SIRT2 regulates high mobility group protein B1 nucleoplasmic shuttle and degradation via deacetylation in microglia.

High mobility group protein B1 (HMGB1) acts as a pathogenic inflammatory response to mediate ranges of conditions such as epilepsy, septic shock, ischemia, traumatic brain injury, Parkinson's disease, Alzheimer's disease and mass spectrometry. HMGB1 promotes inflammation during sterile and infectious damage and plays a crucial role in disease development. Mobilization from the nucleus to the cytoplasm is the first important step in the release of HMGB1 from activated immune cells. Here, we demonstrated that Sirtuin 2 (SIRT2) physically interacts with and deacetylates HMGB1 at 43 lysine residue at nuclear localization signal locations, strengthening its interaction with HMGB1 and causing HMGB1 to be localized in the cytoplasm. These discoveries are the first to shed light on the SIRT2 nucleoplasmic shuttle, which influences HMGB1 and its degradation, hence revealing novel therapeutic targets and avenues for neuroinflammation treatment.

Gamma-glutamyl transferase secreted by Helicobacter pylori promotes the development of gastric cancer by affecting the energy metabolism and histone methylation status of gastric epithelial cells.

BACKGROUND: Helicobacter pylori (H. pylori) infection is critical in the development and occurrence of gastric cancer. H. pylori secretes gamma-glutamyl transferase (GGT), which affects energy metabolism and histone methylation in mesenchymal stem cells. However, its effect on human gastric epithelial cells remains unclear. This study aimed to investigate the effects of GGT on energy metabolism and histone methylation in gastric epithelial cells and determine its role in the development and progression of H. pylori-induced gastric cancer. METHODS: A GGT knockout H. pylori strain and mouse gastric cancer model were constructed, and alpha-ketoglutarate (α-KG) was added. The underlying mechanism was investigated using proteomics, immunohistochemistry, Western blotting, and other experimental assays. RESULTS: H. pylori can colonize the host's stomach and destroy the gastric epithelium. GGT secreted by H. pylori decreased the concentration of glutamine in the stomach and increased H3K9me3 and H3K27me3 expression, which promoted the proliferation and migration of gastric epithelial cells. Additionally, α-KG reversed this effect. GGT increased the tumorigenic ability of nude mice. GGT, secreted by H. pylori, promoted the expression of ribosomal protein L15 (RPL15), while GGT knockout and supplementation with α-KG and trimethylation inhibitors reduced RPL15 expression and Wnt signaling pathway expression. CONCLUSIONS: H. pylori secreted GGT decreased the expression of glutamine and α-KG in gastric epithelial cells, increased the expression of histones H3K9me3 and H3K27me3, and activated the Wnt signaling pathway through RPL15 expression, ultimately changing the biological characteristics of the gastric epithelium and promoting the occurrence of gastric cancer. Altered energy metabolism and histone hypermethylation are important factors involved in this process.

The Efficacy and Safety of Regimens for Helicobacter pylori Eradication Treatment in China: A Systemic Review and Network Meta-Analysis.

BACKGROUND: With Helicobacter pylori's increasing antibiotic resistance, evidence of more effective treatments is lacking in China, where H. pylori prevalence is nearly 50%. Thus, we performed a network meta-analysis to compare therapeutic regimens. METHODS: Data extracted from eligible randomized controlled trials from January 2000 to September 2021 were entered into a Bayesian hierarchical random-effects model to evaluate the efficacy and safety of H. pylori eradication regimens. RESULTS: This study included 101 trials involving 21,745 patients. Vonoprazan-bismuth-containing quadruple therapy (VBQT) ranked the highest [surfaces under cumulative ranking curve (SUCRA), 83.64%], followed by high-dose amoxicillin dual therapy (HDDT) [SUCRA, 79.70%, odds ratio (OR)=1.31, 95% credible interval (CrI) (0.36, 4.72)] and proton pump inhibitor-based bismuth-containing quadruple therapy (BQT) [SUCRA, 63.59%, OR=1.59, 95% CrI (0.48, 5.24)]. HDDT [OR=2.47, 95% CrI (1.51, 4.06)], BQT [OR=2.04, 95% CrI (1.69, 2.47)], concomitant quadruple nonbismuth therapy (CT) [OR=1.93, 95% CrI (1.19, 3.15)], and sequential therapy (ST) [OR=1.86, 95% CrI (1.50, 2.32)] had higher eradication rates than standard triple therapy (TT). ST (SUCRA, 82.52%) and VBQT (SUCRA, 83.89%) had the highest eradication rate before and after 2010 in the effectiveness ranking, respectively. Furthermore, the H. pylori eradication rate of patients receiving 14-day BQT treatment was higher than that of 10-day BQT regimen [OR=2.55, 95% CI (1.84, 3.53)] and 7-day BQT regimen [OR=3.64, 95% CI (2.64, 5.01)]. CONCLUSIONS: The TT regimen was not an optimal choice in China for H. pylori eradication; VBQT, HDDT, and BQT showed better efficacy. After 2010, there is a trend toward significance that VBQT provided a higher H. pylori eradication rate in China, but with only 1 randomized controlled trial. Thus, more supportive real-world data are needed to confirm its effectiveness.

