Objective:To screen key genes of renal clear cell carcinoma based on bioinformatics methods, identify possible microRNA (miRNA)-mRNA action axis, and explore the expression of related genes in clear cell renal cell carcinoma tissues and cells.Methods:Gene expression profiles of GSE40435 and GSE71302 datasets were obtained from the Gene Expression Omnibus (GEO) database. TCGA-KIRC datasets were obtained from The Cancer Genome Atlas (TCGA) database. R software was used to identify the differentially expressed mRNA and miRNA, and the functional enrichment analysis was performed. STRING database and Cytoscape software were used to perform the protein interaction analysis. The prognosis-related differentially expressed miRNA was evaluated by the Oncomir database. The potential targeted genes regulated by miRNA were determined by using TargetScan and miRDB targeted gene prediction tools. The tissue samples and clinicopathological features of 34 patients with clear cell renal cell carcinoma in the First Hospital of Shanxi Medical University from June to December 2021 were collected, and normal renal cell line 293T and clear cell renal cell carcinoma cell line 786O were selected. The real-time fluorescence quantitative polymerase chain reaction (qRT-PCR), was used to detect the relative expression of genes; Western blotting and immunohistochemical staining were used to detect the expression levels of the targeted proteins. The dual luciferase reporter gene assay was carried out to verify the targeting relationship between genes.Results:A total of 1 351 differentially expressed mRNA and 50 differentially expressed miRNA were screened and identified. The result of functional enrichment analysis suggested that the fatty acid metabolism pathway and xenobiotic metabolism pathway were suppressed in clear cell renal cell carcinoma, while the apoptosis and immune response pathways were activated. Protein interaction analysis suggested that the signal transduction and protein ubiquitination pathways might play a key role in clear cell renal cell carcinoma. The screening results showed that miRNA-224-5p (miR-224-5p) was most closely associated with clear cell renal cell carcinoma progression and was highly expressed in tumor tissues, and its prognosis-related target gene was NEDD4L. The relative expression of NEDD4L mRNA in clear cell renal cell carcinoma tissues and paraneoplastic tissues were 0.138±0.103 and 1.000±0.026 ( t = 46.23, P < 0.05), and the relative expression of miR-224-5p was 1.000±0.043 and 0.129±0.108 ( t = 45.28, P < 0.05). The differences of NEDD4L mRNA and miR-224-5p expressions in different grades and stages of clear cell renal cell carcinoma tissues were statistically significant (all P < 0.05). The expression of NEDD4L protein was decreased in clear cell renal cell carcinoma. The relative expression of NEDD4L gene in 293T and 786O cells were 1.000±0.125 and 0.210±0.044 ( t = 17.52, P < 0.05); the relative expressions of miR-224-5p gene were 0.209±0.049 and 1.000±0.234 ( t = 10.61, P < 0.05). The relative expressions of NEDD4L mRNA in miRNA mimic group and negative control group were 0.236±0.062 and 1.000±0.024, and the difference was statistically significant ( t = 43.56, P < 0.05). NEDD4L protein expression was reduced in the miRNA mimic group. Dual luciferase reporter gene assay suggested that NEDD4L was a direct target gene of miR-224-5p. Conclusions:In clear cell renal cell carcinoma, miR-224-5p targets and regulates NEDD4L expression, and this mechanism may be related to carcinogenesis and progression of clear cell renal cell carcinoma.

