To study the effect of physiological conditions on spidroins, we analyzed NTR1SR2CT module secondary structure, aggregation and silk-formation influenced by different salts (in different concentration intervals). According to the full-length Araneus ventricosus MiSp sequence, NTR1SR2CT module was constructed and expressed in Escherichia coli BL21 (DE3), and the recombinant proteins were purified by denaturation method mediated by 8 mol/L urea. Random coil and helix are the main secondary structures of NTR1SR2CT and could be induced into beta-sheet by drying natively and lyophilization, where methanol can be used as a promoter. Furthermore, potassium and phosphate cations can cause significant NTR1SR2CT protein aggregation and silk-formation. The results could be a basis for the determination of silk-formation mechanism, and also useful for industrialized generation of high performance spider silk-like fibers.
Animals ; Fibroins ; chemistry ; Protein Structure, Secondary ; Salts ; chemistry ; Spiders

Objective To design and prepare an RNA interfering vector for effectively inhibiting the cellular expression of zinc finger protein A20 and observe the effect of A20 gene silence on cellular inflammatory response. Methods Specific RNA interfering oligonucleotide fragments (ASRF) were designed and synthesized artificially and the A20 RNA interfering vector pSUPER-EGFP-A20 siRNA constructed. Human monocyte cell line THP1 was used to infect the pSUPER-EGFP-A20 siRNA by means of genetic transfection technique; then, silence rate of cellular A20 was analyzed by real-time polymerase chain reaction (PCR). In the meantime, the activity of nuclear transcription factor nuclear factor-κB (NF-κB) and the level of tumor necrosis factor-α (TNF-α) in culture supernatant were measured by ELISA. Results Of two specific inhibitory oligonueleotide fragments of A20, the fragment M59465-385R/F had a higher inhibition to A20 expression, with rate of A20 gene silence of 83.86%. Preliminary application showed that after A20 gene silence, the activity of NF-κB was increased by 78.13% and the level of TNF-α in cell culture supernatant was increased by 49.30%. Conclusions Vector of A20gene silence with a high efficiency is obtained successfully. Preliminary application indicates that the expression of A20 can down-regulate the degree of cellular inflammatory responses.

According to the characteristics of the Laboratory Medicine and our twenty years' medical educational experience ,we explore to improve the quality of education and cultivate the students' practical ability and creative ability. And we construct an educational mode for clinical practice, laboratory medical practice and research project.

Objective: To investigate the innate immune response of influenza virus-infected glial cells,the transcription levels in chemokines in mouse microglia and astrocytes were detected which pre-infected by human H1N1 or avian H5N1 influenza viruses.Methods: The glial cells isolated from neonatal mice cerebral cortex were cultured and further microglia and astrocytes were purified.The primary mouse microglia and astrocytes were infected in vitro by H1N1 or H5N1 influenza viruses in a multiplicity of infection (MOI) 2.Eight hours post infection,the influenza virus nucleoprotein (NP) was detected by immunofluorescence to identify the proportion of infected cells.The cellular RNA were extracted at 6 h and 24 h to detect the transcriptional level of chemokines by semi-quantitative RT-PCR.Results: More than 95% of the microgha and astrocytes which isolated from mice were infected.The transcription levels of CCL-3,CCL-5,CXCL-2,CXCL-9 and CXCL-10 from infected microglia and astrocytes were upregulated.Futhermore,the mRNA level of CXCL-10 increased much more.In addition,avian H5N1 influenza virus could induce more stronger upregulation of those chemokines than human H1N1 did.Conclusion: The mouse microglia and astro cytes which are infected by H1N1 influenza virus or H5N1 influenza virus could induce upregulation of transcription level of chemokines.

Objective To investigate the clinical significance of the expression of XIAP mRNA and Survivin mRNA in non-small cell lung cancer and their relationship. Methods RT-PCR was used to evaluate the expression of XIAP mRNA and Survivin mRNA in 59 cases of NSCLC and corresponding normal tissues.Results There was a significant difference in XIAP mRNA expression in primary lung and corresponding normal tissues (61.0 % vs 30.5 %, P<0.05), whereas there were no significant correlation between the XIAP mRNA expression and gender, age, smoking history, histological subtype, T stage, lymph node metastatic status and TNM stage. There was a significant difference in Survivin mRNA expression in primary lung and corresponding normal tissues (81.4 % vs 23.7 %, P<0.05), whereas there were no significant correlation between the Survivin mRNA expression and gender, age, smoking history, histological subtype, T stage, lymph node metastatic status and TNM stage. Conclusion The significant difference of XIAP mRNA and Survivin mRNA expression between the tumor and corresponding normal tissues implies they might play important roles in the carcinogenesis and progress of NSCLC and might become marker for the diagnosis and target for treatment of NSCLC.

