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1.
J Orthop Sci ; 25(1): 161-166, 2020 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-30902537

RESUMEN

PURPOSE: The purpose of this study was to investigate the biomechanical properties of load distribution following a centralization procedure for extruded lateral menisci with posterior root deficiency in a porcine model. METHODS: Six porcine knee joints were analyzed in a universal tester, as follows: 1) Intact; 2) Extrusion (meniscus extrusion was created by resecting the posterior root of the lateral meniscus, as well as the posterior synovial capsule); and 3) Centralization (two anchors were inserted at the lateral tibial plateau, and the meniscus was sutured to secure it close to the original position). Meniscus extrusion was evaluated using two markers put on the posterior cruciate ligament and the lateral meniscus, and the load distribution were assessed using a pressure mapping sensor system after applying a loading force of 200 N to the knee joint. RESULTS: Distance between two markers (mm, Average; 95% CI) was larger in the extrusion group (21.9; 17.8, 25.6) than in the intact (18.1; 15.1, 22.7) or the centralization (15.3; 12.9, 18.0) groups. The contact area (mm2) in the middle of the meniscus was significantly smaller in the extrusion group (45.8; 18.5, 73.2) than in the intact (85.7; 72.1, 99.2) or the centralization (98.3; 88.8, 107.8) groups. The maximum contact pressure (MPa) in the tibial plateau was significantly higher in the extrusion group (0.37; 0.35, 0.40) than in the intact (0.29; 0.21, 0.37) or the centralization (0.29; 0.22, 0.36) groups. CONCLUSIONS: The centralization procedure enabled a reduction of the meniscus extrusion in the lateral meniscus with posterior root deficiency and restored the maximum load and contact pressure to values close to those of the normal knee joint.

2.
Sci Rep ; 9(1): 16835, 2019 Nov 14.
Artículo en Inglés | MEDLINE | ID: mdl-31728017

RESUMEN

Mesenchymal stem cells from the synovium (synovial MSCs) are attractive for cartilage and meniscus regeneration therapy. We developed a software program that can distinguish individual colonies and automatically count the cell number per colony using time-lapse images. In this study, we investigated the usefulness of the software and analyzed colony formation in cultured synovial MSCs. Time-lapse image data were obtained for 14-day-expanded human synovial MSCs. The cell number per colony (for 145 colonies) was automatically counted from phase-contrast and nuclear-stained images. Colony growth curves from day 1 to day 14 (for 140 colonies) were classified using cluster analysis. Correlation analysis of the distribution of the cell number per colony at 14 days versus that number at 1-14 days revealed a correlation at 7 and 14 days. We obtained accurate cell number counts from phase-contrast images. Individual colony growth curves were classified into three main groups and subgroups. Our image analysis software has the potential to improve the evaluation of cell proliferation and to facilitate successful clinical applications using MSCs.

3.
Cell Transplant ; 28(11): 1445-1454, 2019 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-31313604

RESUMEN

Complex degenerative tears of the medial meniscus in the knee are usually treated using meniscectomy. However, this procedure increases the risk of osteoarthritis, while other treatments aimed at meniscal repair remain challenging due to the high possibility of failure. The use of synovial mesenchymal stem cells (MSCs) is an attractive additional approach for meniscal repair, as these cells have high proliferative and chondrogenic potential. In this case report, we surgically repaired a complex degenerative tear of the medial meniscus and then transplanted autologous synovial MSCs. We evaluated clinical outcomes at 2 years and assessed adverse events. We enrolled patients with clinical symptoms that included a feeling of instability in addition to pain caused by their complex degenerative tears of the medial meniscus. Two weeks after surgical repair of the torn meniscus, autologous synovial MSCs were transplanted onto the menisci of five patients. The total Lysholm knee score, the Knee Injury and Osteoarthritis Outcome Scale scores for "pain," "daily living," "sports activities," and the Numerical Rating Scale were significantly increased after 2 years. Three adverse events, an increase in c-reactive protein, joint effusion, and localized warmth of the knee were recorded, although these could have been due to the meniscal repair surgery. This first-in-human study confirmed that the combination of surgical repair and synovial MSC transplantation improved the clinical symptoms in patients with a complex degenerative tear of the medial meniscus. No adverse events occurred that necessitated treatment discontinuation. These findings will serve as pilot data for a future prospective study.