Objective:To investigate the value of a cuproptosis-related differential long non-coding RNA (lncRNA) scoring formula related to the prognosis of clear cell renal cell carcinoma (ccRCC) patients in the clinical diagnosis, prognosis prediction and treatment options based on bioinformatics.Methods:Gene matrix and clinical data of ccRCC patients were obtained from the Cancer Genome Atlas (TCGA) database (update to 29 March, 2022). The expression data of 539 ccRCC tissues and 72 paracancerous normal tissues were collected from gene matrix; the data of 530 ccRCC were collected from clinical data. Pearson correlation analysis, Wilcoxon signed rank test and univariate Cox proportional risk model were used to analyze the screened cuproptosis-related differential lncRNA related to the prognosis. R software was used to randomly divide 530 ccRCC patients with survival data into training set (266 cases) and validation set (264 cases) according to approximate 1∶1 ratio. LASSO regression analysis was used to construct a cuproptosis-related differential lncRNA scoring formula and cross-validation was performed. Receiver operating characteristic (ROC) curve analysis was used to evaluate the specificity and sensitivity of cuproptosis-related differential lncRNA scoring formula, and the area of the curve (AUC) was calculated. According to the median risk value, all patients were divided into the low-risk group and high-risk group; Kaplan-Meier method was used to analyze the difference in the overall survival (OS) of patients in the low-risk group and high-risk group. T test was used to detect the differences in the risk value of patients with different clinicopathological characteristics. R package rms was used to construct the nomogram for predicting 1-year, 3-year, 5-year OS rates of ccRCC patients, R package pRRophetic was used to predict the half-inhibitory concentration ( IC50) of common targeted drugs such as sorafenib and sunitinib in clinical treatment of ccRCC patients, and IC50 value of patients in low-risk group and high-risk group was compared by using Wilcoxon signed rank test. Tissue samples of 20 ccRCC patients who underwent radical nephrectomy and were diagnosed with pathology and the matched paracancerous normal tissues were collected from the First Hospital of Shanxi Medical University between June 2021 and December 2021. Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of key lncRNA in ccRCC tissues. Results:Based on the expression matrix of 10 cuproptosis genes (FDX1, LIAS, LIPT1, DLD, DLAT, PDHA1, PDHB, MTF1, GLS, CDKN2A) of ccRCC patients in TCGA database, 153 cuproptosis-related differential lncRNA related to the prognosis were identified. According to LASSO regression analysis, a scoring formula of 4 cuproptosis-related differential lncRNA related to the prognosis was obtained, risk value was calculated as 0.020×AC015912.3+0.011×AC026401.3+0.063×AC103706.1+(-0.076)×EPB41L4A-DT. All patients were divided into high-risk group (≥0.76) and low-risk group (<0.76) based on the median value (0.76). ROC curve analysis showed that the scoring formula had good prediction accuracy in 1-year, 3-year, 5-year OS rates. In training set, validation set, the total cohort, the OS of patients in the high-risk group was worse than that in the low-risk group (all P < 0.001). The age, pathological degree, tumor staging, risk value calculated by cuproptosis-related differential lncRNA were independent influencing factors of OS (all P < 0.001). There were statistically significant differences in the risk value calculated by cuproptosis-related differential lncRNA scoring formula among patients with different pathological degree, tumor staging, T staging, N staging, M staging (all P < 0.01), while there were no statistically significant differences among patients with different gender and age (all P > 0.05). The established nomogram had good prediction accuracy in the 1-year, 3-year, 5-year OS rates. Sunitinib and sirolimus showed higher sensitivity in the high-risk group; axitinib, sorafenib and pazopanib showed higher sensitivity in the low-risk group. qRT-PCR results showed that relative expression level of AC015912.3 in ccRCC tissues was up-regulated compared with paracancerous tissues (1.00±0.04 vs. 0.68±0.24, t = 6.37, P < 0.01); the relative expression level of AC026401.3 in ccRCC tissues was up-regulated compared with paracancerous tissues (1.00±0.05 vs. 0.64±0.22, t = 7.29, P < 0.01); the relative expression level of AC103706.1 in ccRCC tissues was up-regulated compared with paracancerous tissues (1.00±0.04 vs. 0.64±0.21, t = 7.49, P < 0.01); the relative expression level of EPB41L4A-DT in ccRCC tissues was up-regulated compared with paracancerous tissues (1.00±0.06 vs. 0.73±0.10, t = 10.68, P < 0.01). Conclusions:Cuproptosis-related differential lncRNA scoring formula based on TCGA database can be used as a new marker for clinical diagnosis and prognosis prediction of ccRCC patients, which can help guide the clinical drug treatment of patients and facilitate accurate diagnosis and treatment.

