Dataset Information

Synoviocyte-Derived Extracellular Matrix and bFGF Speed Human Chondrocyte Proliferation While Maintaining Differentiation Potential.

ABSTRACT: Improving the ability of human chondrocytes to proliferate, while maintaining their differentiation potential, has presented a great challenge in cartilage tissue engineering. In this study, human chondrocytes were cultured under four unique growth conditions at physiologic oxygen tension: tissue culture plastic (TCP) only, synoviocyte matrix (SCM)-coated flasks only, SCM-coated flasks with bFGF media supplement, and TCP with bFGF media supplement. The results indicated that, compared to standard TCP, all test conditions showed significantly increased cell expansion rates and an increase in both glycosaminoglycan (GAG) and collagen content during redifferentiation culture. Specifically, the combined SCM + bFGF growth condition showed an additive effect, with an increase of approximately 36% more cells per passage (5-7 days) when compared to the SCM alone. In conclusion, the results of this study demonstrate that bFGF and SCM can be used as supplements to enhance the growth of human chondrocytes both as individual enhancers and as a combined additive.

SUBMITTER: Truong RD

PROVIDER: S-EPMC9171110 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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Synoviocyte-Derived Extracellular Matrix and bFGF Speed Human Chondrocyte Proliferation While Maintaining Differentiation Potential.

Truong Rachel D RD Bernier Megan A MA Dennis James E JE Kean Thomas J TJ

Frontiers in bioengineering and biotechnology 20220524

Improving the ability of human chondrocytes to proliferate, while maintaining their differentiation potential, has presented a great challenge in cartilage tissue engineering. In this study, human chondrocytes were cultured under four unique growth conditions at physiologic oxygen tension: tissue culture plastic (TCP) only, synoviocyte matrix (SCM)-coated flasks only, SCM-coated flasks with bFGF media supplement, and TCP with bFGF media supplement. The results indicated that, compared to standar ...[more]

PMID: 35685088

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Project description:BackgroundCurrent tissue engineering methods are insufficient for total joint resurfacing, and chondrocytes undergo de-differentiation when expanded on tissue culture plastic. De-differentiated chondrocytes show poor re-differentiation in culture, giving reduced glycosaminoglycan (GAG) and collagen matrix accumulation. To address this, porcine synoviocyte-derived extracellular matrix and low (5%) oxygen tension were assessed for their ability to enhance human articular chondrocyte expansion and maintain re-differentiation potential.MethodsPorcine synoviocyte matrices were devitalized using 3 non-detergent methods. These devitalized synoviocyte matrices were compared against tissue culture plastic for their ability to support human chondrocyte expansion. Expansion was further compared at both low (5%), and atmospheric (20%) oxygen tension on all surfaces. Expanded cells then underwent chondrogenic re-differentiation in aggregate culture at both low and atmospheric oxygen tension. Aggregates were assessed for their GAG and collagen content both biochemically and histologically.ResultsHuman chondrocytes expanded twice as fast on devitalized synoviocyte matrix vs. tissue culture plastic, and cells retained their re-differentiation capacity for twice the number of population doublings. There was no significant difference in growth rate between low and atmospheric oxygen tension. There was significantly less collagen type I, collagen type II, aggrecan and more MMP13 expression in cells expanded on synoviocyte matrix vs. tissue culture plastic. There were also significant effects due to oxygen tension on gene expression, wherein there was greater collagen type I, collagen type II, SOX9 and less MMP13 expression on tissue culture plastic compared to synoviocyte matrix. There was a significant increase in GAG, but not collagen, accumulation in chondrocyte aggregates re-differentiated at low oxygen tension over that achieved in atmospheric oxygen conditions.ConclusionsSynoviocyte-derived matrix supports enhanced expansion of human chondrocytes such that the chondrocytes are maintained in a state from which they can re-differentiate into a cartilage phenotype after significantly more population doublings. Also, low oxygen tension supports GAG, but not collagen, accumulation. These findings are a step towards the production of a more functional, tissue engineered cartilage.

Project description:BackgroundThe non-canonical NF-?B activating kinase IKK?, encoded by CHUK (conserved-helix-loop-helix-ubiquitous-kinase), has been reported to modulate pro- or anti- inflammatory responses, cellular survival and cellular differentiation. Here, we have investigated the mechanism of action of IKK? as a novel effector of human and murine chondrocyte extracellular matrix (ECM) homeostasis and differentiation towards hypertrophy.Methodology/principal findingsIKK? expression was ablated in primary human osteoarthritic (OA) chondrocytes and in immature murine articular chondrocytes (iMACs) derived from IKK?(f/f):CreERT2 mice by retroviral-mediated stable shRNA transduction and Cre recombinase-dependent Lox P site recombination, respectively. MMP-10 was identified as a major target of IKK? in chondrocytes by mRNA profiling, quantitative RT-PCR analysis, immunohistochemistry and immunoblotting. ECM integrity, as assessed by type II collagen (COL2) deposition and the lack of MMP-dependent COL2 degradation products, was enhanced by IKK? ablation in mice. MMP-13 and total collagenase activities were significantly reduced, while TIMP-3 (tissue inhibitor of metalloproteinase-3) protein levels were enhanced in IKK?-deficient chondrocytes. IKK? deficiency suppressed chondrocyte differentiation, as shown by the quantitative inhibition of.Alizarin red staining and the reduced expression of multiple chondrocyte differentiation effectors, including Runx2, Col10a1 and Vegfa,. Importantly, the differentiation of IKK?-deficient chondrocytes was rescued by a kinase-dead IKK? protein mutant.Conclusions/significanceIKK? acts independent of its kinase activity to help drive chondrocyte differentiation towards a hypertrophic-like state. IKK? positively modulates ECM remodeling via multiple downstream targets (including MMP-10 and TIMP-3 at the mRNA and post-transcriptional levels, respectively) to maintain maximal MMP-13 activity, which is required for ECM remodeling leading to chondrocyte differentiation. Chondrocytes are the unique cell component in articular cartilage, which are quiescent and maintain ECM integrity during tissue homeostasis. In OA, chondrocytes reacquire the capacity to proliferate and differentiate and their activation results in pronounced cartilage degeneration. ????, our findings are also of potential relevance for defining the onset and/or progression of OA disease.

Dataset Information

Synoviocyte-Derived Extracellular Matrix and bFGF Speed Human Chondrocyte Proliferation While Maintaining Differentiation Potential.

Publications

Synoviocyte-Derived Extracellular Matrix and bFGF Speed Human Chondrocyte Proliferation While Maintaining Differentiation Potential.

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