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Ankylosing spondylitis (AS) is characterized by abnormal bone formation in the spine and the sacroiliac joints.  In vitro  assays demonstrate that carbonic anhydrase I (CA1) promotes calcium precipitation. This study investigated the function of CA1 for bio-mineralization and determined if common polymorphisms in the CA1 gene might contribute to AS risk.    Methods  Calcification was induced in Saos-2 cells, a human osteosarcoma cell line, with ascorbic acid and β-glycerophosphate. Calcification was determined by Alizarin Red-S (AR-S) staining. Expressions of CA1, alkaline phosphatase (ALP), bone sialoprotein (BSP), osteocalcin (OCN), osterix (OSX) and runt-related transcription factor-2 (Runx2) were determined by real-time PCR and western blotting. The cells were also treated with acetazolamide, an anti-carbonic anhydrase drug. Genotyping was performed using Illumina VeraCode microarray in a case-control study including 51 AS patients, 267 rheumatoid arthritis (RA) patients and 160 healthy controls. The result was confirmed by Taqman assay, including 258 AS patients, 288 RA patients and 288 healthy controls.    Results  Following the induction of calcification, Saos-2 cells produced large amounts of calcium-rich deposits. Increased transcriptions of CA1, ALP, BSP, OCN, OSX and Runx2, essential genes for ossification, were detected in the cultured cells. Following treatmen with acetazolamide, the expression of CA1 obviously declined and mineralized nodule formation was also decreased. Illumina microarray indicates that SNP at rs7841425 also showed significant differences in allelic frequency ( P  = 0.01396) and genotypic frequency ( P  = 0.005902) between AS cases and controls. In addition, SNP at rs7827474 showed significant differences in allelic frequency ( P  = 5.83E-04) and genotypic frequency ( P  = 0.000186) between RA cases and controls ( P  values were adjusted to multiple comparisons). The Taqman assay revealed that rs725605 demonstrated statistically significant evidence of allele frequency ( P  = 0.022307) and gene frequency ( P  = 0.007731) for association with AS. This SNP did not show significant differences in allelic frequencies and gene frequencies between RA patients and controls.    Conclusions  CA1 may play an essential role in bio-mineralization and new bone formation. The gene encoding CA1 is susceptible to AS.

Carbonic anhydrase I (CA1) is involved in the process of bone formation and is susceptible to ankylosing spondylitis