ABSTRACT
【Objective】 To determine molecular basis of a rare HLA-A typing results carrying triple A alleles in potential allo-HSCT donor and her family. 【Methods】 HLA-A, -B, -C, -DRB1, -DQB1, -E, -F, -G of 5 members in the family were genotyped at a high-resolution level using next-generation sequencing (NGS). HLA-A of probosita was re-checked using polymerase chain reaction-sequence-based typing (PCR-SBT), and SNP oligonucleotide probes (SNP-array)were scanned with genomic DNA of probosita. 【Results】 There was 162.9Kb duplication in 6p22.1(29, 803, 377-29, 966, 301)of probosita who carried triple A alleles A*02∶01∶01, A*11∶01∶01, A*24∶02∶01. Other two family members were found to carry this haplotype: A*02∶01∶01, A*24∶02∶01, B*54∶01∶01, C*01∶02∶01, DRB1*04∶05∶01, DQB1*04∶01∶01, E*01∶01∶01∶03, F*01∶01∶01, G*01∶01∶01∶01, which as a Mendelian gene was segregated and stably transmitted through two generations. 【Conclusion】 Tiny gene duplication induces one haplotype carries two HLA-A alleles in a potential healthy donor for allo-transplantaion and stably transmits through two generations.Routine HLA typing laboratories should pay more attention to this situation and accurately report.
ABSTRACT
Japanese encephalitis virus (JEV) is a single-stranded and positive-sense RNA, which has a single ORF (open reading frame), encoding a polyprotein precursor. Non-structural protein 3 (NS3) plays an important role in processing the polyprotein precursor and has become an important drug target of flavivirus. In this study, NS2BH-NS3 gene was amplified by PCR and subcloned to the prokaryotic expression plasmid, resulting pET30a-NS2BH-NS3. The fusion protein was expressed in Escherichia coli BL21 (DE3) in soluble form after induction by Isopropyl beta-D-1-Thiogalactopyranoside (IPTG). The recombinant protein was purified by Ni-NTA affinity column. Then a fluorescence resonance energy transfer (FRET) method was used to determine enzymatic activity and the assay conditions were optimized. After screening 113 compounds, we found two compounds inhibiting the activity of NS2BH-NS3. This study provides a convenient and cost-effective method for screening of JEV NS3 protease inhibitor.
Subject(s)
Encephalitis Virus, Japanese , Escherichia coli , Metabolism , High-Throughput Screening Assays , Protease Inhibitors , Chemistry , RNA Helicases , Metabolism , Recombinant Fusion Proteins , Metabolism , Serine Endopeptidases , Metabolism , Viral Nonstructural Proteins , MetabolismABSTRACT
Background:Capsule endoscopy( CE)has been widely used for the diagnosis of small intestinal diseases. How to enhance the complete examination rate( CER)has attracted more and more attention in recent years. Aims:To investigate the effect of different intervention on gastric transit time( GTT)and small bowel transit time( SBTT)in CE. Methods:Ninety patients undergoing CE from January 2012 to May 2013 at Changhai Hospital were enrolled and randomly divided into control group,right lateral position group and motion group. The patients in control group were allowed to keep quiet, patients in right lateral position group were asked to lie on their right side,and patients in motion group were asked to take rapid pace walking after swallowing the capsule until the capsule passing pylorus. GTT,SBTT,CER and disease diagnosis rate among the three groups were compared. Results:Compared with control group,GTT was significantly decreased in right lateral position group(P﹤0. 05),however,no significant difference in SBTT was found between right lateral position group and control group(P ﹥0. 05). GTT and SBTT were both significantly decreased in motion group than in control group(P﹤0. 05). No significant differences in CER and disease diagnosis rate were found among the three groups(P﹥0. 05). Conclusions:Right lateral position can shorten GTT,rapid pace walking can shorten GTT and SBTT,and both these two intervention have substantial clinical values in CE.
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Objective To investigate the feasibility,safety,and short term efficacy of endoscopicultrasound guided radiofrequency ablation (EUS-RFA).Methods Three patients with advanced,inoperable pancreatic carcinoma underwent EUS-RFA.Under the guidance of EUS,a 22 gauge fine needle was punctured into the pancreatic carcinoma,and 1 Fr radiofrequency (RF) catheter was placed into the cancer through this 22 gauge needle.RFA was applied at 10 watts for 2 minutes,and then 15 watts for another 2 minutes.The second needle's tract was selected and punctured about 1 ～ 1.5 cm apart from the first needle tract,and the ablation was repeated.Results The average age of these 3 patients was 63 years old.Two patients were diagnosed to have pancreatic tail cancer and one was pancreatic body cancer,and the mean diameter of the tumor was 3.6 cm.Pre-operative EUS-FNA confirmed the presence of malignant cells.EUS-RFA was performed three times at an interval of 2 weeks in the first patient and one time in the last two patients.The average session of EUS-RFA of each lesion was 3.67 times.After 2 weeks,EUS showed that the diameter of the lesions was reduced by 13.9％ on average.There were vacuolar degenerations of variable sizes in those lesions.The serum level of CA19-9 was averagely decreased by 46.5％.Abdominal pain was not obviously aggravated.There were no episodes of pancreatitis,perforation or bleeding within 48 hours after the procedure.The mean follow-up time was 49 days,and no complications occurred.Conclusions EUS-RFA can reduce the diameter of pancreatic carcinoma and decrease the serum level of CA19-9,and it is feasible and safe.
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Objective To evaluate significance of the quantification of bcr-abl mRNA in diagnosis and therapy of chronic myeloid leukemia (CML),essentiality significance for monitoring minimal residual disease. Methods Bcr-abl mRNA of 518 CML patients were detected using real-time PCR. Results Expression of bcr-abl mRNA was gradually increased among blastic phase (BP) (12.6 %),accelerated phase (AP) (25.4 %) and chronic phase (CP) (57.2 %) (P<0.05). Quantification of bcr-abl mRNA was cut down gradually after allotransplantation in the patients and becomes normal after treatment for 6 months. But quantification of bcr-abl mRNA inpatients treated with imatinib mesylate became normal after 12 months. Conclusion Real-time PCR was reliable and can be used for diagnosis,monitoring the treatment outcome,detecting the minimal residual disease,and predicting blast crisis.
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Objective To evaluate the status of P-selectin and platelet activating factor(PAF) in vivo in patients with deep venous thrombosis(DVT) and to observe their changes under interneving of medicines.Methods P-selectin and PAF of fourty patients and twenty normal subjects were fluorescence labled with corresponding monoclonal antibodies by flow cytometry( FCM ) and immunologic method respectively. Results P-selectin and PAF in patients with DVT were higher than that in normal subjects in early period of the disease, and they significantly decreased in different time after patients were treated. P-selectin was significantly different between patients who received sodium ozagrel treatment and those who not (P ＜0.05 ), but PAF was similar( P ＞ 0. 05 )after fourteen days. One month later, P-selectin and D-dimer in DVT patients were lower than before. However, the positive rate of P-selectin of DVT was still higher than normal subjects. Conclusions The platelet is activated in vivo in patients with DVT, so does fibrinolysis. Sodium ozagrel can decrease activity of platelet. P-selectin and PAF may be used diagnostic markers. Post-discharge patients are still at high-risk and must be regularly followed-up.