Green and efficient recycling method for spent Ni-Co-Mn lithium batteries utilizing multifunctional deep eutectic solvents.

Given the growing volume of discarded lithium-ion batteries (LIBs), the extraction and recovery of valuable metals through environmentally-friendly solvent processes have become crucial, but they remain challenging tasks. Deep eutectic solvent (DES), an innovative and green solvents, have demonstrated significant promise in the extraction of valued metal elements from spent LIBs. This work employed a multifunctional DES based on natural molecules dimethyl-beta-propiothetin (DMPT) and ethylene glycol (EG) for the efficient leaching of transition metal ions. Under the reduction effect of EG and the action of carboxyl groups and chloride ions in DMPT, the leaching rate of Li, Ni, Co, and Mn can reach 99.59%, 99.28%, 99.04%, and 99.45%, respectively. Furthermore, DFT calculations were employed to explore the microstructure of DES and its interactions with metal ions. The main active site in the DES molecule is near the chloride ion, and DES binds most strongly to Mn, followed by Co, and weakest to Ni. This work avoids the use of volatile acids and demonstrates great potential in extracting valuable metals, providing a sustainable and environment-friendly alternative for the efficient recycling of waste LIBs.

The E3 ubiquitin ligase TRIM31 attenuates NLRP3 inflammasome activation in Helicobacter pylori-associated gastritis by regulating ROS and autophagy.

BACKGROUND: The NLRP3 inflammasome activation is the molecular basis of Helicobacter pylori (Hp)-associated gastritis. Tripartite motif (TRIM) 31 is involved in diverse pathological events. However, whether TRIM31 plays a role in the activation of NLRP3 inflammasome in Hp infection is not clarified. METHODS: A mouse model of chronic Hp infection was established, and the gastric tissues were subjected to the polymerase chain reaction, western blotting, histopathological analysis, and RNA sequencing. The mitochondrial membrane potential and ROS in the human gastric epithelium GES-1 cells with or without Hp infection were measured by flow cytometry. GES-1 cells with or without TRIM31 knockdown were transfected with mCherry-EGFP-LC3 adenovirus. After rapamycin and bafilomycin A1 stimulation, autophagy flux in the above primed GES-1 cells was assessed by laser confocal microscope. Lysosomal acidification and expression levels of cathepsin B and cathepsin D in GES-1 cells with Hp infection were measured. RESULTS: NLRP3 inflammasome was activated in the gastric tissues of mice with chronic Hp infection in vivo and the GES-1 cells with Hp infection in vitro. TRIM31 was downregulated in Hp infection. TRIM31 negatively regulated the NLRP3 inflammasome activation. Enhanced ROS, impaired autophagy flux, and decreased expression of lysosomal cathepsin B and cathepsin D were observed in TRIM31-deficient GES-1 cells with Hp infection. In turn, inhibition of ROS led to the decreased expression of NLRP3 inflammasome. CONCLUSIONS: Together, our data identified that TRIM31 negatively regulated the activation of NLRP3 inflammasome in Hp-associated gastritis by affecting ROS and autophagy of gastric epithelial cells. Video abstract.

L-Fucose promotes enteric nervous system regeneration in type 1 diabetic mice by inhibiting SMAD2 signaling pathway in enteric neural precursor cells.

