Objective:To establish a digital reverse transcription polymerase chain reaction（dRT-PCR）detection method for hepatitis E virus（HEV）using nanoplates，and to provide technical reference for HEV monitoring in shellfish by combining virus enrichment pretreatment methods.Methods:The annealing temperature，primer and probe concentrations of HEV dRT-PCR were optimized，and the specificity of the method was evaluated；the sensitivity of this method for detecting HEV in water samples and oyster extracts was compared. The inhibition rate and recovery rate of HEV detection in artificially contaminated oyster samples were calculated，commercially available oyster samples were tested，and compare them with real-time fluorescence quantitative RT-PCR（qRT-PCR）method.Results:The optimized annealing temperature for HEV dRT-PCR was determined to be 60 ℃，and the final concentrations of primers and probes were 0.4 μmol/L，0.4 μmol/L，and 0.2 μmol/L，respectively，indicating good specificity. The sensitivity of both methods for detecting HEV RNA in water samples was higher than that in oyster extracts. The recovery rates of HEV in oyster specimens contaminated with HEV fecal suspension by dRT-PCR and qRT-PCR were 18.76% and 18.36%，respectively，with no statistically significant difference（ P>0.05）；the inhibition rates were 17.26% and 9.58%，respectively，with statistically significant differences（ P<0.05）；55 commercially available oyster samples were tested，and both methods detected HEV RNA positivity in the same sample. Conclusion:The dRT-PCR method established in this study，combined with “proteinase K digestion，PEG/NaCl precipitation，and chloroform/n-butanol extraction” pretreatment，has a good recovery effect on HEV in shellfish food containing a large amount of PCR inhibitors，and can achieve absolute quantification. It has certain application value in monitoring and risk assessment of HEV in shellfish food.

Objective:To establish a digital reverse transcription polymerase chain reaction（dRT-PCR）detection method for hepatitis E virus（HEV）using nanoplates，and to provide technical reference for HEV monitoring in shellfish by combining virus enrichment pretreatment methods.Methods:The annealing temperature，primer and probe concentrations of HEV dRT-PCR were optimized，and the specificity of the method was evaluated；the sensitivity of this method for detecting HEV in water samples and oyster extracts was compared. The inhibition rate and recovery rate of HEV detection in artificially contaminated oyster samples were calculated，commercially available oyster samples were tested，and compare them with real-time fluorescence quantitative RT-PCR（qRT-PCR）method.Results:The optimized annealing temperature for HEV dRT-PCR was determined to be 60 ℃，and the final concentrations of primers and probes were 0.4 μmol/L，0.4 μmol/L，and 0.2 μmol/L，respectively，indicating good specificity. The sensitivity of both methods for detecting HEV RNA in water samples was higher than that in oyster extracts. The recovery rates of HEV in oyster specimens contaminated with HEV fecal suspension by dRT-PCR and qRT-PCR were 18.76% and 18.36%，respectively，with no statistically significant difference（ P>0.05）；the inhibition rates were 17.26% and 9.58%，respectively，with statistically significant differences（ P<0.05）；55 commercially available oyster samples were tested，and both methods detected HEV RNA positivity in the same sample. Conclusion:The dRT-PCR method established in this study，combined with “proteinase K digestion，PEG/NaCl precipitation，and chloroform/n-butanol extraction” pretreatment，has a good recovery effect on HEV in shellfish food containing a large amount of PCR inhibitors，and can achieve absolute quantification. It has certain application value in monitoring and risk assessment of HEV in shellfish food.

Objective To investigate the risk factors associated with the presence of multiple Lugol-voiding lesions(LVLs)in patients with early esophageal cancer and the correlation with alcohol dehy-drogenase 1B(ADH1B)and aldehyde dehydrogenase 2(ALDH2)polymorphisms.Methods Patients who underwent endoscopic submucosal dissection due to early esophageal cancer were divided into group with multiple LVLs and group without multiple LVLs based on their endoscopic features.Their clinical data and the genotype of ADH1B and ALDH2 were collect-ed and SPSS 27.0 was used to statistically analyze the above data.Results A total of 83 subjects were included in the study,23 had multiple LVLs,most of them were seen in males,alcohol drinkers,and smokers with smoking index≥1 000,and multivariate analysis showed that alcohol consumption was an independent risk factor for it(OR=6.215,P=0.008).The gene polymorphism of ADH1B and ALDH2 and their interactions did not have any sig-nificant correlation with multiple LVLs.However,among alcohol drinkers,there was a 12-fold increased risk of multiple LVLs in patients carrying the ALDH2 A allele and drinking≥50 g per day compared to those carrying the ALDH2 GG genotype and drinking<50 g per day(P=0.045).Conclusion Alcohol consumption is an inde-pendent risk factor of multiple LVLs of the esophageal mucosa in patients with early esophageal cancer,and heavy alcohol consumption in carriers of the ALDH2 A allele will significantly increase the risk of multiple LVLs,and such patients should be closely followed up with endoscopy.

