Objective To investigate the distribution of ompA gnne in 15 Chinese reference standard strains belonging to 15 serogroups of Leptospira interrogate, and to express recombinant OmpA ( rOmpA ) and to identify immunogenicity and immunoprotection of rOmpA. Methods Genomic DNAs from different leptospiral strains were extracted by phenol-chloroform method. Entire ompA gene fragments from the strains were amplified by PCR and then sequenced after T-A cloning. A prokaryotic expression system of ompA gene from L. interrogans strain 56601 was constructed, and the expression and yield of rOmpA were determined by SOS-PAGE plus Bio-Rad Agarose Image Analyser. Rabbits were immunized with rOmpA for obtaining antiserum, and immunodiffusion test was used to measure the antiserum's titer. Western blot assay was performed to determine the immunoreaetivity of rOmpA with the antiserum against rOmpA and antiserum against whole cell of L. interrogans strain 56601, while mi-croscopic agglutination test (MAT) was applied to detect the cross agglutination to the 15 L. interrogans strains. A leptospire adhering cell model and a leptospire infecting guinea pigs model were used to determine the adhesion-bloc-king effect of rOmpA antiserum and immunoprotection of rOmpA. Results All the 15 L. interrogans strains, but not L. biflexa strain Patoe Ⅰ , had sequence conserved ompA genes. The yield of rOmpA was approximate 20% of the total bacterial proteins, rOmpA could induce rabbits to produce antibody and immunodiffusion titer of the anti-serum was 1:4. Both antisera against rOmpA and against whole cell of L. interrogans strain 56601 were able to pro-duce positive Western blot signs to rOmpA, and the former offered 1 : 20-1 : 320 MAT titers to the 15 L. interrogans strains. 1: 10-1:160 dilutions of rOmpA antiserum could efficiently block L. interrogans strain 56601 adhering to J774A. 1 cells, and 100 μg and 200 μg rOmpA displayed 50.0% and 75.0% immunoprotective rates in the infee-ted guinea pigs. Conclusion ompA gene only exists in genomes of different pathogenic L. interrogans serogroups. rOmpA has relatively stronger antigenicity, cross immunoreactivity and certain immunoprotection, implying that this recombinant protein may be used as a candidate antigen for developing universal genetic engineering vaccine of L. interrogans.

OBJECTIVE:To establish the method for simultaneous determination of risperidone (RIS) and 9-hydroxyrisperi-done (9-OH-RIS) in human plasma. METHODS:After liquid-liquid extraction of plasma sample,using AF2672 as internal stan-dard(IS),LC-MS/MS was adopted. The determination was performed on XtimateTM C18 column with mobile phase consisted of ace-tonitrile-10 mmol/L ammonium acetate solution(containing 0.1％ formic acid)(37:63,V/V,pH=3.2)at the flow rate of 0.25 mL/min. The column temperature was 40 ℃,and the sample size was 6 μL. The ESI was equipped and quantitative analysis was operated in positive ion and MRM mode. The mass transition ion-pairs were followed as m/z 411.26→191.02 for RIS,m/z 427.21→206.91 for 9-OH-RIS and m/z 418.00→175.95 for IS. RESULTS:The linear range of RIS was 0.2-50.0 ng/mL(r=0.9997)and 9-OH-RIS was 1.0-200.0 ng/mL (r=0.9987). RSDs of inter-day and intra-day were all lower than 15％,and the method recoveries were 92.42％-104.81％ and 94.51％-100.57％;matrix effects were 98.33％-107.09％ and 91.05％-105.80％;extraction recoveries were 78.11％-92.62％ and 76.32％-85.09％,respectively. The plasma concentrations of RIS and 9-OH-RIS in 78 schizophrenic patients were(13.58±8.31)and(25.62±15.52)ng/mL. CONCLUSIONS:The developed method is simple,sensitive and specific,which is suitable for routine drug monitoring and acute toxic analysis in patients receiving risperidone.

