Erastin Disrupts Mitochondrial Permeability Transition Pore (mPTP) and Induces Apoptotic Death of Colorectal Cancer Cells.

We here evaluated the potential anti-colorectal cancer activity by erastin, a voltage-dependent anion channel (VDAC)-binding compound. Our in vitro studies showed that erastin exerted potent cytotoxic effects against multiple human colorectal cancer cell lines, possibly via inducing oxidative stress and caspase-9 dependent cell apoptosis. Further, mitochondrial permeability transition pore (mPTP) opening was observed in erastin-treated cancer cells, which was evidenced by VDAC-1 and cyclophilin-D (Cyp-D) association, mitochondrial depolarization, and cytochrome C release. Caspase inhibitors, the ROS scavenger MnTBAP, and mPTP blockers (sanglifehrin A, cyclosporin A and bongkrekic acid), as well as shRNA-mediated knockdown of VDAC-1, all significantly attenuated erastin-induced cytotoxicity and apoptosis in colorectal cancer cells. On the other hand, over-expression of VDAC-1 augmented erastin-induced ROS production, mPTP opening, and colorectal cancer cell apoptosis. In vivo studies showed that intraperitoneal injection of erastin at well-tolerated doses dramatically inhibited HT-29 xenograft growth in severe combined immunodeficient (SCID) mice. Together, these results demonstrate that erastin is cytotoxic and pro-apoptotic to colorectal cancer cells. Erastin may be further investigated as a novel anti-colorectal cancer agent

Prediction and Analysis of Key Genes in Glioblastoma Based on Bioinformatics

Understanding the mechanisms of glioblastoma at the molecular and structural level is not only interesting for basic science but also valuable for biotechnological application, such as the clinical treatment. In the present study, bioinformatics analysis was performed to reveal and identify the key genes of glioblastoma multiforme (GBM). The results obtained in the present study signified the importance of some genes, such as COL3A1, FN1, and MMP9, for glioblastoma. Based on the selected genes, a prediction model was built, which achieved 94.4% prediction accuracy. These findings might provide more insights into the genetic basis of glioblastoma

Erastin induces ROS production and caspase-dependent apoptosis in cultured colorectal cancer cells.

<p>Colorectal cancer cells (HT-29, DLD-1 and Caco-2 lines) or NCM460 colon epithelial cells were treated with vehicle control (0.1% DMSO, “Ctrl”) or indicated concentrations of erastin for applied time, cell apoptosis was examined by listed assays (A-C, G and H); ROS production was also examined (D and I). HT-29 cells were pre-treated with z-DEVD-fmk (“zDEVD”, 50 μM), z-LEHD-fmk (“zLEHD”, 50 μM) or MnTBAP (10 μM) for 1 hour prior to applied erastin stimulation, cell survival and cell death were tested by MTT assay (E) and trypan blue assay (F), respectively. For each assay, n = 5. The data presented were mean ± SD. Experiments were repeated three times with similar results obtained. * p < 0.05 vs. group of “Ctrl”. ^# p < 0.05 vs. group of erastin only (E and F).</p

VDAC-1 over-expression potentiates erastin’s cytotoxicity.

<p>Stably HT-29 cells or NCM460 colon epithelial cells expressing empty vector (pSuper-puro, “Vec”) or VDAC-1 cDNA (“VDAC-1”) were treated with designed erastin for applied time, VDAC-1 expression, cell survival and apoptosis were tested by Western blotting assay (A and F), MTT assay (B and G) and histone DNA ELISA assay (C), respectively; ROS production (D) and JC-10 intensity (E) were also analyzed. For each assay, n = 4. The data presented were mean ± SD. Experiments were repeated three times with similar results obtained. VDAC-1 expression was quantified (F).* p < 0.05 vs. Ctrl group of “Vec” cells (B-E). ^# p < 0.05 vs. erastin group of “Vec” cells (B-E). “Trans” stands for transfection control (D and E).</p

Erastin induces mPTP opening in cultured colorectal cancer cells.