Objective:To compare the clinical efficacy of anchor repair versus screw fixation in the treatment of posterior malleolar fracture with distal tibiofibular syndesmosis injury.Methods:PubMed, Medline, Web of Science, ScienceDirect, CNKI, Wanfang, and Chinese Medical Journal Full-text Database were searched for articles on anchor repair versus screw fixation in the treatment of posterior malleolar fracture with distal tibiofibular syndesmosis injury. The search time was from the establishment of each database to April 2023. Literature screening, data extraction and literature quality assessment were performed independently by two researchers according to the inclusion and exclusion criteria, and meta-analysis of the included literature was performed.Results:A total of 7 articles were included in the meta-analysis, including 3 randomized controlled trials and 4 case-control studies. There were 280 cases treated with anchor repair and 312 cases treated with screw fixation. The results of meta-analysis showed that the number of fluoroscopy [ MD=-5.08, 95% CI (-9.20, -0.96), P=0.020], postoperative anterior inferior tibiofibular space [ MD=-0.93, 95% CI (-1.06, -0.81), P<0.001] and incidence of malposition [ OR=0.21, 95% CI (0.10, 0.46), P<0.001] in the anchor repair group were smaller than that in the screw fixation group, while postoperative recovery time were earlier than that in the screw fixation group [ MD=-2.22, 95% CI (-2.68, -1.75), P<0.001], postoperative ankle plantarflexion angle [ MD=2.77, 95% CI (0.28, 5.25), P=0.030], and postoperative 6 months of American Orthopedic Foot and Ankle Society score [ MD=5.85, 95% CI (2.05, 9.64), P=0.003] were greater than those of the screw fixation group. The operation time [ MD=-10.45, 95% CI (-24.25, 3.35), P=0.140], the American Orthopaedic Foot and Ankle Society score at 6 months after operation [ MD=0.09, 95% CI (-0.94, 1.11), P=0.860] and the postoperative ankle dorsiflexion angle [ MD=0.66, 95% CI (-0.75, 2.88), P=0.360] were not statistically different. Conclusion:Compared with screw fixation, fixation of anterior inferior tibiofibular ligament with anchor fixation has the advantages of less fluoroscopy, faster recovery time, better reduction quality, and higher ankle function score.

Objective:To investigate the effect of endometrial thickness (EMT) on the clinical outcome of hormone replacement frozen-thawed embryo transfer (HRT-FET) cycle in different body mass index (BMI) groups, and to analyze the threshold and optimal EMT and EMT interval corresponding to the ideal clinical pregnancy rate.Methods:A retrospective cohort study was conducted on 10 239 HRT-FET cycles in the Reproductive Medicine Center of Henan Provincial People's Hospital from January 2013 to December 2017, and they were divided into low weight group (BMI<18.5 kg/m 2), normal weight group (BMI=18.5-24.9 kg/m 2), overweight group (BMI=25.0-29.9 kg/m 2) and obese group (BMI≥30.0 kg/m 2). Four subgroups were divided according to EMT, respectively EMT<8.0 mm, 8.0 mm≤EMT<10.0 mm, 10.0 mm≤EMT<12.0 mm, EMT≥12.0 mm. The clinical characteristics and outcome indicators of different EMT subgroups in different BMI groups were compared. To achieve the ideal clinical pregnancy rate, multiple regression analysis, curve fitting and threshold effect analysis were used to find the best EMT and thickness interval. Results:1) After adjusting for confounding factors, multiple regression analysis showed that, there were no significant differences in clinical pregnancy rate and live birth rate among subgroups with the increase of EMT (all groups P>0.05). The clinical pregnancy rate and the live birth rate increased with the increase of EMT between subgroups of normal body weight group and super-recombinant subgroups (all P<0.001 for normal body weight subgroups, P=0.123, P=0.009, P=0.016 and all P<0.001 for super-recombinant subgroups). In the obesity group, with the increase of EMT, the clinical pregnancy rate did not increase significantly except EMT≥12.0 mm subgroup ( P=0.449, P=0.279, P=0.021), while the live birth rate increased significantly ( P=0.014, P=0.005, P<0.001). 2) Curve fitting showed that in the population of low weight and obese, influence of EMT on clinical pregnancy rate was a straight line, in the population of normal weight and overweight, influence of EMT on clinical pregnancy rate was a curve, as EMT increased the clinical pregnancy rate raised and then decreased, the impact on the live birth rate appeared similar. 