Schisantherin D from Schisandra chinensis (Turcz.) Baill. exhibits anti-liver fibrosis capacity via modulating ETBR involved signaling, an in vitro and in vivo study.

Excess levels of chemical hepatotoxicants (alcohol, aflatoxin B1), oxidative drugs (acetaminophen) and some cytokines (ET-1, TGF-ß1) can induce chronic or acute liver injury. After these, the severe hepatic disease, especially the liver fibrosis (LF) occurs without taking measures, which brings threat to human health. The dibenzocyclooctadiene lignans of S. chinensis (SCDLs) were found to act as the hepatoprotective components via blocking endothelin B receptor (ETBR). While study on its anti-LF mechanisms especially for its refined compound of schisantherin D (SC-D) is still a lack. So this study aims to investigate the anti-fibrosis effect of SC-D with in vitro and in vivo assays. Bioinformatics analysis revealed the close relations of ETBR to Smad2, Smad3, Nrf2, etc. in LF-related signaling pathways (such as TGF-ß/Smad and Nrf2/ARE). Histopathological staining on livers showed the recovery trend in SC-D treated LF mice. SC-D also modulated expressions of ETBR and fibrosis or anti-oxidative related proteins (such as TIMP1, p-Smad2/3, Nrf2, Smad7, etc.) in LF mice livers. Serum levels of TNF-α, COLI, ALT, AST and LDH in SC-D treated mice were also downregulated compared with LF mice, and upregulated expression of GSH. In vitro studies, SC-D also modulated expressions of LF-related proteins to the normal tendency in LX-2 cell, while weakened its anti- LX-2 proliferation effect by transfections of si-Smad7 or si-Nrf2. Accordingly the anti-LF approach of SC-D showed relations with modulating ETBR linked fibrosis and anti-oxidative related signaling. Also, Smad7 and Nrf2 might be the key factors for SC-D mediated anti-LF effect.

Effect of thermophilic bacterium HB27 manganese superoxide dismutase in a rat model of chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS).

We investigated the therapeutic effects of superoxide dismutase (SOD) from thermophilic bacterium HB27 on chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) and its underlying mechanisms. A Sprague-Dawley rat model of CP/CPPS was prepared and then administered saline or Thermus thermophilic (Tt)-SOD intragastrically for 4 weeks. Prostate inflammation and fibrosis were analyzed by hematoxylin and eosin staining, and Masson staining. Alanine transaminase (ALT), aspartate transaminase (AST), serum creatinine (CR), and blood urea nitrogen (BUN) levels were assayed for all animals. Enzyme-linked immunosorbent assays (ELISA) were performed to analyze serum cytokine concentrations and tissue levels of malondialdehyde, nitric oxide, SOD, catalase, and glutathione peroxidase. Reactive oxygen species levels were detected using dichlorofluorescein diacetate. The messenger ribonucleic acid (mRNA) expression of tissue cytokines was analyzed by reverse transcription polymerase chain reaction (RT-PCR), and infiltrating inflammatory cells were examined using immunohistochemistry. Nuclear factor-κB (NF-κB) P65, P38, and inhibitor of nuclear factor-κBα (I-κBα) protein levels were determined using western blot. Tt-SOD significantly improved histopathological changes in CP/CPPS, reduced inflammatory cell infiltration and fibrosis, increased pain threshold, and reduced the prostate index. Tt-SOD treatment showed no significant effect on ALT, AST, CR, or BUN levels. Furthermore, Tt-SOD reduced inflammatory cytokine expression in prostate tissue and increased antioxidant capacity. This anti-inflammatory activity correlated with decreases in the abundance of cluster of differentiation 3 (CD3), cluster of differentiation 45 (CD45), and macrophage inflammatory protein 1α (MIP1α) cells. Tt-SOD alleviated inflammation and oxidative stress by reducing NF-κB P65 and P38 protein levels and increasing I-κBα protein levels. These findings support Tt-SOD as a potential drug for CP/CPPS.

The protective effect of manganese superoxide dismutase from thermophilic bacterium HB27 on hydrochloric acid-induced chemical cystitis in rats.

