Central role of intestinal epithelial glucocorticoid receptor in alcohol- and corticosterone-induced gut permeability and systemic response.

Corticosterone, the stress hormone, exacerbates alcohol-associated tissue injury, but the mechanism involved is unknown. We examined the role of the glucocorticoid receptor (GR) in corticosterone-mediated potentiation of alcohol-induced gut barrier dysfunction and systemic response. Hepatocyte-specific GR-deficient (GRΔHC ) and intestinal epithelial-specific GR-deficient (GRΔIEC ) mice were fed ethanol, combined with corticosterone treatment. Intestinal epithelial tight junction integrity, mucosal barrier function, microbiota dysbiosis, endotoxemia, systemic inflammation, liver damage, and neuroinflammation were assessed. Corticosterone potentiated ethanol-induced epithelial tight junction disruption, mucosal permeability, and inflammatory response in GRΔHC mouse colon; these effects of ethanol and corticosterone were absent in GRΔIEC mice. Gut microbiota compositions in ethanol-fed GRΔHC and GRΔIEC mice were similar to each other. However, corticosterone treatment in ethanol-fed mice shifted the microbiota composition to distinctly different directions in GRΔHC and GRΔIEC mice. Ethanol and corticosterone synergistically elevated the abundance of Enterobacteriaceae and Escherichia coli and reduced the abundance of Lactobacillus in GRΔHC mice but not in GRΔIEC mice. In GRΔHC mice, corticosterone potentiated ethanol-induced endotoxemia and systemic inflammation, but these effects were absent in GRΔIEC mice. Interestingly, ethanol-induced liver damage and its potentiation by corticosterone were observed in GRΔHC mice but not in GRΔIEC mice. GRΔIEC mice were also resistant to ethanol- and corticosterone-induced inflammatory response in the hypothalamus. These data indicate that the intestinal epithelial GR plays a central role in alcohol- and corticosterone-induced gut barrier dysfunction, microbiota dysbiosis, endotoxemia, systemic inflammation, liver damage, and neuroinflammation. This study identifies a novel target for potential therapeutic for alcohol-associated tissue injury.

Prevention and mitigation of alcohol-induced neuroinflammation by Lactobacillus plantarum by an EGF receptor-dependent mechanism.

Neuroinflammation is implicated in the pathogenesis of alcohol use disorders. We investigated the role of Gut-Brain interactions in alcohol-induced neuroinflammation by probiotic-mediated manipulation of intestinal dysbiosis in mice. Chronic ethanol feeding induced dysbiosis, as evidenced by an increase in Firmicutes/Bacteroidetes ratio and depletion of Lactobacillus species in the colon. Ethanol increased the levels of IL-1ß, IL-6, and TNFα in plasma and the mRNA for IL-1ß, IL-6, TNFα, and MCP1 genes in the cerebral cortex and hippocampus. Ethanol feeding increased inulin flux from the circulation into different brain regions, accompanied by the increase in TLR4 mRNA levels in the cerebral cortex and hippocampus. The immunofluorescence confocal microscopy showed that ethanol elevates the expression of microglial activation marker TMEM119 in the cerebral cortex. Feeding L. plantarum suppressed the ethanol-induced dysbiosis to some extent, as evidenced by attenuation of ethanol effects on Firmicutes/Bacteroidetes ratio and abundance of Lactobacillus spp. L. plantarum blocked ethanol-induced elevation of plasma cytokines, inulin permeability to the brain, mRNA for TLR4, IL-1ß, IL-6, TNFα, and MCP1 in brain regions, and the expression of TMEM119 in the cerebral cortex. The L. plantarum effect was absent in mice that express a dominant-negative EGFR, suggesting that the EGFR receptor plays an essential role in the protective effect of L. plantarum against ethanol-induced neuroinflammation. L. plantarum, when administered after chronic ethanol-induced injury, rescued the ethanol-induced systemic inflammation and neuroinflammation. This study demonstrates that L. plantarum in the gut prevents and mitigates ethanol-induced neuroinflammation by an EGFR-dependent mechanism.

TEM study of chronic alcoholism effects on early carcinogenesis by probing the nanoscale structural alterations of cell nuclei.

