To construct a secretory-expression vector of antimicrobial peptide Bactenecin 7(Bac7),and identify the secretory-expression product in L.lactis MG1363 and its bioactivity.The splicing primers of regulation elements and Bac7 gene,which designed according to codon usage preferences of L.lactis MG1363,were chemically synthesized,and the overlap-extension PCR method was used to splice the full length of Bac7 gene.Then the Bac7 gene was linked to expression vector pMG36e to construct pMG36e/Bac7 vector,and pMG36e/Bac7 was transformed into L.lactis MG1363 by electrophoration.RT-PCR and Western blot assays were applied to investigate the expression of the Bac7 gene in L.lactis,and bioactivity of Bac7 in culture supernatant of L.lactis was tested with plate-diffusion method.The results showed that the Bac7 gene and its regulation elements was amplified and cloned in the vector pMG36e successfully,The secretory-expressed Bac7 in L.lactis MG1363 harboring pMG36e/Bac7 was identified by Western blot,and it had high bacteriostatic activity against E.coli.These results indicate that the recombinant L.lactis MG1363 could express bioactive Bac7,which lays a foundation for further study of oral administration of a Bac7-secreting L.lactis to treat intestinal bacteria infection.

The effect of acupuncture cooperated with low-frequency repetitive transcranial magnetic stimulation (rTMS) on chronic insomnia was explored.Seventy-eight patients with chronic insomnia were randomly allocated into two groups:treatment group and control group.In the treatment group,the patients received acupuncture combined with rTMS treatment,and those in the control group were given acupuncture cooperated with sham rTMS treatment,3 days per week for 4 weeks.Before and after treatment,the primary outcomes including the scores on Insomnia Severity Index (ISI) and Pittsburgh Sleep Quality Index (PSQI) and the secondary outcomes including total sleep time (TST),sleep onset latency (SOL),wake after sleep onset (WASO),sleep efficiency (SE％) recorded by sleeping diary and actigraphy were observed in both groups.Seventy-five participants finished the study (38 in treatment group and 37 in control group respectively).After treatment,the scores in the two groups were improved significantly,more significantly in the treatment group than in the control group.It can be inferred that acupuncture cooperated with rTMS can effectively improve sleep quality,enhance the quality of life of patients and has less side effects.

OBJECTIVETo investigate GNE gene mutations in 5 Chinese patients with distal myopathy with rimmed vacuoles (DMRV).

METHODSFive patients with typical clinical and pathological features of DMRV were studied. All the 11 coding exons and the flanking intron sequences of GNE gene were amplified by PCR and sequenced. Four family members of case 5 were also examined for GNE gene mutations.

RESULTSAll the patients were identified to have different GNE gene mutations: Cases 1-4 had complex heterozygous mutations and case 5 had homozygous mutation. Six reported mutations had been identified, including 1 nonsense mutation (p.R8X) and 5 missense mutations (p.D176V, p.I298T, p.A591T, P.A631V, and p.V696M). A novel mutation (c.317T>C, p.I106T) was identified in case 2.

CONCLUSIONThis is the first report of p.R8X, p.I298T, p.A591T and p.V696M mutations in GNE gene in Chinese population, and a novel mutation p.I106T was identified. These findings further expand the clinical and genetic spectrum of DMRV in China.

Adult ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; DNA Mutational Analysis ; Distal Myopathies ; enzymology ; genetics ; Female ; Humans ; Male ; Molecular Sequence Data ; Multienzyme Complexes ; genetics ; Mutation ; genetics ; Mutation, Missense ; genetics ; Young Adult

OBJECTIVETo explore the linkage of the polymorphism at D17S1878 site with susceptible gene of essential hypertension.

METHODSForty-five pedigrees from the high prevalence region of the hypertension were collected. The polymorphism of D17S1878 site was genotyped with genetic analyzer and gene-scan software. Discordant sib pair analysis and affected sib pair analysis were used in linkage analysis.

RESULTSThere were significant differences in the age, male, alcohol-consuming, over-salt intake, average systolic pressure, average diastolic pressure, waist-to-hip-ratio, total cholesterol, high-density lipoprotein, and low density lipoprotein between the hypertensive sib and the normotensive sib (P < 0.05). There were eleven alleles at D17S1878 site, and the allele frequency was significantly different between the hypertensive and normotensive sibs (P < 0.05). Forty-three pedigrees were analyzed with affected sib pair analysis (t = 3.05, P < 0.05).

CONCLUSIONThe polymorphism of D17S1878 may be linked with susceptible genes of essential hypertension.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Case-Control Studies ; Female ; Genetic Linkage ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Hypertension ; genetics ; Male ; Microsatellite Repeats ; genetics ; Middle Aged ; Phenotype ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Potassium Channels, Inwardly Rectifying ; genetics

BACKGROUNDMounting evidence suggests that tumors are histologically heterogeneous and are maintained by a small population of tumor cells termed cancer stem cells. CD133 has been identified as a candidate marker of cancer stem cells in laryngeal carcinoma. This study aimed to analyze the chemoresistance of CD133(+) cancer stem cells.