Objective To observe the effect of a rotating magnetic field in preventing and treating irradiation-induced esophagitis in rats. Methods Forty female Wistar rats were randomized into 5 groups:a non-irradiated control group,an irradiation group,an amifostine treatment group ( amifostine group ),a 90 min magnetic field treatment group (90 min magnetic group) and a 120 min magnetic field treatment group ( 120 min magnetic group),with 8 rats in each group.The esophaguses of all rats except those in the control group were exposed to a single irradiation with 6 MV X-rays from a linear accelerator at a dosage of 43 Gy.Four rats in each group were randomly chosen to be observed 1 and 2 weeks after the irradiation.Blood cytokines were detected in their arterial blood.Any pathological changes of the esophagus were observed with HE staining under a light microscope at the same time. Results Irradiation-induced esophagitis was observed in the irradiation group 7 days after irradiation,with obvious exfoliation and necrosis of the esophagal epithelium mucosae.The submucosa were hyperaemic and dropsical with abundant inflammatory cell infiltration.The pathological changes of the esophagus were similar at 7 and 14 days after irradiation.However,the irradiation-induced esophagitis of rats in the amifostine group,the 90 min magnetic group and the 120 min magnetic group were relatively slighter and the blood leucocytes and neutrophis in those 3 groups were significantly lower than those in the irradiation group,while a tendency toward repair of the mucosa of the esophagus was detected.Serum TNF-α,IL-1 and IL-6 in the 90 min magnetic group and 120 min magnetic group were significantly lower than those in the irradiation group. Conclusions Both a rotating magnetic field and amifostine can help prevent and treat irradiation-induced esophagitis.Their therapeutic efficacy is similar.Exposure to a rotating magnetic field could inhibit the expression of inflammatory factors,and thus lessen the inflammatory reaction of acute irradiation-induced esophagitis.

Objective To investigate the association of HLA-DRB31*03,*04 and *11 alleles with alopecia areata(AA)in Han Nationality in East China.Methods Polymerase chain reaction-sequence specific primer(PCR-SSP)method was conducted in 158 Chinese Han patients with AA as well as in 172 healthy human controls in East China.The relationships of HLA-DRB1 polymorphism to age of onset,episode frequency,clinical course,family history,and severity of AA were evaluated.Results No significant differences were observed for the frequency of HLA DRB1*03,*11 alleles between the patients and human controls,while increased frequency of HLA-DRB1*04 was observed in patients(OR=1.99,Pc=0.01).Multiple logistic regression analysis revealed that HLA-DRB1*04 was more prevalent in patients with an onset after 16 years of age(OR=1.94,Pc=0.02),those without family history(OR=1.97,Pc=0.02),those with recurrent AA(OR=2.49,Pc=0.02),those with a clinical course of more than 1 year(OR=2.94,Pc=0.01),those with severe AA(OR=3.53,Pc=0.00)and tbose with single episode of AA(OR=1.83,Pc=0.04)in comparison with the normal human controls.Conclusion This study demonstrates that HLA-DRB1*04 allele is associated with the occurrence and clinical types of AA in Han Nationality in East China.

Objective : To examine the diagnostic and prognostic value of long non-coding RNA (lncRNA) JPX in mesothelioma, so as to provide insights into diagnosis and prognosis of mesothelioma. Methods: Patients with clinically definitive diagnosis of mesothelioma from 2015 to 2019 that were sampled from asbestos processing plants in Zhejiang Province from 2015 to 2019 were recruited in the mesothelioma group, while healthy residents without asbestos exposure or asbestos-related diseases in the same area served as controls. Participants' demographics, pathologic diagnosis and imaging features were collected, and the expression of blood lncRNA JPX was detected using lncRNA microarrays. The diagnostic value of lncRNA JPX for mesothelioma was evaluated using the receiver operating characteristic (ROC) curve, and the correlation between lncRNA JPX expression and prognosis was examined among mesothelioma patients using survival analysis. Results: There were 17 subjects in the mesothelioma group, with a mean age of (65.71±8.36) years, and 34 subjects in the controls, with a mean age of (64.24±8.70) years. LncRNA microarray detected significantly high lncRNA JPX expression in mesothelioma patients, and higher blood lncRNA JPX expression was detected in the mesothelioma group than in the control group [median (interquartile range), 1.10 (1.31) vs. 0.89 (0.54); t'=-2.300, P=0.034]. The area under the ROC curve was 0.673 (95%CI: 0.507-0.839, P=0.046), and if the cutoff was 1.759, the sensitivity and specificity were 35.3% and 100.0%, respectively. Survival analysis showed no significant difference in the survival rate of mesothelioma patients between the high lncRNA JPX expression group and the low expression group (χ2=0.212, P=0.645). Conclusions LncRNA JPX overexpression is detected in the blood of patients with mesothelioma, and lncRNA JPX expression presents a diagnostic value for mesothelioma; however, it shows little prognostic value for mesothelioma.

OBJECTIVETo detect the expression of histone deacetylase 6 (HDAC6) in laryngeal squamous cell carcinoma, and to analyze the effects of downregulation of HDAC6 expression on cell cycle, proliferation and migration of laryngeal squamous cell carcinoma cell line Hep-2 cells, and to explore their possible molecular mechanisms.

