scholarly journals Marine Collagen Hydrolysates Promote Collagen Synthesis, Viability and Proliferation While Downregulating the Synthesis of Pro-Catabolic Markers in Human Articular Chondrocytes

2021 ◽  
Vol 22 (7) ◽  
pp. 3693
Author(s):  
Bastien Bourdon ◽  
Frédéric Cassé ◽  
Nicolas Gruchy ◽  
Pierre Cambier ◽  
Sylvain Leclercq ◽  
...  
Keyword(s):  
Dietary Supplements ◽  
Type I Collagen ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Preventive Therapy ◽  
Human Articular Chondrocytes ◽  
Cartilage Tissue ◽  
Type I ◽  
Repair Capacity ◽  
Collagen Hydrolysates

Cartilage is a non-innervated and non-vascularized tissue. It is composed of one main cell type, the chondrocyte, which governs homeostasis within the cartilage tissue, but has low metabolic activity. Articular cartilage undergoes substantial stresses that lead to chondral defects, and inevitably osteoarthritis (OA) due to the low intrinsic repair capacity of cartilage. OA remains an incurable degenerative disease. In this context, several dietary supplements have shown promising results, notably in the relief of OA symptoms. In this study, we investigated the effects of collagen hydrolysates derived from fish skin (Promerim®30 and Promerim®60) and fish cartilage (Promerim®40) on the phenotype and metabolism of human articular chondrocytes (HACs). First, we demonstrated the safety of Promerim® hydrolysates on HACs cultured in monolayers. Then we showed that, Promerim® hydrolysates can increase the HAC viability and proliferation, while decreasing HAC SA-β-galactosidase activity. To evaluate the effect of Promerim® on a more relevant model of culture, HAC were cultured as organoids in the presence of Promerim® hydrolysates with or without IL-1β to mimic an OA environment. In such conditions, Promerim® hydrolysates led to a decrease in the transcript levels of some proteases that play a major role in the development of OA, such as Htra1 and metalloproteinase-1. Promerim® hydrolysates downregulated HtrA1 protein expression. In contrast, the treatment of cartilage organoids with Promerim® hydrolysates increased the neosynthesis of type I collagen (Promerim®30, 40 and 60) and type II collagen isoforms (Promerim®30 and 40), the latter being the major characteristic component of the cartilage extracellular matrix. Altogether, our results demonstrate that the use of Promerim® hydrolysates hold promise as complementary dietary supplements in combination with the current classical treatments or as a preventive therapy to delay the occurrence of OA in humans.

Effect of seeding density on stability of the differentiated phenotype of pig articular chondrocytes in culture

1988 ◽  
Vol 89 (3) ◽  
pp. 373-378
Author(s):  
F.M. Watt
Keyword(s):  
Type I Collagen ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
High Density ◽  
Seeding Density ◽  
Type I ◽  
Low Density ◽  
Keratan Sulphate ◽  
Chondrocyte Phenotype ◽  
Culture Cells

Articular chondrocytes are known to be phenotypically unstable in culture. One condition that has been reported to suppress dedifferentiation is cultivation at high density on tissue-culture plastic. The aim of the experiments described here was to study the effect of seeding density on chondrocyte proliferation and 35SO4 incorporation, and on the types of collagen and proteoglycan synthesized. I found that cells seeded at low or high density reached the same final density at confluence, and that 35SO4 incorporation, while initially higher (per cell) in high-density cultures, fell under both conditions, reaching the same low level after 3 weeks. The proportion of cells expressing keratan sulphate fell in low- but not high-density cultures and the decline was not prevented by inhibition of cell division. In all the cultures cells expressing keratan sulphate tended to have a rounded morphology. After 21 days in culture, chondrocytes grown at high density expressed predominantly large proteoglycans that aggregated with hyaluronic acid, whereas in low-density cultures a smaller, non-aggregating form was also present. By 21 days in culture cells at both high and low density were expressing type I collagen, although the high-density cells also had an extensive extracellular matrix of type II collagen. These observations support the conclusion that high seeding density stabilizes the chondrocyte phenotype to a greater extent than low seeding density. They also suggest that enhanced dedifferentiation at low density may be due to cell spreading, rather than to selective proliferation of a phenotypically unstable subpopulation of cells.

