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.

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.

Epidemiologic trends and survival of early-onset gastroenteropancreatic neuroendocrine neoplasms.

Background: The epidemiologic trends and survival related to early-onset gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs) have not been well explored. Methods: Trends in the incidence and incidence-based mortality of early-onset GEP-NENs between 1975 and 2018 were obtained from the Surveillance, Epidemiology, and End Results database, and were stratified by age, sex, race, tumor site, stage, and grade. Associated population data were used to determine overall survival (OS) and independent prognostic factors for patients with early-onset GEP-NENs. Results: A total of 17299 patients diagnosed with early-onset GEP-NENs were included in this study. Results revealed an increase in the incidence (5.95% per year, 95% confidence interval (CI), 5.75-6.14%) and incidence-based mortality (4.24% per year, 95% CI, 3.92-4.56%) for early-onset GEP-NENs from 1975 to 2018, with higher rates of increase than those of later-onset GEP-NENs (incidence: 4.45% per year, 95% CI, 4.38-4.53; incidence-based mortality: 4.13% per year, 95% CI, 3.89-4.37; respectively). Increases in incidence were observed across all age, races, tumor sites, grades, and stages, except for patients with unknown stage. Compared to those with later-onset GEP-NENs, a higher proportion of female gender (54.5% vs. 49.0%, p <0.001), well-differentiated tumor (31.1% vs. 28.0%, p <0.05), and localized disease (55.2% vs. 46.7%, p <0.05) were observed in the cohort of patients with early-onset GEP-NENs. Moreover, early-onset GEP-NENs exhibited a superior overall survival in comparison to later-onset GEP-NENs, irrespective of tumor site, grade, or stage (p <0.0001). Multivariable survival analysis identified that race, marital status, stage, grade, chemotherapy, and primary site were significantly correlated with OS in individuals with early-onset GEP-NENs. Conclusions: The incidence and incidence-based mortality rates of early-onset GEP-NENs have steadily increased over time, with higher rates of increase than those of later-onset GEP-NENs. The clinical characteristics and survival were different between early-onset and later-onset GEP-NENs groups. Race, marital status, stage, grade, chemotherapy, and primary site were independent prognostic factors for early-onset GEP-NENs. Further investigations are warranted to better understand the characteristics of this disease subgroup.

Predicting the survival of patients with pancreatic neuroendocrine neoplasms using deep learning: A study based on Surveillance, Epidemiology, and End Results database.

BACKGROUND: The study aims to evaluate the performance of three advanced machine learning algorithms and a traditional Cox proportional hazard (CoxPH) model in predicting the overall survival (OS) of patients with pancreatic neuroendocrine neoplasms (PNENs). METHOD: The clinicopathological dataset obtained from the Surveillance, Epidemiology, and End Results database was randomly assigned to the training set and testing set at a ratio of 7:3. The concordance index (C-index) and integrated Brier score (IBS) were used to compare the predictive performance of the models. The accuracy of the model in predicting the 5-year and 10-year survival rates was compared using the receiver operating characteristic curve, decision curve analysis (DCA) and calibration curve. RESULTS: This study included 3239 patients with PNENs in total. The DeepSurv model had the highest C-index of 0.7882 in the testing set and training set and the lowest IBS of 0.1278 in the testing set compared with the CoxPH, neural multitask logistic and random survival forest models (C-index = 0.7501, 0.7616, and 0.7612, respectively; IBS = 0.1397, 0.1418, and 0.1432, respectively). Moreover, the DeepSurv model had the highest accuracy in predicting 5- and 10-year OS rates (area under the curve: 0.87 and 0.90). DCA showed that the DeepSurv model had high potential for clinical decisions in 5- and 10-year OS models. Finally, we developed an online application based on the DeepSurv model for clinical use (https://whuh-ml-neuroendocrinetumor-app-predict-oyw5km.streamlit.app/). CONCLUSIONS: All four models analyzed above can predict the prognosis of PNENs well, among which the DeepSurv model has the best prediction performance.

Bone mesenchymal stem cells promote gastric cancer progression through TGF-ß1/Smad2 positive feedback loop.

