Endoscopic construction of an antireflux mucosal barrier for the treatment of GERD: a pilot study (with video).

BACKGROUND AND AIMS: Based on the porcine natural antireflux mechanism, we developed a novel endoscopic procedure to build an antireflux mucosal flap to block acid reflux and treat GERD. METHODS: The antireflux mucosal valvuloplasty (ARMV) procedure is performed by releasing and reconstructing three-fourths of the circumference of cardiac mucosa at the lesser curvature side into a double-layer mucosal flap. The mucosal flap works together with cardiac scarring to block reflux. We retrospectively reviewed 30 patients who underwent ARMV from 2019 to 2021. Subjective and objective data evaluating GERD were collected before and after ARMV. RESULTS: All 30 ARMV procedures were performed successfully, with a mean operation time of 72.6 ± 20.3 minutes. One patient had postoperative bleeding that required endoscopic hemostasis. The mean follow-up time was 28.9 ± 13.9 months. Twenty-five of 30 patients (83.3%) and 23 of 26 patients (88.5%) reported discontinuation or reduction in proton pump inhibitor therapy 3 months and 1 year after ARMV, respectively. GERD questionnaire and GERD Health-Related Quality of Life questionnaire scores improved significantly from 14.0 ± 2.6 and 48.7 ± 15.0, respectively, before ARMV to 7.7 ± 2.5 and 10.2 ± 5.9, respectively, 12 months after ARMV (P < .0001 in both comparisons). Eleven patients received 24-hour esophageal pH monitoring before and after ARMV. The mean acid exposure time and DeMeester score dropped from 56.9% ± 23.7% and 167.1 ± 80.1, respectively, before ARMV to 5.5% ± 3.0% and 18.6 ± 11.9, respectively, after ARMV (P < .0001 in both comparisons). CONCLUSIONS: This pilot study showed that ARMV is a safe, feasible, and effective procedure for GERD patients. Further prospective and comparative trials are needed to confirm its role among endoscopic antireflux therapies.

A Neural Network Based Approach to Inverse Kinematics Problem for General Six-Axis Robots.

Inverse kinematics problems (IKP) are ubiquitous in robotics for improved robot control in widespread applications. However, the high non-linearity, complexity, and equation coupling of a general six-axis robotic manipulator pose substantial challenges in solving the IKP precisely and efficiently. To address this issue, we propose a novel approach based on neural network (NN) with numerical error minimization in this paper. Within our framework, the complexity of IKP is first simplified by a strategy called joint space segmentation, with respective training data generated by forward kinematics. Afterwards, a set of multilayer perception networks (MLP) are established to learn from the foregoing data in order to fit the goal function piecewise. To reduce the computational cost of the inference process, a set of classification models is trained to determine the appropriate forgoing MLPs for predictions given a specific input. After the initial solution is sought, being improved with a prediction error minimized, the refined solution is finally achieved. The proposed methodology is validated via simulations on Xarm6-a general 6 degrees of freedom manipulator. Results further verify the feasibility of NN for IKP in general cases, even with a high-precision requirement. The proposed algorithm has showcased enhanced efficiency and accuracy compared to NN-based approaches reported in the literature.

Pelvic peritoneum closure reduces postoperative complications of laparoscopic abdominoperineal resection: 6-year experience in single center.

BACKGROUND: To investigate feasibility of laparoscopic abdominoperineal resection with pelvic peritoneum closure (LAPR-PPC) for lower rectal cancer. METHODS: LAPR-PPC has been used for lower rectal cancer in our institution since 2014. In this study, we retrospectively analyzed the data from 86 patients who underwent LAPR-PPC and compared with the data from 96 patients who underwent laparoscopic APR without PPC (LAPR) from January 2013 to December 2018. RESULTS: The rate of perineal surgical site infection (SSI) (18.75% (18/96) vs. 5.81% (5/86), p < 0.01), delayed (> 4 weeks) perineal healing (12.50% (12/96) vs. 3.49% (3/86), p = 0.027), ileus (7.29% (7/96) vs 1.16% (1/86), p = 0.044), and postoperative perineal hernia (PPH, 5.21% (5/96) vs. 0% (0/86), p = 0.032) were significantly lower in LAPR-PPC group than LAPR group. The patients in LAPR-PPC group had shorter hospitalization time (21.32 ± 11.95 days vs. 13.93 ± 11.51 days, p < 0.01). CONCLUSIONS: PPC procedure enabled the reduction in perineal wound complications, ileus, PPH, and consequently shortened hospitalization time. LAPR-PPC is beneficial for the patients with lower rectal cancer.

