Bile salts inhibit growth and induce apoptosis of culture human normal esophageal mucosal epithelial cells

AIM： To investigate the effect of six bile salts： glycocholate （GC）, glycochenodeoxycholate （GCDC）, glycodeoxycholate （GDC）, taurocholate （TC）, taurochenodeoxycholate （TCDC）, taurodeoxycholate （TDC）, and their mixture on cultured human normal esophageal rnucosal epithelial cells. METHODS： Human normal esophageal mucosal epithelial cells were cultured with serum-free keratinocyte medium. 3-[4,5-Dimethylthiaolyl]-2,5- diphenyl-tetrazolium bromide assay was applied to the detection of cell proliferation. Apoptotic morphology was observed by phase-contrast video microscopy and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling （TUNEL） assay. Sub-G1 DNA fragmentations and early apoptotic cells were assayed by flow cytometry （FCI） with propidium iodide （PI） staining and annexin V-FITC conjugated with PI staining. Apoptotic DNA ladders on agarose gel electrophoresis were observed. RESULTS： Except for GC, GCDC, GDC, TC, TCDC, TDC and their mixture could initiate growth inhibition of esophageal mucosal epithelial cells in a dose- and time-dependent manner. TUNEL and FCM assays demonstrated that the bile salts at 500 μmol/L and their mixture at 1 500 μmol/L induced apoptosis except for GC. The percentage of sub-G1 detected by FCM with PI staining was 83.5% in cells treated with 500 μmol/L TC for 2 h, and 19.8%, 20.4%, 25.6%, 13.5%, and 75.8% in cells treated with 500 μmol/L GCDC, TCDC, GDC, TDC, and 1 500 μmol/L mixture for 24 h, respectively, which were higher than that of the control （1.5%）. The percentage was 1.4% in cells with 500 μmol/L GC for 24 h. DNA ladders on agarose gel electrophoresis were seen in cells treated with 500 μmol/L TC for 2 h and i 500 μmnol/L mixture for 24 h. CONCLUSION： All GCDC, GDC, TC, TCDC, TDC and their mixture can inhibit growth and induce apoptosis of cultured human normal esophageal mucosal epithelial cells, but GC is well tolerated by the cells.

Effect of bile salts and bile acids on human gastric mucosal epithelial cells

Objective：To explore the effect of bile salt and bile acid on cultured eternalized human gastric mucosa epithelium GES-1 cells. Methods：Cultured eternalized human gastric mucosa epithelium GES-1 cells were treated with media containing 6 different kinds of bile salts and 3 different kinds of bile acids and their mixture with different concentrations： GCDC（glycochenodeoxychoμte）, GDC （glycodeoxychoμte）, GC（glycochoμte）, TCDC（taurochenodeoxychoμte）, TDC（taurodeoxychoμte）, TC （taurochoμte）, LCA （lithocholicacid）, CA（cholic acid）, DCA（deoxycholic acid）（50 μ mol/L,250 μ mol/L,500 μ mol/L,1000 μ mol/L）, DY（mixture of bile salts） and DS（mixture of bile acids）（250 μ mol/L,500 μ mol/L,1000 μ mol/L,1500 μ mol/L, 2000 μ mol/L）, in comparison with the control group（in normal media without bile salts and bile acids）. Cell proliferation was assessed by MTT（3-[4,5-Dimethylthiaolyl]-2,5- diphenyl-tetrazolium bromide） assay for 72 hours with different concentrations and the apoptotic cells were assayed by flow cytometry （FCM） with Annex V-FITC conjugated with propidium iodide（PI） staining for 24 hours with different concentrations（1500,2000 μt mol/L）. Results：There was no significant difference in morphology and cell proliferation in GC group after 24-72 h. Low concentration（50 μ mol/L） of GCDC, GDC, TCDC, TDC and TC accelerated gastric epithelial cell growth in a dosage-time dependent manner. At middle concentration （250-500 μ mol/L）, it showed positive effect after 24-48 h, while negative effect after 72 h. At high concentration（1000 μ mol/L）, it accelerated gastric epithelial cell growth after 24h and show consistent inhibition even leading to necrosis after 48-72 h. LCA and CA showed a positive effect on the concentration of 50 μ mol/L after 24-72 h, while 250-1000 μ mol/L showed a trend towards apoptosis after 24-72 h. At 50-500 μmol/L, DCA showed proliferation after 24 h and apoptosis after 48-72 h, but showed necrosis after 24-72 h at 1000 μmol/L. DY and DS could facilitate normal gastric mucosa epithelial cell growth at low concentration （250-500 μ mol/L）, however at 1000-2000 μ mol/L the trend shifted from apoptosis to necrosis. FCM with Annexin-V conjugated with PI staining revealed that GCDC, GDC, GC, TCDC, TDC, TC, LCA, CA, DCA, DY and DS induced apoptosis of human gastric mucosal epithelial cells. They were all significantly higher than that of the control（P 〈 0.05）, but there was no significant difference in GC group （P 〉 0.05）. The bile salts induced apoptosis in a time-dose-dependent manner. Conclusion：Our results suggested that bile acid and bile salt is the trigger of injury in human gastric mucosal epithelial cells.

