| Background and purposeGestational Diabetes Mellitus(GDM)is a common complication during pregnancy and is extremely harmful to the health of mothers and infants.The pathogenesis of GDM is currently unclear and is likely to be related to insulin resistance.Complement-Clq/TNF-related protein 12(CTRP12)is an adipokine,which is mainly expressed in white adipose tissue,and has a role in improving glucose metabolism,reducing insulin resistance,and reducing inflammation in adipose tissue.During pregnancy,the mother is in the process of rapid accumulation of body fat.Whether CTRP12 is related to the occurrence of gestational diabetes and the related mechanisms have not been reported.In insulin-sensitive tissues,glucose transporter 4(GLUT4)is the main glucose transporter.Under insulin stimulation,vesicles containing GLUT4 are transported from the inside of the cell to the cell membrane for glucose uptake.Soluble N-ethylmaleamide-sensitive factor attachment protein receptor(Soluble NSF Attachment Protein Receptor,SNARE)is the main protein that mediates membrane fusion.The vesicle-related proteins STX4,SNAP23 and the target membrane-related protein VAMP2 form a SNARE trimer complex,which promotes the fusion of GLUT4 vesicles with the plasma membrane and regulates the glucose transport rate.The related regulatory proteins MUNC18C and SYNIP can change their molecular conformation through insulin,affecting the formation of the above-mentioned trimeric complexes.In this study,the expression of CTRP12 and the aforementioned vesicle transporter-encoding genes in serum,placenta,and adipose tissue of GDM and PGDM patients was measured to further analyze the relationship between the adipokine CTRP12 and the incidence of gestational diabetes,and whether CTRP12 affects glucose uptake through regulating vesicle transport-related genes,aims to further elucidate the pathogenesis of GDM.Part 1 Changes and significance of serum CTRP12 and TNF-α in patients with GDMObjectives and methodsA total of 109 pregnant women undergoing perinatal health care and childbirth at the Second Affiliated Hospital of Zhengzhou University from December 2018 to June 2019 were selected.All pregnant women were routinely checked for antenatal,and 75g OGTT were checked at 24-28 weeks.According to the diagnostic criteria of GDM and PGDM,all pregnant women were divided into 34 cases of normal control group(NGT group),39 cases of gestational diabetes group(GDM group)and 36 cases of pre-pregnancy diabetes(PGDM group).In the early morning fasting state,6 mL of cubital venous blood was drawn,and centrifuged at 3000 r/min for 10 min to separate serum,took the supernatant and aliquoted,one part was used for the detection of blood biochemical indicators,and the remaining part was stored at-80℃ for testing.Record the subject’s age,gestational week,height,pre-pregnancy weight,prenatal weight,systolic blood pressure,diastolic blood pressure,and calculate BMI(weight/height2)and other general conditions.The levels of fasting blood glucose(FPG),glycated hemoglobin(HbA1c),total cholesterol(TC),triglycerides(TG),high-density lipoprotein(HDL-C),and low-density lipoprotein(LDL-C)were measured with a fully automatic biochemical analyzer.The serum CTRP12 and TNF-α levels were determined by ELISA double antibody sandwich method.SPSS 23.0 software was used to analyze and process the data,and the measurement data were expressed as mean ± standard deviation.The comparison between the three groups was performed by ANOVA one-way analysis of variance,and the pairwise comparison was performed by LSD test.Correlation analysis used Pearson correlation analysis.The difference was statistically significant at P<0.05.Results(1)Comparative analysis of general informationComparison and analysis of general data of three groups of pregnant women,there was no statistical difference in the age,gestational week,height,pre-pregnancy weight,prenatal weight,Δ body weight,pre-pregnancy BMI,prenatal BMI,ΔBMI,systolic and diastolic blood pressure among the three groups(P>0.05).(2)Comparative analysis of blood biochemical indicatorsAmong the glucose metabolism indicators,FPG and HbAlc in pregnant women with PGDM were higher than those in GDM and healthy pregnant women,and the difference was statistically significant(FPG:7.2 ± 1.1mmol/L vs.4.8±0.5mmol/L,P=0;7.2 ± 1.1mmol/L vs.4.24±0.60 mmol/L,P=0)(HbA1c:6.0 ± 0.7%vs.5.5 ±0.2%,P=0;6.0±0.7%vs.5.2 ± 0.4%,P=0).The FPG and HbAlc of GDM pregnant women were also higher than those of healthy pregnant women,and the difference was statistically significant(FPG:4.8±0.5mmol/L vs.4.24±0.60mmol/L,P=0.046;HbA1c:5.5 ± 0.2%vs.5.2±0.4%,P=0.043).Among the lipid metabolism indicators,PGDM pregnant women had higher TG levels than healthy pregnant women,and HDL-C levels were lower than healthy pregnant women.The difference was statistically significant(TG:4.1±1.3mmol/L vs.2.5±0.9mmol/L,P<0.05;HDL-C:2.0± 0.5mmol/L vs.2.3 ± 0.4mmol/L,P=0.039);GDM pregnant women had higher TG levels than healthy pregnant women,and LDL-C levels were lower than healthy pregnant women,the difference was statistically significant(TG:3.8 ± 3.9mmol/L vs.2.5 ± 0.9mmol/L,P<0.05;LDL-C:3.3 ± 0.7mmol/L vs.3.7±0.9 mmol/L,P=0.042).There were no statistical differences in the remaining indicators(P>0.05).