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OBJECTIVE@#To investigate the effect of estradiol (E2)/estrogen receptor 1 (ESR1) on the proliferation of human chondrocytes and explore the molecular mechanism.@*METHODS@#The Ad-Easy adenovirus packaging system was used to construct and package the ESR1-overexpressing adenovirus Ad-ESR1. Western blotting and qPCR were used to detect the expression of ESR1 protein and mRNA in human chondrocyte C28I2 cells. In the cells treated with different adenoviruses, the effects of E2 were tested on the expressions of proteins related with cell autophagy and apoptosis and the phosphorylation of ERK signaling pathway using Western blotting. Immunofluorescence assay was used to observe the intracellular autophagic flow, flow cytometry was performed to analyze the cell apoptosis rate and the cell cycle changes, and qPCR was used to detect the expressions of PCNA, cyclin B1 and cyclin D1 mRNAs. The inhibitory effect of the specific inhibitor of ERK on the expressions of autophagy- and apoptosis-related genes at both the protein and mRNA levels were detected using Western blotting and qPCR.@*RESULTS@#Transfection with the recombinant adenovirus overexpressing ESR1 and E2 treatment of C28I2 cells significantly enhanced the expressions of autophagy-related proteins LC3, ATG7, promoted the colocalization of LC3 and LAMP1 in the cytoplasm, increased the expressions of the proliferation-related marker genes PCNA, cyclin B1 and cyclin D1, and supressed the expressions of cleaved caspase-3, caspase-12 and pERK. RNA interference of ESR1 obviously lowered the expression levels of autophagy-related proteins in C28I2 cells, causing also suppression of the autophagic flow, increments of the expressions of apoptosis-related proteins and pERK, and down-regulated the expressions of the proliferation marker genes. Blocking ERK activation with the ERK inhibitor obviously inhibited the effects of E2/ESR1 on autophagy, proliferationrelated gene expressions and cell apoptosis.@*CONCLUSIONS@#The targeted binding of E2 with ESR1 promotes the proliferation of human chondrocytes possibly by inhibiting the activation of ERK signaling pathway to promote cell autophagy and induce cell apoptosis.
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Humans , Adenoviridae , Metabolism , Apoptosis , Autophagy , Autophagy-Related Protein 7 , Metabolism , Cell Line , Cell Proliferation , Chondrocytes , Cell Biology , Metabolism , Estradiol , Metabolism , Estrogen Receptor alpha , Metabolism , Lysosome-Associated Membrane Glycoproteins , Metabolism , MAP Kinase Signaling System , Microtubule-Associated Proteins , Metabolism , TransfectionABSTRACT
Objective To evaluate the role of mitochondrial ATP sensitive potassium ( mito-KATP ) channel in dexmedetomidine-induced inhibition of subarachnoid hemorrhage ( SAH )-caused programmed cell death ( PCD) in cardiomyocytes of rats. Methods On hundred and twenty clean-grade healthy male Sprague-Dawley rats, aged 9-10 weeks, weighing 350-400 g, were divided into 5 groups ( n=24 each) using a random number table method: sham operation group ( group Sham ) , SAH group, SAH plus dexmedetomidine group ( group SD) , 5-HD plus SAH and dexmedetomidine group ( group HSD) and 5-HD plus SAH group ( group HS) . The rats were subjected to SAH by intracranial vascular puncture after being anesthetized with pentobarbital sodium. Dexmedetomidine 5 μg∕kg was infused for 10 min via the jugular vein starting from the time point after intracranial vascular puncture, followed by a continuous infusion of 5μg·kg-1 ·h-1 for 1 h in SD and HSD groups. 5-HD 30 mg∕kg was intraperitoneally injected at 1 h before intracranial vascular puncture in HSD and HS groups. Blood samples were collected from the abdominal aor-ta at 24 h after intracranial vascular puncture for determination of serum cardiac troponin I ( cTnI) concen-trations. The animals were then sacrificed, and myocardial specimens were collected for determination of PCD rate ( by TUNEL) , reactive oxygen species ( ROS) activity ( by DCFH-DA assay) , and expression of cleaved caspase-3, cleaved caspase-1 and interleukin-1beta ( IL-1β) ( by Western blot) . Results Com-pared with group Sham, the serum concentrations of cTnI, PCD rate and ROS activity were significantly in-creased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas up-regulated in SAH, SD, HSD and HS groups ( P<0. 05) . Compared with group SAH, the serum concentrations of cTnI, PCD rate and ROS activity were significantly decreased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas down-regulated in group SD, and the serum concentrations of cTnI, PCD rate and ROS activity were significantly increased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas up-regulated in group HS ( P<0. 05) . Compared with group SD, the serum concentrations of cT-nI, PCD rate and ROS activity were significantly increased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1β was up-regulated in group HSD ( P<0. 05) . Compared with group HSD, the serum concentrations of cTnI, PCD rate and ROS activity were significantly increased, and the expression of cleaved caspase-3, cleaved caspase-1 and IL-1βwas up-regulated in group HS ( P<0. 05) . Conclusion mito-KATP channel is involved in dexmedetomidine-induced inhibition of PCD in cardiomyocytes of rats with SAH.
