Objective To knock out Asxl2 gene in murine embryonic fibroblast cell line NIH3T3 using CRISPR/Cas9n system. Methods A pair of sgRNAs which targeted exon 5 of Asxl2 gene were designed and subcloned into the pX462 vec?tor. The recombined plasmids were verified by sequencing and transfected into NIH3T3 cell line. Single cells were isolated through serial dilutions, followed by an expansion period to obtain new monoclonal cell lines. The genomic DNA of the new monoclonal cell lines was extracted and a DNA fragment flanked the target site was amplified by genotyping PCR then se?quenced. Lastly, western blotting were applied to confirm whether Asxl2 was successfully knocked out. Results The CRIS?PR/Cas9n plasmids that targeted Asxl2 were successfully constructed. NIH3T3 cells were co-transfected with the two recom?binant constructs. After puromycin selection, subclonal cell lines were obtained and one of them was validated by genotyping PCR-sequencing. Western blotting also confirmed that Asxl2 was completely depleted in the NIH3T3 cell line. Conclu?sion CRISPR/Cas9n plasmids that targeted Asxl2 were successfully constructed therefore a Asxl2 knockout NIH3T3 stable cell line was established via this system.

Objective: To analyze the semen quality parameters of sperm donor volunteers, so as to provide insights into male infertility control and related research. Methods: A total of 155 sperm donors were recruited from the Human Sperm Bank of Zhejiang Province using the convenience sampling method from January to March 2021. Demographic information were collected through questionnaire surveys. Semen were collected, and parameters including semen volume, sperm concentration, total number of sperm, forward motility rate and total sperm viability were measured. Semen quality was evaluated according to the WHO Laboratory Manual for the Examination and Processing of Human Semen. : Results Conclusion There were 20.65% of the 155 sperm donors with unqualified semen, and the unqualified rates of forward motility rate and total sperm viability were relatively high.

Objective To investigate the predictive value of Rho-associated coiled-coil contai-ning kinase1(ROCK1)and ROCK2 for no reflow in patients with acute ST segment elevation myocar-dial infarction(STEMI)undergoing percutaneous coronary intervention(PCI).Methods A total of 168 STEMI patients who received PCI were selected as study objects,and were divided into no reflow group and normal blood flow group based on whether no reflow occurred,were divided into high ex-pression group and low expression group based on the expression of ROCK1 and ROCK2.Enzyme linked immunosorbent assay was used to detect the levels of serum ROCK1 and ROCK2.The differ-ences in serum levels of ROCK1 and ROCK2 between the no reflow group and the normal blood flow group were analyzed,and the risk factors for no reflow in STEMI patients undergoing PCI treatment were analyzed,the predictive value of ROCK1 and ROCK2 for no reflow in STEMI patients undergoing PCI treatment were analyzed.Results Of 168 STEMI patients,no reflow occurred in 46 cases(27.38％).The Killip grade,time from onset to hospital admission,proportion of patients who did not use proph-ylactic no reflow,serum ROCK1 level and serum ROCK2 level in the no reflow group were higher or longer than those in the normal blood flow group(P＜0.05).The incidence of no reflow in ROCK1 high expression group was higher than that in ROCK1 low expression group,and the incidence of no reflow in ROCK2 high expression group was higher than that in ROCK2 low expression group,the differences were statistically significant(P＜0.05).The incidence rates of no reflow in the ROCK1 high expression group and ROCK2 high expression group were higher than that in the ROCK1 low expression group and ROCK2 low expression group(P＜0.05).Multiple Logistic regression analy-sis showed that Killip grade of Ⅲ to Ⅳ,longer onset to admission time,no using prophylactic no re-flow drugs,and higher serum levels of ROCK1 and ROCK2 were all risk factors for no reflow in STEMI patients undergoing PCI(P＜0.05).Receiver operating characteristic curve showed that ROCK1 and ROCK2 had high predictive value for PCI in STEMI patients without reflow,and the predictive value was increased after the combination of ROCK1 and ROCK2.The area under the curve was 0.789(95％CI,0.711 to 0.867).Conclusion High serum levels of ROCK1 and ROCK2 are both risk factors for no reflow in STEMI patients undergoing PCI,and their combination has high predictive value for no reflow.

