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Enhancement of cartilage repair through the addition of growth plate chondrocytes in an immature skeleton animal model.

ABSTRACT: BACKGROUND:The treatment of articular cartilage damage is a major clinical problem. More often, this clinical issue affects children, which forces doctors to find the best treatment method. METHODS:The aim of this experimental study on 2-month-old Landrace pigs was to compare the results of two cartilage defect treatments: (1) filling the cartilage defect with a scaffold incubated with bone marrow aspirate supplemented with growth plate chondrocytes (the CELLS group) and (2) filling the cartilage defect with an empty scaffold implanted after drilling the subchondral bone (the CTRL group). The treatment outcomes were assessed macroscopically and microscopically. RESULTS:Based on the macroscopic evaluation, all animals showed a nearly normal morphology, with an average of 9.66/12 points (CTRL) and 10.44/12 points (CELLS). Based on the microscopic evaluation, 1 very good result and 8 good results were obtained in the CTRL group, with an average of 70.44%, while 5 very good results and 4 good results were obtained in the CELLS group, with an average of 79.61%. CONCLUSIONS:(1) Growth plate chondrocytes have high chondrogenic potential and thus offer new possibilities for cartilage cell therapy. (2) The implantation of a scaffold loaded with bone marrow-derived MSCs (mesenchymal stem cells) and growth plate chondrocytes into a cartilage defect is a good therapeutic method in immature patients. (3) Cartilage repair based on a scaffold with bone marrow aspirate-derived cells supplemented with autologous growth plate chondrocytes achieves better results than repair with marrow stimulation and a hyaluronic acid-based scaffold (overall microscopic rating). (4) Chondrocyte clustering is a manifestation of the cartilage repair process but requires further observation.

SUBMITTER: Tomaszewski R

PROVIDER: S-EPMC6694631 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Enhancement of cartilage repair through the addition of growth plate chondrocytes in an immature skeleton animal model.

Tomaszewski Ryszard R Wiktor Łukasz Ł Gap Artur A

Journal of orthopaedic surgery and research 20190815 1

<h4>Background</h4>The treatment of articular cartilage damage is a major clinical problem. More often, this clinical issue affects children, which forces doctors to find the best treatment method.<h4>Methods</h4>The aim of this experimental study on 2-month-old Landrace pigs was to compare the results of two cartilage defect treatments: (1) filling the cartilage defect with a scaffold incubated with bone marrow aspirate supplemented with growth plate chondrocytes (the CELLS group) and (2) filli ...[more]

PMID: 31416470

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Project description:Prostaglandin E2 (PGE2) plays an important role in bone development and metabolism. To interfere therapeutically in the PGE2 pathway, however, knowledge about the involved enzymes (cyclooxygenases) and receptors (PGE2 receptors) is essential. We therefore examined the production of PGE2 in cultured growth plate chondrocytes in vitro and the effects of exogenously added PGE2 on cell proliferation. Furthermore, we analysed the expression and spatial distribution of cyclooxygenase (COX)-1 and COX-2 and PGE2 receptor types EP1, EP2, EP3 and EP4 in the growth plate in situ and in vitro. PGE2 synthesis was determined by mass spectrometry, cell proliferation by DNA [3H]-thymidine incorporation, mRNA expression of cyclooxygenases and EP receptors by RT-PCR on cultured cells and in homogenized growth plates. To determine cellular expression, frozen sections of rat tibial growth plate and primary chondrocyte cultures were stained using immunohistochemistry with polyclonal antibodies directed towards COX-1, COX-2, EP1, EP2, EP3, and EP4. Cultured growth plate chondrocytes transiently secreted PGE2 into the culture medium. Although both enzymes were expressed in chondrocytes in vitro and in vivo, it appears that mainly COX-2 contributed to PGE2-dependent proliferation. Exogenously added PGE2 stimulated DNA synthesis in a dose-dependent fashion and gave a bell-shaped curve with a maximum at 10-8 M. The EP1/EP3 specific agonist sulprostone and the EP1-selective agonist ONO-D1-004 increased DNA synthesis. The effect of PGE2 was suppressed by ONO-8711. The expression of EP1, EP2, EP3, and EP4 receptors in situ and in vitro was observed; EP2 was homogenously expressed in all zones of the growth plate in situ, whereas EP1 expression was inhomogenous, with spared cells in the reserve zone. In cultured cells these four receptors were expressed in a subset of cells only. The most intense staining for the EP1 receptor was found in polygonal cells surrounded by matrix. Expression of receptor protein for EP3 and EP4 was observed also in rat growth plates. In cultured chrondrocytes, however, only weak expression of EP3 and EP4 receptor was detected. We suggest that in growth plate chondrocytes, COX-2 is responsible for PGE2 release, which stimulates cell proliferation via the EP1 receptor.