AIMS: Bone marrow-derived mesenchymal stem cells (BMSCs) have been proven to be recruited into the tumor microenvironment and contribute to gastric cancer (GC) progression, but the underlying mechanism is still unclear. The purpose of this study is to explore the exact role and potential mechanism of BMSCs in the progression of GC. MATERIALS AND METHODS: Bioinformatics analyzed were used to clarify the correlation between TGF-ß1 and prognosis of gastric cancer. Cell co-culture were used to explore the interaction between gastric cancer cells (GCs) and BMSCs. Quantitative real time-PCR and Western blot assay were used to detect gene and protein expression, respectively. The biological characteristics of GCs and BMSCs were detected by immunofluorescence, Transwell migration, Elisa and invasion assay. Xenograft models in nude mice were constructed to evaluate GC development in vivo. KEY FINDINGS: TGF-ß1 was overexpressed in GC cells and tissues, and is positively related to the poor prognosis of patients. TGF-ß1 from GCs activated the Smad2 pathway in BMSCs, promoting their differentiation into carcinoma-associated fibroblasts (CAFs) and TGF-ß1 expression. Concomitantly, TGF-ß1 secreted by CAFs activate Smad2 signaling in GC cells, thus inducing their epithelial-mesenchymal transition (EMT) and TGF-ß1 secretion. BMSCs can dramatically promote the proliferation, migration, and invasion of GCs while blocking TGF-ß1/Smad2 positive feedback loop can reverse these effects. SIGNIFICANCE: The TGF-ß1/Smad2 positive feedback loop between GCs and BMSCs, promotes the CAFs differentiation of BMSCs and the EMT of GCs, resulting in the progression of GC.

Nomograms for Predicting Survival Outcomes in Patients with Neuroendocrine Neoplasms of the Gallbladder Undergoing Primary Tumor Resection: A Population-Based Study.

BACKGROUND: Neuroendocrine neoplasms of the gallbladder (GB-NENs) are a rare group of histologically heterogeneous tumors, and surgical resection of the primary tumor is the mainstream treatment at the moment. The current study aimed to establish and validate novel nomograms for patients with GB-NENs undergoing primary tumor resection to predict the 6-, 12-, and 18-month overall survival (OS) and cancer-specific survival (CSS). METHODS: Clinicopathological information of patients with GB-NENs undergoing primary tumor resection between 2004 and 2018 was derived from the Surveillance, Epidemiology, and End Results (SEER) database. Candidate prognostic factors were selected by Cox regression analyses, and the nomograms were constructed. Finally, concordance index (C-index), calibration plot, area under the curve from the receiver operating characteristic curve (AUC), and decision curve analysis (DCA) were utilized to assess the effective performance of the nomograms. RESULTS: A total of 221 patients with GB-NENs undergoing resection were enrolled in this retrospective study. Using the Cox regression analyses, age, pathological classification, tumor size, and SEER stage were identified as the independent prognostic factors of patients with GB-NENs undergoing resection, and nomograms were constructed. The C-indexes of OS and CSS in training dataset were 0.802 (95% CI: 0.757-0.848) and 0.846 (95% CI: 0.798-0.895), while those of internal validation dataset were 0.862 (95% CI: 0.802-0.922) and 0.879 (95% CI: 0.824-0.934), respectively. CONCLUSIONS: Taken together, the nomograms are accurate enough to predict the prognostic factors of GB-NEN patients undergoing resection, allowing for treatment decision-making and clinical monitoring for future clinical work.

Cardiovascular-Specific Mortality among Gastrointestinal Stromal Tumor Patients: A Population-Based Analysis.

Background: The overall risk of cardiovascular mortality (CVM) in cancer survivors has increased with time. The trend of CVM in patients with gastrointestinal stromal tumors (GISTs) remains unclear. This study is aimed at assessing the risks and independent predictors of CVM in GIST patients. Methods: Data of the GIST patients were extracted from the Surveillance, Epidemiology, and End Results (SEER) database (2000-2019). The standardized mortality ratio (SMR) was used to evaluate the risk of CVM, and a multivariate competing risk model was utilized to identify the predictors for CVM. Results: A total of 12,058 patients with GIST were included in this study, of whom 477 (4.0%) patients died of cardiovascular disease (CVD). The SMR for CVM among GIST patients was significantly higher than in the general population (SMR, 3.23, 95% CI: 2.97-3.52), and all categories of CVD were associated with a significantly elevated SMR. The cumulative mortality of CVD was the lowest among all causes of death, while the CVM was the second most common cause of death in patients ≥ 80 years when stratified by age at diagnosis. Furthermore, male sex, older age at diagnosis, White race, unmarried, earlier year of diagnosis, and not receiving chemotherapy were the poor prognostic factors for CVM. Conclusions: The CVM risk in GIST patients was significantly higher relative to the general US population. Timely screening and cardioprotective interventions should be implemented to prevent the occurrence of CVM in patients with GIST.

