Proteomic Characterization Identifies Clinically Relevant Subgroups of Gastrointestinal Stromal Tumors.

BACKGROUND & AIMS: Gastrointestinal stromal tumor (GIST) is the most common mesenchymal tumor of the gastrointestinal tract, and it has high metastatic and recurrence rates. We aimed to characterize the proteomic features of GIST to understand biological processes and treatment vulnerabilities. METHODS: Quantitative proteomics and phosphoproteomics analyses were performed on 193 patients with GIST to reveal the biological characteristics of GIST. Data-driven hypotheses were tested by performing functional experiments using both GIST cell lines and xenograft mouse models. RESULTS: Proteomic analysis revealed differences in the molecular features of GISTs from different locations or with different histological grades. MAPK7 was identified and functionally proved to be associated with tumor cell proliferation in GIST. Integrative analysis revealed that increased SQSTM1 expression inhibited the patient response to imatinib mesylate. Proteomics subtyping identified 4 clusters of tumors with different clinical and molecular attributes. Functional experiments confirmed the role of SRSF3 in promoting tumor cell proliferation and leading to poor prognosis. CONCLUSIONS: Our study provides a valuable data resource and highlights potential therapeutic approaches for GIST.

Loratadine as an Anti-inflammatory Agent Against Clostridium difficile Toxin B.

BACKGROUND: Clostridium difficile infection (CDI) is a debilitating nosocomial infection. C. difficile produces toxins A and B, which cause inflammation. Existing therapies have issues with recurrence, cost, and safety. We aim to discover a safe, effective, and economical nonmicrobiological therapeutic approach against CDI. METHODS: We included human primary peripheral blood mononuclear cells (PBMCs), fresh human colonic explants, and humanized HuCD34-NCG mice. Toxin A+B+ VPI 10463 and A-B+ ribotype 017 C. difficile strains were used. We used single-cell RNA profiling and high-throughput screening to find actionable toxin B-dependent pathways in PBMCs. RESULTS: Histamine 1 receptor-related drugs were found among the hit compounds that reversed toxin-mediated macrophage inflammatory protein (MIP) 1α expression in PBMCs. We identified loratadine as the safest representative antihistamine for therapeutic development. Loratadine inhibited toxin B-induced MIP-1α secretion in fresh human colonic tissues. Oral loratadine (10 mg/kg/d) maintained survival, inhibited intestinal CCl3 messenger RNA expression, and prevented vancomycin-associated recurrence in the VPI 10463-infected mice and ribotype 017-infected hamsters. Splenocytes from loratadine-treated mice conferred anti-inflammatory effects to the VPI 10463-infected T/B-cell--deficient Rag-/- mice. Oral loratadine suppressed human MIP-1α expression in monocytes/macrophages in toxin B-expressing ribotype 017-infected humanized HuCD34-NCG mice. CONCLUSIONS: Loratadine may be repurposed to optimize existing therapies against CDI.

Loratadine, a Food and Drug Administration­approved antihistamine, inhibits toxin B­mediated proinflammatory macrophage inflammatory protein 1α secretion from immune cells. Its anti-inflammatory effect ameliorates intestinal inflammation in Clostridium difficile­infected animals. Loratadine may be repurposed to optimize existing therapies.

Fusobacterium periodonticum BCT protein targeting glucose metabolism to promote the epithelial-mesenchymal transition of esophageal cancer cells by lactic acid.

BACKGROUND: The cancer microbiota was considered the main risk factor for cancer progression. We had proved that Fusobacterium periodonticum (F.p) was higher abundance in Esophageal cancer(EC)tissues. Bioinformation analysis found that BCT was a key virulence protein of F.p. However, little is known about the role and mechanism of BCT in EC. This study aimed to recognize the key virulence protein of F.p and explore the mechanism of BCT in promoting EC. METHODS: We constructed a eukaryotic expression vector and purified the recombinant protein BCT. CCK8 used to analyzed the activity of EC after treated by different concentration of BCT. UPLC-MS/MS and ELISA used to detect the metabonomics and metabolites. The ability of migration and invasion was completed by transwell assay. RT-QPCR, WB used to analyze the expression of relevant genes. RESULTS: Our data showed that BCT was higher expression in EC tumor tissues (p < 0.05) and BCT in 20 µg/mL promoted the survival, invasion and migration of EC cells (EC109) (p < 0.05). Meanwhile, UPLC-MS/MS results suggested that BCT resulted in an augmentation of hypotaurine metabolism, arachidonic acid metabolism, glycolysis/gluconeogenesis, tryptophan metabolism, citrate cycle activity in EC109. The metabolic changes resulted in decreasing in glucose and pyruvate levels but increase in lactate dehydrogenase (LDH) activity and lactic acid (LA) as well as the expression of glucose transporter 1, Hexokinase 2, LDH which regulated the glycolysis were all changed (p < 0.05). The BCT treatment upregulated the expression of TLR4, Akt, HIF-1α (p < 0.05) which regulated the production of LA. Furthermore, LA stimulation promoted the expression of GPR81, Wnt, and ß-catenin (p < 0.05), thereby inducing EMT and metastasis in EC109 cells. CONCLUSION: Altogether, these findings identified that impact of BCT in regulation of glycolysis in EC109 and its involves the TLR4/Akt/HIF-1α pathway. Meanwhile, glycolysis increasing the release of LA and promote the EMT of EC109 by GPR81/Wnt/ß-catenin signaling pathway. In summary, our findings underscore the potential of targeting BCT as an innovative strategy to mitigate the development of EC.