Nanoscale structural alteration in the nuclei of cells with the progression of carcinogenesis is due to the rearrangements of the basic building blocks in the cell such as DNA, RNA, lipids, etc. Although epigenetic modifications underlie the development of cancer, exposure to carcinogenic chemicals such as alcohol also enhances the development of cancer. We report the effects of chronic alcoholism on early-carcinogenesis based on changes in the degree of nanoscale structural alterations (L d) in nuclei. For this, transmission electron microscopy (TEM) imaging of the nuclei of colonic cells is performed for the following four mouse models: control mice; chronic alcoholic mice treated with ethanol (i.e., EtOH mice); mice treated with colonic carcinogen azoxymethane (AOM) and dextran sulfate sodium (DSS) that induced colitis (i.e., AOM + DSS mice); and chronic alcoholic or EtOH treated mice, together with AOM and DSS treatment (i.e., AOM + DSS + EtOH mice). The disordered optical lattices are constructed from their respective TEM images of thin colonic cell nuclei and the L d values are calculated using the inverse participation ratio (IPR) technique from the spatially localized eigenfunctions of these lattices. Results show no significant difference in the average L d value of the colon cell nuclei of alcohol treated mice relative to its control [i.e., L d(C) ∼ L d(EtOH)]; however, an increase in the L d value of alcohol treated precancerous cells [i.e., L d(AOM + DSS + EtOH) > L d(AOM + DSS)], indicating that alcohol accelerates the early carcinogenic process.

LPAR2 receptor activation attenuates radiation-induced disruption of apical junctional complexes and mucosal barrier dysfunction in mouse colon.

The tight junction (TJ) and barrier function of colonic epithelium is highly sensitive to ionizing radiation. We evaluated the effect of lysophosphatidic acid (LPA) and its analog, Radioprotein-1, on γ-radiation-induced colonic epithelial barrier dysfunction using Caco-2 and m-ICC12 cell monolayers in vitro and mice in vivo. Mice were subjected to either total body irradiation (TBI) or partial body irradiation (PBI-BM5). Intestinal barrier function was assessed by analyzing immunofluorescence localization of TJ proteins, mucosal inulin permeability, and plasma lipopolysaccharide (LPS) levels. Oxidative stress was analyzed by measuring protein thiol oxidation and antioxidant mRNA. In Caco-2 and m-ICC12 cell monolayers, LPA attenuated radiation-induced redistribution of TJ proteins, which was blocked by a Rho-kinase inhibitor. In mice, TBI and PBI-BM5 disrupted colonic epithelial tight junction and adherens junction, increased mucosal permeability, and elevated plasma LPS; TJ disruption by TBI was more severe in Lpar2-/- mice compared to wild-type mice. RP1, administered before or after irradiation, alleviated TBI and PBI-BM5-induced TJ disruption, barrier dysfunction, and endotoxemia accompanied by protein thiol oxidation and downregulation of antioxidant gene expression, cofilin activation, and remodeling of the actin cytoskeleton. These data demonstrate that LPAR2 receptor activation prevents and mitigates γ-irradiation-induced colonic mucosal barrier dysfunction and endotoxemia.

Influence of imidacloprid on bacterial community diversity of mango orchard soil assessed through 16S rRNA sequencing-based metagenomic analysis.

Imidacloprid, used against mango hopper, is a persistent insecticide in soil. Microbes have the ability to remove toxic pesticides from soil surface. Metagenomic is an approach for understanding the diversity and related metabolic activities in any environmental sample without culturing the microbes. Metagenomic analysis of mango orchard soil was carried out using 16S rRNA gene sequencing to understand the impact of imidacloprid on soil microbial population. In control and imidacloprid applied soil samples, representative sequences clustered were 0.142930 and 0.082320 million, respectively. At the kingdom level, 85 and 88 percent represented to bacteria, 2 and 1 percent to archaea, and 13 and 11 percent to unassigned for control and treated metagenomes, respectively. At phylum level, 16 and 17 percent of OTUs (operational taxonomic units) were assigned with Proteobacteria, while 13 and 11 percent of OTUs were unassigned in control and imidacloprid-treated samples, respectively. The other abundant phyla in both the samples were Planctomycetes, Bacteroidetes, and Actinobacteria. At class level, 9 and 11 percent of OTUs were assigned with Planctomycetia in control as well as imidacloprid-treated samples, respectively. A number of OTUs present in control and imidacloprid applied samples are 31,173 and 21,909, respectively, with 18,018 number of OTUs shared between the two samples. The genus Gemmata totally disappeared in imidacloprid applied soil, while those belonging to class Phycisphaerae, genus Prevotella and species copri were identified in imidacloprid treatment. Bacterial community transformation was evident from this study indicating possible microbial bioremediation of imidacloprid in mango orchard soil.