3) According to the curve fitting, the threshold effect analysis of the normal weight group showed that the endometrial inflection point of EMT on the clinical pregnancy rate and the live birth rate was 10.0 mm. When EMT was lower than 10.0 mm, the clinical pregnancy rate and the live birth rate increased by 20% and 19% for every 1.0 mm increase in endometrial thickness ( OR=1.20, 95% CI=1.13-1.26; OR=1.13,95% CI=1.13-1.26). In overweight group, the inflection point of EMT on the clinical pregnancy rate and the live birth rate was also 10.0 mm. When EMT was lower than 10.0 mm, the clinical pregnancy rate and the live birth rate increased by 24% and 26% for every 1.0 mm increase in EMT ( OR=1.24, 95% CI=1.13-1.26; OR=1.26, 95% CI=1.14-1.40). When EMT exceeded 10.0 mm, the clinical pregnancy rate and the live birth rate did not increase significantly with the increase of EMT. Conclusion:In HRT-FET cycle, the endometrial thickness has an effect on the clinical pregnancy rate and the live birth rate in the normal weight group and the overweight group. The clinical pregnancy rate and the live birth rate were the best when the EMT was between 10.0-13.5/10.0-12.7 mm and 10.0-14.0/10.0-12.5 mm, respectively. Whether the endometrium was too thin or too thick would affect the clinical pregnancy outcome. The influence of EMT on clinical pregnancy rate and live birth rate was linear between the low weight group and the obese group, but further study is needed.

Objective:To investigate the effect of cesarean scar defect and endometrial cavity fluid on the pregnancy outcomes of infertility patients with previous cesarean scar uterus undergoing in vitro fertilization-embryo transfer (IVF-ET). Methods:This was a retrospective cohort study. Totally 732 cases of patients with previous cesarean scar uterus were selected from frozen-thawed embryo transfer (FET) cycles in the Fertility Center, Shenzhen Zhongshan Urology Hospital from January 2019 to March 2020. They were divided into four groups: group A ( n=39) including patients with previous cesarean scar defect and without endometrial cavity fluid; group B ( n=82) including patients with previous cesarean scar defect and with endometrial cavity fluid; group C ( n=495) including patients without previous cesarean scar defect and without endometrial cavity fluid; group D ( n=116) including patients without previous cesarean scar defect and with endometrial cavity fluid. The general data and pregnancy outcomes were compared among these groups. Multivariate logistics regression analysis of pregnancy outcome indexes was performed. Results:The transplantation age of group A was higher than that of group B [(38.33±3.55) years vs. (36.93±3.59) years, P=0.045], the endometrial thickness of luteal transformation day and the rate of good-quality embryo transplantation of group C were higher than those of group D [(9.40±1.56) mm vs. (9.03±1.59) mm, P=0.025; 75.76% (375/495) vs. 65.52% (76/116), P=0.024]. The egg retrieval age of group A was higher than that of group C [(37.72±3.55) years vs. (36.25±4.52) years, P=0.049], but the endometrial thickness of luteal transformation day was thinner [(8.74±1.58) mm vs. (9.40±1.56) mm, P=0.012], and the differences were statistically significant. The implantation rate, the biochemical pregnancy rate and the clinical pregnancy rate of group C were higher than those of group D [34.23% (230/672) vs. 22.58% (35/155), P=0.007; 48.28% (239/495) vs. 37.93% (44/116), P=0.044; 42.83% (212/495) vs. 30.17% (35/116), P=0.012]. The early abortion rate in group D was higher than that in group B [40.00% (14/35) vs. 17.24 % (5/29), P=0.047], the difference was statistically significant. Multivariate logistics regression analyses were used with adjustment for possible confounders: the maternal age at embryo transfer, the maternal age at egg retrieval, anti-Müllerian hormone (AMH),antral follicle count (AFC), basic follicle-stimulating hormone (FSH), endometrial CD138 results, FET protocol, and embryo attributes, the number of embryos transferred, embryo quality, the endometrial thickness and the progesterone value on the day of luteal transformation, the result showed that the implantation rate, the biochemical pregnancy rate and the clinical pregnancy rate of group D were lower than those of