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Objective:To investigate the effects of ceftriaxone(CTX) on nuclear factor erythroid 2-related factor 2(Nrf2)/glutathione peroxidase 4(GPX4) pathway and ferroptosis in early brain injury in rats with subarachnoid hemorrhage(SAH).Methods:Forty-eight clean grade male SD rats were randomly divided into sham operation group (Sham group), SAH group, SAH+ CTX group and SAH+ CTX+ Nrf2 inhibitor group (SAH+ CTX+ ML385 group) according to the random number table with 12 rats in each group.Seven days before modeling, rats in SAH+ CTX+ ML385 group were injected intraperitoneally with ML385 (30 mg · kg -1) once a day for consecutive 7 days.And 5 days before modeling, rats in SAH+ CTX group and SAH+ CTX+ ML385 group were treated with CTX(200 mg · kg -1) by intraperitoneal injection once a day for five consecutive days.Rats in Sham group and SAH group were intraperitoneally injected with the same amount of 0.9% sodium chloride solution.After 24 hours of modeling, the neurological function score and brain tissue water content of rats in each group were measured.HE staining was used to observe the morphology of neurons in CA1 and CA3 regions of hippocampus.Prussian blue staining was used to observe the iron deposition in cerebral cortex.Spectrophotometer was used to determine the iron content, malonic dialdehyde(MDA) content, glutathione(GSH) content and GPX4 activity in cerebral cortex.Western blot was used to detect the expression levels of Nrf2 and GPX4 proteins in cerebral cortex.SPSS 23.0 was used for statistical analysis.One-way ANOVA was used to compare the mean of multiple groups of samples, and Dunnett- t test was used for further pairwise comparison between groups. Results:There was a statistically significant difference in the neurological function scores of rats in the four groups 24 hours after SAH ( F=48.40, P<0.001). The neurological function score of rats in the SAH group 24 hours after SAH was significantly lower than those in Sham group and SAH+ CTX group (both P<0.05). The brain water content of rats in the four groups 24 h after SAH was statistically significant ( F=49.61, P<0.001). The brain water content of rats in the SAH group 24 h after SAH was significantly higher than that in Sham group and SAH+ CTX group(both P<0.05). There was statistically significant differences in the number of neuronal necrosis in CA1 and CA3 regions of hippocampus in the four groups 24 hours after SAH ( F=17.44, 246.50, both P<0.001). The numbers of neuronal necrosis in CA1 and CA3 regions of hippocampus in SAH group were significantly higher than those in Sham group and SAH+ CTX group, and the numbers of neuronal necrosis in CA1 and CA3 regions of hippocampus in SAH+ CTX+ ML385 group were significantly higher than those in SAH+ CTX group (all P<0.05). Twenty-four hours after SAH, the amount of iron deposited in the cerebral cortex of rats in the four groups was statistically significant ( F=2 363.0, P<0.001). The iron deposition in the cerebral cortex of rats in the SAH group was significantly higher than those in Sham group and SAH+ CTX group (both P<0.05). There were significant differences in iron content, MDA content, GSH content and GPX4 activity in the cerebral cortex of the four groups 24 h after SAH( F=2 380.0, 1 322.0, 789.1, 815.5, all P<0.001). The content of iron and MDA in the cerebral cortex of rats in SAH group were significantly higher than those in Sham group, while the content of GSH and the activity of GPX4 were significantly lower than those in Sham group (all P<0.05). The content of iron and MDA in the cerebral cortex of rats in SAH+ CTX group were lower than those in SAH group, and the content of GSH and the activity of GPX4 were higher than those in SAH group (all P<0.05). At 24 h after SAH, the expression levels of Nrf2 and GPX4 protein in the cerebral cortex of the four groups were statistically significant ( F=888.7, 1 556.0, both P<0.001). The protein expression levels of Nrf2 (0.382±0.014) and GPX4 (0.329±0.019) in the cerebral cortex in SAH group were lower than those in Sham group ((0.746±0.009), (0.953±0.009)) (both P<0.05). The expression levels of Nrf2 (0.631±0.006) and GPX4 (0.833±0.008) protein in the cerebral cortex in the SAH+ CTX group were significantly higher than those in the SAH group (both P<0.05). The expression levels of Nrf2 (0.427±0.009) and GPX4 (0.525±0.011) protein in the cerebral cortex in SAH+ CTX+ ML385 group were significantly lower than those in SAH+ CTX group (both P<0.05). Conclusion:Ceftriaxone may inhibit ferroptosis during EBI in SAH rats by regulating Nrf2/GPX4 signal axis.