Phosphorylation hotspot in the C-terminal domain of occludin regulates the dynamics of epithelial junctional complexes.

The apical junctional complex (AJC), which includes tight junctions (TJs) and adherens junctions (AJs), determines the epithelial polarity, cell-cell adhesion and permeability barrier. An intriguing characteristic of a TJ is the dynamic nature of its multiprotein complex. Occludin is the most mobile TJ protein, but its significance in TJ dynamics is poorly understood. On the basis of phosphorylation sites, we distinguished a sequence in the C-terminal domain of occludin as a regulatory motif (ORM). Deletion of ORM and expression of a deletion mutant of occludin in renal and intestinal epithelia reduced the mobility of occludin at the TJs. ORM deletion attenuated Ca2+ depletion, osmotic stress and hydrogen peroxide-induced disruption of TJs, AJs and the cytoskeleton. The double point mutations T403A/T404A, but not T403D/T404D, in occludin mimicked the effects of ORM deletion on occludin mobility and AJC disruption by Ca2+ depletion. Both Y398A/Y402A and Y398D/Y402D double point mutations partially blocked AJC disruption. Expression of a deletion mutant of occludin attenuated collective cell migration in the renal and intestinal epithelia. Overall, this study reveals the role of ORM and its phosphorylation in occludin mobility, AJC dynamics and epithelial cell migration.

Photodynamic activity attained through the ruptured π-conjugation of pyridyl groups with a porphyrin macrocycle: synthesis and the photophysical and photobiological evaluation of 5-mono-(4-nitrophenyl)-10,15,20-tris-[4-(phenoxymethyl)pyridine]-porphyrin and its Zn(ii) complex.

This article compares a reported hydrophobic and photobiologically inert porphyrin synthon, (NPh)TPyP, bearing a single meso-4-nitrophenyl group and three meso-pyridyl groups (A3B type) with a new photobiologically active metal-free porphyrin, P3N, and its zinc-complex, P3NZn, which bear a meso-4-nitrophenyl group along with three distal pyridyl groups. Both P3N and P3NZn experience ruptured π-conjugation with the porphyrin macrocycle and attain hydrophilicity, as indicated via density functional theory (DFT) calculations, becoming photobiologically active under in vitro conditions. The non-invasive photodynamic activity (PDA) predominantly shown by the zinc-complex P3NZn (with higher hydrophilicity) towards KRAS-mutated human lung-cancer cells (A549) was studied. The results indicate the existence of intracellular singlet oxygen inflicted anticancer PDA, which is apparent through the upregulation of intracellular reactive oxygen species (ROS) and the downregulation of both intracellular superoxide dismutase (SOD) and intracellular reduced glutathione (GSH) levels. The trends obtained from both SOD and GSH assays were indicators of therapeutic defence against oxidative stress via neutralizing superoxide anions (SOA).

Biochemical regulation and structural analysis of copper-transporting ATPase in a human hepatoma cell line for Wilson disease.

Hepatic copper levels differ among patients with Wilson disease (WD) and normal individuals depending on the dietary intake, copper bioavailability, and genetic factors. Copper chloride (CuCl2 ) caused dose-dependent reduction in cell viability of human teratocarcinoma (HepG2) cell line, measured using the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Cells were exposed to different concentrations of CuCl2 in log doses and maximum cell viability reduction was recorded at 15 µg/mL. Toxic dose of CuCl2 is potent inducer of reactive oxygen species (ROS). Apoptosis as a pattern of cell death was confirmed through sub-G1 fraction and morphological changes such as mitochondrial depolarization, endoplasmic reticulum and lysosomal destabilization, phosphatidylserine translocation, and DNA damage. Our transcriptional and translational results strongly support apoptotic cell death. Using the available data present in dbSNP and bioinformatics tools, three nonsynonymous single nucleotide polymorphisms (nsSNPs) were identified as deleterious, reducing the stability of protein ATP7B. Structural analysis of native and mutant ATP7B proteins was investigated using molecular dynamics simulation (MDS) approach. Mutation in ATP7B gene might disturb the structural conformation and catalytic function of the ATP7B protein may be inducing WD. Hence, excess dietary intake of copper chloride must be avoided for safety of health to prevent from WD.