group C [22.58% (35/155) vs. 34.23% (230/672), P=0.006; 37.93% (44/116) vs. 48.28% (239/495), P=0.047; 30.17% (35/116) vs. 42.83% (212/495), P=0.022] and the differences were statistically significant. The implantation rate, the biochemical pregnancy rate and the clinical pregnancy rate of group B were higher than those of group D [29.52% (31/105) vs. 22.58% (35/155), P=0.049; 48.78% (40/82) vs. 37.93% (44/116), P=0.012; 35.37% (29/82) vs. 30.17% (35/116), P=0.030] and the differences were statistically significant. Conclusion:Endometrial cavity fluid is the main factor that obviously affects the pregnancy outcome of infertility patients with previous cesarean scar uterus undergoing FET cycles.

Objective:To investigate the effect of endometrial thickness (EMT) on the clinical outcome of hormone replacement frozen-thawed embryo transfer (HRT-FET) cycle in different body mass index (BMI) groups, and to analyze the threshold and optimal EMT and EMT interval corresponding to the ideal clinical pregnancy rate.Methods:A retrospective cohort study was conducted on 10 239 HRT-FET cycles in the Reproductive Medicine Center of Henan Provincial People's Hospital from January 2013 to December 2017, and they were divided into low weight group (BMI<18.5 kg/m 2), normal weight group (BMI=18.5-24.9 kg/m 2), overweight group (BMI=25.0-29.9 kg/m 2) and obese group (BMI≥30.0 kg/m 2). Four subgroups were divided according to EMT, respectively EMT<8.0 mm, 8.0 mm≤EMT<10.0 mm, 10.0 mm≤EMT<12.0 mm, EMT≥12.0 mm. The clinical characteristics and outcome indicators of different EMT subgroups in different BMI groups were compared. To achieve the ideal clinical pregnancy rate, multiple regression analysis, curve fitting and threshold effect analysis were used to find the best EMT and thickness interval. Results:1) After adjusting for confounding factors, multiple regression analysis showed that, there were no significant differences in clinical pregnancy rate and live birth rate among subgroups with the increase of EMT (all groups P>0.05). The clinical pregnancy rate and the live birth rate increased with the increase of EMT between subgroups of normal body weight group and super-recombinant subgroups (all P<0.001 for normal body weight subgroups, P=0.123, P=0.009, P=0.016 and all P<0.001 for super-recombinant subgroups). In the obesity group, with the increase of EMT, the clinical pregnancy rate did not increase significantly except EMT≥12.0 mm subgroup ( P=0.449, P=0.279, P=0.021), while the live birth rate increased significantly ( P=0.014, P=0.005, P<0.001). 2) Curve fitting showed that in the population of low weight and obese, influence of EMT on clinical pregnancy rate was a straight line, in the population of normal weight and overweight, influence of EMT on clinical pregnancy rate was a curve, as EMT increased the clinical pregnancy rate raised and then decreased, the impact on the live birth rate appeared similar. 3) According to the curve fitting, the threshold effect analysis of the normal weight group showed that the endometrial inflection point of EMT on the clinical pregnancy rate and the live birth rate was 10.0 mm. When EMT was lower than 10.0 mm, the clinical pregnancy rate and the live birth rate increased by 20% and 19% for every 1.0 mm increase in endometrial thickness ( OR=1.20, 95% CI=1.13-1.26; OR=1.13,95% CI=1.13-1.26). In overweight group, the inflection point of EMT on the clinical pregnancy rate and the live birth rate was also 10.0 mm. When EMT was lower than 10.0 mm, the clinical pregnancy rate and the live birth rate increased by 24% and 26% for every 1.0 mm increase in EMT ( OR=1.24, 95% CI=1.13-1.26; OR=1.26, 95% CI=1.14-1.40). When EMT exceeded 10.0 mm, the clinical pregnancy rate and the live birth rate did not increase significantly with the increase of EMT. Conclusion:In HRT-FET cycle, the endometrial thickness has an effect on the clinical pregnancy rate and the live birth rate in the normal weight group and the overweight group. The clinical pregnancy rate and the live birth rate were the best when the EMT was between 10.0-13.5/10.0-12.7 mm and 10.0-14.0/10.0-12.5 mm, respectively. Whether the endometrium was too thin or too thick would affect the clinical pregnancy outcome. The influence of EMT on clinical pregnancy rate and live birth rate was linear between the low weight group and the obese group, but further study is needed.

