Objective To research the homology and cross reaction characteristics of human papillomavirus(HPV)16 type L2 N-terminal(1-200)protein in clinical common HPV infection types.Methods The amino acid sequences of the common HPV infection types(6,11,16,18 ,etc.)were blasted and it was found that 1-200 N-terminal sequence of L2 protein was highly homologous.The gene of HPV16 L2(1-200)was amplificated from tissue sample of cervical cancer patient and inserted into the prokaryotic expression vector PGEX-4T-1 to construct the recombinant plasmid PGEX-4T-1-HPV16 L2(1-200).After sequencing identification,the recombinant plasmid was tranformed into E.coli BL21(DE3).Induced by IPTG,the fusion protein containing HPV16 L2(1-200)was expressed and analyzed by both SDS-PAGE and Western blot.Furthermore,the specific binding capacity of the fusion protein to the HPV 6,11,16 and 18 DNA positive patient serums were analyzed by Western blot.The fusion protein was purified with Ni-NTA Agarose Kit and coated with ELISA reaction plates.The specific serum IgG of 98 condyloma acuminatum patients,135 cervix cancer patients and 96 healthy control subjects were detected respectively by indirect ELISA.Results After comparing the amino acid sequences of the common HPV infection types(HPV6,11,16,18,etc.),We found that the homology of HPV L2(1-200)reached 52.7%-74.3%.The recombinant plasmid PGEX-4T-1-HPV 16 L2(1-200)was constructed successfully.Highly expressed HPV16 L2(1-200)fusion protein was obtained and the expression level was account for up to about 22.6% of total bacterial protein.The relative molecular mass(Mr)of the fusion protein is about 49×103,which matches up to the expected Mr Meanwhile,the serums of HPV 6,11,16,18 DNA positive patients were used as the first antibody and the specific band was detected respectively at about 49 × 103 by Western blot.Indirect ELISA showed that the A490 values of the specific IgG of condyloma acuminatum group,cervical cancer group and healthy control subjects were 0.753 ± 0.262,0.756 ± 0.274 and 0.178 ± 0.157 with the positive rate were 89.8%,88.9% and 9.4% respectively.There was no significance of the specific IgG between condyloma acuminatum group and cervical cancer group(P＞0.05),but it was significant among the three groups(P＜0.001).Conclusion The N-terminal 1-200 amino acids of HPV L2 has high homology and there exits cross reaction among the most common HPV infection types.

Objective:To investigate the effect of miR-369-3p targeting ACTN4 expression on proliferation and apoptosis of hepatocellular carcinoma cells.Methods:Real-time quantitative polymerase chain reaction (RT-qPCR) and western blot were used to detect the expression levels of miR-369-3p and ACTN4 in hepatocarcinoma tissues and adjacent tissues. MiR-369-3p mimics, miR-negative control (NC), si-ACTN4, and si-NC were transfected into hepatocellular carcinoma MHCC97H cells by liposome method. Cell proliferation was detected by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-dipheny-ltetrazolium bromide (MTT) assay. Flow cytometry was used to detect cell cycle and apoptotic rates. The dual luciferase reporter assay was used to verify the targeted regulation of ACTN4 by miR-369-3p. Western blot was used to detect the expressions of cyclin D1, p21, Bcl-2 and Bax.Results:The expression level of miR-369-3p in liver cancer tissue was lower than that in adjacent tissues [(0.46±0.04) vs (1.00±0.08), P<0.001)], while the expression level of ACTN4 was higher than that in adjacent tissues [mRNA (3.12±0.29) vs (1.01±0.09); protein (0.61±0.06) vs (0.25±0.03), P<0.001]. Overexpression of miR-369-3p significantly decreased the cell viability[(0.71±0.06) vs (1.26±0.11), P<0.001)], increased cell apoptosis rate [(20.16±2.11)% vs (6.25±0.64)%, P<0.001], increased the proportion of cells in G 1 phase [(31.14±3.36)% vs (51.56±5.23)%, P<0.001], decreased the proportion of cells in S phase [(32.44±3.56)% vs (14.33) ±1.45)%, P<0.001], increased the levels of p21 and Bax protein ( P<0.001), and decreased the levels of cyclin D1 and Bcl-2 protein ( P<0.001). Inhibition of the expression of ACTN4 significantly reduced the cell viability [(0.78±0.07) vs (1.24±0.12), P<0.001], increased the apoptosis rate [(6.58±0.66)% vs (18.32±1.82)%, P<0.001], increased the proportion of cells in G 1 phase [(48.69±4.21)% vs (30.33±3.01)%, P<0.001], decreased the proportion of cells in S phase [(36.21±3.42)% vs (18.54±1.61)%, P<0.001], increased the protein levels of p21 and Bax ( P<0.001), and decreased the levels of cyclin D1 and Bcl-2 protein ( P<0.001). Compared with the miR-369-3p+ pcDNA group, overexpression of ACTN4 increased the proliferation ability of hepatocellular carcinoma MHCC97H cells at 72 hours of culture[(1.12±0.11) vs (0.68±0.06), P<0.001], significantly reduced the proportion of cells in G 1 stage [(38.81±3.24)% vs (51.80±4.57)%, P<0.001], significantly increased the proportion of S-phase cells [(31.65±3.11)% vs (15.69±1.44)%, P<0.001], decreased cell apoptosis rate [(13.86±1.37)% vs (22.69±2.24)%, P<0.001], increased protein expressions of cyclin D1 and Bcl-2 ( P<0.001), decreased the protein expressions of p21 and Bax ( P<0.001). Conclusion:MiR-369-3p can induce cell cycle arrest in G 1 phase, inhibit the proliferation and promote apoptosis of liver cancer cells by regulating the expression of ACTN4.

