Figure 5. The sPLA₂-IIA–mediated increased susceptibility to arthritis is dependent upon the intestinal flora.

(A) Twelve-week-old WT and sPLA₂-IIA^TGN mice housed in an Elite SPF+ animal facility were administered broad-spectrum antibiotics for 23 days. On experimental days 14 and 16, mice were injected i.p. with 150 μL of K/B×N serum (black arrows) to induce arthritis, and the disease severity was monitored daily by measuring ankle thickness (n = 16–30 from 2 separate experiments). (B and C) Quantification of sPLA₂-IIA by time-resolved fluoroimmunoassay in serum (n = 5–10) and the intestinal compartments of arthritic sPLA₂-IIA^TGN mice treated or not with antibiotics (n = 4–5). Dotted line indicates mean concentration in sPLA₂-IIA^TGN mice. (D) The severity of serum-transferred arthritis was evaluated in 12-week-old transgenic mice depleted of IL-17A (IL-17A^−/−) (n = 8–10). (E) Assessment of intestinal permeability in arthritic and nonarthritic mice by quantification of serum 4 kDa FITC-Dextran (FD4) translocated to the circulation following administration by oral gavage (n = 5). (F and G) Ten-week-old WT and sPLA₂-IIA^TGN mice were administered antibiotics for 1 week to deplete their microbiota. On day 7, mice were administered a polyethylene-glycol–based laxative to empty their bowels, and a fecal microbiota transplantation (FMT) was performed. In brief, fresh fecal matter solution was administered by oral gavage to mice once a day for 3 consecutive days. Mice were then allowed to rest for 10 days before arthritis was induced by injection of K/B×N serum (black arrows). The severity of the disease was monitored daily (n = 10). Data from 1 (C–G) to 2 (A and B) separate experiments are presented as mean ± SEM. Statistical analysis included the following: (A, D, and G) repeated-measures 2-way ANOVA evaluating the statistical variation between groups. (B, C, and E) One-way ANOVA with Tukey’s multiple comparisons test. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.