Objective:To investigate the effect of cesarean scar defect and endometrial cavity fluid on the pregnancy outcomes of infertility patients with previous cesarean scar uterus undergoing in vitro fertilization-embryo transfer (IVF-ET). Methods:This was a retrospective cohort study. Totally 732 cases of patients with previous cesarean scar uterus were selected from frozen-thawed embryo transfer (FET) cycles in the Fertility Center, Shenzhen Zhongshan Urology Hospital from January 2019 to March 2020. They were divided into four groups: group A ( n=39) including patients with previous cesarean scar defect and without endometrial cavity fluid; group B ( n=82) including patients with previous cesarean scar defect and with endometrial cavity fluid; group C ( n=495) including patients without previous cesarean scar defect and without endometrial cavity fluid; group D ( n=116) including patients without previous cesarean scar defect and with endometrial cavity fluid. The general data and pregnancy outcomes were compared among these groups. Multivariate logistics regression analysis of pregnancy outcome indexes was performed. Results:The transplantation age of group A was higher than that of group B [(38.33±3.55) years vs. (36.93±3.59) years, P=0.045], the endometrial thickness of luteal transformation day and the rate of good-quality embryo transplantation of group C were higher than those of group D [(9.40±1.56) mm vs. (9.03±1.59) mm, P=0.025; 75.76% (375/495) vs. 65.52% (76/116), P=0.024]. The egg retrieval age of group A was higher than that of group C [(37.72±3.55) years vs. (36.25±4.52) years, P=0.049], but the endometrial thickness of luteal transformation day was thinner [(8.74±1.58) mm vs. (9.40±1.56) mm, P=0.012], and the differences were statistically significant. The implantation rate, the biochemical pregnancy rate and the clinical pregnancy rate of group C were higher than those of group D [34.23% (230/672) vs. 22.58% (35/155), P=0.007; 48.28% (239/495) vs. 37.93% (44/116), P=0.044; 42.83% (212/495) vs. 30.17% (35/116), P=0.012]. The early abortion rate in group D was higher than that in group B [40.00% (14/35) vs. 17.24 % (5/29), P=0.047], the difference was statistically significant. Multivariate logistics regression analyses were used with adjustment for possible confounders: the maternal age at embryo transfer, the maternal age at egg retrieval, anti-Müllerian hormone (AMH),antral follicle count (AFC), basic follicle-stimulating hormone (FSH), endometrial CD138 results, FET protocol, and embryo attributes, the number of embryos transferred, embryo quality, the endometrial thickness and the progesterone value on the day of luteal transformation, the result showed that the implantation rate, the biochemical pregnancy rate and the clinical pregnancy rate of group D were lower than those of group C [22.58% (35/155) vs. 34.23% (230/672), P=0.006; 37.93% (44/116) vs. 48.28% (239/495), P=0.047; 30.17% (35/116) vs. 42.83% (212/495), P=0.022] and the differences were statistically significant. The implantation rate, the biochemical pregnancy rate and the clinical pregnancy rate of group B were higher than those of group D [29.52% (31/105) vs. 22.58% (35/155), P=0.049; 48.78% (40/82) vs. 37.93% (44/116), P=0.012; 35.37% (29/82) vs. 30.17% (35/116), P=0.030] and the differences were statistically significant. Conclusion:Endometrial cavity fluid is the main factor that obviously affects the pregnancy outcome of infertility patients with previous cesarean scar uterus undergoing FET cycles.

