The ribosomal DNA fragment was amplified with PCR utilizing synthetic primers with host bacteria chromosomal DNA as template. The resultant probes were labeled with Digoxigenin and were applied in the dot blot of DNA vaccine samples. The host bacteria genomic DNA in DNA vaccine against hepatitis B was less than 15 ng/?g. Digoxigenin labeled probes proved sensitive and reliable in determining genomic DNA.

OBJECTIVE: To observe the protective effect of Huaxia shallot preparation on human umbilical vein endothelial cell (HUVEC) injury induced by oxidized low density lipoprotein (Ox-LDL) in vitro. METHODS: Ox-LDL was prepared and identified, and HUVECs were cultured. After 2-hour intervention of different drugs and 24-hour following intervention of Ox-LDL, the number of HUVECs was observed by phase contrast optical microscope and the activity of the HUVECs was observed by methyl thiazolyl tetrazolium (MTT) technique. Superoxide dismutase (SOD) activity and nitric oxide (NO) content were assayed by respective kit. The protein expressions and mRNA levels of peroxisome proliferators activated receptor gamma(PPAR-gamma) and endothelial nitric oxide synthase (eNOS) were measured by western blot technique and reverse transcription polymerase chain reaction (RT-PCR). RESULTS: Ox-LDL could increase the apoptosis rate of the HUVECs and decrease the NO release as compared with the blank control group (P<0.05). These effects induced by Ox-LDL were all significantly inhibited by Huaxia shallot preparation. It could up-regulate the protein expressions and mRNA levels of PPAR-gamma and eNOS significantly (P<0.05). Huaxia shallot preparation could decrease the apoptosis rate of the HUVECs. CONCLUSION: Ox-LDL may be involved in the initiation and progression of atherosclerosis by injuring the endothelial cells directly and may cause the endothelial dysfunction. Huaxia shallot preparation can protect against Ox-LDL induced endothelial cell injury by up-regulating the protein expressions and mRNA levels of PPAR-gamma and eNOS. It suggests that Huaxia shallot preparation may play a role in the prevention and treatment of cardiovascular disease.

Objective To investigate the effects of protocadherin 20 (PCDH20) in the spinal cord in the development of bone cancer pain (BCP) in rats.Methods Thirty-six SPF female Sprague-Dawley rats,weighing 180-200 g,were randomly divided into 4 groups (n =9 each):sham operation group (group S),BCP group,lentivirus control group (group LC) and PCDH20 siRNA lentivirus group (group P).Control lentivirus and lentivirus containing PCDH20 siRNA 4 μl were injected into the ipsilateral spinal cord in groups LC and P,respectively.One week later,BCP was induced by injection of Walker 256 breast cancer cells into the upper segment of bone marrow of right tibia.The mechanical paw withdrawal threshold (MWT) was measured at 1 day before injection of lentivirus (T1),1 day before BCP (T2),and 7,14 and 21 days after BCP (T3-5).Three rats in each group were sacrificed after measurement of the MWT at 21 day after BCP and the tibia on the operated side was obtained for examination of invasion of the cancer cells with light microscope.The spinal cord was removed for determination of the expression of PCDH20 and postsynaptic density 95 (PSD95) protein (by Western blot) and mRNA (by RTPCR).Results In groups BCP,LC and P,the cancer cells grew out of the bone and destroyed the cortical bone seriously.Compared with group S,the MWT was significantly decreased at T3-5 in groups BCP,LC and P,the expression of PCDH20 and PSD95 protein and mRNA was up-regulated in groups BCP and LC,and the expression of PCDH20 was up-regulated in group P (P ＜ 0.05).Compared with BCP group,no significant change was found in the MWT and expression of PCDH20 and PSD95 protein and mRNA in group LC (P ＞ 0.05),and the MWT was significantly increased at T4,5 and the expression of PCDH20 and PSD95 protein and mRNA was down-regulated in group P (P ＜ 0.05).Conclusion PCDH20 is involved in the development of BCP through regulating the expression of PSD95 in the spinal cord and adjusting the function of excitatory synapse in rats.

