MicroRNA expression profiles in early mycosis fungoides / 中华皮肤科杂志

Objective To screen microRNAs(miRNAs)related to early mycosis fungoides(MF). Methods A high?throughput miRNA PCR array was used to determine miRNA expression profiles in skin lesions of 6 patients with early MF (early MF group) and 6 patients with lichen planus (control group), followed by screening of differentially expressed miRNAs between the two groups. Then, real?time fluorescence?based quantitative PCR(RT?qPCR)was performed to verify the differentially expressed miRNAs in lesional specimens from 13 patients with early MF and 13 patients with eczema or lichen planus, as well as in Myla cells and normal human T?lymphocytes. Results The high?throughput miRNA PCR array showed that the expressions of hsa?miR?378a?5p, hsa?miR?107 and hsa?miR?302c?3p were significantly higher in the early MF group than in the control group(all P<0.05). For skin lesions, the results from RT?qPCR were similar to those from the miRNA array assay. Compared with normal human peripheral blood T?lymphocytes, Myla cells showed significantly increased expressions of hsa?miR?378a?5p and hsa?miR?107, which was consistent with the results from the miRNA array assay. However, no significant difference was observed in the expression of hsa?miR?302c?3p between the two kinds of cells. Conclusion MiRNA expression profiles in early MF are different from those in inflammatory skin diseases.

Associations of apparent diffusion coefficient value from MR diffusion-weighted imaging with Ki-67 expression and differentiation grade in gastric cancer / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To examine the associations of apparent diffusion coefficient (ADC) value from MR diffusion-weighted imaging (DWI) with Ki-67 expression and differentiation grade in gastric cancer.</p><p><b>METHODS</b>Images and pathologic data of 68 gastric cancer patients between September 2013 and February 2015 in Affiliated Changshu Hospital of Soochow University were analyzed retrospectively. The expression of Ki-67 antigen in cancer tissue sample was determined by immunohistochemistry. Ki-67 labeling index(LI) was calculated to divide the cases into low Ki-67 group(Ki-67 LI <50%) and high Ki-67 group (Ki-67 LI ≥ 50%). Associations of ADC value with differentiation grade and Ki-67 LI were examined.</p><p><b>RESULTS</b>Mean ADC value of low Ki-67 LI group was significantly higher than that of high Ki-67 LI group [(0.977 ± 0.100) × 10(-3) mm(2)/s vs. (0.859 ± 0.064) × 10(-3) mm(2)/s, P=0.000]. The ADC value was negatively correlated with Ki-67 LI (r=-0.685, P=0.000). Mean ADC value of well differentiated adenocarcinoma, moderately differentiated adenocarcinoma, poorly differentiated adenocarcinoma, and signet-ring cell carcinoma was (1.124 ± 0.080) × 10(-3) mm(2)/s, (0.950 ± 0.064) × 10(-3) mm(2)/s, (0.899 ± 0.091) × 10(-3) mm(2)/s, and (0.894 ± 0.081) × 10(-3) mm(2)/s respectively. Difference of ADC value among differentiation grades was significantly different (F=11.405, P=0.000). Difference of ADC value between well differentiated adenocarcinoma and non-well differentiated adenocarcinoma was significantly different(P=0.000).</p><p><b>CONCLUSION</b>ADC value is associated with differentiation grade and Ki-67 LI of gastric cancer, which may be used as a noninvasive predictor for evaluating the proliferation and differentiation grade of gastric cancer.</p>

Screening for and identification of targets of let-7a microRNA in A375 melanoma cells by using iTRAQ technology / 中华皮肤科杂志