Characteristics of early gastric tumors with different differentiation and predictors of long-term outcomes after endoscopic submucosal dissection

BACKGROUND Gastric cancer is a common malignant tumor of the digestive tract,and endosco-pic submucosal dissection(ESD)is the preferred treatment for early-stage gastric cancer.The analysis of the epidemiological characteristics of gastric mucosal tumors with different differentiation degrees and the influencing factors of long-term ESD efficacy may have certain significance for revealing the development of gastric cancer and ESD.AIM To analyze the features of gastric mucosal tumors at different differentiation levels,and to explore the prognostic factors of ESD.METHODS We retrospectively studied 301 lesions in 285 patients at The Second Affiliated Hospital of Xi'an Jiaotong University from 2014 to 2021,according to the latest Japanese guidelines(sixth edition),and divided them into low-grade intrae-pithelial neoplasia(LGIN),high-grade intraepithelial neoplasia(HGIN),and computed tomography at 3,6 and 12 months after ESD.We compared clinicopathologic characteristics,ESD efficacy,and complications with different degrees of differentiation,and analyzed the related factors associated with ESD.RESULTS HGIN and differentiated carcinoma patients were significantly older compared with LGIN patients(P<0.001)and accounted for more 0-IIc(P<0.001),atrophic gastritis was common(P<0.001),and irregular microvascular patterns(IMVPs)and demarcation lines(DLs)were more obvious(P<0.001).There was more infiltration in the undifferentiated carcinoma tissue(P<0.001),more abnormal folds and poorer mucosal peristalsis(P<0.001),and more obvious IMVPs,irregular microsurface patterns and DLs(P<0.05)than in the LGIN and HGIN tissues.The disease-free survival rates at 2,5,and 8 years after ESD were 95.0%,90.1%,and 86.9%,respectively.Undifferen-tiated lesions(HR 5.066),white moss(HR 7.187),incomplete resection(HR 3.658),and multiple primary cancers(HR 2.462)were significantly associated with poor prognosis.CONCLUSION Differentiations of gastric mucosal tumors have different epidemiological and endoscopic characteristics,which are closely related to the safety and efficacy of ESD.

Prevention of esophageal strictures after endoscopic submucosal dissection

Endoscopic mucosal resection (EMR) and endoscopic submucosal dissection (ESD) have recently been accepted as less invasive methods for treating patients with early esophageal cancers such as squamous cell carcinoma and dysplasia of Barrett&#x02019;s esophagus. However, the large defects in the esophageal mucosa often cause severe esophageal strictures, which dramatically reduce the patient&#x02019;s quality of life. Although preventive endoscopic balloon dilatation can reduce dysphagia and the frequency of dilatation, other approaches are necessary to prevent esophageal strictures after ESD. This review describes several strategies for preventing esophageal strictures after ESD, with a particular focus on anti-inflammatory and tissue engineering approaches. The local injection of triamcinolone acetonide and other systemic steroid therapies are frequently used to prevent esophageal strictures after ESD. Tissue engineering approaches for preventing esophageal strictures have recently been applied in basic research studies. Scaffolds with temporary stents have been applied in five cases, and this technique has been shown to be safe and is anticipated to prevent esophageal strictures. Fabricated autologous oral mucosal epithelial cell sheets to cover the defective mucosa similarly to how commercially available skin products fabricated from epidermal cells are used for skin defects or in cases of intractable ulcers. Fabricated autologous oral-mucosal-epithelial cell sheets have already been shown to be safe.

