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1.
Connect Tissue Res ; 62(1): 115-132, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32683988

RESUMO

Purpose/Aim: We recently found that blocking CCN2 signaling using a monoclonal antibody (FG-3019) may be a novel therapeutic strategy for reducing overuse-induced tissue fibrosis. Since CCN2 plays roles in osteoclastogenesis, and persistent performance of a high repetition high force (HRHF) lever pulling task results in a loss in trabecular bone volume in the radius, we examined here whether blocking CCN2 signaling would reduce the early catabolic effects of performing a HRHF task for 3 weeks. Materials and Methods: Young adult, female, Sprague-Dawley rats were operantly shaped to learn to pull at high force levels, before performing the HRHF task for 3 weeks. HRHF task rats were then left untreated (HRHF Untreated), treated in task weeks 2 and 3 with a monoclonal antibody that antagonizes CCN2 (HRHF+FG-3019), or treated with an IgG (HRHF+IgG), while continuing to perform the task. Non-task control rats were left untreated. Results: In metaphyseal trabeculae of the distal radius, HRHF Untreated and HRHF-IgG rats showed increased osteoblast numbers and other indices of bone formation, compared to controls, yet decreased trabecular bone volume, increased osteoclast numbers, and increased serum CTX-1 (a serum biomarker of bone resorption). HRHF+FG-3019 rats also showed increased osteoblast numbers and bone formation, but in contrast to HRHF Untreated and HRHF-IgG rats, showed higher trabecular bone volume, and reduced osteoclast numbers and serum CTX-1 levels (and statistically similar to Control levels). Conclusions: HRHF loading increased bone formation in each task group, yet blocking CCN2 dampened trabecular bone catabolism by reducing osteoclast numbers and activity.

2.
Orthop J Sports Med ; 8(9): 2325967120939001, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32953920

RESUMO

Background: The retear rate after rotator cuff repair remains unacceptably high. Various biological engineered scaffolds have been proposed to reduce the retear rate. We have developed a double rip-stop repair with medial row knot (DRSK) technique to enhance suture-tendon strength and a novel engineered tendon-fibrocartilage-bone composite (TFBC) for rotator cuff repair. Hypothesis: DRSK rotator cuff repair augmented with TFBC will have better biomechanical properties than that of DRSK repair with an acellular dermal graft (DG). Study Design: Controlled laboratory study. Methods: Fresh-frozen canine shoulders (n = 30) and knees (n = 10) were used. TFBCs were harvested from the patellar tendon-tibia complex and prepared for rotator cuff repair. The infraspinatus tendon was sharply detached from its bony attachment and randomly assigned to the (1) control group: DRSK repair alone, (2) TFBC group: DRSK repair with TFBC, and (3) DG group: DRSK repair with DG. All specimens were tested to failure, and videos were recorded. The footprint area, tendon thickness, load to create 3-mm gap formation, failure load, failure modes, and stiffness were recorded and compared. Data were recorded as mean ± SD. Results: The mean load to create a 3-mm gap in both the control group (206.8 ± 55.7 N) and TFBC group (208.9 ± 39.1 N) was significantly higher than that in the DG group (157.7 ± 52.3 N) (P < .05 for all). The failure load of the control group (275.7 ± 75.0 N) and TFBC group (275.2 ± 52.5 N) was significantly higher compared with the DG group (201.5 ± 49.7 N) (P < .05 for both comparisons). The stiffness of the control group (26.4 ± 4.7 N/mm) was significantly higher than of the TFBC group (20.4 ± 4.4 N/mm) and the DG group (21.1 ± 4.8 N/mm) (P < .05 for both comparisons). Conclusion: TFBC augmentation showed superior biomechanical performance to DG augmentation in rotator cuff tears repaired using the DRSK technique, while there was no difference between the TFBC and control groups. Clinical Relevance: TFBC may help to reduce retear or gap formation after rotator cuff repair using the DRSK technique.