4.
BMC Musculoskelet Disord ; 20(1): 316, 2019 Jul 06.
Artículo en Inglés | MEDLINE | ID: mdl-31279341

RESUMEN

BACKGROUND: Synovial mesenchymal stem cells (MSCs) are an attractive cell source for cartilage and meniscus regeneration. The optimum cryopreservation medium has not been determined, but dimethylsulfoxide (DMSO) should be excluded, if possible, because of its toxicity. The purposes of our study were to examine the possible benefits of higher concentrations of serum and the effectiveness of 100% serum (without DMSO) for the cryopreservation of synovial MSCs. METHODS: Human synovium was harvested from the knees of four donors with osteoarthritis during total knee arthroplasty. Synovial MSCs (8 × 105 cells) were suspended in 400 µL medium and used as a Time 0 control. The same number of synovial MSCs was also suspended in 400 µL α-MEM medium containing 10% fetal bovine serum (FBS) (5% DMSO, and 1% antibiotic), 95% FBS (and 5% DMSO), or 100% FBS (no DMSO) and cryopreserved at - 80 °C for 7 days. After thawing, the cell suspensions (1.5 µL; 3 × 103 cells) were cultured in 60 cm2 dishes for 14 days for colony formation assays. Additional 62.5 µL samples of cell suspensions (1.25 × 105 cells) were added to tubes and cultured for 21 days for chondrogenesis assays. RESULTS: Colony numbers were significantly higher in the Time 0 and 95% FBS groups than in the 10% FBS group (n = 24). Colony numbers were much lower in the 100% FBS group than in the other three groups. The cell numbers per dish reflected the colony numbers. Cartilage pellet weights were significantly heavier in the 95% FBS group than in the 10% FBS group, whereas no difference was observed between the Time 0 and the 95% FBS groups (n = 24). No cartilage pellets formed at all in the 100% FBS group. CONCLUSION: Synovial MSCs cryopreserved in 95% FBS with 5% DMSO maintained their colony formation and chondrogenic abilities to the same levels as observed in the cells before cryopreservation. Synovial MSCs cryopreserved in 100% FBS lost their colony formation and chondrogenic abilities.


Asunto(s)
Condrogénesis/efectos de los fármacos , Criopreservación/métodos , Crioprotectores/farmacología , Células Madre Mesenquimatosas , Membrana Sinovial/citología , Anciano , Técnicas de Cultivo de Célula , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Crioprotectores/química , Dimetilsulfóxido/química , Dimetilsulfóxido/farmacología , Femenino , Humanos , Articulación de la Rodilla/citología , Trasplante de Células Madre Mesenquimatosas , Osteoartritis/terapia , Suero/química
5.
J Orthop Res ; 37(11): 2466-2475, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-31115925

RESUMEN

The meniscus functions as a load distributor and secondary stabilizer in the knee, and the loss of the meniscus increases the risk of osteoarthritis. Freeze-thawed menisci are used in clinical practice to replace defective menisci; however, the disadvantages of freeze-thawed tissues include disease transmission and immune rejection. In this study, we decellularized menisci using high hydrostatic pressure (HHP) and compared the decellularized menisci with freeze-thawed menisci. Porcine menisci were either pressurized at 1,000 MPa for 10 min and then washed with DNase solution or frozen at -80°C for 2 days and thawed. These menisci then underwent in vitro histological, biochemical, and biomechanical comparisons with native menisci. The HHP-treated and freeze-thawed menisci were also subcutaneously implanted in a pig, and later harvested for histological analysis. The numbers of histologically detected cells were significantly lower and the amount of biochemically detected DNA was approximately 100-fold lower in HHP-treated than in native and freeze-thawed menisci. The compression strength of the HHP-decellularized menisci was decreased after 1 and 50 cycles at 20% strain but was unchanged in the freeze-thawed menisci. After implantation, the numbers of multinucleated giant cells were significantly lower around the HHP-treated menisci than around the freeze-thawed menisci. Recellularization of the HHP-decellularized menisci was confirmed. Thus, although the HHP-decellularized menisci were mechanically inferior to the freeze-thawed meniscus in vitro, they were immunologically superior. Our study is the first to demonstrate the use of HHP for decellularization of the meniscus. © 2019 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res 37:2466-2475, 2019.


Asunto(s)
Aloinjertos , Meniscos Tibiales/trasplante , Animales , Congelación , Presión Hidrostática , Porcinos
6.
J Orthop Res ; 37(6): 1350-1357, 2019 06.
Artículo en Inglés | MEDLINE | ID: mdl-29737046

RESUMEN

In vitro chondrogenesis of mesenchymal stem cells (MSCs) mimics in vivo chondrogenesis of MSCs. However, the size of the cartilage pellets that can be attained in vitro is limited by current methods; therefore, some modifications are required to obtain larger pellets. Petaloid pieces of recombinant peptide (petaloid RCP) have the advantage of creating spaces between cells in culture. The RCP used here is based on the alpha-1 sequence of human collagen type I and contains 12 Arg-Gly-Asp motifs. We examined the effect and mechanisms of adding petaloid RCP on the in vitro chondrogenesis of human synovial MSCs by culturing 125k cells with or without 0.125 mg petaloid RCP in chondrogenic medium for 21 days. The cartilage pellets were sequentially analyzed by weight, sulfated glycosaminoglycan content, DNA retention, and histology. Petaloid RCP significantly increased the weight of the cartilage pellets: The petaloid RCP group weighed 7.7 ± 1.2 mg (n = 108), whereas the control group weighed 5.3 ± 1.6 mg. Sulfated glycosaminoglycan and DNA contents were significantly higher in the petaloid RCP group than in the control group. Light and transmission electron microscopy images showed that the petaloid RCP formed the framework of the pellet at day 1, the framework was broken by production of cartilage matrix by the synovial MSCs at day 7, and the cartilage pellet grew larger, with diffuse petaloid RCP remaining, at day 21. Therefore, petaloid RCP formed a framework for the pellet, maintained a higher cell number, and promoted in vitro cartilage formation of synovial MSCs. © 2018 The Authors. Journal of Orthopaedic Research® Published by Wiley Periodicals, Inc. J Orthop Res 37:1350-1357, 2019.


