Study On Expression Of ANXA1、GATA-3and T-bet In Th1/Th2Cells From Burn Sepsis Mouse At Early Stage

Purposes:To observe the change of the Thl, Th2in the peripheral blood and the expression of the ANXA1, GATA-3, T-bet proteins and mRNA of the Kunming mice in pathological conditions of burns, sepsis and burn sepsis, It tries to discuss the immune regulation mechanism of ANXA1, GATA-3in burn sepsis, find the critical factor in the immune suppression of burn sepsis and provide research basis in order to find the new drug targets and make effective prevention and treatment on burn sepsis.MethodsFirstly, it used the male Kunming mice in SPF level, with the weight of35-40g. They were divided into4groups:SHAM group,burn group, endotoxin injection group and burn sepsis group. The model of20%deep II degree burns was constructed. Then it took the peripheral blood of mice for detected in12h,24h,48h and72h after injured.Secondly, CD4+IFN-γ+and CD4+IL-4+was used to mark the Thl and Th2cells, and the proportions of Thl and Th2cells and the condition of Th1/Th2balance displacement was observed. Then it supposed the gate of Th1/Th2cells to detect the ANXA1, GATA-3and T-bet.Thirdly, the method of RT-PCR was used to detect the expression of ANXA1, GATA-3, T-bet and mRNA in the peripheral blood lymphocytes.Fourthly, the data was put into software Excel to make statistical processing, and the software SPSS was used to process experimental data. Results1. Detection of flow cytometryThe proportion of Th1and Th2cells in burns group was gradually decreasing in12h,24h after injured, and slowly recovery in48h and72h after injured. While the proportions of Thl and Th2cells in endotoxin injection group and burns sepsis group were gradually decreasing, and the Thl/Th2balance shifted direction of Th2. It has significant different with the SHAM group (P<0.05).The expression rate of the ANXA1and GATA-3in Thl, Th2cells in peripheral blood of the burns group from24h after injured was lower than SHAM group (P<0.05), and then changed rising. The expression rate of the T-bet decreased in12h and24h after injured, but was higher than SHAM group in48h and72h after injured.The ANXA1and GATA-3in Th1, Th2cells in the endotoxin injection group have not appeared the rise trend like the burns group, but gradually decreased after injured. The expression rate of the T-bet was begun significantly rising (P<0.05).The expression rates of ANXA1, GATA-3and T-bet in Th1, Th2cells were comprehensive declined after injured (P<0.05).2. PT-PCR resultThe expression quantity of the ANXA1, GATA-3and T-bet mRNA in the burns group was begun to decrease after injured. It reached minimum in24h after injured, then expressed recovery. But the ANXA1and GATA-3in all subsequent phase points were still lower than in the SHAM group, while the expression quantity of T-bet was significant higher than SHAM group in next two phase points (P<0.05). The expression quantity of the ANXA1, GATA-3and mRNA in the endotoxin injection group was in downward trend (P<0.05), while the expression quantity of T-bet gradually increased since12h after injured.The expression quantity of the ANXA1, GATA-3and T-bet mRNA in the burn sepsis group was gradually declined from12h to72h after injured (P<0.05).3. Correlation analysisIt use the correlation analysis of Pearson to research the expression rate of the ANXA1and GATA-3in Thl, Th2cells in the detection of flow cytometry and expression quantity of the ANXA1and GATA-3mRNA in peripheral blood in the PCR detection. The former correlation coefficients are0.747and0.787respectively; the latter correlation coefficient is0.862(P<0.05).Conclusion1. The Thl/Th2balance in the burns group, endotoxin injection group and burn sepsis group shifted direction of Th2. The trend was more obvious in burn sepsis group.4.2. The expression of ANXA1, GATA-3and mRNA in the burns group, endotoxin injection group and burn sepsis group was decreased, while the burns group was gradually increased since24h after injured. But the expression in the endotoxin injection group and burn sepsis was declined continually, and more obvious in burn sepsis group. The T-bet and it’s mRNA appeared increasing in the burn and endotoxin injection group, but they were declined continually in burn sepsis group.3. The expression changes of ANXA1, GATA-3and mRNA were relevant; It demonstrated that they may join the pathophysiological process of Th1/Th2balance offset towards the Th2.