Objective To compare the safely between 3 ways of inserting the anteroinferior iliac spine channel screws using computer simulation.Methods The spiral pelvic CT data of 100 patients were collected who had sought medical treatment at General Hospital of The Northern Theater Command from October 2017 to October 2018.They were 61 males and 39 females,aged from 20 to 60 years (average,47.5 years).The data were imported into Mimics (Materi-alise,Belgium) software to create three-dimensional models of the pelvis.The pelvic modeling data were then imported into 3-Matic (Materi-alise,Belgium) software for post-processing.Three cylinders with a diameter of 6.7 mm were created,taking the anteroinferior iliac spine as the entry point and taking the posterosuperior iliac spine,the posteroinferior iliac spine and the midpoint between the 2 spines as the 3 exit points.The insertion of anteroinferior iliac spine channel screws was simulated to observe the screw penetration.Results In the channel from the anteroinferior iliac spine to the posterosuperior iliac spine,penetration occurred in 41 cases out of the medial ilium and in 2 cases out of the lateral ilium,giving a penetration rate of 43％ (43/100);in the channel from the anteroinferior iliac spine to the midpoint between the 2 spines,penetration occurred in 16 cases out of the medial ilium and in 2 cases out of the lateral ilium,giving a penetration rate of 18％ (18/100);in the channel from the anteroinferior iliac spine to the posteroinferior iliac spine,penetration occurred in 6 cases out of the medial ilium,in 2 cases out of the lateral ilium,in 60 cases out of the greater sciatic notch and in 8 cases out of both the medial ilium and greater sciatic notch,giving a penetration rate of 76％ (76/100).There were significant differences between the 3 ways of insertion in the screw penetration (x2 =68.219,P ＜ 0.001).The rate of screw penetration in the channel from the anteroinferior iliac spine to the posteroinferior iliac spine was significantly higher than that in the channel from the anteroinferior iliac spine to the posterosuperior iliac spine which was significantly higher than that in the channel from the anteroinferior iliac spine to the midpoint between the 2 spines (P ＜ 0.05).Conclusions The channel from the anteroinferior iliac spine to the midpoint between the posterosuperior iliac spine and the posteroinferior iliac spine may lead to a lower rate of screw penetration while the channel from the anteroinferior iliac spine to the posteroinferior iliac spine may lead to a higher rate of screw penetration.

Objective To investigate the effects of Lactobacillus rhamnosus GG (LGG) colonization in early life on intestinal barrier and intestinal development in offspring mice and its possible mechanism.Methods Six C57BL/6 pregnant mice with the same conception time of 6 weeks were selected and randomly divided into experiment group given 108 cfu/ml LGG live bacteria and control group given LGG inactivated bacteria by gavage from the 18th day of pregnancy until natural birth.The progeny mice in the two groups were continued to be gavaged with 107 cfu/ml of LGG live bacteria or LGG inactivated bacteria on days 1-5 of birth.The body weight changes of 3 week'progeny mice were recorded.The colonization of LGG bacteria in offspring mice was detected at 2nd and 3rd weeks.The mRNA of intestinal proinflammatory cytokines and tight junction molecules were evaluated by real-time PCR method.HE,immunohistochemistry,immunofluorescence staining and enzyme-linked immunosorbent assay were used to evaluate the intestinal barrier of 3-week old off spring mice.Results Compared with the control group,the progeny mice of the experiment group showed no significant difference in body weight at the first week,and the body weight increased at the second week and the third week [2ndweek:(3.790±0.240) g vs.(4.326±0.140) g,t=3.707,P=0.006;3rd week:(7.295±0.326) g vs.(8.040±0.370) g,t=3.130,P=0.011].LGG colonization can be detected only in the feces of progeny mice in the experiment group.Intestinal colonization can promote the growth of small intestine villi and colon crypt depth [jejunum:(320.000±22.514) μm vs.(265.100±15.611) μm,t=8.258,P＜0.001;ileum:(150.500±13.099) μm vs.(111.000±11.308) μm,t=9.958,P＜0.001;colon:(295.000±15.209) μm vs.(233.100±6.678) μm,t=9.129,P＜0.001].Compared with the control group,the number of goblet cells in the colonic crypt of the experiment group increased (11.62 ± 0.780 vs.35.24 ±1.370,t=15.000,P＜0.001),and the relative mRNA expression levels of pro-inflammatory factors as IFN-γ (1.280±0.232 vs.0.512±0.206,t=4.970,P=0.001),IL-6 (1.364±0.271 vs.0.941±0.215,t=2.452,P=0.040),IL-10 (1.341±0.320vs.0.744±0.294,t=2.762,P=0.025)andTNF-α (3.702±0.150 vs.2.581±0.500,t=2.553,P=0.034) in the experiment group decreased;the expression levels of the intimate tight junction molecules (Claudin3) (1.283±0.152 vs.1.881±0.172,t=4.932,P=0.001) and the atresia protein molecule (Occludin) (1.164±0.342 vs.0.812±0.224,t=3.67,P=0.016) significantly increased.Conclusion Early life LGG colonization protects the intestinal barrier by inhibiting lowgrade intestinal inflammation.This study will lay the experimental foundation for the supplementation of probiotics in early life so as to prevent intestinal diseases.