PURPOSE: To evaluate the effects of manganese superoxide dismutase (Mn-SOD) from thermophilic bacterium HB27 (name as Tt-SOD) on chemical cystitis. METHODS: Control and experimental rats were infused by intravesical saline or hydrochloric acid (HCl) on the first day of the experiments. Saline, sodium hyaluronate (SH) or Tt-SOD were infused intravesically once a day for three consequent days. On the fifth day, the rats were weighted and sacrificed following a pain threshold test. The bladder was harvested for histological and biochemical analyses. RESULTS: Tt-SOD could reduce the bladder index, infiltration of inflammatory cells in tissues, serum inflammatory factors and SOD levels, mRNA expression of inflammatory factors in tissues, and increase perineal mechanical pain threshold and serum MDA and ROS levels in HCl-induced chemical cystitis. Furthermore, Tt-SOD alleviated inflammation and oxidative stress by the negative regulation of the NF-κB p65 and p38 MAPK signaling pathway. CONCLUSIONS: Intravesical instillation of Tt-SOD provides protective effects against HCl-induced cystitis.

Anthelmintic niclosamide attenuates pressure-overload induced heart failure in mice.

The heart is a high energy demand organ and enhancing mitochondrial function is proposed as the next-generation therapeutics for heart failure. Our previous study found that anthelmintic drug niclosamide enhanced mitochondrial respiration and increased adenosine triphosphate (ATP) production in cardiomyocytes, therefore, this study aimed to determine the effect of niclosamide on heart failure in mice and the potential molecular mechanisms. The heart failure model was induced by transverse aortic constriction (TAC) in mice. Oral administration of niclosamide improved TAC-induced cardiac hypertrophy, cardiac fibrosis, and cardiac dysfunction in mice. Oral administration of niclosamide reduced TAC-induced increase of serum IL-6 in heart failure mice. In vitro, niclosamide within 0.1 µM increased mitochondrial respiration and ATP production in mice heart tissues. At the concentrations more than 0.1 µM, niclosamide reduced the increased interleukin- 6 (IL-6) mRNA expression in lipopolysaccharide (LPS)-stimulated RAW264.7 and THP-1 derived macrophages. In cultured primary cardiomyocytes and cardiac fibroblasts, niclosamide (more than 0.1 µM) suppressed IL-6- and phenylephrine-induced cardiomyocyte hypertrophy, and inhibited collagen secretion from cardiac fibroblasts. In conclusion, niclosamide attenuates heart failure in mice and the underlying mechanisms include enhancing mitochondrial respiration of cardiomyocytes, inhibiting collagen secretion from cardiac fibroblasts, and reducing the elevated serum inflammatory mediator IL-6. The present study suggests that niclosamide might be therapeutic for heart failure.

Injectable pegylated niclosamide (polyethylene glycol-modified niclosamide) for cancer therapy.

Niclosamide is an antihelminthic drug. Recent studies show that niclosamide exerts antitumor activity through inhibiting multiple signals including Wnt/ß-catenin, mTORC1, signal transducer and activator of transcription 3, NF-κB, notch signals; however, the insolubility and poor bioavailability limits its potential clinic use, the aim of the present work is to synthesize an injectable pegylated niclosamide (polyethylene glycol-modified niclosamide) and investigate its antitumor activity in vitro and in vivo. The pegylated niclosamide (mPEG5000-Nic) was synthesized and the chemical structure was identified by Fourier transform infrared spectra and 1 H nuclear magnetic resonance spectra. The antitumor activity was evaluated in CT26 and HCT116 colon cancer cells in vitro and nude mouse xenograft model of CT26 cells in vivo. The water solubility of niclosamide in mPEG5000-Nic was significantly increased. Niclosamide could be released from mPEG5000-Nic nanoparticles in PBS solution. mPEG5000-Nic inhibited the cell viability of CT26 and HCT116 cells in vitro. No animal death was observed in mice with intraperitoneal injection of mPEG5000-Nic (equivalent to 1000 mg/kg niclosamide) within 24 hr, indicating that mPEG5000-Nic was less toxic. In nude mouse, xenograft model of CT26 colon carcinoma, intraperitoneal injection of mPEG5000-Nic (equivalent to niclosamide 50 mg/kg) inhibited tumor growth but had no effect on animal body weight and heart, liver, kidney, and lung weight in vivo. Meanwhile, in the same model, intraperitoneal injection of the positive clinic drug 5-fluorouracil not only inhibited the tumor growth, but also reduced the animal body weight. Our study demonstrates that pegylated niclosamide is novel niclosamide delivery system with clinical perspective for cancer therapy.