Objective To explore the causes of Fra-1 high expression in breast cancer cells and molecular mechanism of Fra-1 high transcription activity .Methods Two human breast cancer cell lines MDA231 and MCF-7 and 1 human umbilical vein endothelial cell ECV304 were selected as the research objects .Real time fluorescent quantitative reverse transcription -polymerase chain re-action and Western blotting were used to detect the Fra-1 mRNA and protein expression ;the Fra-1 dual fluorescence reporter vector was constructed ,and the Fra-1 promotor transcription activity was detected ;the Fra-1 promotor short-cut reporter vector and mutation reporter vector at related loci were constructed ,and the transcription activity of short-cut reporter vector and mutation reporter vector was detected ;the binding situation of specific probe marked by biotin with activator protein-1(AP1) was observed by using the gel migration block test .Results The relative expression of mRNA and protein and promotor whole length reporter vector transcription activity of Fra-1 in human breast cancer cell lines MDA231 and MCF-7 cells were higher than that in human umbilical vein endothelial cell MCV304 ,the difference was statistically significant (P＜0 .05);in constructed series Fra-1 promotor shortcut reporter vectors ,only the activity of pGL3B-256 was obviously decreased ,the difference was statistically significant (P＜0 .05);the activity of AP1 mutation reporter vector pGL3B-M2 and SP1 and AP1 dual mutation reporter vector pGL3B-M3 was significantly lower than that of wild repoter vector pGL 3B-M1 ,the difference was statistically significant (P＜0 .05);the gel migration block test found that the binding of specific probe marked by biotin with AP1 had obvious blocking effect .Conclusion In in vitro cultured breast cancer cell lines MDA231 and MCF-7 ,the trans-acting factor AP1 up-regulates its expression by activating Fra-1 gene promoter .

To explore the automated immunostainer screening anaplastic lymphoma kinase (ALK) gene fusion non-small cell lung cancer (NSCLC) and clinicopathological characteristics of the molecular subtype lung cancers. Methods Five hundred and sixty-six cases of NSCLC were collected over a 16 month period. The test for ALK was performed by Ventana automated immunostainer with anti-ALK D5F3. The histological features, treatment and outcome of patients were assessed. Results Thirty-eight cases (6.7%, 38/566) of NSCLC showed ALK gene fusion. The frequency of ALK gene fusion was higher in male (7.1%, 25/350) than that in female (6.0%, 13/216) patients, but not achieving statistical significance (chi2 = 0.270, P = 0.604). ALK + NSCLC was more significantly more frequent in patients < or = 60 years (9.9%, 28/282) than >60 years (3.5% , 10/284) of age. Histologically, the ALK + NSCLCs were mostly adenocarcinoma (81.6%, 31/38) , among which eighteen cases were solid predominant subtype with mucin production; nine cases were acinar predominant subtype; one case was papillary predominant subtype and three cases were invasive mucinous adenocarcinoma. The ALK + non-adenocarcinoma included three cases of squamous cell carcinoma, three cases of adenosquamous carcinoma and one case of pleomorphic carcinoma. Among the ALK + NSCLC patients, the number of non/light cigarette smokers (86. 8% , 33/38) was more than that of heavy smokers. Twenty-nine cases were stages III and IV; twenty-nine cases showed lymph node metastasis; twenty cases showed metastases mostly to brain and bone; and one case showed EGFR gene mutation coexisting with ALK gene fusion. Twelve of fifteen patients received crizotinib therapy and remained stable. Conclusions NSCLC with ALK gene rearrangement shows distinctive clinical and histological features. Ventana-IHC may he a feasible and valid technique for detection of ALK rearrangement in NSCLC.
Adenocarcinoma ; genetics ; pathology ; Carcinoma, Adenosquamous ; genetics ; pathology ; Carcinoma, Non-Small-Cell Lung ; genetics ; pathology ; Carcinoma, Squamous Cell ; genetics ; pathology ; Female ; Gene Fusion ; Gene Rearrangement ; Humans ; Lung Neoplasms ; genetics ; pathology ; Male ; Middle Aged ; Protein Kinase Inhibitors ; therapeutic use ; Pyrazoles ; therapeutic use ; Pyridines ; therapeutic use ; Receptor Protein-Tyrosine Kinases ; genetics ; Sex Factors