| Part ?: clinical manifestations of preeclampsia in c57 b mice induced by placenta-derived extracellular vesicles in preeclampsia patientsObjective: to investigate the potential pathogenesis of preeclampsia in placenta-derived extracellular vesicles.Methods: By Zhong Fu Hui international peace maternal and child health care ethics committee approval,selected from October to December,2017 in China benefits the international peace maternity and child care hospital line cutting palace of preeclampsia patients and normal pregnant women in childbirth each 3 cases of placenta,the villi tissue culture in vitro of placenta prepared source extracellular vesicles,three-step ultracentrifugation after PBS suspension EVs,BCA method to quantitatively determine its protein concentration,adjust the concentration of 1 mg/ml.C57 b female randomly divided into three groups namely blank(blank)and normal group(control)group and pre-eclampsia(PE),6 mice in each group,respectively in experimental D1,D3,D5,D7 by tail vein injection of 100 ul different solution,injection of PBS blank group,control group injected with normal pregnant women sources of placenta EVs that injection of preeclampsia eclampsia group source of placenta source sex EVs(each placenta preparation of EVs are two mice injection,close observation during the tail intravenous pregnant rat spirit,diet and activity.The blood pressure and heart rate of mice in each group were measured and recorded by tail artery non-invasive blood pressure instrument for D0,D2,D4,D6 and D8 days.The urine of mice in each group for 24 hours was collected by using the mouse metabolic cage.The serum albumin and creatinine levels in the urine were detected by ELISA,and the ratio of the two was calculated.Serum of mice was collected from the epicanthic vein of D10,and serum human placental alkaline phosphatase(PLAP),endothelin-1(ET-1),interleukin-1?(IL-1?),angiotensin 2(ANGII)and other indicators were determined by ELISA.The glomerular pathological changes were observed by HE staining.Conclusion: placenta-derived EVs in preeclampsia patients can cause the typical clinical symptoms of preeclampsia in c57 b female rats,such as increased blood pressure and increased urine protein,while placenta-derived EVs in normal pregnant women cannot cause the typical symptoms of preeclampsia in c57 b female rats.Placenta-derived EVs in preeclampsia patients may induce endothelin-1 synthesis and secretion by causing vascular endothelial dysfunction in c57 b female rats,thereby causing increased blood pressure.Placenta-derived EVs in patients with preeclampsia can lead to glomerular hyperplasia in c57 b female rats,resulting in increased glomerular plasma albumin leakage and thus increased urinary protein.Placenta-derived EVs in patients with preeclampsia may stimulate immune cells to release inflammatory substances such as IL-1?,leading to increased systemic inflammatory response.Part ?: correlation between galectin14 in plasma and symptoms of preeclampsia in preeclampsia patientsObjective: to investigate the potential pathogenesis of preeclampsia in placenta-derived extracellular vesicles.Methods: the source of normal pregnant women and patients with preeclampsia source of placenta source sex cells vesicles,3 patients for LS-MS protein spectrum analysis and qualitative analysis,find out the differences in protein source(preeclampsia with the normal source without),from the differences in protein found in the placenta,unique primate animal protein,by western blot test in different sources of placenta,placenta outside source sex secretion body of content.Collection in July 2018 to October 2018 in Zhong Fu Hui international peace maternal and child health care delivery in preeclampsia patients plasma(33 cases)and normal pregnancy pregnant women plasma(35 cases),collect the basic data of two groups of pregnant women and related clinical data,using ELISA method to detect the plasma concentrations of galactose lectin 14,through statistical methods to analyze plasma galactose lectin 14 pregnant women with preeclampsia correlation between the severity of symptoms such as high blood pressure,albuminuria.Results: a total of 184 differentially expressed proteins were obtained by comparing the qualitative analysis of the protein spectra of placenta-derived EVs between the two groups.The GO analysis results indicated that these differentially expressed proteins were widely involved in the process of cell growth and differentiation,metabolism and biological regulation.Pubmed search of relevant databases found that only galectin14 was specifically expressed in the placental tissues of primates among 184 differentially expressed proteins.Western blot results indicated that the expression level of galectin14 in placenta and placenta-derived EVs of preeclampsia was significantly higher than that in normal pregnant women,and the difference was statistically significant.Plasma galectin14 concentration in preeclampsia patients was significantly higher than that in normal pregnant women,and the difference was statistically significant.The concentration of plasma galectin14 was positively correlated with the increase of blood pressure and urine protein,and negatively correlated with the concentration of plasma albumin in pregnant women and the birth weight of the fetus.The above correlation