Objective To screen for and identify targets of let-7a microRNA (miRNA) in A375 melanoma cells by using isobaric tags for relative and absolute quantitation (iTRAQ) technology,and to explore mechanisms underlying the tumor-suppressing effect of let-7a.Methods Cultured A375 cells were classified into two groups to be transfected with 100 nmol/L hsa-1et-7a mimics (hsa-let-7a mimics group) or negative control mimic (NC group).After 54-hour incubation,A375 cells were collected and total proteins were collected.iTRAQ technology was used to analyze and identify differentially expressed proteins,bioinformatic analysis was performed to assess let-7a candidate targets and their functions,and a dual-luciferase reporter system was utilized to verify let-7a targets.Results As mass spectrometry showed,a total of 327 differentially expressed proteins were identified in the hsa-1et-7a mimics group compared with the NC group,including 151 up-regulated proteins with iTRAQ ratio ＞ 1.2 and 176 down-regulated proteins with iTRAQ ratio ＜ 0.8.Of 176 down-regulated proteins,47 were predicted as miRNA targets by the miRWalk software.The dual-luciferase reporter system showed that the relative luciferase activity of the 3' untranslated region (UTR) of the wild-type HMGA2 and THOC2 genes were reduced by 64.3％ and 46.4％,respectively,in the hsa-1et-7a mimics group compared with the NC group.Conclusion A total of 47 candidate let-7a targets were screened out in A375 melanoma cells by using iTRAQ technology and bioinformatic analysis,and HMGA2 and THOC2 genes were identified as direct targets of let-7a.

Effect of T-cell immunoglobulin and mucin domain-3 on TRP-2180-188 peptide-stimulated murine spleen lymphocytes co-cultured with B16F10 murine melanoma cells / 中华皮肤科杂志

Objective To evaluate the effect ofT-cell immunoglobulin and mucin domain-3 (TIM-3) on TRP-2180-188 peptide-stimulated murine spleen lymphocytes co-cultured with B16F10 murine melanoma cells.Methods A recombinant plasmid pFUSE-TIM-3-mIgG2Aae1-Fc2 encoding TIM-3 was constructed.Then,the recombinant plasmid and an empty plasmid pFUSE-mIgG2Aae1-Fc2 were transfected into human 293T epithelial cells followed by 48-hour culture for the preparation of supernatants containing TIM-3 and Ig-tail respectively.C57BL/6 mice were immunized with the TRP-2180-188 peptide vaccine for 4 sessions.One week after the last vaccination,C57BL/6 mice were sacrificed,and spleen lymphocytes were collected and then cultured with the TRP-21180-188 peptide and interleukin-2 (IL-2) for 5 days,with lymphocytes untreated with the TRP-2180-188 peptide or IL-2 serving as the control group.Mitomycin-treated B16F10 murine melanoma cells and TRP-2180-188 peptide-stimulated lymphocytes were co-cultured with the presence of supernatants of 293T cells that had been cultured for 48 hours (blank control group),TIM-3-containing supernatants (TIM-3 group) and Ig-tail-containing supernatants (negative control group) separately.After 24 and 48 hours of co-culture,cell counting kit-8 (CCK-8) assay was performed to estimate the proliferative activity of lymphocytes,enzyme-linked immunosorbent assay (ELISA) to determine the supernatant levels of interferon (INF)-γ and tumor necrosis factor (TNF)-α,flow cytometry to determine the percentage of CD8 + T cells in the co-culture system.Results Enzyme digestion and sequence analysis showed that the TIM-3 gene was successfully inserted into the eukaryotic expression plasmid.After 48-hour culture,TIM-3 and Ig-tail expressions were detected in the supernatants of 293T cells transfected with the recombinant plasmid and empty plasmid respectively.As CCK-8 assay showed,the proliferative activity of lymphocytes was significantly lower in the TIM-3 group than in the blank control group and negative control group after 24-and 48-hour culture (78.06％ ± 6.37％ vs.100.00％ ± 10.42％ and 108.70％ ± 9.90％ at 24 hours,42.93％ ± 5.93％ vs.100.00％ ± 6.24％ and 168.00％ ± 2.98％at 48 hours,all P ＜ 0.05),so was the ratio of cellular proliferative activity at 48 hours to that at 24 hours (all P ＜ 0.05).Compared with the blank control group and negative control group,the TIM-3 group showed significantly decreased supernatant levels of IFN-γ and TNF-α after 24-hour (IFN-γ:192.96 γ 5.05 ng/L vs.216.44 ± 7.85 ng/L and 223.67 ±7.79 ng/L,both P＜ 0.05;TNF-α:58.43 ± 0.26 ng/L vs.26.43 ± 0.01 ng/L and 86.85 ± 1.12 ng/L,both P＜ 0.05) and 48-hour culture (IFN-γ:54.95 ± 0.57 ng/L vs.230.06 ± 4.23 ng/L and 167.24 ± 3.33 ng/L,both P ＜ 0.05;TNF-α:30.23 ±0.26 ng/L vs.26.84 ± 0.20 ng/L and 45.34 ± 0.22 ng/L,both P ＜ 0.05).In addition,the median percentage of CD8+ T cells was significantly increased in the TIM-3 group compared with the blank control group and negative control group after 24-and 48-hour culture (3.30％ vs.0.421％ and 2.22％ at 24 hours,4.06％ vs.0.577％ and 0.691％ at 48 hours,all P＜ 0.05).Conclusion TIM-3 in vitro can suppress the proliferative activity of and secretion of IFN-γand TNF-α by lymphocytes,but increase the percentage of CD8 + T cells in the co-culture system of TRP-2180-188 peptide-stimulated lymphocytes and B16F10 cells.