Asunto(s)
Condrogénesis/efectos de los fármacos , Células Madre Mesenquimatosas/citología , Péptidos/farmacología , Membrana Sinovial/citología , Anciano , Anciano de 80 o más Años , Diferenciación Celular , Femenino , Glicosaminoglicanos/metabolismo , Humanos , Proteínas Recombinantes/farmacología
7.
PLoS One ; 13(8): e0202922, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30138399

RESUMEN

Osteoarthritis (OA), a common chronic joint disorder in both humans and canines, is characterized by a progressive loss of articular cartilage. Canines can serve as an animal model of OA for human medicine, and this research can simultaneously establish effective veterinary treatments for canine OA. One attractive treatment that can lead to cartilage regeneration is the use of mesenchymal stem cells (MSCs). However, for canine OA, little information is available regarding the best source of MSCs. The purpose of this study was to identify a promising MSC source for canine cartilage regeneration. We collected synovial, infrapatellar fat pad, inguinal adipose, and bone marrow tissues from six canines and then conducted a donor-matched comparison of the properties of MSCs derived from these four tissues. We examined the surface epitope expression, proliferation capacity, and trilineage differentiation potential of all four populations. Adherent cells derived from all four tissue sources exhibited positivity for CD90 and CD44 and negativity for CD45 and CD11b. The positive rate for CD90 was higher for synovium-derived than for adipose-derived and bone marrow-derived MSCs. Synovium-derived and infrapatellar fat pad-derived MSCs displayed substantial proliferation ability, and all four populations underwent trilineage differentiation. During chondrogenesis, the wet weight was heavier for cartilage pellets derived from synovium MSCs than from the other three sources. The synovium is therefore a promising source for MSCs for canine cartilage regeneration. Our findings provide useful information about canine MSCs that may be applicable to regenerative medicine for treatment of OA.


Asunto(s)
Tejido Adiposo/metabolismo , Cartílago/metabolismo , Condrogénesis/fisiología , Células Madre Mesenquimatosas/metabolismo , Membrana Sinovial/metabolismo , Tejido Adiposo/citología , Tejido Adiposo/fisiología , Animales , Técnicas de Cultivo de Célula/veterinaria , Diferenciación Celular , Proliferación Celular , Tratamiento Basado en Trasplante de Células y Tejidos/veterinaria , Perros , Femenino , Masculino , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/fisiología , Osteoartritis/terapia , Osteoartritis/veterinaria , Regeneración , Medicina Regenerativa , Membrana Sinovial/citología
8.
Stem Cell Res Ther ; 9(1): 123, 2018 05 02.
Artículo en Inglés | MEDLINE | ID: mdl-29720268

RESUMEN

BACKGROUND: Synovial mesenchymal stem cells (MSCs) are an attractive cell source for cartilage and meniscus regeneration. Synovial tissue can be histologically classified into three regions; surface, stromal and perivascular region, but the localization of synovial MSCs has not been fully investigated. We identified markers specific for each region, and compared properties of MSCs derived from each region in the synovium. METHODS: The intensity of immunostaining with 19 antibodies was examined for surface, stromal, and perivascular regions of human synovium from six osteoarthritis patients. Specific markers were identified and synovial cells derived from each region were sorted. Proliferation, surface marker expression, chondrogenesis, calcification and adipogenesis potentials were compared in synovial MSCs derived from the three regions. RESULTS: We selected CD55+ CD271- for synovial cells in the surface region, CD55- CD271- in the stromal region, and CD55- CD271+ in the perivascular region. The ratio of the sorted cells to non-hematopoietic lineage cells was 5% in the surface region, 70% in the stromal region and 15% in the perivascular region. Synovial cells in the perivascular fraction had the greatest proliferation potential. After expansion, surface marker expression profiles and adipogenesis potentials were similar but chondrogenic and calcification potentials were higher in synovial MSCs derived from the perivascular region than in those derived from the surface and stromal regions. CONCLUSIONS: We identified specific markers to isolate synovial cells from the surface, stromal, and perivascular regions of the synovium. Synovial MSCs in the perivascular region had the highest proliferative and chondrogenic potentials among the three regions.


Asunto(s)
Biomarcadores/metabolismo , Células Madre Mesenquimatosas/metabolismo , Microscopía Electrónica de Transmisión/métodos , Membrana Sinovial/metabolismo , Anciano , Anciano de 80 o más Años , Diferenciación Celular , Citometría de Flujo , Humanos , Persona de Mediana Edad
9.
J Orthop Sci ; 23(4): 676-681, 2018 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-29724468