The different response of cardiomyocytes and cardiac fibroblasts to mitochondria inhibition and the underlying role of STAT3.

Cardiomyocyte loss and cardiac fibrosis are the main characteristics of cardiac ischemia and heart failure, and mitochondrial function of cardiomyocytes is impaired in cardiac ischemia and heart failure, so the aim of this study is to identify fate variability of cardiomyocytes and cardiac fibroblasts with mitochondria inhibition and explore the underlying mechanism. The mitochondrial respiratory function was measured by using Oxygraph-2k high-resolution respirometry. The STAT3 expression and activity were evaluated by western blot. Cardiomyocytes and cardiac fibroblasts displayed different morphology. The mitochondrial respiratory function and the expressions of mitochondrial complex I, II, III, IV, and V of cardiac fibroblasts were lower than that of cardiomyocytes. Mitochondrial respiratory complex I inhibitor rotenone and H2O2 (100 µM, 4 h) treatment induced cell death of cardiomyocyte but not cardiac fibroblasts. The function of complex I/II was impaired in cardiomycytes but not cardiac fibroblasts stimulated with H2O2 (100 µM, 4 h) and in ischemic heart of mice. Rotenone and H2O2 (100 µM, 4 h) treatment reduced STAT3 expression and activity in cardiomyocytes but not cardiac fibroblasts. Inhibition of STAT3 impaired mitochondrial respiratory capacity and exacerbated H2O2-induced cell injury in cardiomycytes but not significantly in cardiac fibroblasts. In conclusion, the different susceptibility of cardiomyocytes and cardiac fibroblasts to mitochondria inhibition determines the cell fate under the same pathological stimuli and in which STAT3 plays a critical role.

Characterizations of mitochondrial uncoupling induced by chemical mitochondrial uncouplers in cardiomyocytes.

Induction of mild mitochondrial uncoupling is protective in a variety of disorders; however, it is unclear how to recognize the mild mitochondrial uncoupling induced by chemical mitochondrial uncouplers. The aim of the present study is to identify the pharmacological properties of mitochondrial uncoupling induced by mitochondrial uncouplers in cardiomyocytes. Neonatal rat cardiomyocytes were cultured. Protein levels were measured by using western blot technique. The whole cell respiratory function was determined by using high-resolution respirometry. The typical types of chemical mitochondrial uncouplers, carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone (FCCP), niclosamide, and BAM15, induced biphasic change of STAT3 activity in cardiomyocytes, activating STAT3 at low dose and inhibiting STAT3 at high dose, though the dose range of these drugs was distinct. Low-dose uncouplers induced STAT3 activation through the mild increase of mitochondrial ROS (mitoROS) generation and the subsequent JAK/STAT3 activation in cardiomyocytes. However, high-dose uncouplers induced inhibition of STAT3, decrease of ATP production, and cardiomyocyte death. High-dose uncouplers induced STAT3 inhibition through the excessive mitoROS generation and the decreased ATP -induced AMPK activation. Low-dose mitochondrial uncouplers attenuated doxorubicin (DOX)-induced STAT3 inhibition and cardiomyocyte death, and activated STAT3 contributed to the cardioprotection of low-dose mitochondrial uncouplers. Uncoupler-induced mild mitochondrial uncoupling in cardiomyocytes is characterized by STAT3 activation and ATP increase whereas excessive mitochondrial uncoupling is characterized by STAT3 inhibition, ATP decrease and cell injury. Development of mitochondrial uncoupler with optimal dose window of inducing mild uncoupling is a promising strategy for heart protection.

Niclosamide inhibits vascular smooth muscle cell proliferation and migration and attenuates neointimal hyperplasia in injured rat carotid arteries.