(3)Analysis of serum CTRP12 and TNF-α levelsSerum CTRP12 level was statistically different among the three groups(P=0.038).The serum CTRP12 levels of pregnant women in the PGDM group were lower than healthy pregnant women(27.9 ± 21.8pg/mL vs.48.7 ± 34.1 pg/mL,P=0.016).Serum TNF-α levels were statistically different between the three groups(P=0.002),and serum TNF-α levels in pregnant women with PGDM were higher than those in GDM and healthy pregnant women(118.8 ± 41.2pg/mL vs.68.8±33.1 pg/mL,P=0.001;118.8 ± 41.2pg/mL vs.73.1 ± 31.4pg/mL,P=0.002).(4)Correlation analysisCorrelation analysis showed that serum CTRP12 levels of PGDM pregnant women were negatively correlated with Δbody weight and ΔBMI(r=-0.668,P=0.035;r=-0.641,P=0.046),and negatively correlated with prenatal fasting blood glucose(r=-0.430,P=0.028),no correlation with other clinical indicators.Summary(1)The decrease of serum CTRP12 level in PGDM patients may lead to insulin resistance;(2)Serum CTRP12 level may be related to weight gain during pregnancy.Part 2 Expression and significance of CTRP12 and vesicle transport-related genes in placenta and adipose tissue of patients with GDMMaterials and methodsThirty-nine GDM pregnant women(GDM group)and 30 healthy pregnant women(NGT group)who underwent perinatal health care and hospitalized cesarean delivery at the Second Affiliated Hospital of Zhengzhou University from December 2018 to June 2019 were selected as the research subjects.During the cesarean section and within half an hour after delivery,two pieces of subcutaneous fat and the central maternal surface tissue of the placenta were taken,each about 1 cm in diameter.Try to avoid fascia,blood vessels and calcification points.Real-time quantitative,PCR was used to detect CTRP12 mRNA,SYNIP mRNA,SNAP23 mRNA,MUNC18C mRNA,VAMP2 mRNA,and STX4 mRNA in placenta and adipose tissue,and the relative expression levels of genes were calculated using 2-ΔΔCT method.SPSS 23.0 software was used to analyze and process the data,and the measurement data were expressed as mean ± standard deviation.Comparison between the two groups was performed using t test.Correlation analysis used Pearson correlation analysis.The difference was statistically significant at P<0.05.Results(1)Analysis of CTRP12 mRNA and vesicle transporter-related gene expression in placentaThe relative expression level of CTRP12 mRNA in placenta tissue of GDM pregnant women was lower than that of healthy pregnant women,the difference was statistically significant(0.15±0.05 vs.1.07±0.34,P<0.01).The relative expression levels of vesicle protein-related gene SNAP23 mRNA and regulatory protein-related gene MUNC18C mRNA were also lower than those of healthy pregnant women,and the difference was statistically significant(SNAP23 mRNA:0.43±0.16 vs.1.26 ± 0.7,P=0.004;MUNC18C mRNA:0.1±0.03 vs.1.21±0.45,P<0.01).There were no statistical differences between the target membrane-associated gene VAMP2 mRNA,the vesicle protein-related gene STX4 mRNA,and the regulatory protein-related gene SYNIP mRNA between the two groups(P>0.05).(2)Analysis of CTRP12 mRNA and vesicle transporter-related gene expression in adipose tissueIn adipose tissue,the relative expression of mRNA of CTRP12 and vesicle transporter-related genes between pregnant women with GDM and healthy pregnant women was not statistically different(P>0.05).(3)Correlation analysisThe expression of CTRP12 mRNA in placental tissue of pregnant women with GDM was positively correlated with HDL-C(r=0.636,P=0.015),and had no correlation with serum CTRP12,TNF-α levels,and other serum indicators(P>0.05).The relative expression level of CTRP12 mRNA in placental tissue of pregnant women with GDM was positively correlated with SNAP23 mRNA(r=0.562,P=0.025),and there was no correlation between the expression levels of other genes in the placental tissue(P>0.05).The expression of CTRP12 mRNA in adipose tissue of pregnant women with GDM was not related to the expression of CTRP12 mRNA in placental tissue and the expression of genes related to vesicle transport in adipose tissue(P<0.05).SummaryThe expression of CTRP12 in the placental tissue of GDM patients is reduced,and the low expression of CTRP12 may reduce the formation of SNARE complex by down-regulating the expression of SNAP23,thereby reducing glucose uptake and leading to insulin resistance.ConclusionCTRP12 levels are reduced in the serum of PGDM patients and placental tissues of GDM patients and may lead to insulin resistance.The mechanism may be that the low expression of CTRP12 down-regulates the expression of vesicle-related gene SNAP23 and reduces the formation of SNARE complex,resulting in a reduction in glucose uptake. |
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