2.10 Statistical analysis
The Prism version 6 software (GraphPad Software, San Diego, CA, USA) was
used for statistical analysis. Data were expressed as the mean ±
standard error of the mean (SEM) or standard deviation (SD). All P
values were calculated using the Student’s t -test or one-way
analysis of variance (ANOVA). Statistical significance was set at P
< 0.05.
3.
Results
3.1
Expression of IL-35 was significantly increased in
patie nts
with psoriasis
To investigate the involvement of IL-35 in skin diseases, we first
examined IL35 expression in the serum of healthy controls and patients
with psoriasis. We recruited patients (n = 53) and age/sex-matched
healthy donors (n = 20); the demographics are summarised in Table 1.
Using ELISA, we found that the levels of IL-35 were significantly
elevated in the serum from patients with psoriasis compared with those
in healthy controls
(Fig.
1A, p = 0.0089). Moreover, we observed that as the symptoms of
psoriasis worsened (increased PASI score), the expression of IL-35 was
increased. More specifically, the expression of IL-35 in specimens with
a PASI score
greater
than or equal to 3 points was significantly higher than that in
specimens with a PASI score less than 3 points (Fig. 1A). We also found
increased populations of
CD4+EBI3+p35+ and
CD19+EBI3+p35+cells in the peripheral blood of patients with psoriasis via flow
cytometry (Fig. 1B-D). We demonstrated that IL-35 secreted by both T and
B lymphocytes was significantly increased (p < 0.01 and p
< 0.05, respectively). Moreover, immunofluorescence staining
results showed that the expression of IL-35 in the skin tissue of
patients with psoriasis increased significantly (Fig. 1E).
3.2 M-MDSCs were
significantly expanded in patients with psoriasis
We
examined whether the number of M-MDSCs is altered in
patients
with psoriasis. Using flow cytometry, we confirmed that the
number
of human M-MDSCs
(CD11b+CD14+HLA-DR-)
was remarkably increased in the peripheral blood of patients with
psoriasis
(Fig.
2A-B, p = 0.0065). Next, immunofluorescence staining of
CD14+HLA-DR- M-MDSCs in the inflamed
skin of patients with psoriasis revealed that the infiltration of M-MDSC
cells in psoriatic skin tissue increased significantly compared to that
in healthy controls (p < 0.01).