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Objective To investigate the clinico-pathological features of idiopathic membranous nephropathy (IMN ) and the expression of phospholipase A2 receptor (PLA2R ) in elderly patients. Methods A total of 109 elderly patients with IMN confirmed by renal biopsy at Wuxi People's Hospital from July 2008 to February 2015 were included.Data were retrospectively collected. Results (1)Participating patients with IMN had a mean age of (67.3 ± 5.4)years ,and 67.9% of them had hypertension and 65.1% had nephrotic syndrome.Compared with non-elderly patients ,elderly patients had a higher proportion with hypertension (67.9% vs.25.2%)(P＝0.000) ,higher systolic pressure[(143.1 ± 15.2)mmHg vs. (127.3 ± 13.3)mmHg](P＝ 0.000) ,higher diastolic pressure [(88.4 ± 10.0)mmHg vs. (80.2 ± 8.4)mmHg](P＝ 0.000) ,more severe tubulointerstitial lesions [(3.1±1.9)points vs.(2.0±1.9)points](P＝0.000),and lower eGFR[(70.9±22.9)ml·min－1· 1.73 m －2vs. (90.6 ± 27.1 ) ml·min－1·1.73 m －2] ( P ＝ 0.000 ). (2 ) There were more severe tubulointerstitial lesions[(4.7 ± 1.8)points vs. (2.4 ± 1.7)points ,2.9 ± 1.6 points](P ＝ 0.000 , 0.000)and lower eGFR[(50.4 ± 17.4)ml·min－1·1.73 m －2vs. (80.3 ± 19.7)ml·min－1·1.73 m －2, (72.3 ± 21.4)ml·min－1·1.73 m －2](P＝0.000 ,0.000)in elderly patients of pathological stage Ⅱ, compared with patients of pathological stages Ⅰ and Ⅰ-Ⅱ. (3)The rate of positive PLA2R was 82.4%.Patients with positive PLA2R had higher proteinuria[(4.5 ± 2.3)g vs. (2.9 ± 1.1)g](P＝0.042) ,lower eGFR[(66.8 ± 21.8)ml·min－1·1.73 m －2vs. (97.7 ± 16.0)ml·min－1·1.73 m －2](P＝0.000) ,and more severe tubulointerstitial lesions [(3.1 ± 2.0)points vs. (1.7 ± 1.1)points](P＝0.037)than patients with negative PLA2R. (4)Multiple regression analysis showed that PLA2R positive rate(P＝0.008) ,tubulointerstitial lesion(P＝0.000) ,and level of cholesterol(P＝0.025)were negatively correlated with eGFR (R2＝0.572). Conclusions Compared with non-elderly patients , elderly patients with IMN have poorer prognosis as a result of higher blood pressure and more severe tubulointerstitial lesions.Elderly patients with IMN of advanced pathological stages and positive PLA2R have more severe kidney injury and tubulointerstitial lesions ,resulting in poor prognosis.
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Objective To establish a time-resolved fluoroimmunoassay (TRFIA) method for detecting M-type phospholipase A2 receptor (PLA2R) antibody and to investigate the diagnostic value of serum PLA2R antibody for idiopathic membranous nephropathy (IMN).Methods The microplates coated with recombined PLA2R antigen and Eu3+-streptavidin labeled PLA2R antigen were used to establish a dual-antigen sandwich-type TRFIA for PLA2R antibody detection (anti-PLA2R-TRFIA).The serum concentrations of PLA2R antibody in 63 IMN patients (36 males,27 females,age 25-75 years) and 90 healthy volunteers (30 males,60 females,age 22-53 years) were quantitatively analyzed.Kruskal-Wallis H test and MannWhitney u test were used to analyze the data.Results The range of anti-PLA2R-TRFIA was 0-10 mg/L and the sensitivity was 5 μg/L,while ED20,ED50 and ED80 of the standard curve were (0.144±0.012),(0.707±0.029) and (3.466±0.098) mg/L,respectively.The CV of inter-and intra-assay were 4.7％ and 5.1％,respectively.The average concentration of serum PLA2R antibody in healthy volunteers was 0.455(0.320,0.593) mg/L,but in IMN patients it was 2.690(0.717,7.750) mg/L (z=-3.688,P＜0.05).Meanwhile,the serum levels of PLA2R antibody in IMN patients were significantly different between different stages and ages (x2 values:10.328,9.716,both P＜0.05).According to the receiver operating characteristic (ROC) curve,when the diagnostic cut-off was set at 0.80 mg/L for IMN detection,the sensitivity and specificity of anti-PLA2R-TRFIA were 73.0％ (46/63) and 95.6％ (86/90),respectively.Conclusions AntiPLA2R-TRFIA has been well established.This quick and easy-performance method could increase the diagnostic accuracy for IMN.
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Objective To investigate the features and correlation analysis of clinico-pathological and expression of phospholipase A2 receptor(PLA2R)in idiopathic membranous nephropathy. Methods A number of 244 patients of IMN proved by renal biopsy were recruited in Wuxi People's Hospital from July 2008 to February 2015. Data were restrospectively collected. Results In the 244 IMN patients(mean age 54.07 ± 15.22 years,130 males and 114 females),44.3% had hypertension and 62.7% had nephrotic syndrome. Compared with female patients ,male patients had more severe proteinuria and lower eGFR(P 8 g/24 h had higher level of cholesterol and blood pressure ,lower eGFR and more severe tubulointerstitial lesions(P < 0.05). Pathological stage Ⅰ and Ⅱaccounted for 98%,and there were more severe tubulointerstitial lesions and lower eGFR in the patients of stage Ⅱthan that in the patients of stageⅠandⅠ~Ⅱ(P<0.05). The positive rate of PLA2R accounted for 84.3%. Lower eGFR and more severe tubulointerstitial lesions were found in PLA2R-positive patients than those in PLA2R-neg-ative patients(P < 0.05). Multiple regression analysis showed that tubulointerstitial lesions(B =-7.253),hyper-tension ratio(B=-10.726)and the level of cholesterol(B=-2.077)had negative correlations with eGFR(P<0.01, R2=0.470). Conclusions IMN patients of male gender,grave proteinuria,high pathological stage and positive PLA2R should be treated more actively , since severe tubulointerstitial lesions and kidney injury were more common in those patients.