<p>HT-29 cells were treated with applied erastin for indicated time, mPTP opening was evidenced by mitochondrial VDAC-1-ANT-1 association (A), cytochrome C (“Cyto-C”) release (B) and JC-10 intensity increase (C). HT-29 cells were pre-treated with sanglifehrin A (SfA, 2.5 μM), cyclosporin A (CsA, 0.5 μM) or bongkrekic acid (BA, 5 μM) prior to erastin (10 μM) treatment, cell survival (D) and apoptosis (E) were analyzed afterwards. Stably HT-29 cells expressing VDAC-1 shRNA-1/-2 or scramble control shRNA (“scr shRNA”) were treated with erastin (10 μM), VDAC-1 expression (F), cell survival (G) and apoptosis (H) were tested. ANT-1-assocaited Cyp-D (A), cytosol cytochrome C expression (B) and VDAC-1 expression (F) were quantified. For each assay, n = 4. The data presented were mean ± SD. Experiments were repeated three times with similar results obtained. * p < 0.05 vs. group of “Ctrl”. ^# p < 0.05 vs. group of erastin only (D and E) or the “scr shRNA” group (G and H). “Trans” stands for transfection control (F-H).</p

Erastin exerts cytotoxic, but not cytostatic, effects to cultured colorectal cancer cells.

<p>Colorectal cancer cells (HT-29, DLD-1 and Caco-2 lines) or NCM460 colon epithelial cells were treated with vehicle control (0.1% DMSO, “Ctrl”) or indicated concentrations of erastin for applied time, cell survival was tested by MTT assay (A and E) and colony formation assay (C); The percentage of trypan blue positive (“dead” cells) was recorded (B); Cell proliferation was tested by BrdU incorporation assay (D and F). For each assay, n = 5. The data presented were mean ± SD. Experiments were repeated three times with similar results obtained. * p < 0.05 vs. group of “Ctrl”.</p

Erastin administration inhibits HT-29 xenograft growth in SCID mice.

<p>HT-29 tumor bearing SCID mice were intraperitoneally administrated with erastin (10/30 mg/kg body weight or “bw”, daily, for 4 weeks) or vehicle (Saline) control, tumor volumes (A) and mice body weights (B) were recorded weekly for five weeks; Daily tumor growth was also calculated (C). At the termination of experiments, all the xenografts were isolated and weighted (D). For each assay, n = 10. The data presented were mean ± SD. Experiments were repeated twice with similar results obtained. * p < 0.05 vs. group of “Vehicle”. ^# p < 0.05 vs. group of erastin at 10 mg/kg of body weight.</p

COX-2/sEH Dual Inhibitor PTUPB Alleviates CCl 4 -Induced Liver Fibrosis and Portal Hypertension.

Background: 4-(5-phenyl-3-{3-[3-(4-trifluoromethylphenyl)-ureido]-propyl}-pyrazol-1-yl) -benzenesulfonamide (PTUPB), a dual cyclooxygenase-2 (COX-2)/soluble epoxide hydrolase (sEH) inhibitor, was found to alleviate renal, pulmonary fibrosis and liver injury. However, few is known about the effect of PTUPB on liver cirrhosis. In this study, we aimed to explore the role of PTUPB in liver cirrhosis and portal hypertension (PHT). Method: Rat liver cirrhosis model was established via subcutaneous injection of carbon tetrachloride (CCl4) for 16 weeks. The experimental group received oral administration of PTUPB (10 mg/kg) for 4 weeks. We subsequently analyzed portal pressure (PP), liver fibrosis, inflammation, angiogenesis, and intra- or extrahepatic vascular remodeling. Additionally, network pharmacology was used to investigate the possible mechanisms of PTUPB in live fibrosis. Results: CCl4 exposure induced liver fibrosis, inflammation, angiogenesis, vascular remodeling and PHT, and PTUPB alleviated these changes. PTUPB decreased PP from 17.50 ± 4.65 to 6.37 ± 1.40 mmHg, reduced collagen deposition and profibrotic factor. PTUPB alleviated the inflammation and bile duct proliferation, as indicated by decrease in serum interleukin-6 (IL-6), liver cytokeratin 19 (CK-19), transaminase, and macrophage infiltration. PTUPB also restored vessel wall thickness of superior mesenteric arteries (SMA) and inhibited intra- or extrahepatic angiogenesis and vascular remodeling via vascular endothelial growth factor (VEGF), von Willebrand factor (vWF), etc. Moreover, PTUPB induced sinusoidal vasodilation by upregulating endothelial nitric oxide synthase (eNOS) and GTP-cyclohydrolase 1 (GCH1). In enrichment analysis, PTUPB engaged in multiple biological functions related to cirrhosis, including blood pressure, tissue remodeling, immunological inflammation, macrophage activation, and fibroblast proliferation. Additionally, PTUPB suppressed hepatic expression of sEH, COX-2, and transforming growth factor-β (TGF-β). Conclusion: 4-(5-phenyl-3-{3-[3-(4-trifluoromethylphenyl)-ureido]-propyl}-pyrazol-1-yl)- benzenesulfonamide ameliorated liver fibrosis and PHT by inhibiting fibrotic deposition, inflammation, angiogenesis, sinusoidal, and SMA remodeling. The molecular mechanism may be mediated via the downregulation of the sEH/COX-2/TGF-β