BACKGROUND: Diabetes can lead to extensive damage to the enteric nervous system (ENS), causing gastrointestinal motility disorders. However, there is currently a lack of effective treatments for diabetes-induced ENS damage. Enteric neural precursor cells (ENPCs) closely regulate the structural and functional integrity of the ENS. L-Fucose, is a dietary sugar that has been showed to effectively ameliorate central nervous system injuries, but its potential for ameliorating ENS damage and the involvement of ENPCs in this process remains uncertain. METHODS: Genetically engineered mice were generated for lineage tracing of ENPCs in vivo. Using diabetic mice in vivo and high glucose-treated primary ENPCs in vitro, the effects of L-Fucose on the injured ENS and ENPCs was evaluated by assessing gastrointestinal motility, ENS structure, and the differentiation of ENPCs. The key signaling pathways in regulating neurogenesis and neural precursor cells properties, transforming growth factor-ß (TGF-ß) and its downstream signaling pathways were further examined to clarify the potential mechanism of L-Fucose on the injured ENS and ENPCs. RESULTS: L-Fucose improved gastrointestinal motility in diabetic mice, including increased defecation frequency (p < 0.05), reduced total gastrointestinal transmission time (p < 0.001) and bead expulsion time (p < 0.05), as well as enhanced spontaneous contractility and electric field stimulation-induced contraction response in isolated colonic muscle strips (p < 0.001). The decrease in the number of neurons and glial cells in the ENS of diabetic mice were reversed by L-Fucose treatment. More importantly, L-Fucose treatment significantly promoted the proportion of ENPCs differentiated into neurons and glial cells both in vitro and in vivo, accompanied by inhibiting SMAD2 phosphorylation. CONCLUSIONS: L-Fucose could promote neurogenesis and gliogenesis derived from ENPCs by inhibiting the SMAD2 signaling, thus facilitating ENS regeneration and gastrointestinal motility recovery in type 1 diabetic mice. Video Abstract.

Microvesicles from bone marrow-derived mesenchymal stem cells promote Helicobacter pylori-associated gastric cancer progression by transferring thrombospondin-2.

BACKGROUND: Our previous study found that bone marrow-derived mesenchymal stem cells (BMSCs) promote Helicobacter pylori (H pylori)-associated gastric cancer (GC) progression by secreting thrombospondin-2 (THBS2). Extracellular vesicles (EVs) are important carriers for intercellular communication, and EVs secreted by BMSCs have been shown to be closely related to tumor development. The aim of this study was to investigate whether BMSC-derived microvesicles (MVs, a main type of EV) play a role in H. pylori-associated GC by transferring THBS2. METHODS: BMSCs and THBS2-deficient BMSCs were treated with or without the supernatant of H. pylori for 12 h at a multiplicity of infection of 50, and their EVs were collected. Then, the effects of BMSC-derived MVs and THBS2-deficient BMSC-derived MVs on the GC cell line MGC-803 were assessed by in vitro proliferation, migration, and invasion assays. In addition, a subcutaneous xenograft tumor model, a nude mouse intraperitoneal metastasis model, and a tail vein injection metastasis model were constructed to evaluate the effects of BMSC-derived MVs and THBS2-deficient BMSC-derived MVs on GC development and metastasis in vivo. RESULTS: BMSC-derived MVs could be readily internalized by MGC-803 cells. BMSC-derived MVs after H. pylori treatment significantly promoted their proliferation, migration and invasion in vitro (all P < 0.05) and promoted tumor development and metastasis in a subcutaneous xenograft tumor model, a nude mouse intraperitoneal metastasis model, and a tail vein injection metastasis model in vivo (all P < 0.05). The protein expression of THBS2 was significantly upregulated after H. pylori treatment in BMSC-derived MVs (P < 0.05). Depletion of the THBS2 gene reduces the tumor-promoting ability of BMSC-MVs in an H. pylori infection microenvironment both in vitro and in vivo. CONCLUSION: Overall, these findings indicate that MVs derived from BMSCs can promote H. pylori-associated GC development and metastasis by delivering the THBS2 protein. Video Abstract.

Artificial intelligence-based diagnosis of abnormalities in small-bowel capsule endoscopy.