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Objective To investigate the effects of Wuling Powder on insulin resistance of C57BL/6J mice induced by high lipid diet, and discuss the mechanism. Methods C57BL/6J mice were randomly divided into six groups:normal group, model group, rosiglitazone group, Wuling Powder in low, middle, and high dose groups, 10 mice per group. Besides the normal group, other five groups were fed with high fat and sugar diet, with a purpose to establish insulin resistance model. Normal group and model group were given pure water. Rosiglitazone group received a gavage with rosiglitazone of 0.75 mg/kg. Wuling Power low, middle, and high groups received gavage with Wuling Power of 1.23, 3.69, 11.07 g/kg, respectively, the does volume was 0.2 mL/10 g, once a day. The weight and abdominal girth were detected every week. At the end of the sixth week, mice were given 12-hour fasting, and their eyeball were taken for blood. The body weight, length, and fat in abdomen and both kidneys were detected. Paraffin section was made with HE staining. FPG and FINS of each group were detected. ISI and IRI were calculated, and TC and TG were detected. Results Compared with the model group, Wuling Powder can significantly reduce the body weight and abdominal girth of mice (P<0.01, P<0.001), improve liver fatty degeneration, lower the FPG, FINS, TCH, TG, IRI, and increase the ISI in mice (P<0.05, P<0.01). Conclusions Wuling Powder has the effect of preventing insulin resistance of C57BL/6J mice induced by high lipid diet.
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Objective To investigate the role of fibroblast growth factor-23 (FGF23) in secondary hyperparathyroidism (SHPT). Methods (1)Serum FGF23 and intact parathyroid hormone (iPTH)from 38 maintenance hemodialysis (MHD) patients were measured by ELISA and chemiluminescence enzyme immunoassay respectively.(2) Parathyroid cells from six SHPT patients underwent parathyroidectomy with forearm autotrlansplantation were cultured for 24 h,then were induced by 0.1 mg/L FGF23.The supernatant was collected at 0.6,12,24 and 48 h respectively. The concentration of iPTH was measured by chemiluminescence enzyme immunoassay. (3)Protein expression of Klotho,FGFR1,FGFR3,GATA-3 and PCNA in parathyroid tissue from 33 SHPT eases and 3 healthy people were detected by immunohistochemistry SP and PV methods respectively. Positive cell rate and absorbance were calculated. Results (1) Serum FGF23 [(3901.85±2618.11) ng/L] was positively correlated with serum iPTH [(460.00±489.77) ng/L] in MHD patients. (2) 0.1 mg/L FGF23 suppressed iPTH secretion of parathyroid cells only at 24 h time point in vitro (P<0.05). (3) Expression of GATA-3, FGFR3, Klotho and PCNA was significantly increased and FGFRl was signiticantly decreased in parathyroid tissue of SHPT-patients as compared to healthy people. (4) Positive cell rate of GATA-3 was positively correlated with iPTH (r~2=0.1901, P=0.0425) and PCNA (r~2=0.2584, P=0.0025). Klotho was positively correlated with FGFRI and FGFR3 (r~2=0.2046, P=0.0082;r~2=0.2833, P=0.0014). PCNA was negatively correlated with FGFR1 (r~2=0.1292, P=0.0399) and positively correlated with FGFR3 (r~2=0.1226, P=0.0457). FGFR1 was negatively correlated with serum phosphate (r~2=0.2329, P=0.0044) and positively correlated with serum calcium (r~2=0.1422, P=0.0305). Conclusions FGF23 level is positively correlated with iPTH level in MHD patients. FGF23 can inhibit iPTH secretion of parathyroid cells in a weak and short way, which may be associated with the proliferation of GATA-3 positive cells and parathyroid cells, the up-regulation of FGFR3 and the down-regulation of FGFR1 expression.
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Objective:To observe the effect of Busheng Yiqi Huoxue Capsule on the lipid peroxide, lipofuscin, lipid of aged rats and on the lipid, platelet aggregation and prostaglandin of the experimental hyperlipidemic rats. Methods: Adopting the aged rats and dividing randomly into groups, the indicators of MDA. lipofuscin, lipid were determined; Adopting the hyperlipidemic rats fed by high lipid diet and dividing randomly into groups, the indicators of lipid, platelet aggregation, 6 tone PGF 1? and TXB 2 were determined. Results: Busheng Yiqi Huoxue Capsule can reduce the level of lipid peroxide in the heart, brain, liver and plasma, the level of lipofuscin in the heart and brain and the level of lipid in the aged rats ( P