Objective:To express,purify,prepare antibodies,and analyze the subcellular localization of Toxoplasma gondii thioredoxin 20(Trx20),and to provide the reference for the development of Toxoplasma gondii vaccine.Methods:Bioinformatics-related websites and software were used to perform bioinformatics analysis of the Trx20 protein;specific primers were designed to amplify the target fragment and construct the prokaryotic expression vector;the protein was expressed in vitro and purified;experimental animals were immunized to prepare antibodies;enzyme-linked immunosorbent assay(ELISA)method was used to detect the titer of the polyclonal antibodies;Western blotting method was used to verify the specificity and sensitivity of the antibodies and to determine the natural expression of the protein;immunofluorescence assay(IFA)was used to analyze the subcellular localization of the protein.Results:The bioinformatics analysis results showed that Trx20 protein was a relatively stable hydrophilic protein with a molecular formula of C2172H3412N548O616S20,containing 424 amino acids,a predicted relative molecular mass of 47 700,and a theoretical isoelectric point of 8.55;it was predicted that the protein had one signal peptide,no transmembrane region,contained one domain named"Thioredoxin like Superfamily",and had 35 phosphorylation sites,one N-glycosylation site,and 17 antigenic determinants;in the secondary structure,alpha-helices accounted for 41.51％of the total amino acids,and random coils accounted for 39.86％;the recombinant plasmids pET-28a-Trx20 and pGEX-4T-1-Trx20 were successfully constructed,and the soluble recombinant protein was expressed and purified;polyclonal antibodies were successfully prepared with a titer as high as 1:64 000,and they specifically recognized the endogenous Trx20 protein in Toxoplasma gondii;the subcellular localization results showed that Trx20 protein was widely distributed in the cytoplasm of the parasite.Conclusion:Toxoplasma gondii Trx20 protein is a secretory protein containing phosphorylation/glycosylation modification sites and a thioredoxin domain,and it is localized in the cytoplasm of the parasite.

Objective:To screen the host interaction proteins of the severe fever with thrombocytopenia syndrome virus(SFTSV)nonstructural protein(NSs)by immunoprecipitation combined with mass spectrometry analysis,to discuss the functions,subcellular localization,and biological pathways of these interaction proteins,and to provide the basis for clarifying the replication and pathogenic mechanism of SFTSV.Methods:The eukaryotic expression vectors pSFTSV-NSs-Flag(experimental group)and Flag-CMV-3(negative group)were transfected into the human embryonic kidney 293T cells,and contorl group(no treatment)was set up.The lysates of the cells in various groups were collected,and the expression and localization of SFTSV NSs in the host cells were verified by indirect immunofluorescence and Western blotting methods.The protein lysates were treated with protein A/G and immunoprecipitation was used to enrich host proteins binding to NSs.The captured interaction proteins were initially analyzed by silver staining and Coomassie brilliant blue staining to observe the differential protein bands in various groups;liquid chromatography-tandem mass spectrometry was used to obtain the information of protein sequences;the reliable proteins were retained and searched by UniProt database;Gene Ontology(GO)functional enrichment analysis,IPR,eukaryotic orthologous groups(KOGs)functional annotation,Kyoto Encyclopedia of Genes and Genomes(KEGG)signaling pathway enrichment analysis,subcellular localization,and transcription factor(TF)functional annotation were used to determine the subcellular structure,gene functions,and biological processes of the interaction proteins.Results:The immunofluorescence results showed that the SFTSV NSs expressed a single specific band at relative molecular mass 33 000 and was localized in the cytoplasm in a granular inclusion body-like manner.The silver staining and Coomassie brilliant blue staining results showed there were significant differential protein bands between experimental group and negative group.The mass spectrometry results identified 46 potential interaction proteins.The GO functional enrichment analysis,KOGs functional annotation,and KEGG signaling pathway enrichment analysis results showed that the biological pathways related to viral translation,cellular metabolism,and protein transport were enriched with a considerable number of proteins.Eight annotated proteins had intermediate filament domains.The highest percentage of subcellular localization was cytoplasmic proteins,consistent with the NSs localization site.The TF functional annotation analysis results showed one protein from the NF-Y family.Conclusion:The interaction proteins play roles in assisting the proper protein folding,participating in the cribosome translation,and forming the cytoskeleton,which may be involved in antiviral replication.These proteins can be used as candidate proteins for further study on the replication mechanism of SFTSV.