Role of blood lipids in mediating the effect of dietary factors on gastroesophageal reflux disease: a two-step mendelian randomization study.

BACKGROUND: Growing studies have indicated an association between dietary factors and gastroesophageal reflux disease (GERD). However, whether these associations refer to a causal relationship and the potential mechanism by which dietary factors affect GERD is still unclear. METHODS: A two-step mendelian randomization analysis was performed to obtain causal estimates of dietary factors, blood lipids on GERD. Independent genetic variants associated with 13 kinds of dietary factors and 5 kinds of blood lipids at the genome-wide significance level were selected as instrumental variables. The summary statistics for GERD were obtained from European Bioinformatics Institute, including 129,080 cases and 473,524 controls. Inverse variance weighted was utilized as the main statistical method. MR-Egger intercept test, Cochran's Q test, and leave-one-out analysis were performed to evaluate possible heterogeneity and pleiotropy. And the potential reverse causality was assessed using Steiger filtering. RESULTS: The results of the inverse variance weighted method indicated that genetically predicted total pork intake (OR = 2.60, 95% CI: 1.21-5.58, p = 0.0143), total bread intake (OR = 0.68, 95% CI: 0.46-0.99, p = 0.0497), total cereal intake (OR = 0.42, 95% CI: 0.31-0.56, p = 2.98E-06), and total cheese intake (OR = 0.41, 95% CI: 0.27-0.61, p = 1.06E-05) were associated with the risk of GERD. Multivariable Mendelian randomization analysis also revealed a negative association between total cereal intake, total cheese intake and the risk of GERD, but the effect of total pork intake and total bread intake on GERD disappeared after adjustment of smoking, alcohol consumption, use of calcium channel blockers, BMI, physical activity levels, and biological sex (age adjusted). Furthermore, the concentration of low-density lipoprotein cholesterol (LDL-C) is negatively correlated with total cheese intake, which mediates the impact of total cheese intake on GERD. The proportion mediated by LDL-C is 2.27% (95%CI: 1.57%, 4.09%). CONCLUSIONS: This study provides evidence that an increase in total cereal intake and total cheese intake will decrease the risk of GERD. Additionally, LDL-C mediates the causal effect of total cheese intake on GERD. These results provide new insights into the role of dietary factors and blood lipids in GERD, which is beneficial for disease prevention.

Causal relationships between coffee intake, apolipoprotein B and gastric, colorectal, and esophageal cancers: univariable and multivariable Mendelian randomization.

