BACKGROUND: Epoxyeicosatrienoic acids (EETs), which are metabolites of arachidonic acid catalyzed by cytochrome P450 epoxygenase, are degraded into inactive dihydroxyeicosatrienoic acids by soluble epoxide hydrolase (sEH). Many studies have revealed that sEH gene deletion exerts protective effects against diabetes. Vascular calcification is a common complication of diabetes, but the potential effects of sEH on diabetic vascular calcification are still unknown. METHODS: The level of aortic calcification in wild-type and Ephx2-/- C57BL/6 diabetic mice induced with streptozotocin was evaluated by measuring the aortic calcium content through alizarin red staining, immunohistochemistry staining, and immunofluorescence staining. Mouse vascular smooth muscle cell lines (MOVAS cells) treated with β-glycerol phosphate (0.01 mol/L) plus advanced glycation end products (50 mg/L) were used to investigate the effects of sEH inhibitors or sEH knockdown and EETs on the calcification of vascular smooth muscle cells, which was detected by Western blotting, alizarin red staining, and Von Kossa staining. RESULTS: sEH gene deletion significantly inhibited diabetic vascular calcification by increasing levels of EETs in the aortas of mice. EETs (especially 11,12-EET and 14,15-EET) efficiently prevented the osteogenic transdifferentiation of MOVAS cells by decreasing nidogen-2 (NID2) expression. Interestingly, suppressing sEH activity by small interfering ribonucleic acid or specific inhibitors did not block osteogenic transdifferentiation of MOVAS cells induced by β-glycerol phosphate and advanced glycation end products. NID2 overexpression significantly abolished the inhibitory effect of sEH gene deletion on diabetic vascular calcification. Moreover, NID2 overexpression mediated by adeno-associated virus 9 vectors markedly increased insulin-like growth factor 2 (IGF2) and phospho-ERK1/2 expression in MOVAS cells. Overall, sEH gene knockout inhibited diabetic vascular calcification by decreasing aortic NID2 expression and, then, inactivating the downstream IGF2-ERK1/2 signaling pathway. CONCLUSIONS sEH gene deletion markedly inhibited diabetic vascular calcification through repressed osteogenic transdifferentiation of vascular smooth muscle cells mediated by increased aortic EET levels, which was associated with decreased NID2 expression and inactivation of the downstream IGF2-ERK1/2 signaling pathway.
Animals ; Mice ; Vascular Calcification/metabolism* ; Mice, Inbred C57BL ; Epoxide Hydrolases/metabolism* ; Diabetes Mellitus, Experimental/genetics* ; Male ; Gene Deletion ; MAP Kinase Signaling System/genetics* ; Cell Line ; Immunohistochemistry ; Muscle, Smooth, Vascular/metabolism* ; Signal Transduction/genetics* ; Mice, Knockout

As a new clinical trial mode,decentralized & digitalized clinical trial(DCT)is based on digital health equipment and uses internet and artificial intelligence technologies to complete the screening,registration,randomization,intervention,evaluation and follow-up of subjects,which is helpful to improve efficiency and reduce trial costs.The DCT mode has been applied to evaluate the treatment and management effects of cardiovascular diseases such as atrial fibrilla-tion,heart failure,coronary heart disease,and hypertension,showing broad development prospects and application space.This article will provide a brief introduction to representative DCT in the global cardiovascular disease field,and look for-ward to the application prospects of this model,providing reference and guidance for accelerating the development of cardio-vascular DCT in China.

ObjectiveTo explore the effect of repetitive peripheral magnetic stimulation on upper limb motor function rehabilitation of stroke patients after contralateral seventh cervical nerve transfer (CC7). MethodsFrom May, 2020, to May, 2022, 34 stroke patients with hemiplegia underwent CC7 in Jing'an District Centre Hospital of Shanghai were randomly divided into control group (n = 17) and observation group (n = 17). Both groups received conventional rehabilitation. The observation group accepted repetitive peripheral magnetic stimulation, and the control group received sham stimulation, for eight weeks. They were assessed with Fugl-Meyer Assessment-Upper Extremities (FMA-UE) and Hua-Shan Grading of Upper Extremity (H-S grading) before and after treatment. ResultsTwo cases dropped down in each group. There was difference in gender between two groups (χ² = 6.136, P < 0.05). After treatment, the scores of FMA-UE and H-S grading significantly improved in both groups (t > 4.000, P < 0.01), and the improvement was better in the observation group than in the control group (t > 2.362, P < 0.05). ConclusionRepetitive peripheral magnetic stimulation could improve the motor function of upper limb and hand of stroke patients with hemiplegia after CC7.

