This study explored the effect of the Mycobacterium tuberculosis(Mtb)PE/PPE family protein PE_PGRS37 on the growth of Mycobacterium smegmatis(Ms)and macrophage autophagy during Mtb infection.The pe_pgrs37 gene was amplified from Mtb genome through PCR,and the recombinant vector pAIN-PE_PGRS37 was successfully constructed through homologous recombi-nation.pAIN-PE_PGRS37 and pAIN were integrated into Ms through electroshock to construct pAIN-PGRS37/Ms and pAIN/Ms re-combinant bacteria.Western blotting indicated that the PE_PGRS37 protein was correctly expressed in pAIN-PE_PGRS37/Ms.The re-combinant bacteria were inoculated in 7H9/7H10 medium,and their colony morphology and growth curves were observed.No signifi-cant difference in colony morphology was observed between pAIN-PE_PGRS37/Ms and pAIN/Ms.The growth rate significantly in-creased between 10 and 16 h,and a plateau was reached at 26 h.After infection of U937 cells with pAIN-PE_PGRS37/Ms and pAIN/Ms,macrophage autophagy flow was detected with western blotting and immunofluorescence.In the pAIN-PE_PGRS37/Ms-infected group,compared with the pAIN/Ms-infected group,macrophage LC3-II and p62 protein expression was significantly up-regulated(P<0.001)and inhibited autophagosome and lysosome fusion.The intracellular survival of the recombinant bacteria was detected through colony counting,and pAIN-PE_PGRS37/Ms showed significantly greater survival in macrophages at 12 h,24 h,and 48 h than pAIN/Ms(P<0.05).Our results suggested that PE_PGRS37 protein promotes Mycobacterium survival in macrophages by blocking macro-phage autophagy flow,thus inhibiting macrophage autophagy.

This study explored the effect of the Mycobacterium tuberculosis(Mtb)PE/PPE family protein PE_PGRS37 on the growth of Mycobacterium smegmatis(Ms)and macrophage autophagy during Mtb infection.The pe_pgrs37 gene was amplified from Mtb genome through PCR,and the recombinant vector pAIN-PE_PGRS37 was successfully constructed through homologous recombi-nation.pAIN-PE_PGRS37 and pAIN were integrated into Ms through electroshock to construct pAIN-PGRS37/Ms and pAIN/Ms re-combinant bacteria.Western blotting indicated that the PE_PGRS37 protein was correctly expressed in pAIN-PE_PGRS37/Ms.The re-combinant bacteria were inoculated in 7H9/7H10 medium,and their colony morphology and growth curves were observed.No signifi-cant difference in colony morphology was observed between pAIN-PE_PGRS37/Ms and pAIN/Ms.The growth rate significantly in-creased between 10 and 16 h,and a plateau was reached at 26 h.After infection of U937 cells with pAIN-PE_PGRS37/Ms and pAIN/Ms,macrophage autophagy flow was detected with western blotting and immunofluorescence.In the pAIN-PE_PGRS37/Ms-infected group,compared with the pAIN/Ms-infected group,macrophage LC3-II and p62 protein expression was significantly up-regulated(P<0.001)and inhibited autophagosome and lysosome fusion.The intracellular survival of the recombinant bacteria was detected through colony counting,and pAIN-PE_PGRS37/Ms showed significantly greater survival in macrophages at 12 h,24 h,and 48 h than pAIN/Ms(P<0.05).Our results suggested that PE_PGRS37 protein promotes Mycobacterium survival in macrophages by blocking macro-phage autophagy flow,thus inhibiting macrophage autophagy.