Clinicopathological characteristics of mixed neuroendocrine-non-neuroendocrine neoplasms in gastrointestinal tract.

BACKGROUND: In 2019, the World Health Organization (WHO) classification system updated the definition of mixed neuroendocrine non-neuroendocrine neoplasms (MiNENs), previously known as mixed adenoneuroendocrine carcinomas (MANECs). The clinicopathological characteristics of this new definition remains to be clarified. METHODS: We analyzed the clinical data of 43 patients diagnosed with MiNENs in Wuhan Union Hospital from 2011 to 2020 according to the definition of MiNENs proposed in 2019. RESULTS: Among the 43 patients with MiNENs, the top two most common sites were stomach and colon, and 69.8% were males. Nearly half (21/43) of the patients were diagnosed at TNM stage III, and about 53.5% (23/43) of patients were the neuroendocrine neoplasm dominant type. Among the non-neuroendocrine tumor components of 43 MiNENs patients, adenocarcinoma accounted for 95.3% (41/43) and squamous cell carcinoma accounted for 4.7% (2/43)；95.3% (41/43) of the neuroendocrine neoplasm components were neuroendocrine carcinoma (NEC) and 4.7% (2/43) were neuroendocrine tumor (NET). Approximately 60.5% (26/43) neuroendocrine components had a Ki-67 index ≥ 55%. In addition, we further compared the prognosis of different subtypes of the MiNENs based on the neuroendocrine neoplasm component and non-neuroendocrine neoplasm component, and the results showed that there was no significant difference in survival between different subtypes of MiNENs (P > 0.05). CONCLUSIONS: MiNENs can exhibit diverse clinicopathological characteristics, and there is no significant difference in prognosis among MiNENs subtypes, indicating that the definition of MiNENs can well summarize the prognosis of this type of tumor.

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.

BMSCs Promote Differentiation of Enteric Neural Precursor Cells to Maintain Neuronal Homeostasis in Mice With Enteric Nerve Injury.

BACKGROUND & AIMS: Our previous study showed that transplantation of bone marrow-derived mesenchymal stem cells (BMSCs) promoted functional enteric nerve regeneration in denervated mice but not through direct transdifferentiation. Homeostasis of the adult enteric nervous system (ENS) is maintained by enteric neural precursor cells (ENPCs). Whether ENPCs are a source of regenerated nerves in denervated mice remains unknown. METHODS: Genetically engineered mice were used as recipients, and ENPCs were traced during enteric nerve regeneration. The mice were treated with benzalkonium chloride to establish a denervation model and then transplanted with BMSCs 3 days later. After 28 days, the gastric motility and ENS regeneration were analyzed. The interaction between BMSCs and ENPCs in vitro was further assessed. RESULTS: Twenty-eight days after transplantation, gastric motility recovery (gastric emptying capacity, P < .01; gastric contractility, P < .01) and ENS regeneration (neurons, P < .01; glial cells, P < .001) were promoted in BMSCs transplantation groups compared with non-transplanted groups in denervated mice. More importantly, we found that ENPCs could differentiate into enteric neurons and glial cells in denervated mice after BMSCs transplantation, and the proportion of Nestin+/Ngfr+ cells differentiated into neurons was significantly higher than that of Nestin+ cells. A small number of BMSCs located in the myenteric plexus differentiated into glial cells. In vitro, glial cell-derived neurotrophic factor (GDNF) from BMSCs promotes the migration, proliferation, and differentiation of ENPCs. CONCLUSIONS: In the case of enteric nerve injury, ENPCs can differentiate into enteric neurons and glial cells to promote ENS repair and gastric motility recovery after BMSCs transplantation. BMSCs expressing GDNF enhance the migration, proliferation, and differentiation of ENPCs.

Glutamate regulates gliosis of BMSCs to promote ENS regeneration through α-KG and H3K9/H3K27 demethylation.