Objective To study the effects of monosialoganglioside combined with edaravone in the treatment of cerebral ischemic neuronal injury in rats.Methods Healthy SD rats were randomly divided into model group and experimental group (monosialin ganglioside 20 mg · kg-1 + edaravone 3 mg · kg-1),each group of 21 rats.The experimental group was injected one,twice a day respectively.The model group using the same amount of 1％ saline,the two groups were administered 14 days.Animal model of focal cerebral ischemia-reperfusion was made by middle cerebral artery occlusion.Using the image analysis system,according to the method of positioning the cerebral ischemic semi-dark bag in the same field of vision,the upper middle cerebral artery occlusion of the same side of the parietal lobe of the upper part of the cortex is defined as a semi-dark band.The silt was observed by immunity.Comparison of the average absorbance and positive area of 3-phosphoinositidedependent(PDK1),phosphatidylinositol-3-kinase (PI3 K),serine-threonine protein kinase (Akt) and glycogen synthase kinase-3β (GSK-3β) protein immunoreactive cells in histochemical method.Results The expression of the PI3K protein,Aktprotein,PDK1 protein were increased while expression of the GSK3β protein was decreased in the model group and the experimental group.After 7 days,positive unit area of PI3K protein,Aktprotein,PDK1 protein and GSK3β protein in the model group and the experimental group were (19.58 ± 1.13),(26.91 ± 0.90) mm2;(15.98 ± 0.48),(22.87 ± 1.20)mm2;(17.97 ± 0.58),(26.02 ± 0.54) mm2;(22.03 ± 0.92),(14.02 ± 0.45) mm2,the difference between two groups was significant (all P ＜ 0.05).The average absorbance density of the four protein in the model group and the experimental group were 0.19 ±0.01,0.28 ±0.02;0.16 ±0.01,0.25 ±0.01;0.16 ±0.01,0.25 ±0.01;0.22 ±0.01,0.14 ±0.01,the difference between two groups was statistically significant (all P ＜0.05).Conclusion Single sialic acid ganglioside combined with edaravone against cerebral ischemia injury of nerve cells in the region.

Objective:To determine risk factors for postoperative esophageal refractory stenosis after endoscopic submucosal dissection (ESD) of large-scale early esophageal carcinomas and precancerous lesions.Methods:Two hundred and twelve early esophageal carcinomas or precancerous lesions in 186 patients who underwent ESD larger than 3/4 the total esophageal circumference in Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, between July 2013 and December 2017 were divided into two groups according to session number of endoscopic balloon dilatation (EBD), the refractory stenosis group ( n=69, ≥6 EBD sessions) and non-refractory stenosis group ( n=117, ≤5 EBD sessions). Student′s t-test or Mann-Whitney U test was used for univariate analysis and χ2 test and Fisher exact test were used for comparison of categorical variables. Logistic regression was used for multivariate analysis. Results:Compared with the non-refractory stenosis group, the refractory stenosis group had statistically significant differences in the longitudinal diameter of lesions, the longitudinal diameter of artificial ulcer, lesion location, the circumferential range of lesions and the composition of the muscular layer injury (all P<0.05). After eliminating the factor of the vertical diameter of artificial ulcer (because there was significant correlation between the vertical diameter of artificial ulcer and the longitudinal diameter of lesion in clinical practice), multivariate logistic regression analysis showed that the longitudinal diameter of lesion>5 cm (VS ≤5 cm: P=0.003, OR=3.531, 95% CI:1.547-8.060), the location of lesion in the upper thoracic segment (VS lower thoracic segment: P=0.001, OR=36.720, 95% CI:4.233-318.551), in the cervical segment (VS lower thoracic segment: P=0.003, OR=24.959, 95% CI:2.927-212.795), the whole circumferential lesion (VS ≥3/4 but not the whole circumference: P<0.001, OR=10.082, 95% CI:4.196-24.226) and the presence of muscular layer injury ( P<0.001, OR=7.128, 95% CI:2.748-18.486) were more likely to lead to esophageal refractory stenosis after ESD. Conclusion:The longitudinal lesion diameter of more than 5 cm, the circumferential extent of esophageal ESD, cervical or upper-thoracic esophageal lesions, and muscular layer damage are independent risk factors for postoperative esophageal refractory stenosis after ESD for large-scale esophageal cancer and precancerous lesions.