Objective To investigate the correlation between the expression of long non-coding RNA lymph node metastasis-associated suppressor(LNMAS)mRNA and tight junction protein claudin 4(CLDN4)mRNA and epithelial-mesenchymal transition(EMT)in cervical cancer and the clinical significance.Methods A total of 96 patients with cervical cancer who were first diagnosed and treated at Cangzhou Maternal and Child Health Hospital from June 2018 to June 2020 were selected.Real-time fluorescent quantitative PCR was used to detect the expressions of LNMAS mRNA,CLDN4 mRNA and EMT-related genes.Pearson correlation analysis was used to analyze the correlation between LNMAS mRNA,CLDN4 mRNA and VIM mRNA,E-cad mRNA,N-cad mRNA.The impact of LNMAS mRNA and CLDN4 mRNA expression on the 3-year survival rate of cervical cancer patients was analyzed using the K-M curve.Cox regression was used to analyze prognostic factors for cervical cancer.Results Compared to adjacent tissues,the expressions of LNMAS mRNA(1.13±0.28 vs.1.97±0.37)and E-cadherin(E-cad)mRNA(1.02±0.33 vs 2.29±0.56)in cervical cancer tissues were lower(t=17.738,19.144),while CLDN4 mRNA(3.14±0.52 vs 1.19±0.28),N-cadherin(N-cad)mRNA(2.60±0.36 vs 1.02±0.33)and Vimentin(VIM)mRNA(2.84±0.46 vs 1.25±0.33)were higher(t=32.351,26.951,27.518),with significant differences(all P<0.05).LNMAS mRNA showed a positive correlation with E-cad mRNA(r=0.712,P<0.05),but a negative correlation with VIM mRNA and N-cad mRNA in cervical cancer tissues(r=-0.654,-0.589,all P<0.05).Additionally,the expression of CLDN4 mRNA was negatively correlated with E-cad mRNA(r=-0.668,P<0.05),but positively correlated with VIM mRNA and N-cad mRNA(r=0.714,0.749,all P<0.05).Compared with patients with FIGO stage ⅠA～ⅠB1 and no lymph node metastasis,patients with FIGO stage ⅠB2-ⅡA and lymph node metastasis exhibited a lower LNMAS mRNA expression and a higher CLDN4 mRNA expression with significant differences(t=12.288,10.228,13.636,11.771).Comparison Log-Rank x2 square test,the 3-year overall survival rates for high and low expression groups of LNMAS mRNA were 92.00％(46/50)and 60.87％(28/46),respectively;the 3-year overall survival rates for high and low expression groups of CLDN4 mRNA were 59.18％(29/49)and 95.74％(45/47),respectively,and the difference between the two groups were significant(log-Rank x2=11.030,15.600,all P<0.001).FIGO stage ⅠB2～ⅡA(HR=1.119,95％CI:1.148～1.830),lymph node metastasis(HR=1.442,95％CI:1.124～1.850)and CLDN4 mRNA(HR=1.637,95％CI:1.124～1.850)were risk factors affecting the prognosis of cervical cancer patients,while LNMAS mRNA(HR=0.637,95％CI:0.465～0.873)was a protective factor(all P<0.05).Conclusion The expression of LNMAS mRNA is decreased,while the expression of CLDN4 mRNA is increased in cervical cancer.The expressions of LNMAS mRNA and CLDN4 mRNA were associated with the process of EMT,which may be markers for evaluating the prognosis of patients with cervical cancer.

Objective:To establish a standardized information management system (IMS) for preserving, managing, querying, and performing statistics on biospecimens and their clinical data, which is conducive to improving the utilization of biobank.Methods:Under the premise of ensuring operating environment and data security, a database-based data logic relationship model is created and applied to the IMS to manage and analyze biospecimens and their supporting clinical information of patients enrolled in the biobank of our center.Results:To ensure the establishment of the follow-up cohort, biospecimens and clinical information of inpatients and outpatients were continuously collected in the biobank of our center. Since December 2014, more than 270 000 biospecimens from inpatient, outpatient, and scientific research have been preserved. The IMS optimized by this model efficiently completes the basic work of the biobank. At the same time, the data can be queried jointly and in batches, and then converted into a report format for statistical analysis.Conclusions:The IMS of our center is suitable for application and popularization as a construction and management model for the hospital-level biobank, which meets the daily work of the biobank and diverse research needs, and provides a convenient platform and rich resources for the development of precision medicine.

Objective To explore the strategies of constructing clinical biobank based on the real world study.Methods Biospecimens have been collected since November 2014,as well as continuingly construction of the biobank resources to match the implenmentation of real world study.Related measurements adopted including standardized ethical management,quality control,automated data management,and follow-up management.Results Biospecimens and clinical data information were collected with high quality according to standard SOPs.Sophisticated equipment and data automation management mode were used to make sure the protection of patient privacy and data security.At the same time,biospecimens and data information management system were improved gradually to fulfill the integration and sharing of biospecimen data resources.Conclusions Biorepositories are the key to translational medical research,biospecimens collected in our biobank will link with clinical data information simultaneously,which increase the resource pool that can be applied in real world study.