Expression of cartilage-specific molecules is retained on long-term culture of human articular chondrocytes

1995 ◽  
Vol 108 (5) ◽  
pp. 1991-1999 ◽  
Author(s):  
E. Kolettas ◽  
L. Buluwela ◽  
M.T. Bayliss ◽  
H.I. Muir
Keyword(s):  
Type I Collagen ◽  
Articular Chondrocytes ◽  
Culture Conditions ◽  
Human Articular Chondrocytes ◽  
Type I ◽  
Human Articular Cartilage ◽  
Long Term Culture ◽  
Chondrocyte Phenotype ◽  
Link Protein

Normal human adult articular chondrocytes were used to determine how the chondrocyte phenotype is modulated by culture conditions following long-term culture. We report here for the first time that human articular chondrocytes have a lifespan in the range of 34–37 population doublings. While chondrocytes cultured as monolayers displayed a fibroblastoid morphology and grew faster, those cultured as suspensions over agarose adopted a round morphology and formed clusters of cells reminiscent of chondrocyte differentiation in intact cartilage, with little or no DNA synthesis. These morphologies were independent of the age of the culture. Despite, these morphological differences, however, chondrocytes expressed markers at mRNA and protein levels characteristic of cartilage: namely, types II and IX collagens and the large aggregating proteoglycans, aggrecan, versican and link protein, but not syndecan, under both culture conditions. However, they also expressed type I collagen alpha 1(I) and alpha 2(I) chains. It has been suggested that expression of collagen alpha 1(I) by chondrocytes cultured as monolayers is a marker of the loss of the chondrocyte phenotype. However, we show here, using reverse transcriptase/polymerase chain reaction, that normal fresh intact human articular cartilage expresses collagen alpha 1(I). The data show that following long-term culture human articular chondrocytes retain their differentiated characteristics and that cell shape does not correlate with the expression of the chondrocyte phenotype. It is proposed that loss of the chondrocyte phenotype is marked by the loss of one or more cartilage-specific molecules rather than by the appearance of non-cartilage-specific molecules.

Evaluation of a Collagen-Hyaluronate Bilayer Matrix for Bone and Cartilage Repair

2000 ◽  
Vol 662 ◽  
Author(s):  
Lin-Shu Liu ◽  
Andrea Thompson ◽  
Robin Daverman ◽  
James W. Poser ◽  
Robert C. Spiro
Keyword(s):  
Growth Factor ◽  
Type I Collagen ◽  
Human Growth ◽  
Type Ii Collagen ◽  
Cartilage Tissue ◽  
Type I ◽  
Collagen Layer ◽  
And Cartilage

AbstractWe have developed a novel bilayer matrix composed of a porous type I collagen layer that transitions into a hyaluronate gel layer. This study evaluates the potential of the bilayer matrix to support the in vitro and in vivo formation of both bone and cartilage tissue. In the presence of recombinant human growth and differentiation factor-5, fetal rat calvarial cells cultured in the HA layer grew in a round, aggregated, chondrocyte-like morphology, while those in the collagen layer grew flattened and spread. Biochemical analysis demonstrated that cells in the collagen layer expressed higher levels of alkaline phosphatase activity, and lower levels of sulfated glycosaminoglycans and type II collagen when compared to cells in the HA layer. Intramuscular implants of the bilayer matrix with growth factor retrieved at 28 days revealed the presence of bone and cartilage tissue in the collagen and hyaluronate layers, respectively. These results demonstrate that the differentiation of cells in response to a single growth factor can be guided by specific compositional changes of the extracellular matrix.

scholarly journals Efficacy of collagen and alginate hydrogels for the prevention of rat chondrocyte dedifferentiation