Value of diffusion-weighted MRI in the diagnosis of T staging for rectal cancer / 中华胃肠外科杂志

<p><b>OBJECTIVE</b>To investigate the value of diffusion-weighted imaging (DWI) in the diagnosis of T staging for rectal cancer.</p><p><b>METHODS</b>Clinicopathologic data and MR images of 46 patients with rectal cancer in our hospital from July 2013 to September 2014 were retrospectively analyzed. The diagnostic sensitivity, specificity and accuracy of T2WI were compared with those of T2WI plus DWI in T staging for rectal cancer. The relationship of mean apparent diffusion coefficient (ADC) value with different T stages of rectal cancer was analyzed.</p><p><b>RESULTS</b>There were no significant differences in the diagnostic sensitivity, specificity and accuracy between T2WI and T2WI plus DWI (all P>0.05). The mean ADC value of DWI performed in pathologic T2, T3a, T3b, T3c and T4 stage was (1.110 ± 0.117) × 10⁻³ mm²/s, (1.035 ± 0.121) × 10⁻³ mm²/s, (0.948 ± 0.109) × 10⁻³ mm²/s, (0.932 ± 0.122) × 10⁻³ mm²/s and (0.843 ± 0.050) × 10⁻³ mm²/s, respectively (F=6.972, P=0.000).</p><p><b>CONCLUSION</b>DWI can serve as a complement for T2WI in the diagnosis of T stage patients with rectal cancer, and its ADC value presents a downward trend with the advance of T stage.</p>

The efficacy of intratympanic dexamethasone injection for the moderate and severe sudden deafness with BPPV / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To evaluate the efficacy of intratympanic dexamethasone injection for the moderate and severe sudden deafness with BPPV.@*METHOD@#A total of 63 patients diagnosed with sudden sensorineural hearing loss with BPPV were treated through OPD. Patients were divided into three groups: 20 cases in intratympanic dexamethasone injection as initial treatment (group A); 18 cases in systemic hormone therapy group (group B); 25 cases in intratympanic dexamethasone injection as salvage treatment (group C). In addition, routine drugs were used to all patients.@*RESULT@#The overall effective rate of group A, B and C in hearing recovery was 60.0%, 38.9% and 48.0%, respectively: (1) No significant difference of hearing recovery was observed among three groups (P > 0.05); (2) A significant difference of hearing recovery was evidenced between group A and C (P 0.05).@*CONCLUSION@#Our data showed that intratympanic dexamethasone should be used as initial therapy for treating the moderate and severe sudden deafness with BPPV.

The roles of otolith organs in the recurrence primary benign paroxysmal positional vertigo / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To explore the roles of otolith organs in the occurrence and recurrence of primary benign paroxysmal positional vertigo (BPPV) by vestibular evoked myogenic potential (VEMP) test.@*METHOD@#We enrolled 17 recurrent primary BPPV patients and 42 non-recurrent primary BPPV patients between September 2014 and November 2014. All patients underwent VEMP tests, including cervical vestibular evoked myogenic potential (cVEMP and ocular vestibular evoked myogenic potential (oVEMP) tests. The abnormal case was defined as non-elicitation or asymmetry rate between bilateral sides is larger than 29%.@*RESULT@#Significant difference was found in abnormal rate between cVEMP and oVEMP (P 0.05). No significant difference was found in sex and age between recurrent and non-recurrent groups (P > 0.05).@*CONCLUSION@#The impairment of otolith organs, especially the utricle, is related to primary BPPV. Dysfunction of utricle may play a role in recurrence of BPPV. Recurrence of BPPV is not correlated with sex and age.

