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In Vitro and In Vivo Comparison of Different Types of Rabbit Mesenchymal Stem Cells for Cartilage Repair.

ABSTRACT: Objective:Systematic studies indicate a growing number of clinical studies that use mesenchymal stem cells (MSCs) for the treatment of cartilage lesions. The current experimental and preclinical study aims to comparatively evaluate the potential of MSCs from a variety of tissues for the treatment of cartilage defect in rabbit's knee which has not previously been reported. Materials and Methods:In this experimental study, MSCs isolated from bone marrow (BMMSCs), adipose (AMSCs), and ears (EMSCs) of rabbits and expanded under in vitro culture. The growth rate and differentiation ability of MSCs into chondrocyte and the formation of cartilage pellet were investigated by drawing the growth curve and real-time polymerase chain reaction (RT-PCR), respectively. Then, the critical cartilage defect was created on the articular cartilage (AC) of the rabbit distal femur, and MSCs in collagen carrier were transplanted. The studied groups were as the control (only defect), sham (defect with scaffold), BMMSCs in the scaffold, EMSCs in the scaffold, and EMSCs in the scaffold with cartilage pellets. Histological and the gene expression analysis were performed following the transplantation. Results:Based on our comparative in vitro investigation, AMSCs possessed the highest growth rate, as well as the lowest chondrogenic differentiation potential. In this context, MSCs of the ear showed a significantly higher growth rate and cartilage differentiation potential than those of bone marrow tissue (P<0.05). According to our in vivo assessments, BMMSC- and EMSC-seeded scaffolds efficiently improved the cartilage defect 4 weeks post-transplantation, while no improvement was observed in the group contained the cartilage pellets. Conclusion:It seems that the ear contains MSCs that promote cartilage regeneration as much as the conventional MSCs from the bone marrow. Considering a high proliferation rate and easy harvesting of MSCs of the ear, this finding could be of value for the regenerative medicine.

SUBMITTER: Khalilifar MA

PROVIDER: S-EPMC6397606 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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In Vitro and In Vivo Comparison of Different Types of Rabbit Mesenchymal Stem Cells for Cartilage Repair.

Khalilifar Mohammad Ali MA Baghaban Eslaminejad Mohamad Reza MR Ghasemzadeh Mohammad M Hosseini Samaneh S Baharvand Hossein H

Cell journal 20190220 2

<h4>Objective</h4>Systematic studies indicate a growing number of clinical studies that use mesenchymal stem cells (MSCs) for the treatment of cartilage lesions. The current experimental and preclinical study aims to comparatively evaluate the potential of MSCs from a variety of tissues for the treatment of cartilage defect in rabbit's knee which has not previously been reported.<h4>Materials and methods</h4>In this experimental study, MSCs isolated from bone marrow (BMMSCs), adipose (AMSCs), an ...[more]

PMID: 30825288

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Project description:This study compares the collagen types present in rabbit ear cartilage with those synthesized by dissociated chondrocytes in cell culture. The cartilage was first extracted with 4M-guanidinium chloride to remove proteoglycans. This step also extracted type I collagen. After pepsin solubilization of the residue, three additional, genetically distinct collagen types could be separated by fractional salt precipitation. On SDS (sodium dodecyl sulphate)/polyacrylamide-gel electrophoresis they were identified as type II collagen, (1 alpha, 2 alpha, 3 alpha) collagen and M-collagen fragments, a collagen pattern identical with that found in hyaline cartilage. Types I, II, (1 alpha, 2 alpha, 3 alpha) and M-collagen fragments represent 20, 75, 3.5, and 1% respectively of the total collagen. In frozen sections of ear cartilage, type II collagen was located by immunofluorescence staining in the extracellular matrix, whereas type I collagen was closely associated with the chondrocytes. Within 24h after release from elastic cartilage by enzymic digestion, auricular chondrocytes began to synthesize type III collagen, in addition to the above-mentioned collagens. This was shown after labelling of freshly dissociated chondrocytes with [3H]proline 1 day after plating, fractionation of the pepsin-treated collagens from medium and cell layer by NaCl precipitation, and analysis of the fractions by CM(carboxymethyl)-cellulose chromatography and SDS/polyacrylamide-gel electrophoresis. The 0.8 M-NaCl precipitate of cell-layer extracts consisted predominantly of type II collagen. The 0.8 M-NaCl precipitate obtained from the medium contained type I, II, and III collagen. In the supernatant of the 0.8 M-NaCl precipitation remained, both in the cell extract and medium, predominantly 1 alpha-, 2 alpha-, and 3 alpha-chains and M-collagen fragments. These results indicate that auricular chondrocytes are similar to chondrocytes from hyaline cartilage in that they produce, with the exception of type I collagen, the same collagen types in vivo, but change their cellular phenotype more rapidly after transfer to monolayer culture, as indicated by the prompt onset of type III collagen synthesis.

Project description:ObjectiveLubricin 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.MethodsFor 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.ResultsIn 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.ConclusionCartilage 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.

Dataset Information

In Vitro and In Vivo Comparison of Different Types of Rabbit Mesenchymal Stem Cells for Cartilage Repair.

Publications

In Vitro and In Vivo Comparison of Different Types of Rabbit Mesenchymal Stem Cells for Cartilage Repair.

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