MoS₂/NiSe₂/rGO Multiple-Interfaced Sandwich-like Nanostructures as Efficient Electrocatalysts for Overall Water Splitting

Constructing a heterogeneous interface using different components is one of the effective measures to achieve the bifunctionality of nanocatalysts, while synergistic interactions between multiple interfaces can further optimize the performance of single-interface nanocatalysts. The non-precious metal nanocatalysts MoS2/NiSe2/reduced graphene oxide (rGO) bilayer sandwich-like nanostructure with multiple well-defined interfaces is prepared by a simple hydrothermal method. MoS2 and rGO are layered nanostructures with clear boundaries, and the NiSe2 nanoparticles with uniform size are sandwiched between both layered nanostructures. This multiple-interfaced sandwich-like nanostructure is prominent in catalytic water splitting with low overpotential for oxygen evolution reaction (OER) and hydrogen evolution reaction (HER) and almost no degradation in performance after a 20 h long-term reaction. In order to simulate the actual overall water splitting process, the prepared nanostructures are assembled into MoS2/NiSe2/rGO||MoS2/NiSe2/rGO modified two-electrode system, whose overpotential is only 1.52 mV, even exceeded that of noble metal nanocatalyst (Pt/C||RuO2~1.63 mV). This work provides a feasible idea for constructing multi-interface bifunctional electrocatalysts using nanoparticle-doped bilayer-like nanostructures

Inhibition of the gut enzyme intestinal alkaline phosphatase may explain how aspartame promotes glucose intolerance and obesity in mice

Diet soda consumption has not been associated with tangible weight loss. Aspartame (ASP) commonly substitutes sugar and one of its breakdown products is phenylalanine (PHE), a known inhibitor of intestinal alkaline phosphatase (IAP), a gut enzyme shown to prevent metabolic syndrome in mice. We hypothesized that ASP consumption might contribute to the development of metabolic syndrome based on PHE’s inhibition of endogenous IAP. The design of the study was such that for the in vitro model, IAP was added to diet and regular soda, and IAP activity was measured. For the acute model, a closed bowel loop was created in mice. ASP or water was instilled into it and IAP activity was measured. For the chronic model, mice were fed chow or high-fat diet (HFD) with/without ASP in the drinking water for 18 weeks. The results were that for the in vitro study, IAP activity was lower (p < 0.05) in solutions containing ASP compared with controls. For the acute model, endogenous IAP activity was reduced by 50% in the ASP group compared with controls (0.2 ± 0.03 vs 0.4 ± 0.24) (p = 0.02). For the chronic model, mice in the HFD + ASP group gained more weight compared with the HFD + water group (48.1 ± 1.6 vs 42.4 ± 3.1, p = 0.0001). Significant difference in glucose intolerance between the HFD ± ASP groups (53 913 ± 4000.58 (mg·min)/dL vs 42 003.75 ± 5331.61 (mg·min)/dL, respectively, p = 0.02). Fasting glucose and serum tumor necrosis factor-alpha levels were significantly higher in the HFD + ASP group (1.23- and 0.87-fold increases, respectively, p = 0.006 and p = 0.01). In conclusion, endogenous IAP’s protective effects in regard to the metabolic syndrome may be inhibited by PHE, a metabolite of ASP, perhaps explaining the lack of expected weight loss and metabolic improvements associated with diet drinks.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author