Lactobacillus plantarum prevents and mitigates alcohol-induced disruption of colonic epithelial tight junctions, endotoxemia, and liver damage by an EGF receptor-dependent mechanism.

Pathogenesis of alcohol-related diseases such as alcoholic hepatitis involves gut barrier dysfunction, endotoxemia, and toxin-mediated cellular injury. Here we show that Lactobacillus plantarum not only blocks but also mitigates ethanol (EtOH)-induced gut and liver damage in mice. L. plantarum blocks EtOH-induced protein thiol oxidation, and down-regulation of antioxidant gene expression in colon L. plantarum also blocks EtOH-induced expression of TNF-α, IL-1ß, IL-6, monocyte chemotactic protein 1 ( MCP1), C-X-C motif chemokine ligand ( CXCL)1, and CXCL2 genes in colon. Epidermal growth factor receptor (EGFR) signaling mediates the L. plantarum-mediated protection of tight junctions (TJs) and barrier function from acetaldehyde, the EtOH metabolite, in Caco-2 cell monolayers. In mice, doxycycline-mediated expression of dominant negative EGFR blocks L. plantarum-mediated prevention of EtOH-induced TJ disruption, mucosal barrier dysfunction, oxidative stress, and inflammatory response in colon. L. plantarum blocks EtOH-induced endotoxemia as well as EtOH-induced pathologic lesions, triglyceride deposition, oxidative stress, and inflammatory responses in the liver by an EGFR-dependent mechanism. L. plantarum treatment after injury accelerated recovery from EtOH-induced TJ, barrier dysfunction, oxidative stress, and inflammatory response in colon, endotoxemia, and liver damage. Results demonstrate that L. plantarum has both preventive and therapeutic values in treatment of alcohol-induced tissue injury, particularly in alcoholic hepatitis.-Shukla, P. K., Meena, A. S., Manda, B., Gomes-Solecki, M., Dietrich, P., Dragatsis, I., Rao, R. Lactobacillus plantarum prevents and mitigates alcohol-induced disruption of colonic epithelial tight junctions, endotoxemia, and liver damage by an EGF receptor-dependent mechanism.

Calcium-mediated oxidative stress: a common mechanism in tight junction disruption by different types of cellular stress.

The role of reactive oxygen species (ROS) in osmotic stress, dextran sulfate sodium (DSS) and cyclic stretch-induced tight junction (TJ) disruption was investigated in Caco-2 cell monolayers in vitro and restraint stress-induced barrier dysfunction in mouse colon in vivo Live cell imaging showed that osmotic stress, cyclic stretch and DSS triggered rapid production of ROS in Caco-2 cell monolayers, which was blocked by depletion of intracellular Ca2+ by 1,2-bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Knockdown of CaV1.3 or TRPV6 channels blocked osmotic stress and DSS-induced ROS production and attenuated TJ disruption and barrier dysfunction. N-Acetyl l-cysteine (NAC) and l-NG-Nitroarginine methyl ester (l-NAME) blocked stress-induced TJ disruption and barrier dysfunction. NAC and l-NAME also blocked stress-induced activation of c-Jun N-terminal kinase (JNK) and c-Src. ROS was colocalized with the mitochondrial marker in stressed cells. Cyclosporin A blocked osmotic stress and DSS-induced ROS production, barrier dysfunction, TJ disruption and JNK activation. Mitochondria-targeted Mito-TEMPO blocked osmotic stress and DSS-induced barrier dysfunction and TJ disruption. Chronic restraint stress in mice resulted in the elevation of intracellular Ca2+, activation of JNK and c-Src, and disruption of TJ in the colonic epithelium. Furthermore, corticosterone administration induced JNK and c-Src activation, TJ disruption and protein thiol oxidation in colonic mucosa. The present study demonstrates that oxidative stress is a common signal in the mechanism of TJ disruption in the intestinal epithelium by different types of cellular stress in vitro and bio behavioral stress in vivo.

Occludin deficiency promotes ethanol-induced disruption of colonic epithelial junctions, gut barrier dysfunction and liver damage in mice.