3.
J Orthop Sci ; 2020 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-32814661

RESUMO

BACKGROUND: The purpose of this study was to determine the effect of fibrinogen concentration on cell viability and migration in a tissue culture tendon healing model. METHODS: Forty-eight canine flexor digitorum profundus tendons were randomly divided into three groups. In each group the tendons were lacerated and repaired augmented with a canine bone marrow stromal cell seeded fibrin interposition patch using either 5 mg/ml fibrinogen and 25 U/ml thrombin (physiological as a control), 40 mg/ml fibrinogen and 250 U/ml thrombin (low adhesive), or 80 mg/ml fibrinogen and 250 U/ml thrombin (high adhesive). The sutured tendons were cultured for two or four weeks. RESULTS: Failure load was not significantly different among the groups. Cell-labeling staining showed that the stromal cells migrated across the gap in the control and low adhesive groups, but there was no cell migration in the high adhesive group at two weeks. CONCLUSION: A high fibrinogen concentration in a fibrin patch or glue may impede early cell migration. LEVEL OF EVIDENCE: Not applicable because this study was a laboratory study.

4.
Bone Joint Res ; 9(6): 285-292, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32728429

RESUMO

Aims: Many biomechanical studies have shown that the weakest biomechanical point of a rotator cuff repair is the suture-tendon interface at the medial row. We developed a novel double rip-stop (DRS) technique to enhance the strength at the medial row for rotator cuff repair. The objective of this study was to evaluate the biomechanical properties of the DRS technique with the conventional suture-bridge (SB) technique and to evaluate the biomechanical performance of the DRS technique with medial row knots. Methods: A total of 24 fresh-frozen porcine shoulders were used. The infraspinatus tendons were sharply dissected and randomly repaired by one of three techniques: SB repair (SB group), DRS repair (DRS group), and DRS with medial row knots repair (DRSK group). Specimens were tested to failure. In addition, 3 mm gap formation was measured and ultimate failure load, stiffness, and failure modes were recorded. Results: The mean load to create a 3 mm gap formation in the DRSK and DRS groups was significantly higher than in the SB group. The DRSK group had the highest load to failure with a mean ultimate failure load of 395.0 N (SD 56.8) compared to the SB and DRS groups, which recorded 147.1 N (SD 34.3) and 285.9 N (SD 89.8), respectively (p < 0.001 for both). The DRS group showed a significantly higher mean failure load than the SB group (p = 0.006). Both the DRS and DRSK groups showed significantly higher mean stiffness than the SB group. Conclusion: The biomechanical properties of the DRS technique were significantly improved compared to the SB technique. The DRS technique with medial row knots showed superior biomechanical performance than the DRS technique alone.

5.
J Orthop Sci ; 2020 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-32317146

RESUMO

BACKGROUND: Carpal Tunnel Syndrome (CTS) is an idiopathic fibrotic disorder. Fibrosis in the subsynovial connective tissues (SSCT) of CTS and many other fibrotic diseases is mediated by Transforming growth factor ß (TGF-ß). Recently monocyte chemoattractant protein-1 (MCP-1) a cytokine involved in cellular recruitment has been suggested to regulate TGF-ß activity. It is related to the onset of diseases which are caused by fibrosis, such as idiopathic pulmonary fibrosis, renal fibrosis, and systemic scleroderma. In this study, we evaluated the effect of the MCP-1 synthesis inhibitor, Bindarit, on primary cultures of fibroblasts from the SSCT of five CTS patients. METHODS: Fibroblasts were treated with Bindarit (10 µM, 50 µM, 100 µM, or 300 µM). Responses to inhibitors were evaluated by regulation of CTS fibrosis-associated genes, fibrosis gene array and Smad luciferase reporter assay. We also assessed the combination effect of Bindarit and SD208, a TGF-ß receptor type 1 inhibitor on TGF-ß signaling. RESULTS: Collagen type III A1 (Col3), connective tissue growth factor (CTGF), and SERPINE1 expression were significantly down-regulated by Bindarit (300 µM) compared to vehicle control. In the fibrosis array, expression of inhibin beta E chain precursor (INHBE), beta actin (ACTB), endothelin 1 (EDN1) and hypoxanthine phosphoribosyltransferase 1 (HPRT1) were significantly down-regulated, and integrin beta-3 (ITGB3) was significantly up-regulated by Bindarit (300 µM). Smad signal transduction activation was significantly down-regulated by Bindarit (300 µM) and/or SD208 (1 µM) with TGF-ß1 compared to vehicle control with TGF-ß1. CONCLUSIONS: These results suggest that Bindarit in combination with SD208 may be beneficial as medical therapy for the SSCT fibrosis associated with CTS.