Objective:To study the relationship between red blood cell distribution width(RDW)and short-term mortality in elderly patients with hip fragility fractures.Methods:The clinical data and blood routine test at admission of 205 elderly patients with brittle hip fractures who were admitted to our hospital from 2020 to 2021 and were followed up for one year were retrospectively analyzed.The comorbid conditions, RDW and cumulative mortality at 6 months and 1 year after fractures were counted, and the relationship between RDW and short-term mortality were analyzed.Results:The 6-month(6.7% and 20.8%, χ2=8.591, P=0.003)and 1-year(6.7% and 26.7%, χ2=14.818, P<0.001)mortality of patients with ≤1 comorbidity were significantly lower than those of patients with ≥2 comorbidities.Moreover, the 6-month and 1-year mortality in patients with RDW>13.5% were significantly higher than those of patients with RDW ≤ 13.5%.The proportion of RDW>13.5 % in patients with at least two comorbidities was significantly higher than that in patients with ≤1 comorbidity.Taking RDW=13.6% as the cut-off value of 6-month and 1-year mortality, the sensitivity and specificity for predicting 6-month mortality were 71.4 % and 59.9 %, respectively, and the sensitivity and specificity for predicting 1-year mortality were 64.7 % and 59.6 %, respectively. Conclusions:Red cell distribution width is associated with short-term mortality, and higher RDW is associated with a higher risk of mortality among elderly patients with brittle hip fractures.

Background::High-dose dual therapy (HDDT) with proton pump inhibitors (PPIs) and amoxicillin has attracted widespread attention due to its favorable efficacy in eradicating Helicobacter pylori ( H. pylori). This study aimed to compare the efficacy and safety of high-dose PPI–amoxicillin dual therapy and bismuth-containing quadruple therapy for H. pylori rescue treatment. Methods::This was a prospective, randomized, multicenter, non-inferiority trial. Patients recruited from eight centers who had failed previous treatment were randomly (1:1) allocated to two eradication groups: HDDT (esomeprazole 40 mg and amoxicillin 1000 mg three times daily; the HDDT group) and bismuth-containing quadruple therapy (esomeprazole 40 mg, bismuth potassium citrate 220 mg, and furazolidone 100 mg twice daily, combined with tetracycline 500 mg three times daily; the tetracycline, furazolidone, esomeprazole, and bismuth [TFEB] group) for 14 days. The primary endpoint was the H. pylori eradication rate. The secondary endpoints were adverse effects, symptom improvement rates, and patient compliance. Results::A total of 658 patients who met the criteria were enrolled in this study. The HDDT group achieved eradication rates of 75.4% (248/329), 81.0% (248/306), and 81.3% (248/305) asdetermined by the intention-to-treat (ITT), modified intention-to-treat (MITT), and per-protocol (PP) analyses, respectively. The eradication rates were similar to those in the TFEB group: 78.1% (257/329), 84.2% (257/305), and 85.1% (257/302). The lower 95% confidence interval boundary (–9.19% in the ITT analysis, –9.21% in the MITT analysis, and –9.73% in the PP analysis) was greater than the predefined non-inferiority margin of –10%, establishing a non-inferiority of the HDDT group vs. the TFEB group. The incidence of adverse events in the HDDT group was significantly lower than that in the TFEB group (11.1% vs. 26.8%, P < 0.001). Symptom improvement rates and patients’ compliance were similar between the two groups. Conclusions::Fourteen-day HDDT is non-inferior to bismuth-containing quadruple therapy, with fewer adverse effects and good treatment compliance, suggesting HDDT as an alternative for H. pylori rescue treatment in the local region. Trial registration::Clinicaltrials.gov, NCT04678492.