Objective:To investigate the clinical efficacy of modified biliary-intestinal anasto-mosis by pancreaticoduodenectomy and influencing factors of postoperative biliary leakage.Methods:The propensity score matching and retrospective cohort study was conducted. The clinicopatholo-gical data of 165 patients with benign and malignant diseases around the ampullary who underwent pancreaticoduodenectomy in the Affiliated Hospital of Inner Mongolia Medical University from June 2014 to October 2020 were collected. There were 92 males and 73 females, aged (59±10)years. Of the 165 patients, 44 patients undergoing modified biliary-intestinal anastomosis within pancreatico-duodenectomy were divided into the modified group, and 121 patients undergoing traditional biliary-intestinal anastomosis within pancreaticoduodenectomy were divided into the traditional group. Observation indicators: (1) propensity score matching and comparison of general data of patients between the two groups after matching; (2) intraoperative and postoperative situations; (3) analysis of influencing factors of biliary leakage after pancreaticoduodenectomy. Propensity score matching was done by the 1:1 nearest neighbor matching method, with the caliper setting as 0.05. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the t test. Measurement data with skewed distribution were represented as M( Q1, Q3), and comparison between groups was conducted using the Mann-Whitney U test. Count data were described as absolute numbers, and comparison between groups was conducted using the chi-square test or Fisher exact probability. Univariate analysis was conducted using the corresponding statistical methods based on data type. All indicators in univariate analysis were included in multivariate analysis. Multivariate analysis was conducted using the Logistic regression model. Results:(1) Propensity score matching and comparison of general data of patients between the two groups after matching. Of the 165 patients, 72 cases were successfully matched, including 36 cases in the modified group and 36 cases in the traditional group, respectively. The elimination of jaundice, preoperative reduction of jaundice and hypertension confounding bias ensured comparability between the two groups after propensity score matching. (2) Intraoperative and postoperative situations. All patients in the two groups underwent surgery successfully. The operation time, postoperative pathological type (lower bile duct cancer, pancreatic head cancer, pancreatic cystic tumor, chronic pancreatitis, duodenal cancer), time of no drainage fluid in the drainage tube around biliary-intestinal anastomosis were 371(270,545)minutes, 6, 12, 1, 2, 15, (12±7)days in patients of the modified group, versus 314(182,483) minutes, 13, 14, 1, 4, 4, (16±8)days in patients of the traditional group, showing significant differences in the above indicators between the two groups ( Z=-3.54, χ2=10.01, t=-2.34, P<0.05). Cases with postoperative grade A biliary leakage was 0 in patients of the modified group, versus 6 in patients of the traditional group, showing a significant difference between the two groups ( P<0.05). Cases with postoperative grade B biliary leakage, cases with postoperative grade B pancreatic fistula, cases with postoperative bleeding, cases with abdominal infection, cases with incision infection, cases with delayed gastric emptying, cases undergoing unplanned readmission were 1, 0, 1, 4, 1, 5, 1 in patients of the modified group, versus 0, 1, 2, 5, 2, 5, 2 in patients of the traditional group, showing no significant difference in the above indicators between the two groups ( P>0.05). Cases with postoperative grade A pancreatic fistula, cases with overall complications, cases with Clavien-Dindo grade Ⅰ-Ⅱ complications, cases with Clavien-Dindo grade Ⅲ-Ⅳ complications were 6, 12, 6, 6 in patients of the modified group, versus 7, 14, 8, 6 in patients of the traditional group, showing no significant difference in the above indicators between the two groups ( χ2=0.09, 0.24, 0.36, 0.00, P>0.05). None of patient in the two groups had postoperative grade C biliary leakage and postoperative grade C pancreatic fistula. (3) Analysis of influencing factors of biliary leakage after pancreaticoduodenectomy. Results of multivariate analysis showed that preoperative reduction of jaundice and traditional biliary-intestinal anastomosis were independent risk factors for biliary leakage after pancreaticoduodenectomy ( odds ratio=11.37, 12.27, 95% confidence interval as 1.76-73.35, 1.14-131.23, P<0.05). Conclusions:Compared with traditional biliary-intestinal anastomosis, modified biliary-intestinal anastomosis within pancreaticoduodenectomy is safe and feasible. Preoperative reduction of jaundice and traditional biliary-intestinal anastomosis are independent risk factors for biliary leakage after pancreaticoduodenectomy.

