The Role Of IL-10on The Abnormal Pregnant Outcome Induced By Toxoplasma Gondii Infection During Early Pregancy

Objective:To investigate the potential role of IL-10for the abnormal pregnancy outcome of C57BL/6mice induced by T. gondii infection. Methods:Establish the early pregnancy infection model of C57BL/6mice in vivo. Pregnant mouse were divided into control group (CG), infected group (IG), infected plus IL-10treated group (IPTG). Pregnant mice of IG and IPTG were infected intraperitoneally with400T. gondii tachyzoites per mice on gestation day (GD)8. IPTG mice were injected with lug recombinant mouse IL-10per mice on GD10and GD12respectively. Dams were executed on gestation15, pregnant outcome were recorded; the ratio of NKG2A and Qa-1positive cells of placenta were analyzed with flow cytometry; the mRNA expression of NKG2A and Qa-1were analyzed with real-time RT-PCR; the protein expressions of IL-4, IL-10and IFN-r of placenta were analyzed with ELISA. BeWo cells and human trophoblast cells were infected in vitro with T. gondii tachyzoites of54h-post infection at ratio of1:3(cell:T. gondii). Recombinant human IL-10was added into the supernatant at the concentration of50ng/ml. Cells were collected at16,24,36,48h and60h post infection separately. The protein expressions of HLA-G and cell apoptosis ratio of both cells were analyzed with flow cytometry. The mRNA expression of HLA-G, caspase3, caspase8and c-FLIP in BeWo were analyzed with real-time PCR. Results:Pregnant mice of CG showed nice mental and agile action whereas those of IG showed drooping spirit and low temperature. IPTG showed better than IG. The placentas and fetuses of CG showed flesh red and normal blood supply, and the average weight of placenta of CG was95mg. The placentas and fetuses of IG were both smaller than those of CG, the average weight of IG was40mg, and showed severe extravasated blood and the fetuses showed fatty generation and bad blood supply. The placentas and fetuses of IPTG were bigger than those of IG, the average weight of placenta of IPTG was70mg, extravasated blood was less-common and the fetus showed better blood supply and more normal colour. The fetal resorption ratio of IG was hIGher compared to CG and the ratio in IPTG was sIGnificantly lower compared to IG; NKG2A positive cells ratio in IG was increased compared to CG and the ratio in IPTG was increased compared to IG; Qa-1positive cells ratio in IG was increased compared to CG and increased in IPTG compared to IG; The mRNA expression of NKG2A in IG was increased compared to CG and the expression in IPTG was increased compared to IG; The mRNA expression of Qa-1in IG was increased compared to CG and increased in IPTG compared to IG; IL-4and IL-10in IG were both decreased compared to CG and were both increased in IPTG compared to IG The concentration of IFN-y in IG was increased compared to CG and decreased in IPTG compared to IG IFN-r/IL-4and IFN-r/IL-10were both decreased in IPTG compared to IG The apoptosis of BeWo and Tro of CG were both low during experimental time. Obvious apoptosis in IG and IPTG were observed and increased with the infection time (all p<0.05). IPTG showed lower apoptosis compared to IG (p<0.05). The MFI (mean fluorescence intensity) of HLA-G were sIGnificantly increased during16-36h and decreased during48-60h compared to CG (p<0.05). IPTG showed hIGher HLA-G MFI during48-60h, interesting, lower HLA-G MFI were detected during16-36h compared to IG(all p<0.05). The mRNA levels of HLA-G showed similar trend to the protein expression. Caspase3and caspase8were both up-regulated in IG compared to control (p<0.05) and down-regulated in IPTG compared to IG (p<0.05). C-FLIP was down-regulated in IG compared to CG (p<0.05) and were up-regulated in IPTG compared to IG (p<0.05). Conclusion:T. gondii infection induces severe adverse pregnant outcomes in C57BL/6mice. The inhibitory receptor of NK cells NKG2A and its1IGand-the non-classical MHC-I molecule Qa-1at the maternal-fetal interface were both up-regulated; the imbalance of Th1/Th2due to T. gondii infection was corrected evidenced by down-regulated Thl type cytokines and up-regulated Th2cytokines following IL-10treatment, and this was why the abnormal pregnant outcomes resulted from was improved by IL-10. HLA-G was up-regulated following IL-10treatment during late infection period in BeWo and Tro cells infected with T. gondii, maybe enhancing the combination with the inhibitory receptors of NK cell in vivo and maintaining the immunu tolerance in contation of T. gondii infection. The apoptosis of BeWo and Tro cells were both reduced by IL-10, maybe as a consequence of the down-regulated caspase3and caspase8and up-regulated c-FLIP following IL-10treatment which was another reason that IL-10could improve the abnormal pregnancy outcome resulted from T. gondii infection. This study certainly will provide the theory for exploring the preventive and curative method for clinical T. gondii infection during gestation and immit the new method and thought into the investIGation to relieve the adverse pregnant outcome induced by T. gondii infection.