Jun-Kai Yan

Eriocalyxin B induces apoptosis of t(8;21) leukemia cells through NF-|[kappa]|B and MAPK signaling pathways and triggers degradation of AML1-ETO oncoprotein in a caspase-3-dependent manner

Diterpenoids isolated from Labiatae family herbs have strong antitumor activities with low toxicity. In this study, Eriocalyxin B (EriB), a diterpenoid extracted from Isodon eriocalyx, was tested on human leukemia/lymphoma cells and murine leukemia models. Acute myeloid leukemia cell line Kasumi-1 was most sensitive to EriB. Significant apoptosis was observed, concomitant with Bcl-2/Bcl-XL downregulation, mitochondrial instability and caspase-3 activation. AML1-ETO oncoprotein was degraded in parallel to caspase-3 activation. EriB-mediated apoptosis was associated with NF-κB inactivation by preventing NF-κB nuclear translocation and inducing IκBα cleavage, and disturbance of MAPK pathway by downregulating ERK1/2 phosphorylation and activating AP-1. Without affecting normal hematopoietic progenitor cells proliferation, EriB was effective on primary t(8;21) leukemia blasts and caused AML1-ETO degradation. In murine t(8;21) leukemia models, EriB remarkably prolonged the survival time or decreased the xenograft tumor size. Together, EriB might be a potential treatment for t(8;21) leukemia by targeting AML1-ETO oncoprotein and activating apoptosis pathways.

Interactions between Th1 cells and Tregs affect regulation of hepatic fibrosis in biliary atresia through the IFN-|[gamma]||[sol]|STAT1 pathway

Regulatory T cells (Tregs) and CD4+ T helper (Th) cells have important roles in bile duct injury of biliary atresia (BA). However, their impacts on liver fibrosis are undefined. Between 2013 and 2016, 146 patients with various stages of BA were enrolled in this study. Peripheral blood, liver biopsy and lymph node samples were collected. Flow cytometry, magnetic cell sorting and immunostaining were used to characterize lymphocytes from BA patients. Deficiency of Tregs was observed along with increased Th1, Th2 and Th17 frequencies in the peripheral blood and livers of BA patients. The levels of peripheral and intrahepatic Th1 cells positively correlated with the stage of liver fibrosis. Furthermore, Th1 cells were located in close proximity to activated hepatic stellate cells (HSCs) and areas of fibrosis in BA livers. In culture, Th1 cells accelerated the proliferation and secretion of profibrogenic markers of HSCs through the IFN-γ/STAT1 pathway. Of note, Tregs blocked the Th1-stimulated effects on HSCs by inhibiting Th1-induced activation of STAT1. Consistent with the results of in vitro study, intrahepatic IFN-γ/STAT1 levels increased in relation to the severity of liver fibrosis in BA patients, and the altered balance between MMP2 and TIMP1 expressions in livers may contribute to increased deposition of extracellular matrix and fibrosis. Finally, to identify the effects of Th1 cells on Tregs, we demonstrated that Th1 cells upregulated the proportion of aTreg cells by secreting IFN-γ cytokine. Thus, aberrant Th1 immune responses in BA promote the proliferation and secretion of HSCs through the IFN-γ/STAT1 pathway. The regulation of HSCs by the interactions between Tregs and Th1 cells might be part of the mechanism underlying progressive liver fibrosis and may be a suitable target for therapy.

P38 MAPK Pharmacological Inhibitor SB203580 Alleviates Total Parenteral Nutrition-Induced Loss of Intestinal Barrier Function but Promotes Hepatocyte Lipoapoptosis

Background & Aims: Our previous studies have provided evidence that p38 mitogen-activated protein kinase (MAPK) is involved in total parenteral nutrition (TPN)-associated complications, but its exact effects and mechanisms have not been fully understood. This study aimed to evaluate the roles of p38 MAPK inhibitor SB203580 in the TPN-induced loss of intestinal barrier function and liver disease. Methods: A rodent model of TPN was used to analyze the roles of SB203580 in TPN-associated complications.Intestinal barrier function was evaluated by transepithelial electrical resistance (TER) and paracellular permeability in Caco-2 cells. The palmitic acid (PA) was used to induce hepatic lipoapoptosis in vitro. The lipoapoptosis was detected using Caspase-3/7 and lipid staining. Results: In the present study, we showed that SB203580 treatment significantly suppressed TPN-mediated intestinal permeability in rats. SB203580 treatment significantly inhibited IL-1β-induced an increase in tight junction permeability of Caco-2 cells via repressing the p38/ATF-2 signaling. Unexpectedly, SB203580 treatment enhanced hepatic lipoapoptosis in the model of TPN. Palmitic acid (PA)-induced hepatic lipoapoptosis in human liver cells was significantly augmented by the SB203580 treatment. Conclusions: We demonstrate that the p38 MAPK inhibitor SB203508 ameliorates intestinal barrier function but promotes hepatic lipoapoptosis in model of TPN.