Objective:To investigate the effect of miR-369-3p targeting ACTN4 expression on proliferation and apoptosis of hepatocellular carcinoma cells.Methods:Real-time quantitative polymerase chain reaction (RT-qPCR) and western blot were used to detect the expression levels of miR-369-3p and ACTN4 in hepatocarcinoma tissues and adjacent tissues. MiR-369-3p mimics, miR-negative control (NC), si-ACTN4, and si-NC were transfected into hepatocellular carcinoma MHCC97H cells by liposome method. Cell proliferation was detected by 3-(4, 5-Dimethylthiazol-2-yl)-2, 5-dipheny-ltetrazolium bromide (MTT) assay. Flow cytometry was used to detect cell cycle and apoptotic rates. The dual luciferase reporter assay was used to verify the targeted regulation of ACTN4 by miR-369-3p. Western blot was used to detect the expressions of cyclin D1, p21, Bcl-2 and Bax.Results:The expression level of miR-369-3p in liver cancer tissue was lower than that in adjacent tissues [(0.46±0.04) vs (1.00±0.08), P<0.001)], while the expression level of ACTN4 was higher than that in adjacent tissues [mRNA (3.12±0.29) vs (1.01±0.09); protein (0.61±0.06) vs (0.25±0.03), P<0.001]. Overexpression of miR-369-3p significantly decreased the cell viability[(0.71±0.06) vs (1.26±0.11), P<0.001)], increased cell apoptosis rate [(20.16±2.11)% vs (6.25±0.64)%, P<0.001], increased the proportion of cells in G 1 phase [(31.14±3.36)% vs (51.56±5.23)%, P<0.001], decreased the proportion of cells in S phase [(32.44±3.56)% vs (14.33) ±1.45)%, P<0.001], increased the levels of p21 and Bax protein ( P<0.001), and decreased the levels of cyclin D1 and Bcl-2 protein ( P<0.001). Inhibition of the expression of ACTN4 significantly reduced the cell viability [(0.78±0.07) vs (1.24±0.12), P<0.001], increased the apoptosis rate [(6.58±0.66)% vs (18.32±1.82)%, P<0.001], increased the proportion of cells in G 1 phase [(48.69±4.21)% vs (30.33±3.01)%, P<0.001], decreased the proportion of cells in S phase [(36.21±3.42)% vs (18.54±1.61)%, P<0.001], increased the protein levels of p21 and Bax ( P<0.001), and decreased the levels of cyclin D1 and Bcl-2 protein ( P<0.001). Compared with the miR-369-3p+ pcDNA group, overexpression of ACTN4 increased the proliferation ability of hepatocellular carcinoma MHCC97H cells at 72 hours of culture[(1.12±0.11) vs (0.68±0.06), P<0.001], significantly reduced the proportion of cells in G 1 stage [(38.81±3.24)% vs (51.80±4.57)%, P<0.001], significantly increased the proportion of S-phase cells [(31.65±3.11)% vs (15.69±1.44)%, P<0.001], decreased cell apoptosis rate [(13.86±1.37)% vs (22.69±2.24)%, P<0.001], increased protein expressions of cyclin D1 and Bcl-2 ( P<0.001), decreased the protein expressions of p21 and Bax ( P<0.001). Conclusion:MiR-369-3p can induce cell cycle arrest in G 1 phase, inhibit the proliferation and promote apoptosis of liver cancer cells by regulating the expression of ACTN4.