6.
J Cell Physiol ; 235(7-8): 5679-5688, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31975377

RESUMO

Deletion of TGFß inducible early gene-1 (TIEG) in mice results in an osteopenic phenotype that exists only in female animals. Molecular analyses on female TIEG knockout (KO) mouse bones identified increased expression of sclerostin, an effect that was confirmed at the protein level in serum. Sclerostin antibody (Scl-Ab) therapy has been shown to elicit bone beneficial effects in multiple animal model systems and human clinical trials. For these reasons, we hypothesized that Scl-Ab therapy would reverse the low bone mass phenotype of female TIEG KO mice. In this study, wildtype (WT) and TIEG KO female mice were randomized to either vehicle control (Veh, n = 12/group) or Scl-Ab therapy (10 mg/kg, 1×/wk, s.c.; n = 12/group) and treated for 6 weeks. Following treatment, bone imaging analyses revealed that Scl-Ab therapy significantly increased cancellous and cortical bone in the femur of both WT and TIEG KO mice. Similar effects also occurred in the vertebra of both WT and TIEG KO animals. Additionally, histomorphometric analyses revealed that Scl-Ab therapy resulted in increased osteoblast perimeter/bone perimeter in both WT and TIEG KO animals, with a concomitant increase in P1NP, a serum marker of bone formation. In contrast, osteoclast perimeter/bone perimeter and CTX-1 serum levels were unaffected by Scl-Ab therapy, irrespective of mouse genotype. Overall, our findings demonstrate that Scl-Ab therapy elicits potent bone-forming effects in both WT and TIEG KO mice and effectively increases bone mass in female TIEG KO mice.

7.
J Orthop Res ; 38(8): 1845-1855, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-31930553

RESUMO

Flexor tendon injuries and tendinopathy are very common but remain challenging in clinical treatment. Exosomes-based cell-free therapy appears to be a promising strategy for tendon healing, while limited studies have evaluated its impacts on tenocyte biology. The objective of this study was to characterize a novel purified exosome product (PEP) derived from plasma, as well as to explore its cellular effects on canine tenocyte biology. The transmission electron microscope revealed that exosomes of PEP present cup-shaped structures with the diameters ranged from 80 to 141 nm, and the NanoSight report presented that their size mainly concentrated around 100 nm. The enzyme-linked immunosorbent assay kits analysis showed that PEP was positive for CD63 and AChE expression, and the cellular uptake of exosomes internalized into tenocyte cytoplasm was observed. The cell growth assays displayed that tenocyte proliferation ability was enhanced by PEP solution in a dose-dependent manner. Tenogenic phenotype was preserved as is evident by that tendon-related genes expression (SCX, COL1A, COL3A1, TNMD, DCN, and MKX) were expressed insistently in a high level, while tenocytes were treated with 5% PEP solution. Furthermore, migration capability was maintained and total collagen deposition was increased. More interesting, dexamethasone-induced cellular apoptosis was attenuated during the incubation of tenocytes with a 5% PEP solution. These findings will provide the basic understandings about the PEP, and support the potential use of this biological strategy for tendon healing.

8.
J Orthop Translat ; 19: 58-67, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31844614

RESUMO

Background/objective: Stem cell-based therapy has been applied to accelerate the revitalization of allograft tendon into a viable and functional tendon. Although many authors have proposed different methods to help the seeded stem cell distribution in the decellularized allograft, limited success has been achieved as tendon is a high dense connective tissue. We hypothesized that bone marrow stromal cells (BMSCs), seeded through the lateral slit, can regenerate the decellularized tendon (DCT) graft. The cell proliferation, cell viability, and tendon-specific gene expression are increased with the seeded cell density. Methods: Eighty-seven flexor digitorum profundus tendons were equally and randomly divided into 6 treatment groups that were seeded with low-density (2 × 107 cells/mL) and high-density (5 × 107 cells/mL) BMSCs through lateral slits cultured for 2 and 4 weeks, DCT without cells, and fresh live tendons. Tendons were evaluated for cell distribution, cell proliferation, cell viability, gene expression of Collagen I and Collagen III, tenogenic markers, and MMPs. Results: Histologic evaluation revealed BMSCs distributed from the lateral slit to the whole DCT. BMSCs were proliferated and kept viable in lateral slit decellularized tendon (LSDCT) in both seeded cell density groups after 2 and 4 weeks of culture. However, no significant differences in the cell proliferation between both cell density groups at 2 and 4 weeks of culture were observed. The lowest cell viability was found in the high-density group after 4 weeks of culture. BMSCs in LSDCT showed a significant tendency of higher gene expression of Collagen I, Collagen III, tenascin C, MMP2, MMP9, and MMP13 compared to normal tendons in both cell density groups at 2 and 4 weeks of culture. Conclusion: BMSCs proliferated and remained viable after 2 and 4 weeks of culture with distribution throughout the lateral slits. Lateral slit preparation allows for the effective delivery and maintenance of mesenchymal cells with proliferation and generating a tenogenic behaviour of DCT in both the low and high cell densities in an in vitro model. The translation potential of this article: Revitalizing the implanted decellularized allograft is important for clinical application. In this study, we demonstrated that the DCT, with lateral slits, could harbour the seeded stem cell and stimulate proliferation with collagen synthesis. This evidence was presented for clinical application of the lateral slit technique, in DCT grafts, which would repopulate the seeded BMSCs during tendon and ligament reconstruction.

