Objective To investigate the mechanism of hearing loss after focal cerebral ischemia-reperfusion injury in rats.Methods A total of 60 healthy male adult SD rats were included in this study and randomly divided into 2 groups, ischemia-reperfusion(I/R) group and sham operated control, with 30 rats in each group.The rats in I/R group were operated for suture-occluded method to establish middle cerebral artery occlusion(MCAO)model, with ischemia for 60 mins followed by reperfusion for 24 hrs.The control group was only to be isolated cervical vessels, with no thread embolism inserted.The auditory brainstem response (ABR) was tested before operation and at 24 hrs post-operation respectively.At 24 hrs post-operation, we scored neurological functions, measured the changes of water content in the brain using the dry-wet weight method, and determined the infarct volume through TTC method.We also evaluated the integrity of blood-brain barrier(BBB) by viewing the exudation of Evans blue and observed the apoptosis of neurocyte by TUNEL method to conclude apoptotic index(AI).The expression of MMP-9,Claudin-5,Occludin and CX-43 were detected by Western blot.Results Compared with the sham group,the neurological function scores,the infarct volume and water content of the brain increased, with the elevated thresholds of ABR significantly and AI went up in I/R group.The expression levels of MMP-9 and CX-43 were significantly up-regulated,but the expressions of Claudin-5 and Occludin were obviously down-regulated.All of the differences above had statistical significances.Conclusion The mechanism of hearing loss after focal cerebral ischemia-reperfusion injury in rats is possibly related to MMPs activation,neurocyte apoptosis,BBB breakage and gap junction damage.

Aim To investigate the effect of curcumin on radiosensitivity of radioresistant nasopharyngeal carcinoma cell line CNE-2R and its mechanism.Methods The concentration of curcumin was screened by MTT assay.Dose-survival curves were obtained according to the colony forming test for L-Q matching and multitarget-single hitting matching,while SF2 and the correlation parameters of radiation biology were calculated.The changes of cell cycle in CNE-2R cells caused by curcumin were also tested by flow cytometry(FCM).The differential expression of genes related to cell cycle and DNA damage repair were detected by RT-qPCR.Results CNE-2R cells could not be inhibited by 10 μmol·L-1 curcumin.Dealt with 10 μmol·L-1 curcumin for 24 h,the value of α/β increased to 1 596 from 6.56;the value of SF2 decreased to 0.361 Gy from 1.93 Gy;the value of N decreased to 1.06 from 1.60;the value of D0 decreased to 2.12 from 3.27;the value of Dq decreased to 0.12 from 1.53.FCM showed that the cells in G2 phase had a significant increase and the cells in S phase had a significant decrease after dealt with 10 μmol·L-1 curcumin for 24 h.The expression of CDK4 was significantly up-regulated and GADD45g,BRCA1 were significantly down-regulated.Conclusion Curcumin radiosensitizes nasopharyngeal carcinoma cell line CNE-2R by changing cell cycle and affecting DNA damage repair through regulating the expression of CDK4,GADD45 g and BRCA1.

Objective:To analyze the differences in esophageal motility between patients with gastroesophageal reflux disease (GERD) and functional esophageal disorders by comparing the esophagogastric junction (EGJ) barrier function, esophageal body motor function and results of provocative test.Methods:From December 2016 to December 2018, the 100 patients with typical symptoms of GERD, who visited The First Aftiliated Hospifal of Zhejiang Chinese Medical Universtiy wese selected. According to the Rome Ⅳ standand and the Lyon consensus, and the results of endoscopic examination, 24 h multichannel intratuminal impedante pH (MII-pH) monitoring and esophageal high resolution manometry (HRM), the patients were divided into GERD group ( n=32), hypersensitivity(RH) group ( n=33) and functional heartburn(FH) group ( n=35). According to the results of esophageal HRM, the differences in esophageal dynamics among the groups were analyzed. T test, variance analysis and Chi square test were performed for statistical analysis. Results:The lower esophageal sphincter (LES) resting pressure of GERD group was lower than that of FH group ((19.37±7.92) mmHg vs. (25.35±12.38) mmHg (1 mmHg=0.133 kPa)); the EGJ-contractile integral of GERD group and RH group was lower than that of FH group ((20.84±21.52) mmHg·cm and (20.72±19.35) mmHg·cm vs. (35.93±36.82) mmHg·cm), and the level of distal contractile integral of GERD group was lower than that of FH group and RH group ((802.35±496.86) mmHg·s·cm vs. (1 316.84±853.92) mmHg·s·cm and (1 141.65±607.93) mmHg·s·cm), and the differences were statistically significant ( t=-2.377, -2.069, -2.149, -3.045 and -2.467, all P<0.05). There were no major motility disorders in patients of the three groups. The incidence of ineffective oesophageal motility of GERD group was higher than that of RH group and FH group (62.5%(20/32) vs. 39.4%(13/33) and 25.7%(9/35)), and the differences were statistically significant ( χ2=4.440 and 9.214, both P<0.05). Conclusions:GERD patients have abnormal esophageal motility, which is mainly manifested by reduced LES resting pressure, decreased EGJ barrier function, and abnormal peristalsis of the esophagus. Patients with RH also have decreased EGJ barrier function.

