Objective To sequence and analyze the full-length genome of one HEV strain,W2-1 isolated from a sporadic hepatitis E patient hospitalized in 1999 in Xinjiang,China.Methods Nested RT-PCR assays with 4 sets primers were used to amplify the entire genome.The PCR products were purified and sequenced.The full-length genome was acquired by assembling the fragmental sequences using the DNAstar 5.01 software.The genome of W2-1 was analyzed by comparing with the reference HEVs from GenBank.Results The complete genome of W2-1 is 7212 nt in length,including three open reading frames (ORF1-3) with 5079,1980 and 345 nt respectively,27 nt 5'UTR and 83 nt 3'UTR,and a 3' poly A tail.Phylogenetic analysis based on full-length genome showed that W2-1 belonged to genotype 1,subtype 1b.W2-1 had high homology with the HEV strains isolated in the large hepatitis E epidemic in Xinjiang in 1987-1989,sharing 97.2％-98.5％ nucleotide identity in the full length genome.W2-1 also showed high homology with 1b strains isolated in China after 2000,with 97.6％-99.2％ nucleotide identity.The specific amino acid sites in ORF1-3 proteins that distinct between genotype 1 HEV and the potential zoonotic strains did not change in W2-1.Conclusion W2-1 belongs to subtype 1b.The study indicates subtype 1b HEV has been circulating in China in a long period after hepatitis E outbreak in Xinjiang in 1986-1989.The amino acids of ORF1-3 of subtype 1b are conserved.

Objective:To evaluate the value of combined measurement of urinary insulin-like growth factor-binding protein 7 (IGFBP7) and urinary metalloproteinase inhibitor-2 (TIMP-2) in the early diagnosis and prognosis of cardiac surgery-associated acute kidney injury (CSA-AKI).Methods:From March 2018 to June 2018, cardiac surgery patients admitted to the cardiac macrovascular surgery department of the First Affiliated Hospital of Nanjing Medical University were prospectively included, and the blood creatinine was monitored to observe the presence of acute kidney injury (AKI). The prognostic information of the patients was collected, including in-hospital dialysis, in-hospital death, complete recovery of kidney function at discharge, death in one year after surgery, and progression to chronic kidney disease. The levels of urine IGFBP7 and TIMP-2 at 6 h, 24 h and 48 h after cardiac surgery were detected by enzyme linked immunosorbent assay (ELISA), and the urine creatinine (Cr) was also measured. Moreover, receiver operating characteristic curves (ROC) were plotted and the areas under the curves ( AUC) were calculated to evaluate the predictive value and prognostic value of urinary [TIMP-2]·[IGFBP7] (T*I for short) and urine T*I/urine Cr 2 in CSA-AKI. Results:A total of 74 patients with age of (58.43±10.91) years old and 47 males, were enrolled in this study, of which 24 cases (32.4%) had AKI and 10 cases (13.5%) had stage 2-3 AKI. Compared with the non-AKI group, the AKI group had significantly higher levels of urine T*I levels at 6 h and 24 h (both P<0.05). The AUC of T*I at 24 h predicting for AKI was 0.71(95% CI 0.59-0.81, P=0.001, cutoff value 0.020, sensitivity 79.2%, specificity 56.0%), while the AUC for stage 2-3 AKI was 0.85 (95% CI 0.75-0.92, P<0.001, cutoff value 0.083, sensitivity 70.0%, specificity 90.6%). Urinary T*I normalized for urinary creatinine excretion did not show better predictive value. In addition, of T*I at 24 h predicting for poor hospitalization outcome, renal recovery, and one year postoperative death, the AUC was 0.82(95% CI 0.71-0.90, P=0.001), 0.80(95% CI 0.66-0.86, P<0.001), and 0.81(95% CI 0.70-0.89, P=0.047), respectively. Conclusion:The combined detection of TIMP-2 and IGFBP7 in urine is expected to be a biomarker for early diagnosis of CSA-AKI and has certain clinical value in predicting the prognosis of CSA-AKI.