METHODSThe response of Hep-2 cells to different chemotherapeutic agents was investigated and the expression of CD133 was studied. Fluorescence-activated cell sorting analysis was used to identify CD133, and the CD133(+) subset of cells was separated and analyzed in colony formation assays, cell invasion assays, chemotherapy resistance studies, and analyzed for the expression of the drug resistance gene ABCG2.

RESULTSAbout 1% - 2% of Hep-2 cells were CD133(+) cells, and the CD133(+) proportion was enriched by chemotherapy. CD133(+) cancer stem cells exhibited higher potential for clonogenicity and invasion, and were more resistant to chemotherapy. This resistance was correlated with higher expression of ABCG2.

CONCLUSIONSThis study suggested that CD133(+) cancer stem cells are more resistant to chemotherapy. The expression of ABCG2 could be partially responsible for this. Targeting this small population of CD133(+) cancer stem cells could be a strategy to develop more effective treatments for laryngeal carcinoma.

AC133 Antigen ; ATP Binding Cassette Transporter, Sub-Family G, Member 2 ; ATP-Binding Cassette Transporters ; genetics ; metabolism ; Antigens, CD ; genetics ; metabolism ; Antineoplastic Agents ; pharmacology ; Blotting, Western ; Carcinoma ; genetics ; metabolism ; Cell Line, Tumor ; Cisplatin ; pharmacology ; Flow Cytometry ; Fluorouracil ; pharmacology ; Glycoproteins ; genetics ; metabolism ; Humans ; Laryngeal Neoplasms ; genetics ; metabolism ; Neoplasm Proteins ; genetics ; metabolism ; Neoplastic Stem Cells ; cytology ; drug effects ; metabolism ; Paclitaxel ; pharmacology ; Peptides ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo evaluate the acceptability of self-management project for patients with hypertension and the short term result, and provide the evidence of preventing and controlling hypertension using chronic disease self-management.

METHODSVoluntary patients with hypertension were grouped into the control and experimental groups after matching age, sex, education levels and the number of chronic diseases between the two groups. Data collected through questionnaire after intervention were compared with the baseline data through analysis of covariance and chi(2) test.

RESULTSRate of awareness for the diagnostic criteria of hypertension, risk factors and associated diseases increased by 56.4%, 50.4% and 37.6% respectively, and the rate of high salt diet intake, body mass index and the times of visiting doctors decreased by 27.1%, 0.8 kg/m(2) and 2.7 times on average in experimental group, as compared with control group. The health status (energy, fatigue, shortness of breathe, pain) improved (P < 0.05). In the similar comparison, the rate in which the hypertensive patients took antihypertensive medicine increased by 20%, and the scores of cognitive symptoms management and of patient communication with physician increased by 3.9 and 1.1 points respectively. The rate of compliance to medication increased by 21.5% compared with themselves.

CONCLUSIONThe awareness rate of hypertensive patients on knowledge and on unhealthy life style improved after implementing the self-management program. Their emotion, healthy behavior, health status and symptoms related to hypertension improved. Data showed that it was effective for hypertensive patients to use chronic diseases self-management.

Aged ; Awareness ; Body Mass Index ; Female ; Humans ; Hypertension ; therapy ; Life Style ; Male ; Middle Aged

Objective To explore the association between subclinical hypothyroidism and the left ventricular functions under conventional 2D Doppler echocardiography and to provide evidence for the protection of heart function.Methods Literatures regarding the association of subclinical hypothyroidism and the left ventricular functions were retrieved in large databases from home and abroad for the last 12 years.The left ventricular systolic function was assessed by left ventricular ejection fraction and the shortening of left ventricular fraction.The left ventricular diastolic function was assessed by left ventricular early diastolic filling flow velocity,late diastolic filling flow velocity,their ratios(E/A),and the left ventricular isovolumic relaxation time.The relationship between subclinical hypothyroidism and the left ventricular functions were assessed by Meta-analysis with Stata 11 software.The weighted mean difference(WMD)and 95％ confidence interval(CI)were calculated,and the publication bias was assessed by Begg' s test.Results 1 3 eligible papers were included.(1)Statistics on the combined data showed that in the evaluation of left ventricular diastolic function indicators.There were significant differences in left ventricular late diastolic filling flow velocity(WMD=4.51,95％CI:2.41 to 6.61)and E/A(WMD=-0.22,95％CI:-0.30 to-0.13),as well as the left ventricular isovolumic relaxation time(WM D=6.13,95％ CI:2.79 to 9.48)between patients with subclinical hypothyroidism and normal controls but,no significant difference was found in left ventricular early diastolic filling flow velocity.Looking at the left ventricular systolic function indicators.There were no significant differences in the left ventricular ejection fraction and left ventricular fractional shortening between patients with subclinical hypothyroidism and normal controls.(2)Data from the subgroup analysis showed that the differences of left ventricular late diastolic filling flow velocity,E/A and left ventricular isovolumic relaxation time were significantly different between patients with subclinical hypothyroidism and normal controls in the mean heart rate ≥72 bpm group.The difference of left ventricular isovolumic relaxation time was significantly different in the mean heart rate ＜72 bpm group,and the difference of left ventricular late diastolic filling flow velocity was significant in the mean age ＜60-year-old group.Conclusion Subclinical hypothyroidism was associated with the left ventricular diastolic dysfunction,but not associated with the left ventricular systolic dysfunction.The results suggested that subclinical hypothyroidism might change the heart function which could be evaluated by Doppler echocardiography.