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Objective To discuss the agreement between pulse pressure variation of radial artery and pulse pressure variation of dorsal pedalartery in neurosurgery.Methods Twenty-five patients undergoing selective craniotomy under general anesthesia were enrolled.The following data were monitored and recorded respectively after tracheal intubation general anesthesia under different time:radial artery pulse pressure variability (PPV1) and dorsalis pedis pulse pressure variation (PPV2).Tidal volume was set to 8 ml/kg.Bland-Altman plots were created to assess agreement between PPV1 and PPV2.Results The mean differences and the limits of agreement between PPV1 and PPV2 are 20 min after induction of anesthesia 0.5% (-1.9%-2.8%), boneless flap instantly-0.5% (-3.8%-2.9%), Cut the dura mater instantly-0.1% (-3.2%-3.0%), and bone flap 0.1% (-2.4%-2.6%).Conclusion Dorsal pedal artery pulse pressure variation in neurosurgery craniotomy has certain guiding significance to the monitoring and management.
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Objective To evaluate the role of Toll-like receptor 4 (TLR4) signaling pathway in sevoflurane anesthesia-induced cognitive dysfunction in aged rats.Methods Twenty-seven SPF healthy male Sprague-Dawley rats,aged 20 months,weighing 550-750 g,were divided into 3 groups (n =9 each) using a random number table:control group (C group),sevoflurane anesthesia group (S group) and TLR4 antagonist plus sevoflurane anesthesia group (TS group).TLR4 monoclonal antibody 30 μl was injected into the lateral cerebral ventricle in group TS,and the equal volume of serum containing no antibody was injected into the lateral cerebral ventricle in C and S groups.At 10 min after completion of injection,S and TS groups inhaled the mixture of 4％ sevoflurane and 30％ oxygen for 6 h,and group C only inhaled the mixture of air and oxygen.Morris water maze test was performed at 24 h after the end of sevoflurane anesthesia.The animals were sacrificed after completion of Morris water maze test,brains were removed and hippocampi were isolated for determination of nerve cell apoptosis (using TUNEL) and expression of activated caspase-3 (using immunofluorescent staining).Nerve cell apoptosis rate was calculated.The expression of high-mobility group box 1 protein (HMGB1) mRNA in hippocampi was measured by Northern blot assay at 6 h after the end of sevoflurane anesthesia.The expression of amyloid precursor protein (APP) and amyloid beta protein (Aβ) in hippocampi was assessed by Western blot at 24 h after the end of sevoflurane anesthesia.Results Compared with C group,the escape latency was significantly prolonged,nerve cell apoptosis rate was increased,the expression of activated caspase-3,HMGB1 mRNA,APP and Aβ was up-regulated in group S,and nerve cell apoptosis rate was increased,the expression of activated caspase-3,HMGB1 mRNA,APP and Aβ was up-regulated (P＜0.05),and no significant change was found in the escape latency in group TS (P＞0.05).Compared with S group,the escape latency was significantly shortened,nerve cell apoptosis rate was decreased,and the expression of activated caspase-3,HMGB1 mRNA,APP and Aβ was down-regulated in group TS (P＜0.05).Conclusion Activation of TLR4 signaling pathway is involved in the mechanism of sevoflurane anesthesia-induced cognitive dysfunction in aged rats.