2018 ◽  
Vol 9 ◽  
pp. 204173141880243 ◽  
Author(s):  
Guang-Zhen Jin ◽  
Hae-Won Kim
Keyword(s):  
Tissue Engineering ◽  
Type I Collagen ◽  
Cartilage Tissue Engineering ◽  
Collagen Gel ◽  
Type Ii Collagen ◽  
Cartilage Tissue ◽  
Type I ◽  
Type Ii ◽  
Chondrogenic Phenotype

Dedifferentiation of chondrocytes remains a major problem in cartilage tissue engineering. The development of hydrogels that can preserve chondrogenic phenotype and prevent chondrocyte dedifferentiation is a meaningful strategy to solve dedifferentiation problem of chondrocytes. In the present study, three gels were prepared (alginate gel (Alg gel), type I collagen gel (Col gel), and their combination gel (Alg/Col gel)), and the in vitro efficacy of chondrocytes culture while preserving their phenotypes was investigated. While Col gel became substantially contracted with time, the cells encapsulated in Alg gel preserved the shape over the culture period of 14 days. The mechanical and cell-associated contraction behaviors of Alg/Col gel were similar to those of Alg. The cells in Alg and Alg/Col gels exhibited round morphology, whereas those in Col gel became elongated (i.e. fibroblast-like) during cultures. The cells proliferated with time in all gels with the highest proliferation being attained in Col gel. The expression of chondrogenic genes, including SOX9, type II collagen, and aggrecan, was significantly up-regulated in Alg/Col gel and Col gel, particularly in Col gel. However, the chondrocyte dedifferentiation markers, type I collagen and alkaline phosphatase ( ALP), were also expressed at significant levels in Col gel, which being contrasted with the events in Alg and Alg/Col gels. The current results suggest the cells cultured in hydrogels can express chondrocyte dedifferentiation markers as well as chondrocyte markers, which draws attention to choose proper hydrogels for chondrocyte-based cartilage tissue engineering.

scholarly journals The lncRNA, Nespas, Is Associated with Osteoarthritis Progression and Serves as a Potential New Prognostic Biomarker

Cartilage ◽  
2017 ◽  
Vol 10 (2) ◽  
pp. 148-156 ◽  
Author(s):  
Sujung Park ◽  
Myeungsoo Lee ◽  
Churl-Hong Chun ◽  
Eun-Jung Jin
Keyword(s):  
Noncoding Rna ◽  
Type I Collagen ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Human Chondrocytes ◽  
Type I ◽  
Caspase 1 ◽  
Osteoarthritis Progression ◽  
Polymerase Chain ◽  
Abnormal Lipid Metabolism

Introduction. In this article, we explored the hypothesis that the long noncoding RNA, Nespas, promotes osteoarthritis (OA) by supporting abnormal lipid metabolism in human chondrocytes. Materials and Methods. Human articular chondrocytes from osteoarthritis patients were used and expression level of Nespas were determined by real-time polymerase chain reaction. Introduction of Nespas and Nespas-associated genes/miRNAs were performed by using a lentiviral system. The effect of Nespas on mitochondrial function was determined by staining mitochondria and analyzing mitopotential and mitochondrial genes. Moreover, to identify the responsible molecules in Nespas-induced pathogenesis, profiling of peroxisomal genes and miRNAs were applied and interactome analysis was performed. Results. Highly elevated levels of Nespas and Acyl-CoA synthetase 6 (ACSL6) were observed in OA patients. Both Nespas overexpression and ACSL6 upregulation into human chondrocytes induced typical OA characteristics, such as downregulation of type II collagen; upregulation of type I collagen, metalloproteinase 13, and caspase-1 and -3; and dysfunction of mitochondria and peroxisome. Co-expression of Nespas and ACSL6 siRNA reduced caspase-1 and -3 levels. Moreover, Nespas overexpression significantly suppressed levels of miR-291a-3p, -196a-5p, -23a-3p, -24-3p, and let-7a-5p, and these miRs are known to potentially target ACSL6 according to ingenuity pathway analysis. We also confirmed that these miRs were significantly suppressed in human OA chondrocytes. Overexpression of miR-291a-3p, -196a-5p, -23a-3p, -24-3p, or let-7a-5p in the presence of Nespas suppressed levels of ACSL6, caspase-1 and -3. Discussion. Overall, we suggest that elevated Nespas level in OA are associated with OA pathogenesis by suppressing miRs targeting ACSL6 and subsequent ACSL6 upregulation.