Project description:Articular cartilage is not a physiologically self-renewing tissue. Injury of cartilage often progresses from the articular surface to the subchondral bone, leading to pathogenesis of tissue degenerative diseases, such as osteoarthritis. Therapies to treat cartilage defects using autologous chondrocyte-based tissue engineering have been developed and used for more than 20 years; however, the challenge of chondrocyte expansion in vitro remains. A promising cell source, cartilage stem/progenitor cells (CSPCs), has attracted recent attention. Because their origin and identity are still unclear, the application potential of CSPCs is under active investigation. Here we have captured the emergence of a group of stem/progenitor cells derived from adult human chondrocytes, highlighted by dynamic changes in expression of the mature chondrocyte marker, COL2, and mesenchymal stromal/stem cell (MSC) marker, CD146. These cells are termed chondrocyte-derived progenitor cells (CDPCs). The stem cell-like potency and differentiation status of CDPCs were determined by physical and biochemical cues during culture. A low-density, low-glucose 2-dimensional culture condition (2DLL) was critical for the emergence and proliferation enhancement of CDPCs. CDPCs showed similar phenotype as bone marrow mesenchymal stromal/stem cells but exhibited greater chondrogenic potential. Moreover, the 2DLL-cultured CDPCs proved efficient in cartilage formation both in vitro and in vivo and in repairing large knee cartilage defects (6-13 cm(2)) in 15 patients. These findings suggest a phenotype conversion between chondrocytes and CDPCs and provide conditions that promote the conversion. These insights expand our understanding of cartilage biology and may enhance the success of chondrocyte-based therapies.Injury of cartilage, a non-self-repairing tissue, often progresses to pathogenesis of degenerative joint diseases, such as osteoarthritis. Although tissue-derived stem cells have been shown to contribute to tissue renewal and homeostasis, the derivation, biological function, and application potential of stem/progenitor cells found in adult human articular cartilage are incompletely understood. This study reports the derivation of a population of cartilage stem/progenitor cells from fully differentiated chondrocytes under specific culture conditions, which have the potential to reassume their chondrocytic phenotype for efficient cartilage regeneration. These findings support the possibility of using in vitro amplified chondrocyte-derived progenitor cells for joint cartilage repair.

Project description:We used human fetal bone marrow-derived mesenchymal stromal cells (hfMSCs) differentiating towards chondrocytes as an alternative model for the human growth plate (GP). Our aims were to study gene expression patterns associated with chondrogenic differentiation to assess whether chondrocytes derived from hfMSCs are a suitable model for studying the development and maturation of the GP. hfMSCs efficiently formed hyaline cartilage in a pellet culture in the presence of TGFB3 and BMP6. Microarray and principal component analysis were applied to study gene expression profiles during chondrogenic differentiation. A set of 232 genes was found to correlate with in vitro cartilage formation. Several identified genes are known to be involved in cartilage formation and validate the robustness of the differentiating hfMSC model. KEGG pathway analysis using the 232 genes revealed 9 significant signaling pathways correlated with cartilage formation. To determine the progression of growth plate cartilage formation, we compared the gene expression profile of differentiating hfMSCs with previously established expression profiles of epiphyseal GP cartilage. As differentiation towards chondrocytes proceeds, hfMSCs gradually obtain a gene expression profile resembling epiphyseal GP cartilage. We visualized the differences in gene expression profiles as protein interaction clusters and identified many protein clusters that are activated during the early chondrogenic differentiation of hfMSCs showing the potential of this system to study GP development. To determine the progression of growth plate cartilage formation, we compared the gene expression profile of differentiating hfMSCs with previously established expression profiles of epiphyseal GP cartilage. As differentiation towards chondrocytes proceeds, hfMSCs gradually obtain a gene expression profile resembling epiphyseal GP cartilage. We visualized the differences in gene expression profiles as protein interaction clusters and identified many protein clusters that are activated during the early chondrogenic differentiation of hfMSCs showing the potential of this system to study GP development.

Dataset Information

Enhancement of cartilage repair through the addition of growth plate chondrocytes in an immature skeleton animal model.

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Enhancement of cartilage repair through the addition of growth plate chondrocytes in an immature skeleton animal model.

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