The Study In The Effect Of TLR4 Acetylation On Pathogenesis Of Gestional Diabetes Mellitus

Background:Gestational diabetes mellitus(GDM) are common complications during pregnancy, which bring great harm to the health of pregnant women and the fetus,including infection, polyhydramnios, macrosomia, fetal growth restriction, fetal malformation rate increase, etc. Now insulin resistance(IR) and impaired pancreatic islet function which could lead to insufficient insulin secretion are considered to be the reasons of GDM. The chronic low-grade inflammation in the gravidabodyinvolved in the formation of islet function damage and IR.In recent years,the role of inflammation on the pathogenesis of GDM has become a hot research topic.In 2006, MBL gene mutation detection was applied to complete the preliminary study of the pathogenesis of GDM inflammation and the study indicated that thepoint mutation of the 54 th codon could reduce theconcentration of MBL in serum, low MBL concentration might be involved to the pathogenesis of GDM, and then we discovered that the activation of LPS- TLR4- NF-κB pathwaymediated the release of inflammatory cytokines which prompted the pathogenesis of GDMfrom the study of TLR4 m RNA, NF-κB m RNA,expression of inflammatory factors and the corresponding study.Objective:The expression of TLR4 acetylation in the peripheral blood mononuclear cells(PBMC)in normal pregnant women and pregnant women with GDM was studied on the basis of previous research which was a further research on the inflammatory pathogenesis of GDM from the start point of LPS- TLR4- NF-κB pathway.Methods:15 mlvenous blood in the morning was extracted from normal pregnantwomen and GDM pregnant women, 30 cases were selected for each type. Ficoll lymphocyte separation medium was centrifuged by density gradient centrifugation to separate PBMC and serum was collected to detect the content of bacterial endotoxin. 1 ug/ml lipopolysaccharide(LPS) was added into PBMC culturing medium to intervene the culture after a 48 h incubation for PBMC, and inflammationmodel was established.Cells and supernatant were harvested after 24 h intervention cultivation. Immune sedimentation and Western blot were used to detect the expression of TLR4 acetylation.Western blot and ELISA were used to measure theexpression of NF-κB p65 and the content of inflammatory factors TNF-α, IL-1, IL-10 in thesupernatant, respectively.Results:1. Serum levels of LPS(0.92±0.03 IU/ml) in pregnant women with GDMwere higher than that in normal pregnant women(0.53 ±0.02IU/ml), difference had statistic significance(P < 0.05).2. TLR4 acetylation was detected in peripheral blood mononuclear cells in pregnant women with GDM,but was negative in normal pregnant women.3. After the intervention cultivation with LPS,inflammation model was established.The increased expression of NF-κB p65 and TLR4 acetylation were observed in normal +LPS group and GDM + LPS group, and the latterwas significantly higher than GDM group and normal + LPS group. The protein expression of NF-κB p65 and TLR4 acetylation were positively correlated(r GDM group = 0.901, r normal + LPS group = 0.870, r GDM +LPS group= 0.942).4.The difference of expression levels of inflammatory markers in normal group, GDM group, normal + LPS group and GDM + LPS group was statistically significant.Corresponding to these groups, figures were displayed as follow.TNF-alphaexpression: 45.49±0.81, 55.97±2.84, 52.48±1.39 and 62.55±0.87, respectively;The expression of IL-1: 53.82±5.13,75.92±1.18,73.60±0.99 and89.08±2.93, respectively;The expression of IL-1:21.01±0.51,31.42±1.52,28.77±0.63 and 39.51±3.81,respectively.Conclusion:LPS is the ligand of TLR4,regulating the extent of TLR4 acetylation. a The activation of LPS- TLR4- NF-κB pathway was influenced by TLR4 acetylation which might promote the inflammatory cytokines release in downstream of the pathways, which contributed to the development of GDM.