BACKGROUND: Disruption of epithelial tight junctions (TJ), gut barrier dysfunction and endotoxemia play crucial role in the pathogenesis of alcoholic tissue injury. Occludin, a transmembrane protein of TJ, is depleted in colon by alcohol. However, it is unknown whether occludin depletion influences alcoholic gut and liver injury. METHODS: Wild type (WT) and occludin deficient (Ocln(-/-)) mice were fed 1-6% ethanol in Lieber-DeCarli diet. Gut permeability was measured by vascular-to-luminal flux of FITC-inulin. Junctional integrity was analyzed by confocal microscopy. Liver injury was assessed by plasma transaminase, histopathology and triglyceride analyses. The effect of occludin depletion on acetaldehyde-induced TJ disruption was confirmed in Caco-2 cell monolayers. RESULTS: Ethanol feeding significantly reduced body weight gain in Ocln(-/-) mice. Ethanol increased inulin permeability in colon of both WT and Ocln(-/-) mice, but the effect was 4-fold higher in Ocln(-/-) mice. The gross morphology of colonic mucosa was unaltered, but ethanol disrupted the actin cytoskeleton, induced redistribution of occludin, ZO-1, E-cadherin and ß-catenin from the junctions and elevated TLR4, which was more severe in Ocln(-/-) mice. Occludin knockdown significantly enhanced acetaldehyde-induced TJ disruption and barrier dysfunction in Caco-2 cell monolayers. Ethanol significantly increased liver weight and plasma transaminase activity in Ocln(-/-) mice, but not in WT mice. Histological analysis indicated more severe lesions and fat deposition in the liver of ethanol-fed Ocln(-/-) mice. Ethanol-induced elevation of liver triglyceride was also higher in Ocln(-/-) mice. CONCLUSION: This study indicates that occludin deficiency increases susceptibility to ethanol-induced colonic mucosal barrier dysfunction and liver damage in mice.

Quantitative analysis of nanoscale intranuclear structural alterations in hippocampal cells in chronic alcoholism via transmission electron microscopy imaging.

Chronic alcoholism is known to alter the morphology of the hippocampus, an important region of cognitive function in the brain. Therefore, to understand the effect of chronic alcoholism on hippocampal neural cells, we employed a mouse model of chronic alcoholism and quantified intranuclear nanoscale structural alterations in these cells. Transmission electron microscopy (TEM) images of hippocampal neurons were obtained, and the degree of structural alteration in terms of mass density fluctuation was determined using the light-localization properties of optical media generated from TEM imaging. The results, which were obtained at length scales ranging from ~30 to 200 nm, show that 10-12 week-old mice fed a Lieber-DeCarli liquid (alcoholic) diet had a higher degree of structural alteration than control mice fed a normal diet without alcohol. The degree of structural alteration became significantly distinguishable at a sample length of ~100 nm, which is the typical length scale of the building blocks of cells, such as DNA, RNA, proteins and lipids. Interestingly, different degrees of structural alteration at such length scales suggest possible structural rearrangement of chromatin inside the nuclei in chronic alcoholism.

Physiological performance of two contrasting rice varieties under water stress.

Two rice varieties PR-115 and Super-7 were imposed to water stress and different physiological traits were monitored to evaluate the performance of these varieties under drought. Under water stress condition although the relative water content, osmotic potential, chlorophyll content, photosynthesis rate, carbon discrimination and biomass decreased in both the varieties however, the reduction was more pronounced in Super-7 variety. Oryzanol a trans-ester of ferulic acid functions as antioxidant and it increased along with total phenolic and anthocyanin content in both the varieties under drought stress. However, gallic acid, 4 hydroxy benzoic acid, syringic acid and chlorogenic acid showed differential pattern in both of the varieties under water limiting conditions. Under drought, grain yield was penalized by 17 and 54% in PR-115 and Super-7 varieties, respectively in comparison to watered plants. Super-7 variety showed pronounced electrolyte leakage and MDA enhancement under water stress condition. High non photochemical quenching and reduction in Y(NO) and Y(II) indicated balanced energy management in tolerant PR-115 variety. The studies showed that PR-115 is a drought tolerant variety while Super-7 is drought sensitive in nature.

Rapid disruption of intestinal epithelial tight junction and barrier dysfunction by ionizing radiation in mouse colon in vivo: protection by N-acetyl-l-cysteine.

