Vibrio parahaemolyticus has been considered as one of the most important foodborne bacterial pathogens.The loop-mediated isothermal amplification(LAMP) that amplifies DNA with high specificity and rapidity under an isothermal condition was applied for rapid detection of this pathogen for the first time.A set of four primers,two outer and two inner primers,was designed specifically to recognize the thermolabile hemolysin gene(tlh) of V.parahaemolyticus.The LAMP reaction mix was optimized.The most optimal reaction temperature and time of the LAMP assay for the tlh gene were 60℃ and 60min,respectively.Genomic DNAs from 28 bacterial strains including 14 V.parahaemolyticus strains were amplified using LAMP,and no amplicon was observed in other bacterial strains.The detection limit of this LAMP assay was around 90 fg of V.parahaemolyticus genomic DNA and 24 colonies forming units for pure cultures.In addition,this method was applied to detect artificially contaminated food samples,and the detection limit was 89 cfu/g for non-cultured artificially contaminated food samples.These results suggested that detection of V.parahaemolyticus by LAMP is an effective and low-cost procedure with high specificity and sensitivity that requires no specialized equipment.This assay is expected to become a valuable tool for rapid detection and identification of V.parahaemolyticus.

Objective To investigate the change of CD8+T lymphocyte non-cytotoxic antivirus response(CNAR)in slow progressors infected by HIV.Methods Applying with density gradient and immunomagnetic beads methods to purify the CD4+T lymphocyte from the healthy person and CD8+T lymphocyte from HIV-infected individuals.The CD4+T cell was infected by HIV(SF-33)virus and cocuhured with CD8+T cell.The culture supernatant was collected and the p24 value was detected by ELISA method.Results Our study showed that the CNAR function decreased by turns of slow progressors(SP),typical progressors(TP),health control group and AIDS group.There was significant difference between groups(P<0.01).We found a significant positive correlation between the CIM+T cell ture count and the CNAR function.The virus load didn't statistically correlate with the CNAR function.Conclusion The CNAR function possibly protected the HIV-infected individuals from progression.

Objective To investigate the underlying mutation in a late-onset Chinese patient with classic Bartter syndrome. Methods The mutation analysis of CLCNKB gene was performed by the PCR direct sequencing. The patient's parents and siblings were studied as well. Fifty normal volunteers were analyzed as control group. Results The heterozygous deletion mutation cDNA 753delG and heterozygous missense mutation G433E were detected in the patient. Her father was found to carry heterozygous G433E and her mother to carry cDNA 753delG mutation respectively. Her brother carried heterozygous G433E and her sister was normal. Conclusions Two mutations of the CLCNKB gene in this Chinese patient with late-onset classic Bartter syndrome are identified. The cDNA 753delG mutation has not been reported previously.

objective To investigate the clinicopathological features of mini-cancer of the stomach. Method In this study,out of 296 early gastric cancer cases there were 34 cases of early gastric cancer in which tumor diameter was≤10 mm,among those there were 5 cases with tumor size≤2 mm and 29 cases of the size 2～10 mm. Result Mini-cancer accunted for 2% of all early gastric cancers in this series:All these mini-cancers were of intramucosal cancer(100%),while that took up to 45%in control group in which tumors were between≥2 mm and≤10 mm:Tumors were high or moderately differentiated pathologically in 100%of mini-cancers and 55%in control group.None of mini-cancer patients had lymph node metastasis,however,1 of 29 patients in control group had lymph node metastasis.Both groups had no blood vessel and lymphatic vessel invasion:The differentiation concordance rate between superficial lesions and invasive fronts in mini-cancer was 100%,higher than 86%in control group. Conclusion Gastric mini-cancer is usually of high differentiation,low tumor invasion and low rate of lympy node metastasis than control group.Endoscopic therapy is applicable for most gastric mini-cancers.