9.
J Orthop Res ; 37(6): 1419-1428, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30548886

RESUMO

Flexor tendon injury is often associated with suboptimal outcomes and results in substantial digit dysfunction. Stem cells have been isolated from several experimental animals for the growing interest and needs of utilizing cell-based therapies. Recently, turkey has been developed as a new large animal model for flexor tendon research. In the present study, we reported the isolation and characterization of bone marrow-derived mesenchymal stem cells (BMSCs) from 8- to 12-month-old heritage-breed turkeys. The isolated cells demonstrated fibroblast-like morphology, clonogenic capacity, and high proliferation rate. These cells were positive for surface antigens CD90, CD105, and CD44, but were negative for CD45. The multipotency of turkey BMSCs was determined by differentiating cells into osteogenic, adipogenic, chondrogenic, and tenogenic lineages. There was upregulated gene expression of tenogenic markers, including mohawk, tenomodulin, and EGR1 as well as increased collagen synthesis in BMP12 induced cells. The successful isolation and verification of bone marrow-derived MSCs from turkey would provide opportunities of studying cell-based therapies and developing new treatments for tendon injuries using this novel preclinical large animal model. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 37:1419-1428, 2019.


Assuntos
Separação Celular/métodos , Células-Tronco Mesenquimais/citologia , Adipogenia , Animais , Células da Medula Óssea/citologia , Diferenciação Celular , Proliferação de Células , Condrogênese , Osteogênese , Tendões/citologia , Perus
10.
Biomaterials ; 192: 189-198, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30453215

RESUMO

Reducing rotator cuff failure after repair remains a challenge due to suboptimal tendon-to-bone healing. In this study we report a novel biomaterial with engineered tendon-fibrocartilage-bone composite (TFBC) and bone marrow-derived mesenchymal stem cell sheet (BMSCS); this construct was tested for augmentation of rotator cuff repair using a canine non-weight-bearing (NWB) model. A total of 42 mixed-breed dogs were randomly allocated to 3 groups (n = 14 each). Unilateral infraspinatus tendon underwent suture repair only (control); augmentation with engineered TFBC alone (TFBC), or augmentation with engineered TFBC and BMSCS (TFBC + BMSCS). Histomorphometric analysis and biomechanical testing were performed at 6 weeks after surgery. The TFBC + BMSCS augmented repairs demonstrated superior histological scores, greater new fibrocartilage formation and collagen fiber organization at the tendon-bone interface compared with the controls. The ultimate failure load and ultimate stress were 286.80 ± 45.02 N and 4.50 ± 1.11 MPa for TFBC + BMSCS group, 163.20 ± 61.21 N and 2.60 ± 0.97 MPa for control group (TFBC + BMSCS vs control, P = 1.12E-04 and 0.003, respectively), 206.10 ± 60.99 N and 3.20 ± 1.31 MPa for TFBC group (TFBC + BMSCS vs TFBC, P = 0.009 and 0.045, respectively). In conclusion, application of an engineered TFBC and BMSCS can enhance rotator cuff healing in terms of anatomic structure, collagen organization and biomechanical strength in a canine NWB model. Combined TFBC and BMSCS augmentation is a promising strategy for rotator cuff tears and has a high potential impact on clinical practice.