Objective:To study the safety and efficacy of using the ultrasonic lithotripsy system (ULS) in assisting percutaneous nephroscopic retroperitoneal pancreatic necrosectomy in patients with acute necrotizing pancreatitis (ANP) extending to both sides of the retroperitoneal regions.Methods:The clinical data of 47 patients with extensive ANP who underwent video-assisted retroperitoneal debridement (VARD) from January 2017 to October 2022 at the Affiliated Hangzhou First People's Hospital, Zhejiang University School of Medicine, were analyzed retrospectively. There were 32 males and 15 females, aged [ M( Q1, Q3)] 60(43, 75) years old. The patients were divided into two groups based on the debridement methods: patients treated by nephroscopic pancreatic necrosectomy (NPN) were included in the NPN group ( n=22), while patients treated by the ULS-assisted treatment were included in the ULS group ( n=25). The surgical debridement time, operation time and complications of the two groups were compared. Follow up on recurrence and death of patients was done by telephone, outpatient and/or re-hospitalization records. Results:All patients underwent the VARD operation successfully, without any need for conversion to laparotomy, transfer to intensive care unit and death related to the operations. The pancreatic exocrine function was not damaged in both groups. When compared with the ULS group, the NPN group required significantly more debridement time [3(2, 4) times/person vs. 2(1, 2) times/person], longer operation time [65(40, 85) min vs. 35(30, 50) min] and longer hospitalization time [91(76, 130) d vs. 72(62, 102) d, all P<0.05]. No complications occurred in the ULS group. In the NPN group, postoperative hemorrhage occurred in 3 patients, colon fistula in 1 patient, and delayed viral encephalitis in 1 patient. The incidence of postoperative complications in the NPN group was significantly higher than that in the ULS group [22.7%(5/22) vs. 0(0/25), P=0.032]. All the 47 patients with extensive ANP were followed up for a median of 28 months (range 3 to 60 months), and there were no patients who developed residual recurrence and death. Conclusion:For patients with extensive ANP, ULS-assisted nephroscopic pancreatic necrosectomy was safe and feasible. When compared with NPN, the ULS-assisted procedure showed more advantages in debridement efficiency, operation time and hospital stay.

Objective:To grasp the distribution of fine antigenic epitope profiles of nucleoprotein (NP) and glycoprotein (GP) fragments of Crimean-Congo hemorrhagic fever virus (CCHFV) and to clarify the value of dominant antigenic epitopes in laboratory testing of Crimean-Congo hemorrhagic fever (CCHF).Methods:In a minimal synthetic short peptide consisting of 8 amino acids was segmentally expressed by CCHFV YL04057 strain using a modified bio-peptide synthesis method from 2014 to 2021 in the laboratory of Xinjiang University, College of Life Sciences. Using CCHFV polyclonal antibody or monoclonal antibody 14B7 (IgM) or CCHFV-positive sheep serum as antibodies, the minimal antigenic epitopes (BCEs) with antigenic activity on NP and GP fragments were identified by immunoblotting, and the obtained BCEs with sequence polymorphism were spatially clustered with CCHFV from different regions using the neighbor-joining method to determine the combination mode of BCEs with geographical correlation of regional distribution, to explore its application in establishing serological diagnosis. A prokaryotic expression plasmid (pET-32a), an E. coli expression plasmid (pGEX-KG) and a prokaryotic expression plasmid with an incomplete glutathione (GST188) tag (pXXGST-ST-1) were used to construct and express six dominant antigenic epitopes of different peptide lengths on NP fragments, and an indirect Enzyme-linked immunosorbent assay (ELISA) was established. CCHF sheep serum identified by immunofluorescence assay (IFA) was used as a control, and the specificity, sensitivity and overall compliance of the recombinant proteins with different peptide lengths of antigenic epitopes with IFA assay results were statistically analyzed. Results:CCHFV, NP and GP fragments had a total of 30 antigenically active BCEs, among which the core intermediate fragment NP2 (aa 170 th-305 th), which had a concentration of antigenic epitopes in the NP fragment, has 6 BCEs, and the NP1 (aa 1 st-200 th) and NP3 (aa 286 th-482 nd) at both ends have 9 BCEs; the Gc (aa 1 st-558 th) and Gn (aa 533 th-708 th) fragments of the GP fragment have 14 BCEs and a long antigenic peptide (AP) containing 15 amino acids, and the amino acid sequence homology of the NP fragment BCEs was 97.1% and that of the GP fragment BCEs was 89.1%. There was a significant difference ( P=0.0281, P<0.05). Among the 9 BCEs with sequence polymorphism in the GP fragment, 6 combined BCEs from GnEc1, GnE2, GnE4, GcE3, GcE6 and GcAP-4 (Ap) could cluster 15 CCHFV strains from different regions of the world into 5 geographical taxa, AsiaⅠ, AsiaⅡ, AficaⅠ, AficaⅡ and Europe. The constructs expressing PET-32a-NP (full length), PGEX-KG-NP2 (aa 170 th-305 th), pGEX-KG-NP2-1 (aa 235 th-275 th), PGEX-KG-NP2-1-1 (aa 237 th-256 th), pXXGST-1-NP2-1-2 (aa 250 th-265 th) and PGEX KG-NP2-1-3 (aa 260 th-276 th), six recombinant proteins CCHFV NP rabbit polyclonal antiserum (pAb) Western Blotting reaction positive, 33 sheep sera tested by IFA XHF as a reference, the sensitivity of the assay established by indirect ELISA using the recombinant proteins constructed from two fragments of NP2 and NP2-1 as antigens. The sensitivity, specificity and overall compliance were the best, with 73.4% (11/15) and 66.7% (10/15) for sensitivity, 100% (18/18) and 94.4% (17/18) for specificity, and 87.9% (29/33) and 81.8% (27/33) for overall compliance. Conclusion:CCHFV NP and GP are distributed with a high number of BCEs with antigenic immunoreactivity, among which the dominant antigenic epitopes are of high value in the laboratory serological diagnosis of CCHF.