BACKGROUND: There is a lack of effective therapies for enteric nervous system (ENS) injury. Our previous study showed that transplanted bone marrow-derived mesenchymal stem cells (BMSCs) play a "glia-like cells" role in initiating ENS regeneration in denervated mice. Cellular energy metabolism is an important factor in maintaining the biological characteristics of stem cells. However, how cellular energy metabolism regulates the fate of BMSCs in the ENS-injured microenvironment is unclear. METHODS: The biological characteristics, energy metabolism, and histone methylation levels of BMSCs following ENS injury were determined. Then, glutamate dehydrogenase 1 (Glud1) which catalyzes the oxidative deamination of glutamate to α-KG was overexpressed (OE) in BMSCs. Further, OE-Glud1 BMSCs were targeted-transplanted into the ENS injury site of denervated mice to determine their effects on ENS regeneration. RESULTS: In vitro, in the ENS-injured high-glutamate microenvironment, the ratio of α-ketoglutarate (α-KG) to succinate (P < 0.05), the histone demethylation level (P < 0.05), the protein expression of glial cell markers (P < 0.05), and the gene expression of Glud1 (P < 0.05) were significantly increased. And the binding of H3K9me3 to the GFAP, S100B, and GDNF promoter was enhanced (P < 0.05). Moreover, α-KG treatment increased the monomethylation and decreased the trimethylation on H3K9 (P < 0.01) and H3K27 (P < 0.05) in BMSCs and significantly upregulated the protein expression of glial cell markers (P < 0.01), which was reversed by the α-KG competitive inhibitor D-2-hydroxyglutarate (P < 0.05). Besides, overexpression of Glud1 in BMSCs exhibited increases in monomethylation and decreases in trimethylation on H3K9 (P < 0.05) and H3K27 (P < 0.05), and upregulated protein expression of glial cell markers (P < 0.01). In vivo, BMSCs overexpressing Glud1 had a strong promotion effect on ENS regeneration in denervated mice through H3K9/H3K27 demethylation (P < 0.05), and upregulating the expression of glial cell protein (P < 0.05). CONCLUSIONS: BMSCs overexpressing Glud1 promote the expression of glial cell markers and ENS remodeling in denervated mice through regulating intracellular α-KG and H3K9/H3K27 demethylation.

Eradication of Helicobacter pylori alleviates lipid metabolism deterioration: a large-cohort propensity score-matched analysis.

BACKGROUND: The impact of Helicobacter pylori (H. pylori) eradication on metabolism of lipid and the potential predictor of such changes remain unclear. METHODS: This study retrospectively included subjects who underwent at least two 13C urea breath tests between 2015 and 2019 at Wuhan Union Hospital. Based on two H. pylori 13C examination results, subjects were divided into propensity score-matched persistently negative (HPN), persistently positive (HPP), and eradication (HPE) groups. The changes in lipid measurements from before to after H. pylori eradication, including high-density lipoprotein (HDL), low-density lipoprotein (LDL), total cholesterol, and triglycerides, were compared within and between groups. Forty-two candidate factors were tested for their ability to predict lipid metabolism changes after H. pylori eradication. RESULTS: After propensity score matching, 3412 matched cases were analyzed. Within-group comparisons showed significantly decreased HDL (P <  0.001) and increased LDL (P <  0.001) at the second examination in both the HPE and HPP groups. Between-group comparisons showed that the HDL decrease of the HPE group was significantly larger and smaller when compared with the HPN (P = 0.001) and HPP (P = 0.004) group, respectively. Uni- and multivariate analyses showed that low diastolic blood pressure (DBP) (P = 0.002) and high mean platelet volume (MPV) (P = 0.001) before eradication were associated with increased HDL after eradication. Low total protein (TP) (P <  0.001) was associated with decreased LDL after eradication. CONCLUSIONS: Compared with sustained H. pylori infectious states, H. pylori eradication alleviated the lipid metabolism deterioration but did not restore it to the uninfected level within 1.5 years after eradication. Patients with low DBP, high MPV, and low TP may reap a greater lipid-metabolism benefit from H. pylori eradication.

Confocal Laser Endomicroscopy Can Improve the Diagnosis Rate and Range Assessment of Patients With Conflicting Chronic Atrophic Gastritis Results of White Light Endoscopic and Pathological Diagnosis.