Objective To study the differentially expressed microRNA (miRNA) between pancreatic ductal adenocarcinoma (PDAC) and para-cancerous tissues,and determine related target genes.Methods Nine fresh PDAC tumor tissues and 3 adjacent normal pancreatic tissues were collected,then Agilent miRNAmicroarray with 713 miRNA loci was used to identify the differentially expressed miRNA.Real-time PCR method was applied to verify the up-regulated miRNA.TargetScan 5.1 and miRandaV5 software were used to analyze the related target genes.Results miRNA microarray identified 11 PDAC related miRNAs,among them,the expressions of miR-194*,miR-192*,miR-602,miR-194 were up-regulated,while the expressions of miR-139-3p,miR-513a-5p,miR-630,miR-30c-1 *,miR-887,miR-508-5p,miR-516a-5p were down-regulated.The expressions of miR-192,miR-194 and their homolog were verified in 31 PDAC tumor tissues.After software analysis,it was found that target genes of miR-192 were ZEB2,CXCL-2,EEF1A1,ERCC3,and target genes of miR-192 * were DCC,SMAD4,FAS,and target genes of miR-194 included DACHI,IGSF11,PTPN2,RBBP4,while target genes of miR-194 * included CD40LG,CIDEB,FHL1.Conclusions Eleven differentially expressed miRNAs are present in PDC,and they may be involved in the occurrence and development of PDC.

Objective:Recent studies have shown that in contrast to decrease in distal gastric adenocarcinoma (DGA), incidence of adenocarcinoma of the esophagogastric junction (AEG) has increased noticeably in numerous counties. However, the reasons remain unclear. This study evaluated the possible differences in the expression of KLF4, SP1, and Cyclin D1 in AEG and DGA, and explored the potential carcinogenesis of AEG. Methods:Immunohistochemistry was performed on paraffin-embedded tissues to evaluate the pu-tative differences in the expressions of KLF4, SP1, and Cyclin D1 at protein level between AEG (n=58) and DGA (n=47). The patholog-ical significance of these markers between the two groups was also compared and analyzed. Results:The percentage of positive KLF4 expression was significantly lower in DGA than in AEG (P<0.05). Lower KLF4 expression was found both in well-or moderately dif-ferentiated cases and in poorly differentiated cases with DGA compared with their AEG counterparts (P<0.05). However, positive stain-ing for SP1 was significantly higher in DGA (P<0.05). No significant difference was found in the expression of Cyclin D1 between the two groups. Further analysis showed that in DGA, the positive expression of KLF4, SP1, and Cyclin D1 were significantly correlated with lymph node metastasis. In AEG, only Cyclin D1 expression was correlated with lymph node metastasis (P<0.05). No correlation was found among the expression of KLF4, SP1, and Cyclin D1 in AEG. In DGA, KLF4 was inversely correlated with SP1 and Cyclin D1 (r=-0.334 and r=-0.341, respectively, P<0.05), and SP1 was positively correlated with Cyclin D1 expression (r=0.340, P<0.05).Conclusion:Different expression patterns and clinicopathological significance of KLF4, SP1, and Cyclin D1 were observed between AEG and DGA, suggesting the putative difference in the carcinogenesis and progression of AEG and DGA.

Objective To investigate the expression of HSPB1 in pancreatic cancer and its relationship with clinicopathological characterization.Methods Pathological specimens from 47 cases of pancreatic cancers,13 cases of para-cancerous normal pancreatic tissues and 3 cases of chronic pancreatitis tissues were selected,and tissue microarray construction instrument was used to prepare tissue microarray,then the expression of HSPB1 was determined by immunohistochemistry SP method.The scores of the proportion of positive cells and staining intensity were multiplied to make the judgment.Results The expression of HSPB1 in malignant,para-cancerous and chronic pancreatitis tissues was 80.9％ (38/47),12.5％ (2/16),respectively; and the difference between the two groups was statistically significant (X2 =24.058,P =0.000).The expression of HSPB1 in pancreatic cancer tissue was not significantly related to sex,age,location,differentiation degree and nerve infiltration of tumor ( P ＞ 0.05 ).Conclusions The expression of HSPB1 is related to development and progression of pancreatic cancer,and may be an early molecular event.

Objective To analyze the practicability of using ELISA kit for the detection of hepati-tis E virus antigen ( HEV-Ag) in plasma donations and Biomex HEV seroconversion panels. Methods The HEV-Ag positive samples were screened out from 36 340 donated blood plasma samples. Real-time fluores-cent PCR was performed for the detection of HEV RNA in HEV-Ag positive samples. The open reading frame 2 (ORF2) in HEV RNA was amplified by nested RT-PCR and the amplified products were confirmed by sequencing analysis. Phylogenetic tree was constructed for HEV genotyping. Five Biomex HEV serocon-version panels were used in this study for the detection of HEV-Ag, anti-HEV antibody and HEV RNA as well as the correlation analysis between HEV-Ag and HEV RNA. Results Twenty-six out of 36 340 plasma samples (0. 07%) were positive for HEV-Ag. Of the 26 samples, 25 samples were positive for HEV RNA as indicated by the results of nested RT-PCR and 23 positive samples were confirmed by sequencing analysis. The positive rate of HEV RNA in blood plasma donators was 1 ∶ 1 580 (0. 06%). There were 17 samples of genotype 1 (74%) and 6 samples of genotype 4 (26%) according to the phylogenetic tree analysis. All of the HEV-Ag positive samples were also positive for HEV RNA as indicated by the analysis of Biomex sero-conversion panels. HEV-Ag was consistent with the peak of the HEV RNA concentration. Conclusion A close relationship between HEV-Ag and HEV RNA was observed. HEV-Ag screening could be used as a measure to reduce the risk of HEV transmission by blood transfusion.