Efficacy analysis of Chuanxiongqingnaokeli treatment of children with migraine / 中国综合临床

Objective To observe the clinical efficacy and adverse reaction of Chuanxiongqingnaokeli for children with migraine. Methods One hundred children patients with migraine were randomly divided into treatment group( n = 50 ) and control group( n = 50 ). Patients in treatment group were given Chuanxiongqingnaokeli,10 g/once,and three times one day,while in control group were given Flunarizine Hydrochloride Capsules,2. 5 mg/once,and who's body mass ﹥50 kg with 5. 0 mg/once,and one times each night. Three months as one course of treatment,and compared the efficacy of two groups after tree course of treatment. Results The hemodynamics of two groups all decreased after treatment compared with before treatment,but ACA,MCA,PCABA and VA in treatment group(( 81. 10 ± 11. 95 ),( 93. 3 ± 14. 16 ),( 70. 2 ± 11. 57),(70. 6 ± 13. 02),(65. 5 ± 12. 6)cm/s respectively)decreased more significantly than that of control group(( 104. 2 ± 12. 63 ),( 116. 2 ± 15. 82 ),( 93. 5 ± 11. 91 ),( 93. 5 ± 12. 71 ),87. 4 ± 12. 92 ) cm/s respectively),and the differences were significant( P﹤0. 05). The headache frequency and duration in treatment group were(1. 0 ± 0. 6)and(3. 3 ± 1. 0),less than that of control group((2. 3 ± 0. 9)and(5. 6 ± 1. 7);t= -3. 345,-3. 269;P﹤0. 05). The total effective rate in treatment group was 90. 0%(45/50),higher than that of control group(74. 0%(37/50);χ2 =4. 336,P﹤0. 05). There was no severe adverse reaction in both two groups. Conclusion The Chuanxiongqingnaokeli is safe and effective for treatment of children with migraine.

The preliminary research of progress on sudden deafness with benign paroxysmal positional vertigo patients / 临床耳鼻咽喉头颈外科杂志

OBJECTIVE@#To investigate the prognosis of sudden deafness patients with benign paroxysmal positional vertigo (BPPV).@*METHOD@#The clinical data of 24 sudden deafness patients with BPPV was analyzed. The outcome of 125 sudden deafness patients without BPPV at the same time was compared.@*RESULT@#Hearing improvement after three months treatment was 41.67% and 72.80% in sudden deafness patients with BPPV and sudden deafness patients without BPPV, respectively. The difference was statistically significant (P<0.05).@*CONCLUSION@#The prognosis of hearing in sudden deafness patients with BPPV is worse than that in sudden deafness patients without BPPV. BPPV may predict a poor hearing outcome in sudden deafness.

Regulatory effect of anti-IL-6Rβ(gp130) mAb on IL-6 signaling pathway / 中国免疫学杂志

Objective:To analyse the biological function of anti-IL-6Rβ(gp130) monoclonal antibody and its regulatory effect on IL-6 signaling.Methods:Biological characteristics of anti-IL-6Rβ(gp130) mAb were assessed by Western blot analysis, capture ELISA and peptide ELISA .The phosphorylation of STAT 3 was tested by Western blot analysis in IL-6-stimulated U266/RA-FLS/RA-PBMC with or without anti-IL-6Rβ(gp130) mAb treatment.Results:3 strains of mouse anti-human gp130 mAb were with high affini-ty and different binding epitopes , the kaff of 10A1 was 2.62E-10.In U266, RA-PBMC and RA-SFMC, IL-6 signaling highly activated STAT3 which could be inhibited by anti-gp130 mAb.Conclusion: Anti-IL-6Rβ( gp130 ) mAb might have different binding epitopes and could affect IL-6 stimulated phosphorylation of STAT3, which provides a preliminary experiment for analyse the correlation of IL-6 signaling and RA .