BACKGROUND AND PURPOSE: The anti-helminthic drug niclosamide regulates multiple cellular signals including STAT3, AMP-activated protein kinase (AMPK), Akt, Wnt/ß-catenin and mitochondrial uncoupling which are involved in neointimal hyperplasia. Here we have examined the effects of niclosamide on vascular smooth muscle cell proliferation, migration and neointimal hyperplasia and assessed the potential mechanisms. EXPERIMENTAL APPROACH: Cell migration was measured by using wound-induced migration assay and Boyden chamber assay. Protein levels were measured by using Western blot technique. Neointimal hyperplasia in vivo was induced in rats by balloon injury to the carotid artery. KEY RESULTS: Niclosamide treatment inhibited serum-induced (15% FBS) and PDGF-BB-induced proliferation and migration of vascular smooth muscle cells (A10 cells). Niclosamide showed no cytotoxicity at anti-proliferative concentrations, but induced cell apoptosis at higher concentrations. Niclosamide treatment inhibited serum-induced (15% FBS) and PDGF-BB-induced STAT3 activation (increased protein levels of p-STAT3 at Tyr705 ) but activated AMPK, in A10 cells. Niclosamide exerted no significant effects on ß-catenin expression and the activities of ERK1/2 and Akt in A10 cells. Injection (i.p.) of soluble pegylated niclosamide (PEG5000-niclosamide) (equivalent to niclosamide 25 mg·kg-1 ) attenuated neointimal hyperplasia following balloon-injury in rat carotid arteries in vivo. CONCLUSIONS AND IMPLICATIONS: Niclosamide inhibited vascular smooth muscle cell proliferation and migration and attenuated neointimal hyperplasia in balloon-injured rat carotid arteries through a mechanism involving inhibition of STAT3.

Mitochondrial Fission Inhibitors Suppress Endothelin-1-Induced Artery Constriction.

BACKGROUND/AIMS: Endothelin-1 is implicated in the pathogenesis of hypertension, but the underlying mechanisms remained elusive. Our previous study found that inhibition of mitochondrial fission of smooth muscle cells suppressed phenylephrine- and high K+-induced artery constriction. Here, we studied the effects of mitochondrial fission inhibitors on endothelin-1-induced vasoconstriction. METHODS: The tension of rat mesenteric arteries and thoracic aorta was measured by using a multi-wire myograph system. Mitochondrial morphology of aortic smooth muscle cells was observed by using transmission electron microscopy. RESULTS: Dynamin-related protein-1 selective inhibitor mdivi-1 relaxed endothelin-1-induced constriction, and mdivi-1 pre-treatment prevented endothelin-1-induced constriction of rat mesenteric arteries with intact and denuded endothelium. Mdivi-1 had a similar inhibitory effect on rat thoracic aorta. Another mitochondrial fission inhibitor dynasore showed similar effects as mdivi-1 in rat mesenteric arteries. Mdivi-1 inhibited endothelin-1-induced increase of mitochondrial fission in smooth muscle cells of rat aorta. Rho-associated protein kinase inhibitor Y-27632 which relaxed endothelin-1-induced vasoconstriction inhibited endothelin-1-induced mitochondrial fission in smooth muscle cells of rat aorta. CONCLUSION: Endothelin-1 increases mitochondrial fission in vascular smooth muscle cells, and mitochondrial fission inhibitors suppress endothelin-1-induced vasoconstriction.

Niclosamide ethanolamine inhibits artery constriction.

We previously demonstrated that the typical mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) inhibited artery constriction, but CCCP was used only as a pharmacological tool. Niclosamide is an anthelmintic drug approved by FDA. Niclosamide ethanolamine (NEN) is a salt form of niclosamide and has been demonstrated to uncouple mitochondrial oxidative phosphorylation. The aim of the present study was to elucidate the vasoactivity of NEN and the potential mechanisms. Isometric tension of rat mesenteric artery and thoracic aorta was recorded by using multi-wire myograph system. The protein levels were measured by using western blot techniques. Niclosamide ethanolamine (NEN) treatment relaxed phenylephrine (PE)- and high K+ (KPSS)-induced constriction, and pre-treatment with NEN inhibited PE- and KPSS-induced constriction of rat mesenteric arteries. In rat thoracic aorta, NEN also showed antagonism against PE- and KPSS-induced constriction. NEN induced increase of cellular ADP/ATP ratio in vascular smooth muscle cells (A10) and activated AMP-activated protein kinase (AMPK) in A10 cells and rat thoracic aorta. NEN-induced aorta relaxation was attenuated in AMPKα1 knockout (-/-) mice. SERCA inhibitors cyclopiazonic acid and thapsigargin, but not KATP channel blockers glibenclamide and 5-hydroxydecanoic acid, attenuated NEN-induced vasorelaxation in rat mesenteric arteries. NEN treatment increased cytosolic [Ca2+]i and depolarized mitochondrial membrane potential in vascular smooth muscle cells (A10). Niclosamide in non-salt form showed the similar vasoactivity as NEN in rat mesenteric arteries. Niclosamide ethanolamine inhibits artery constriction, indicating that it would be promising to be developed as an anti-hypertensive drug or it would induce vasodilation-related side effects when absorbed in vivo.