Objective:To study the rule of spontaneous behavior and to explore the effect on emotion of mice peripherally infected with influenza A WSN33 virus( H1N1).Methods: Mice were intranasal inoculated with H1N1 WSN33 or saline.Then mice bodyweight change,and total distance movement,average movement speed distance in the central area and feces in the open field test in 5 minutes were recorded in two weeks.Results: Mice following WSN33 infection bodyweight declined sharply until day 7 post-inoculation,and mice bodyweight recovered from influenza infection at day 8 post-inoculation.Total distance movement of mice following H1N1 WSN33 infection decreased in the open field test,and difference of the reduction was significant from day 5 to day 10 post-inocu-lation.The average movement speed had no statistical difference.The range of numbers of fecal grains was large, and they were no significant difference.Conclusion:The total distance movement decreased,but average movement speed did not change following mice infected with H1N1 WSN33.They told us that mice infected with H1N1 WSN33 had anxiety,depressed and nervous emotion which is more evident in acute stage and early recovery stage,whereas the change of the nervous emotion was small and not obvious.

Objective T o study the effect of different attentional conditions on the event-related potential (E R P ) com ponents generated by the visual inform ation stim uli related to visual acuity, and provide a theoretical reference for clinical forensic visual objective evaluation. Methods W ith visual acuity opto-types as norm al form of visual inform ation stim uli, 15 volunteers as study subjects w ere supposed to ac-count the visual acuity optotypes under the attentional condition of visual stim uli. Furtherm ore, the sub-jects w ere required to listen to the storytelling carefully under the non-attentional condition of visual stim uli, and after the exam ination, they needed to answ er the story-related questions. A ll the E E G re-sults of tw o different attentional conditions from the subjects w ere recorded by 32 channel E R P system . Results U nder tw o attentional conditions, P 1 and P 300 com ponents w ere evoked by the visual acuity opto-types on supra-threshold and threshold visual acuity levels, w hile only P 1 com ponent w ere evoked by the visual acuity optotypes on sub-threshold levels. In the E R P w aveform s evoked by the visual acuity optotypes on supra-threshold, P 1 and P 300 am plitudes under attentional condition w ere larger than that un-der non-attentional condition. Conclusion A ttentional conditions can influence the detection of visual acuity. P 300 com ponent can be used to distinguish the visual acuity levels w ith supra-threshold and sub-threshold under non-attentional condition.

Objective To investigate the clinical efficacy and safety of oral high-dose methylprednisolone in the treatment of infantile spasms (IS).Methods The clinical data of 38 children with infantile spasms were analyzed retrospectively who treated with oral administration of high-dose methylprednisone in Department of Neurology,Wuhan Children's Hospital,Tongji Medical College,Huazhong University of Science and Technology from January 2016 to April 2017.Results (1) Twenty patients (52.6％) of all the 38 patients were seizure-free after 2 weeks of treatment,and 16 cases (42.1％) were seizure-free at the end of treatment.(2) All the 38 cases were typical or atypical hyperarrhythmia.After treatment of 2 weeks,25 cases (65.8％) of hyperarrhythmia disappeared;at the end of the treatment,30 cases (78.9％) of hyperarrhythmia disappeared.(3) Adverse effects mainly were weight gain,Cushing signs,increased appetite,irritability,drowsiness,co-infection,electrolyte disturbance.(4) Follow-up of 3 to 12 months,the recurrence rate was low and the development quotient had improved.Conclusions Oral high dose methylprednisolone in the treatment of IS is effective,safe and has a low recurrence rate.It can be recommended in clinicalal application.

APOBEC(apolipoprotein B mRNA-editing enzyme catalytic-polypeptide) family members were reported as innate immune molecules with anti-viral activity for many viruses, such as HIV and HBV.In order to understand the function of APOBEC, the APOBEC-3F and-3G were cloned, expressed, and the sub-cellular localization of them was detected.The genes of APBEC-3F and-3G were cloned from PHA-stimulated PMBC and expressed in the MDCK cell by transfection.The sub-cellular localization of APOBEC-3F and-3G were detected by immunofluorescence.APOBEC-3F and-3G were cloned by RT-PCR and confirmed by DNA sequencing.The immunofluorescence indicated APOBEC-3F and-3G were located in the cytosal.APOBEC-3F and-3G could inhibit HBV replication effectively in HepG2.2.15 cell.APOBEC-3F and-3G could not be trans-located into nuclear by nuclear location signal(NLS) or bi-NLS(B-NLS).These results will help the future research on the function of APOBEC.

OBJECTIVETo analyze a fetus with abnormal cardiac ultrasound by using various techniques and explore its genotype-phenotype correlation.

METHODSLymphocytes derived from umbilical cord blood sample were subjected to G-banding analysis. Short tandem repeats quantitative fluorescence PCR (STR-QF-PCR) was used for analysis of fetal DNA as an auxiliary test. Low-coverage whole genome sequencing (WGS) was used to detect chromosomal deletion/duplication which exceeded 100 kb in size.

RESULTSThe karyotype of the fetus was 47,XN,+mar. As detected by STR-QF-PCR, the copy number of GATA178F11 locus on chromosome 18 was 4, and the duplicated fragment was derived from the mother. WGS suggested that the fetus to be 46,XN,dup(18p11.21p11.32).seq [GRCh37/hg19](10 001-15 378 887)× 4, with the duplicated fragment spanning approximately 15.38 Mb.

CONCLUSIONThe cardiac malformation of the fetus may be attributed to the partial duplication of chromosome 18p. Combined cytogenetic and molecular methods can facilitate prenatal detection of genetic abnormalities.