| Part oneThe role of the integrin linked kinase in the pathogenesis of pre-eclampsiaObjectiveTo investigate the expression of integrin-linked kinase in endothelial progenitor cells from patients with pre-eclampsia and the relation to placenta perfusion.MethodsThirty patients with pre-eclampsia and thirty-five normal late pregnant women were investigated. The expression of ILK mRNA and protein was determined with RT-PCR and Western blotting respectively. And the angiogenesis of EPCs was examined by tube formation in vitro.Results1. The values of ILK mRNA in normal group and pre-eclampsia group were0.64±0.05and0.45±0.06; The values of ILK mRNA in mild and severe pre-eclampsia groups were0.47±0.07and0.39±0.08(P<0.05)2. The values of ILK protein in normal group and pre-eclampsia group were31.6±2.3and25.6±1.3; The values of ILK protein in mild and severe pre-eclampsia groups were25.3±1.6and20.2±1.7(P<0.05) 3. The tube formation capacity in pre-eclampsia group (135.29±6.86) was significantly reduced than in normal group (329.58±8.26)(P<0.05); The decrease was even more obvious in severe group (115.72±7.63) than in mild group (148.33±6.34)(P<0.05)4. There was positive correlation between the expression of ILK mRNA and protein and tube formation capacity in normal group,(r=0.69and0.73, P<0.05), as well as in pre-eclampsia group (r=0.67and0.72, P<0.05), in mild pre-eclampsia group (r=0.65and0.68, P <0.05), and in severe pre-eclampsia group (r=0.63and0.74, P<0.05)ConclusionThe study shows that the decrease of ILK in EPCs may be one of the possible reasons for the incidence of pre-eclampsia. Part twoIntegrin-linked kinase improves function of endothelial progenitor cells in vitroObjectiveTo investigate the effect of integrin-linked kinase (ILK) on the proliferation, migration and tube formation capacity of endothelial progenitor cells(EPCs)from patients with pre-eclampsia and the role of ILK in the onset and development of pre-eclampsia.MethodsThe experiment was divided into three groups:groups of transfection, negative control and blank control which corresponded to groups of Ad-GFP-ILK transfection, Ad-GFP transfection and no transfection. Expression of ILK was detected by real-time polymerase chain reaction and western blot analysis. Changes of cell proliferation, migration and tube formation capacity were respectively detected by CCK-8, transwell test and tube formation.Results1. The mRNA and protein levels of ILK control groun and negative control group were 0.831±0.14,0.78±0.12and0.453±0.21,0.34±0.16, respectively, which were both significantly different (P<0.05); the number of blank transfection group was0.391±0.13and0.41±0.21, which had no significant difference compared with negative control group (P>0.05).2. The integral absorbance (A) values of the ILK transfection group and negative control group were2.343±0.027and1.625±0.033respectively, which was significantly different (P<0.05); the (A) value of blank control group was1.461±0.024, which was not significantly different compared with negative group (P>0.05).3. The cell migration number of transfection group and negative control group were138.4±2.6and82.6±3.7respectively, with a statistically significant difference (P<0.05); and the number of blank control group was95.3±4.2, which was not significantly different compared with negative group (P>0.05).4. The number of endothelial-like spindle-shaped cells in transfection group and negative control group were546.1±6.3and323.2±3.3respectively, with a statistically significant difference between them (P<0.05); the number of blank control group was267.4±3.6, which was not significantly different compared with negative group (P>0.05).5. There were no tube-like structures formed when EPCs of three groups seeded on the matrigel. However, the tendency of tube-like formation in ILK group was more clear and obvious than that of blank control group.ConclusionILK is probably involved in the onset of pre-eclampsia by regulating proliferation, migration and tube formation capacity of EPCs. Part threeLocal transfection of ILK to improve placental perfusion in reduction of uterine perfusion pressure rat ObjectiveTo test the hypothesis that enhanced leval of ILK expression might improve placental vasculogenesis.Methods1. Placenta ischimia model was established by reduction of uterine perfusion pressure in pregnant rat;2.30model rat were devided into three groups and were treated with local injection of adenoviral vector expressing ILK on placenta;3. Transfection effect of placenta was observed through frozen sections under fluorescence microscope;4. Placenta weight were measured at day17of gestation;5. Expression of ILK mRNA of transfected placenta was detected by real time quantitative PCR;6. Expression of ILK protein of transfected placenta was detected by western blot;7. Placental vascular density after transfected was detected by immnun ohistochemical assay.Results1. Four dyed from anesthesia accident and infection and one preterm birth of all the30rats;2. Expression of GFP was expected in placenta by adenoviral transfected technology;3. The weight of placenta in ILK group and negative control group were (0.83±0.12g and (0.86.±0.23) g respectively, the weight of placenta in blank control group was (0.79.±0.41)g, which had no significant difference compared between three group(P>0.05);4. The values of ILK mRNA in ILK group and negative control group were respectively1.352±0.31and0.757±0.47, with a statistically significant difference between them(P<0.05); the values of ILK mRNA in blank control group was0.693±0.46, which had no significant difference compared with negative control group (P>0.05);5. The values of ILK protein in ILK group and negative control group respectively were0.93±0.45and0.48±0.32, with a statistically significant difference between them (P<0.05); the values of ILK protein in blank control group was0.54±0.41, which had no significant difference compared with negative control group (P>0.05);6. The number of placental microvessels in ILK group and negative control group were respectively78.4±5.03and64.2±3.15, with a statistically significant difference (P<0.05); the number of placental microvessels in blank control group was60.1±6.34, which had no significant difference compared with negative control group (P>0.05)ConclusionExpression of ILKmRNA and protein in placenta can be elevated efficiently through adenoviral transfection technology and ILK promotes placenta vasculogenesis in experiment reduction of uterine perfusion pressure rat.
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