Diagnosis of one case of Williams-Beuren syndrome presenting with hypothyroidism by low-coverage massively parallel CNV sequencing / 中华内分泌代谢杂志

Objective@#To explore the clinical phenotypes and the genetic causes for a 5 years old boy with unexplained growth retardation, developmental delay, special face, and hypothyroidism.@*Methods@#Routine G-banding was performed to analyze the karyotype of the patient and his parents. In addition, whole exome sequencing and low-coverage massively parallel CNV sequencing (CNV-seq) were used to determine the potentially pathogenic variants as well as the copy number variations (CNVs).@*Results@#The child′s karyotype was 46, XY, and his parents′ karyotypes were normal.However, CNV-seq identified a heterozygous deletion of 1.56 Mb on chromosome region 7q11.23 in the patient, including 24 protein-coding genes, which were associated with Williams-Beuren syndrome. His parents′ results of CNV-seq were normal, indicating a de novo CNVs.@*Conclusion@#A Williams-Beuren syndrome child presenting with hypothyroidism was diagnosed by CNV-seq, which would contribute to further understanding the clinical phenotypes and pathogenesis of this disease.

Diagnosis of one case of Williams-Beuren syndrome presenting with hypothyroidism by low-coverage massively parallel CNV sequencing / 中华内分泌代谢杂志

Objective:To explore the clinical phenotypes and the genetic causes for a 5 years old boy with unexplained growth retardation, developmental delay, special face, and hypothyroidism.Methods:Routine G-banding was performed to analyze the karyotype of the patient and his parents. In addition, whole exome sequencing and low-coverage massively parallel CNV sequencing (CNV-seq) were used to determine the potentially pathogenic variants as well as the copy number variations (CNVs).Results:The child′s karyotype was 46, XY, and his parents′ karyotypes were normal.However, CNV-seq identified a heterozygous deletion of 1.56 Mb on chromosome region 7q11.23 in the patient, including 24 protein-coding genes, which were associated with Williams-Beuren syndrome. His parents′ results of CNV-seq were normal, indicating a de novo CNVs. Conclusion:A Williams-Beuren syndrome child presenting with hypothyroidism was diagnosed by CNV-seq, which would contribute to further understanding the clinical phenotypes and pathogenesis of this disease.

The effect of para-aortic lymph node metastasis on the resectability of pancreatic cancer / 中华外科杂志

Pancreatic cancer has poor prognosis and lymph node metastasis is a poor prognostic factor in patients with resectable pancreatic cancer. The metastatic prevalence of para-aortic lymph node (PALN) ranges from 9.1% to 26.5% and it is listed as the distant metastatic group in pancreatic cancer. Nevertheless, it is controversial whether PALN metastasis is the contraindication of surgery in resectable pancreatic cancer for the shortage of level Ⅰ evidence.This study concluded that PALN metastasis indicated poor prognosis in patients with pancreatic cancer, but some patients with PALN metastases could benefit from surgery and their survival could be much improved after the combination of surgery and adjuvant therapy. Therefore, it is not wise to refuse surgery for all pancreatic cancer patients with PALN metastasis and the clinicians can cautiously choose the patients to do surgery. Besides, there are mainly retrospective studies rather than prospective and multicenter studies to explore the prognosis of pancreatic cancer patients with PALN metastasis. Thus, more prospective and multicenter studies are needed to decide whether PALN metastasis is an independent prognostic factor in patients with resectable pancreatic cancer.

Polymorphisms in endothelial protein C receptor gene and Kawasaki disease susceptibility in a Chinese children / 中南大学学报(医学版)

Objective:To investigate association between the single nucleotide polymorphisms of endothelial protein C receptor (EPCR) gene and the risk of Kawasaki disease (KD) in a Chinese children.Methods:A total of 103 KD patients including 23 patients with coronary artery lesions (CAL) and 158 controls were recruited.Seven tagging SNPs (rs6088738,rs2069940,rs2069945,rs2069952,rs867186,rs9574,and rs1415774) of EPCR gene were selected for TaqMan allelic discrimination assay.The plasma soluble EPCR (sEPCR) levels of 53 KD and 52 healthy children were detected by ELISA.Results:We found a significant association between rs2069952,rs9574 or rs1415774 and higher probability for the occurrence of KD but not CAL formation.Interestingly,males with these 3 SNPs and rs2069945 SNPs bore a much greater risk of KD than females.The level of plasma sEPCR in children with KD didnot predict the formation of CAL.However,the allele G of rs867186 in EPCR was associated with the increased level of plasma sEPCR in KD patients.Conclusion:The SNPs of EPCR are associated with KD susceptibility in a Chinese Han children.

