The role of soluble epoxide hydrolase in the intestine.

The soluble epoxide hydrolase (sEH; encoded by the EPHX2 gene) is an α/ß hydrolase fold protein that is, widely distributed throughout the body. Recent studies have highlighted that sEH, in the metabolism of polyunsaturated fatty acids, plays a part in the pathogenesis of various diseases, including cardiovascular disease, Alzheimer's disease and intestine-associated disease. This review discusses the current findings on the role of sEH in the development of intestine- and intestine-associated diseases, including colitis, colorectal cancer, and other intestinal diseases, as well as the potential underlying mechanisms involved.

Structure Composition and Intracellular Transport of Clathrin-Mediated Intestinal Transmembrane Tight Junction Protein.

Tight junctions (TJs) are located in the apical region of the junctions between epithelial cells and are widely found in organs such as the brain, retina, intestinal epithelium, and endothelial system. As a mechanical barrier of the intestinal mucosa, TJs can not only maintain the integrity of intestinal epithelial cells but also maintain intestinal mucosal permeability by regulating the entry of ions and molecules into paracellular channels. Therefore, the formation disorder or integrity destruction of TJs can induce damage to the intestinal epithelial barrier, ultimately leading to the occurrence of various gastrointestinal diseases, such as inflammatory bowel disease (IBD), gastroesophageal reflux disease (GERD), and irritable bowel syndrome (IBS). However, a large number of studies have shown that TJs protein transport disorder from the endoplasmic reticulum to the apical membrane can lead to TJs formation disorder, in addition to disruption of TJs integrity caused by external pathological factors and reduction of TJs protein synthesis. In this review, we focus on the structural composition of TJs, the formation of clathrin-coated vesicles containing transmembrane TJs from the Golgi apparatus, and the transport process from the Golgi apparatus to the plasma membrane via microtubules and finally fusion with the plasma membrane. At present, the mechanism of the intracellular transport of TJ proteins remains unclear. More studies are needed in the future to focus on the sorting of TJs protein vesicles, regulation of transport processes, and recycling of TJ proteins, etc.

Identification and verification of YBX3 and its regulatory gene HEIH as an oncogenic system: A multidimensional analysis in colon cancer.

The novel gene YBX3 is important for regulating translation and RNA catabolism and encodes a protein with a highly conserved cold-shock domain. However, its pathogenic roles across cancers (e.g., colon cancer) and its regulation remain unclear. We identified the pathogenic roles of YBX3 and its regulatory lncRNA HEIH in various cancers and investigated their effects on tumor progression in colon cancer. Methods including RNA pull-down, MS, and TMA of 93 patients, qPCR of 12 patients with diverse clinicopathologic stages, and western blotting were performed. The pancancer analysis showed that YBX3 expression varies significantly among not only cancer types but also molecular and immune subtypes of the same cancer. Furthermore, its expression in colon cancer is clinically significant, and there is an obvious negative regulatory association between HEIH and YBX3. Among various cancers, especially colon cancer, YBX3 is more related than HEIH expression to the clinical features and prognosis of subgroups. The receiver operating characteristic analysis showed that HEIH and YBX3 have similar predictive capacity in various cancers. The analysis of differentially expressed genes in colon cancer revealed that they have similar hub gene networks, indicating an oncogenic system with a strong overlap. The results also suggest that YBX3 is associated with tumor immune evasion via different mechanisms involving T-cell exclusion in different cancer types and by the tumor infiltration of immune cells. Interestingly, scRNA-seq revealed that HEIH inhibits this phenomenon. Our results also suggest that YBX3 expression is associated with immune or chemotherapeutic outcomes in various cancers, and YBX3 exhibited a higher predictive power than two of seven standardized biomarkers for response outcomes and overall survival of immune checkpoint blockade subcohorts. In colon cancer cell lines, lncRNA-HEIH and YBX3 associate. MS confirmed that YBX3 was pulled down with HEIH, and western blot showed that HEIH knockdown disinhibited YBX3. This study strongly suggests that lncRNA-HEIH/YBX3 is a pancancer immune-oncogenic system and could serve as a biomarker for diagnosis and prognosis and as a therapeutic target, especially in colon cancer.

Feasibility and preliminary experience of single-incision plus one-port laparoscopic total gastrectomy with Overlap esophagojejunostomy for gastric cancer: A study of 10 cases.