RESUMEN

BACKGROUND: Meniscus surgery is the most commonly performed orthopedic surgery, and despite recent emphasis on saving the meniscus, the current status of meniscus surgeries is little known in many countries, including Japan. The National Database of Health Insurance Claims and Specific Health Checkups of Japan and the Statistics of Medical Care Activities in Public Health Insurance track meniscus surgeries through health insurance claims. The National Database provides the numbers for 2014 and 2015, and the Statistics of Medical Care Activities provides the numbers from June 2011 to June 2016. Our aim was to analyze isolated meniscus surgery numbers and meniscus repair ratios by age group based on the National Database and evaluate trends of meniscus repair ratios for the latest six years from the Statistics of Medical Care Activities. METHODS: Meniscus surgeries by age group were counted from the National Database for 2014-2015, and meniscus repair ratios (meniscus repairs/meniscus surgeries) were calculated. The numbers were also counted from the Statistics of Medical Care Activities in 2011-2016. For statistical analysis of annual trends of meniscus repair ratios, the Cochran-Armitage trend test was used. Meniscus surgeries with concomitant knee ligament surgeries were excluded. RESULTS: According to the National Database, isolated meniscus surgeries totaled 34,966 in 2015, with peak ages of patients in their late teens and 60s. The meniscus repair ratio was 19% in 2014 and 24% in 2015. According to the Statistics of Medical Care Activities, the meniscus repair ratio was 9% in 2011 and significantly increased to 25% in 2016 (p = 0.0008). The ratio also increased significantly in each age group between the early 20s and late 70s. CONCLUSIONS: Approximately 35,000 meniscus surgeries are performed in Japan annually, with peak ages in the late teens and 60s. The number of meniscus repairs has increased over the past six years.


Asunto(s)
Meniscectomía/tendencias , Meniscos Tibiales/cirugía , Lesiones de Menisco Tibial/cirugía , Adolescente , Adulto , Distribución por Edad , Anciano , Anciano de 80 o más Años , Estudios de Cohortes , Bases de Datos Factuales , Femenino , Humanos , Incidencia , Japón/epidemiología , Masculino , Meniscectomía/métodos , Meniscos Tibiales/fisiopatología , Persona de Mediana Edad , Complicaciones Posoperatorias/epidemiología , Complicaciones Posoperatorias/fisiopatología , Pronóstico , Estudios Retrospectivos , Medición de Riesgo , Distribución por Sexo , Lesiones de Menisco Tibial/diagnóstico por imagen , Lesiones de Menisco Tibial/epidemiología , Resultado del Tratamiento , Adulto Joven
10.
Clin Calcium ; 28(3): 319-327, 2018.
Artículo en Japonés | MEDLINE | ID: mdl-29512522

RESUMEN

Cartilage injury remains one of the common clinical problems due to its limited regeneration capacities. Meniscectomy commonly performed for meniscus injury leads to osteoarthritis, but the indication of meniscus repair is limited. We have identified that synovial mesenchymal stem cells(MSCs)are superior to MSCs derived from other tissues in proliferation capacity and in vitro/in vivo chondrogenic potentials. When suspension of synovial MSCs was placed on the cartilage defect for 10 minutes, 60% of the cells attached to the defect site. Based on these basic researches, we started a clinical study for cartilage regeneration by arthroscopic transplantation of synovial MSCs. Additionally, we transplanted synovial MSCs for meniscus injury after meniscus repair. We obtained good clinical results of cartilage regeneration and meniscus healing without any serious side effects. Transplantation of synovial MSCs will be useful for cartilage or meniscus injuries.


Asunto(s)
Enfermedades de los Cartílagos/terapia , Homeostasis , Menisco , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas , Regeneración , Animales , Humanos , Membrana Sinovial
11.
BMC Musculoskelet Disord ; 19(1): 78, 2018 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-29523119

RESUMEN

BACKGROUND: Mobilization of mesenchymal stem cells (MSCs) from the synovium was revealed using a "suspended synovium culture model" of osteoarthritis (OA). The pathology of rheumatoid arthritis (RA) differs from that of OA. We investigated whether mobilization of MSCs from the synovium also occurred in RA, and we compared the properties of synovial MSCs collected from suspended synovium culture models of RA and OA. METHODS: Human synovium was harvested during total knee arthroplasty from the knee joints of patients with RA (n = 8) and OA (n = 6). The synovium was suspended in a bottle containing culture medium and a culture dish at the bottom. Cells were harvested from the dish and analyzed. RESULTS: No significant difference was observed between RA and OA in the harvested cell numbers per g of synovium. However, the variation in the number of cells harvested from each donor was greater for RA than for OA. The harvested cells were multipotent and no difference was observed in the cartilage pellet weight between RA and OA. The surface epitopes of the cells in RA and OA were similar to those of MSCs. CONCLUSION: Mobilization of MSCs from the synovium was demonstrated using a suspended synovium culture model for RA. The harvested cell numbers, chondrogenic potentials, and surface epitope profiles were comparable between the RA and OA models.


Asunto(s)
Artritis Reumatoide/patología , Técnicas de Cultivo de Célula/métodos , Células Madre Mesenquimatosas/fisiología , Osteoartritis/patología , Membrana Sinovial/citología , Membrana Sinovial/fisiología , Adulto , Anciano , Células Cultivadas , Femenino , Humanos , Masculino , Persona de Mediana Edad
12.
Stem Cell Res Ther ; 8(1): 144, 2017 06 13.
Artículo en Inglés | MEDLINE | ID: mdl-28610596