Contribution of Common Variants in GABRG2, RELN and NRG3 and Interaction Networks to the Risk of Hirschsprung Disease

Background: Hirschsprung disease (HSCR) is a complex and heterogeneous disorder, characterized by a deficit in enteric nervous system. Genome-wide studies implied GABRG2, RELN and NRG3 might be involved in HSCR etiology. Here, we aimed to assess genetic variants in GABRG2, RELN and NRG3 that may confer susceptibility to HSCR and explore genetic interaction networks in HSCR. Methods: Using a strategy that combined case-control study and gene-gene interaction analysis with the MassArray system, we evaluated 24 polymorphisms within GABRG2, RELN and NRG3 in 104 HSCR cases and 151 normal controls of Han Chinese origin. Results: We observed that seven polymorphisms showed statistically significant differences between HSCR subjects and normal controls. For each of the three genes, the haplotypes which combined eight markers were the most significant. Moreover, we recruited SNPsyn, GO enrichment and MDR analyses to interrogate the interactions among GABRG2, RELN, NRG3 and our previous identified PTCH1 gene. Significant interaction networks were found among GABRG2, RELN, and PTCH1. Conclusion: We provide a first indication that common variants of GABRG2, RELN and NRG3 and the GABRG2-RELN-PTCH1 interaction networks might confer altered susceptibility to HSCR in the Han Chinese population, suggesting a potential mechanism underlying HSCR pathogenesis.

Soybean Oil-Based Lipid Emulsion Increases Intestinal Permeability of Lipopolysaccharide in Caco-2 Cells by Downregulation of P-Glycoprotein via ERK-FOXO 3a Pathway

Background and Aims: Elevated intestinal permeability of lipopolysaccharide (LPS) is a major complication for patients with parenteral nutrition (PN), but the pathogenesis is poorly understood. Intestinal P-glycoprotein (P-gp) is one of the efflux transporters that contribute to restricting the permeability of lipopolysaccharide via transcellular route. P-gp expression may be regulated by PN ingredients, and thus this study sought to investigate the effect of PN on the expression of P-gp and to elucidate the underlying mechanism in vitro. Methods: Caco-2 cells were treated with PN ingredients. Changes in P-gp expression and function were determined and the role of ERK-FOXO 3a pathway was studied. Transport studies of FITC-lipopolysaccharide (FITC-LPS) across Caco-2 cell monolayers were also performed. Results: Among PN ingredients, soybean oil-based lipid emulsion (SOLE) exhibited significant inhibitory effect on P-gp expression and function. This regulation was mediated via activation of ERK pathway with subsequent nuclear exclusion of FOXO 3a. Importantly, P-gp participated in antagonizing the permeation of FITC-LPS (apical to basolateral) across Caco-2 cell monolayers. SOLE significantly increased the permeability of FITC-LPS (apical to basolateral), which was associated with impaired P-gp function. Conclusions: The expression and function of intestinal P-gp is suppressed by SOLE in vitro.

Olive Oil-Supplemented Lipid Emulsion Induces CELF1 Expression and Promotes Apoptosis in Caco-2 Cells

Background and Aims: Parenterally-administered lipid emulsion (LE) is a key cause of enterocyte apoptosis under total parenteral nutrition, yet the pathogenesis has not been fully understood. CUGBP, Elav-like family member 1 (CELF1) has been recently identified as a crucial modulator of apoptosis, and thus this study sought to investigate its role in the LE-induced apoptosis in vitro. Methods: Caco-2 cells were used as an in vitro model. The cells were treated with varying LEs derived from soybean oil, olive oil or fish oil, and changes in the apoptosis and CELF1 expression were assessed. Rescue study was performed using transient knockdown of CELF1 with specific siRNA prior to LE treatment. Regulation of CELF1 by LE treatment was studied using quantitative real-time PCR and Western blotting. Results: All the LEs up-regulated CELF1expression and induced apoptosis, but only olive oil-supplemented lipid emulsion (OOLE)-induced apoptosis was attenuated by depletion of CELF1. Up-regulation of apoptosis-inducing factor (AIF) was involved in OOLE-induced CELF1 dependent apoptosis. The protein expression of CELF1 was up-regulated by OOLE in a dose- and time-dependent manner, but the mRNA expression of CELF1 was unchanged. Analysis by polysomal profiling and nascent protein synthesis revealed that the regulation of CELF1 by OOLE treatment was mediated by directly accelerating its protein translation. Conclusion: OOLE-induces apoptosis in Caco-2 cells partially through up-regulation of CELF1.