Erythropoietin -induced proliferation of gastric mucosal cells

AIM： To analyze the localization of erythropoietin receptor on gastric specimens and characterize the effects of erythropoietin on the normal gastric epithelial proliferation using a porcine gastric epithelial cell culture model. METHODS： Erythropoietin receptor was detected by RT-PCR, Western blotting and immunohistochermistry. Growth stimulation effects of erythropoietin on cultured gastric mucosal cells were determined by ELISA using bromodeoxyuridine （BrdU）. RESULTS： Erythropoietin receptor was detected on cultured porcine gastric mucosal epithelial cells. Erythropoietin receptor was also detected histochemically at the base of gastric mucosal epithelium. BrdU assay demonstrated a dose-dependent increase in growth potential of cultured porcine gastric mucosal epithelial cells by administration of erythropoietin, as well as these effects were inhibited by administration of antierythropoietin antibody （P〈 0.01）. CONCLUSION： These findings indicate that erythropoietin has a potential to proliferate gastric mucosal epithelium via erythropoietin receptor.

Cell-based therapies for limbal stem cell deficiency: a literature review

Background and Objective:Limbal stem cell deficiency(LSCD)is characterized by the insufficiency of limbal stem cells to maintain the corneal epithelium.Severe cases of LSCD may be treated with limbal transplantation from healthy autologous or allogeneic limbal tissue.Multiple cell-based therapies have been studied as alternative treatments to improve success rates and minimize immunosuppressive regimens after allogeneic transplants.In this review,we describe the success rates,and complications of different cell-based therapies for LSCD.We also discuss each therapy’s relative strengths and weaknesses,their history in animal and human studies,and their effectiveness compared to traditional transplants.Methods:PubMed was searched for publications using the terms LSCD,cell-based therapy,cultivated limbal epithelial transplantation(CLET),cultivated oral mucosal epithelial transplantation(COMET),and mesenchymal stem cells from 1989 to August 2022.Inclusion criteria were English language articles.Exclusion criteria were non-English language articles.Key Content and Findings:current cell-based therapies for LSCD are CLET and non-limbal epithelial cells.Non-limbal epithelial cell methods include COMET,conjunctival epithelial autografts,and mesenchymal stem/stromal cells(MSCs).Moreover,several alternative potential sources of non-limbal cells have described,including induced pluripotent stem cells(iPSCs),human embryonic stem cells(hESCs),human dental pulp stem cells,hair follicle bulge-derived epithelial stem cells,amniotic membrane epithelial cells,and human umbilical cord lining epithelial cells.Conclusions:Cell-based therapies are a promising treatment modality for LSCD.While CLET is currently the only approved cell-based therapy and is only approved in the European Union,more novel methods have also been shown to be effective in human or animal studies thus far.Non-limbal epithelial cells such as COMET are also an alternative treatment to allogeneic transplants especially as a surface stabilizing procedure.iPSCs are currently being studied in early phase trials and have the potential to revolutionize the way LSCD is treated.Lastly,cell-based therapies for restoring the limbal niche such as mesenchymal stem cells have also shown promising results in the first human proof-of-concept study.Several potential sources of non-limbal cells are under investigation.

Telomerase and hTERT: Can they serve as markers for gastric cancer diagnosis?

AIM: To investigate telomerase activity and human telomerase reverse transcriptase (hTERT) expression in normal human gastric mucosal epithelial cells (nhGMECs) and fibroblasts (nhGMFs).