Role of neuropeptide Y and peroxisome proliferator-activated receptor γ coactivator-1α in stress cardiomyopathy / 华中科技大学学报（医学）（英德文版）

Death following situations of intense emotional stress has been linked to the cardiac pathology described as stress cardiomyopathy, whose pathomechanism is still not clear. In this study, we sought to determine, via an animal model, whether the transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator-1alpha (PGC-1α) and the amino peptide neuropeptide Y (NPY) play a role in the pathogenesis of this cardiac entity. Male Sprague-Dawley rats in the experimental group were subjected to immobilization in a plexy glass box for 1 h, which was followed by low voltage electric foot shock for about 1 h at 10 s intervals in a cage fitted with metallic rods. After 25 days the rats were sacrificed and sections of their hearts were processed. Hematoxylin-eosin staining of cardiac tissues revealed the characteristic cardiac lesions of stress cardiomyopathy such as contraction band necrosis, inflammatory cell infiltration and fibrosis. The semi-quantitative RT-PCR analysis for PGC-1α mRNA expression showed significant overexpression of PGC1-α in the stress-subjected rats (P<0.05). Fluorescence immunohistochemistry revealed a higher production of NPY in the stress-subjected rats as compared to the control rats (P=0.0027). Thus, we are led to conclude that following periods of intense stress, an increased expression of PGC1-α in the heart and an overflow of NPY may lead to stress cardiomyopathy and even death in susceptible victims. Moreover, these markers can be used to identify stress cardiomyopathy as the cause of sudden death in specific cases.

Role of neuropeptide Y and peroxisome proliferator-activated receptor γ coactivator-1α in stress cardiomyopathy / 华中科技大学学报（医学）（英德文版）

Death following situations of intense emotional stress has been linked to the cardiac pathology described as stress cardiomyopathy, whose pathomechanism is still not clear. In this study, we sought to determine, via an animal model, whether the transcriptional coactivator peroxisome proliferator-activated receptor γ coactivator-1alpha (PGC-1α) and the amino peptide neuropeptide Y (NPY) play a role in the pathogenesis of this cardiac entity. Male Sprague-Dawley rats in the experimental group were subjected to immobilization in a plexy glass box for 1 h, which was followed by low voltage electric foot shock for about 1 h at 10 s intervals in a cage fitted with metallic rods. After 25 days the rats were sacrificed and sections of their hearts were processed. Hematoxylin-eosin staining of cardiac tissues revealed the characteristic cardiac lesions of stress cardiomyopathy such as contraction band necrosis, inflammatory cell infiltration and fibrosis. The semi-quantitative RT-PCR analysis for PGC-1α mRNA expression showed significant overexpression of PGC1-α in the stress-subjected rats (P<0.05). Fluorescence immunohistochemistry revealed a higher production of NPY in the stress-subjected rats as compared to the control rats (P=0.0027). Thus, we are led to conclude that following periods of intense stress, an increased expression of PGC1-α in the heart and an overflow of NPY may lead to stress cardiomyopathy and even death in susceptible victims. Moreover, these markers can be used to identify stress cardiomyopathy as the cause of sudden death in specific cases.

Cloning and expression of lipoxygenase gene from Anabaena sp. PCC 7120 and purification, characterization of the recombinant enzyme / 生物工程学报

We cloned the lipoxygenase gene (ana-LOX) from Anabaena sp. PCC 7120 and expressed it in Escherichia coli BL21 (DE3) pLysS. We determined the active site of the recombinant ana-LOX through site-directed gene mutagenesis and obtained the shortest length of the functional gene. Meanwhile, we studied the properties of recombinant ana-LOX after purification. The C-terminal of the Aos (allene oxide synthase)-LOX fusion gene in Anabaena sp. PCC 7120 genome was found belonging to LOXs family by bioinformatics analysis. Further results of site-directed gene mutagenesis confirmed that the active sites of ana-LOX were His197, His202, His369, Asn373and Ile455. The shortest length of functional gene was identified to be 1 254 bp based on the strategy of shortening the gene length gradually. The highest activity of recombinant ana-LOX of 6 750 U/mL could be achieved when constructed to pET-32a vector and expressed at low temperature 16 degrees C. We purified the enzyme by Ni-NTA chelating affinity chromatography, with 60.89% yield and specific activity of 11.4 x 10(4) U/mg. The optimum reaction temperature and pH for ana-LOX were 45 degrees C and 6.0, respectively. Furthermore, the obtained ana-LOX was stable at room temperature. The effect of metal ions on ana-LOX was determined also. Fe2+, Mg2+ Ca2+ could markedly promote the activity of this enzyme whereas Fe3+ and Cu2+ had a strong inhibitory effect on it. Finally, the ana-LOX could improve the microscopical structure of dough. The results of this study will provide a basis for future improvements and food industrial applications of ana-LOX.