Objective To establish an appropriate diabetic retinopathy (DR) risk assessment model for patients with type 2 diabetes mellitus (T2DM).Methods A retrospective clinical analysis.From January 2016 to December 2017,753 T2DM patients in the Third Affiliated Hospital of Southern Medical University were analyzed retrospectively.Digital fundus photography was taken in all patients.Fasting plasma glucose (FPG),HbA1c,total bilirubin (TB),blood platelet,total cholesterol (TC),triglyceride (TG),high density lipoprotein cholesterol (HDL-c),low density lipoprotein cholesterol (LDL-c),apolipoprotein-A (apoA),apolipoprotein-B (apoB),serum creatinine,blood urea nitrogen (BUN),blood uric acid,fibrinogen (Fg),estimated glomerular filtration (eGFR) were collected.The patients were randomly assigned to model group and testify group,each had 702 patients and 51 patients respectively.Logistic regression was used to screen risk factors of DR and develop an assessment scale that can be used to predict DR.Goodness of fit was examined using the Hosmer-Lemeshow test and the area under the receiver operating characteristic (ROC) curve.Results Among 702 patients in the model group,483 patients were DR,219 patients were NDR.The scores for DR risk were duration of diabetes ≥4.5 years,4 points;total bilirubin ＜6.65 mol/L,2 points;apoA≥ 1.18 g/L,2 points;blood urea≥6.46 mmol/L,1 points;HbA1c ≥7.75％,2 points;HDL-c＜ 1.38 mmol/L,2 points;diabetic neplropathy,3 points;fibrinogen,1 point.The area under the receiver operating characteristic curve was 0.787.The logistic regression analysis showed that the risk factors independently associated with DR were duration of diabetes (β=1.272,OR=3.569,95％CI 2.283-5.578,P＜0.001),TB (β=0.744,OR=2.104,95％CI 1.404-3.152,P＜0.001,BUN (β=0.401,OR=1.494,95％CI 0.996-2.240,P=0.052),HbA1c (β=0.545,OR=1.724,95％CI 1.165-2.55,P=0.006),HDL-c (β=0.666,OR=1.986,95％CI 1.149-3.298,P=0.013),diabetic nephropathy (β=1.151,OR=3.162,95％CI2.080-4.806,P=0.013),Fg (β=0.333,OR=1.396,95％CI 0.945-2.061,P=0.094).The risk model was P=1/[1+exp-(-3.799+1.272X1+0.744X2+0.769X3+0.401X4+0.545X5+0.666X6+1.151X7+0.333X8)].X1=duration of diabetes,X2=TB,X3=apoA,X4=BUN,X5=HbA1c,X6=HDL-c,X7=diabetic nephropathy,X8=Fg.The area under the ROC curve was 0.787 and the Hosmer-Lemeshow test suggested excellent agreement (x2=10.125,df=8,P=0.256) in model group.The area under the ROC curve was 0.869 and the Hosmer-Lemeshow test suggested excellent agreement (x2=5.345,df=7,P=0.618) in model group.Conclusion The area under the ROC curve for DR was 0.787.The duration of diabetes,TB,BUN,HbAlc,HDL-c,diabetic nephropathy,apoA,Fg are the risk factors of DR in T2DM patients.

Objective To observe the efficacy and safety of citalopram in the treatment of behav-ioural and psychological symptoms of dementia(BPSD)and cognitive function of Alzheimer's disease(AD) patients.Methods From April 2015 to February 2016,80 cases of moderate Alzheimer's disease(according to the clinical dementia rating scale,CDR)with symptoms of BPSD in Qingdao Mental Health Center were collected and randomly divided into treatment group and control group.Treatment group was given citalopram (10 to 30 mg/d),the control group was given the same dose of placebo,and the patients in both group were given memantine(10 mg/bid)for 12 weeks.Simple mental state examination(MMSE)was used to measure cognitive function.Neural psychiatric questionnaire(NPI)measurement was used to evaluate BPSD and the TESS was used to assess adverse effects.Results Decreased scores of MMSE between the treatment group and the control group were respectively(0.67±0.77)and(0.26±0.68)after 12 weeks of treatment.There was significant difference in decreased scores of MMSE between the two groups(t=2.49,P=0.02).The scores of NPI in agitation/attack(t=2.986,P=0.04),apathy(t=3.144,P=0.002),indifference/dysthymia (t=6.094,P=0.000)and anxiety(t=6.496,P=0.000)showed statistically significant differences between the two groups.There were no significant difference in TESS scores(P>0.05).The most frequently adverse e-vents in the study included dizziness,headache,fatigue and nausea.QTc interval prolongation were found in participants treated with 30 mg citalopram per day in the study group.Conclusion Citalopram is an effec-tive and safe drug in the treatment of BPSD and cognitive function associated with moderate AD.

