African swine fever(ASF)is an acute,febrile,and highly fatal disease caused by African swine fever virus(ASFV)in pigs.Given the current lack of commercial vaccines and the continu-ous evolution of ASFV in recent years,the emergence of moderately virulent genotype Ⅱ strains and the introduction of genotype Ⅰ attenuated strains have led to persistent and chronic infections in pigs.Therefore,the detection of specific antibodies against ASFV has become imperative.In this study,we established a competitive chemiluminescence immunoassay(p30-cCLIA)for detecting ASFV p30 antibodies using p30 monoclonal antibodies.By detecting sera with clear negative and positive backgrounds,we determined that the Cut-off value of this method was 50％,with both di-agnostic sensitivity(Dsn)and diagnostic specificity(Dsp)reaching 100％.Under optimal reaction conditions,we screened out an enzyme-labeled stabilizer suitable for p30 monoclonal antibody 16-5E7E8-HRP.Furthermore,the sensitivity of the established p30-cCLIA method was higher than that of the commercial blocking ELISA kit(1∶2 048 vs 1∶512)and exhibited good repeatability.Detection of sera positive for other porcine virus infections showed no cross-reactivity.The estab-lishment of this method provides a powerful tool for early diagnosis of ASF.

African swine fever(ASF)is an acute,febrile,and highly fatal disease caused by African swine fever virus(ASFV)in pigs.Given the current lack of commercial vaccines and the continu-ous evolution of ASFV in recent years,the emergence of moderately virulent genotype Ⅱ strains and the introduction of genotype Ⅰ attenuated strains have led to persistent and chronic infections in pigs.Therefore,the detection of specific antibodies against ASFV has become imperative.In this study,we established a competitive chemiluminescence immunoassay(p30-cCLIA)for detecting ASFV p30 antibodies using p30 monoclonal antibodies.By detecting sera with clear negative and positive backgrounds,we determined that the Cut-off value of this method was 50％,with both di-agnostic sensitivity(Dsn)and diagnostic specificity(Dsp)reaching 100％.Under optimal reaction conditions,we screened out an enzyme-labeled stabilizer suitable for p30 monoclonal antibody 16-5E7E8-HRP.Furthermore,the sensitivity of the established p30-cCLIA method was higher than that of the commercial blocking ELISA kit(1∶2 048 vs 1∶512)and exhibited good repeatability.Detection of sera positive for other porcine virus infections showed no cross-reactivity.The estab-lishment of this method provides a powerful tool for early diagnosis of ASF.

Objective : To investigate the effect of long non⁃coding RNA (LncRNA) anoctamin 1 antisense RNA⁃1 (ANO1⁃AS1) on the proliferation and apoptosis of esophageal squamous cell carcinoma (ESCC) cells and its possible mechanisms. Methods : Silenced ANO1⁃AS1 lentivirus was transfected in ESCC cells TE⁃1 and EC109. Subsequently, the expression levels of ANO1⁃AS1 and calcium⁃activated chloride channel protein 1 (ANO1) in the cells were detected by qRT⁃PCR. CCK⁃8 and colony formation assays were used to detect the proliferation of TE⁃1 and EC109 cells. ANO1 positively related expressed genes were obtained from the LinkedOmics database and then the gene set was enriched for pathways and possible pathways were validated. The expression levels of proliferating cell nuclear antigen(PCNA), P53 protein, apoptosis⁃related protein ( Bax and Bcl⁃2), ANO1 protein and phosphatidylinositol⁃3⁃kinase/protein kinase B (PI3K/Akt) pathway⁃related protein were assessed by Western blot. Results: After transfection of lentivirus with silent expression function, the expression level of ANO1⁃AS1 was significantly reduced in TE⁃1 and EC109 cells (P < 0. 05); After down⁃regulation of ANO1⁃AS1, compared with the negative control group, the proliferation ability of ESCC cells was reduced (P < 0. 05) and the rate of clone formation decreased (P < 0. 05); Western blot results showed that, compared with negative controls, the expression of PCNA decreased, the expression of oncogene P53 protein increased ( P < 0. 05 ), the expression of proteins ( Bax) increased, Bcl⁃2 decreased and the levels of phosphorylation of the pathway proteins PI3K and Akt decreased (P < 0. 05) . Conclusion Knockdown of ANO1⁃AS1 can decrease proliferation and promote apoptosis in ESCC, which may be achieved by affecting PI3K/Akt pathway activation.

OBJECTIVETo investigate the effect of intestinal resection on hydrogen sulfide (H2S) biosynthesis and interstitial cells of Cajal(ICC) in mice.

METHODSAfter intestinal resection mouse model was established, the activity of MPO in the proximal anastomosis intestinal tissue were detected. Sensitive sulphur electrode assay was applied to measure the H2S level. RT-PCR technique was employed to investigate the mRNA expression of the endogenous H2S biosynthesis enzymes, cystathionine-b-synthase (CBS) and cystathionine-c-lyase (CSE). Immunofluorescence staining was used to detect the expression of c-kit in order to calculate the area of ICC.

