos Wistar , Glândula Submandibular/metabolismo , Glândula Submandibular/patologiaRESUMO
OBJECTIVE: To study the response of rat bone marrow mesenchymal stem cells (MSCs) to a single period of mechanical strain and expression patterns of transforming growth factor-beta (TGF-beta) and insulin-like growth factor-II (IGF-II) after mechanical stretch. METHODS: Bone marrow MSCs were isolated from SD rats and cultured in vitro. A four-point bending apparatus were used to perform a single period of mechanical strain (2000 microepsilon, 40 min) on MSCs. Cellular proliferation and alkaline phosphatase (ALP) activity of MSCs were examined and gene expression patterns of TGF-beta and IGF-II were detected by SYBR green quantitative real-time RT-PCR. RESULTS: Cell proliferation, ALP activity and expression of TGF-beta and IGF-II were all significantly up-regulated in stretched MSCs when compared with their controls. The mRNA levels of TGF-beta and IGF-II got top increase immediately after mechanical loading and increased about 51.44 and 8.92 folds, respectively, when compared with control cells. Expression of TGF-beta and IGF-II decreased with time and returned to control level at 12 h after mechanical stimulus, despite of a small increase at 6 h. CONCLUSION: The mechanical stretch can promote MSCs proliferation, up-regulate its ALP activity and induce a time-dependent expression increase of TGF-beta and IGF-II which in turn result in osteogenic differentiation of MSCs. Mechanical stimulus is a key stimulator for osteogenic differentiation of MSCs and vital for bone formation in distraction osteogenesis.
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Fator de Crescimento Insulin-Like II , Fator de Crescimento Transformador beta , Animais , Células da Medula Óssea , Diferenciação Celular , Proliferação de Células , Células-Tronco Mesenquimais , Osteogênese , Osteogênese por Distração , RNA Mensageiro , Ratos , Ratos Sprague-Dawley , SomatomedinasRESUMO
OBJECTIVE: To evaluate the osteoblastic differentiation and compare the difference in the gene expression of rat bone marrow mesenchymal stem cells (MSCs) affected by a single period of mechanical strain. METHODS: Bone marrow MSCs were harvested from the femurs and tibiae of SD rats and cultured in vitro. A four-point bending apparatus were used to perform a single 40-minute period of 2,000 microepsilon mechanical strain on these MSCs. The proliferation of the MSCs was tested by MTT on scheduled date, and the osteoblastic differentiation of the MSCs was measured by testing the expression of osteocalcin and alkaline phosphate (ALP) activity of these cells. In addition, we have investigated the possible mechanisms underlying the action of the single 40-minute period of 2,000 microepsilon mechanical strain on these MSCs, after profile blotted and handled by bioinformation, the gene expressions of these two periods of MSCs were examined. RESULTS: The MSCs have grown well in vitro. Our experiment showed that mechanical environment did not weaken the proliferation of the MSCs. However, the ALP activity and the expression of osteocalcin were significantly up-regulated by the 2,000 microepsilon mechanical strain. Using the 27 K Rat Genome Array, 416 different expressions were found. The rate of different genes was 2.8%, of which the expressions of 247 genes increased (61 genes remarkably increased) and 169 genes decreased (74 genes remarkably decreased) in these two periods of MSCs. CONCLUSION: Mechanical strain induced the osteoblastic differentiation of the MSCs, which may be attributed to the different gene levels.
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Células da Medula Óssea , Transcriptoma , Fosfatase Alcalina , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Células-Tronco Mesenquimais , Osteoblastos , Osteocalcina , Ratos , Ratos Sprague-DawleyRESUMO
AIM: Understanding the response of mesenchymal stem cells (MSCs) to mechanical strain and their consequent gene expression patterns will broaden our knowledge of the mechanobiology of distraction osteogenesis. METHODOLOGY: In this study, a single period of cyclic mechanical stretch (0.5 Hz, 2,000 microepsilon) was performed on rat bone marrow MSCs. Cellular proliferation and alkaline phosphatase (ALP) activity was examined. The mRNA expression of six bone-related genes (Ets-1, bFGF, IGF-II, TGF-beta, Cbfa1 and ALP) was detected using real-time quantitative RT-PCR. RESULTS: The results showed that mechanical strain can promote MSCs proliferation, increase ALP activity, and up-regulate the expression of these genes. A significant increase in Ets-1 expression was detected immediately after mechanical stimulation, but Cbfa1 expression became elevated later. The temporal expression pattern of ALP coincided perfectly with Cbfa1. CONCLUSION: The results of this study suggest that mechanical strain may act as a stimulator to induce differentiation of MSCs into osteoblasts, and that these bone-related genes may play different roles in the response of MSCs to mechanical stimulation.