Assuntos
Fibrocartilagem/química , Células-Tronco Mesenquimais/citologia , Manguito Rotador/fisiologia , Tendões/química , Tecidos Suporte/química , Cicatrização , Animais , Materiais Biocompatíveis/química , Osso e Ossos/química , Cães , Transplante de Células-Tronco Mesenquimais , Manguito Rotador/citologia , Engenharia Tecidual
11.
BMC Musculoskelet Disord ; 19(1): 342, 2018 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-30243295

RESUMO

BACKGROUND: Fibroblast behavior and cell-matrix interactions of cells from normal and idiopathic carpal tunnel syndrome (CTS) subsynovial connective tissue (SSCT) with and without Triamcinolone Acetonide (TA) were compared in this study. A cell-seeded gel contraction model was applied to investigate the effect of steroid treatment on SSCT fibroblast gene expression and function. METHODS: SSCT cells were obtained from CTS patients and fresh cadavers. Cells were isolated by mechanical and collagenase digestion. Collagen gels (1 mg/ml) were prepared with SSCT cells (1 × 106/mL). A sterile Petri dish with a cloning ring in the center was prepared. The area between the ring and outer dish was filled with cell-seeded collagen solution and gelled for 1 h. The gel was released from the outer way of the petri dish to allow gel contraction. Cell seeded gels were treated with 10 M triamcinolone acetonide (TA) or vehicle (DMSO) in modified MEM. Every 4 h for 3 days the contracting gels were photographed and areas calculated. Duplicate contraction tests were performed with each specimen, and the averages were used in the analyses, which were conducted using two-factor analysis of variance in a generalized linear model framework utilizing generalized estimating equations (GEE) to account for the correlation between samples. The contraction rate was determined by the area change over time, and the decay time constant was calculated. A customized mechanical test system was used to determine gel stiffness and tensile strength. Gene expression was assessed using Human Fibrosis and Cell Motility PCR arrays. RESULTS: TA-treated gels had a significantly higher contraction rate, tensile strength and stiffness than the untreated gels. Proteinases involved in remodeling had increased expression in TA-treated gels of the patient group. Pro-fibrotic genes and ECM regulators, such as TGF-ß, collagens and integrins, were down-regulated by TA, indicating that TA may work in part by decreasing fibrotic gene expression. CONCLUSIONS: This study showed that TA affects cell-matrix interaction and suppresses fibrotic gene expression in the SSCT cells of CTS patients.


Assuntos
Síndrome do Túnel Carpal/tratamento farmacológico , Fibroblastos/efeitos dos fármacos , Glucocorticoides/farmacologia , Triancinolona Acetonida/farmacologia , Síndrome do Túnel Carpal/metabolismo , Colágeno/metabolismo , Feminino , Fibroblastos/metabolismo , Glucocorticoides/uso terapêutico , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Cultura Primária de Células , Fator de Crescimento Transformador beta/metabolismo , Triancinolona Acetonida/uso terapêutico
12.
Stem Cells Int ; 2018: 3697971, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29977306

RESUMO

Tendon injuries are among the most common and severe hand injuries with a high demand for functional recovery. Stem cells have been identified and isolated from different species and a variety of tissues for the sake of regenerative medicine. Recently, turkey has been suggested as a potential new large animal model for flexor tendon-related research. However, turkey tissue-specific stem cells have not been investigated. Here, we presented the isolation and verification of tendon-derived stem cells (TDSCs) from 6- to 8-month-old heritage-breed turkey. TDSCs were isolated from turkey flexor tendon by plating nucleated cells at the determined optimal density. Approximately 4% of the nucleated cells demonstrated clonogenicity, high proliferation rate, and trilineage differentiation potential after induction culturing. These cells expressed surface antigens CD90, CD105, and CD44, but did not express CD45. There was a high level of gene expression of tenogenic markers in TDSCs, including mohawk, collagen type I, tenascin C, and elastin. Turkey TDSCs also expressed transcription factors PouV, Nanog, and Sox2, which are critically involved in the regulation of stemness. The successful isolation of tendon-derived stem cells from turkey was beneficial for future studies in tendon tissue engineering and would help in the development of new treatment for tendon diseases using this novel animal model.

