Lysophosphatidic acid receptor 1 suppression sensitizes rheumatoid fibroblast-like synoviocytes to tumor necrosis factor-induced apoptosis

• Published in: Arthritis & rheumatology (Hoboken, N.J.) N.J.), 2012-08, Vol.64 (8), p.2460-2470
• Main Authors: Orosa, Beatriz, González, Antonio, Mera, Antonio, Gómez-Reino, Juan J., Conde, Carmen
• Format: Article
• Language: English
• Subjects: Apoptosis; Apoptosis - drug effects; Arthritis; Arthritis, Rheumatoid - metabolism; Arthritis, Rheumatoid - pathology; Biological and medical sciences; CARD Signaling Adaptor Proteins; Cell Proliferation - drug effects; Cells, Cultured; Cytoskeletal Proteins - metabolism; Diseases of the osteoarticular system; Drug therapy; Gene Expression Regulation - drug effects; Humans; Inflammatory joint diseases; Medical research; Medical sciences; Osteoarthritis - metabolism; Osteoarthritis - pathology; Receptors, Lysophosphatidic Acid - deficiency; Receptors, Lysophosphatidic Acid - drug effects; Receptors, Lysophosphatidic Acid - genetics; RNA, Small Interfering - pharmacology; Synovial Membrane - metabolism; Synovial Membrane - pathology; TNF Receptor-Associated Death Domain Protein - metabolism; TNF-Related Apoptosis-Inducing Ligand - metabolism; Tumor Necrosis Factor-alpha - pharmacology
• ISSN: 0004-3591; 2326-5191; 1529-0131; 2326-5205
• DOI: 10.1002/art.34443

Objective To investigate the role of lysophosphatidic acid (LPA) receptors in the proliferation and apoptosis of fibroblast‐like synoviocytes (FLS) from patients with rheumatoid arthritis (RA). Methods Expression of LPA receptors 1–3 was analyzed by real‐time polymerase chain reaction (PCR). LPAR1 and LPAR2 were suppressed in RA FLS by small interfering RNA (siRNA) transfection. Proliferation of RA FLS after tumor necrosis factor (TNF) and LPA stimulation was determined with a luminescent cell viability assay. Apoptosis was analyzed by quantification of nucleosome release and measurement of activated caspase 3/7. Genes involved in the apoptotic response were identified with a human apoptosis PCR array and validated with Western blot assays. The requirement of these genes for apoptosis sensitization was assessed by siRNA transfection. Secretion of mediators of inflammation was analyzed by enzyme‐linked immunosorbent assay. Results Only LPAR1 and LPAR2 were expressed by RA FLS, and their levels were higher than those in osteoarthritis (OA) FLS. Suppression of LPAR1 abrogated TNF‐induced proliferation and sensitized the RA FLS, but not the OA FLS, to TNF‐induced apoptosis. These changes occurred despite an increased early inflammatory response to TNF. Sensitization to apoptosis was associated with changes in expression of multiple apoptosis‐related genes. Three of the up‐regulated proapoptotic genes were further studied to confirm their involvement. In contrast, suppression of LPAR2 showed no effect in any of these analyses. Conclusion LPA1 is an important receptor in RA FLS. Its suppression is accompanied by a global increase in the response to TNF that is ultimately dominated by sensitization to apoptosis.