Alteration of the gut microbiome and immune factors of grass carp infected with Aeromonas veronii and screening of an antagonistic bacterial strain (Streptomyces flavotricini).

Aeromonas veronii is a widely distributed novel pathogen that can affect humans and animals, it can cause sepsis in fish with high mortality and serious economic losses to aquaculture. In the study, the gut microbiome of the infected and uninfected grass carp with Aeromonas veronii were analyzed probiotics and pathogenic bacteria by the Miseq high-throughput sequencing, the results showed that the infected fish were significantly higher in Proteobacteria, Firmicutes, Fusobacteria, and the immune factors in liver and kidney were up-regulated by qRT-PCR. In order to effectively inhibit the pathogen, we screened an actinomycete strain and had good antibacterial effect on Aeromonas veronii. The new antagonistic bacteria was named as Streptomyces flavotricini X101, the whole genome sequencing revealed that the metabolic process was most active. After grass carp was inoculated with the minimum inhibitory concentration of 900 µg/mL of the strain's fermentation supernatant, then Aeromonas veronii was injected, we found that the pathological symptoms such as body surface, anus and abdominal congestion were alleviated by H&E staining. Cellular experiments showed that it wasn't toxic to liver cells of grass carp. Overall, this is the first study of changes in intestinal flora, phenotype, and immune factors in grass crap infected with Aeromonas veronii, it had important theoretical significance and application value for immunization and prevention.

Extracellular matrix dynamics in cell migration, invasion and tissue morphogenesis.

This review describes how direct visualization of the dynamic interactions of cells with different extracellular matrix microenvironments can provide novel insights into complex biological processes. Recent studies have moved characterization of cell migration and invasion from classical 2D culture systems into 1D and 3D model systems, revealing multiple differences in mechanisms of cell adhesion, migration and signalling-even though cells in 3D can still display prominent focal adhesions. Myosin II restrains cell migration speed in 2D culture but is often essential for effective 3D migration. 3D cell migration modes can switch between lamellipodial, lobopodial and/or amoeboid depending on the local matrix environment. For example, "nuclear piston" migration can be switched off by local proteolysis, and proteolytic invadopodia can be induced by a high density of fibrillar matrix. Particularly, complex remodelling of both extracellular matrix and tissues occurs during morphogenesis. Extracellular matrix supports self-assembly of embryonic tissues, but it must also be locally actively remodelled. For example, surprisingly focal remodelling of the basement membrane occurs during branching morphogenesis-numerous tiny perforations generated by proteolysis and actomyosin contractility produce a microscopically porous, flexible basement membrane meshwork for tissue expansion. Cells extend highly active blebs or protrusions towards the surrounding mesenchyme through these perforations. Concurrently, the entire basement membrane undergoes translocation in a direction opposite to bud expansion. Underlying this slowly moving 2D basement membrane translocation are highly dynamic individual cell movements. We conclude this review by describing a variety of exciting research opportunities for discovering novel insights into cell-matrix interactions.

Differential expression of ST6GAL1 in the tumor progression of colorectal cancer.

Elevated expression of ß-galactoside α2,6-sialyltranferase 1 (ST6GAL1) has been observed in colorectal cancer (CRC) and demonstrated to be important for its tumorigenesis. Here, we found that ST6GAL1 expression was significantly higher in non-metastatic tumors (stage I and II) than that in metastatic tumors (stage III and IV) using 62 pair-matched tumor/normal tissues. To elucidate the molecular mechanisms of how ST6GAL1 affected the CRC progression, we performed a global identification of the substrates of ST6GAL1 in the colon adenocarcinoma cell line SW480. A total of 318 membrane proteins were identified differentially affected by ST6GAL1 overexpression using metabolic labeling and proteomic analysis. Subsequent bioinformatic analysis revealed a list of potential substrates that might mediate the different functions of ST6GAL1 in CRC including cell movement, cell death and survival. Taken together, these results indicate a dynamic change in the expression of ST6GAL1 during the CRC progression and provide a list of sialylated proteins potentially relevant to the different functions of ST6GAL1 in CRC.

Targeting the Metastasis Suppressor, N-Myc Downstream Regulated Gene-1, with Novel Di-2-Pyridylketone Thiosemicarbazones: Suppression of Tumor Cell Migration and Cell-Collagen Adhesion by Inhibiting Focal Adhesion Kinase/Paxillin Signaling.