13.
J Tissue Eng Regen Med ; 12(7): 1690-1701, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29763517

RESUMO

Surgical repair of rotator cuff tears presents a significant clinical challenge with high failure rates and inferior functional outcomes. Graft augmentation improves repair outcomes; however, currently available grafting materials have limitations. Although cell-seeded decellularized tendon slices may facilitate cell infiltration, promote tendon incorporation, and preserve original mechanical strength, the unique fibrocartilage zone is yet to be successfully reestablished. In this study, we investigated the biological and mechanical properties of an engineered tendon-fibrocartilage-bone composite (TFBC) with cyclic tension (3% strain; 0.2 Hz). Decellularized TFBCs seeded with bone marrow-derived mesenchymal stem cell (BMSCs) sheets and subjected to mechanical stimulation for up to 7 days were characterised by histology, immunohistochemistry, scanning electron microscopy, mechanical testing, and transcriptional regulation. The decellularized TFBC maintained native enthesis structure and properties. Mechanically stimulated TFBC-BMSC constructs displayed increased cell migration after 7 days of culture compared with static groups. The seeded cell sheet not only integrated well with tendon scaffold but also distributed homogeneously and aligned to the direction of stretch under dynamic culture. Developmental genes were regulated including scleraxis, which was significantly upregulated with mechanical stimulation. The Young's modulus of the cell-seeded constructs was significantly higher compared with the noncell-seeded controls. In conclusion, the results of this study reveal that the TFBC-BMSC composite provides an ideal multilayer construct for cell seeding and growth, with mechanical preconditioning further enhances cell penetration and differentiation. The BMSC cell sheet revitalised TFBC in conjunction with mechanical stimulation could serve as a novel and primed biological patch to improve rotator cuff repair.


Assuntos
Células da Medula Óssea/metabolismo , Fibrocartilagem , Células-Tronco Mesenquimais/metabolismo , Lesões do Manguito Rotador , Manguito Rotador , Engenharia Tecidual , Tecidos Suporte/química , Animais , Células da Medula Óssea/patologia , Movimento Celular , Cães , Fibrocartilagem/química , Fibrocartilagem/metabolismo , Fibrocartilagem/patologia , Células-Tronco Mesenquimais/patologia , Manguito Rotador/química , Manguito Rotador/metabolismo , Manguito Rotador/patologia , Lesões do Manguito Rotador/metabolismo , Lesões do Manguito Rotador/patologia , Lesões do Manguito Rotador/terapia
14.
J Orthop Res ; 2018 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-29575268

RESUMO

This study was to test our hypothesis that flexor tendon reconstruction with an allograft revitalized with bone marrow stromal cells (BMSCs) and synovialized with carbodiimide derivatized autologous synovial fluid (cd-SYN) would result in better digit functional restoration than the conventional allograft tendon. A total of 32 flexor digital profundus tendons from the second and fifth digit of 16 dogs were created a repair failure model first. Then, failed-repaired tendons were reconstructed with either a revitalized-synovialized allograft tendon or a clinical standard autograft tendon (control group). The allograft tendon was seeded with autologous BMSCs in multiple slits and the graft surface was coated with cd-SYN. A 6 weeks after tendon reconstruction, the digits were harvested and evaluated for digit function, adhesion status, tendon gliding resistance, attachment strength, cell viability, and histologic factors. The allograft group had significantly improved digit function compared with the control group through decreased work of flexion, increased digit range of motion under 2-Newton force, and less adhesion score (p < .05). However, the distal attachment-site strength and stiffness in the allograft tendon were significantly weaker than the autografts (p < .05). No significant difference was found for gliding resistance. Histologically, allograft tendons coated with allograft had smoother surfaces and showed tendon-to-bone and tendon-to-tendon incorporation. Viable BMSCs were found in the tendon slits 6 weeks after the graft. In conclusion, cellular lubricant-based modification of allograft tendons improved digit function and reduced the adhesions compared with autograft for flexor tendon reconstruction. However, improvement of graft-to-host tendon healing is still challenging. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.

