Demographic and clinical features of RA patients
60 RA patients as well as 20 age and gender matched OA patients and 20 healthy controls were enrolled in the study. Among 60 RA patietns, there were 12 (20%) males and 48 (80%) females with mean age of 54.8±11.6 years and median disease duration of 6.3 (0.5-34) years. 37 (61.7%) of them were rheumatoid factor (RF) positive and 53 (88.3%) were anti-cyclic citrullinated protein (anti-CCP) positive (Table 1). All the 60 patients were distributed in four different disease activity groups, including remission, low disease activity (LDA), moderate disease activity (MDA) and high disease activity (HDA), evaluated by disease activity score based on 28 joint count-C reactive protein (DAS28-CRP), DAS28-erythrocyte sedimentation rate (DAS28-ESR), simplified disease activity index (SDAI) and clinical disease activity index (CDAI).
The comparison of syndecan-4 in serum, synovial fluid and synovial tissue among RA patients, OA patients and healthy controls
The syndecan-4 concentrations in sera of 60 RA patients, 20 OA patients and 20 healthy controls were 509.3 (345.2-769.8) pg/mL, 363.6±137.6 pg/mL and 204.4±59.1pg/mL, respectively. The serum syndecan-4 concentration was significantly higher in RA patients than in OA patients and healthy controls (p<0.001, p<0.001, respectively). It was also higher in OA patients than in healthy controls (p<0.001). (Figure 1)
In the 23 paired serum and SF samples of RA patients, the syndecan-4 concentrations was significantly lower in SF than in sera (234.3±106.3 pg/mL vs 85.3±61.2 pg/mL, p<0.001).(Figure S1)
The syndecan-4 concentrations was also significantly lower in SF than in sera of OA patients [363.6±137.6 pg/mL vs 51.0 (27.2-163.6) pg/mL, p<0.001], although the sera and SF samples were not paired.
No significant difference of syndecan-4 concentrations in SF was found between the RA and OA patients (p=0.733), although it was slightly higher in RA patients.
Syndecan-4 expression was detected by immunohistochemistry in the lining and sublining layers of all the 5 RA and 5 OA patents (Figure 2). The mean area, mean diameter, mean density and mean integrated optical density (IOD) were comparable between RA and OA patients (Table S1).
Immunofluorescence demonstrated that Syndecan-4 was positively stained in cultured synovial fibroblasts from both of RA and OA patients. The fluorescence intensity was similar between RA and OA patients. (Figure S2)
The comparison of serum syndecan-4 concentration between sera-positive and sera-negative RA patients
The syndecan-4 concentrations in sera were significantly higher in RF positive patients than in RF negative patients [586.2 (390.2-1124.8) pg/mL vs 434.3 (312.0-537.5) pg/mL, (p=0.009)]. The binary logistic regression showed that the OR was 1.003 (1.000-1.005, p=0.022). For anti-CCP antibody positive and negative patients, it was comparable [517.4 (346.2-781.0) pg/mL vs 500.7 (373.8-537.5) pg/mL, p=0.542] (Figure S3).
Then we defined patients with at least one of the above two antibodies positive as “sera positive”, and there were 53 sera positive and 7 sera negative patients. The result was the same as that when stratified by anti-CCP antibody.
The correlation between Syndecan-4 concentration with disease activity of RA patients
Among the 60 RA patients, the syndecan-4 concentration in sera was positively correlated with TJC, SJC, PGA, EGA, and the disease activity scores, including DAS28-ESR, DAS28-CRP, SDAI and CDAI (Table 2). The correlation coefficents (r value) were mild to moderate (Table 2). Based on any of these disease activity scores, the patients in HDA had the highest serum syndecan-4 concentration, compared with those in remission, LDA or MDA (Table 3, Figure 3)
Among the 23 RA patients, the syndecan-4 concentration in SF was positively correlated with PGA, EGA, CRP, DAS28-ESR, DAS28-CRP and SDAI (Table S2). All of the correlation coefficents (r value) were higher than 0.5.