Objective To investigate the effects of microRNA497 (miR-497) on the metastasis of gastric cancer and its possible molecular mechanism. Methods SGC-7901 gastric cancer parent cells were cultured in an ultra-low adhesion environment, and the anoikis resistance model of SGC-7901 cells was created after re-adhesion. Clone formation assay, flow cytometry, Transwell^TM test and scratch healing test were used to detect the differences of biological behavior compared with their parent cells. Fluorescence quantitative PCR was performed to detect the expression of miR-497. Western blot analysis was used to detect the changes of key proteins of Wnt/β-catenin signaling pathway and epithelial mesenchymal transformation (EMT) related proteins such as vimentin and E-cadherin. Parent cells and anoikis resistant SGC-7901 cells were transfected with miR-497 inhibitor or miR-497 mimic, and CCK-8 assay was used to detect the proliferation activity. Transwell^TM invasion assay was performed to detect the invasion ability of cells. Transwell^TM migration test and scratch healing assay was used to determine the migration ability. Western blot analysis was used to detect the expressions of Wnt1, β-catenin, vimentin and E-cadherin. By transfecting miR-497 mimic into the anoikis resistance SGC-7901 cells and inoculating them subcutaneously in nude mice, the changes in the volume and mass of tumor tissues were measured and recorded. Western blot analysis was used to determine the expressions of Wnt1, β-catenin, vimentin and E-cadherin of tumor tissues. Results Compared with the parent cells, the anoikis resistance SGC-7901 gastric cancer cells had faster proliferation rate, stronger colony formation, lower apoptosis rate, stronger invasion and migration ability. The expression of miR-497 was significantly decreased. After down-regulation of miR-497, the proliferation ability, invasion and migration ability were significantly enhanced. The expressions of Wnt1, β-catenin and vimentin increased significantly, while E-cadherin decreased notably. The results of up-regulation miR-497 were the opposite. The tumor growth rate, tumor volume and mass of miR-497 overexpression group were significantly lower than those of control group. The expressions of Wnt1, β-catenin and vimentin decreased significantly, while the expression of E-cadherin increased significantly. Conclusion The expression of miR-497 is low in the anoikis resistance SGC-7901 cells. miR-497 can inhibit the growth and metastasis of gastric cancer cells by blocking Wnt/β-catenin signaling pathway and EMT.
Animals ; Mice ; Humans ; beta Catenin/metabolism* ; MicroRNAs/metabolism* ; Vimentin/metabolism* ; Stomach Neoplasms/pathology* ; Anoikis/genetics* ; Wnt Signaling Pathway/genetics* ; Mice, Nude ; Cell Proliferation/genetics* ; Cadherins/genetics* ; Cell Line, Tumor ; Epithelial-Mesenchymal Transition/genetics* ; Cell Movement/genetics*