RESUMEN

BACKGROUND: In our clinical practice, we perform transplantations of autologous synovial mesenchymal stem cells (MSCs) for cartilage and meniscus regenerative medicine. One of the most important issues to ensuring clinical efficacy involves the transport of synovial MSCs from the processing facility to the clinic. Complete human serum (100% human serum) is an attractive candidate material in which to suspend synovial MSCs for their preservation during transport. The purpose of this study was to investigate whether complete human serum maintained MSC viability and chondrogenic potential and to examine the optimal temperature conditions for the preservation of human synovial MSCs. METHODS: Human synovium was harvested from the knees of 14 donors with osteoarthritis during total knee arthroplasty. Passage 2 synovial MSCs were suspended at 2 million cells/100 µL in Ringer's solution or complete human serum at 4, 13, and 37 °C for 48 h. These cells were analyzed for live cell rates, cell surface marker expression, metabolic activity, proliferation, and adipogenic, calcification, and chondrogenic differentiation potentials before and after preservation. RESULTS: After preservation, synovial MSCs maintained higher live cell rates in human serum than in Ringer's solution at 4 and 13 °C. Synovial MSCs preserved in human serum at 4 and 13 °C also maintained high ratios of propidium iodide- and annexin V- cells. MSC surface marker expression was not altered in cells preserved at 4 and 13 °C. The metabolic activities of cells preserved in human serum at 4 and 13 °C was maintained, while significantly reduced in other conditions. Replated MSCs retained their proliferation ability when preserved in human serum at 4 and 13 °C. Adipogenesis and calcification potential could be observed in cells preserved in each condition, whereas chondrogenic potential was retained only in cells preserved in human serum at 4 and 13 °C. CONCLUSION: The viability and chondrogenic potential of synovial MSCs were maintained when the cells were suspended in human serum at 4 and 13 °C.


Asunto(s)
Diferenciación Celular/genética , Trasplante de Células Madre Mesenquimatosas , Osteoartritis/genética , Membrana Sinovial/trasplante , Cartílago Articular/citología , Cartílago Articular/crecimiento & desarrollo , Condrocitos/citología , Condrocitos/metabolismo , Condrogénesis/genética , Humanos , Células Madre Mesenquimatosas/citología , Osteoartritis/patología , Osteoartritis/terapia , Medicina Regenerativa , Membrana Sinovial/citología
13.
Stem Cell Res Ther ; 8(1): 115, 2017 05 16.
Artículo en Inglés | MEDLINE | ID: mdl-28511664

RESUMEN

BACKGROUND: Mesenchymal stem cells derived from the synovial membrane (synovial MSCs) are a candidate cell source for regenerative medicine of cartilage and menisci due to their high chondrogenic ability. Regenerative medicine can be expected for RA patients with the inflammation well-controlled as well as OA patients and transplantation of synovial MSCs would also be a possible therapeutic treatment. Some properties of synovial MSCs vary dependent on the diseases patients have, and whether or not the pathological condition of RA affects the chondrogenesis of synovial MSCs remains controversial. The purpose of this study was to compare the properties of primary synovial MSCs between RA and OA patients. METHODS: Human synovial tissue was harvested during total knee arthroplasty from the knee joints of eight patients with RA and OA respectively. Synovial nucleated cells were cultured for 14 days. Total cell yields, surface markers, and differentiation potentials were analyzed for primary synovial MSCs. RESULTS: Nucleated cell number per 1 mg synovium was 8.4 ± 3.9 thousand in RA and 8.0 ± 0.9 thousand in OA. Total cell number after 14-day culture/1 mg synovium was 0.7 ± 0.4 million in RA and 0.5 ± 0.3 million in OA, showing no significant difference between in RA and OA. Cells after 14-day culture were mostly positive for CD44, CD73, CD90, CD105, negative for CD45 both in RA and OA. There was no significant difference for the cartilage pellet weight and sGAG content per pellet between in RA and OA. Both oil red O-positive colony rate and alizarin red-positive colony rate were similar in RA and OA. CONCLUSIONS: Yields, surface markers and chondrogenic potential of primary synovial MSCs in RA were comparable to those in OA. Synovium derived from RA patients can be the cell source of MSCs for cartilage and meniscus regeneration.


Asunto(s)
Artritis Reumatoide/patología , Condrogénesis , Células Madre Mesenquimatosas/patología , Osteoartritis/patología , Membrana Sinovial/patología , Adipogénesis , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/metabolismo , Calcificación Fisiológica , Recuento de Células , Núcleo Celular/metabolismo , Forma de la Célula , Ensayo de Unidades Formadoras de Colonias , Demografía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Tamaño de los Órganos
14.
J Orthop Sci ; 22(3): 542-548, 2017 May.
Artículo en Inglés | MEDLINE | ID: mdl-28351717

RESUMEN

BACKGROUND: Meniscus extrusion often observed in knee osteoarthritis has a strong correlation with the progression of cartilage degeneration and symptom in the patients. We recently reported a novel procedure "arthroscopic centralization" in which the capsule was sutured to the edge of the tibial plateau to reduce meniscus extrusion in the human knee. However, there is no animal model to study the efficacy of this procedure. The purposes of this study were [1] to establish a model of centralization for the extruded medial meniscus in a rat model; and [2] to investigate the chondroprotective effect of this procedure. METHODS: Medial meniscus extrusion was induced by the release of the anterior synovial capsule and the transection of the meniscotibial ligament. Centralization was performed by the pulled-out suture technique. Alternatively, control rats had only the medial meniscus extrusion surgery. Medial meniscus extrusion was evaluated by micro-CT and macroscopic findings. Cartilage degeneration of the medial tibial plateau was evaluated macroscopically and histologically. RESULTS: By micro-CT analysis, the medial meniscus extrusion was significantly improved in the centralization group in comparison to the extrusion group throughout the study. Both macroscopically and histologically, the cartilage lesion of the medial tibial plateau was prevented in the centralization group but was apparent in the control group. CONCLUSIONS: We developed medial meniscus extrusion in a rat model, and centralization of the extruded medial meniscus by the pull-out suture technique improved the medial meniscus extrusion and delayed cartilage degeneration, though the effect was limited. Centralization is a promising treatment to prevent the progression of osteoarthritis.