Objective To investigate the effect of improving immunity in the treatment of chronic brucellosis,and to analyze and evaluate its clinical curative effect.Methods A patient with chronic brucellosis was treated with Mongolian medicine combined with chemical drugs to enhance immunity.The clinical symptoms,serological antibodies,Brucella DNA and immune function were compared before and after treatment.The specific antibody against Brucella in serum was detected by tube agglutination test (SAT) and tiger red plate agglutination test (RBPT).Brucella DNA in serum and blood cells was detected by PCR,and the peripheral blood lymphocyte subsets were detected by flow cytometry.Immuno-luminescence technique was used to detect serum immunoglobulin and complement components.Results After treatment,the clinical symptoms such as cold back,fatigue,and joint pain disappeared completely,and the results of serum specific antibodies against Brucella were SAT 1 ∶ 50 (++)and RBPT (+) with no changes before and after treatment,and the results of cells and serum were both negative after treatment though the results of DNA detection of Brucella were cell positive and seronegative before treatment.The results of immunological function test showed that γδT cells decreased to 9.50％ after treatment compared to 14.00％ before treatment,and the percentage of monocytes and Treg cells were 5.59％ and 7.33％ after treatment,which were higher than 3.35％ and 4.72％ of before treatment,and the level of complement C3 was 0.79 g/L before treatment and 0.91 g/L after treatment that was returned to normal reference range (0.88 ～ 2.01 g/L).Conclusion The patients with chronic brucellosis can improve their clinical treatment by improving immunity.

Objective:To analyze the differences in the contraction pattern of esophageal body in patients with different types of non-cardiac chest pain (NCCP).Methods:From January 1, 2019 to December 31, 2020, 46 NCCP patients visited the First Affiliated Hospital of Zhejiang Chinese Medical University were selected. According to the Lyon consensus and Rome Ⅳ dignostic criteria, combined with the results of gastr oscopy and 24 h muitichannel intraluminal impedance combined with pH detection monitoring, 27 patients were finally included. The 27 patients were divided into functional chest pain group (12 cases) and gastroesophageal reflux disease (GERD) group (15 cases). The differences in contraction pattern of esophageal body between the two groups were analyzed according to the results of high-resolution esophageal manometry (the maximal wave amplitude of each contraction segment (S1, S2, S3), average contraction amplitude, contraction transmission time, segment lengths, distal contractile integral (DCI) and the DCI ratio of S2 to S3). Independent sample t test and chi-square test were used for statistical analysis. Results:The segment length and contraction transmission time of S3 in GERD group were shorter than those in functional chest pain group, the DCI of S3 in GERD group was lower than that in functional chest pain group, and the DCI ratio of S2 to S3 was higher than that of functional chest pain group ((5.69±0.55) cm vs. (6.61±0.99) cm, (3.45±0.49) s vs. (4.15±0.90) s, (798.88±354.70) mmHg·s·cm (1 mmHg=0.133 kPa) vs. (1 421.45±802.47) mmHg·s·cm, 0.99±0.44 vs. 0.67±0.17), and the differences were statistically significant ( t=2.682, 2.249, 2.308 and -2.616, all P<0.05). In GERD group, the transmitted segment length of S2 was longer than that of S3 ((7.02±1.40) cm vs. (5.69±0.55) cm), the contraction time of S2 of functional chest pain group was shorter than that of S3 ((3.29±0.80) s vs. (4.15±0.90) s), and the differences were statistically significant ( t=3.413 and -2.269, both P<0.05). Conclusion:High-amplitude contraction of S3 mainly occurs in patients with functional chest pain rather than GERD patients, suggesting that it may have a certain value in differential diagnosis of functional chest pain and GERD.