scholarly journals PTH decreases in vitro human cartilage regeneration without affecting hypertrophic differentiation

2019 ◽  
Author(s):  
Marijn Rutgers ◽  
Frances Bach ◽  
Luciënne Vonk ◽  
Mattie van Rijen ◽  
Vanessa Akrum ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Parathyroid Hormone ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Human Articular Chondrocytes ◽  
Type I ◽  
Human Articular Cartilage ◽  
Type Ii ◽  
Collagen Deposition ◽  
Healthy Human

AbstractRegenerated cartilage formed after Autologous Chondrocyte Implantation may be of suboptimal quality due to postulated hypertrophic changes. Parathyroid hormone-related peptide, containing the parathyroid hormone sequence (PTHrP 1-34), enhances cartilage growth during development and inhibits hypertrophic differentiation of mesenchymal stromal cells (MSCs) and growth plate chondrocytes. This study aims to determine whether human articular chondrocytes respond correspondingly. Healthy human articular cartilage-derived chondrocytes (n=6 donors) were cultured on type II collagen-coated transwells with/without 0.1 or 1.0 μM PTH from day 0, 9, or 21 until the end of culture (day 28). Extracellular matrix production, (pre)hypertrophy and PTH signaling were assessed by RT-qPCR and/or immunohistochemistry for collagen type I, II, X, RUNX2, MMP13, PTHR1 and IHH and by determining glycosaminoglycan production and DNA content. The Bern score assessed cartilage quality by histology. Regardless of the concentration and initiation of supplementation, PTH treatment significantly decreased DNA and glycosaminoglycan content and reduced the Bern score compared with controls. Type I collagen deposition was increased, whereas PTHR1 expression and type II collagen deposition were decreased by PTH supplementation. Expression of the (pre)hypertrophic markers MMP13, RUNX2, IHH and type X collagen were not affected by PTH. In conclusion, PTH supplementation to healthy human articular chondrocytes did not affect hypertrophic differentiation, but negatively influenced cartilage quality, the tissues’ extracellular matrix and cell content. Although PTH may be an effective inhibitor of hypertrophic differentiation in MSC-based cartilage repair, care may be warranted in applying accessory PTH treatment due to its effects on articular chondrocytes.

scholarly journals Direct Perfusion Improves Redifferentiation of Human Chondrocytes in Fibrin Hydrogel with the Deposition of Cartilage Pericellular Matrix

2021 ◽  
Vol 11 (19) ◽  
pp. 8923
Author(s):  
Alexandre Dufour ◽  
Frédéric Mallein-Gerin ◽  
Emeline Perrier-Groult
Keyword(s):  
Type I Collagen ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Type I ◽  
Pericellular Matrix ◽  
Fibrin Gels ◽  
Matrix Quality ◽  
The Matrix ◽  
Cartilage Injuries ◽  
Main Component

Articular cartilage has limited potential for self-repair, and cell-based strategies combining scaffolds and chondrocytes are currently used to treat cartilage injuries. However, achieving a satisfying level of cell redifferentiation following expansion remains challenging. Hydrogels and perfusion bioreactors are known to exert beneficial cues on chondrocytes; however, the effect of a combined approach on the quality of cartilage matrix deposited by cells is not fully understood. Here, we combined soluble factors (BMP-2, Insulin, and Triiodothyronine, that is, BIT), fibrin hydrogel, direct perfusion and human articular chondrocytes (HACs) to engineer large cartilage tissues. Following cell expansion, cells were embedded in fibrin gels and cultivated under either static or perfusion conditions. The nature of the matrix synthesized was assessed by Western blotting and immunohistochemistry. The stability of cartilage grafts and integration with native tissue were also investigated by subcutaneous implantation of human osteochondral cylinders in nude mice. Perfusion preconditioning improved matrix quality and spatial distribution. Specifically, perfusion preconditioning resulted in a matrix rich in type II collagen but not in type I collagen, indicating the reconstruction of hyaline cartilage. Remarkably, the production of type VI collagen, the main component of the pericellular matrix, was also increased, indicating that chondrocytes were connecting to the hyaline matrix they produced.