Asunto(s)
Cartílago Articular/diagnóstico por imagen , Meniscos Tibiales/diagnóstico por imagen , Lesiones de Menisco Tibial/diagnóstico , Animales , Artroscopía/métodos , Cartílago Articular/cirugía , Modelos Animales de Enfermedad , Imagen por Resonancia Magnética , Masculino , Ratas , Ratas Endogámicas Lew , Lesiones de Menisco Tibial/metabolismo
15.
BMC Musculoskelet Disord ; 18(1): 36, 2017 01 25.
Artículo en Inglés | MEDLINE | ID: mdl-28122526

RESUMEN

BACKGROUND: It is still debated whether strenuous running in the inflammatory phase produces beneficial or harmful effect in rat knees. We examined (1) the dropout rate of rats during a 30-km running protocol, (2) influences of strenuous running and/or low amounts of mono-iodoacetate injection on cartilage, and (3) the effect of strenuous running on synovitis. METHODS: Rats were forced to run 30 km over 6 weeks and the dropout rate was examined. One week after 0.1 mg mono-iodoacetate was injected into the right knee, rats were forced to run either 15 km or not run at all over 3 weeks, after which knee cartilage was evaluated. Synovium at the infrapatellar fat pad was also examined histologically. RESULTS: Even though all 12 rats run up to 15 km, only 6 rats completed 30 km of running. Macroscopically, 0.1 mg mono-iodoacetate induced erosion at the tibial cartilage irrespective of 15 km of running. Histologically, 0.1 mg mono-iodoacetate induced loss of cartilage matrix in the tibial cartilage, and an additional 15 km of strenuous running significantly exacerbated the loss. Synovitis caused by mono-iodoacetate improved after running. CONCLUSIONS: Only 50% of rats completed 30 km of running because of foot problems. Strenuous running further exacerbated tibial cartilage erosion but did not influence synovitis induced by mono-iodoacetate.


Asunto(s)
Cartílago Articular/patología , Yodoacetatos/toxicidad , Articulación de la Rodilla/patología , Carrera/tendencias , Animales , Cartílago Articular/efectos de los fármacos , Inyecciones Intraarticulares , Yodoacetatos/administración & dosificación , Articulación de la Rodilla/efectos de los fármacos , Masculino , Ratas , Ratas Wistar , Estrés Mecánico
16.
Arthroscopy ; 33(4): 800-810, 2017 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-28043752

RESUMEN

PURPOSE: To develop an in vitro model, the "suspended synovium culture model," to demonstrate the mobilization of mesenchymal stem cells (MSCs) from the synovium into a noncontacted culture dish through culture medium. In addition, to examine which synovium, fibrous synovium or adipose synovium, released more MSCs in the knee with osteoarthritis. METHODS: Human synovial tissue was harvested during total knee arthroplasty from knee joints of 34 patients with osteoarthritis (28 patients: only fibrous synovium, 6 patients: fibrous and adipose synovium). One gram of synovium was suspended with a thread in a bottle containing 40 mL of culture medium and a 3.5-cm-diameter culture dish at the bottom. After 7 days, the culture dish in the bottle was examined. For the cells harvested, multipotentiality and surface epitopes were analyzed. The numbers of colonies derived from fibrous synovium and adipose synovium were also compared. RESULTS: Colonies of spindle-shaped cells were observed in the culture dish in all 28 donors. Colonies numbered 26 on average, and the cells derived from colony-forming cells had multipotentiality for chondrogenesis, adipogenesis, calcification, and surface epitopes similar to MSCs. The number was colonies was significantly higher in fibrous synovium than in adipose synovium (P < .05, n = 6). CONCLUSIONS: We developed a suspended synovium culture model. Suspended synovium was able to release MSCs into a noncontacted culture dish through medium in a bottle. Fibrous synovium was found to release greater numbers of MSCs than adipose synovium in our culture model. CLINICAL RELEVANCE: This model could be a valuable tool to screen drugs capable of releasing MSCs from the synovium into synovial fluid.


Asunto(s)
Tejido Adiposo/patología , Células Madre Mesenquimatosas/patología , Osteoartritis de la Rodilla/patología , Membrana Sinovial/patología , Adipogénesis/fisiología , Anciano , Anciano de 80 o más Años , Diferenciación Celular , Células Cultivadas , Condrogénesis/fisiología , Ensayo de Unidades Formadoras de Colonias , Medios de Cultivo , Femenino , Fibrosis , Humanos , Articulación de la Rodilla/patología , Masculino , Células Madre Mesenquimatosas/fisiología , Persona de Mediana Edad , Osteoartritis de la Rodilla/metabolismo , Osteoartritis de la Rodilla/cirugía , Técnicas de Cultivo de Tejidos
17.
BMC Musculoskelet Disord ; 17: 188, 2016 Apr 27.
Artículo en Inglés | MEDLINE | ID: mdl-27118194