Background: This study aimed to explore the feasibility and safety of single-incision plus one-port laparoscopic total gastrectomy (SITG + 1) with Overlap esophagojejunostomy (SITG + 1-Overlap) and to share preliminary experiences. Methods: This retrospective study included 10 patients with gastric cancer located in the cardia or body who underwent SITG + 1-Overlap between August 2020 and October 2021.The demographics, tumor characteristics, postoperative outcomes, and short-term complications of all the enrolled patients were summarized and statistically analyzed. Data were expressed as mean ± standard deviation (SD) if they were normally distributed. Otherwise, Median (Quartile1, Quartile3) was used. Results: In the collective perioperative data of these 10 patients who underwent radical gastrectomy, the median of the length of transumbilical incision and blood loss were 3.0 cm and 100.0 ml respectively, and the mean operation time and 385.5 ± 51.6 min. Postoperative data indicated that the gastric tube was removed on 2.0 (2.0, 3.0) days, and the timing of first feeding, activity, flatus, and defecation was 1.5 (1.0, 2.0) days, 2.0 (2.0, 2.0) days, 3.0 (2.0, 3.0) days, and 3.8 ± 0.6 days, respectively. The timing of drainage tube removal was 4.6 ± 1.0 days after operation. The duration of hospital stay was 7.5 ± 1.2 days and the VAS pain scores for the 3 days following surgery were 3.0 (2.0, 3.3), 2.0 (2.0, 3.0), and 1.5 (1.0, 2.0) respectively. The mean number of retrieved lymph nodes was 30.7 ± 13.2. Most biochemical indicators gradually normalized with the recovery of the patients after surgery. No 30-day postoperative complications were noted. Conclusions: For the first time, our preliminary data indicate the feasibility and safety of Overlap esophagojejunostomy in SITG + 1 surgery. This modified Overlap procedure has the potential to simplify the reconstruction procedure and lower the technical challenge of SITG + 1 radical gastrectomy for cardia or upper gastric cancer in the early and advanced stages.

Single-incision plus one-port laparoscopic gastrectomy versus conventional multi-port laparoscopy-assisted gastrectomy for gastric cancer: a retrospective study.

BACKGROUND: We compared short-term perioperative outcomes after single-incision plus one-port laparoscopic gastrectomy (SILG+1) and conventional multi-port laparoscopy-assisted gastrectomy (C-LAG) for gastric cancer. METHODS: The work was conducted between August 2017 and October 2019. A total of 90 patients with early or advanced gastric cancer were retrospectively analyzed: 43 patients of which underwent SILG+1, and 47 of which underwent C-LAG, respectively. These were divided into two groups: the total gastrectomy group (SILT+1 and C-LATG) and the distal gastrectomy group (SILD + 1 and C-LADG). The demographics, tumor characteristics, postoperative outcomes, and short-term complications of all enrolled patients were summarized and statistically analyzed. RESULTS: The mean incision length in SILT+1 group was 5.40 cm shorter than that in C-LATG group (3.15 ± 0.43 vs. 8.55 ± 2.72, P < 0.001). This comparison between the SILD + 1 and the C-LADG group produced comparable results. The SILT+1 group underwent a 56.32 min longer operation than the C-LATG group (273.03 ± 66.80 vs. 216.71 ± 82.61, P = 0.0205). SILG+1 group had better postoperative visual analog scale (VAS) and cosmetic score than those of the C-LATG group (P < 0.05). There were no significant differences in preoperative demographics or 30-day postoperative complication rates between the SILG+1 and C-LAG groups. Tumor-related index, including mass size, histological type, number of retrieved lymph nodes, pathological tumor-node-metastasis (TNM) stage, and proximal and distal edges were all equivalent between the SILG+1 and the C-LAG group. CONCLUSIONS: This retrospective study demonstrates the safety and feasibility of SILG+1 with D1+ or D2 lymphadenectomy for the treatment of early and advanced gastric cancers, compared with C-LAG.

SCFAs induce autophagy in intestinal epithelial cells and relieve colitis by stabilizing HIF-1α.