METHODSImmunohistochemistry was used to detect the expression of HDAC6 protein in 55 cases of laryngeal squamous cell carcinoma and 20 cases of normal laryngeal mucosa. HDAC6 siRNA and control siRNA were transfected into Hep-2 cells via lipofectamine 2000, and the interfering effect was analyzed using Western blotting. The effects of downregulation of HDAC6 expression on cell cycle, proliferation and migration were determined by cell counting kit-8 (CCK-8), flow cytometry and Boyden chamber, respectively. Finally, Western blotting was used to detect the expressions of cell cycle, proliferation and migration related proteins.

RESULTSThere was a high level expression of HDAC6 protein in laryngeal squamous cell carcinoma, and its expression was not related to age and sex of the patients (P > 0.05), but closely associated with the degree of histological differentiation, TNM staging and lymph node metastasis (P < 0.05). HDAC6 siRNA effectively down-regulated the expression of HDAC6 protein in laryngeal squamous cell carcinoma cell line Hep-2 cells, and downregulation of its expression obviously inhibited cell proliferation, arrested cell cycle at G(0)/G(1) phase and decreased cell migration ability in Hep-2 cells. Additionally, the downregulation of HDAC6 protein expression markedly decreased the expressions of cyclin D1, cyclin E, cdk2 and MMP-9 proteins, but increased the expressions of p21 and E-cadherin proteins.

CONCLUSIONSHDAC6 may play a pivotal role in the carcinogenesis and development of laryngeal squamous cell carcinoma. The downregulation of HDAC6 expression-mediated cell proliferation inhibition, cell cycle arrest and decreased cell migration ability may be closely associated with the decrease of cyclin D1, cyclin E, cdk2 and MMP-9 proteins and increase of p21 and E-cadherin proteins.

Adult ; Aged ; Cadherins ; metabolism ; Carcinoma, Squamous Cell ; genetics ; metabolism ; pathology ; Cell Cycle ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Cyclin D1 ; metabolism ; Cyclin E ; metabolism ; Cyclin-Dependent Kinase 2 ; metabolism ; Down-Regulation ; Female ; Histone Deacetylase 6 ; Histone Deacetylases ; genetics ; metabolism ; Humans ; Laryngeal Neoplasms ; genetics ; metabolism ; pathology ; Lymphatic Metastasis ; Male ; Matrix Metalloproteinase 9 ; metabolism ; Middle Aged ; Neoplasm Staging ; Oncogene Proteins ; metabolism ; Proto-Oncogene Proteins p21(ras) ; metabolism ; RNA, Small Interfering ; genetics ; Transfection

Objective To explore the association between tumor necrosis factor receptor associated factor 6 (TRAF6) polymorphisms and the susceptibility to sepsis. Method Haplotype tagging single nucleotide polymorphisms (htSNPs) were selected from the HapMap database. The htSNPs were genotyped in 255 patients with sepsis and 260 control subjects by the Beckman SNP stream genotyping platform. The association with susceptibility to and severity of sepsis were estimated by logistic regression with adjustment for age, sex, smoking, drinking,chronic diseases status, APACHE Ⅱ score and critical illness. Results Of 13 TRAF6 ANPs, 7 were tagged by htSNPs. Of them, 5 htSNPs (rs5030496, rs5030411, rs5030416, rs5030445 and rs3740961) were used for final genotyping analysis. Genotype frequencies of those htSNPs were conformed to the Hardy-Weinberg law in both patients and controls. There were no significant association found between the 5 htSNPs and susceptibility to sepsis.Also, there was no significant association between the TRAF6 polymorphisms and the septic shock, death from sepsis as well as organ dysfunction. Conclusions The TRAF6 gene polymorphisms might not play a major role in severity of sepsis.

Objective To investigate the prevalence and pattern of androgenetic alopecia (AGA) in Shanghai through a community-based survey. Methods A cluster sampling survey was done among the residents in Beixinjing Community, Changning District, Shanghai. All the subjects were asked to fill a questionnaire to provide their general information, including sex, age, native place, physical status, life habit, family history, etc. The diagnosis of AGA was made by dermatologists. To determine the pattern of hair loss,Norwood-Hamilton classification system and Ludwig classification system were used for male AGA and female AGA, respectively. All the data were statistically analyzed by EpiData and SPSS11.5 software. Results Totally, 7056 subjects completed the questionnaire, including 3519 males and 3537 females, and the response rate was 72.5%. AGA was diagnosed in 809 patients, consisting of 701 males aging from 19 to 91 years (mean 64.16±11.9 years) and 108 females aging from 35 to 91 years (mean 70.46±18.89 years). The standardized prevalence (SP) was 9.47% in total, 15.73% in males and 2.73% in females; the difference was significant between males and females (χ2=356.00, P<0.001). A family history of AGA was observed in 52.7% of all subjects including 391 (55.78%) males and 35 (32.41%) females. Type Ⅲ vertex involvement was the most common type in men aging from 20 to 70 years old, and type Ⅵ in those over 70 years old. Grade Ⅰ and Ⅱ predominated in female AGA. Conclusions The results of this survey indicate that the prevalence of AGA is remarkably higher in men than that in women. Furthermore, the prevalence is steadily increased with advancing age in Shanghai.