Objective:To establish autoverification rules for coagulation tests in multicenter cooperative units, in order to reduce workload for manual review of suspected results and shorten turnaround time (TAT) of test reports, while ensure the accuracy of results.Methods:A total of 14 394 blood samples were collected from fourteen hospitals during December 2019 to March 2020. These samples included: Rules Establishment Group 11 230 cases, including 1 182 cases for Delta check rules; Rules Validation Group 3 164 cases, including 487cases for Delta check; Clinical Application Trial Group 77 269 cases. Samples were analyzed for coagulation tests using Sysmex CS series automatic coagulation analyzers, and the clinical information, instrument parameters, test results, clinical diagnosis, medication history of anticoagulant and other relative results such as HCT, TG, TBIL, DBIL were summarized; on the basis of historical data, the 2.5 and 97.5 percentile of all data arranged from low to high were initially accumulated; on the basis of clinical suggestions, critical values and specific drug use as well as relative guidelines, autoverification rules and limits were established.The rules were then input into middleware, in which Stage I/Stage II validation was done. Positive coincidence, negative coincidence, false negative, false positive, autoverification pass rate, passing accuracy (coincidence of autoverification and manual verification) were calculated. Autoverification rules underwent trial application in coagulation results reports.Results:(1) The autoverification algorisms involve 33 rules regarding PT/INR, APTT, FBG, D-dimer, FDP,Delta check, reaction curve and sample abnormalities; (2)Autoverification Establishment Group showed autoverification pass rate was 68.42% (7 684/11 230), the false negative rate was 0%(0/11230), coincidence of autoverification and manual verification was 98.51%(11 063/11 230), in which positive coincidence and negative coincidence were respectively 30.09% (3 379/11 230) and 68.42%(7 684/11 230); Autoverification Validation Group showed autoverification pass rate was 60.37%(1 910/3 164), the false negative rate was 0%(0/11 230), coincidence of autoverification and manual verification was 97.79%(3 094/3 164), in which positive coincidence and negative coincidence were respectively 37.42%(1 184/3 164) and 60.37%(1 910/3 164); (3) Trialed implementation of these autoverification rules on 77 269 coagulation samples showed that the average TAT shortened by 8.5 min-83.1 min.Conclusions:This study established 33 autoverification rules in coagulation tests. Validation showedthese rules could ensure test quality while shortening TAT and lighten manual workload.

Objective To explore the effects of miRNA-204 targeted LC3B expression on Ang Ⅱ induced cardiomyocytes hypertrophy.Methods The primary neonatal rat cardiomyocytes served as the research objects and divided into the control group,AngⅡ group,combination-treated group 1 (cardiomyocytes were given Ang Ⅱ stimulation,meanwhile infected by negative control lentivirus vector),combination-treated group 2 (cardiomyocytes were given Ang Ⅱ stimulation,meanwhile infected by lentivirus carrying miRNA-204 overexpression vector) according to different treatments.About 48 h to 72 h after intervention treatment,the cardiomyocyte hypertrophy change was detected by confocal microscopy,the expression of miRNA-204 was analyzed by real time PCR,the protein expression of LC3B was measured by Western blot and targeted gene of miRNA-204 was demonstrated by dual-luciferase reporter assay system.Results Compared with the control group,the cardiomyocyte relative surface area in the Ang Ⅱ group was significantly enlarged,the protein expression of LC3B was significantly increased,the expression of miRNA-204 was upregulated,the differences were statistically significant (P＜0.05).Whereas comparing the combination-treated group 1 with combination-treated group 2,the protein expression of LC3B in the latter was down-regulated and the cell area was reduced (P＜0.05).The further luciferase activity report gene experiment results suggested that miRNA-204 was able to bind to LC3B 3'-UTR and decreased the luciferase activities (P＜0.05),but not to bind its mutated fragment for inactivating luciferase activity(P＞0.05).Conclusion miRNA-204 is able to inhibit Ang Ⅱ induced cardiomyocytes hypertrophy,its action is realized by targeting the expression of LC3B.