BACKGROUND : Further development of deep learning-based artificial intelligence (AI) technology to automatically diagnose multiple abnormalities in small-bowel capsule endoscopy (SBCE) videos is necessary. We aimed to develop an AI model, to compare its diagnostic performance with doctors of different experience levels, and to further evaluate its auxiliary role for doctors in diagnosing multiple abnormalities in SBCE videos. METHODS : The AI model was trained using 280 426 images from 2565 patients, and the diagnostic performance was validated in 240 videos. RESULTS : The sensitivity of the AI model for red spots, inflammation, blood content, vascular lesions, protruding lesions, parasites, diverticulum, and normal variants was 97.8 %, 96.1 %, 96.1 %, 94.7 %, 95.6 %, 100 %, 100 %, and 96.4 %, respectively. The specificity was 86.0 %, 75.3 %, 87.3 %, 77.8 %, 67.7 %, 97.5 %, 91.2 %, and 81.3 %, respectively. The accuracy was 95.0 %, 88.8 %, 89.2 %, 79.2 %, 80.8 %, 97.5 %, 91.3 %, and 93.3 %, respectively. For junior doctors, the assistance of the AI model increased the overall accuracy from 85.5 % to 97.9 % (P  < 0.001, Bonferroni corrected), comparable to that of experts (96.6 %, P > 0.0125, Bonferroni corrected). CONCLUSIONS : This well-trained AI diagnostic model automatically diagnosed multiple small-bowel abnormalities simultaneously based on video-level recognition, with potential as an excellent auxiliary system for less-experienced endoscopists.

Novel scheme for non-invasive gut bioinformation acquisition with a magnetically controlled sampling capsule endoscope.

OBJECTIVE: Intestinal flora and metabolites are associated with multiple systemic diseases. Current approaches for acquiring information regarding microbiota/metabolites have limitations. We aimed to develop a precise magnetically controlled sampling capsule endoscope (MSCE) for the convenient, non-invasive and accurate acquisition of digestive bioinformation for disease diagnosis and evaluation. DESIGN: The MSCE and surgery were both used for sampling both jejunal and ileal GI content in the control and antibiotic-induced diarrhoea groups. The GI content was then used for microbiome profiling and metabolomics profiling. RESULTS: Compared with surgery, our data showed that the MSCE precisely acquired data regarding the intestinal flora and metabolites, which was effectively differentiated in different intestinal regions and disease models. Using MSCE, we detected a dramatic decrease in the abundance of Bacteroidetes, Patescibacteria and Actinobacteria and hippuric acid levels, as well as an increase in the abundance of Escherichia-Shigella and the 2-pyrrolidinone levels were detected in the antibiotic-induced diarrhoea model by MSCE. MSCE-mediated sampling revealed specific gut microbiota/metabolites including Enterococcus, Lachnospiraceae, acetyl-L-carnitine and succinic acid, which are related to metabolic diseases, cancers and nervous system disorders. Additionally, the MSCE exhibited good sealing characteristics with no contamination after sampling. CONCLUSIONS: We present a newly developed MSCE that can non-invasively and accurately acquire intestinal bioinformation via direct visualization under magnetic control, which may further aid in disease prevention, diagnosis, prognosis and treatment.

Exploration of an effective training system for diagnosis of superficial esophageal squamous cell carcinoma with magnifying narrow-band imaging: Prospective research.

BACKGROUND AND AIMS: The aim was to explore an effective training system for diagnosis of superficial esophageal squamous cell carcinoma (SESCC) and its staging with magnifying narrow-band imaging (M-NBI). PATIENTS AND METHODS: Fifteen endoscopists with no or less M-NBI experience participated in this training, which consisted of four stages and five teaching methods (M-NBI classification criterion, case analysis, hands-on operation, error correction and SESCC pathological knowledge). M-NBI images were evaluated and diagnostic accuracy was analyzed. RESULTS: After training, the accuracy of distinguishing neoplastic esophageal from non-neoplastic (0.58 ± 0.16 vs. 0.95 ± 0.05, P = 0.000) and diagnosing SESCC staging (0.25 ± 0.26 vs. 0.89 ± 0.08, P = 0.000) with M-NBI were significantly increased. Participants with no M-NBI experience achieve equivalent diagnostic accuracy with less experienced trainees after the training (0.91 ± 0.08 vs. 0.92 ± 0.04, P = 0.816). Besides, diagnosis of MM (muscularis mucosa)/SM1 (submucosal) staging tumors (Stage I, 0.47 ± 0.15; Stage II-III-IV, 0.76 ± 0.12) with M-NBI was difficult for trainees and should be the focus of this training. Every teaching method could improve the diagnostic accuracy for esophageal lesions, especially for case analysis (from 0.59 ± 0.10 to 0.85 ± 0.08, P = 0.000). In addition, the average operation score for trainees was significantly increased after hands-on teaching (60.40 ± 11.11 vs. 91.80 ± 4.28, P = 0.0001). CONCLUSIONS: For novices, this training system showed efficient performance for diagnosing SESCC staging with M-NBI. Diagnosing MM/SM1 staging SESCC was difficult for beginners, and should be the focus of training.

