Overexpression of the transmembrane carbonic anhydrase isoforms IX and XII in the inflamed synovium

Figures & data

Table 1. Levels of carbonic anhydrase present in the lysed samples of blood, serum, synovial fluid and synoviocytes (from healthy controls or JIA patients) and inhibition data with the standard sulfonamide inhibitor acetazolamide (AAZ) and CA IX/CA XII-selective Coumarin inhibitor C (Figure 1) by a stopped flow CO₂ hydrase assay.

Sample	% CA activity
	Healthy donors	JIA patients
Blood	100 ± 3.9	100 ± 6.2
Serum	100 ± 5.1	100 ± 4.8
Synovial fluid	NO	135.4 ± 12
Synoviocytes	100 ± 4.7	129.2 ± 8.1
Blood + AAZ, 10 μM	6.8 ± 0.2	5.9 ± 0.5
Serum + AAZ, 10 μM	6.3 ± 0.5	6.1 ± 0.4
Synovial fluid + AAZ, 10 μM	NO	5.5 ± 0.2
Synoviocytes + AAZ, 10 μM	6.0 ± 4.2	5.9 ± 0.7
Blood + Coumarin C, 10 μM	100 ± 5.1	100 ± 4.5
Serum + C, 10 μM	100 ± 3.9	100 ± 4.5
Synovial fluid + C, 10 μM	NO	25.8 ± 3.4
Synoviocytes + C, 10 μM	16.0 ± 33.8	23.4 ± 2.3

Figure 1. The pan-inhibitor of CAs acetazolamide (AAZ) and its inhibition constants against isoforms CA I, II, IX and XII, and the newly developed, isoform-selective Coumarin inhibitor C, and its inhibitory activity against the same isoforms.

Figure 2. The CA IX and CA XII mRNA expression levels were quantified in healthy and JIA synovial fibroblasts using real time-PCR. 18s was used as housekeeping gene. Results of three independent experiments performed in triplicates are expressed as fold change according to 2^−ΔΔCT method.

Figure 3. Western blotting detection of CA IX and CA XII in synovial fibroblasts from healthy donors and from JIA patients. Tubulin expression and Ponceau staining are used as a control for protein loading. Histograms on the right report band densitometry.