Objective:To explore the incidence and risk factors of parastomal hernia after colostomy.Methods:The retrospective cohort study was conducted. The clinicopathological data of 145 patients undergoing colostomy in Xinxiang Central Hospital from January 2015 to January 2019 were collected. There were 86 males and 59 females, aged(59±11) years. Patients received pelvic and abdominal computed tomography once every 6 months after colostomy to detect the occurrence of parastomal hernia. Measurement data with normal distribution were represented as Mean± SD, and the independent sample t test was used for comparison between groups. Measurement data with skewed distribution were represented as M(range). Count data were represented as absolute numbers, and chi-square test or Fisher exact probability was used for comparison between groups. Kaplan-Meier method was used to analyze the cumulative annual incidence of parastomal hernia. Logarithmic rank test was used to analyze the cumulative incidence based on clinical variables. COX proportional hazard regression model was used for univariate and multivariate analyses. Results:(1) Incidence of parastomal hernia after colostomy. All the 145 patients were followed up for 86(range, 60?108)months after colostomy, of which 46 cases had parastomal hernia and 99 cases had no parastomal hernia. There were significant differences in gender, age, body mass index (BMI) and chronic liver disease between patients with and without parastomal hernia after colostomy ( χ2=23.28, t=13.27, χ2=6.17, 5.82, P<0.05). (2) Annual cumulative incidence of parastomal hernia after colostomy. The 1-, 3-, and 5-year cumulative incidence of parastromal hernia after colostomy was 8.5%, 26.4% and 42.7%, respectively. When the follow-up time is more than 5 years, the incidence of parastromal hernia tended to be stable. The 5-year incidence of parastomal hernia after colostomy in female patients was higher than that in male patients (70.7% vs 20.3%, χ2=12.37, P<0.05). The 5-year incidence of parastomal hernia after colostomy in patients≥60 years old was higher than that in patients under 60 years old (49.8% vs 20.0%, χ2=10.52, P<0.05). The 5-year incidence of parastomal hernia after colostomy in patients with BMI >28 kg/m 2 was higher than that in patients with BMI ≤28 kg/m 2 (55.3% vs 33.2%, χ2=11.76, P<0.05). The 5-year incidence of parastomal hernia after colostomy in patients with chronic liver disease was higher than that in patients with non-chronic liver disease (45.2% vs 32.4%, χ2=15.32, P<0.05). (3) Analysis of risk factors for parastomal hernia after colostomy. Results of multivariate analysis showed that female, age >60 years old, BMI ≥28 kg/m 2 and chronic liver disease were independent risk factors for parastomal hernia after colostomy ( hazard ratio=2.70, 2.51, 1.85, 5.88, 95% confidence intervals as 1.39?6.74, 1.01?4.59, 1.02?4.87, 1.05?8.24, P<0.05). Conclusions:The incidence of parastomal hernia after colostomy is increasing year by year, and tends to be stable after 5 years. Female, age >60 years old, BMI≥28 kg/m 2, and chronic liver disease are independent risk factors for parastomal hernia after colostomy.

Objective:To explore the feasibility of applying quantitative flow ratio(QFR) to assess the degree of coronary artery functional stenosis before surgery, and to guide coronary artery bypass grafting(CABG) revascularization strategy.Methods:The study prospectively included a total of 154 patients who were electively treated with CABG in the 11th ward of the Department of Cardiac Surgery of Beijing Anzhen Hospital from January 2019 to September 2020, and their coronary angiography visually showed stenosis of the coronary artery to perform QFR analysis to know the diseased blood vessels. For functional stenosis, the surgeon was blinded to the results of QFR analysis before surgery. Collect its baseline data, perioperative data and recent clinical outcomes for summary analysis.Results:One year later, the coronary artery CTA showed that the occlusion rate of functionally significant disease(QFR<0.8) was 5.5%, and that of non-functionally significant disease(QFR≥0.8) was 15.6%. There was no difference in angina class or repeat interventions between patients with or without occluded bypass grafts.Conclusion:According to QFR analysis, coronary arteries with functional non-significant disease have a higher risk of grafts failure than those with functionally significant disease. For coronary arteries with negative QFR lesions, the risk of occlusion of arterial grafts is higher than that of venous. However, this finding is not significantly related to clinical prognosis, because patients with patency or occlusion of the grafts in non-significant lesions have not found excessive angina pectoris or repeated coronary interventions. QFR-guided selection of coronary surgery strategies is safe and feasible.