OBJECTIVE: To investigate the inhibitory effects of dihydromyricetin on the proliferation and migration of gastric cancer BGC-823 cells and explore the molecular mechanisms. METHODS: BGC-823 cells in routine culture were treated with different concentrations of dihydromyricetin (0, 40, 60, 80, 100, and 120 μg/mL) for 24 h, and the changes in cell viability were detected using CCK-8 assay; colony forming assay and Transwell assay were performed to assess the changes in colonyforming and migration abilities of the cells, respectively. The levels of MMP-2 and MMP-9 in the treated cells were determined using ELISA, and Western blotting was used to detect the expressions of E-cadherin, N-cadherin, cyclin D1, cyclin E1, HSP70 and HMGB1 and the phosphorylation levels of Akt and Stat3. RESULTS: CCK-8 assay showed that dihydromyricetin treatment dose-dependently inhibited the viability of BGC-823 cells ( CONCLUSIONS Dihydromyricetin inhibits the proliferation and migration of BGC-823 cells through suppressing the activation of Akt/stat3 signaling pathways and HMGB1 expression.
Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Flavonols ; HMGB1 Protein/metabolism* ; Humans ; Proto-Oncogene Proteins c-akt/metabolism* ; STAT3 Transcription Factor ; Stomach Neoplasms

Objective:To explore the surgical technique and efficacy of pure retroperitoneoscopic extravesical standardized seeable (P.R.E.S.S.) technique for bladder cuff excision (BCE).Methods:Ninety five patients with UTUC from five domestic centers (30 cases from Changzheng Hospital, 21 cases from Peking University First Hospital, 20 cases from Yuhuangding Hospital, 21 cases from Dalian Medical University affiliated No.2 Hospital and 3 cases from General Hospital of Eastern Theater Command)between August 2017 and December 2020 were retrospectively analyzed. There were 57 males and 38 females with a mean age of (67.7±10.0) years and median tumor size of 3.0 cm. All patients underwent pure retroperitoneoscopic radical nephroureterectomy with a single surgical position and four (36 cases) or five (59 cases) trocar layout according to the surgeon’s prefer habit and experience. The demographics of the two groups were the age of [(66.3±11.2)years vs. (68.6±9.1)years], male/female ratio of (25/11 cases vs. 32/27 cases), body mass index of [(25.0± 3.0)kg/m 2 vs. (24.8±3.4)kg/m 2], tumor maximum diameter of [2.8(1.6, 3.5)cm vs. 3.0(2.0, 4.0)cm], left/right side tumor of(19/17 cases vs. 34/25 cases), T 1-2/T 3-4/Tis stage of(25/10/4 cases vs. 49/10/0 cases), and multifocal tumors of(3 cases vs. 2 cases), and the difference was not statistically significant( P>0.05). On the other hand, the differences of hydronephrosis of the operated kidney(13 cases vs. 39 cases, P=0.004), and tumor location (in renal pelvis or calyx or upper/middle/lower ureter being 23/9/4 cases vs. 35/4/20 cases, P=0.005), were statistically significant. The umbilical artery cord was used as anatomical landmark in the process of P. R.E.S.S. bladder cuff excision. The pelvic floor and extraperitoneal space around the ureter were expanded, the bladder wall was opened to form pneumovesicum, and a sufficient bladder cuff resection and exact bladder cuff closure was performed. Perioperative outcomes and follow-up data were analyzed, and the clinical outcomes between the four and five trocars were compared to evaluate the impact of trocar layout on the surgical outcomes. Results:There were 91(95.8%) cases successfully undergoing P. R.E.S.S. BCE technique, with one case converted to open BCE due to bleeding and three cases converted to distal ureter Hem-o-lok clipping because of poor exposure. Median operative time was 180(125, 230)min, and estimated blood loss was 100(50, 100)ml. The overall complication rate was 10.5%(10/95), including 2 cases(2.1%) of intraoperative bleeding, with 1 case treated by transfusion (400 ml), the other case converted to open surgery without transfusion. There were 8 cases of postoperative complications(8.4%), including 7 cases of Clavien-Dindo grade Ⅱ(3 cases of secondary hemorrhage, one case for each of drug allergy, acute renal insufficiency, blood creatinine increased to 490 μmol/L, or lung infection with lymphatic leakage), 1 case of grade Ⅲa(intestinal obstruction treated with insertion of the intestinal obstruction catheter under local anesthesia), and all these patients were discharged smoothly. The difference between the four and five trocars was not statistically significant in the following variables, including the rate of surgical conversion(8.3% vs. 1.7%), estimated intraoperative blood loss(100 ml vs. 60 ml), ratio of intraoperative lymph node dissection (25.0% vs.20.3%), P. R.E.S.S. bladder cuff excision success rate(91.7% vs.98.3%), the incidence of intraoperative and postoperative complications (13.8% vs.8.5%), pT 1-2/pT 3-4/pTis stage(22/11/3 cases vs.37/19/3 cases) and the proportion of recurrence or metastasis(8.3% vs.3.4%)(all P>0.05). However, the differences in the operation time(190 min vs.170 min, P=0.011)and postoperative hospital stay(5 d vs.6 d, P=0.005) were statistically significant. Conclusions:P. R.E.S.S. bladder cuff resection technique is safe and feasible during the procedure of pure retroperitoneoscopic radical nephroureterectomy by a single surgical position and facilitates seeable adequate bladder cuff excision by establishing an enlarged pelvic lateral extraperitoneal space and pneumovesicum. Five-trocar technique is more suitable for patients with lower ureteral tumors but may be associated with a longer postoperative hospital stay compared with the four-trocar technique.