Clinical features and etiology of cerebral infarction in children / 中国小儿急救医学

Objective To summarize the clinical features and etiology of cerebral infarction in children.Methods The clinical data of 47 children with cerebral infarction who were hospitalized in Shengjing Hospital of China Medical University from Jan 2009 to Jul 2011 were analyzed retrospectively.Results There were 30 boys and 17 girls in all the 47 children.The median age of onset was 3.1 years(ranged from 2 months to 11 years old).Among 47 cases,the common neurological manifestations included limb paralysis in 32 cases(68.1％ ),central facial paralysis in 15 cases(31.9％ ),convulsion in 12 cases(25.5％ ),disturbance of consciousness in 10 cases(21.3％ ),and language disorders in 10 cases(21.3％ ).Among 47 cases,31 cases had basal ganglia infarction with neuronal imaging( CT or MRI),of whom 4 cases accompanied with other location infarction.Several lobes of infarction in 5 cases,hemispheric infarction in 3 cases,parietal infarction in 2 cases,frontal lobe infarction in 2 cases,temporal lobe infarction in 2 cases,and thalamic infarction in 2 cases.Nineteen cases were carried out blood vessel imageology examination,11 cases showed abnormality,the most common affected cerebral blood vessel were middle cerebral artery(5 cases).The common causes of 47 cases were trauma ( 19 cases,40.4％ ),infection( 12 cases,25.5％ ) and moyamoya disease (5 cases,10.6％ ).Ten children (21.3％) had no identifiable cause.Conclusion The common period of cerebral infarction is in infancy.The most frequent neurological symptom is hemiplegia.The most common region of infarction is in basal ganglia with neuronal imaging.The common causes of cerebral infarction are trauma,infection and moyamoya disease.

Expression, purification, and bio-activity analysis of fusion protein HBx-EGFP-TLM / 生物工程学报

Hepatitis B virus X protein (HBx) has various functions and plays a crucial role in the development of hepatocellular carcinoma (HCC). However, due to different transfection efficiency levels and experimental approaches, it is difficult to correlate the exact functions of HBx to HBV-associated HCC. In this study, we constructed two prokaryotic expression vectors, pGEX-HBx-EGFP-TLM and pGEX-EGFP-TLM, which expressed HBx-EGFP-TLM and EGFP-TLM fusion proteins respectively. Both vectors contained a coding sequence of TLM transduction motif derived from the PreS2-domain of Hepatitis B Virus surface antigens. In addition, EGFP was expressed as a reporter reflecting the transduction efficiency of TLM. The fusion protein HBx-EGFP-TLM or EGFP-TLM purified from Escherichia coli BL21(DE3) by AKTA Purifier system was incubated with AML12 and SMMC-7721 cells. Both Western blotting and laser confocal results indicated that the translocation motif TLM could lead HBx-EGFP and EGFP into the cytoplasm. Dual-Luciferase Reporter Assay revealed that the activity of mEZH2 promoter could be up-regulated by the recombinant HBx. In conclusion, we expressed a cell-permeable HBx, which could provide a new method to study the functions of HBx.

Composition of and drug resistance in bacteria isolated from the lesions of patients with squamous cell carcinoma of the face and head / 中华皮肤科杂志

Objective To analyze the composition of and drug resistance in bacteria isolated from the lesions of patients with squamous cell carcinoma of the face and head.Methods Lesional tissue or discharges were obtained from 246 patients with squamous cell carcinoma of the face and head,and subjected to conventional bacterial culture.The isolated bacteria were identified by VITEK TWO automated microbiology system.Antimicrobial susceptibility testing was carried out by Kirby-bauer method.WHONET 5.3 software was utilized for statistical analysis.Results Totally,294 bacterial strains were isolated,including 168 Gram-negative bacteria (57.1％)and 126 Gram-positive bacteria(42.9％).The bacterial isolates were predominated by Staphylococcus aureus(21.4％),followed by Escherichia coli(20.4％),Staphylococcus epidermidis(18.4％),Klebsiellapneumoniae(15.4％)and Pseudomonas aeruginosa(9.5％).The prevalence was 40％,26.7％,42.9％ and 55.6％ respectively for extended spectrum β lactamases-producing E.coli and K.pneumoniae,methicillinresistant staphylococcus aureus(MRSA)and methicillin-resistant coagulase-negative S.epidermidis(MRCNS)respectively.P.aeruginosa,E.coli and K.pneumoniae were highly susceptible to imipenem and meropenem,and favorably sensitive to β-lactam and β-1actamase inhibitor combination.No resistance was observed for vancomycin,teicoplanin or linezolid in staphylococci.Conclusions The bacterial isolates from squamous cell carcinoma tissue on the head and neck are predominated by conditional pathogenic bacteria,and the proportion of Gram-negative bacteria is higher than that of Gram-positive bacteria.These isolates seem to be highly resistant to common antibiotics.