Ferroptosis is a unique iron-dependent cell death pattern,a novel death phenotype distinct from apoptosis,va-rious forms of necrosis,and autophagy.Numerous active ingredients extracted from traditional Chinese medicine have been found to exert anti-cancer effects by inducing ferroptosis in various cancers.An increasing number of studies have found that the regulation of ferroptosis can influence the sensitivity of tumor cells to drugs and even reverse drug resistance.When combined with chemo-therapy drugs such as cisplatin,5-FU and gemcitabine,some natural products enhance cancer cells'sensitivity to chemothera-peutic drugs by inducing ferroptosis.This paper mainly summarizes traditional Chinese medicine and its natural products that can exert anti-cancer effects by inducing ferroptosis,providing new insights for cancer treatment and drug resistance reversal.Addition-ally,it contributes to exploring the potential advantages of traditional Chinese medicine,thereby expanding its scope of applica-tion.

ObjectiveTo investigate the effect of Gegen Qinliantang （GGQLT）-medicated serum on free fatty acid （FFA）-induced nonalcoholic steatohepatitis （NASH） in vitro model of human hepatoma cells HepG2. MethodNASH model of HepG2 cells was established in vitro， and the cells were intervened with different volume fractions of GGQLT-medicated serum and resveratrol. Intracellular lipid deposition in each group was detected by oil red O staining， the level of reactive oxygen species （ROS） in each group were detected by flow cytometry， the levels of glutathione peroxidase （GSH-Px）， superoxide dismutase （SOD）， triglyceride （TG） and malondialdehyde （MDA） in each group were detected by kits. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to measure the mRNA expression levels of nuclear transcription factor （NF）E₂-related factor 2 （Nrf2）， heme oxygenase-1 （HO-1）， quinone oxidoreductase 1 （NQO1）， Kelch-like epichlorohydrin-associated protein-1 （Keap1）， NF-κB， thioredoxin interacting protein （TXNIP）， interleukin-1β （IL-1β） in HepG2 cells of each group. The protein expression of Nrf2， TXNIP in cells of each group was detected by Western blot. ResultFFA induced large accumulation of intracellular lipids. Compared with the normal group， the activities of GSH-Px and SOD were significantly decreased （P<0.01） and the contents of TG， ROS and MDA were significantly increased （P<0.05， P<0.01） in the model group. Compared with the model group， all GGQLT groups and resveratrol group could elevate intracellular SOD activity to different degrees （P<0.05， P<0.01） and significantly reduce the levels of intracellular ROS and MDA （P<0.05， P<0.01）， GGQLD high- and medium-dose groups and resveratrol group significantly elevated GSH-Px activity （P<0.01）， GGQLD medium- and low-dose groups and resveratrol group significantly decreased TG content （P<0.05， P<0.01）. Compared with the model group， GGQLT high- and medium-dose groups and resveratrol group could significantly upregulate the mRNA expression levels of Nrf2， HO-1 and NQO1 （P<0.01）， all GGQLT groups and resveratrol group could significantly downregulate the TXNIP protein expression level， as well as significantly downregulate the mRNA expression levels of Keap1， NF-κB （P<0.05， P<0.01）. Nrf2-siRNA transfection of cells revealed that Nrf2 expression was significantly downregulated （P<0.01） in the Nrf2-siRNA group of cells by comparing with NC-siRNA group at the corresponding dose of drugs， and the inhibitory effects of GGQLT and resveratrol on TXNIP， IL-1β were attenuated. ConclusionFFA induces the production of ROS and inflammatory factors in HepG2 cells， and GGQLT can improve the anti-inflammatory and antioxidant capacities of cells， and its mechanism may be related to the regulation of Nrf2/TXNIP signaling pathway， so as to improve NASH.

