Identification of known and novel genes in activated monocytes from patients with rheumatoid arthritis

Objective To define gene activation patterns of monocytes (MO) in patients with rheumatoid arthritis (RA). Methods A complementary DNA (cDNA) library was constructed from first‐leukapheresis MO obtained from an RA patient with active disease; 32 P‐labeled cDNA from first‐leukapheresis MO (activated pool) and third‐leukapheresis MO (nonactivated pool) were used as probes for differential hybridization. Reverse transcriptase–polymerase chain reaction (RT‐PCR) was used to assess gene activation in MO from an additional 26 RA patients and 6 normal controls. Results Subtraction of genes from first‐ and third‐leukapheresis MO resulted in 482 differentially expressed clones. In first‐leukapheresis MO, these clones included the following: 1) interleukin‐1α (IL‐1α), IL‐1β, IL‐6, tumor necrosis factor α, growth‐related oncogene α (GROα)/melanoma growth‐stimulatory activity, macrophage inflammatory protein 2/GROβ, ferritin, α 1 ‐antitrypsin, lysozyme, transaldolase, Epstein‐Barr virus–encoded RNA 1 (EBER‐1)/EBER‐2–associated‐protein, thrombospondin 1, an angiotensin receptor II (ATRII) C‐terminal homolog, and RNA polymerase II elongation factor (elongin); 2) two clones homologous to functionally undefined genes (BSK‐67 and BSK‐83); and 3) three unknown cDNA sequences (BSK‐66, 80, 89). In third‐leukapheresis MO, the clones included differentiation genes (HOX‐B3, thymosin‐β 4 , PU.1, glucocerebrosidase, MEL‐18, and glucose‐6‐phosphate dehydrogenase) and 3 unknown/functionally undefined sequences. Differential expression of most genes from the activated pool was confirmed in leukapheresis samples from 2 additional RA patients. In MO from RA patients, not only were IL‐1β and the ATRII homolog significantly overexpressed (maximum 36‐fold), but also 4 of the unknown/functionally undefined genes (maximum 102‐fold). Notably, messenger RNA levels of BSK‐89 correlated positively with the erythrocyte sedimentation rate (ESR), whereas those of BSK‐83 correlated negatively with the ESR and C‐reactive protein level. Conclusion The combined strategy of gene subtraction and semiquantitative RT‐PCR may allow the definition of MO activation patterns during different disease phases (including therapy‐induced remission) and the identification of novel MO genes with pathogenetic relevance in RA.