Gastroenterologist-Level Identification of Small-Bowel Diseases and Normal Variants by Capsule Endoscopy Using a Deep-Learning Model.

BACKGROUND & AIMS: Capsule endoscopy has revolutionized investigation of the small bowel. However, this technique produces a video that is 8-10 hours long, so analysis is time consuming for gastroenterologists. Deep convolutional neural networks (CNNs) can recognize specific images among a large variety. We aimed to develop a CNN-based algorithm to assist in the evaluation of small bowel capsule endoscopy (SB-CE) images. METHODS: We collected 113,426,569 images from 6970 patients who had SB-CE at 77 medical centers from July 2016 through July 2018. A CNN-based auxiliary reading model was trained to differentiate abnormal from normal images using 158,235 SB-CE images from 1970 patients. Images were categorized as normal, inflammation, ulcer, polyps, lymphangiectasia, bleeding, vascular disease, protruding lesion, lymphatic follicular hyperplasia, diverticulum, parasite, and other. The model was further validated in 5000 patients (no patient was overlap with the 1970 patients in the training set); the same patients were evaluated by conventional analysis and CNN-based auxiliary analysis by 20 gastroenterologists. If there was agreement in image categorization between the conventional analysis and CNN model, no further evaluation was performed. If there was disagreement between the conventional analysis and CNN model, the gastroenterologists re-evaluated the image to confirm or reject the CNN categorization. RESULTS: In the SB-CE images from the validation set, 4206 abnormalities in 3280 patients were identified after final consensus evaluation. The CNN-based auxiliary model identified abnormalities with 99.88% sensitivity in the per-patient analysis (95% CI, 99.67-99.96) and 99.90% sensitivity in the per-lesion analysis (95% CI, 99.74-99.97). Conventional reading by the gastroenterologists identified abnormalities with 74.57% sensitivity (95% CI, 73.05-76.03) in the per-patient analysis and 76.89% in the per-lesion analysis (95% CI, 75.58-78.15). The mean reading time per patient was 96.6 ± 22.53 minutes by conventional reading and 5.9 ± 2.23 minutes by CNN-based auxiliary reading (P < .001). CONCLUSIONS: We validated the ability of a CNN-based algorithm to identify abnormalities in SB-CE images. The CNN-based auxiliary model identified abnormalities with higher levels of sensitivity and significantly shorter reading times than conventional analysis by gastroenterologists. This algorithm provides an important tool to help gastroenterologists analyze SB-CE images more efficiently and more accurately.

Digestive Symptoms in COVID-19 Patients With Mild Disease Severity: Clinical Presentation, Stool Viral RNA Testing, and Outcomes.

OBJECTIVES: Coronavirus disease 2019 (COVID-19) most commonly presents with respiratory symptoms, including cough, shortness of breath, and sore throat. However, digestive symptoms also occur in patients with COVID-19 and are often described in outpatients with less severe disease. In this study, we sought to describe the clinical characteristics of COVID-19 patients with digestive symptoms and mild disease severity. METHODS: We identified COVID-19 patients with mild disease and one or more digestive symptoms (diarrhea, nausea, and vomiting), with or without respiratory symptoms, and compared them with a group presenting solely with respiratory symptoms. We followed up patients clinically until they tested negative for COVID-19 on at least 2 sequential respiratory tract specimens collected ≥24 hours apart. We then compared the clinical features between those with digestive symptoms and those with respiratory symptoms. RESULTS: There were 206 patients with low severity COVID-19, including 48 presenting with a digestive symptom alone, 69 with both digestive and respiratory symptoms, and 89 with respiratory symptoms alone. Between the 2 groups with digestive symptoms, 67 presented with diarrhea, of whom 19.4% experienced diarrhea as the first symptom in their illness course. The diarrhea lasted from 1 to 14 days, with an average duration of 5.4 ± 3.1 days and a frequency of 4.3 ± 2.2 bowel movements per day. Concurrent fever was found in 62.4% of patients with a digestive symptom. Patients with digestive symptoms presented for care later than those with respiratory symptoms (16.0 ± 7.7 vs 11.6 ± 5.1 days, P < 0.001). Nevertheless, patients with digestive symptoms had a longer duration between symptom onset and viral clearance (P < 0.001) and were more likely to be fecal virus positive (73.3% vs 14.3%, P = 0.033) than those with respiratory symptoms. DISCUSSION: We describe a unique subgroup of COVID-19 patients with mild disease severity marked by the presence of digestive symptoms. These patients are more likely to test positive for viral RNA in stool, to have a longer delay before viral clearance, and to experience delayed diagnosis compared with patients with only respiratory symptoms.