Objective To investigate the predictive value of Rho-associated coiled-coil contai-ning kinase1(ROCK1)and ROCK2 for no reflow in patients with acute ST segment elevation myocar-dial infarction(STEMI)undergoing percutaneous coronary intervention(PCI).Methods A total of 168 STEMI patients who received PCI were selected as study objects,and were divided into no reflow group and normal blood flow group based on whether no reflow occurred,were divided into high ex-pression group and low expression group based on the expression of ROCK1 and ROCK2.Enzyme linked immunosorbent assay was used to detect the levels of serum ROCK1 and ROCK2.The differ-ences in serum levels of ROCK1 and ROCK2 between the no reflow group and the normal blood flow group were analyzed,and the risk factors for no reflow in STEMI patients undergoing PCI treatment were analyzed,the predictive value of ROCK1 and ROCK2 for no reflow in STEMI patients undergoing PCI treatment were analyzed.Results Of 168 STEMI patients,no reflow occurred in 46 cases(27.38％).The Killip grade,time from onset to hospital admission,proportion of patients who did not use proph-ylactic no reflow,serum ROCK1 level and serum ROCK2 level in the no reflow group were higher or longer than those in the normal blood flow group(P＜0.05).The incidence of no reflow in ROCK1 high expression group was higher than that in ROCK1 low expression group,and the incidence of no reflow in ROCK2 high expression group was higher than that in ROCK2 low expression group,the differences were statistically significant(P＜0.05).The incidence rates of no reflow in the ROCK1 high expression group and ROCK2 high expression group were higher than that in the ROCK1 low expression group and ROCK2 low expression group(P＜0.05).Multiple Logistic regression analy-sis showed that Killip grade of Ⅲ to Ⅳ,longer onset to admission time,no using prophylactic no re-flow drugs,and higher serum levels of ROCK1 and ROCK2 were all risk factors for no reflow in STEMI patients undergoing PCI(P＜0.05).Receiver operating characteristic curve showed that ROCK1 and ROCK2 had high predictive value for PCI in STEMI patients without reflow,and the predictive value was increased after the combination of ROCK1 and ROCK2.The area under the curve was 0.789(95％CI,0.711 to 0.867).Conclusion High serum levels of ROCK1 and ROCK2 are both risk factors for no reflow in STEMI patients undergoing PCI,and their combination has high predictive value for no reflow.

{L-End}Objective To analyze the effects of night shift work and overweight/obesity on blood pressure of workers in chemical fiber industry. {L-End}Methods A total of 1 004 workers of a chemical fiber factory were selected as the study subjects using convenient sampling method, and their blood pressure and body mass index were measured. Multiple linear regression model was used to analyze the relationship between night shift work and blood pressure, and multiple logistic regression was used to assess the independent impact and combined impact of night shifts and overweight/obesity on the risk of hypertension. {L-End}Results Compared with the non-night shift workers, the prevalence of hypertension in night shift workers was increased (5.3% vs 13.0%, P<0.05), with elevated systolic and diastolic blood pressure (both P<0.05). The results of multiple linear regression analysis showed that the systolic blood pressure and diastolic blood pressure of the night shift workers were higher than those of the non-night shift workers (both P<0.05), and the systolic blood pressure and diastolic blood pressure of overweight/obesity workers were higher than those of non-overweight/obesity workers (both P<0.01). The results of multiple logistic regression analysis showed that the risk of hypertension in night shift workers and overweight/obesity workers was higher than that in non-night shift workers and non-overweight/obesity workers [odds ratio (OR) and 95% confidence interval (CI) were 2.49 (1.04-5.99) and 2.65 (1.77-3.95), both P<0.05]. Night shift work and overweight/obesity showed a synergistic effect on blood pressure of workers. Compared to non-overweight/obesity non-night shift workers, overweight/obesity night shift workers had a higher risk of hypertension (OR=4.93, 95%CI: 1.70-14.29, P<0.01). {L-End}Conclusion Night shift work could lead to elevated blood pressure in workers in the chemical fiber industry, which is a potential risk factor for hypertension. The synergistic effect of night shift work and overweight/obesity may contribute to the increased risk of hypertension.

OBJECTIVETo analyze the geographic spatial patterns and risk areas of main digestive system cancers in Yiwu city.

METHODSNewly diagnosed cases of esophageal, gastric and colorectal cancer during 2010-2014 were obtained from Yiwu Center for Disease Control and Prevention (CDC). The household registration population data in 2013 were obtained from public security bureau. Hierarchy clustering and partitioning regionalization method was used to generate geographic units. Global Moran's I was used to evaluate whether cancer incidence was significantly clustered in space, Anselin Local Moran's I was used to identify statistically significant hot spots, cold spots, and spatial outliers, and Spatial Scan Statistics was implemented to analyze the relative risk of cancers in different areas.

RESULTSThe 5-year average incidence of esophageal, gastric and colorectal cancers were 9.99/100 000, 34.01/100 000 and 31.46/100 000, respectively. Males showed significantly higher incidence than females. The incidence was heterogeneous throughout the study area. Spatial Scan analysis revealed that southern Yiwu presented a significantly higher male esophageal cancer (=1.78) and gastric cancer (=1.87) risk. The central area of Yiwu showed a significantly lower female esophageal cancer risk (=0.00) and male stomach cancer risk (=0.63) and the northern Yiwu exhibited a significantly lower female colorectal cancer risk (=0.48).

CONCLUSIONSThe incidence of main digestive tract cancers shows a heterogeneous distribution in Yiwu city.