Denys-Drash Syndrome ; diagnosis ; Humans ; Infant ; Male

AIM: To compare the effectiveness of botulinus toxin of type A and complete resection of the periorbital muscle on idiopathic blepharospasm.
● METHODS: Patients with idiopathic blepharospasm and having undergone either of two procedures from Dec. 2010 to Jun. 2015 were selected ( 60 patients ) . Among them, group A (30 patients, 60 eyes) underwent botulinus toxin of type A, group B (30 patients, 60 eyes) underwent complete resection of the periorbital muscle.
●RESULTS: ln group A, the patients with complete response, obvious response, partial response, and no response were 36(60. 0%), 20(33. 3%), 2(3. 3%) and 2 (3. 3%) cases respectively. ln group B, the patients with complete response, obvious response, partial response, and no response were 16(26. 7%), 24(40. 0), 12(20. 0%) and 8 ( 13. 3%) cases respectively. The difference was statistically significant ( Z = - 2. 968, P = 0. 003 ). The relapse rate of group A and group B were 93. 3% and 20. 0% after 6mo, the difference was statistically significant (χ2=32. 851, P<0. 001).
●CONCLUSION: The botulinus toxin injection of type A is effective for idiopathic blepharospasm. But recurrence rate is high after 6mo. Complete resection of the periorbital muscle have long-term efficacy for idiopathic blepharospasm. It′s a supplementary therapy after idiopathic blepharospasm recurrence.

Objective To perform a prenatal diagnosis for the second fetuses from 28 pedigrees with proband of mitochondrial disease due to mitochondrial DNA (mtDNA) mutation.Methods From April 2011 to November 2015,peripheral blood samples of 28 probands and their parents,urine samples of these probands and their mothers as well as amniotic fluid samples of the second fetuses from the 28 pedigrees were collected in Peking University First Hospital.DNA sequencing was used to identify mtDNA mutations.Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed to verify mutation sites,calculate mutation loads,and further confirm the diagnosis after birth.Microsatellite maker analysis was also performed on five short tandem repeats located in nuclear genes to exclude maternal contamination.Statistical analysis was carried out using independent t-test.Results In the 15 pedigrees carrying A3243G mutation,13 mothers and nine fetuses carried A3243G mutation.Neither the other two mothers nor their fetuses were positive for A3243G mutation.Among the 12 pedigrees with T8993G mutation,there were eight mothers carrying T8993G mutation and all of their fetuses carried the same mutation;and the other four mothers and their fetuses were negative for T8993G mutation.T10191C mutation was only found in one proband and the second fetus of that pedigree,but not in the mother.None of the fathers had mtDNA mutation.Results of PCR-RFLP were consistent with those of DNA sequencing.Short tandem repeat analysis demonstrated that amniocyte samples were from fetuses without maternal contamination.No mtDNA mutations were found in the six newborns who were negative for mtDNA mutations in prenatal diagnosis.The mean mutation load in urine samples of the six mothers without A3243G mutation in amniocytes was significantly lower than that of the nine mothers with A3243G mutation [(10.1 ±4.8) ％ vs (28.2 ± 15.1) ％,t=2.290,P=0.043].Conclusions The lower the mtDNA mutation load in maternal urine samples,the less the possibility she bears a child with mtDNA mutation.However,prenatal diagnosis of mitochondrial disease is necessary.