KHZ1 and KHZ2, novel members of the autonomous pathway, repress the splicing efficiency of FLC pre-mRNA in Arabidopsis.

As one of the most important events during the life cycle of flowering plants, the floral transition is of crucial importance for plant propagation and requires the precise coordination of multiple endogenous and external signals. There have been at least four flowering pathways (i.e. photoperiod, vernalization, gibberellin, and autonomous) identified in Arabidopsis. We previously reported that two Arabidopsis RNA-binding proteins, KHZ1 and KHZ2, redundantly promote flowering. However, the underlying mechanism was unclear. Here, we found that the double mutant khz1 khz2 flowered late under both long-day and short-day conditions, but responded to vernalization and gibberellin treatments. The late-flowering phenotype was almost completely rescued by mutating FLOWERING LOCUS C (FLC) and fully rescued by overexpressing FLOWERING LOCUS T (FT). Additional experiments demonstrated that the KHZs could form homodimers or interact to form heterodimers, localized to nuclear dots, and repressed the splicing efficiency of FLC pre-mRNA. Together, these data indicate that the KHZs could promote flowering via the autonomous pathway by repressing the splicing efficiency of FLC pre-mRNA.

Helicobacter pylori infection progresses proximally associated with pyloric metaplasia in age-dependent tendency: a cross-sectional study.

BACKGROUND: The elderly population presents higher morbidity of H. pylori associated diseases in proximal stomach. The specific pathogenesis and mechanism have not been clearly addressed. The gastric environment for H. pylori colonization is dynamic with increasing age. The aim of present study is to investigate the correlation among the distribution of H. pylori, mucosal inflammation, gastric microenvironment and age. METHODS: A total of 180 patients with dyspepsia symptoms were divided into young, middle-aged and elderly groups. Biopsies were obtained from each patient in five locations: great curvature (mid-corpus, mid-antrum), lesser curvature (mid-corpus, mid-antrum) and incisura angularis (IA), analyzed for H. pylori density, mucosal inflammation and histopathology. RESULTS: The infection rate of H. pylori increased linearly with age (p <  0.001) in corpus, but not in antrum and IA. The H. pylori density was significantly aggravated in IA (p = 0.002) and corpus (p <  0.001) in elderly patient, but not in antrum. The mucosa inflammation scores were consistent with the severity of H. pylori colonization among three age groups. In elderly patients, the pyloric glands present more frequently in corpus, comparing with young and middle-aged group. A significant positive correlation among aggravating severity of H. pylori infection, mucosal inflammation and pyloric metaplasia in corpus with increasing age (p <  0.001) was occurred. CONCLUSIONS: With increasing age, both topographic distribution of H. pylori and the expansion of pyloric glands increased in a distal-to-proximal gastric direction. Pyloric metaplasia in corpus was correlated with the risk of aggravated H. pylori colonization and associated inflammation in elderly population.

Variability of the Ki-67 proliferation index in gastroenteropancreatic neuroendocrine neoplasms - a single-center retrospective study.

BACKGROUND: The Ki-67 index in gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs) may change throughout the disease course. However, the definitive effect of Ki-67 variability on GEP-NENs remains unknown. The aims of this study were to evaluate changes in Ki-67 levels throughout the disease course and investigate the role of Ki-67 index variability in GEP-NENs. METHODS: Specimens with multiple pathologies were evaluated from 30 patients who were selected from 514 patients with GEP-NENs, being treated at Wuhan Union Hospital from July 2009 to February 2018. The Ki-67 index was evaluated among multiple specimens over the disease course. Univariable and multivariable Cox proportional hazards regression analyses were performed to assess the prognostic significance of various clinical and histopathologic features. RESULTS: Among the 514 patients with GEP-NENs, metastases were seen in 182 (35.41%). Among the 30 patients from whom specimens with multiple pathologies were obtained, 24 were both primary and metastatic specimens and six were specimens collected over the course of the disease. Changes in Ki-67 levels were detected in 53.3% of the patients, of whom 40% had up-regulated Ki-67 levels, and 13.3% had down-regulated Ki-67 levels. Kaplan-Meier survival analysis showed that the group with Ki-67 variability had a shorter overall survival (p = 0.0297). The Cox regression analysis indicated that Ki-67 variability (p = 0.038) was the only independent prognostic factor for overall survival. CONCLUSIONS: Our data suggest that patients with GEP-NENs and Ki-67 variability had a poorer prognosis. The re-assessment of Ki-67 at sites of metastasis or during the disease course might play a role in predicting the prognosis of patients with GEP-NENs. This finding could have implications for how GEP-NENs are monitored and treated.