Objective To evaluate the dietary iodine intake level of population in coastal and inland areas of Zhejiang province.Methods The cluster sampling method was applied to select Zhoushan,Ningbo and Taizhou cities from Zhejiang province as coastal areas,to select Jinhua,Quzhou and Lishui cities from Zhejiang province as inland areas,and two counties(districts) were randomly selected in each chosen city,three sub-districts(towns) were randomly selected in each chosen counties (districts),then one community (village) was randomly selected in each chosen sub-districts(towns).Adult residents aged greater than 18 were selected as investigation subjects,their dietary iodine intake in the past 24 hours was investigated,and the Food composition table” was inquired to get the data of dietary iodine intake.One hundred copies of residential edible salt samples and drinking water samples in each of the selected community (village) were collected to detect salt iodine and water iodine by direct titrimetric and spectrophotometric method,respectively.Results The mean of adult residents' dietary iodine intake in Zhejiang coastal area was 267.76 μg/d,which was less than that in inland area (429.05 μg/d,t =- 6.90,P ＜0.05),and the means of adult residents' dietary iodine intake from drinking water,laver and fish(5.75,69.72,5.61 μg/d,respectively) in coastal area were higher than those in inland areas(3.25,35.27,3.43 μg/d,respectively,t =21.73,3.92,4.08,all P ＜ 0.05),however,the means of adult residents' dietary iodine intake from salt,kelp and other food (166.81,3.04,16.82 μg/d,respectively) in coastal areas were less than those in inland areas (355.15,6.14,25.81 μg/d,respectively,t =- 8.76,- 5.49,- 18.56,all P ＜ 0.05).In coastal areas,the proportion which was less than estimated average requirement of iodine(EAR,120 μg/d) was 46.48％ (1029/2214),the ratio which was higher than the maximum tolerable intake of iodine(UL,1000 μg/d) was 3.34％ (74/2214),and the average contribution rate of dietary intake of iodine in salt was 62.30％(166.81/267.76).While in inland areas,the corresponding proportions were 7.61％ (171/2246),2.80％ (63/2246) and 82.78％ (355.15/429.05),respectively.Conclusions The dietary iodine intake in Zhejiang inland areas has reached the recommended nutrient intake levels of the Chinese Nutrition Society,but there is a certain degree of insufficient iodine intake in population of the coastal areas.

OBJECTIVETo investigate the effects of chronic lead exposure on expression of autophagy-associated proteins in rat hippocampus.

METHODSSD rats were randomly divided into three groups: control group was given distilled water, lead-exposed groups were given 0.5 g/L (low-dose) or 2.0 g/L(high-dose) lead acetate solution in drinking water. The rat pups started to drink the lead content water until 60 d maturity. The lead contents in blood and brain samples were analyzed by graphite furnace atomic absorption spectrophotometry. The expressions of Beclin 1, LC3, LAMP2 and cathepsin B proteins were detected by Western blot and immunohistochemistry.

RESULTSCompared with control group, the contents of lead were significantly higher in blood and hippocampus samples in chronic lead-exposed rats (P<0.01). Western blot showed that the expression of Beclin 1 and LC3-II/LC3-I increased significantly in high dose lead-exposed group compared with control group (P<0.05 or P<0.001). The confocal laser immunostaining results demonstrated that increased immunofluorescence staining of cathepsin B in hippocampal neurons compared with control animals.

CONCLUSIONThe disturbance of autophagy-lysosome signaling molecules might be partially contribute to neurotoxicity of chronic lead exposure.

Animals ; Apoptosis Regulatory Proteins ; metabolism ; Autophagy ; drug effects ; physiology ; Beclin-1 ; Cathepsin B ; metabolism ; Chronic Disease ; Disease Models, Animal ; Female ; Hippocampus ; drug effects ; metabolism ; pathology ; Lead Poisoning ; metabolism ; pathology ; Lysosomal-Associated Membrane Protein 2 ; metabolism ; Male ; Microtubule-Associated Proteins ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects