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Objective:To evaluate the role of caspase-3 in aggravation of the oxidative stress injury in stored red blood cells (sRBCs) by cardiopulmonary bypass (CPB).Methods:Eight patients with type O blood undergoing valve replacement with CPB were selected, blood samples 7 ml were collected through the central venous catheter before CPB, blood samples 20 ml were collected at 2 h after CPB, and the plasma before and after CPB was obtained after centrifugation. Eight samples of sRBCs 14 ml of blood type O stored for 7-14 days were collected and each sample was divided into A, B, C and D groups with 3.5 ml in each group. Normal saline 30 μl was added to group A and group B, 10% dimethyl sulfoxide 30 μl was added to group C, and 3.5 mmol/L Z-DEVD-fmk solution 30 μl was added to group D. The sRBCs were pretreated in a 37 ℃ water bath for 2 h in the four groups. Then group A was incubated with plasma before CPB, group B, C and D were incubated with plasma at 2 h of CPB, and four groups were incubated for 48 h in a thermostatic oscillator at 37 ℃ and 80 rpm. At 2, 24 and 48 h of incubation, the activity of caspase-3 and concentration of ATP were determined by enzyme-linked immunosorbent assay, the concentrations of glutathione (GSH) and free hemoglobin (FHb) were measured by colorimetry, and the exposure rate of cell membrane phosphatidylserine (PS) was detected by flow cytometry.Results:With the prolongation of incubation time, the activity of caspase-3, exposure rate of PS at cell membrane and concentration of FHb were gradually increased, and the concentrations of ATP and GSH were gradually decreased in the four groups ( P<0.05). Compared with group A, the activity of caspase-3 was significantly increased at each time point of incubation in group B and group C, the activity of caspase-3 was increased at 24 and 48 h of incubation in group D, and the concentration of ATP was decreased at 24 and 48 h of incubation, and the concentration of GSH was decreased and the concentration of FHb was increased at each time point of incubation in group B, group C and group D ( P<0.05). Compared with group B and group C, the activity of caspase-3 was significantly decreased, the concentrations of ATP and GSH were increased, and the exposure rate of PS at cell membrane and concentration of FHb were decreased at 24 and 48 h of incubation in group D ( P<0.05). There was no significant difference in the parameters mentioned above between group B and group C ( P>0.05). Conclusions:Caspase-3 is involved in aggravation of oxidative stress injury in sRBCs by CPB.
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Biejiajian Wan， a classical formula for liver diseases， originated from Synopsis of the Golden Chamber. It has been applied in clinical settings for more than 2 000 years. According to modern pharmacological studies， it has anti-tumor， anti-fibrosis， and immunity-enhancing effects and thus is widely used for the treatment of liver fibrosis， hepatitis， liver injury， and other diseases. In recent years， accumulating evidence has proven the efficacy of this formula in the treatment of malignant tumors， especially liver cancer. This paper summarizes relevant papers in the last 20 years and summed up the anti-liver cancer mechanisms of Biejiajian Wan as regulating biological behaviors of liver cancer cells， anti-precancerosis， inhibiting tumor angiogenesis， modulating signaling pathways， suppressing activity of relevant enzymes， and regulating immunity. Moreover， this prescription can inhibit the proliferation， invasion， and metastasis of liver cancer cells， promote apoptosis of cells， suppress tumor angiogenesis， and boost immunity. In addition， it regulates Wnt/β-catenin， interleukin-6/signal transducer and activator of transcription 3 （IL-6/STAT3）， NOD-like receptor protein 3 （NLRP3）， Janus kinase-signal transducers and activators of transcription （JAK-STAT）， Delta-like ligand 4-Notch （DLL4-Notch）， nuclear factor-κB （NF-κB）， Rho-associated kinase （Rho/ROCK）， transforming growth factor-β/Smad （TGF-β/Smad）， phosphatidylinositol 3-kinase/protein kinase B/glycogen synthase kinase-3β （PI3K/Akt/GSK-3β）， and other signaling pathways. Thus， Biejiajian Wan is confirmed to have anti-liver cancer effect based on the molecular mechanisms. According to the summary of Biejiajian Wan in anti-liver cancer treatment， Biejiajian Wan alone or in combination with other drugs can significantly alleviate the symptoms， reduce adverse reactions， prolong the survival time with definite efficacy. Thus， it is safe with no adverse reactions in long-term use， which should be further promoted in clinical application. This paper analyzed Biejiajian Wan in the treatment of liver cancer based on the molecular mechanisms and clinical studies， summarized the limitations in current research， and put forward suggestions， which can lay a basis for the future in-depth research on and clinical application of Biejiajian Wan and development of anti-tumor drugs.
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Pigs are considered as ideal donors for xenotransplantation because they have many physiological and anatomical characteristics similar to human beings. However, antibody-mediated immunity, which includes both natural and induced antibody responses, is a major challenge for the success of pig-to-primate xenotransplantation. Various genetic modification methods help to tailor pigs to be appropriate donors for xenotransplantation. In this study, we applied transcription activator-like effector nuclease (TALEN) to knock out the porcine α-1, 3-galactosyltransferase gene GGTA1, which encodes Gal epitopes that induce hyperacute immune rejection in pig-to-human xenotransplantation. Meanwhile, human leukocyte antigen-G5 gene HLA-G5, which acts as an immunosuppressive factor, was co-transfected with TALEN into porcine fetal fibroblasts. The cell colonies of GGTA1 biallelic knockout with positive transgene for HLA-G5 were chosen as nuclear donors to generate genetic modified piglets through a single round of somatic cell nuclear transfer. As a result, we successfully obtained 20 modified piglets that were positive for GGTA1 knockout (GTKO) and half of them expressed the HLA-G5 protein. Gal epitopes on the cell membrane of GTKO/HLA-G5 piglets were completely absent. Western blotting and immunofluorescence showed that HLA-G5 was expressed in the modified piglets. Functionally, the fibroblasts from the GTKO/HLA-G5 piglets showed enhanced resistance to complement-mediated lysis ability compared with those from GTKO-only or wild-type pigs. These results indicate that the GTKO/HLA-G5 pigs could be a valuable donor model to facilitate laboratory studies and clinics for xenotransplantation.
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Animals , Humans , Animals, Genetically Modified , Gene Knockout Techniques , HLA Antigens , Nuclear Transfer Techniques , Swine , Transplantation, HeterologousABSTRACT
MicroRNA is small,noncoding,single-stranded RNA,which represses gene expression by mRNA destabilization and translational inhibition.With the further study of microRNA,it is found to be related to allergic disease in children, including allergic rhinitis, atopic dermatitis, eosinophilic esophagitis,asthma,milk protein allergy,etc.In recent years,with the change of the living environment and diet structure,the incidence of children allergic disease is increasing.The emergence of microRNA provides a new diagnosis method for children allergic disease.This article summarizes the research progress of microRNA in allergic disease in children.
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Objective To construct the recombinant plasmid of PA-ⅠL and express in E.coli BL 21 (DE3), and evaluate the biological activity of recombinant protein.Methods PA-ⅠL gene was amplified by PCR using primers designed according to Pseudomonas aeruginosa genome sequences and then cloned to the vector pET -28a ( +).The recombinant plasmid was transformed into E.coli BL21(DE3) and induced to express by IPTG.The recombinant protein was purified by nickel affinity chromatography.The binding activity of recombinant PA-ⅠL with Gb3/CD77 was evaluated by flow cytometry.The function of recombinant PA-ⅠL on the binding of bacteria with host cells was evaluated by colony plate counting.Results and Conclusion The recombinant PA-ⅠL protein was highly expressed in E.coli BL21(DE3) and protein purity by SDS-PAGE analysis was high after nickel affinity chromatography .Besides, the recombinant PA-ⅠL had binding activity to Gb3/CD77 and inhibited the binding of PAO1 to host cells in a dose-dependent manner.
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Objective To explore the interaction of streptococcus suis serotype 2 recombinant suilysin ( SLY ) with platelets, and provide the theoretical basis for clinic treatment of patients infected with S.suis.Methods The nickel column affinity chromatography was used to purify the recombinant SLY.The hemolytic acivity was identified by optical density before the platelets aggregation induced by a SLY was detected by a platelet aggregometer or electron microscope and the effect of aspirin on platelets aggregation was analyzed.The impact of wild type 05ZYH33 and sly-deficient mutant strainΔSLY on platelets of mice was compared to predict the interaction of the SLY with platelets in vivo.Results and Conclusion Hemolytic activity of recombinant SLY was 2000 hemolytic units( HU) and platelets aggregation was induced at 1 μg/ml.The aggregation can be inhibited by aspirin in 5 mmol/L.SLY can also increase the volume and reduce the amount of platelets in mice.
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Objective To investigate the effect of Dihuang Guanshitong Oral Liquids on expression of E-cadherin of esophageal carcinoma rats. Methods: esophageal membranous epithelium were examined for E-Cadherin by Western blotting. Results: E-cadherin showed obvious low expression in the model group, compared with the normal group (P
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Objective: To establish the standard for the quality control of Dougenguanshitong granule.Methods: The TCL method was used to identify the Radix Ootoginseng and Dioscorea Bulbifera in the granule and the content of matrine in the granule was determined by the HPLC method.Results: The linear relationship of matrine was the range of 0.68-10.2?g(r=0.9997,n=6).The average recovery was 100.23% and RSD was 1.54%(n=6).Conclusion:The method is convenient,rapid,accurate and suitable for the quality control of Dougenguanshitong granule.
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Objective To observe the effects of all-trans retinoic acid (ATRA) and oxaliplatin (L-OHP) on the proliferation of human gastric cancer BGC-823 cells. Methods Human gastric cancer cells BGC-823 were treated with ATRA and/or L-OHP,respectively. The cell proliferative activity was assessed by MTT assay. Cell morphological changes were observed under inverted microscope while apoptosis rate and cell cycle were assayed by flow cytometry. The expressions of Bcl-2 and Survivin protein were detected by immunocytochemistry. Results The proliferation of the cells treated with ATRA was inhibited obviously and the morphology of the cells changed. The apoptosis rate of BGC-823 cells increased gradually and the effect was enhanced when ATRA was combined with L-OHP. After treatment with ATRA,the expressions of Bcl-2 and Survivin protein in BGC-823 cells were both down-regulated obviously. With the combination of ATRA and L-OHP,the expressions both further decreased. Conclusion ATRA can inhibit the proliferation of human gastric cancer BGC-823 cells,and the inhibitory effect is synergistic when ATRA is combined with L-OHP. The mechanism might be related to the down-regulation of Bcl-2 and Survivin protein expressions.