Background and Aims: Chronic atrophic gastritis (CAG) is closely related to the development of gastric cancer. However, the diagnostic accuracy of white light endoscopy (WLE) biopsy for CAG is poor. The diagnostic role and efficacy of confocal laser endomicroscopy (CLE) in CAG missed under WLE biopsy remain unclear. Methods: This study is a single-center prospective study that included 21 patients from 1,349 patients who underwent WLE and biopsy and whose WLE results confirmed CAG, but pathological results did not. Then, all these patients received CLE examination and underwent targeted biopsies and five-point standard biopsies. The sensitivity, specificity, and accuracy of CLE diagnosis and targeted biopsy were analyzed. Results: The pathological results of five-point standard biopsies in 21 patients confirmed CAG, and 17 patients (81.0%) were confirmed to have intestinal metaplasia (IM). According to the image diagnosis of CLE, there were 19 cases (90.5%) of CAG and 14 cases (66.7%) of IM among these 21 patients. According to the targeted biopsy of CLE, 17 cases (81.0%) of CAG and 14 cases (66.7%) of IM were diagnosed. There was no significant difference between CLE image diagnosis and five-point standard biopsies in terms of atrophy severity score (p = 0.927), IM severity score (p = 0.250), atrophy scope score (p = 0.781), and IM scope score (p = 0.195). For CAG, the sensitivity and accuracy of CLE image diagnosis were higher than those of CLE targeted biopsies (90.5% vs. 81.0%, p = 0.331), but for IM, the diagnosis was the same. Conclusions: CLE can improve the diagnosis rate of CAG and can increase the comprehensive assessment of the scope and severity of CAG.

THBS4/integrin α2 axis mediates BM-MSCs to promote angiogenesis in gastric cancer associated with chronic Helicobacter pylori infection.

BACKGROUND: BM-MSCs contribute to Helicobacter pylori (H. pylori)-induced gastric cancer, but their mechanism is still unclear. The aim of our study was to investigate the specific role and mechanism of BM-MSCs in H. pylori-induced gastric cancer. MAIN METHODS: Mice received total bone marrow transplants and were then infected with H. pylori. BM-MSCs were extracted and transplanted into the gastric serosal layer of mice chronically infected with H. pylori. Hematoxylin and eosin staining, immunohistochemistry staining and immunofluorescence were performed to detect tumor growth and angiogenesis in mouse stomach tissues. Chicken chorioallantoic membrane assays, xenograft tumor models, and human umbilical vein endothelial cell tube formation assays were used for in vivo and in vitro angiogenesis studies. THBS4 was screened from RNA-seq analysis of gastric tissues of BM-MSCs transplanted into H. pylori-infected mice. RESULTS: BM-MSCs can migrate to the site of chronic mucosal injury and promote tumor angiogenesis associated with chronic H. pylori infection. Migration of BM-MSCs to the site of chronic mucosal injury induced the upregulation of THBS4, which was also evident in human gastric cancer and correlated with increased blood vessel formation and worse outcome. The THBS4/integrin α2 axis promoted angiogenesis by facilitating the PI3K/AKT pathway in endothelial cells. CONCLUSIONS: Our results revealed a novel proangiogenic effect of BM-MSCs in the chronic H. pylori infection microenvironment, primarily mediated by the THBS4/integrin α2 axis, which activates the PI3K/AKT pathway in endothelial cells and eventually induces the formation of new tumor vessels.

Bone marrow-derived mesenchymal stem cells promote Helicobacter pylori-associated gastric cancer progression by secreting thrombospondin-2.

OBJECTIVES: Bone marrow-derived cells (BMDCs), especially mesenchymal stem cells (MSCs), may be involved in the development of Helicobacter pylori-associated gastric cancer (GC) in mice, but the specific mechanism remains unclear, and evidence from human studies is lacking. MATERIALS AND METHODS: To verify the role of BM-MSCs in H pylori-associated GC, green fluorescent protein (GFP)-labelled BM-MSCs were transplanted into the subserosal layers of the stomach in a mouse model of chronic H pylori infection. Three months post-transplantation, the mice were sacrificed, and the gastric tissues were subjected to histopathological and immunofluorescence analyses. In addition, we performed fluorescence in situ hybridization (FISH) and immunofluorescence analyses of gastric tissue from a female patient with H pylori infection and a history of acute myeloid leukaemia who received a BM transplant from a male donor. RESULTS: In mice with chronic H pylori infection, GFP-labelled BM-MSCs migrated from the serous layer to the mucosal layer and promoted GC progression. The BM-MSCs differentiated into pan-cytokeratin-positive epithelial cells and α-smooth muscle actin-positive cancer-associated fibroblasts (CAFs) by secreting the protein thrombospondin-2. FISH analysis of gastric tissue from the female patient revealed Y-chromosome-positive cells. Immunofluorescence analyses further confirmed that Y-chromosome-positive cells showed positive BM-MSCs marker. These results suggested that allogeneic BMDCs, including BM-MSCs, can migrate to the stomach under chronic H pylori infection. CONCLUSIONS: Taken together, these findings imply that BM-MSCs participate in the development of chronic H pylori-associated GC by differentiating into both gastric epithelial cells and CAFs.