PURPOSE: Coffee intake and apolipoprotein B levels have been linked to gastric, colorectal, and esophageal cancers in numerous recent studies. However, whether these associations are all causal remains unestablished. This study aimed to assess the potential causal associations of apolipoprotein B and coffee intake with the risk of gastric, colorectal, and esophageal cancers using Mendelian randomization analysis. METHODS: In this study, we utilized a two-sample Mendelian randomization analysis to access the causal effects of coffee intake and apolipoprotein B on gastric, colorectal, and esophageal cancers. The summary statistics of coffee intake (n = 428,860) and apolipoprotein B (n = 439,214) were obtained from the UK Biobank. In addition, the summary statistics of gastric cancer, colorectal cancer, and esophageal cancer were obtained from the FinnGen biobank (n = 218,792). Inverse variance weighted, MR-Egger, weighted median, and weighted mode were applied to examine the causal relationship between coffee intake, apolipoprotein B and gastric, colorectal, and esophageal cancers. MR-Egger intercept test, Cochran's Q test, and leave-one-out analysis were performed to evaluate possible heterogeneity and pleiotropy. Steiger filtering and bidirectional mendelian randomization analysis were performed to evaluate the possible reverse causality. RESULTS: The result of the inverse variance weighted method indicated that apolipoprotein B levels were significantly associated with a higher risk of gastric cancer (OR = 1.392, 95% CI 1.027-1.889, P = 0.0333) and colorectal cancer (OR = 1.188, 95% CI 1.001-1.411, P = 0.0491). Furthermore, multivariable Mendelian randomization analysis also revealed a positive association between apolipoprotein B levels and colorectal cancer risk, but the effect of apolipoprotein B on gastric cancer risk disappeared after adjustment of coffee intake, body mass index or lipid-related traits. However, we did not discover any conclusive evidence linking coffee intake to gastric, colorectal, or esophageal cancers. CONCLUSIONS: This study suggested a causal association between genetically increased apolipoprotein B levels and higher risk of colorectal cancer. No causal relationship was observed between coffee intake and gastric, colorectal, or esophageal cancers.

The effect of weak π-π interactions on single-molecule electron transport properties of the tetraphenylethene molecule and its derivatives: a first-principles study.

Intramolecular π-π interactions are a significant research focus in fields such as chemistry, biology, and materials science. Different configurations of benzene-benzene moieties within a molecule can affect the magnitude of their π-π interactions, consequently influencing the electronic transport capabilities of the molecule. In this study, we designed three π-conjugated molecules, TPEM, TPEEM, and TEEPM, based on tetraphenylethene (TPE). These three molecules exhibit three distinct π-conjugated structures: linear cis-π-conjugation, linear trans-π-conjugation, and cross-π-conjugation. Thereinto, TPEM and TPEEM molecules share the same TPE core, with identical π-π interaction distances, while the TEEPM molecule has acetylene groups between the TPE units, thereby increasing the π-π interaction distances between the benzene moieties. Using density functional theory calculations combined with non-equilibrium Green's function (DFT+NEGF), our results reveal that the conductance order of different π-conjugated structures in TPEM and TPEEM molecules is as follows: cis > cross ≈ trans. Through analysis of transmission spectra, transmission pathways, and the innermost π orbitals, we find that in TPEM and TPEEM molecules, the cis- and cross-π-conjugated structures exhibit π-π interactions between benzene moieties and provide special through-space electron transport pathways, enhancing their electronic transport capabilities in coordination with the bonded molecular framework, whereas their trans-conjugated structures only allow electron transport along the molecular backbone. In contrast, in TEEPM molecule, due to the absence of π-π interactions, the conductance of different π-conjugated structures is primarily determined by the molecular backbone and follows the order: trans > cis > cross. These findings provide a theoretical basis for designing single-molecule electronic devices with multiple electron channels based on intramolecular π-π interactions.

Association between monocyte-to-high-density lipoprotein-cholesterol ratio and gallstones in U.S. adults: findings from the National Health and Nutrition Examination Survey 2017-2020.

BACKGROUND: Studies have indicated that monocyte-to-high-density lipoprotein cholesterol ratio (MHR) can be a reliable indicator of various diseases. However, the association between MHR and gallstone prevalence remains unclear. Therefore, this study aimed to explore any potential association between MHR and gallstone prevalence. METHODS: This study used data from the National Health and Nutrition Examination Survey (NHANES) 2017-March 2020. MHR was calculated as the monocyte count ratio to high-density lipoprotein cholesterol levels. Multiple logistic regression models, Cochran-Armitage trend test, and subgroup analyses were used to examine the association between MHR and gallstones. RESULTS: This study included 5907 participants, of whom 636 (10.77%) were gallstone formers. The study participants had a mean age of 50.78 ± 17.33 years. After accounting for multiple covariables, the multiple logistic regression model showed a positive linear association between MHR and gallstone odds. The subgroup analyses and interaction testing results revealed that the association between MHR and gallstones was statistically different across strata, including sex, smoking, asthma, and hypertension. CONCLUSIONS: Gallstone prevalence positively associated with elevated MHR, indicating that MHR can be employed as a clinical indicator to assess gallstone prevalence.

Increasing CRISPR/Cas9-mediated gene editing efficiency in T7 phage by reducing the escape rate based on insight into the survival mechanism.