Hypoxia-inducible factor-1α (HIF-1α) is a critical regulator of barrier integrity during colonic mucosal injury. Previous works have shown that the absence of autophagy is implicated in the development of inflammatory bowel disease (IBD). Additionally, changes in bacterial profiles in the gut are intimately associated with IBD. Although HIF-1α, autophagy, microbiota, and their metabolites are all involved in the pathogenesis of IBD, their roles are not known. In this study, we investigated the relationship between HIF-1α and autophagy in healthy and inflammatory states using transgenic mice, colitis models, and cell culture models. We confirmed that the absence of intestinal epithelial HIF-1α changed the composition of the intestinal microbes and increased the susceptibility of mice to dextran sodium sulfate (DSS)-induced colitis. In addition, autophagy levels in the intestinal epithelial cells (IECs) were significantly reduced in IEC-specific HIF-1α-deficient (HIF-1α∆IEC) mice. Moreover, in the cell culture models, butyrate treatment significantly increased autophagy in HT29 cells under normal conditions, whereas butyrate had little effect on autophagy after HIF-1α ablation. Furthermore, in the DSS-induced colitis model, butyrate administration relieved the colonic injury and suppressed inflammation in Cre-/HIF-1α- (HIF-1αloxP/loxP) mice. However, the butyrate-mediated protection against colonic injury was considerably diminished in the HIF-1α∆IEC mice. These results show that HIF-1α, autophagy, and intestinal microbes are essential for the maintenance of intestinal homeostasis. Butyrate can alleviate DSS-induced colitis by regulating autophagy via HIF-1α. These insights may have important implications for the development of therapeutic strategies for IBD. KEY MESSAGES: • The absence of intestinal epithelial HIF-1α leads to downregulation of autophagy in mice. • The absence of intestinal epithelial HIF-1α exacerbates DSS-induced colitis. • Short-chain fatty acids (SCFAs) can alleviate DSS-induced colitis by regulating autophagy via HIF-1α.

Mutual regulation between butyrate and hypoxia-inducible factor-1α in epithelial cell promotes expression of tight junction proteins.

Inflammatory bowel disease is a kind of multi-aetiological chronic disease that is driven by multidimensional factors. Hypoxia-inducible factor-1α (HIF-1α) plays an important role in anti-inflammatory and cellular responses to hypoxia. Previous studies have found that B or T-cell-specific HIF-1α knock out mice exhibit severe colonic inflammation. However, we know very little about other functions of HIF-1α in intestinal epithelial cells (IECs). In our study, HIF-1αΔIEC mice were used to study the function of HIF-1α in IECs. HIF-1α was knocked down in Caco-2 cells by transfection with a small interfering (si) RNA. Immunohistochemical staining and western blotting were used to detect the expression of zonula occluden-1 (ZO-1) and Occludin. The content of colon was harvested for high-performance liquid chromatography analysis to examine the levels of butyrate in the gut. Our research found that HIF-1α played a protective role in dextran sulphate sodium-induced colitis, which was partly due to its regulation of tight junction (TJ) protein expression. Further study revealed that HIF-1α mediated TJ proteins levels by moderating the content of butyrate. Moreover, we found that butyrate regulated TJ protein expression, which is dependent on HIF-1α. These results indicated that there is a mutual regulatory mechanism between butyrate and HIF-1α, which has an important role in the maintenance of barrier function of the gastrointestinal tract.

KGF inhibits hypoxia-induced intestinal epithelial cell apoptosis by upregulating AKT/ERK pathway-dependent E-cadherin expression.

OBJECTIVE: Intestinal ischemia-reperfusion (I/R) causes direct cellular damage, and the potential injury to the mucosal structure and barrier function. Keratinocyte growth factor (KGF) is highly expressed in gastrointestinal tract and exerts beneficial effects for intestinal epithelial growth and maintenance. E-cadherin plays an important role in intestinal epithelium renewal. However, the regulatory role of KGF on E-cadherin levels and I/R-induced apoptosis remain to be explored. The present study aimed to identify the effect of KGF on E-cadherin expression and I/R-induced intestinal epithelial cell apoptosis. METHODS: Caco2 cells were treated with KGF (100 ng/ml) for 0, 4, 8, 12, and 24 h under hypoxia or normoxia. An E-cadherin-knockdown model was successfully established by treatment with E-cadherin RNAi. Western blotting and immunofluorescence labeling were performed to assess E-cadherin expression. Levels of PI3K|[sol]|Akt/mitogen-activated protein kinases (MAPKs), phosphoinositide 3-kinase (PI3K|[sol]|Akt)/PI3K|[sol]|Akt pathway-related proteins, and apoptosis-related proteins were also detected by western blot. Finally, a rat model of acute intestinal I/R was established and treated with KGF. Hematoxylin-eosin (HE), terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL), and immunofluorescence staining were performed to detect morphological changes in intestinal mucosal epithelium and Caco2 cell apoptosis. RESULTS: KGF enhanced E-cadherin expression in differentiated intestinal epithelial cells under hypoxia via AKT/extracellular-regulated kinase (ERK) pathway regulation. In vitro, E-cadherin downregulation aggravates hypoxia-induced intestinal epithelial cell apoptosis. In the rat model, KGF increased E-cadherin expression, which was associated with the reduced apoptosis. CONCLUSIONS: KGF exerts protective effects on intestinal epithelial cells under hypoxia by elevating E-cadherin levels or activating AKT/ERK signaling.