The goals of this study were to evaluate the effects of ionizing radiation on apical junctions in colonic epithelium and mucosal barrier function in mice in vivo. Adult mice were subjected to total body irradiation (4 Gy) with or without N-acetyl-l-cysteine (NAC) feeding for 5 days before irradiation. At 2-24 h postirradiation, the integrity of colonic epithelial tight junctions (TJ), adherens junctions (AJ), and the actin cytoskeleton was assessed by immunofluorescence microscopy and immunoblot analysis of detergent-insoluble fractions for TJ and AJ proteins. The barrier function was evaluated by measuring vascular-to-luminal flux of fluorescein isothiocyanate (FITC)-inulin in vivo and luminal-to-mucosal flux in vitro. Oxidative stress was evaluated by measuring protein thiol oxidation. Confocal microscopy showed that radiation caused redistribution of occludin, zona occludens-1, claudin-3, E-cadherin, and ß-catenin, as well as the actin cytoskeleton as early as 2 h postirradiation, and this effect was sustained for at least 24 h. Feeding NAC before irradiation blocked radiation-induced disruption of TJ, AJ, and the actin cytoskeleton. Radiation increased mucosal permeability to inulin in colon, which was blocked by NAC feeding. The level of reduced-protein thiols in colon was depleted by radiation with a concomitant increase in the level of oxidized-protein thiol. NAC feeding blocked the radiation-induced protein thiol oxidation. These data demonstrate that radiation rapidly disrupts TJ, AJ, and the actin cytoskeleton by an oxidative stress-dependent mechanism that can be prevented by NAC feeding.

Chronic ethanol feeding promotes azoxymethane and dextran sulfate sodium-induced colonic tumorigenesis potentially by enhancing mucosal inflammation.

BACKGROUND: Alcohol consumption is one of the major risk factors for colorectal cancer. However, the mechanism involved in this effect of alcohol is unknown. METHODS: We evaluated the effect of chronic ethanol feeding on azoxymethane and dextran sulfate sodium (AOM/DSS)-induced carcinogenesis in mouse colon. Inflammation in colonic mucosa was assessed at a precancerous stage by evaluating mucosal infiltration of neutrophils and macrophages, and analysis of cytokine and chemokine gene expression. RESULTS: Chronic ethanol feeding significantly increased the number and size of polyps in colon of AOM/DSS treated mice. Confocal microscopic and immunoblot analyses showed a significant elevation of phospho-Smad, VEGF and HIF1α in the colonic mucosa. RT-PCR analysis at a precancerous stage indicated that ethanol significantly increases the expression of cytokines IL-1α, IL-6 and TNFα, and the chemokines CCL5/RANTES, CXCL9/MIG and CXCL10/IP-10 in the colonic mucosa of AOM/DSS treated mice. Confocal microscopy showed that ethanol feeding induces a dramatic elevation of myeloperoxidase, Gr1 and CD68-positive cells in the colonic mucosa of AOM/DSS-treated mice. Ethanol feeding enhanced AOM/DSS-induced suppression of tight junction protein expression and elevated cell proliferation marker, Ki-67 in the colonic epithelium. CONCLUSION: This study demonstrates that chronic ethanol feeding promotes colonic tumorigenesis potentially by enhancing inflammation and elevation of proinflammatory cytokines and chemokines.

Host-mediated gene silencing of a single effector gene from the potato pathogen Phytophthora infestans imparts partial resistance to late blight disease.

RNA interference (RNAi) has proved a powerful genetic tool for silencing genes in plants. Host-induced gene silencing of pathogen genes has provided a gene knockout strategy for a wide range of biotechnological applications. The RXLR effector Avr3a gene is largely responsible for virulence of oomycete plant pathogen Phytophthora infestans. In this study, we attempted to silence the Avr3a gene of P. infestans through RNAi technology. The P. infestans inoculation resulted in lower disease progression and a reduction in pathogen load, as demonstrated by disease scoring and quantification of pathogen biomass in terms of Pi08 repetitive elements, respectively. Transgenic plants induced moderate silencing of Avr3a, and the presence and/or expression of small interfering RNAs, as determined through Northern hybridization, indicated siRNA targeted against Avr3a conferred moderate resistance to P. infestans. The single effector gene did not provide complete resistance against P. infestans. Although the Avr3a effector gene could confer moderate resistance, for complete resistance, the cumulative effect of effector genes in addition to Avr3a needs to be considered. In this study, we demonstrated that host-induced RNAi is an effective strategy for functional genomics in oomycetes.

