The role of interleukin-35-producing regulatory B cells in immune pathogenesis of Kawasaki disease

Abstract

Objective To investigate the role of IL-35-producing regulatory B cells(IL-35+ Breg)in immunological pathogenesis of Kawasaki disease (KD). Methods Thirty-two children with KD and 28 age-matched healthy children were allowed to participate in this study.Flow cytometry was performed to evaluate the proportions of IL-35+ Breg as well as requlatory T cells (Treg)and expression levels of associated molecules such as programmed death-ligand 1(PD-L1), CD169, programmed death 1(PD-1), CD43, IL-12p35, epstein-Barr virus induced 3(IL-27EBI3). IL-12 receptor beta 2(IL-12Rβ2), IL-27 receptor alpha(IL-27Rα), phosphated signal transdu-cer and activator of transcription 1 (pSTAT1)and phosphated signal transducer and activator of transcription 3 (pSTAT3). Transcription levels of the Src homology 2 domain-containing protein tyrosine phosphatase 2 (SHP-2), phosphatase and tensin homolog (PTEN), vav1 guanine nucleotide exchange factor(Vav)in CD19+ B cells were determined by using quantitative real-time PCR.Plasma concentrations of IL-35, tumor necrosis factor α(TNF-α) and IL-12 were measured by adopting enzyme-linked immunosorbent assay. Results (1)The proportions of IL-35+ Breg and its expressions of IL-12p35, IL-27EBI3 and IL-10 in patients with acute KD were lower than those of healthy controls [IL-35+ Breg: (5.79±2.60)% vs (12.65±5.34)%; F=19.23, 9.70, 14.30.7.08; all P<0.05], but they were significantly increased after intravenous immune globulin(IVIG) treatment [IL-35+ Breg: (10.52±4.95)%; all P<0.05]. (2)The proportions of Treg and its transcriptional levels of IL-12p35 and IL-27EBI3 were down-regulated during acute KD[Treg: (4.12±1.51)% vs (8.06±3.32)%; F=19.70, 17.69, 38.22; all P<0.05], but were increased after therapy[Treg: (7.39±2.85)%; P<0.05]. A positive correlation was found between the proportions of Treg and IL-35+ Breg during acute KD(r=0.69, P<0.05). Meanwhile, plasma concentrations of IL-35 and expression levels of IL-12Rβ2, IL-27Rα, pSTAT1 and pSTAT3 in CD19+ B cells were significantly down-regulated in children with acute KD, but they were increased after treatment(F=8.09, 7.54, 7.69, 5.89, 12.59, all P<0.05). (3)Compared with healthy controls, expressions of PD-L1 and CD169 on CD14+ cells and plasma concentrations of TNF-α and IL-12 were elevated during acute KD(F=24.94, 16.53, 34.71, 19.51; all P<0.05). Expression levels of PD-1, CD43 and its downstream molecules (SHP-2, PTEN, Vav)in CD19+ B cells were down-regulated during acute KD(F=6.43, 5.57, 19.52, 10.37, 11.37; all P<0.05), and restored remarkably after therapy(all P<0.05). Conclusion Insufficiency of IL-35+ Breg and its expression of IL-35 may be the important factors contributing to immunological dysfunction in KD. Key words: Interleukin-35; Regulatory B cell; Kawasaki disease; Regulatory T cell; Immune regulation