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BCRU is specialized in the selection (screening) of active anti-osteoarthritis compounds or drugs. Our laboratory has developed several culture models for studying mechanisms of action of these compounds on human chondrocytes, osteoblasts and synoviocytes. It is a partner of choice for pharmaceutical and food processing sectors active in osteo-articular pathologies.

 

Chondrocyte metabolism

 

Chondrocyte culture in monolayer

In this model, human or bovine articular chondrocytes (P0) are cultured in monolayer. This model allow the selection of potential anti-osteoarthritic compounds. Cellular proliferation, apoptosis and gene involved in cartilage remodeling can be assessed, in the presence or not of IL-1b. In vitro, IL-1 is a potent inhibitor of proteoglycans and collagen type II synthesis, and a stimulator of metalloproteases, IL-6, IL-8, PGE2, NO and reactive oxygen species (ROS) production. For these reasons, IL-1b is commonly used in culture models in order to mimic the circumstances leading to in vivo cartilage degradation

 

Alginate beads chondrocyte culture

Alginate is a linear polysaccharide isolated from brown algae. In the presence of divalent cations such as Ca++, this polymer gelled instantly and forms a porous matrix around chondrocytes. Under these conditions, chondrocytes maintain their phenotype for at least five weeks.

This model offers many other advantages:

- The matrix provides a newly synthesized compartmentalization identical to that found in the hyaline cartilage in vivo. Electron microscopy can distinguish clearly two compartments. The matrix surrounding the cell or "cell-associated matrix" (CM), which corresponds to the matrix pericellular and territorial matrix of cartilage in vivo, and secondly the matrix contained in alginate or "Further-removed matrix" ( FRM) which corresponds to the inter-territorial matrix of cartilage.

- The cells and the extracellular matrix newly synthesized can be easily recovered after degelation of the alginate by adding a calcium chelator (i.e. citrate). It is therefore possible to perform quantitative and qualitative analysis of the CM and FRM fractions.

- The volumes occupied by different compartments of the alginate beads correspond to those observed in normal cartilage.

- The half-life (t1 / 2) of aggrecan is longer than that recorded in other crop models, and approaches the value observed in vivo.

This model is relevant for the study of substances on the formation of extracellular matrix of cartilage.

Cartilage in explants culture

allows the study of homeostasis of the cartilage matrix. In this model, interactions between the chondrocyte and pericellular matrix are preserved. The extracellular matrix may be marked by the incorporation of radio-elements (35S ou 3H-proline). It is therefore possible to study the degradation of proteoglycans and collagen in basal conditions or in the presence of one or more cytokines that stimulate matrix proteolysis (eg IL-1).

This model is suitable for studying the degradation of the extracellular matrix.

 

Culture of chondrocytes under tensile stress.

During the movement, chondrocytes are subjected to tensile stress resulting from sliding of articular surfaces.

The system Flexercell allows to cultured chondrocytes in monolayer under tension. The force exerted on the chondrocyte and frequency of application of these forces can be modulated. This model allows the study of traction on the metabolism of chondrocytes.

 

 

Osteoblasts metabolism

Human subchondral osteoblasts culture

In this model, human OA subchondral osteoblasts coming from the sclerotic (SC) or the not sclerotic (NSC) area are cultured in monolayer [11].

 

Mechanical stress on osteoblasts

Recently, we developed a new three dimensional in vitro model of Ob loading, allowing the study of cyclic compression on osteoblasts embedded in a newly synthesized extracellular matrix. In this model, cell/matrix interactions are conserved and fluid flow through a three dimensional extracellular matrix is allowed. So, compression, tension and fluid shear stimuli occurred in our model. [12] .

 

Bone/cartilage crosstalk

Osteoblasts/chondrocytes co-culture

In this model, OA chondrocytes were cultured in alginate beads and osteoblasts from sclerotic or not sclerotic areas in monolayer. Both cell types are separated by a porous membrane allowing the passage of molecules of low molecular weight. This model is particularly well suited to the study of communication between chondrocytes and osteoblasts [13]

 

Synoviocytes metabolism

Synoviocytes cultures

We isolate and cultivate synoviocytes, from inflamed areas or not of OA synovial membrane. This model allows us to study the influence of molecules on inflammation of the synovial membrane, as well as angiogenesis.