Cardiac Catheterization ; Ductus Arteriosus, Patent ; therapy ; Heart Septal Defects, Atrial ; therapy ; Humans ; Printing, Three-Dimensional ; Vena Cava, Inferior

This study was aimed to construct the targeting AATF shRNA eukaryotic expression vector and establish the stably transfected U937 cell lines. The sequence of AATF mRNA was obtained from GenBank. After excluding homology, three plasmid expression vectors coding shRNA targeting 228 ∼ 249, 303 ∼ 324 and 443 ∼ 464 of AATF gene sequence were synthesized. Two terminals of shRNA carried BamHI and HindIII restriction sites. The selected nucleotides were cloned into the plasmid pSilencer 3.1-H1 neo respectively, and the resultant recombinant plasmids were named as pSA-1, pSA-2, pSA-3. The sequences of the recombinant plasmids were identified by DNA sequencing. The recombinant plasmids were transfected into the cell line U937 by electroporation with Neon(TM) Transfection System. The transfected cells were persistently screened under G418 (500 mg/L), and isolated with a limited dilution for 8 weeks. The inhibition of AATF mRNA and protein expression was respectively detected by RT-PCR and Western blot. The results indicated that RNAi eukaryotic expression vectors targeting AATF had correct reading frame and nucleotide sequence. Real-time PCR revealed that AATF shRNA effectively silenced mRNA expression of AATF. Western blot analysis found that AATF shRNA obviously suppressed protein expression of AATF (P < 0.05). It is concluded that the shRNA eukaryotic expression vector has been successfully constructed which can inhibit the expression of AATF, and the establishment of stably transfected U937 cell lines provide a original route for exploring the mechanism of AATF in human Leukemia further.
Apoptosis Regulatory Proteins ; genetics ; Gene Expression ; Genetic Vectors ; Humans ; Plasmids ; RNA Interference ; RNA, Messenger ; RNA, Small Interfering ; genetics ; Repressor Proteins ; genetics ; Transfection ; U937 Cells

BACKGROUND:It is of great significance to find new diagnostic markers of the disease and molecular targets for the treatment of the disease and the alleviation of organ injury.Ferroptosis is a newly discovered form of cell death.Overactivation of ferroptosis in animal models of sepsis is associated with the activation of inflammatory response and the injury of the liver,heart,kidney and other important organs,but the relationship between ferroptosis and bloodstream infection is not very clear. OBJECTIVE:To study the changes and biological significance of ferroptosis in a mouse model of blood stream infection induced by different bacteria. METHODS:Blood stream infection models induced by gram negative bacteria Escherichia coli,Klebsiella pneumoniae and gram positive bacteria Staphylococcus aureus and Enterococcus faecalis were established in SPF-grade ICR male mice,with 42 mice in each group.The mRNA expression levels of ferroptosis marker genes transferrin receptor 1 and glutathione peroxidase 4 in the liver,myocardium and kidney were detected at 0.5,1,3,6,12,24 and 48 hours after modeling.Another 18 SPF-grade ICR male mice were selected and randomly divided into dimethyl sulfoxide(DMSO)control group,DMSO+Klebsiella pneumoniae group,and Ferrostatin-1+Klebsiella pneumoniae group,with 6 mice in each group.In the latter two groups,animal models of Klebsiella pneumoniae bloodstream infection were established by tail vein injection of Klebsiella pneumoniae suspension,and 5 mg/kg Ferrostatin-1 and an equal dose of DMSO were given intraperitoneally 1 hour prior to the modeling of bloodstream infection,respectively.Serum levels of alanine aminotransferase,aspartate aminotransferase,blood creatinine,blood urea nitrogen,phosphocreatine kinase isoenzyme,lactate dehydrogenase,and mRNA expression levels of ferroptosis marker genes in various tissues were assayed at 6 hours after modeling. RESULTS AND CONCLUSION:After bloodstream infection modeling,the mRNA expression levels of transferrin receptor 1 in the liver,myocardium and kidney of bloodstream infection mice with different bacteria increased first and then decreased;and the mRNA expression level of glutathione peroxidase 4 decreased first,then increased,and reached the peak at 6 hours after modeling.The changes in transferrin receptor 1 and glutathione peroxidase 4 mRNA levels in bloodstream infection mice induced by gram-negative bacteria were more significant than those in blood stream infection mice induced by gram-positive bacteria,especially in bloodstream infection mice induced by Klebsiella pneumoniae.At 6 hours after bloodstream infection induced by Klebsiella pneumoniae,the levels of alanine aminotransferase,aspartate aminotransferase,serum creatinine,blood urea nitrogen,creatine phosphate kinase isoenzyme,lactate dehydrogenase in mice were significantly increased.Before modeling,Ferrostatin-1 intervention significantly reduced the levels of alanine aminotransferase,aspartate aminotransferase,serum creatinine,blood urea nitrogen,creatine phosphate kinase isoenzyme,and lactate dehydrogenase.All these findings indicate that the activation of ferroptosis in bloodstream infection mice induced by different bacteria is obvious,and the activation of ferroptosis in bloodstream infection mice induced by gram-negative bacteria is more obvious.Inhibition of iron death significantly attenuates liver,myocardial,and kidney injury in the mouse model of bloodstream infection induced by Klebsiella pneumoniae.