Bacteriophages have been used across various fields, and the utilization of CRISPR/Cas-based genome editing technology can accelerate the research and applications of bacteriophages. However, some bacteriophages can escape from the cleavage of Cas protein, such as Cas9, and decrease the efficiency of genome editing. This study focuses on the bacteriophage T7, which is widely utilized but whose mechanism of evading the cleavage of CRISPR/Cas9 has not been elucidated. First, we test the escape rates of T7 phage at different cleavage sites, ranging from 10 -2 to 10 -5. The sequencing results show that DNA point mutations and microhomology-mediated end joining (MMEJ) at the target sites are the main causes. Next, we indicate the existence of the hotspot DNA region of MMEJ and successfully reduce MMEJ events by designing targeted sites that bypass the hotspot DNA region. Moreover, we also knock out the ATP-dependent DNA ligase 1. 3 gene, which may be involved in the MMEJ event, and the frequency of MMEJ at 4. 3 is reduced from 83% to 18%. Finally, the genome editing efficiency in T7 Δ 1. 3 increases from 20% to 100%. This study reveals the mechanism of T7 phage evasion from the cleavage of CRISPR/Cas9 and demonstrates that the special design of editing sites or the deletion of key gene 1. 3 can reduce MMEJ events and enhance gene editing efficiency. These findings will contribute to advancing CRISPR/Cas-based tools for efficient genome editing in phages and provide a theoretical foundation for the broader application of phages.

Hsa_circ_0001707 regulates endothelial-mesenchymal transition in esophageal squamous cell carcinoma via miR-203a-3p/Snail2 pathway.

Esophageal squamous cell carcinoma (ESCC) is a malignant tumor with high mortality and poor prognosis. Despite intensive research focused on tumor suppression, the 5-year survival rate of ESCC is lower than 15%. Therefore, investigate fundamental mechanisms involved in ESCC is on-demand crucial for diagnostics and developing targeted therapeutic drugs. Circular RNAs (circRNAs), as an emerging class of non-coding RNA, have been elucidated that circRNAs participated in regulating a variety of pathological processes and tumorigenesis. Nevertheless, the functional role of circRNAs in the occurrence and development of ESCC remains unclear. We identify a novel circRNA (hsa_circ_0001707), which was highly expressed in ESCC patients' tissues and cell lines. Furthermore, gain- and loss-of-function assays were performed and found that overexpression of hsa_circ_0001707 significantly promote tumor proliferation, metastasis, and invasion. By functioning as a competing endogenous RNA (ceRNA), the dual-luciferase activity assay verified that hsa_circ_0001707 can endogenously bind with miR-203a-3p and regulate its downstream gene Snail2. Rescue assay further confirms that hsa_circ_0001707 downregulation could partially attenuate the facilitation effect of miR-203a-3p, thereby inhibiting the endothelial-mesenchymal transition (EMT) process of ESCC. Our results suggested that hsa_circ_0001707 play an oncogenic role in the pathogenesis of ESCC, which might be a potential biomarker for diagnostics and targeting therapy.

Genistein Inhibits Clostridioides difficile Infection via Estrogen Receptors and Lysine-Deficient Protein Kinase 1.

BACKGROUND: Clostridioides difficile infection (CDI) is a debilitating nosocomial disease. Postmenopausal women may have an increased risk of CDI, suggesting estrogen influence. Soybean products contain a representative estrogenic isoflavone, genistein. METHODS: The anti-inflammatory and antiapoptotic effects of genistein were determined using primary human cells and fresh colonic tissues. The effects of oral genistein therapy among mice and hamsters were evaluated. RESULTS: Within 10 days of CDI, female c57BL/6J mice in a standard environment (regular diet) had a 50% survival rate, while those with estrogen depletion and in an isoflavone-free environment (soy-free diet) had a 25% survival rate. Oral genistein improved their 10-day survival rate to 100% on a regular diet and 75% in an isoflavone-free environment. Genistein reduced macrophage inflammatory protein-1α (MIP-1α) secretion in fresh human colonic tissues exposed to toxins. Genistein inhibited MIP-1α secretion in primary human peripheral blood mononuclear cells, abolished apoptosis and BCL-2-associated X (BAX) expression in human colonic epithelial cells, and activated lysine-deficient protein kinase 1 (WNK1) phosphorylation in both cell types. The anti-inflammatory and antiapoptotic effects of genistein were abolished by inhibiting estrogen receptors and WNK1. CONCLUSIONS: Genistein reduces CDI disease activity by inhibiting proinflammatory cytokine expression and apoptosis via the estrogen receptor/G-protein estrogen receptor/WNK1 pathways.