Synthesis, Cytotoxic, and Antibacterial Evaluation of Quinazolinone Derivatives with Substituted Amino Moiety.

A series of novel quinazolinone derivatives containing a substituted amino moiety were synthesized, evaluated for their cytotoxic and antibacterial activities. The results of MTT assay showed that all synthesized target compounds 5A - 5O showed potent cytotoxicity against SGC-7901 (IC50 , 0.72 - 1.41 µm). Moreover, the compounds 5D, 5I, and 5K showed better selectivity as compared with positive controls pemetrexed and MTX due to weak cytotoxicity against normal tissue cell line HUVSMC. Among synthesized compounds, the compounds 5E, 5J, 5L, and 5N showed broad-spectrum cytotoxic activities against at least four cancer cell lines at a micromolar level. The results of antibacteria evaluation revealed that all synthesized compounds showed good to moderate antibacterial activities against Gram-negative bacteria Escherichia coli. Among them, the MIC values of the compounds 5C, 5F, and 5M were 0.31 µg/mL.

Arabidopsis KHZ1 and KHZ2, two novel non-tandem CCCH zinc-finger and K-homolog domain proteins, have redundant roles in the regulation of flowering and senescence.

KEY MESSAGE: The two novel CCCH zinc-finger and K-homolog (KH) proteins, KHZ1 and KHZ2, play important roles in regulating flowering and senescence redundantly in Arabidopsis. The CCCH zinc-finger proteins and K-homolog (KH) proteins play important roles in plant development and stress responses. However, the biological functions of many CCCH zinc-finger proteins and KH proteins remain uncharacterized. In Arabidopsis, KHZ1 and KHZ2 are characterized as two novel CCCH zinc-finger and KH domain proteins which belong to subfamily VII in CCCH family. We obtained khz1, khz2 mutants and khz1 khz2 double mutants, as well as overexpression (OE) lines of KHZ1 and KHZ2. Compared with the wild type (WT), the khz2 mutants displayed no defects in growth and development, and the khz1 mutants were slightly late flowering, whereas the khz1 khz2 double mutants showed a pronounced late flowering phenotype. In contrast, artificially overexpressing KHZ1 and KHZ2 led to the early flowering. Consistent with the late flowering phenotype, the expression of flowering repressor gene FLC was up-regulated, while the expression of flowering integrator and floral meristem identity (FMI) genes were down-regulated significantly in khz1 khz2. In addition, we also observed that the OE plants of KHZ1 and KHZ2 showed early leaf senescence significantly, whereas the khz1 khz2 double mutants showed delayed senescence of leaf and the whole plant. Both KHZ1 and KHZ2 were ubiquitously expressed throughout the tissues of Arabidopsis. KHZ1 and KHZ2 were localized to the nucleus, and possessed both transactivation activities and RNA-binding abilities. Taken together, we conclude that KHZ1 and KHZ2 have redundant roles in the regulation of flowering and senescence in Arabidopsis.

Na+/H+ Exchanger Regulates Amino Acid-Mediated Autophagy in Intestinal Epithelial Cells.

BACKGROUND/AIMS: Dysfunctional autophagy has been reported to be associated with aberrant intestinal metabolism. Amino acids can regulate autophagic activity in intestinal epithelial cells (IECs). Na+/H+-exchanger 3 (NHE3) has been found to participate in the absorption of amino acids in the intestine, but whether NHE3 is involved in the regulation of autophagy in IECs is unclear. METHODS: In the present study, an amino acid starvation-induced autophagic model was established. Then, the effects of alanine and proline with or without the NHE inhibitor 5-(N-ethyl-N-isopropyl) amiloride (EIPA) were evaluated. Autophagy was examined based on the microtubule-associated light chain 3 (LC3) levels, transmission electron microscopy (TEM), tandem GFP-mCherry-LC3 construct, sequestosome-1 (SQSTM1, P62) mRNA and protein levels, and autophagy-related gene (ATG) 5, 7, and 12 expression levels. The autophagic flux was evaluated as the ratio of yellow (autophagosomes) to red (autolysosomes) LC3 puncta. RESULTS: Following amino acid starvation, we found the LC3-II and ATG expression levels were enhanced in the IEC-18 cells. An increase in the number of autophagic vacuoles was concomitantly observed by TEM and confocal microscopy. Based on the results, supplementation with either alanine or proline depressed autophagy in the IEC-18 cells. Consistent with the elevated LC3-II levels, ATG expression increased upon NHE3 inhibition. Moreover, the mCherry-GFP-LC3 autophagic puncta representing both autophagosomes and autolysosomes per cell increased after EIPA treatment. CONCLUSIONS: These results demonstrate that NHE (most likely NHE3) may participate in the amino acid regulation of autophagy in IECs, which would aid in the design of better treatments for intestinal inflammation.