Cholesterol is an important lipid in the body of mammals and an essential component of membrane structures. Cholesterol homeostasis is critical for the maintenance of cellular and body activities, and is mainly regulated by the balance of cholesterol biosynthesis and the exogenous cholesterol uptake. Aberrantly regulated cholesterol metabolism promotes tumor cell proliferation，survival，invasion and metastasis，and their adaptability into the tumor microenvironment. Therefore，targeting cholesterol biosynthesis and reduction of plasma cholesterol levels and cholesterol esterification will provide new strategies for cancer treatment. This review summarizes the current understanding in cholesterol homeostasis regulation and its function in the occurence and development of cancer，as well as current metabolism-targeted cancer treatments.
Animals ; Cell Proliferation ; Cholesterol ; Homeostasis ; Lipid Metabolism ; Neoplasms ; Tumor Microenvironment

Objective:To determine the association between recreational drug usage， high risk sexual behavior， and HIV infection among men who have sex with men（MSM） in Jinhua. Methods:A cross-sectional survey was conducted to recruit MSM for anonymous questionnaire survey and serological examination. We used EpiData3.1 for data entry and SPSS 19.0 for statistical analysis. Results:A total of 368 MSM were surveyed， in which the proportion of recreational drug usage was 13.3% （ 49 / 368 ）， with the HIV infection of 10.3% （38/ 368 ） and syphilis infection of 8.9% （25/368）. Risk factors associated with recreational drug usage were determined to be part-time job or being unemployed （OR=5.26； 95%CI： 2.10-13.18； P<0.001）， average monthly income ≥CNY 5 000 （OR=6.45； 95%CI： 2.87-14.61； P<0.001）， education level of high school or above （OR=1.56； 95%CI： 1.37-3.57； P=0.037）， sexual orientation being homosexual（OR=3.52； 95%CI： 1.60-7.33； P=0.002）， number of sexual partners >1 （OR=4.37； 95%CI： 1.76-10.82； P=0.001）， engaged in group sex （OR=7.90； 95%CI： 2.11-29.55； P=0.002） and previously diagnosed sexually transmitted diseases （OR=4.76； 95%CI： 1.29-17.65；P=0.019）. Conclusion:Prevalence of recreational drug usage among MSM in Jinhua is relatively low. MSM with part-time or unemployed status， monthly income ≥CNY 5 000， higher education level， homosexual orientation， multiple sexual partners， sexually transmitted diseases and group sex behavior are the risk factors associated with recreational drug usage in Jinhua city. Targeted countermeasures should be considered for intervention.