Metastasis is a complex process that is regulated by multiple signaling pathways, with the focal adhesion kinase (FAK)/paxillin pathway playing a major role in the formation of focal adhesions and cell motility. N-myc downstream regulated gene-1 (NDRG1) is a potent metastasis suppressor in many solid tumor types, including prostate and colon cancer. Considering the antimetastatic effect of NDRG1 and the crucial involvement of the FAK/paxillin pathway in cellular migration and cell-matrix adhesion, we assessed the effects of NDRG1 on this important oncogenic pathway. In the present study, NDRG1 overexpression and silencing models of HT29 colon cancer and DU145 prostate cancer cells were used to examine the activation of FAK/paxillin signaling and the formation of focal adhesions. The expression of NDRG1 resulted in a marked and significant decrease in the activating phosphorylation of FAK and paxillin, whereas silencing of NDRG1 resulted in an opposite effect. The expression of NDRG1 also inhibited the formation of focal adhesions as well as cell migration and cell-collagen adhesion. Incubation of cells with novel thiosemicarbazones, namely di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone and di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone, that upregulate NDRG1 also resulted in decreased phosphorylation of FAK and paxillin. The ability of these thiosemicarbazones to inhibit cell migration and metastasis could be mediated, at least in part, through the FAK/paxillin pathway.

Facilitating retroflexed endoscopic full-thickness resection through loop-mediated or rope-mediated countertraction (with videos).

BACKGROUND AND AIMS: Endoscopic full-thickness resection (EFTR) is difficult to perform in a retroflexed fashion in the gastric fundus and lesser curvature. Here we describe two simple methods to provide countertraction and thereby facilitate dissection. METHODS: In this retrospective cohort study, 62 patients diagnosed as having gastric submucosal tumors in the fundus or in the lesser curvature received EFTR with or without countertraction methods. For the clip-with-thread method, a clip tied with surgical suture was anchored on the distal edge of the tumor to provide countertraction; for the loop-assisted method, a snare placed on the transparent cap beforehand was adopted to grasp the tumor to provide countertraction. RESULTS: Mean operative time was significantly reduced in the thread-with-clip group and loop-assisted group (45 minutes, 40 minutes, respectively) compared with the time needed in the traditional EFTR group (85 minutes). Intraoperative pneumoperitoneum occurred regularly among the 3 groups because of iatrogenic perforation, but fewer patients in the thread-with-clip group and loop-assisted group (23%, 18%, respectively) needed abdominal puncture to relieve free air and stabilize life signs compared with patients in the traditional EFTR group (63%). A reduced occurrence of high fever after surgery may contribute as another advantage from accelerated dissection. Both techniques did not jeopardize oncologic safety during short-term follow-up. CONCLUSIONS: Both the thread-with-clip method and loop-assisted method provide effective countertraction and offer faster and safer gastric EFTR in difficult anatomic locations.

Transcardiac tunneling technique for endoscopic submucosal dissection of gastric fundus tumors arising from the muscularis propria.

The promising endoscopic resection techniques for upper gastrointestinal submucosal tumors (SMTs) are challenged when performed in the gastric fundus. Here, we report on the development of a transcardiac endoscopic tunneling technique (TCTT) for the resection of tumors in this area. A total of 18 patients with gastric fundus SMTs arising from the muscularis propria on endoscopic ultrasound underwent TCTT. The procedure involved the excavation of a submucosal tunnel from the esophagus, through the cardia, to the gastric SMT for resection. The tunnel was closed by clips after retrieval of the tumor. The mean tumor size was 2.1 cm (range 0.8 - 5.0 cm). The mean procedure time was 75.1 minutes (range 40 - 100 minutes). Complete resection was achieved in all cases. Iatrogenic perforation occurred in one case. This and one other patient developed mild pneumoperitoneum on the day after the procedure; symptoms resolved under conservative management. No patient developed gastrointestinal leakage, delayed bleeding, or secondary infection. Therefore, in this pilot study, TCTT provided a definitive histological diagnosis as well as a feasible, safe, and easy therapeutic approach for gastric fundus SMTs arising from muscularis propria in the circular area within 8 cm below the cardia.

Endoscopic management of upper gastrointestinal submucosal tumors arising from muscularis propria.