CD44 fucosylation on bone marrow-derived mesenchymal stem cells enhances homing and promotes enteric nervous system remodeling in diabetic mice.

BACKGROUND: Diabetes can cause extensive enteric nervous system (ENS) injuries and gastrointestinal motility disorder. In developing possible treatments, researchers have engaged in tissue regeneration engineering with the very promising bone marrow-derived mesenchymal stem cells (BMSCs). However, BMSCs have poor homing ability to the targeted tissues after intravenous injection. Thus, we aimed to investigate whether enhancing the expression of E-selectin ligand on BMSCs could improve their homing ability and subsequently influence their role in ENS remodeling in diabetic mice. METHODS: First, we constructed the fucosylation modification of CD44 on BMSCs through a fucosyltransferase VII (FTVII) system to generate a Hematopoietic Cell E-/L-selectin Ligand (HCELL) property, a fucosylated sialyllactosaminyl glycovariant of CD44 that potently binds E-selectin. Next, FTVII-modified and unmodified BMSCs labeled with green fluorescent protein (GFP) were injected into diabetic mice through the tail vein to compare their homing ability to the gastrointestinal tract and their effect on ENS remodeling, respectively. A bioluminescent imaging system was used to evaluate the homing ability of GFP-labeled BMSCs with and without FTVII modification, to the gastrointestinal tract. Gastrointestinal motility was assessed by gastrointestinal transient time, defecation frequency, stool water content and colon strips contractility. Immunofluorescence staining and western blotting were used to assess the expression levels of protein gene product 9.5 (PGP9.5), glial fibrillary acidic protein (GFAP) and glial cell line-derived neurotrophic factor (GDNF). RESULTS: The FTVII-mediated α(1,3)-fucosylation modification of CD44 on BMSCs generated a HCELL property. Bioluminescent imaging assays showed that FTVII-modified BMSCs had enhanced homing ability to gastrointestinal tract, mainly to the colon, 24 h after injection through the tail vein. Compared with diabetic mice, FTVII-modified BMSCs significantly promoted the gastrointestinal motility and the ENS remodeling, including intestinal peristalsis (P < 0.05), increased feces excretion (P < 0.05) and the water content of the feces (P < 0.05), restored the spontaneous contraction of the colon (P < 0.05), and upregulated the protein expression levels of PGP9.5 (P < 0.01), GFAP (P < 0.001), and GDNF (P < 0.05), while unmodified BMSCs did not (P > 0.05). CONCLUSIONS: CD44 fucosylation modification on murine BMSCs promotes homing ability to the gastrointestinal tract and ENS remodeling in diabetic mice.

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.

Concordance Between the Ki-67 Index Cutoff Value of 55% and Differentiation in Neuroendocrine Tumor and Neuroendocrine Carcinoma in Grade 3 Pancreatic Neuroendocrine Neoplasms.

OBJECTIVE: In 2017 and 2019, the World Health Organization defined grade 3 neuroendocrine tumors (G3 NETs) and neuroendocrine carcinoma (G3 NEC) in the pancreas. The validity of this classification remains to be verified. METHODS: Clinical data were collected and analyzed for 39 G3 pancreatic neuroendocrine neoplasms (PanNENs) patients between 2009 and 2018. RESULTS: The tumor-node-metastasis stage (P = 0.0260), differentiation (P = 0.0115), and Ki-67 index (P = 0.0371) are prognostic factors for G3 PanNENs by Kaplan-Meier survival analysis. Among 39 patients, 18 had a Ki-67 index of less than 55% and well-differentiated morphology (G3 NET) and 16 had a Ki-67 index of 55% or greater and poorly differentiated morphology (G3 NEC). Grade 3 neuroendocrine tumor had a significant better prognosis than G3 NEC (median overall survival time, 25 months [95% confidence interval, 10.854-39.146 months] vs 12 months [95% confidence interval, 6.316-17.684 months], P = 0.0164). Based on Cox regression analyses, tumor-node-metastasis stage (P = 0.016) was identified as the independent prognostic factor for G3 PanNENs. CONCLUSIONS: The upper Ki-67 index cutoff of 55% might be the best cutoff value to define G3 NETs and G3 NECs for G3 PanNENs. The World Health Organization 2017 and 2019 classification system for G3 PanNENs can identify high-risk patients with G3 PanNENs.