Mitochondrial Fission of Smooth Muscle Cells Is Involved in Artery Constriction.

Mitochondria are dynamic organelles and continuously undergo fission and fusion processes. Mitochondrial fission is involved in multiple physiological or pathological processes, but the role of mitochondrial fission of smooth muscle cells in artery constriction is unknown. The role of mitochondrial fission of smooth muscle cells in arterial function was investigated by measuring the tension of rat mesenteric arteries and thoracic aorta and by evaluating mitochondrial fission, mitochondrial reactive oxygen species, and cytosolic [Ca2+]i in rat vascular smooth muscle cells. Mitochondrial fission inhibitors mdivi-1 and dynasore antagonized phenylephrine- and high K+-induced constriction of rat mesenteric arteries. Mdivi-1 relaxed phenylephrine-induced constriction, and mdivi-1 pretreatment prevented phenylephrine-induced constriction in mice, rat aorta, and human mesenteric arteries. Phenylephrine- and high K+-induced increase of mitochondrial fission in smooth muscle cells of rat aorta and the increase was inhibited by mdivi-1. Mdivi-1 inhibited high K+-induced increases of mitochondrial fission, mitochondrial reactive oxygen species, and cytosolic [Ca2+]i in rat vascular smooth muscle cells. Prechelation of cytosolic Ca2+ prevented high K+-induced cytosolic [Ca2+]i increase, mitochondrial fission, and mitochondrial reactive oxygen species overproduction. Mitochondria-targeted antioxidant mito-TEMPO antagonized phenylephrine- and high K+-induced constriction of rat mesenteric arteries. Nitroglycerin and ROCK (Rho-associated protein kinase) inhibitor Y27632, the 2 vasodilators with different vasorelaxant mechanisms, relaxed high K+-induced vasoconstriction and inhibited high K+-induced mitochondrial fission. In conclusion, the mitochondrial fission of smooth muscle cells is involved in artery constriction.

Mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone induces vasorelaxation without involving KATP channel activation in smooth muscle cells of arteries.

BACKGROUND AND PURPOSE: The effects and mechanisms of chemical mitochondrial uncouplers on vascular function have never been identified. Here, we characterized the effects of the typical mitochondrial uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP) on vascular function in rat mesenteric arteries and aorta and elucidated the potential mechanisms. EXPERIMENTAL APPROACH: Isometric tension of mesenteric artery and thoracic aorta was recorded by using a multiwire myograph system. Protein levels were measured by western blot analyses. Cytosolic [Ca2+ ]i , mitochondrial ROS (mitoROS) and mitochondrial membrane potential of smooth muscle cells (A10) were measured by laser scanning confocal microscopy. KEY RESULTS: Acute treatment with CCCP relaxed phenylephrine (PE)- and high K+ (KPSS)-induced constriction of rat mesenteric arteries with intact and denuded endothelium. Pretreatment with CCCP prevented PE- and KPSS-induced constriction of rat mesenteric arteries with intact and denuded endothelium. Similarly, CCCP prevented PE- and KPSS-induced constriction of rat thoracic aorta. CCCP increased the cellular ADP/ATP ratio in vascular smooth muscle cells (A10) and activated AMPK in A10 cells and rat thoracic aorta tissues. CCCP-induced aorta relaxation was attenuated in AMPK α1 knockout (-/-) mice. SERCA inhibitors thapsigargin and cyclopiazonic acid (CPA) but not the KATP channel blocker glibenclamide partially inhibited CCCP-induced vasorelaxation in endothelium-denuded rat mesenteric arteries. CCCP increased cytosolic [Ca2+ ]i , mitoROS production and depolarized mitochondrial membrane potential in A10 cells. FCCP, the analogue of CCCP, had similar vasoactivity as CCCP in rat mesenteric arteries. CONCLUSIONS AND IMPLICATIONS: CCCP induces vasorelaxation by a mechanism that does not involve KATP channel activation in smooth muscle cells of arteries.