Disseminated cryptococcosis characterized by obstructive jaundice and convulsions: a case report / 临床儿科杂志

Objective To explore the diagnosis and treatment of disseminated cryptococcosis involving the biliary tract and the central nervous system. Methods The clinical data of one child with disseminated cryptococcosis involving biliary tract and central nervous system were analyzed retrospectively. The related literatures were reviewed. Results A 6-year-old boy presented with jaundice, hepatic dysfunction and convulsions. The surgical treatment was performed. The pathological biopsy showed cryptococcal infection, cryptococcal capsular polysaccharide antigen latex agglutination test in serum and cerebrospinal fluid were positive and cerebrospinal fluid ink staining was also positive. After antifungal treatment, the symptoms were relieved and no sequelae existed during the follow-up period. Conclusion Disseminated cryptococcosis simultaneously involving the biliary tract and the central nervous system is rare in children.

Interactions of highly pathogenic avian influenza H5N1 nucleoprotein with NF-κB-inducing kinase(NIK) / 军事医学

Objective To study the interactions between the highly pathogenic avian influenza H5N1 nucleoprotein (H5N1 NP) and NF-κB-inducing kinase (NIK),and to reveal the effect of H5N1 NP on NIK-induced NF-κB transcriptional activity.Methods The gene encoding NIK protein was amplified by RT-PCR from total RNA of HeLa cell line.Eukaryotic expression plasmid pCMV-Myc-NIK and prokaryotic expression plasmid pGEX-4T-1-NP (GST-NP) were constructed by cloning from HeLa cell cDNA and pcDNA3-Flag-NP vector,respectively.Co-immunoprecipitation (co-IP) and GST pull-down were used to test the interactions between H5N1 NP and NIK.Dual-luciferase reporter gene analysis system was used to test the effect of H5N1 NP on NIK-induced NF-κB transcriptional activity.Results Co-IP and GST pull-down showed that pCMV-Myc-NIK and pGEX-4T-1-NP (GST-NP) could express Myc tagged NIK protein and GST tagged NP protein in HEK293T cells and E.coli,respectively,and that H5N1 NP was associated with NIK in vivo and in vitro.Dual-luciferase reporter gene analysis suggested that H5N1 could inhibit NIK-induced NF-κB transcriptional activity.Conclusion H5N1 NP interacts with NIK and inhibits NIK-induced NF-κB transcriptional activity.This finding can facilitate further study of H5N1.

Disseminated cryptococcosis characterized by obstructive jaundice and convulsions: a case report / 临床儿科杂志

Objective To explore the diagnosis and treatment of disseminated cryptococcosis involving the biliary tract and the central nervous system. Methods The clinical data of one child with disseminated cryptococcosis involving biliary tract and central nervous system were analyzed retrospectively. The related literatures were reviewed. Results A 6-year-old boy presented with jaundice, hepatic dysfunction and convulsions. The surgical treatment was performed. The pathological biopsy showed cryptococcal infection, cryptococcal capsular polysaccharide antigen latex agglutination test in serum and cerebrospinal fluid were positive and cerebrospinal fluid ink staining was also positive. After antifungal treatment, the symptoms were relieved and no sequelae existed during the follow-up period. Conclusion Disseminated cryptococcosis simultaneously involving the biliary tract and the central nervous system is rare in children.

Construction and preliminary study of biological functions of O157∶H7 typeⅢsecre-tion system effector NleF gene knockout mutant / 军事医学

Objective To construct Escherichia coli O157∶H7 T3SS effector NleF gene knockout mutant and its com-plementary strain, and probe its effects on bacterial growth and cell death .Methods T3SS Effector NleF gene knockout mutant ΔnleF was constructed with λ-Red homologous recombination .Complementary strain ΔnleF/NleF was constructed by transferring pET-24a(+)-NleF into ΔnleF competent cells.Wild type,ΔnleF and ΔnleF/NleF were cultured in LB and DMEM(10%FBS) respectively,D600 was measured every hour , and the growth curve was drawn .HeLa cells were infected with three kinds of strains , the supernatant of LDH release was detected with cytotoxicity detection kit ,and the cytotoxicity was calculated .Results ΔnleF and ΔnleF/NleF were constructed .The growth rates of wild type , ΔnleF and ΔnleF/NleF was not significantly different .Wild type O157 infection induced cell death .Cytotoxicity was increased as much in ΔnleF in-fected cells as in ΔnleF/NleF infected cells.Conclusion EHEC O157∶H7 T3SS Effector NleF has no significant effect on bacterial growth ,but might inhibit host cell death caused by bacterial infection .