Clinicopathological heterogeneity between primary and metastatic sites of gastroenteropancreatic neuroendocrine neoplasm.

BACKGROUND: Chromogranin A (CgA), synaptophysin (Syn) and the Ki-67 index play significant roles in diagnosis or the evaluation of the proliferative activity of gastroenteropancreatic neuroendocrine neoplasms (GEP-NENs). However, little is known about whether these biological markers change during tumor metastasis and whether such changes have effect on prognosis. METHODS: We analyzed 35 specimens of both primary and metastatic tumor from 779 patients who had been diagnosed as GEP-NENs at Wuhan Union Hospital from August 2011 to October 2019. The heterogeneity of CgA, Syn and Ki-67 index was evaluated by immunohistochemical analysis. RESULTS: Among these 779 patients, the three most common sites of NENs in the digestive tract were the pancreas, rectum and stomach. Metastases were found in 311 (39.9%) patients. Among the 35 patients with both primary and metastatic pathological specimens, differences in the Ki-67 level were detected in 54.3% of the patients, while 37.1% showed a difference in CgA and only 11.4% showed a difference in Syn. Importantly, due to the difference in the Ki-67 index between primary and metastatic lesions, the WHO grade was changed in 8.6% of the patients. In addition, a Kaplan-Meier survival analysis showed that patients with Ki-67 index variation had a shorter overall survival (p = 0.0346), while neither Syn variation nor CgA variation was related to patient survival (p = 0.7194, p = 0.4829). CONCLUSIONS: Our data indicate that primary and metastatic sites of GEP-NENs may exhibit pathological heterogeneity. Ki-67 index variation is closely related to the poor prognosis of patients with tumor metastasis, but neither Syn variation nor CgA variation is related to patient prognosis. Therefore, clinicopathologic evaluation of the primary tumor and metastatic sites could be helpful for predicting the prognosis.

Do neuroendocrine carcinomas and mixed neuroendocrine-non-neuroendocrine neoplasm of the gastrointestinal tract have the same prognosis? A SEER database analysis of 12,878 cases.

BACKGROUND: Neuroendocrine carcinomas (NECs) and mixed neuroendocrine-non-neuroendocrine neoplasm (MiNEN) in the gastrointestinal (GI) tract are both rare and malignant; however, it is unclear whether their prognosis is the same. METHODS: In this cross-sectional study, a total of 12,878 patients with NEC or MiNEN in the GI tract were reviewed retrospectively by searching the Surveillance, Epidemiology, and End Results (SEER) program database. Next, we compared the characteristics and survival between patients with NEC or MiNEN and further analyzed the prognostic factors for the patients. RESULTS: The data showed that patients with MiNEN had a worse prognosis as compared with patients with pure NEC in the small intestine (SI) and appendix, whereas there was no significant survival difference between NEC and MiNEN in the other parts of the GI system. On the whole, age ⩾55 years (p < 0.0001), male (p = 0.002), being diagnosed at TNM Stage II-IV (p < 0.0001) or not receiving surgical treatment (p < 0.0001) were the independent negative prognostic factors for NEC patients, whereas age ⩾55 years (p = 0.003), being diagnosed at TNM Stage III-IV (p < 0.001) or not receiving surgical treatment (p < 0.001) were identified as the independent negative prognostic factors for the MiNEN patients. Furthermore, when NECs or MiNENs were classified based on the primary tumor site, the results showed that the prognostic factors for NEC and MiNEN varied between the tumor sites. CONCLUSION: The prognostic differences between NECs and MiNENs in the GI tract are heterogeneous and site-related. Patients with appendiceal or SI MiNEN have a poorer prognosis than patients with pure appendiceal or SI NEC. Therefore, we should pay more attention to patients with MiNEN in the SI and appendix and monitor them more closely.