Supplementary choline attenuates olive oil lipid emulsion-induced enterocyte apoptosis through suppression of CELF1/AIF pathway

Enterocyte apoptosis induced by lipid emulsions is a key cause of intestinal atrophy under total parenteral nutrition (TPN) support, and our previous work demonstrated that olive oil lipid emulsion (OOLE) could induce enterocyte apoptosis via CUGBP, Elav‐like family member 1 (CELF1)/ apoptosis‐inducing factor (AIF) pathway. As TPN‐associated complications are partially related to choline deficiency, we aimed to address whether choline supplementation could attenuate OOLE‐induced enterocyte apoptosis. Herein we present evidence that supplementary choline exhibits protective effect against OOLE‐induced enterocyte apoptosis both in vivo and in vitro. In a rat model of TPN, substantial reduction in apoptotic rate along with decreased expression of CELF1 was observed when supplementary choline was added to OOLE. In cultured Caco‐2 cells, supplementary choline attenuated OOLE‐induced apoptosis and mitochondria dysfunction by suppressing CELF1/AIF pathway. Compared to OOLE alone, the expression of CELF1 and AIF was significantly decreased by supplementary choline, whereas the expression of Bcl‐2 was evidently increased. No obvious alterations were observed in Bax expression and caspase‐3 activation. Mechanistically, supplementary choline repressed the expression of CELF1 by increasing the recruitment of CELF1 mRNA to processing bodies, thus resulting in suppression of its protein translation. Taken together, our data suggest that supplementary choline exhibits effective protection against OOLE‐induced enterocyte apoptosis, and thus, it has the potential to be used for the prevention and treatment of TPN‐induced intestinal atrophy.

Urinary glutamine/glutamate ratio as a potential biomarker of pediatric chronic intestinal pseudo-obstruction

Chronic intestinal pseudo-obstruction (CIPO) is a rare intestinal motility disorder with significant morbidity and mortality in pediatric patients. The diagnosis of CIPO is difficult, because it is clinically based on the symptoms and signs of bowel obstruction which are similar to the clinical manifestations of other gastrointestinal diseases like short bowel syndrome (SBS). Therefore, it is desirable to identify and establish new laboratory diagnostic markers for CIPO that are reliable and easily accessible. In our study we have identified the ratio of the urinary glutamine and glutamic acid as a promising biomarker for distinguishing suspected CIPO cases and simple SBS cases. The area under ROC curve was 0.83, at cutoff value = 7.04 with sensitivity of 65% and specificity of 92%.

Butyrate stimulates the growth of human intestinal smooth muscle cells by activation of Yes-Associated Protein†

Intestinal smooth muscle cells play a critical role in the remodeling of intestinal structure and functional adaptation after bowel resection. Recent studies have shown that supplementation of butyrate (Bu) contributes to the compensatory expansion of a muscular layer of the residual intestine in a rodent model of short‐bowel syndrome (SBS). However, the underlying mechanism remains elusive. In this study, we found that the growth of human intestinal smooth muscle cells (HISMCs) was significantly stimulated by Bu via activation of Yes‐Associated Protein (YAP). Incubation with 0.5 mM Bu induced a distinct proliferative effect on HISMCs, as indicated by the promotion of cell cycle progression and increased DNA replication. Notably, YAP silencing by RNA interference or its specific inhibitor significantly abolished the proliferative effect of Bu on HISMCs. Furthermore, Bu induced YAP expression and enhanced the translocation of YAP from the cytoplasm to the nucleus, which led to changes in the expression of mitogenesis genes, including TEAD1, TEAD4, CTGF, and Cyr61. These results provide evidence that Bu stimulates the growth of human intestinal muscle cells by activation of YAP, which may be a potential treatment for improving intestinal adaptation.