 

Bibliography

[1] Bassleer CT, Henrotin YE, Reginster JL, Franchimont PP. Effects of tiaprofenic acid and acetylsalicylic acid on human articular chondrocytes in 3-dimensional culture. J Rheumatol 1992;19: 1433-8. [Pubmed]
[2] Bassleer CT, Franchimont PP, Henrotin YE, Franchimont NM, Geenen VG, Reginster JY. Effects of ipriflavone and its metabolites on human articular chondrocytes cultivated in clusters. Osteoarthritis Cartilage 1996;4: 1-8. [Pubmed]
[3] Bassleer C, Henrotin Y, Franchimont P. Effects of ximoprofen and acetylsalicylic acid on human articular chondrocytes in three-dimensional culture. Drug Invest 1993;5: 11-18.
[4] Henrotin Y, Labasse A, Zheng SX, Galais P, Tsouderos Y, Crielaard JM, Reginster JY. Strontium ranelate increases cartilage matrix formation. J Bone Miner Res 2001;16: 299-308. [Pubmed]
[5] Sanchez C, Mateus MM, Defresne MP, Crielaard JM, Reginster JY, Henrotin YE. Metabolism of human articular chondrocytes cultured in alginate beads. Longterm effects of interleukin 1beta and nonsteroidal antiinflammatory drugs. J Rheumatol 2002;29: 772-82. [Pubmed]
[6] Sanchez C, Mathy-Hartert M, Deberg MA, Ficheux H, Reginster JY, Henrotin YE. Effects of rhein on human articular chondrocytes in alginate beads. Biochem Pharmacol 2003;65: 377-88. [Pubmed]
[7] Henrotin YE, Sanchez C, Deberg MA, Piccardi N, Guillou GB, Msika P, Reginster JY. Avocado/soybean unsaponifiables increase aggrecan synthesis and reduce catabolic and proinflammatory mediator production by human osteoarthritic chondrocytes. J Rheumatol 2003;30: 1825-34. [Pubmed]
[8] Henrotin YE, Labasse AH, Jaspar JM, De Groote DD, Zheng SX, Guillou GB, Reginster JY. Effects of three avocado/soybean unsaponifiable mixtures on metalloproteinases, cytokines and prostaglandin E2 production by human articular chondrocytes. Clin Rheumatol 1998;17: 31-9. [Pubmed]
[9] Henrotin YE, Labasse AH, Simonis PE, Zheng SX, Deby GP, Famaey JP, Crielaard JM, Reginster JY. Effects of nimesulide and sodium diclofenac on interleukin-6, interleukin-8, proteoglycans and prostaglandin E2 production by human articular chondrocytes in vitro. Clin Exp Rheumatol 1999;17: 151-60. [Pubmed]
[10] Henrotin YE, Zheng SX, Labasse AH, Deby GP, Crielaard JM, Reginster JY. Modulation of human chondrocyte metabolism by recombinant human interferon. Osteoarthritis Cartilage 2000;8: 474-82. [Pubmed]
[11] Sanchez C., Deberg M. A., Bellahcene A., Castronovo V., Msika P., Delcour J. P., Crielaard J. M. and Henrotin Y. E. Phenotypic characterization of osteoblasts from the sclerotic zones of osteoarthritic subchondral bone. Arthritis Rheum 2008;58 (2): 442-455. [Pubmed]
[12] Sanchez C., Gabay O., Salvat C., Henrotin Y. E. and Berenbaum F. Mechanical loading highly increases IL-6 production and decreases OPG expression by osteoblasts. Osteoarthritis Cartilage 2009;17 (4): 473-81. [Pubmed]
[13] Henrotin YE, Deberg MA, Crielaard JM, Piccardi N, Msika P, Sanchez C. Avocado/soybean unsaponifiables prevent the inhibitory effect of osteoarthritic subchondral osteoblasts on aggrecan and type II collagen synthesis by chondrocytes. J Rheumatol 2006;33: 1668-78. [Pubmed]