Constructing nurse′s ability of scientific research assessment system / 中国实用护理杂志

Objective To construct a scientific and objective assessment to value nurse′s ability of scientific research in order to provide an objective unified standard and operable method to value nurse′s ability of scientific research and provide a standard for medical institutions to select nurses of scientific research. Methods After searching literatures and interviewing 7 experts, Delphi method was used to construct primary variables of nurse′s ability of scientific research. Then a second interview was given to each of 18 experts. The variables were weighted using analytic hierarchy process. Results Variables to assess nurse′s ability of scientific research included 2 first grade indices, 9 second grade indices and 35 third grade indices. Questionnaire return rates for two rounds of interviews were 84.00%(21/25) and 85.71%(18/21), respectively. Degree of authority Cr was 0.938. Scores of variables′importance ranged at 4.06-4.96. Ratios of full scores ranged at 33.33%-94.44%. Variable coefficients ranged at 0.05-0.25. The weight coefficients of 3 first grade indices (scientific research accumulation, explicit ability of scientific research, and intrinsic ability of scientific research) were 0.475 and 0.525, respectively. Conclusions The nurse′s ability of scientific research assessment system based on Delphi method and analytic hierarchy process is scientific and objective. This system combines quantitative and qualitative variables. It thus can calculate the ability of nurse′s scientific research according to the three grade indices. It is operable to be used to assess the ability of nurse′s scientific research.

Effect of typeⅢsecretion system deficient mutant of E.coli O157∶H7 on attachment of HeLa cells / 军事医学

Objective To construct the typeⅢ secretion system (T3SS) deficient mutant of O157∶H7 EDL933, and to detect its ability of attachment after infection with HeLa cells.Methods The recombinant DNA fragments obtained kana-mycin resistant gene were constructed by overlap extension PCR and transferred into EDL933/pKD46 to replace escR gene by homologous recombination.Results The T3SS-deficient mutant (ΔescR) was successfully construted.Compared with the wild-type EDL933, the growth rate and attachment on HeLa cells of ΔescR were significantly decreased.Conclusion The deletion of escR gene, which encodes T3SS structural protein EscR, affects the attachment of EDL933, suggesting a better basis for future studies of T3SS.

Prokaryotic expression, purification, identification, and polyclonal antibody prepa-ration of enterohemorrhagic Escherichia coli effector NleB1 / 军事医学

Objective To construct a prokaryotic plasmid expressing the recombinant protein of enterohemorrhagic Escherichia coli(EHEC) effector NleB1 and to prepare the polyclonal antibody of mouse anti-NleB1.Methods The nleB1 (990 bp) gene was amplified from the genome EHEC O157∶H7 and cloned into the expression plasmid pET24a to construct the recombinant plasmid pET24a-nleB1 that was transformed into E.coli BL21(DE3).After induction with isopropylthio-gelactoside( IPTG) , the His-tag fusion proteins were purified by Ni+affinity chromatography and gel slices.The polyclonal antibody was prepared by immunizing BALB/c mice with purified recombinant proteins and analyzed by Western blotting and ELISA.Results The pET24a-nleB1 recombinant plasmid was successfully constructed, the fusion protein was ex-pressed and purified,and the polyclonal antibody was obtained by immunizing mice with purified fusion protein.Western blotting and ELISA staining demonstrated that the polyclonal antibody was successfully obtained.Conclusion The prepara-tion of the polyclonal antibody against EHEC O157∶H7 NleB1 will be of help for further studies on the function of NleB1 protein.

Biosynthesis of amorpha-4,11-diene, a precursor of the antimalarial agent artemisinin, in Escherichia coli through introducing mevalonate pathway / 生物工程学报

Artemisinin-based combination therapies (ACTs) are recommended to be the most effective therapies for the first-line treatment of uncomplicated falciparum malaria. However, artemisinin is often in short supply and unaffordable to most malaria patients, which limits the wide use of ACTs. Production of amorpha-4,11-diene, an artemisinin precursor, was investigated by engineering a heterologous isoprenoid biosynthetic pathway in Escherichia coli. The production of amorpha-4,11-diene was achieved by expression of a synthetic amorpha-4,11-diene synthase gene in Escherichia coli DHGT7 and further improved by about 13.3 fold through introducing the mevalonate pathway from Enterococcus faecalis. After eliminating three pathway bottlenecks including amorpha-4,11-diene synthase, HMG-CoA reducase and mevalonate kinase by optimizing the metabolic flux, the yield of amorpha-4,11-diene was increased by nearly 7.2 fold and reached at 235 mg/L in shaking flask culture. In conclusion, an engineered Escherichia coli was constructed for high-level production of amorpha-4,11-diene.