Reduced PICD in Monocytes Mounts Altered Neonate Immune Response to Candida albicans

BackgroundInvasive fungal infections with Candida albicans (C. albicans) occur frequently in extremely low birthweight (ELBW) infants and are associated with poor outcome. Phagocytosis of C.albicans initializes apoptosis in monocytes (phagocytosis induced cell death, PICD). PICD is reduced in neonatal cord blood monocytes (CBMO).HypothesisPhagocytosis of C. albicans causes PICD which differs between neonatal monocytes (CBMO) and adult peripheral blood monocytes (PBMO) due to lower stimulation of TLR-mediated immune responses.MethodsThe ability to phagocytose C. albicans, expression of TLRs, the induction of apoptosis (assessment of sub-G1 and nick-strand breaks) were analyzed by FACS. TLR signalling was induced by agonists such as lipopolysaccharide (LPS), Pam3Cys, FSL-1 and Zymosan and blocked (neutralizing TLR2 antibodies and MYD88 inhibitor).ResultsPhagocytic indices of PBMO and CBMO were similar. Following stimulation with agonists and C. albicans induced up-regulation of TLR2 and consecutive phosphorylation of MAP kinase P38 and expression of TNF-α, which were stronger on PBMO compared to CBMO (p ConclusionOur data suggest direct involvement of TLR2-signalling in C. albicans-induced PICD in monocytes and an alteration of this pathway in CBMO.【研究背景】白假丝酵母菌（Candida albicans, C. albicans）侵袭性真菌感染在极低出生体重（extremely low birthweight, ELBW）婴儿中高发，且与不良预后密切相关。白假丝酵母菌的吞噬作用可诱导单核细胞发生凋亡，即吞噬诱导细胞死亡（phagocytosis induced cell death, PICD）。新生儿脐带血单核细胞（cord blood monocytes, CBMO）的PICD水平显著降低。 【研究假说】白假丝酵母菌的吞噬作用可诱发PICD，由于Toll样受体（TLR）介导的免疫应答刺激不足，新生儿CBMO与成人外周血单核细胞（peripheral blood monocytes, PBMO）的PICD水平存在差异。 【实验方法】采用流式细胞术（FACS）检测各组单核细胞的白假丝酵母菌吞噬能力、TLR表达水平以及凋亡诱导情况（通过检测亚G1期细胞与DNA断链进行评估）。使用脂多糖（lipopolysaccharide, LPS）、Pam3Cys、FSL-1以及酵母聚糖（Zymosan）等激动剂激活TLR信号通路，并通过抗TLR2中和抗体与MYD88抑制剂阻断该通路。 【实验结果】PBMO与CBMO的吞噬指数无显著差异。经激动剂与白假丝酵母菌刺激后，TLR2表达上调，继而P38丝裂原活化蛋白激酶发生磷酸化，肿瘤坏死因子-α（TNF-α）表达升高；上述变化在PBMO中的幅度显著高于CBMO（P< 【研究结论】本研究结果表明，TLR2信号通路直接参与单核细胞中白假丝酵母菌诱导的PICD过程，且CBMO中的该通路存在功能异常。