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.

scholarly journals Maintenance of chondrocyte phenotype during expansion on PLLA microtopographies

2018 ◽  
Vol 9 ◽  
pp. 204173141878982 ◽  
Author(s):  
Elisa Costa ◽  
Cristina González-García ◽  
José Luis Gómez Ribelles ◽  
Manuel Salmerón-Sánchez
Keyword(s):  
Lactic Acid ◽  
Type I Collagen ◽  
Collagen Type ◽  
Articular Chondrocytes ◽  
Collagen Type I ◽  
Cell Phenotype ◽  
Type I ◽  
Type Ii ◽  
Collagen Type Ii ◽  
Chondrocyte Phenotype

Articular chondrocytes are difficult to grow, as they lose their characteristic phenotype following expansion on standard tissue culture plates. Here, we show that culturing them on surfaces of poly(L-lactic acid) of well-defined microtopography allows expansion and maintenance of characteristic chondrogenic markers. We investigated the dynamics of human chondrocyte dedifferentiation on the different poly(L-lactic acid) microtopographies by the expression of collagen type I, collagen type II and aggrecan at different culture times. When seeded on poly(L-lactic acid), chondrocytes maintained their characteristic hyaline phenotype up to 7 days, which allowed to expand the initial cell population approximately six times without cell dedifferentiation. Maintenance of cell phenotype was afterwards correlated to cell adhesion on the different substrates. Chondrocytes adhesion occurs via the α5 β1 integrin on poly(L-lactic acid), suggesting cell–fibronectin interactions. However, α2 β1 integrin is mainly expressed on the control substrate after 1 day of culture, and the characteristic chondrocytic markers are lost (collagen type II expression is overcome by the synthesis of collagen type I). Expanding chondrocytes on poly(L-lactic acid) might be an effective solution to prevent dedifferentiation and improving the number of cells needed for autologous chondrocyte transplantation.

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.

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.

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.

Collagen Expression in Tissue Engineered Cartilage of Aged Human Articular Chondrocytes in a Rotating Bioreactor

2003 ◽  
Vol 26 (4) ◽  
pp. 319-330 ◽  
Author(s):  
S. Marlovits ◽  
B. Tichy ◽  
M. Truppe ◽  
D. Gruber ◽  
W. Schlegel
Keyword(s):  
Electron Microscopy ◽  
Collagen Type ◽  
Articular Chondrocytes ◽  
Three Dimensional ◽  
Collagen Type I ◽  
Human Articular Chondrocytes ◽  
Type I ◽  
Transmission Electron ◽  
Low Shear Stress ◽  
Engineered Cartilage

This study describes the culture and three-dimensional assembly of aged human articular chondrocytes under controlled oxygenation and low shear stress in a rotating-wall vessel. Chondrocytes cultured in monolayer were released and placed without any scaffold as a single cell suspension in a rotating bioreactor for 12 weeks. Samples were analyzed with immunohistochemistry, molecular biology and electron microscopy. During serial monolayer cultures chondrocytes dedifferentiated to a “fibroblast-like” structure and produced predominantly collagen type I. When these dedifferentiated cells were transferred to the rotating bioreactor, the cells showed a spontaneous aggregation and formation of solid tissue during the culture time. Expression of collagen type II and other components critical for the extracellular cartilage matrix could be detected. Transmission electron microscopy revealed a fine network of randomly distributed collagen fibrils. This rotating bioreactor proves to be a useful tool for providing an environment that enables dedifferentiated chondrocytes to redifferentiate and produce a cartilage-specific extracellular matrix.

scholarly journals Effects of denosumab in Japanese rheumatoid arthritis patients treated with conventional anti-rheumatic drugs: 36-month extension of a phase 3 study

2021 ◽  
pp. jrheum.201376
Author(s):  
Yoshiya Tanaka ◽  
Tsutomu Takeuchi ◽  
Satoshi Soen ◽  
Hisashi Yamanaka ◽  
Toshiyuki Yoneda ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Type I Collagen ◽  
Joint Destruction ◽  
Term Treatment ◽  
Drug Discontinuation ◽  
Type I ◽  
Mineral Density ◽  
Phase 3 ◽  
Long Term Treatment

Objective To evaluate safety and efficacy of long-term denosumab 60 mg every 6 (Q6M) or 3 months (Q3M) in rheumatoid arthritis (RA) patients. Methods This 12-month, randomised, double-blind, placebo-controlled, multicentre phase 3 trial with an open-label extension period from 12 to 36 months (DESIRABLE) enrolled Japanese RA patients treated with placebo for 12 months then denosumab Q6M (P/Q6M) or denosumab Q3M (P/Q3M); denosumab Q6M for 36 months (Q6M/Q6M); or denosumab Q3M for 36 months (Q3M/Q3M). Efficacy was assessed by van der Heijde modified total Sharp (mTSS), bone erosion (ES), and joint space narrowing (JSN) scores. Results Long-term treatment better maintained mTSS and ES suppression in the P/Q3M and Q3M/Q3M versus P/Q6M and Q6M/Q6M groups; changes from baseline in total mTSS at 36 months were 2.8 (standard error 0.4), 1.7 (0.3), 3.0 (0.4), and 2.4 (0.3), respectively; corresponding changes in ES were 1.3 (0.2), 0.4 (0.2), 1.4 (0.2), and 1.1 (0.2). No JSN effect was observed. Bone mineral density consistently increased in all groups after denosumab initiation, regardless of concomitant glucocorticoid administration. Serum C-telopeptide of type I collagen decreased rapidly at 1-month post-denosumab administration (both in the initial 12- month [Q3M, Q6M groups] and long-term treatment [P/Q3M, P/Q6M groups] phases). Adverse event incidence leading to study drug discontinuation was similar across treatment groups. Conclusion Denosumab treatment maintained inhibition of progression of joint destruction up to 36 months. Based on effects on ES progression, higher dosing frequency at an earlier treatment stage may be needed to optimise treatment. Denosumab was generally well tolerated.

Sign in / Sign up

Export Citation Format

Share Document