Figure 3.

IGFBP2 production is induced by IGF-1 and is functionally active in psoriatic keratinocytes. IGFBP2 release was analysed by ELISA in protein lysates and supernatants (sups) of untreated healthy and pso KC cultures (A) or in sups of healthy and pso KC stimulated or not with 10 ng/ml IGF-1 or IGF-2 for 24 hours. (B) Data are expressed as mean of ng/10⁶ cells ± SD of three different experiments carried out on different strains (n = 3). *p ≤ 0.05, as assessed by unpaired Student’s t test; **p ≤ 0.01, as calculated by the Mann–Whitney U test, comparing IGFBP2 production between healthy and pso KC groups. (C) BrdU incorporation was evaluated in healthy KC grown on coverslips and treated with complete medium alone (i) or 10-fold concentrated sup. of healthy KC (ii) or of pso KC (Pso) (iii). In parallel, healthy KC were treated with IGF-1 (10 ng/ml) alone (iv), or in presence of 10-fold concentrated healthy KC sups (v), or pso KC sups (vi). In two experimental conditions, 1 μg/ml of neutralizing anti-IGFBP2 Ab (α-IGFBP2) was added (vii, viii). Images are relative to one of three independent experiments performed on three different healthy KC strains. (D) Graphs represent the number of BrdU-positive cells counted in high power fields and expressed as cells per area unit ± SD (n [microscopic fields per slide] = 6); H, healthy KC sups; Pso, psoriatic KC sups. (E) CXCL8 mRNA levels were detected by Real-time PCR in healthy KC stimulated for 8 hours with IGF-1 (10 ng/ml) alone or in combination with M4, including IFN-γ (200 U/ml), TNF-α (50 ng/ml), IL-17A (50 ng/ml) and IL-22 (50 ng/ml), in presence or not of healthy (H) or psoriatic (Pso) supernatants. In two experimental settings, 1 μg/ml of neutralizing anti-IGFBP2 Ab (α-IGFBP2) was added. Data shown are means of relative mRNA expression (normalized to GAPDH) of three independent experiments. (D, E), *p ≤ 0.05, as calculated by unpaired Student’s t test.