Small (<2 cm) or incidentally found gastrointestinal submucosal tumors pose challenges to gastroenterologists due to the potential malignancy of tumors and current suboptimal diagnostic methods. On the basis of the development of endoscopic submucosal dissection, recent technical advances, including endoscopic submucosal excavation, submucosal tunneling and endoscopic resection, and endoscopic full-thickness resection, have been able to resect submucosal tumors in the muscularis propria. These techniques provide definitive histologic diagnosis as well as a minimally invasive therapeutic approach to such tumors. Current studies, despite with promising results, are limited to small, retrospective studies, focusing on introducing new methods and varied in technical details. This review provides a thorough discussion of technical details of each endoscopic method, and compares different methods in terms of tumor sites, to ensure safety and optimize treatment outcomes.

Endoscopic resection of submucosal tumors in muscularis propria: the choice between direct excavation and tunneling resection.

BACKGROUND: Endoscopic submucosal excavation (ESE) and submucosal tunneling endoscopic resection (STER) are nowadays both adopted by endoscopists to resect upper gastrointestinal submucosal tumors (SMTs) arising from muscularis propria. This study aimed to compare the efficacy and safety of these two techniques. METHODS: Seventy-seven patients were included in this retrospective study, among them, 35 received ESE and 42 received STER. Main outcome measurements included complete resection rate, perforation rate, adverse events, and tumor recurrence. Subgroup analysis based on tumor size was also performed. RESULTS: In general, ESE and STER had similar complete resection rate (94.7 vs. 97.7%), perforation rate (10.5 vs. 13.3%), and incidence of postoperative air leakage symptoms (13.2 vs. 2.2%). No tumor recurrence occurred in any group. Subgroup analysis revealed that for tumors <10 mm, both techniques achieved satisfactory therapeutic effects (100 % complete resection, no adverse events); for tumors >10 mm, perforation rate increased in both ESE and STER (16 vs. 18.2%), yet incidence of air leakage symptoms was significantly low in STER (3%) compared to it in ESE (20%). CONCLUSIONS: For SMTs <10 mm, both ESE and STER have satisfactory therapeutic results, it is not necessary to pursue a difficult procedure when a simple method is available. For SMTs >10 mm, STER is a preferable choice in terms of preventing air leakage symptoms, especially, when perforation is likely to happen.

CRISPR/Cas9-Mediated Gene Knockout in Cells and Tissues Using Lentivirus.

CRISPR-Cas9 has become a powerful and popular gene editing tool. However, successful application of this tool in the lab can still be quite daunting to many newcomers to molecular biology, mostly because it is a relatively lengthy process involving multiple steps with variations of each step. Here, we provide a reliable, stepwise, and newcomer-friendly protocol to knock out a target gene in wild-type human fibroblasts. This protocol involves sgRNA design using CRISPOR, construction of an "all-in-one" vector expressing both sgRNA and Cas9 using Golden Gate cloning, streamlined production of high-titer lentiviruses in 1 week after molecular cloning, and transduction of cells to generate a knockout cell pool. We further introduce a protocol for lentiviral transduction of ex vivo mouse embryonic salivary epithelial explants. In summary, our protocol is useful for new researchers to apply CRISPR-Cas9 to generate stable gene knockout cells and tissue explants using lentivirus. Published 2023. This article is a U.S. Government work and is in the public domain in the USA. Basic Protocol 1: sgRNA design Basic Protocol 2: Cloning sgRNA in plasmid vector containing Cas9 encoding sequence using golden gate cloning Basic Protocol 3: Lentivirus packaging Basic Protocol 4: Lentivirus transduction of cells Basic Protocol 5: Lentivirus transduction of salivary gland epithelial buds.

Benefits of precise endoscopic incision on post-dilation mucosal scars to treat refractory esophageal stricture after endoscopic submucosal dissection.

Endoscopic dilation (ED) is the mainstream treatment for esophageal stricture after endoscopic submucosal dissection (ESD). However, some complex esophageal strictures do not respond well to dilation. Endoscopic radial incision (ERI) has proved to be effective in treating anastomotic strictures, but it is rarely used to treat post-ESD esophageal strictures due to technical difficulties and risks, not to mention the optimal method and timing to perform ERI. Here, we developed an integrated procedure in which ED was performed first, followed by ERI on the stiff scars that remained intact after dilation. The ED + ERI procedure resulted in complete, uniform expansion of the esophageal lumen. Between 2019 and 2022, 5 post-ESD patients who received a median number of 11 sessions of ED (range, 4-28) of ED over a period of 322 days (range, 246-584) but still had moderate to severe dysphagia were admitted. 2 or 3 sessions of ED + ERI were performed for each patient interspersed with ED. After a median number of 4 treatments (range, 2-9), all patients were symptom-free or had minimal symptoms. No serious complications occurred in any patients who underwent ED + ERI. Therefore, ED + ERI is safe, feasible, and may serve as a useful therapeutic method for refractory esophageal stricture after ESD.