Analysis of PARP10 tissue expression profile, interactive protein and UV stress reaction / 生物工程学报

One pair of primers were designed and synthesized based on the cDNA sequence encoding Homo sapiens poly (ADP-ribose) polymerase family, member 10 (PARP10) reported on the GenBank. The cDNA sequence encoding PARP10 was cloned from 293FT cell by RT-PCR. Then the RT-PCR product was cloned into pCMV-Myc and pEGFP-C1 plasmids. The interaction between PARP10 and beta-actin was identified through immuno-precipitation and laser confocal microscopy. Extensive expression of PARP10 in mouse tissues was confirmed by RT-PCR. Besides, Western blotting analysis indicated that cell injury caused by UV treatment could promote the expression of PARP10. The results in this paper would benefit further study of PARP10.

Generation of mouse UBE2W antibody and analysis of UBE2W expression in mouse tissues / 生物工程学报

Ubiquitin conjugating enzyme functions as the second enzyme required for protein ubiquitination and plays an important role in ubiquitin transferring and substrate specific recognition. UBE2W, a newly described member of E2 family, was formerly reported probably involving in phototransduction or retinal degeneration in Drosophila. In this study, we report that murine UBE2W harbors a typical UBC domain and is highly conserved in different vertebrate homologues. GST-tagged UBE2W was expressed in E. coli BL21 (DE3) and purified with GST affinity chromatography. Using this antigen, we generated and further separated rabbit polyclonal antibody of UBE2W, of which the activity and specificity were confirmed by immunoblotting of transiently expressed myc-UBE2W fusion protein. Wide expression of UBE2W was found in brain, muscle, heart, lung, liver, spleen, kidney and testis of mouse with the generated antibody, indicating the functional importance of this novel protein. Furthermore, the UBE2W highly expression was confined to the adult testis and was developmental stage-specific.

Effect of influenza virus NS1 protein on host cell / 生物工程学报

NS1 is a non-structural protein of the influenza A virus, which could only be expressed when cells are infected. The effect of NS1 protein on host cell is still not clear. To understand the role of NS1 protein in cell infection, recombinant plasmid pCMV-myc-NS1 was constructed, and then transfected into A549 cells. Two-dimensional electrophoresis was employed to analyze proteins regulated by NS1 that could reflect the interaction between influenza virus and host cells at the protein level. The influence of NS1 on cell proliferation and cell cycle was also studied. The result showed that not only could NS1 remarkably affect metabolism, but it could also slow down cell proliferation through blocking cell cycle.

Selecting functional siRNA target sites of hUBE2W based on H1-U6 dual promoter RNAi plasmid / 生物工程学报

hUBEW, a newly identified class I ubiquitin conjugating enzyme, probably plays an important role in tumorigenesis and DNA repair processes. RNA interference (RNAi) is a process in cells to degrade specific homologous mRNA by forming duplex RNA and has been developed into a powerful tool to study gene functions. In this study, the H1-U6 dual promoter RNAi plasmid was constructed and the target sequence for hUbe2w could be transcribed from both strands and form a double stranded RNA with two 5'Uridine overhangs, which closely resembles endogenous functional siRNA. The hUbe2w cDNA was amplified from reverse transcription of the 293FT total RNA by RT-PCR, and then cloned into the pGL3-Control, pCMV-myc and pDsRed-express-C1 plasmids respectively, which were selected as report vectors to detect the RNAi effects. The plasmids were co-transfected into HEK293FT cells, and then the luciferase activity and hUBE2W protein expression were measured respectively. The Resulted reduction of mRNA and protein level demonstrate that the targets of 125 and 259 could significantly inhibit the hUbe2w expression.