The provenances of Rongshui miniature pigs ( RMPs) were purchased from Rongshui, Guangxi, in 2012.130 RMPs were transported to Sanshui, Guangdong,China, which were breed according to the laboratory animal standards.83 RMPs were selected randomly from the first filial generations ( F1 ).The basic data were collected including breed characteristics, reproductive performance, grow curve, hematology, biochemical markers of serum and urine, organ coefficient, Chromosome analysis.According to the national and local standards, the quality control standards of RMP were set up including microbiological, parasitic, environment and facilities, fodder, pathology, genetic.The results showed that RMPs adapt to the climate of Guangdong.The natural mating and conceive rate was 88.3% with the pregnancy of 112 days.The average number of firstborn and still-born was 6.1 and 7.9 respectively.RMP was small body size with the adult body weight of 17.21 ±5.20 kg and 16.35 ±5.23 kg in female and male respectively.RMP was very tame.The mitochondrial genome analysis suggested RMP belonged a typical miniature pig breed in China, which is ancient than Lanyu pig.RMP could be breed as a new kind of laboratory animal.

Objective To investigate the isolation rate, distribution and trend of nontuberculous mycobacteria (NTM) in Wenzhou during 2014 to 2016.Methods Sputum or alveolar lavage specimens of patients with suspected pulmonary tuberculosis were collected for mycobacteria culture from January 2014 to December 2016.Mycobacterium culture positive strains were further identified by gene chip, 16S rRNA and hsp65 gene sequencing.SPSS 19.0 software was used to analyze the data.Results After excluding repetitive strains, 3 295 mycobacteria strains (MTB) were isolated from respiratory specimens, included 3 032 mycobacterium tuberculosis complex strains, 238 NTM strains, 20 Gordon genera strains, 3 Nocardia genera strains and 2 Tsukamurella genera strains.The proportion of NTM among confirmed mycobacteria was 8.5％ (86/1 006), 6.7％ (72/1 079) and 6.8％ (80/1 185) in 2014, 2015 and 2016, respectively (x2 =2.459,P ＞ 0.05).The overall prevalence of NTM was 7.3 ％ (238/3 270).There were 15 species of NTM, and the most common NTM strain was Mycobacterium intracellulare (52.5％,125/238), followed by Mycobacterium abscessus (22.7％, 54/238) and Mycobacterium avium (10.1％, 24/238), other species were only accounted for 14.7％ (35/238).The ranking of Mycobacterium avium went up rapidly from the fifth in 2014 to the second in 2016 (x2 =18.259, P ＜ 0.01), while proportion of Mycobacterium abscess, dropped from 34.9％ (30/86) in 2014 to 17.5％ (14/80) in 2016 (x2 =7.335, P＜0.01).Among patients from whom the NTM strains were isolated, 56.7％ (135/238) were male and most of them were aged 45 years or above (79.8％, 190/238).Conclusions In the past three years, the trend of NTM isolation rate in Wenzhou is steady.The most prevalent NTM species is Mycobacterium intracellulare, followed by Mycobacterium abscessus and Mycobacterium avium.Mycobacterium avium shows a continuously upward trend, while the separation of Mycobacterium abscessus shows a downward trend.