15.
J Cell Physiol ; 233(3): 2067-2074, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28294324

RESUMO

Fibrosis of the subsynovial connective tissue (SSCT) in carpal tunnel syndrome (CTS) patients is increasingly recognized as an important aspect of CTS pathophysiology. In this study, we evaluated the effect of blocking profibrotic pathways in fibroblasts from the SSCT in CTS patients. Fibroblasts were stimulated with transforming growth factor ß1 (TGF-ß1), and then treated either with a specific fibrosis pathway inhibitor targeting TGF-ß receptor type 1 (TßRI), platelet-derived growth factor receptor (PDGFR), epidermal growth factor receptor (EGFR), or vascular endothelial growth factor receptor (VEGFR). Fibrosis array and quantitative real-time polymerase chain reaction of fibrotic genes were evaluated. Array gene expression analysis revealed significant down-regulation of multiple fibrotic genes after treatment with TßRI, PDGFR, and VEGFR inhibitors. No array fibrotic genes were significantly down-regulated with EGFR inhibition. Further gene expression analysis of known CTS fibrosis markers collagen type I A2 (Col1), collagen type III A1 (Col3), connective tissue growth factor (CTGF), and SERPINE1 showed significantly down-regulation after TßRI inhibition. In contrast, VEGFR inhibition significantly down-regulated CTGF and SERPINE1, whereas, PDGFR and EGFR inhibition significantly down-regulated Col3. Taken together the inhibition of TßRI appears to be the primary mediator of fibrotic gene expression in fibroblasts from CTS patients. TGF-ß/Smad activity was further evaluated, and as expected inhibition of Smad activity was significantly down-regulated after inhibition of TßRI, but not with PDGFR, VEGFR, or EGFR inhibition. These results indicate that local therapies specifically targeting TGF-ß signaling alone or in combination offer the potential of a novel local antifibrosis therapy for patients with CTS.


Assuntos
Síndrome do Túnel Carpal/tratamento farmacológico , Receptores ErbB/antagonistas & inibidores , Fibrose/patologia , Receptores do Fator de Crescimento Derivado de Plaquetas/antagonistas & inibidores , Receptores de Fatores de Crescimento Transformadores beta/antagonistas & inibidores , Receptores de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Membrana Sinovial/patologia , Fator de Crescimento Transformador beta/metabolismo , Síndrome do Túnel Carpal/patologia , Células Cultivadas , Colágeno Tipo I/biossíntese , Colágeno Tipo I/genética , Colágeno Tipo III/biossíntese , Colágeno Tipo III/genética , Tecido Conjuntivo/patologia , Células do Tecido Conjuntivo/citologia , Fator de Crescimento do Tecido Conjuntivo/biossíntese , Fator de Crescimento do Tecido Conjuntivo/genética , Fibroblastos/metabolismo , Fibrose/tratamento farmacológico , Humanos , Inibidor 1 de Ativador de Plasminogênio/biossíntese , Inibidor 1 de Ativador de Plasminogênio/genética , Membrana Sinovial/citologia
16.
Sci Rep ; 7(1): 16192, 2017 11 23.
Artigo em Inglês | MEDLINE | ID: mdl-29170419

RESUMO

Fibrosis of the subsynovial connective tissue (SSCT) is a pathognomonic change in carpal tunnel syndrome (CTS). Identification of molecular targets and anti-fibrotic therapies could provide new treatment strategies for CTS. The contribution of SSCT cells to fibrosis and the signaling pathways that initiate and aggravate fibrosis in CTS remain unknown. Here we report that platelet-derived growth factor receptor alpha (PDGFRα) positive ( + ) cells accumulate in CTS SSCT and that the presence of fibrotic growth factor, PDGF-AA, results in increased proliferation of PDGFRα+ cells via PI3K/Akt signaling pathway. Although PI3K inhibition decreased proliferation, there was no change in fibrosis-related gene expression. Indeed, protein levels of fibrosis signaling mediator TGF-ß remained the same and the second messenger, Smad2/3, accumulated in the nucleus. In contrast AMP-activated protein kinase (AMPK) activation, which can be induced with metformin and AICAR inhibited proliferation, TGF-ß expression, and altered cell morphology in SSCT cells. Further we show that AMPK activation by metformin reduced collagen III levels and the ratio of Collagen I to Collagen III. Both AICAR and metformin reduced F-actin and significantly reduced the fiber cross alignment. Our results suggest that PDGFRa signaling may be an important fibrosis target and that activators of AMPK, may be an important therapeutic approach for treating CTS.