scholarly journals Synthesis of cartilage matrix by mammalian chondrocytes in vitro. II. Maintenance of collagen and proteoglycan phenotype.

1982 ◽  
Vol 93 (3) ◽  
pp. 751-757 ◽  
Author(s):  
K E Kuettner ◽  
V A Memoli ◽  
B U Pauli ◽  
N C Wrobel ◽  
E J Thonar ◽  
...  
Keyword(s):  
Type I Collagen ◽  
Articular Chondrocytes ◽  
Type Ii Collagen ◽  
Type I ◽  
Type Ii ◽  
Roller Bottle ◽  
The Media ◽  
Bovine Articular Chondrocytes ◽  
Associated Matrix

The in vitro phenotype of bovine articular chondrocytes is described. Chondrocytes plated at high density in roller-bottle and dish cultures were maintained in vitro. The major matrix macromolecules, collagen and proteoglycan, synthesized by these cells were characterized during the course of the culture period. The chondrocytes synthesized mainly Type II collagen, which was found predominantly in the cell-associated matrix. The media contained a mixture of Type II and Type III collagens. Type I collagen was detectable in neither the medium nor the cell-associated matrix. The proteoglycan monomers found in media and cell-associated matrix had the same hydrodynamic sizes as monomers synthesized by cartilage slices or those extracted from adult articular cartilage. The majority of proteoglycans synthesized by the cells were found in high molecular weight aggregates which were readily recovered from the media and were extractable from cell-associated matrix with low ionic strength buffers. The results demonstrate the long-term in vitro phenotypic stability of the bovine articular chondrocytes. The advantages of the in vitro system as a model for studying the effects of external agents, such as drugs and vitamins, are discussed.

scholarly journals Marine Collagen Hydrolysates Downregulate the Synthesis of Pro-Catabolic and Pro-Inflammatory Markers of Osteoarthritis and Favor Collagen Production and Metabolic Activity in Equine Articular Chondrocyte Organoids

2021 ◽  
Vol 22 (2) ◽  
pp. 580
Author(s):  
Bastien Bourdon ◽  
Romain Contentin ◽  
Frédéric Cassé ◽  
Chloé Maspimby ◽  
Sarah Oddoux ◽  
...  
Keyword(s):  
Dietary Supplements ◽  
Metabolic Activity ◽  
Articular Chondrocytes ◽  
Articular Chondrocyte ◽  
Repair Capacity ◽  
Mechanical Constraints ◽  
Chondral Defects ◽  
Inflammatory Conditions ◽  
Collagen Hydrolysates

Articular cartilage experiences mechanical constraints leading to chondral defects that inevitably evolve into osteoarthritis (OA), because cartilage has poor intrinsic repair capacity. Although OA is an incurable degenerative disease, several dietary supplements may help improve OA outcomes. In this study, we investigated the effects of Dielen® hydrolyzed fish collagens from skin (Promerim®30 and Promerim®60) and cartilage (Promerim®40) to analyze the phenotype and metabolism of equine articular chondrocytes (eACs) cultured as organoids. Here, our findings demonstrated the absence of cytotoxicity and the beneficial effect of Promerim® hydrolysates on eAC metabolic activity under physioxia; further, Promerim®30 also delayed eAC senescence. To assess the effect of Promerim® in a cartilage-like tissue, eACs were cultured as organoids under hypoxia with or without BMP-2 and/or IL-1β. In some instances, alone or in the presence of IL-1β, Promerim®30 and Promerim®40 increased protein synthesis of collagen types I and II, while decreasing transcript levels of proteases involved in OA pathogenesis, namely Htra1, and the metalloproteinases Mmp1-3, Adamts5, and Cox2. Both Promerim® hydrolysates also decreased Htra1 protein amounts, particularly in inflammatory conditions. The effect of Promerim® was enhanced under inflammatory conditions, possibly due to a decrease in the synthesis of inflammation-associated molecules. Finally, Promerim® favored in vitro repair in a scratch wound assay through an increase in cell proliferation or migration. Altogether, these data show that Promerim®30 and 40 hold promise as dietary supplements to relieve OA symptoms in patients and to delay OA progression.