RESULTSThe mRNA expression of CSE was detected in the small intestine tissue of mice, while no CBS mRNA was found. The mRNA expression of CSE in proximal anastomotic stoma increased in time-dependent manner in the model group. CSE mRNA expression began to increase 1 hour after operation, reached the peak at 6th hour, then decreased gradually, and was similar to the control group at postoperative 24th hour. Compared to the model group, in the intestinal tissues of proximal 3 cm to anastomotic stoma, the mRNA expression of CSE (1.16 ± 0.18 vs. 1.63 ± 0.13, P<0.05), the activity of MPO [(0.54 ± 0.07) U/g vs. (0.83 ± 0.09) U/g, P<0.05], the H2S level [(36.1 ± 6.1) nmol/mg vs. (5.3 ± 5.6) nmol/mg, P<0.05] were significantly reduced in the PPG group. Meanwhile, average percentage of positive ICC area in the PPG groups was significantly higher [(2.26 ± 0.19)% vs. (1.65 ± 0.24)%, P<0.05].

CONCLUSIONSInflammatory reaction in muscular layer induced by intestinal resection up-regulates the mRNA expression of CSE proximal to anastomotic stoma, generates excess H2S to damage ICC leading to intestinal motor dysfunction. Preoperative inhibition of endogenous H2S generation may protect the ICC.

Animals ; Cystathionine gamma-Lyase ; Disease Models, Animal ; Hydrogen Sulfide ; Inflammation ; Interstitial Cells of Cajal ; Intestines ; Mice ; Proto-Oncogene Proteins c-kit ; RNA, Messenger

Objective To investigate the effect of intestinal resection on hydrogen sulfide (H2S) biosynthesis and interstitial cells of Cajal (ICC) in mice. Methods After intestinal resection mouse model was established , the activity of MPO in the proximal anastomosis intestinal tissue were detected. Sensitive sulphur electrode assay was applied to measure the H2S level. RT-PCR technique was employed to investigate the mRNA expression of the endogenous H2S biosynthesis enzymes , cystathionine-b-synthase (CBS) and cystathionine-c-lyase (CSE). Immunofluorescence staining was used to detecte the expression of c-kit in order to calculate the area of ICC. Results The mRNA expression of CSE was detected in the small intestine tissue of mice , while no CBS mRNA was found. The mRNA expression of CSE in proximal anastomotic stoma increased in time-dependent manner in the model group. CSE mRNA expression began to increase 1 hour after operation , reached the peak at 6th hour, then decreased gradually, and was similar to the control group at postoperative 24th hour. Compared to the model group, in the intestinal tissues of proximal 3 cm to anastomotic stoma, the mRNA expression of CSE (1.16 ± 0.18 vs. 1.63±0.13, P<0.05), the activity of MPO [(0.54 ± 0.07) U/g vs. (0.83 ± 0.09) U/g, P<0.05], the H2S level [(36.1±6.1) nmol/mg vs. (5.3±5.6) nmol/mg, P<0.05] were significantly reduced in the PPG group. Meanwhile, average percentage of positive ICC area in the PPG groups was significantly higher[(2.26±0.19)%vs. (1.65±0.24)%, P<0.05]. Conclusions Inflammatory reaction in muscular layer induced by intestinal resection up-regulates the mRNA expression of CSE proximal to anastomotic stoma, generates excess H2S to damage ICC leading to intestinal motor dysfunction. Preoperative inhibition of endogenous H2S generation may protect the ICC.

Objective To investigate the effect of intestinal resection on hydrogen sulfide (H2S) biosynthesis and interstitial cells of Cajal (ICC) in mice. Methods After intestinal resection mouse model was established , the activity of MPO in the proximal anastomosis intestinal tissue were detected. Sensitive sulphur electrode assay was applied to measure the H2S level. RT-PCR technique was employed to investigate the mRNA expression of the endogenous H2S biosynthesis enzymes , cystathionine-b-synthase (CBS) and cystathionine-c-lyase (CSE). Immunofluorescence staining was used to detecte the expression of c-kit in order to calculate the area of ICC. Results The mRNA expression of CSE was detected in the small intestine tissue of mice , while no CBS mRNA was found. The mRNA expression of CSE in proximal anastomotic stoma increased in time-dependent manner in the model group. CSE mRNA expression began to increase 1 hour after operation , reached the peak at 6th hour, then decreased gradually, and was similar to the control group at postoperative 24th hour. Compared to the model group, in the intestinal tissues of proximal 3 cm to anastomotic stoma, the mRNA expression of CSE (1.16 ± 0.18 vs. 1.63±0.13, P<0.05), the activity of MPO [(0.54 ± 0.07) U/g vs. (0.83 ± 0.09) U/g, P<0.05], the H2S level [(36.1±6.1) nmol/mg vs. (5.3±5.6) nmol/mg, P<0.05] were significantly reduced in the PPG group. Meanwhile, average percentage of positive ICC area in the PPG groups was significantly higher[(2.26±0.19)%vs. (1.65±0.24)%, P<0.05]. Conclusions Inflammatory reaction in muscular layer induced by intestinal resection up-regulates the mRNA expression of CSE proximal to anastomotic stoma, generates excess H2S to damage ICC leading to intestinal motor dysfunction. Preoperative inhibition of endogenous H2S generation may protect the ICC.