Efficacy of Mesenchymal Stromal Cells for Fistula Treatment of Crohn's Disease: A Systematic Review and Meta-Analysis.

BACKGROUND AND AIM: The introduction of mesenchymal stromal cells (MSCs) has changed the management of Crohn's fistula, while it remains controversial. The aim of this study was to provide an overview of efficacy and optimum state of MSCs treatment on Crohn's fistula. METHODS: Studies reporting MSCs treatment on Crohn's fistula were searched and included. A fixed-effects model was used to assess the efficacy of MSCs, and outcomes of healing and recurrence were used to evaluate the best states of MSCs intervention. RESULTS: Fourteen articles were enrolled (n = 477). Pooled analysis showed MSCs had a significant efficacy compared to other treatments [risk difference: 0.21 (0.09, 0.32), P = 0.000]. Notably, after MSCs treatment, the group of Crohn's disease activity index (CDAI) baseline >150 group had a higher healing rate (HR) and a clinical response (a change in CDAI of >50 points) (79.17 ± 8.78 vs. 47.54 ± 15.90, P = 0.011) compared to CDAI baseline of <150. The duration time of CD and fistulas had a negative correlation with HR accompanied by MSC therapy (r = -0.900, -0.925). Then, a moderate dose MSCs (2-4 × 107 cells/ml) had a higher HR (80.07%) and lower recurrence rate (RR 13.98%) compared to other dosages. Moreover, adipose-derived MSCs therapy had an advantage over bone marrow-derived MSCs in terms of low RR (7.4 ± 4.28 vs. 13.39 ± 0.89). CONCLUSIONS: The evidence supported the effect of MSCs at a more appropriate time of Crohn's fistula. And CDAI baseline (the points >150) has been a candidate for evaluating effectiveness of MSCs application on Crohn's fistula.

The Role of Probiotics in Lipopolysaccharide-Induced Autophagy in Intestinal Epithelial Cells.

BACKGROUND/AIMS: Dysfunction of autophagy has been associated with loss of intestinal homeostasis. Lipopolysaccharide (LPS) from Gram-negative bacteria is known to be a major initiator of intestinal epithelial cell (IEC) autophagy. Although probiotics have been recognized to be involved in many therapeutic properties and participate in host defense responses, the molecular mechanisms by which probiotics exert these positive effects remain unknown. This study assessed the effect of probiotics on LPS-induced physical barrier dysfunction and the underlying mechanism of probiotic action in IECs with a focus on autophagy. METHODS: A LPS-induced autophagic model was established in rat IEC18 cells wherein cells were treated with culture medium supernatants of Bifidobacteria following LPS intervention at indicated times. Autophagosomes in IEC18 cells were visualized by confocal microscopy after transfection with a tandem GFP-mCherry-LC3 construct and also by transmission electron microscopy. Autophagy-associated protein levels were analyzed by western blot and transepithelial electrical resistance (TEER) was measured using an epithelial voltohmmeter. RESULTS: Probiotic treatment could effectively inhibit LPS-induced autophagy, as evidenced by the decreased ratio of microtubule-associated light chain 3 (LC3)-II/LC3-I, fewer autophagic vacuoles, and reduced punctate distribution of GFP-mCherry-LC3. In addition, probiotics prevented chloroquine (CQ) inhibition of autophagic flux and autophagolysosomal fusion as indicated by a failure to recruit LAMP1 and cathepsin D to lysosomes. Interestingly, ATG16L1 knockdown did not inhibit the effect of probiotics on LPS-induced autophagy. Furthermore, the diminished barrier function could be prevented by probiotics. CONCLUSIONS: We provide evidence that autophagy mediation by probiotics may be involved in enteroprotection against LPS-induced intestinal epithelial toxicity, and could serve as a novel mechanism through which probiotics promote and maintain gut homeostasis.