Objective:To study the effect of peroxisome proliferator activated receptor- γ (PPAR-γ) agonists rosiglitazone on the hepatocytes of Sprague Dawley (SD) rats with severe acute pancreatitis (SAP) and the regulatory mechanism.Methods:Seventy two healthy male SD rats, weighing 255-315 g, aged 49-56 days, were randomly divided into SAP model group ( n=24, SAP model preparation), rosiglitazone group ( n=24, rosiglitazone intravenous injection after SAP model preparation) and sham operation group ( n=24, normal saline injection only). After 6 h, 12h and 24 hours of injection, 8 rats were treated at each time point. HE staining was used to study the liver tissue structure and detect the levels of serum tumor necrosis factor-α(TNF-α), interleukin (IL) -1β, IL-6, AST, ALT and lactate dehydrogenase (LDH) in the rats. Western blot was used to detect the expression of high mobility group box-1 protein (HMGB1), Janus activated kinase (JAK)2 and signal transducer and activator of transcription (STAT)3. Results:The levels of serum TNF-α , IL-1β, IL-6, AST, ALT, LDH in SAP model group and rosiglitazone group were significantly higher than those in the sham operation group (all P<0.05). IL-1β at 6, 12h and 24 h in rosiglitazone group was (226.5±52.1)ng/L, (458.2±82.3)ng/L, (556.4±83.4) ng/L, ALT was (158. 3±39.2) U/L, (235.0±44.6)U/L, (298.4±56.6) U/L, which was lower than that in SAP model group (443. 5±62.3) ng/L, (622.6±78.3) ng/L, (789.1±105.7) ng/L and (198.4±42.5)U/L, (253.8±47.0)U/L, (337.2±60.1) U/L, the differences were statistically significant (all P<0.05). AST and LDH in rosiglitazone group were also lower than those in SAP model group at each time point, and the differences were statistically significant (all P<0.05). HE staining showed that there were less inflammation, hemorrhage and necrosis in rosiglitazone group than those in SAP model group. Expression of STAT3 in liver of rosiglitazone group at 6 h, 12 h and 24 h was (0.22±0.03), (0.30±0.04), (0.31±0.06), lower than SAP model group (0. 28±0.04), (0.38±0.05), (0.40±0.06), the differences were statistically significant (all P<0.05). Expression of JAK2 and HMGB1 in rosiglitazone group at 12 h and 24 h was also lower than that in SAP model group (all P<0.05). Conclusion:PPAR-γ agonists rosiglitazone can protect the SAP rats suffering from hepatocyte injury and inflammation, through JAK2/STAT3 pathway.

Objective:Explore the open sharing mode of large-scale scientific instruments in specialized laboratories, consider how to effectively improve the use efficiency of instruments.Methods:Based on the analysis of the problems existing in the management of large scientific instruments in specialized laboratories of a general hospital, the open sharing platform of specialized laboratory instruments was constructed with the help of laboratory information management system, and relevant management suggestions and thoughts were put forward.Results:Depending on the system design, specialized laboratories can use the instrument sharing platform to implement the " special management sharing" of large scientific instruments and improve the efficiency of instrument use.Conclusions:The construction of large scientific research instrument sharing platform for specialized laboratories in general hospitals is an effective means to solve the instrument management problems in specialized laboratories at present, but to give full play to the effect, it is necessary to constantly improve the platform system construction and technical personnel training.

Objective:To analyse the use of nephroscopic peripancreatic necrotic tissue removal through drainage tube sinus tracts without the use of anesthesia.Methods:The data of 17 patients with severe acute pancreatitis (SAP) were treated from August 2016 to December 2019 at the Affiliated Hangzhou First People’s Hospital, Zhejiang University School of Medicine were studied. There were 5 males and 12 females, age ranged from 24 to 80 years (mean 49.3 years). These 17 patients underwent nephroscopic pancreatic necrosis tissue removal through drainage tube sinus tracts of drains using a dual-scope necrosectomy approach without the use of any anesthesia. The surgical related and follow-up data were analysed.Results:Of 17 patients who underwent pancreatic necrosis removal as the first operation, one underwent a further postoperative nephroscopic resection without anesthesia, while 10 patients underwent three nephroscopic resections without anesthesia, and 6 patients underwent four nephroscopic resections without anesthesia. The operative durations of nephroscopic resection without anesthesia was 26 to 37 min. All patients had smooth surgical procedures and they were fed on the same day after surgery. There were no postoperative complications on follow-up after discharge from hospital for 1 to 40 months.Conclusion:Transcatheter sinusoidal nephroscopic pancreatic necrotic tissue removal without anesthesia was safe. There was no need for fasting. The procedure is minimally invasive and it can be repeated, especially for those patients who cannot tolerate any form of anesthesia. This is the least invasive procedure for treatment of SAP.