AhR­E2F1­KGFR signaling is involved in KGF­induced intestinal epithelial cell proliferation.

Keratinocyte growth factor (KGF) stimulates intestinal epithelial cell proliferation upon binding to the KGF receptor (KGFR). The activated aryl hydrocarbon receptor (AhR) serves an important role in the development of tissues by promoting the expression of AhR receptors, which can regulate cell proliferation. In the present study, the signaling pathway between AhR and KGFR in investigated with regards to KGF­induced intestinal epithelial cell proliferation. Male C57BL/6J wild type and AhR­/­ mice, were randomized into four groups: Control, KGF, AhR­/­ + KGF and AhR­/­ (n=6 per group). The small bowel was harvested on day 5 post­treatment. LoVo cells were used to study signaling pathways in vitro and were divided into the following four treatment groups: DMSO, KGF, KGF + small­interfering (si)AhR and siAhR. In vivo, knockdown of AhR mRNA transcripts may abolish KGF­induced intestinal epithelial cell proliferation. Furthermore, KGFR expression was downregulated following knockdown or silencing of AhR expression in vivo and in vitro. The present study identified that the transcription factor E2F1 could regulate KGFR expression, and that siAhR treatment led to reduced expression of E2F1 in the nucleus and inhibited KGF­induced cell proliferation. In conclusion, the current results demonstrated that the AhR­E2F1­KGFR pathway is involved in KGF­induced intestinal epithelial cell proliferation.

The AhR is involved in the regulation of LoVo cell proliferation through cell cycle-associated proteins.

Some ingredients in foods can activate the aryl hydrocarbon receptor (AhR) and arrest cell proliferation. In this study, we hypothesized that 6-formylindolo [3, 2-b] carbazole (FICZ) arrests the cell cycle in LoVo cells (a colon cancer line) through the AhR. The AhR agonist FICZ and the AhR antagonist CH223191 were used to treat LoVo cells. Real-time PCR and Western blot analyses were performed to detect the expression of the AhR, CYP1A1, CDK4, cyclinD1, cyclin E, CDK2, P27, and pRb. The distribution and activation of the AhR were detected with immunofluorescence. A 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and flow cytometric analysis were performed to measure cell viability, cell cycle stage, and apoptosis. Our results show that FICZ inhibited LoVo cell proliferation by inducing G1 cell cycle arrest but had no effect on epithelial apoptosis. Further analysis found that FICZ downregulated cyclinD1 and upregulated p27 expression to arrest Rb phosphorylation. The downregulation of cyclinD1 and upregulation of p27 were abolished by co-treatment with CH223191. We conclude that the AhR, when activated by FICZ (an endogenous AhR ligand), can arrest the cell cycle and block LoVo cell proliferation.

Keratinocyte Growth Factor Regulation of Aryl Hydrocarbon Receptor Activation in Colorectal Cancer Cells.

BACKGROUND: Keratinocyte growth factor (KGF) stimulates normal growth, development and intestinal epithelial cell proliferation. Cyclin D1 promotes the cell cycle by inhibiting retinoblastoma protein (RB1). The activated aryl hydrocarbon receptor (AhR) has an important influence on the development of tumors through its interactions with the cell cycle. AIM: The aim of the present study was to explore a new role for AhR in KGF-induced colon cancer cell growth. MATERIALS AND METHODS: Real-time PCR, western blot or immunofluorescence analysis were used to detect the expression of KGF, AhR, cyclin D1 and CYP1A1. Immunohistochemistry was used to observe the localization of AhR. MTT assay and flow cytometric analyses were performed to measure cell viability and the cell cycle. RESULTS: Real-time PCR analysis revealed that KGF, AhR, and CYP1A1 mRNAs were overexpressed in colorectal cancer tissues. Meanwhile, overexpression of AhR was primarily observed in epithelial cells. In in vitro assay, KGF promoted colon cancer cell growth, as well as up-regulated and activated AhR. At the same time, AhR-knockdown colon cancer cells were less responsive to KGF. Western blot analysis, real-time PCR, or immunofluorescence data indicated that cyclin D1 expression was up-regulated by KGF but this up-regulation was compromised when AhR was silenced, and the cell cycle was arrested in the G0/G1 stage in these cells. CONCLUSIONS: Our study suggests that KGF, AhR, and CYP1A1 are overexpressed in colorectal cancer tissues. Moreover, we reveal a new mechanism by which KGF promotes cell proliferation through the AhR-cyclin D1 pathway in colon cancer cells.