Azo polymeric micelles designed for colon-targeted dimethyl fumarate delivery for colon cancer therapy.

UNLABELLED: Colon-targeted drug delivery and circumventing drug resistance are extremely important for colon cancer chemotherapy. Our previous work found that dimethyl fumarate (DMF), the approved drug by the FDA for the treatment of multiple sclerosis, exhibited anti-tumor activity on colon cancer cells. Based on the pharmacological properties of DMF and azo bond in olsalazine chemical structure, we designed azo polymeric micelles for colon-targeted dimethyl fumarate delivery for colon cancer therapy. We synthesized the star-shape amphiphilic polymer with azo bond and fabricated the DMF-loaded azo polymeric micelles. The four-arm polymer star-PCL-azo-mPEG (sPCEG-azo) (constituted by star-shape PCL (polycaprolactone) and mPEG (methoxypolyethylene glycols)-olsalazine) showed self-assembly ability. The average diameter and polydispersity index of the DMF-loaded sPCEG-azo polymeric micelles were 153.6nm and 0.195, respectively. In vitro drug release study showed that the cumulative release of DMF from the DMF-loaded sPCEG-azo polymeric micelles was no more than 20% in rat gastric fluid within 10h, whereas in the rat colonic fluids, the cumulative release of DMF reached 60% in the initial 2h and 100% within 10h, indicating that the DMF-loaded sPCEG-azo polymeric micelles had excellent colon-targeted property. The DMF-loaded sPCEG-azo polymeric micelles had no significant cytotoxicity on colon cancer cells in phosphate buffered solution (PBS) and rat gastric fluid. In rat colonic fluid, the micelles showed significant cytotoxic effect on colon cancer cells. The blank sPCEG-azo polymeric micelles (without DMF) showed no cytotoxic effect on colon cancer cells in rat colonic fluids. In conclusion, the DMF-loaded sPCEG-azo polymeric micelles show colon-targeted DMF release and anti-tumor activity, providing a novel approach potential for colon cancer therapy. STATEMENT OF SIGNIFICANCE: Colon-targeted drug delivery and circumventing drug resistance are extremely important for colon cancer chemotherapy. Our previous work found that dimethyl fumarate (DMF), the approved drug by the FDA for the treatment of multiple sclerosis, exhibited anti-tumor activities on colon cancer cells (Br J Pharmacol. 2015 172(15):3929-43.). Based on the pharmacological properties of DMF and azo bond in olsalazine chemical structure, we designed azo polymeric micelles for colon-targeted dimethyl fumarate delivery for colon cancer therapy. We found that the DMF-loaded sPCEG-azo polymeric micelles showed colon-targeted DMF release and anti-tumor activities, providing a novel approach potential for colon cancer therapy.

GDF11/BMP11 activates both smad1/5/8 and smad2/3 signals but shows no significant effect on proliferation and migration of human umbilical vein endothelial cells.

GDF11/BMP11, a member of TGF-ß superfamily, was reported to rejuvenate heart, skeletal muscle and blood vessel architecture in aged mice. However, the rejuvenative effects of GDF11 were questioned recently. Here, we investigated the effects of GDF11 on smad and non-smad signals in human umbilical vein endothelial cells (HUVECs) and the effects of GDF11 on proliferation and migration of HUVECs and primary rat aortic endothelial cells (RAECs). GDF11 factor purchased from two different companies (PeproTech and R&D Systems) was comparatively studied. Western blot was used to detect the protein expressions. The cell viability and migration were examined by using MTT and wound healing assays. Results showed that GDF11 activated both smad1/5/8 and smad2/3 signals in HUVECs. GDF11 increased protein expression of NADPH oxidase 4(NOX4) in HUVECs. GDF11 showed no significant effect on the protein level of p38, p-p38, ERK, p-ERK, Akt, p-Akt (Ser473) and p-Akt(Thr308), but increased the protein level of p-JNK and p-AMPK in HUVECs, and these increases were inhibited by antioxidant mitoTEMPO treatment. GDF11 slightly increased cell viability after short-term treatment and slightly decreased cell viability after long-term treatment. GDF11 showed no significant effect on cell proliferation and migration. These data indicated that the notion of GDF11 as a rejuvenation-related factor for endothelial cells needs to be cautious.