Objective To investigate plasma glutathione S-transferase(GSTs) activity and GSTP1 gene Ile105Val polymorphism in Bijie City, Guizhou Province, a coal-burning fluorosis endemic area. Methods One hundred and sixty villagers from Yachi Twon using non-improved cooking stoves were selected as the non-intervened group in Bijie City, Guizhou Province where coal-burning fluorosis was prevailing; 153 villagers as the intervented group were chosen from Changchun Twon, where cooking stoves were improved; 151 villagers were served as the control group from Baiyunshan Twon, Changshun County without endemic fluorosis. The activity of GSTs was tested by colorimetric analysis with spectrophotometer. The genotype of the GSTP1 gene Ile105Val polymorphism, presenting as either homozygous wild-type (AA), or heterozygous mutation type (AG), or homozygous mutation type (GG), was detected through the PCR-RFLP procedure. Results The activity of GSTs in plasma of non-intervened group [(12.44±4.97) kU/L]was significantly lower than that of intervened group (P < 0.05), and that of intervened group[(20.78±6.20)kU/L]was significantly lower than that of control group[(24.30±6.27)kU/L, P< 0.05]. The difference of the enzyme activity of three groups were statistically significant (F = 51.71, P < 0.05), but this enzyme activity did not vary significantly in each sex of each grnup(P > 0.05). Compared intervened group [AA:67.3%(103/153), AG:29.4%(45/153),GG:3.3%(5/153)]and non-intervened group[AA:66.9%(107/160), AG:30%(48/160), GG:3.1%(5/160)]with control group[AA:74.8%(113/151), AG:25.2%(38/151), GG:0 (0/151)], the Ile105Val polymorphism site of GSTP1 gene had significant difference(χ2= 6.04,6.07, both P< 0.05), but not significant between intervened and non-intervened groups(χ2 = 0.02, P>0.05). Conclusions Fluorosis can decrease the activity of GSTs and introduce the GSTP1 gene Ile105Val polymorphism, intervention with the fluorine intake will improve the effect of fluoride on the body.

Objective: To study the recombinant epitope antigens of hepatitis C virus (HCV), in order to fulfil the requirements of recombinant immunoblot assay kit. Methods: An expressing vector pBVIL1 for expression of recombinant antigens in a fusion manner with IL-1β was constructed. A series of selected genes from the HCV antigens including the C, NS3, NS4 and NS5 were amplified from HCV gene-containing plasmids using PCR and the expression plasmids for these genes were constructed in pBVIL1, respectively. The activity of the purified recombinant antigens were tested against an identified HCV antibody positive and negative panel with ELISA. Results and Conclusions: All the cloned genes of chosen antigen epitopes were highly expressed in pBVIL1 in E.coli. The activity of the C and NS4 antigens were slightly higher than the RIBA3.0 antigens, while the activity of NS3 was slightly lower than the RIBA3.0 antigen. But the total evaluation for the panel was same as RIBA3.0. That means the cloned antigens were suitable for the use in RIBA test kit.

Objective · To identify broad-spectrum bacteriophages against extensively drug-resistant Klebsiella pneumonia and analyze their characteristics by biological and genomic methods. Methods · Multi-drug resistant Klebsiella pneumonia strains collected from a hospital were used as host bacteria to isolate and purify broad-spectrum phages in the wastewater at the same hospital area. The size and shape of phages were observed by transmission electron microscope. Titer, host range, pH stability and thermal stability were measured. Moreover, the DNA extracted from the phage SH-Kp152234 was sequenced and analyzed. Results · One strain of bacteriophage against Klebsiella pneumonia was isolated and named as SH-Kp152234. The electron microscope revealed it belongs to Podoviridae family. Moreover, genome of SH-Kp152234 showed to be a linear double-stranded DNA of 40578 bp with the GC content of 52.85%. It was predicted to have 49 open reading frames with related known functions.Conclusion · SH-Kp152234, with a broad host range and a short latent period, which could exert its activity in a wide range of temperature and pH, is a promising candidate to be exploited in the treatment of multiple drug-resistant Klebsiella pneumonia.