The aim of this study was to develop a blocking enzyme-linked immunosorbent assay (bELISA) based on a biotinylated nanobody target the S1 protein of porcine epidemic diarrhea virus (PEDV) for detecting the anti-PEDV antibodies and evaluating the immune effect of the vaccine. The gene encoding the single-domain antibody sdAb3 target the PEDV S1 protein was amplified and the Avitag sequence was fused at its 3'-end. The PCR product was cloned into the expression vector pET-21b for expression and purification of the sdAb3-Avitag protein. The purified sdAb3-Avitag fusion protein was biotinylated and its activity was determined. Using the recombinant S1 protein as a coating antigen, a bELISA was established and optimized. Serum samples were tested in parallel by the bELISA and a commercial kit. The recombinant vector pET21b-sdAb3-Avitag was constructed to express the tagged sdAb3. After induction for expression, the biotin-labeled sdAb3 (sdAb3-Biotin) with high purity and good activity was obtained. For the optimized bELISA, the coating concentration of the S1 protein was 200 ng/well, the serum dilution was 1:2 and incubated for 2 h, the dilution ratio of the biotinylated sdAb3 was 1:8 000 and incubated for 30 min, the dilution of the enzyme-labeled antibody was 1:5 000 and incubated for 30 min. The bELISA had no cross reaction with the sera of major porcine viruses including transmissible gastroenteritis virus, porcine reproductive and respiratory syndrome virus and showed good specificity and reproducibility. For a total of 54 porcine serum samples tested, the overall compliance rate of the bELISA with a commercial kit was 92.56%. This study developed a rapid and reliable bELISA method, which can be used for serosurveillance and vaccine evaluation for PEDV.
Animals ; Antibodies, Viral ; Coronavirus Infections/veterinary* ; Enzyme-Linked Immunosorbent Assay ; Porcine epidemic diarrhea virus/genetics* ; Reproducibility of Results ; Sensitivity and Specificity ; Single-Domain Antibodies ; Swine ; Swine Diseases

In mammalian cells, long noncoding RNAs (lncRNAs) form complexes with proteins to execute various biological functions such as gene transcription, RNA processing and other signaling activities. However, methods to track endogenous lncRNA dynamics in live cells and screen for lncRNA interacting proteins are limited. Here, we report the development of CERTIS (CRISPR-mediated Endogenous lncRNA Tracking and Immunoprecipitation System) to visualize and isolate endogenous lncRNA, by precisely inserting a 24-repeat MS2 tag into the distal end of lncRNA locus through the CRISPR/Cas9 technology. In this study, we show that CERTIS effectively labeled the paraspeckle lncRNA NEAT1 without disturbing its physiological properties and could monitor the endogenous expression variation of NEAT1. In addition, CERTIS displayed superior performance on both short- and long-term tracking of NEAT1 dynamics in live cells. We found that NEAT1 and paraspeckles were sensitive to topoisomerase I specific inhibitors. Moreover, RNA Immunoprecipitation (RIP) of the MS2-tagged NEAT1 lncRNA successfully revealed several new protein components of paraspeckle. Our results support CERTIS as a tool suitable to track both spatial and temporal lncRNA regulation in live cells as well as study the lncRNA-protein interactomes.