Objective To study the in vivo expression and biodistribution of Ad5-Fluc (Adenovirus carrying firefly luciferase genes) in mice.Methods The recombinant Ad5-Fluc virus was constructed and infected to BALB/c or nude mice through three different routes.The protein expression level,tissue distribution and the characteristics of infection were analyzed by in vivo bioluminescence imaging technology.Results Compared to other two routes,the BALB/c mice infected through muscular route had the longest expression cycle (over 60 days) and the highest expression level,while the virus was transferred into the liver and spleen after infection.The nude mice had a significantly extended expression cycle than BALB/c mice.Moreover,the characteristic of liver tropism was eliminated after Ad5 F35 infection in mice,while maintained similar expression efficiency.Conclusion Due to the highest expression efficiency,the muscular route would be the optimal route for Ad5 vector based vaccination.In addition,Ad5F35 virus could become an ideal alternative vaccine vector for eliminating the liver tropism.

BACKGROUND:Cryotherapy plays a positive role in the treatment of delayed-onset muscle soreness caused by high intense exercise. OBJECTIVE: To investigate the effects of different crypotherapy programs on the levels of interleukin-6 and prostaglandin 2 in long distance race-walkers after 15-day training, and to determine a rational treatment program for delayed-onset muscle soreness. METHODS:Sixteen male race-walkers in Liaoning Province were randomly divided into cryotherapy and cryo/heat therapy groups, and received 10-minute cryotherapy and 2.5-minute cryo/heat therapy (2.5-mintue cryotherapy and 2.5-minute heat therapy alternately for 10 minutes), respectively, after 15-day training. The serum levels of interleukin-6 and prostaglandin 2 were detected at six different time points to compare the efficacy between two methods. RESULTS AND CONCLUSION: Compared with the cryo/heat therapy group, the serum levels of interleukin-6 and prostaglandin 2 in the cryotherapy group were significantly decreased. That is to say, cryotherapy is more available for alleviating delayed-onset muscle soreness after intensive eccentric training or in intensive seasons.

Objective To study the prevalence and genotype of human parvovirus B19 virus among blood products and plasma pools in China. Methods B19 DNA derived from 16 lots of factor Ⅷ concentrate produced by 4 manufactures and 10 lots of plasma pools were detected by nested PCR. Phylogenetic comparison of the partial B19 sequences obtained from positive samples were performed by direct sequencing. Results Twelve of sixteen lots of factor Ⅷ concentrate and all of ten lots of plasma pools were contaminated by B19 DNA. By comparing the partial B19 sequences,all the isolated viruses were genotype Ⅰ and their nucleotides were high conserved with homology of 98. 3%-100%. Conclusion B19 genotype Ⅰ DNA has been detected in high prevalence in factor Ⅷ concentrate and plasma pools. The genetic diversities were shown to be very low.

Objective To investigate the clinical significance of detection of hepatitis E virus (HEV) RNA in sera from patients with acute hepatitis E using real-time reverse transcription (RT)-PCR to detect hepatitis E virus RNA in sera from patients with acute hepatitis E.Methods A real-time RT-PCR assay, which can amplifies and detect the conserved region on ORF3, was used in this study. 434 outpatients and hospitalized patients with acute HEV infection was enrolled into this study.Simultaneously,the serum samples from 40 patients with HAV infection, 100 patients with HBV infection and 110 healthy blood donors were collected as the control The real-time RT-PCR was performed to detect HEV RNA in all these sera.Results 232 sera (53.5%) were positive for HEV RNA by real-time RT-PCR and all of the control were negative.The results of real-time RT-PCR and anti-HEV IgM (ELISA) were concordant in 67.1% samples.There was significant difference between the two methods ( Kappa = 0.308, P = 0.000 ).The first serum sample from five serum samples of the patients was positive for HEV RNA and negative for anti-HEV IgM.Follow-up studies showed all the five sera samples were positive for anti-HEV IgM.HEV RNA in serum could be detected between 2 and 10 days.Conclusions The real-time fluorescent RT-PCR method has high specificity, and can be applied to the qualitative detection of the serum with genotypes Ⅰ and Ⅳ of hepatitis E virus.Its clinical use can improve the early diagnosis of HEV.