Objective To improve the understanding,the incidence and imaging findings of tuberous sclerosis complex (TSC) combined with cardiac rhabdomyomas (CRs) in fetuses and infants.Methods The imaging findings of 9 infants with TSC combined with CRs and 4 fetuses with TSC combined with CRs from our hospital between June,2006 and November,2013 were retrospectively reviewed.Results The brain MRI of 9 with TSC combined with CRs showed bilateral subependy-mal nodules,subcortical white matter and cortical tubers.Subependymal nodules were isointense or hypointense on spin-echo T1WI and hypointense or hyperintense on spin-echo T2WI.Subcortical white matter and cortical tubers were hypointense or hyperintense on T1WI and hypointense or hyperintense on T2WI.There was varying contrast enhancement.Three of 9 infants presented single cardiac tumor and 6 of 9 infants presented multiply cardiac lesions.CRs on contrast cardiac MRI showed round solid masses in ventricular septums,ventricular outflow tract,ventricle or atrial free walls.The masses were isointense relative to the cardiac muscles on T1WI,T2WI and B-TFE sequence.There was varying contrast enhancement.Four fetuses with TSC on ultrafast MRI showed bilateral multiply subependymal nodules,the nodules were isointense or hyperintense on TFE T1WI and isointense or hypointense signals on SSTSE or B-FFE sequence,Four fetuses with CRs showed isointense to hyperintense solid masses in ventricular septums on ultrafast MRI,ventricle or atrial free walls on B-FFE sequence and SSTSE sequence images.Conclusions TSC in infant and fetus is a kind of neurocutaneous syndrome,usually combines with CRs.Fetal ultrafast and routine MRI is a useful method to make a definite diagnosis for cranial and cardiac lesions.The development of MRI might improve the timeliness and accuracy of the assessment for this disease.

Objective To build the experimental basement for the clinical use of cytokines induced killer(CIK)cells from the cord blood mononuclear cells(CBMNC)in tumor adoptive cellular immunotherapy, an effective protocol for their proliferation in vitro and cytotoxicity of CIK cells was established.Methods The lymphocytes from umbilical cord blood were isolated by density gradient centrifugation and suspended in medium with CD_3 mAb,rIL-2,rIL-1 and IFN-? as inducing agents to prepare CIK cells.At the same time, the lymphokine activated killer(LAK)and CBMNC were set as controls,which were only added IL-2 and not any cytokines during the whole culture.The changes of CIK cells before and after induction were observed with microscope and the phenotypes of the cells were analyzed by using flow cytometry.The proliferation of CIK cells were determined by trypan blue exclusion assay and the cytotoxic activity to lung cancer cell were tested with MTF method.Results According to the experiment,combining use of four types of cytokines could generate a great deal of CIK cells possessing highly cytotoxicity.From day 5 CIK cells became to prolif- erate and reached the peak at day 14.During the whole period,the relative percentage of CD_3~+ CD_(56)~+ cells in- creased significantly.Compared with LAK cells,which reached the proliferation peak at day 7 and then showed no evident proliferation.The control cells(CBMNC)showed no evident change of phenotypes and proliferation.CIK cells showed a higher antitumor activity on the tumor cells than LAK cells and CBMNC in vitro.Conclusion Umbilical cord blood can generate a great deal of CIK cells combining used with cy- tokines.Compared with classic LAK cells,umbilical cord blood CIK cells have the advantages of rapid prolif- eration speed and powerful cytotoxicity.CIK cells will be promising as a new strategy for the adoptive cellular immunotherapy of tumor.

This study aims to investigate the preparation process and in vitro release behavior of artesunate polylactic acid microspheres, in order to prepare an artesunate polylactic acid (PLA) administration method suitable for hepatic arterial embolization. With PLA as the material and polyvinyl alcohol (PVA) as the emulsifier, O/W emulsion/solvent evaporation method was adopted to prepare artesunate polylactic acid microspheres, and optimize the preparation process. With drug loading capacity, encapsulation efficiency and particle size as indexes, a single factor analysis was made on PLA concentration, PVA concentration, drug loading ratio and stirring velocity. Through an orthogonal experiment, the optimal processing conditions were determined as follows: PLA concentration was 9. 0% , PVA concentration was 0. 9% , drug loading ratio was 1:2 and stirring velocity was 1 000 r x min(-1). According to the verification of the optimal process, microsphere size, drug loading and entrapment rate of artesunate polylactic acid microspheres were (101.7 +/- 0.37) microm, (30.8 +/- 0.84)%, (53.6 +/- 0.62)%, respectively. The results showed that the optimal process was so reasonable and stable that it could lay foundation for further studies.
Artemisinins ; chemistry ; Calibration ; Drug Compounding ; methods ; Lactic Acid ; chemistry ; Microspheres ; Polyesters ; Polymers ; chemistry ; Polyvinyl Alcohol ; chemistry