RESUMEN

BACKGROUND: Cross-linked hyaluronan--also called Hylan G-F 20--is a medical device developed to treat osteoarthritis of the knee. However, it is still controversial whether Hylan G-F 20 has a cartilage protective effect in trauma-induced osteoarthritis. We investigated whether Hylan G-F 20 delayed osteoarthritis progression in a partial meniscectomized rat model. METHODS: Lewis rats were used for the experiments. The anterior medial meniscus was resected at the level of the medial collateral ligament in both knees. From 1 week after the surgery, 50 µl of Hylan G-F 20 was injected weekly into the left knee and phosphate buffered saline was injected into the right knee. Cartilage was evaluated for macroscopic findings, histology with safranin-o, and expression of type II collagen at 2, 4, and 8 weeks. Synovitis was also evaluated, and immunohistochemical analysis was performed for ED1. RESULTS: Macroscopic findings demonstrated that India ink positive area, representing fibrillated cartilage, was significantly smaller in the Hylan G-F 20 group than in the control group at 2, 4, and 8 weeks (n = 5). There were no significant differences in osteophyte score between the Hylan G-F 20 group and the control group at 2, 4, and 8 weeks. Histologically, the cartilage in the medial tibial plateau was destroyed at 8 weeks in the control group, while type II collagen expression was still observed at 8 weeks in the Hylan G-F 20 group. OARSI score for cartilage histology was significantly lower in the Hylan G-F 20 group than in the control group at 4 and 8 weeks (n = 5). There were no significant differences in synovial cell number or modified synovitis score between the Hylan G-F 20 group and the control group at 2, 4, and 8 weeks (n = 5). In the Hylan G-F 20 group, foreign bodies surrounded by ED1 positive macrophages were observed in the synovium. CONCLUSION: Weekly injections of Hylan G-F 20 starting 1 week after surgery delayed cartilage degeneration after meniscectomy in a rat model. Synovitis induced by meniscectomy was not alleviated by Hylan G-F 20. Insoluble gels were observed in the synovium after the Hylan G-F 20 injection.


Asunto(s)
Materiales Biocompatibles/administración & dosificación , Ácido Hialurónico/análogos & derivados , Meniscos Tibiales/cirugía , Osteoartritis de la Rodilla/prevención & control , Osteoartritis de la Rodilla/cirugía , Animales , Enfermedades de los Cartílagos/patología , Enfermedades de los Cartílagos/prevención & control , Enfermedades de los Cartílagos/cirugía , Esquema de Medicación , Ácido Hialurónico/administración & dosificación , Articulación de la Rodilla/efectos de los fármacos , Articulación de la Rodilla/patología , Articulación de la Rodilla/cirugía , Masculino , Meniscos Tibiales/efectos de los fármacos , Meniscos Tibiales/patología , Osteoartritis de la Rodilla/patología , Ratas , Ratas Endogámicas Lew
18.
PLoS One ; 11(2): e0148777, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26867127

RESUMEN

OBJECTIVE: Lubricin expression in the superficial cartilage will be a crucial factor in the success of cartilage regeneration. Mesenchymal stem cells (MSCs) are an attractive cell source and the use of aggregates of MSCs has some advantages in terms of chondrogenic potential and efficiency of cell adhesion. Lubricin expression in transplanted MSCs has not been fully elucidated so far. Our goals were to determine (1) whether cartilage pellets of human MSCs expressed lubricin in vitro chondrogenesis, (2) whether aggregates of human MSCs promoted lubricin expression, and (3) whether aggregates of MSCs expressed lubricin in the superficial cartilage after transplantation into osteochondral defects in rats. METHODS: For in vitro analysis, human bone marrow (BM) MSCs were differentiated into cartilage by pellet culture, and also aggregated using the hanging drop technique. For an animal study, aggregates of BM MSCs derived from GFP transgenic rats were transplanted to the osteochondral defect in the trochlear groove of wild type rat knee joints. Lubricin expression was mainly evaluated in differentiated and regenerated cartilages. RESULTS: In in vitro analysis, lubricin was detected in the superficial zone of the pellets and conditioned medium. mRNA expression of Proteoglycan4 (Prg4), which encodes lubricin, in pellets was significantly higher than that of undifferentiated MSCs. Aggregates showed different morphological features between the superficial and deep zone, and the Prg4 mRNA expression increased after aggregate formation. Lubricin was also found in the aggregate. In a rat study, articular cartilage regeneration was significantly better in the MSC group than in the control group as shown by macroscopical and histological analysis. The transmission electron microscope showed that morphology of the superficial cartilage in the MSC group was closer to that of the intact cartilage than in the control group. GFP positive cells remained in the repaired tissue and expressed lubricin in the superficial cartilage. CONCLUSION: Cartilage derived from MSCs expressed lubricin protein both in vitro and in vivo. Aggregation promoted lubricin expression of MSCs in vitro and transplantation of aggregates of MSCs regenerated cartilage including the superficial zone in a rat osteochondral defect model. Our results indicate that aggregated MSCs could be clinically relevant for therapeutic approaches to articular cartilage regeneration with an appropriate superficial zone in the future.