AIM:To analyze the correlation between serum angiotensin-converting enzyme 2 (ACE2) levels and different stages of coronary heart disease (CHD), and to explore the change of serum ACE2 level during the development of CHD.METHODS:The control group included 85 non-CHD samples, and 174 CHD samples were divided into light stenosis (ls-CHD, stenosis degree <50%) group, moderate stenosis (ms-CHD, stenosis degree 50%~75%) group and severe stenosis (ss-CHD, stenosis degree ≥75%) group.The ACE2 level in each serum sample was detected by ELISA.The relationship between the ACE2 level and the development of coronary heart disease was explored by statistical analysis of serum ACE2 levels in different stages of CHD.RESULTS:The serum ACE2 levels in ls-CHD group, ms-CHD group and ss-CHD group were all higher than that in control group.The more severe the coronary artery stenosis existed, the higher the ACE2 level was observed.The serum ACE2 level in the males was higher than that in the females.In a single sex, the serum ACE2 levels in ls-CHD group, ms-CHD group and ss-CHD group were higher than that in control group with significant differences.Regression analysis found that sex, diabetes and CHD were associated with the serum ACE2 levels.Among them, sex and CHD were the independent factors to affect serum ACE2 levels.CONCLUSION:The serum ACE2 level of males was higher than that of females.Compared with the non-CHD samples, the serum ACE2 level of CHD patients was higher than that of the non-CHD samples.During the development of coronary heart disease, the serum ACE2 level increased constantly.

Objective To explore the effects of miR-34a and the alteration of AMPK/mTOR signal pathway on angiotensin (Ang)Ⅱ-stimulated cardiomyocytes hypertrophy. Methods Neonatal rat cardiomyocytes were cultrued in vitro and were divided into 3 groups:negative control for lentivirus carrying miR-34a group (NC), AngⅡplus lentivirus carrying negative control group (AngⅡ+NC) and AngⅡplus lentivirus carrying miR-34a group (AngⅡ+miR-34a). The relative cell area was detected by confocal microscopy. The expression of miR-34a and hypertrophy-related genes (ANP and β-MHC) were analyzed by real time PCR. The AMPK/mTOR signal pathway was measured by Western blot assay. Results Compared to NC group, the relative cell area was increased in AngⅡ+NC group (P<0.05). But compared with AngⅡ+NC group, the relative cell area was decreased in AngII+miR-34a group (P<0.05). Moreover, compared with NC group, the expression of miR-34a was decreased, and the expression of hyperthophy-related genes(ANP and β-MHC) was up-regulated in AngⅡ+NC group. However, the expression of miR-34a was decreased, and the expression of hyperthophy-related genes (ANP and β-MHC) was down-regulated (P<0.05). Finally, compared to NC group, the ratio of phosphop-AMPK/AMPK was significantly induced in AngII + NC group, but the ratio of phosphop-mTOR/mTOR was significantly decreased (P<0.05). However, compared to AngⅡ+NC group, the ratio of phosphop-AMPK/AMPK was significantly decreased in AngII + miR-34a group, but the ratio of phosphop-mTOR/mTOR was significantly increased (P<0.05). Conclusion miR-34a is able to inhibit myocardial hypertrophy of neonatal rat cardiomyocytes, and its mechanism is partly carried out by the alteration of the signal pathway of AMPK/mTOR.

BACKGROUND:Degradable polymer materials initiate the degradation process immediately after implantation. How to regulate the degradation of these materials is rarely reported at present. OBJECTIVE:To study the effect of ultrasonic wave on control ing the degradation of polymer materials. METHODS:The sample is made ofε-caprolactone/L-lactide copolymer, and its core was coated with low density polyethylene on the surface with the fol owing four different methods. (1) The core surface was firstly covered with CaCl 2 powder, and then coated with polyethylene. (2) The core was firstly coated with polyethylene and coarsened for 3 hours. (3) The core surface was firstly covered with CaCl 2 powder, and then coated with polyethylene, and coarsened for 3 hours. (4) The core was directly coated with polyethylene. The four kinds of specimens obtained were embedded in pork for ultrasonic bombardment experiment in vitro. RESULTS AND CONCLUSION:In the specimens prepared with methods 1 and 4, the lyophobic layer could protect core materials before ultrasonic treatment, and no absorption peak was found at 631 nm. After ultrasonic treatment, the lyophobic layer was destroyed, toluidine blue dye was released, leading to change the color of immersion solution and increase the absorption peak at 631 nm. In the specimens prepared with methods 2 and 3,the lyophobic layer cannot exhibit the protection effects, the absorption peak was found at 631 nm. Under electron microscope, the appearance of the specimens in four groups was changed obviously. It is feasible to control the starting of the degradation by coating the degradable copolymer with LDPE and using ultrasonic as a trigger.