Objective To clone and express the HIV-1B gp120 genes isolated at different organizations from a patient died of AIDS dementia complex (ADC) in eukaryotic cells.Methods Using the genomic DNA isolated from peripheral lymphnodes,choroid plexus and occipital white matter from a patient died of ADC as the template,HIV-1B gp120 gene was amplified with PCR.After sequenced,HIVIB gp120 was inserted into pcDNA3.1 ( + ) and recombinant expressing vector gp120/pcDNA3.1 ( + ) was constructed succeffuly confirming with sequencing. Then expressing vector was transfected into eukaryotic cells U87 using liposome transfection and expression of HIV-1B gp120 gene was assayed with indirect immunofluorescence.Results HIV-I B gp120 genes isolated from peripheral lymphnodes,choroid plexus and occipital white matter of the ADC patient were successfully cloned and recombinant expressing vector gp120/pcDNA3.i ( + ) could express envelope glycoprotein HIV-1B gp120 in U87 cells.Conclusion All the HIV-1B gp120 gene isolated at the different organizations of the same ADC patient could express in U87 cells,which may supply a valuable basis for studying the neurotoxicity and neurotoxic mechanism of HIV-1 gp120 protein.

Objective To investigate the influences of Lycium barbarum polysaccharide on apoptosis of corneal epithelial cells induced by hypertonicity through regulating adenosine monophosphate activated protein kinase(AMPK)/uncoordinated 51-like kinase 1(ULK1)autophagy pathway.Methods The ophthalmoxerosis cell model was established by osmotic pressure of 500 mOsm·L-1 on corneal epithelial cells.They were divided into model group,positive control group(0.3％sodium hyaluronate eye drops),inhibitor group(1 mg·mL-1 Lycium barbarum polysaccharide+10 μmol·L-1compound C),experimental-L,-H groups(0.5,1.0 mg·mL 1 Lycium barbarum polysaccharide),and normal cultured CRL-11135 cells were taken as blank group(no treatment was performed).Cell apoptosis was detected by flow cytometry.Autophagy was detected by MDC staining.Western blot was used to detect the expressions of p-AMPK/AMPK and p-ULK1/ULK1.Results The apoptosis rates of experimental-L,experimental-H,positive control,inhibitor,model and blank groups were(26.47±2.13)％,(13.68±2.21)％,(12.54±2.16)％,(18.73±2.12)％,(37.56±3.25)％and(6.35±2.14)％;the relative contents of autophagosomes were(59.63±8.14)％,(89.89±9.04)％,(90.31±9.13)％,(62.75±7.26)％,(43.11±6.45)％and(100.00±0.00)％;p-AMPK/AMPK were 0.45±0.07,0.64±0.08,0.66±0.06,0.53±0.04,0.34±0.05 and 0.87±0.06;p-ULKl/ULKl were 0.54±0.06,0.75±0.05,0.77±0.03,0.65±0.04,0.46±0.04 and 0.92±0.08,respectively.The above indexes in experimental-L,-H groups and positive control group were significantly different from those in model group(all P＜0.05);the above indexes in inhibitor group were significantly different from those in experimental-H group(all P＜0.05).Conclusion Lycium barbarum polysaccharide can inhibit the apoptosis of corneal epithelial cells induced by hypertonicity by activating the AMPK/ULK1 autophagy pathway.

Chemotherapy resistance is the main cause of poor prognosis in patients with advanced esophageal squamous cell carcinoma (ESCC). Pyroptosis is one of the anti-tumor mechanisms by chemotherapy drugs. Studies have shown that DEP-domain containing mTOR-interacting protein (DEPTOR) is correlated with sorafenib and gefitnib resistance, which is discovered as a naturally negative regulator of mammalian/mechanistic target of rapamicin (mTOR). In this study, DEPTOR knockdown (shDEPTOR) lentivirus was used to establish the stable DEPTOR knockdown ESCC cell lines. The results showed that knockdown of DEPTOR reduced chemosensitivity to cisplatin in ESCC cells in vitro. The lower expression of DEPTOR caused less extensive morphological characteristics of pyroptosis than that was observed in sh-con cells with the treatment of cisplain. Further studies showed that knockdown of DEPTOR induced downregulation of Caspase-1 expression and reduction of Caspase-1 activation, thereby inhibiting the activation of the classical pathway of pyroptosis. This paper demonstrates that DEPTOR can improve cisplatin chemosensitivity in ESCC cells via inducing Caspase-1-mediated pyroptosis.