Asunto(s)
Células de la Médula Ósea/metabolismo , Cartílago/metabolismo , Regulación de la Expresión Génica , Glicoproteínas/biosíntesis , Células Madre Mesenquimatosas/metabolismo , Animales , Células de la Médula Ósea/citología , Diferenciación Celular , Células Cultivadas , Condrogénesis , Femenino , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Articulación de la Rodilla/fisiopatología , Trasplante de Células Madre Mesenquimatosas/métodos , Microscopía Electrónica de Transmisión , Proteoglicanos/biosíntesis , Proteoglicanos/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Transgénicas , Regeneración , Microtomografía por Rayos X
19.
Stem Cell Res Ther ; 6: 243, 2015 Dec 10.
Artículo en Inglés | MEDLINE | ID: mdl-26652649

RESUMEN

INTRODUCTION: For expansion of human mesenchymal stem cells (MSCs), autologous human serum is safer than fetal bovine serum in clinical situations. One of the problems with the use of autologous human serum is that its proliferative effect on MSCs varies widely between donors. The threefold goals of this study were: (1) to demonstrate an improved method for preparing human serum; (2) to identify growth factors predictive of proliferative potential; and (3) to identify a cytokine to promote MSC proliferation in human serum. METHODS: Fresh blood was collected using a closed bag system containing glass beads. The bag was shaken at 20 °C for 30 minutes for rapid preparation, or kept stationary at 4 °C for 24 hours for slow preparation. Passage 0 synovial MSCs derived from four donors were cultured with 10 % conventional rapid preparation serum or modified slow preparation serum from four different donors. To perform the colony-forming unit assay, synovial MSCs were cultured in these serums. The protein expression profile in serum was analyzed using cytokine array. The candidate proteins were speculated from the correlation between the colony-forming ability and protein expression. As an evaluation of the candidate proteins, proliferation ability, surface marker phenotype and differentiation capability of synovial MSCs were examined. RESULTS: Compared with rapid preparation serum, slow preparation serum resulted in a significantly higher total colony number and twofold higher expression levels of nine proteins (angiopoietin-1, BDNF, EGF, ENA-78, IGFBP-2, platelet-derived growth factor (PDGF)-AA, PDGF-AB/BB, RANTES and TfR). Colony number was positively correlated with PDGF-AA/AB concentrations. Exogenous PDGF-AA significantly promoted proliferation of synovial MSCs, whereas PDGF receptor (PDGFR) inhibitor decreased it. Addition of PDGFs or PDGFR inhibitor did not affect surface epitopes of synovial MSCs. Pretreatment with PDGFs or PDGFR inhibitor did not affect chondrogenic, adipogenic, or calcification potentials of synovial MSCs. CONCLUSION: Slow preparation serum contained higher concentrations of PDGF-AA/AB and increased the colony formation number of synovial MSCs. PDGF-AA/AB were indicators of the proliferative potential of human serum. Exogenous PDGF-AA increased proliferation of synovial MSCs without alteration of surface epitopes and differentiation potentials.


Asunto(s)
Células Madre Mesenquimatosas/citología , Células Madre Mesenquimatosas/metabolismo , Factor de Crecimiento Derivado de Plaquetas/metabolismo , Membrana Sinovial/citología , Membrana Sinovial/metabolismo , Adulto , Diferenciación Celular , Proliferación Celular , Células Cultivadas , Ensayo de Unidades Formadoras de Colonias , Medios de Cultivo/metabolismo , Citocinas/sangre , Humanos , Técnicas In Vitro , Masculino
20.
Stem Cells ; 33(6): 1927-38, 2015 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-25993981

RESUMEN

Although meniscus defects and degeneration are strongly correlated with the later development of osteoarthritis, the promise of regenerative medicine strategies is to prevent and/or delay the disease's progression. Meniscal reconstruction has been shown in animal models with tendon grafting and transplantation of mesenchymal stem cells (MSCs); however, these procedures have not shown the same efficacy in clinical studies. Here, our aim was to investigate the ability of tendon grafts pretreated with exogenous synovial-derived MSCs to prevent cartilage degeneration in a rat partial meniscus defect model. We removed the anterior half of the medial meniscus and grafted autologous Achilles tendons with or without a 10-minute pretreatment of the tendon with synovial MSCs. The meniscus and surrounding cartilage were evaluated at 2, 4, and 8 weeks (n = 5). Tendon grafts increased meniscus size irrespective of synovial MSCs. Histological scores for regenerated menisci were better in the tendon + MSC group than in the other two groups at 4 and 8 weeks. Both macroscopic and histological scores for articular cartilage were significantly better in the tendon + MSC group at 8 weeks. Implanted synovial MSCs survived around the grafted tendon and native meniscus integration site by cell tracking assays with luciferase+, LacZ+, DiI+, and/or GFP+ synovial MSCs and/or GFP+ tendons. Flow cytometric analysis showed that transplanted synovial MSCs retained their MSC properties at 7 days and host synovial tissue also contained cells with MSC characteristics. Synovial MSCs promoted meniscus regeneration augmented by autologous Achilles tendon grafts and prevented cartilage degeneration in rats.


Asunto(s)
Tendón Calcáneo/citología , Diferenciación Celular/fisiología , Células Madre Mesenquimatosas/citología , Regeneración/fisiología , Membrana Sinovial/citología , Animales , Cartílago Articular , Modelos Animales de Enfermedad , Masculino , Trasplante de Células Madre Mesenquimatosas/métodos , Ratas , Medicina Regenerativa/métodos
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