Objective To analyze the clinical efficacy of balloon occlusion of distal abdominal aorta for patients with pernicious placenta previa and placenta accreta.Methods Data of 72 patients with pernicious placenta previa and placenta accreta were retrospectively analyzed.There were 53 cases (occlusion group) reserved balloon occlusion in abdominal aorta before cesarean section,which can temporarily blocked abdominal aortic blood flow during operation.The other 19 cases (non-occlusion group) underwent cesarean section without balloon occlusion of abdominal aorta.The intraoperative,post operative situations and the birth state of newborn of the two groups were compared.Results The bleeding,blood transfusion and hysterectomy rate during the operation in occlusion group were less than those in non-occlusion group (all P＜ 0.05).Differences of the rate of postoperative transferring to intensive care unit (ICU) and the time in ICU were statistically significant between two groups (both P ＜0.05).No statistical difference of operation time,postoperative total hospital stay time and the rate of postoperative infection was found between two groups (both P＞0.05).There was no statistical difference of newborns weight and Apgar scores (5 min and 10 min after birth) between two groups (all P＞0.05).Conclusion The balloon occlusion of distal abdominal aorta in cesarean section for patients with pernicious placenta previa and placenta accreta is safe and feasible,which can effectively reduce the intraoperative bleeding,the blood transfusion and the risk of hysterectomy.

Objective To investigate the HBV reactivation status and clinic outcomes in the renal allograft recipients with inactive HBsAg carriers,and explore the preventive measures.Methods A retrospective analysis of clinical manifestation was processed in 88 cases of inactive HBsAg carriers before and after renal transplantation.Preoperative liver function in all cases was normal and serum HBsAg positive,HBV DNA＜106 copies/L.Tacrolimus (or cyclosporine A) + mycophenolate mofetil (MMF) + prednisone were given in prevention of rejection after transplantation.In 88 cases,56 cases were given nucleoside analogues (acid) for prophylactic antiviral therapy,in which 31 cases were given lamivudine (LAM) (LAM group),25 cases were given entecavir (ETV) (ETV group) ; The rest 32 cases were not given prophylactic antiviral therapy,only receiving routine liver-protecting therapy (inosine,glucurolactone) (control group).Incidence of HBV re-activation,liver function,response to treatment and the pathological changes of hepatic tissue were observed.Results During the follow-up period,the incidence of HBV reactivation in LAM group and ETV group was 45.2％ and 28.0％ respectively,significantly lower than in control group (84.4％,P＜ 0.05).In prophylactic treatment groups,HBV reactivation occurred later,liver function damage was milder,and HBV DNA load peak was lower (P＜0.05).In LAM group,HBV reactivation occurred in 14 cases,including 10 cases occurred during administration of LAM,and ETV treatment was given for about 2 months,serum HBV DNA levels in 7 cases were under detection line;in the rest 4 cases,HBV reactivation occurred in patients with treatment less than 1 year and noncompliance,who withdrew medicine blindly.After the original scheme of antiviral therapy was done,serum HBV DNA levels in 3 cases were under detection line,and the effect was not obvious in one case.In control group,HBV reactivation occurred in 27 cases.Fourteen cases therefore accepted nucleoside (acid) analogs antiviral therapy,and HBV DNA levels in 10 cases were under detection line.Histological examination revealed the liver with fibrotic cholestatic hepatitis changes in 9 patients,including 8 cases in control group,and 1 case in LAM group due to blind withdrawal of medicine.Conclus(i)on LAM and ETV prophylactic use may decrease the HBV reactivation rate in inactive HBsAg carriers after renal transplantation,reduce the severity of liver damage and the occurrence of fibrotic cholestatic hepatitis.