coefficients were statistically significant,and the concentration of plasma galectin14 in pregnant women was not correlated with liver and kidney function and blood coagulation function in pregnant women.The area under the ROC curve(AUC)of the diagnostic value of galectin14 in preeclampsia was 9038.Conclusion: the concentrations of placental EVs and plasma galectin14 in preeclampsia patients were significantly higher than those in normal pregnant women.Plasma galectin14 concentration in pregnant women is positively correlated with the severity of preeclampsia,and positively correlated with the severity of blood pressure,urinary protein and other symptoms,which is a potential new biomarker for the evaluation and prediction of the severity of preeclampsia.Part ?: study on the mechanism of galectin14 induced hypertensionObjective: to investigate whether galectin14 induces preeclampsia in c57 b mice and its mechanism of blood pressure elevation.Methods: e.coli protein expression system was used to synthesize recombinant galectin14.Animal experiments: C57 b female randomly divided into three groups and control group(conntrol),galactose lectin 14 group(galectin14)and galactose lectin14 caffeic acid phenethyl ester group(galectin14 + cape),6 mice in each group,respectively,in the experimental D1,D3,D5,D7 by end 100 ul different solution,given intravenously,blank group injected PBS,galectin14 1 set of injection solution(including restructuring galectin14 16 ug),The galectin14+cape group was injected with solution 2(including recombinant galectin14 16 ug and 10 umol cape),and the pregnant rats' mental state,diet and activity were closely observed during tail intravenous injection.The blood pressure and heart rate of mice in each group were measured and recorded by tail artery non-invasive blood pressure instrument for D0,D2,D4,D6 and D8 days.The urine of mice in each group for 24 hours was collected by using the mouse metabolic cage.The serum albumin and creatinine levels in the urine were detected by ELISA,and the ratio of the two was calculated.Serum of D10 inner canthus vein was collected and serum galectin14,endothelin-1(et-1),interleukin-1(IL-1?)and angiotensin 2(ANGII)were determined by ELISA.The pathological changes of glomerulus were observed by HE staining.In vitro experiment: in vitro umbilical vein endothelial cells(HUVECs),three different processing respectively:(1)different concentrations of restructuring galectin14 including 0 tendency for m L,1 * tendency for 0mmol /ml,1*10-11 mol/ml,1*10-10mol/ml,1*10-9 mol/ml,together with cell culture,some cells co-culture fixed used in immunofluorescence and nucleoprotein extracted after 30 minutes,24 hours the rest of the training to extract protein extraction;(2)Every hole in the cell to join the CAPE,the final concentration ofup to 10 umol/ml,trained after 1 hour,in adding different concentrations of restructuring galectin14 including 0mmol /ml,1*10-11 mol/ml,1*10-10 mol/ml,1*10-9 mol/ml with cell culture,some cells co-culture fixed used in immunofluorescence and nucleoprotein extracted after 30 minutes,24 hours the rest of the training to extract protein extraction?Results: animal experiment: before tail vein injection,there were no statistical differences in systolic blood pressure,diastolic blood pressure,mean arterial pressure and heart rate between the three groups.The systolic blood pressure,diastolic blood pressure and mean arterial pressure of D2 in galectin14 group were significantly higher than those in the other two groups,and the difference was statistically significant.There was no significant difference in heart rate between the three groups.The serum concentrations of et-1 and IL-1? in galectin14 group were higher than those in the other two groups,and the difference was statistically significant.There was no statistical difference in serum ANGII concentration among the three groups.The urinary protein/creatinine ratio and glomerular diameter of galectin14 group were higher than those of the other two groups,and the difference was statistically significant.There were no significant differences in urinary protein,urinary protein/creatinine ratio or glomerular diameter between the other two groups.In vitro cytological experiments: after 30 minutes of co-culture,the NFkb number of HUVECs co-cultured with galectin14 was significantly higher than that of galectin14+CAPE feeding and blank control,and the difference was statistically significant.After 24 hours of culture,the ET-1 synthesis of HUVECs co-cultured with galectin14 increased with the increase of galectin14 concentration.The ET-1synthesis of HUVECs co-cultured with galectin14+CAPE increased with the increase of galectin14 concentration.Conclusion: galectin14 can induce the typical clinical symptoms of preeclampsia such as increased blood pressure,proteinuria and systemic inflammatory response in c57 b female rats.galectin14 stimulates NF b,a transcription factor of vascular endothelium,into the nucleus,causing increased secretion of endothelial vasoconstriction factor endothelin-1 in vascular endothelial cells,thereby causing vascular systolic dysfunction and increased blood pressure.galectin14 May cause renal impairment and systemic inflammatory overreaction through the NFKB signaling pathway. |
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