Assuntos
Síndrome do Túnel Carpal/metabolismo , Tecido Conjuntivo/metabolismo , Tecido Conjuntivo/patologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Síndrome do Túnel Carpal/genética , Proliferação de Células/genética , Proliferação de Células/fisiologia , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Proteína Oncogênica v-akt/genética , Proteína Oncogênica v-akt/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Fator de Crescimento Derivado de Plaquetas/genética , Fator de Crescimento Derivado de Plaquetas/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia
17.
Endocrinology ; 158(10): 3354-3368, 2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28977607

RESUMO

Endoxifen, the primary active metabolite of tamoxifen, is currently being investigated as a novel endocrine therapy for the treatment of breast cancer. Tamoxifen is a selective estrogen receptor modulator that elicits potent anti-breast cancer effects. However, long-term use of tamoxifen also induces bone loss in premenopausal women and is associated with an increased risk of endometrial cancer in postmenopausal women. For these reasons, we have used a rat model system to comprehensively characterize the impact of endoxifen on the skeleton and uterus. Our results demonstrate that endoxifen elicits beneficial effects on bone in ovary-intact rats and protects against bone loss following ovariectomy. Endoxifen is also shown to reduce bone turnover in both ovary-intact and ovariectomized rats at the cellular and biochemical levels. With regard to the uterus, endoxifen decreased uterine weight but maintained luminal epithelial cell height in ovariectomized animals. Within luminal epithelial cells, endoxifen resulted in differential effects on the expression levels of estrogen receptors α and ß as well as multiple other genes previously implicated in regulating epithelial cell proliferation and hypertrophy. These studies analyze the impact of extended endoxifen exposure on both bone and uterus using a Food and Drug Administration-recommended animal model. Although endoxifen is a more potent breast cancer agent than tamoxifen, the results of the present study demonstrate that endoxifen does not induce bone loss in ovary-intact rats and that it elicits partial agonistic effects on the uterus and skeleton in ovariectomized animals.


Assuntos
Remodelação Óssea/efeitos dos fármacos , Osso e Ossos/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Tamoxifeno/análogos & derivados , Útero/efeitos dos fármacos , Animais , Endométrio/efeitos dos fármacos , Endométrio/metabolismo , Endométrio/patologia , Receptor alfa de Estrogênio/efeitos dos fármacos , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/efeitos dos fármacos , Receptor beta de Estrogênio/metabolismo , Feminino , Tamanho do Órgão , Osteoporose/induzido quimicamente , Ovariectomia , Ratos , Moduladores Seletivos de Receptor Estrogênico/efeitos adversos , Tamoxifeno/efeitos adversos , Tamoxifeno/farmacologia , Útero/metabolismo , Útero/patologia
20.
JCI Insight ; 2(7): e90517, 2017 04 06.
Artigo em Inglês | MEDLINE | ID: mdl-28405613

RESUMO

Osteolytic bone diseases, such as osteoporosis, are characterized by diminished bone quality and increased fracture risk. The therapeutic challenge remains to maintain bone homeostasis with a balance between osteoclast-mediated resorption and osteoblast-mediated formation. Osteoclasts are formed by the fusion of monocyte/macrophage-derived precursors. Here we report, to our knowledge for the first time, that receptor-interacting protein 140 (RIP140) expression in osteoclast precursors and its protein regulation are crucial for osteoclast differentiation, activity, and coupled bone formation. In mice, monocyte/macrophage-specific knockdown of RIP140 (mϕRIP140KD) resulted in a cancellous osteopenic phenotype with significantly increased bone resorption and reduced bone formation. Osteoclast precursors isolated from mϕRIP140KD mice had significantly increased differentiation potential. Furthermore, conditioned media from mϕRIP140KD primary osteoclast cultures significantly suppressed osteoblast differentiation. This suppressive activity was effectively and rapidly terminated by specific Syk-stimulated RIP140 protein degradation. Mechanistic analysis revealed that RIP140 functions primarily by inhibiting osteoclast differentiation through forming a transcription-suppressor complex with testicular receptor 4 (TR4) to repress osteoclastogenic genes. These data reveal that monocyte/macrophage RIP140/TR4 complexes may serve as a critical transcription regulatory complex maintaining homeostasis of osteoclast differentiation, activity, and coupling with osteoblast formation. Accordingly, we propose a potentially novel therapeutic strategy, specifically targeting osteoclast precursor RIP140 protein in osteolytic bone diseases.


Assuntos
Diferenciação Celular , Macrófagos/metabolismo , Proteína 1 de Interação com Receptor Nuclear/metabolismo , Osteoclastos/citologia , Osteogênese , Receptores de Esteroides/metabolismo , Receptores dos Hormônios Tireóideos/metabolismo , Animais , Células Cultivadas , Homeostase , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína 1 de Interação com Receptor Nuclear/genética , Receptores de Esteroides/genética , Receptores dos Hormônios Tireóideos/genética
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