Taurochenodeoxycholate relaxes rat mesenteric arteries through activating eNOS: Comparing with glycochenodeoxycholate and tauroursodeoxycholate.

The bile acids (BAs) and their conjugates have vascular activities and the serum levels of BAs and their conjugates are increased in liver diseases. In the present study, we examined the in vitro vasoactivities of BAs conjugates taurochenodeoxycholate (TCDC) (5-80 µM), glycochenodeoxycholate (GCDC) (20-150 µM) and tauroursodeoxycholate (TUDC) (20-150 µM) in rat mesenteric arteries and thoracic aorta. The isometric tension of rat mesenteric arteries and thoracic aorta was recorded by using multi-wire myograph systems. TCDC induced significant concentration-dependent relaxation in endothelium-intact but not endothelium-denuded rat mesenteric arteries pre-contracted with phenylephrine (PE). TCDC also showed vasorelaxant effects on high K(+) induced contraction in rat mesenteric arteries. L-NAME treatment inhibited TCDC-induced relaxation in mesenteric arteries pre-contracted with PE. Acute treatment with TCDC increased protein expression of P-eNOS (ser1177) in human umbilical vein endothelial cells. GCDC dose-dependently relaxed PE-induced vasoconstriction in both endotheium-intact and endothelium-denuded rat mesenteric arteries, but GCDC showed no effect on high K(+)-induced vasoconstriction. Both GCDC and TCDC showed no apparent relaxation on PE and high K(+)-induced vasoconstriction in rat thoracic aorta. TUDC showed no effect on PE and high K(+)-induced vasoconstriction in rat mesenteric arteries and thoracic aorta. The study demonstrates that TCDC relaxes rat mesenteric arteries through activating eNOS. TCDC might be the major BAs conjugate for vasorelaxation in vivo.

Dimethyl fumarate induces necroptosis in colon cancer cells through GSH depletion/ROS increase/MAPKs activation pathway.

BACKGROUND AND PURPOSE: Dimethyl fumarate (DMF) is a newly approved drug for the treatment of relapsing forms of multiple sclerosis and relapsing-remitting multiple sclerosis. Here, we investigated the effects of DMF and its metabolites mono-methylfumarate (MMF and methanol) on different gastrointestinal cancer cell lines and the underlying molecular mechanisms involved. EXPERIMENTAL APPROACH: Cell viability was measured by the MTT or CCK8 assay. Protein expressions were measured by Western blot analysis. LDH release, live- and dead-cell staining, intracellular GSH levels, and mitochondrial membrane potential were examined by using commercial kits. KEY RESULTS: DMF but not MMF induced cell necroptosis, as demonstrated by the pharmacological tool necrostatin-1, transmission electron microscopy, LDH and HMGB1 release in CT26 cells. The DMF-induced decrease in cellular GSH levels as well as cell viability and increase in reactive oxygen species (ROS) were inhibited by co-treatment with GSH and N-acetylcysteine (NAC) in CT26 cells. DMF activated JNK, p38 and ERK MAPKs in CT26 cells and JNK, p38 and ERK inhibitors partially reversed the DMF-induced decrease in cell viability. GSH or NAC treatment inhibited DMF-induced JNK, p38, and ERK activation in CT26 cells. DMF but not MMF increased autophagy responses in SGC-7901, HCT116, HT29 and CT26 cancer cells, but autophagy inhibition did not prevent the DMF-induced decrease in cell viability. CONCLUSION AND IMPLICATIONS: DMF but not its metabolite MMF induced necroptosis in colon cancer cells through a mechanism involving the depletion of GSH, an increase in ROS and activation of MAPKs.