Statistical Insight into China's Indigenous Diagnosis-Related-Group System Evolution.

The use of Diagnosis-Related Groups (DRG) is a prevalent payment system employed to control hospitalization costs and improve medical efficiency. China has developed an indigenized DRG payment system including Single Disease Payment (SDP), DRGs, and Big Data Diagnosis-Intervention Packet (DIP). In this study, we took cholecystitis as an example, drawing on both primary and secondary data to verify the effectiveness of China's indigenized DRG system and to introduce the evolution of DRGs in China. Primary data were gathered from Qilu Hospital in 2019-2021. Secondary data were collected from published literature from 2004-2016. Only studies with both pre-SDP/DRG and post-SDP/DRG groups were included. Among the studies included, 92.9% (13/14) reported a significant reduction in hospitalization costs after the implementation of SDP while other studies identified length of stay (LOS) and age as the most significant influential factors in SDP. Furthermore, we elaborated the efficiency of DRGs using data from 2738 inpatients in Qilu hospital. Moreover, 60% (6/10) of the studies from the databases also showed the efficiency of DRGs in different regions. SDP is efficient in saving hospitalization costs, but its implementation is limited. DRGs have a broader scope of application, but their effectiveness remains to be validated. DIP is a brand new concept in China, and more data are needed to assess its efficiency.

HOXA11-AS facilitates the proliferation, cell cycle process and migration of keloid fibroblasts through sponging miR-188-5p to regulate VEGFA.

BACKGROUND: Abnormal expression of long non-coding RNA (lncRNA) has been proved to be related to the formation of keloid. Homeobox A11 antisense (HOXA11-AS) has been found to be a significantly upregulated lncRNA in keloid tissues. OBJECTIVE: To explore the mechanism of HOXA11-AS regulates keloid formation. METHODS: Primary fibroblasts were isolated from keloid tissues and normal skin tissues. The expression of HOXA11-AS, microRNA (miR)-188-5p and vascular endothelial growth factor A (VEGFA) was determined using quantitative real-time PCR (qRT-PCR). Cell counting kit 8 (CCK8) assay, EdU staining, flow cytometry and wound healing assay were performed to assess the proliferation, cell cycle process, apoptosis and migration of keloid fibroblasts. Importantly, some marker protein levels and VEGFA protein level were examined by western blot (WB) analysis. The interaction between miR-188-5p and HOXA11-AS or VEGFA was confirmed using dual-luciferase reporter assay, RNA pull-down assay and RIP assay. Animal experiments were performed to further confirm the role of HOXA11-AS in keloid growth. RESULTS: HOXA11-AS was markedly upregulated in keloid tissues and fibroblasts. Knockdown of HOXA11-AS repressed the proliferation, cell cycle process, migration and promoted the apoptosis of keloid fibroblasts. Further analysis suggested that HOXA11-AS could sponge miR-188-5p to positively regulate VEGFA. The inhibition of HOXA11-AS silencing on the biological functions of keloid fibroblasts could be reversed by miR-188-5p inhibitor. In addition, VEGFA overexpression also abolished the suppressive effect of miR-188-5p on the biological functions of keloid fibroblasts. Interferences of HOXA11-AS suppressed keloid growth in vivo. CONCLUSION: HOXA11-AS might regulate the miR-188-5p/VEGFA axis to promote keloid formation.

Cell-3D matrix interactions: recent advances and opportunities.

Tissues consist of cells and their surrounding extracellular matrix (ECM). Cell-ECM interactions play crucial roles in embryonic development, differentiation, tissue remodeling, and diseases including fibrosis and cancer. Recent research advances in characterizing cell-matrix interactions include detailed descriptions of hundreds of ECM and associated molecules, their complex intermolecular interactions in development and disease, identification of distinctive modes of cell migration in different 3D ECMs, and new insights into mechanisms of organ formation. Exploring the roles of the physical features of different ECM microenvironments and the bidirectional regulation of cell signaling and matrix organization emphasize the dynamic nature of these interactions, which can include feedback loops that exacerbate disease. Understanding mechanisms of cell-matrix interactions can potentially lead to targeted therapeutic interventions.