Background Rheumatoid arthritis (RA) is a chronic, inflammatory and systemic autoimmune disease that leads to progressive cartilage destruction. Advances in the treatment of RA-related destruction of cartilage require profound insights into the molecular mechanisms involved in cartilage degradation. Until now, comprehensive data about the molecular RA-related dysfunction of chondrocytes have been limited. Hence, the objective of this study was to establish a standardized in vitro model to profile the key regulatory molecules of RA-related destruction of cartilage that are expressed by human chondrocytes.

Nonbiological therapeutics are frequently used for the treatment of patients with rheumatoid arthritis (RA). Because the mechanisms of action of these therapeutics are unclear, the authors aimed to elucidate the molecular effects of typical antirheumatic drugs on the expression profile of RA-related genes expressed in activated synovial fibroblasts. For reasons of standardization and comparability, immortalized synovial fibroblasts derived from RA (RASF) and normal donors (NDSF) were treated with methotrexate, prednisolone, or diclofenac and used for gene expression profiling with oligonucleotide microarrays. The cytotoxicity of the antirheumatic drugs was tested in different concentrations by MTS tetrazolium assay. Genes that were differentially expressed in RASF compared to NDSF and reverted by treatment with antirheumatic drugs were verified by semiquantitative polymerase chain reaction and by chemiluminescent enzyme immunoassay. Treatment with methotrexate resulted in the reversion of the RA-related expression profile of genes associated with growth and apoptosis including insulin-like growth factor binding protein 3, retinoic acid induced 3, and caveolin 2 as well as in the re-expression of the cell adhesion molecule integrin α6. Prednisolone reverted the RA-related profile of genes that are known from inflammation and suppressed interleukins 1β and 8. Low or high doses of diclofenac had no effect on the expression profile of genes related to RA in synovial fibroblasts. These data give the first insight into the mechanisms of action of common antirheumatic drugs used for the treatment of arthritides. Synovial fibroblasts reflect the disease-related pathophysiology and are useful tools for screening putative antirheumatic compounds. (Journal of Biomolecular Screening 2007:328-340)

Introduction Rheumatoid arthritis (RA) leads to progressive destruction of articular cartilage. This study aimed to disclose major mechanisms of antirheumatic drug action on human chondrocytes and to reveal marker and pharmacological target genes that are involved in cartilage dysfunction and regeneration.

Pellet formation of freshly isolated chondrocytes followed a reproducible developmental kinetics and showed typical immature hyaline cartilage properties. Such uniform cartilage pellets are very useful as a substrate for interactive cell culture models that simulate diseases like RA.

Historically, catecholamine-producing chromaffin cells and steroid-producing adrenocortical cells have been regarded as two independent endocrine systems that are united under a common capsule to form the adrenal gland. There is increasing evidence for bidirectional interactions, with regulatory influences of adrenocortical secretory products on adrenomedullary functions and vice versa. However, the direct involvement of chromaffin cells on the regulation and maintenance of cortical function has not yet been demonstrated. Therefore, we analyzed glucocorticoid secretion and P450 messenger RNA (mRNA) expression in bovine adrenocortical cells in cocultures with chromaffin cells compared with those in pure cortical cell cultures.Cortisol release from cortical cells in coculture with chromaffin cells was 10 times as high (mean Ϯ SEM, 1035 Ϯ 119%) as that from the same number of isolated cortical cells (100 Ϯ 11%). By a [3 H]thymidine incorporation assay, it was demonstrated that this effect was not due to a higher proliferation rate. Northern analysis revealed an increasing expression of P450 17␣ mRNA in the coculture from days 1-5, whereas in isolated cortical cells, P450 17␣ mRNA decreased, leading to a 6-fold difference on day 5. Inhibitors of protein (cycloheximide) or RNA (actinomycin D) synthesis completely annulled the observed increase in cortisol release, indicating that de novo protein synthesis is required for this activation of adrenocortical steroidogenesis. Addition of the cyclooxygenase inhibitor indomethacin reduced the stimulatory effect, suggesting that this stimulation is in part mediated by PGs. Locally produced ACTH, catecholamines, and interleukin-1 accounted for 43% of the effect. Secretory products of chromaffin cells that act in concert are believed to be responsible for the stimulation of steroidogenesis in the coculture.The coculture system is an in vitro model that corresponds to the in vivo situation in the intact adrenal gland, where both endocrine cell systems are in close contact. Our data demonstrate the requirement of intraadrenal cellular communication for the full strength of the adrenocortical hormonal response. (Endocrinology 139: [772][773][774][775][776][777][778][779][780] 1998)