scholarly journals Transcriptomics Study to Determine the Molecular Mechanism by which sIL-13Rα2-Fc Inhibits Caudal Intervertebral Disc Degeneration in Rats

2020 ◽  
Vol 2020 ◽  
pp. 1-13
Author(s):  
Xin Wang ◽  
Jianshi Tan ◽  
Junhao Sun ◽  
Pengzhong Fang ◽  
Jinlei Chen ◽  
...  
Keyword(s):  
Intervertebral Disc ◽  
Disc Degeneration ◽  
Intervertebral Disc Degeneration ◽  
Type I Collagen ◽  
Type Ii Collagen ◽  
Type I ◽  
Model Group ◽  
Expression Levels ◽  
Protein Levels ◽  
Collagen Protein

Background. Intervertebral disc degeneration is related to tissue fibrosis. ADAMTS can degrade the important components of the ECM during the process of intervertebral disc degeneration, ultimately resulting in the loss of intervertebral disc function. sIL-13Rα2-Fc can inhibit fibrosis and slow down the degeneration process, but the mechanism involved remains unclear. Objective. To determine the mechanism by which sIL-13Rα2-Fc inhibits ECM degradation and reduces intervertebral disc tissue fibrosis using a transcriptomics analysis. Methods. A rat model of caudal intervertebral disc degeneration was established, and Sirius red staining was used to observe the pathological changes in the caudal intervertebral disc. Transcriptome sequencing was employed to assess the gene expression profiles of the intervertebral disc tissues in the model group and the sIL-13Rα2-Fc-treated group. Differentially expressed genes were identified and analyzed using GO annotation and KEGG pathway analyses. Real-time fluorescence quantitative PCR was used to verify the expression levels of candidate genes. The levels of GAG and HA were quantitatively assessed by ELISA, and the levels of collagen I and collagen II were analyzed by western blotting. Results. Sirius red staining showed that in the model group, the annulus fibrosus was disordered, the number of breaks increased, and the type I collagen protein levels increased, whereas in the sIL-13Rα2-Fc group, the annulus fibrosus was ordered, the number of breaks decreased, and the type II collagen protein levels increased. In comparison with the model group, we identified 58 differentially expressed genes in the sIL-13Rα2-Fc group, and these were involved in 35 signaling pathways. Compared with those in the model group, the mRNA expression levels of Rnux1, Sod2, and Tnfaip6 in the IL-13Rα2-Fc group were upregulated, and the mRNA expression levels of Aldh3a1, Galnt3, Fgf1, Celsr1, and Adamts8 were downregulated; these results were verified by real-time fluorescence quantitative PCR. TIMP-1 (an ADAMTS inhibitor) and TIMP-1 combined with the sIL-13Rα2-Fc intervention increased the levels of GAG and HA, inhibited the expression of type I collagen, and promoted the expression of type II collagen. Conclusion. Adamts8 may participate in the degradation of ECM components such as GAG and HA and lead to an imbalance in the ECM of the intervertebral disc, resulting in intervertebral disc degeneration. sIL-13Rα2-Fc promoted anabolism of the ECM and increased the levels of ECM components by inhibiting the expression of Adamts8, thus maintaining the dynamic equilibrium of the ECM and ultimately delaying intervertebral disc degeneration.

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