Objective To investigate the metabolites of serum samples from liver cirrhotic patients with or without minimal hepatic encephalopathy,and even overt hepatic encephalopathy,then to find out diagnostic markers for minimal hepatic encephalopathy.Methods High performance liquid chromatography-orbit trap mass spectrometry (UPLC/LTQ-Orbit trap MS) technology was applied to analyze the serum metabolites from 38 patients of liver cirrhosiswith hepatitis B and 33 healthy volunteers.Results The serum metabolites of patients with simple liver cirrhosis were different from those of patients with minimal or overt hepatic encephalopathy.The serum metabolites of patients with minimal hepatic encephalopathy was mostly similar with those of overt hepatic encephalopathy patients.Arginase,L-tyrosine,glutamic acid,two L-phenylalanine peptide,homovanillic acid,omithine,L-serine were increased in patients with minimal or overt hepatic encephalopathy,and hypoxanthine decreased in patients with minimal or overt hepatic encephalopathy patients.Conclusions The serum metabolites of patients with minimal hepatic encephalopathy are mostly similar to those of patients with overt hepatic encephalopathy.Arginase,L-tyrosine,glutamic acid,two L-phenylalanine peptide,homovanillic acid,ornithine,L-serine maybe the early metabolites biomarkers to diagnose minimal hepatic encephalopathy.Hypoxanthine is likely to be an effective complement to treat patients with hepatic encephalopathy.

ObjectiveTo study the effect of low molecular weight heparin sodium (LMWHS) therapy for exertional heat stroke (EHS) patients with pre-disseminated intravascular coagulation (pre-DIC).Methods A prospective randomized controlled trial (RCT) was conducted. Thirty-six patients with EHS with pre-DIC admitted to Department of Critical Care Medicine of 180th Hospital of Chinese PLA from April 2012 to November 2014 were divided into heparin sodium group (n = 20) and LMWHS group (n = 16) in accordance with the random number table. All patients received bundle treatment after being admitted to the hospital, including rapid cooling, fluid resuscitation, organ support (mechanical ventilation, hemopurification if necessary), supplement of pro-coagulation factors, etc. The patients in heparin sodium group were treated with continuous heparin sodium 12 500 U throughout 24 hours with intravenous pump for 5 days, and the patients in LMWHS group were given LMWHS 2 500 U subcutaneously, twice a day for 5 days.The incidence of DIC, incidence of bleeding and mortality of two groups were compared.The platelet count (PLT), prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (Fib) and D-dimer of each patient between pre and post treatment times were compared.Results No significant difference was found in the incidence of DIC and mortality between LMWHS group and heparin sodium group (31.2% vs. 30.0%,χ2 =0.007,P = 0.936; 6.2% vs. 5.0%,χ2 = 0.026,P = 0.871). Incidence of bleeding during treatment in LMWHS group was significantly lower than that in heparin sodium group (12.5% vs. 45.0%,χ2 = 4.425,P = 0.035). After treatment,PLT in both LMWHS group and heparin sodium group was significantly increased compared with that before treatment (×109/L: 140.5±17.5 vs. 110.5±16.5, 152.6±21.5 vs. 120.0±20.0, bothP< 0.05) and D-dimer was significantly decreased (mg/L: 0.5±0.1 vs. 3.2±1.2, 0.6±0.2 vs. 4.4±1.8, bothP< 0.05). APTT after treatment in heparin sodium group was significantly prolonged compared with that before treatment (s: 75.3±10.6 vs. 44.1±8.2,P< 0.05) while no change in APTT was found in LMWHS group (s: 38.6±5.5 vs. 42.1±8.4,P> 0.05). No significant difference was found in PT and Fib between pre and post treatment in all the patients.Conclusion When LMWHS was applied in EHS patients in pre-DIC stage, it could not only prevent DIC as efficiently as heparin sodium, but also results in lower incidence of bleeding. So LMWHS is safer.