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Objective:To evaluate the implementation effects of critical illness insurance of New Cooperative Medical System(NCMS) on the occurrence rate and economic burden of major disease expenditure.Methods:Based on the peasant household data of China Family Panel Studies(CFPS) in 2014.the two-part model was applied to analyze the changes in major disease occurrence and burden after the implement of insurance.Results:NCMS critical illness insurance did not reduce the occurrence of critical disease expenditure,but signally cut down the economic burden of serious illness peasants in central and eastern China.Conclusion:The implementation effect of NCMS critical illness insurance was well in central and eastern China,but was poor in western China;the prevention and health care system of NCMS should be built,while the implementation plans and compensation level of critical illness insurance should be improved in western region.
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Objective To investigate the effect of sterol regulating element binding protein (SREBP1c) and its ac-tive form (SREBP1cm) on human protein kinase R-like endoplasmic reticulum kinase (PERK).Methods Re-porter victors of PERK promoter and its truncations were constructed with pGL 3-basic and co-transfected with internal reference pRL-SV40 into cell and luciferase activity was detected .pcDNA3.1 ( -)-SREBP1c or pcDNA3.1 ( -)-SREBP1cm was co-transfected with PERK promoter transcriptional activity core regions and the detection of dual -lu-ciferase reporter gene was used to analyze the regulation of SREBP 1c/1cm on PERK promoter transcriptional activity . The expression level of PERK mRNA and protein were detected by RT-PCR and Western blot .Results PERK pro-moter and truncations were successfully constructed into pGL 3-basic, and PERK promoter core area of transcription-al activity had determined;Dual-luciferase report gene showed that SREBP 1c inhibited PERK promoter transcrip-tional activity and SREBP1cm promoted PERK promoter transcriptional activity .RT-PCR and Western blot showed that SREBP1c decreased PERK mRNA and protein expression , but SREBP1cm increased PERK mRNA and protein expression, which was consistent with the detection of dual-luciferase report gene .Conclusions SREBP1c and SREBP1cm have a opposite regulation effect on PERK promoter transcriptional activity and its expression .
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Objective To explore the effect of erythropoietin (EPO) attenuating apoptosis in old rat hippocampal neuronal cells exposed to sevoflurane and the role of toll like receptor 4.Methods Twenty months old SD rats,male,550-750 g,in accordance with the random number table,were divided into 3 groups (n =9):control group (group C),sevoflurane treatment (group S),and sevoflurane plus EPO treatment (group ES).The rats in group S and ES were subjected to inhale 4％ sevoflurane for 6 h,but the rats in group C were inhaled air-oxygen only.The rats in group ES were injected with EPO into caudal vein at 24 h,48 h,and 72 h after sevoflurane exposure.The cognitive ability was assessed by Morris water maze test;the effects of hippocampal apoptosis were assessed by TUNEL assays;the expressions of TLR4 mRNA was measured by RT-PCR assay;mitochondrial membrane potential (MMP) was assessed by JC-1 fluorescence;the expressions of APP and Aβ were assessed by western blot.Results Compared with group C,there were significant increases of escape latency period,neuronal apoptosis,TLR4 mRNA,and APP and Aβ expression,but a decrease of MMP in group S (P＜0.05).Compared with group S,there were significant decreases of escape latency period,neuronal apoptosis,TLR4 mRNA,and APP and Aβ expression,but a increase of MMP in group ES (P＜0.05).Conclusion The attenuation of rat hippocampal neuronal apoptosis induced by EPO could be associated with inhibition of TLR4,improvement of MMP,as well as inhibition of APP and Aβ activity.
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[ABSTRACT]OBJECTIVETo investigate the phoenix roebelenii pollen as the allergen of allergic rhinitis in Hainan Province and provide guidance for prevention and treatment of allergic rhinitits.METHODSA total of 2054 patients with allergic rhinitis were tested with the allergen of phoenix roebelenii pollen by skin prick test, and then choose 30 positive cases to give the nasal mucosa provocation test. RESULTSThe total positive rate of allergen of phoenix roebelenii pollen by skin prick test was 67.38% (1384/2054). The 30 cases with positive skin prick test were all positive in nasal mucosa provocation test and the cases in control group were all negative.CONCLUSIONPhoenix roebelenii pollen is an important allergen in Hainan Province. There is a correlation between skin prick test and nasal mucosa provocation test. The allergen skin prick test can provide clue for the patients to avoid the pathogenic allergens and for the specific immunotherapy.
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RET gene is an important oncogene,which is closely associated with the development of various types of human tumors. The mainly mechanisms of RET gene associated tumor are mutation and over-expression of wild type. Activated RET protein participates in the proliferation,apoptosis,metastasis through some signal pathways and influences the tumorigenesis and development.