Assuntos
Células da Medula Óssea/fisiologia , Células-Tronco Mesenquimais/fisiologia , Osteogênese por Distração , Fosfatase Alcalina/análise , Animais , Antígenos de Superfície/análise , Fenômenos Biomecânicos , Técnicas de Cultura de Células , Diferenciação Celular/fisiologia , Proliferação de Células , Subunidade alfa 1 de Fator de Ligação ao Core/análise , Fator 2 de Crescimento de Fibroblastos/análise , Fator de Crescimento Insulin-Like II/análise , Osteoblastos/fisiologia , Células-Tronco Pluripotentes/fisiologia , Proteína Proto-Oncogênica c-ets-1/análise , Ratos , Estresse Mecânico , Fator de Crescimento Transformador beta/análise , Regulação para Cima/fisiologiaRESUMO
OBJECTIVE: This study was to observe the effects of bone marrow mesenchymal stem cell transplantation on new bone formation in a rat mandibular osteodistraction model. MATERIAL AND METHODS: Autologous bone marrow stem cells were obtained from tibiae of 40 male rats. Two weeks after cell harvest, the rats underwent right mandibular distraction and were then randomly divided into two groups (group A=20, group B=20). After distraction was complete, the stem cells were injected into the distracted gaps in group A, while the rats in group B only received physiological saline. Twenty rats (10 from each group) were sacrificed on postoperative days 27 and 55, respectively. The distracted mandibles were harvested and processed for radiographic, histological and histomorphometric analysis. RESULTS: The radiodensity of the distraction zone was higher in group A than in group B at both time points. Histologically callus was found in both groups but more bone was formed in group A. Histomorphometric analysis also demonstrated that both new bone volume and thickness of the new trabeculae were significantly greater in group A than in group B. CONCLUSION: The results of this study suggest that autologous bone marrow stem cell transplantation may be considered as a potential method to accelerate bone regeneration in the distraction gap, and enhance consolidation.
Assuntos
Transplante de Medula Óssea/métodos , Regeneração Óssea/fisiologia , Mandíbula/fisiologia , Transplante de Células-Tronco Mesenquimais/métodos , Osteogênese por Distração/métodos , Animais , Calo Ósseo/fisiologia , Bovinos , Masculino , Mandíbula/diagnóstico por imagem , Mandíbula/cirurgia , Modelos Animais , Radiografia , Ratos , Ratos Sprague-DawleyRESUMO
OBJECTIVE: To explore the response of rat bone marrow mesenchymal stem cells (MSCs and calvarial osteoblasts to mechanical strain and the consequent changes of cytoskeleton F-actin. METHODS: Bone marrow MSCs and calvarial osteoblasts were isolated from SD rats and cultured in vitro. Mechanical stretch was performed on passage 3 cells at 2 000 microepsilon for 0, 2, 6 and 12 hours using four-point bending system. The response of cells and the distribution of F-actin were observed using fluorescent staining under laser scanning confocal microscope and the morphological parameters were quantified using image analysis software Laserpix. RESULTS: Under mechanical stretch, the fluorescent staining decreased obviously at both MSCs and osteoblasts, and F-actin filaments were rearranged and became tenuous, thinner, and abnormally distributed. The outline of nucleus became unclear and apoptotic changes were observed at some of both cells. Cellular size decreased more significantly in MSCs than in osteoblasts. Quantity analysis showed that total area of cells, total fluorescent density and green fluorescent density (F-actin) were all significantly decreased in MSCs (P < 0.05 or P < 0.01), and total fluorescent density, green fluorescent density and red fluorescent density (nuclei) did also in osteoblasts (P < 0.05 or P < 0.01). CONCLUSION: Mechanical stretch caused extensive response on both MSCs and osteoblasts which led to the rearrangement of F-actin filament and apoptosis in some of these cells. MSCs were more sensitive to mechanical strain than osteoblasts.
Assuntos
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Células-Tronco Mesenquimais , Osteoblastos , Animais , Células da Medula Óssea , Células Cultivadas , Citoesqueleto , Microtúbulos , Ratos , Estresse MecânicoRESUMO
Objective:To study the effects of BMP-7 gene transfected bone marrow mesenchymal stem cells(MSCs)on callus acceleration in rat mandibular distraction osteogenesis.Methods:Forty-eight adult male SD rats were randomly divided into experimental and control groups.MSCs were obtained from individual rat and transfected by pEGF-P-BMP7 for the 24 experimental rats and by the empty vector pEGFP-N1 for the 24 control rats.The rats were underwent right mandibular distraction.1?106/100 ?l BMP-7 gene transfected cells in 10 ?l of normal saline were injected into the distraction gap in each experimental rat,while the same number of empty vector transfected cells in each control rat.The distracted mandibles were harvested 2,4,and 8 weeks respectively after cell injection and evaluated by radiological,histological and histomorphometric analysis.Results:Radiological and histological examinations showed that more new bone was formed in experimental group than in control.Histomorphometric analysis also demonstrated that both new bone volume(NBV1 and NBV2)and the thickness of new trabeculae(TNT)were significantly higher in experimental rats than those in control(P