ALDH2 Deficiency Promotes Ethanol-Induced Gut Barrier Dysfunction and Fatty Liver in Mice.

BACKGROUND: Acetaldehyde, the toxic ethanol (EtOH) metabolite, disrupts intestinal epithelial barrier function. Aldehyde dehydrogenase (ALDH) detoxifies acetaldehyde into acetate. Subpopulations of Asians and Native Americans show polymorphism with loss-of-function mutations in ALDH2. We evaluated the effect of ALDH2 deficiency on EtOH-induced disruption of intestinal epithelial tight junctions and adherens junctions, gut barrier dysfunction, and liver injury. METHODS: Wild-type and ALDH2-deficient mice were fed EtOH (1 to 6%) in Lieber-DeCarli diet for 4 weeks. Gut permeability in vivo was measured by plasma-to-luminal flux of FITC-inulin, tight junction and adherens junction integrity was analyzed by confocal microscopy, and liver injury was assessed by the analysis of plasma transaminase activity, histopathology, and liver triglyceride. RESULTS: EtOH feeding elevated colonic mucosal acetaldehyde, which was significantly greater in ALDH2-deficient mice. ALDH2(-/-) mice showed a drastic reduction in the EtOH diet intake. Therefore, this study was continued only in wild-type and ALDH2(+/-) mice. EtOH feeding elevated mucosal inulin permeability in distal colon, but not in proximal colon, ileum, or jejunum of wild-type mice. In ALDH2(+/-) mice, EtOH-induced inulin permeability in distal colon was not only higher than that in wild-type mice, but inulin permeability was also elevated in the proximal colon, ileum, and jejunum. Greater inulin permeability in distal colon of ALDH2(+/-) mice was associated with a more severe redistribution of tight junction and adherens junction proteins from the intercellular junctions. In ALDH2(+/-) mice, but not in wild-type mice, EtOH feeding caused a loss of junctional distribution of tight junction and adherens junction proteins in the ileum. Histopathology, plasma transaminases, and liver triglyceride analyses showed that EtOH-induced liver damage was significantly greater in ALDH2(+/-) mice compared to wild-type mice. CONCLUSIONS: These data demonstrate that ALDH2 deficiency enhances EtOH-induced disruption of intestinal epithelial tight junctions, barrier dysfunction, and liver damage.

Retraction Note to: Pathogen virulence of Phytophthora infestans: from gene to functional genomics.

[This retracts the article DOI: 10.1007/s12298-012-0157-z.].

Sex-Specific Effect of Ethanol on Colon Content Lipidome in a Mice Model Using Nontargeted LC/MS.

Consumption of alcohol has widespread effects on the human body. The organs that are most significantly impacted are the liver and digestive system. When alcohol is consumed, it is absorbed in the intestines and processed by the liver. However, excessive alcohol use may affect gut epithelial integrity, microbiome composition, and lipid metabolism. Despite past studies investigating the effect of ethanol on hepatic lipid metabolism, the focus on colonic lipid metabolism has not been well explored. In this study, we investigated the sex-specific effect of ethanol on the colonic content lipidome in a mouse model using nontargeted liquid chromatography-mass spectrometry. Comprehensive lipidome analysis of colonic flush samples was performed using ethanol-fed (EF) and pair-fed (PF) mice of each sex. Partial least-squares discriminant analysis revealed that ethanol altered colonic lipid composition largely in male mice compared with female mice. A significant increase in free fatty acids, ceramides, and hexosylceramides and decreased phosphatidylglycerols (PG) was observed in the EF group compared to the PF group in male mice. Phosphatidylethanolamine (PE) levels were increased significantly in the EF group of both sexes compared to the PF group. The volcanic plot shows that PG (O-15:1/15:0) and PE (O-18:2/15:0) are common markers that are increased in both sexes of the EF group. In addition, decreased fatty acid esters of hydroxy fatty acids (FAHFA) were observed specifically in the EF group of female mice. Overall, a significant variation in the mice colonic content lipidome between the EF and PF groups was observed. Target pathways, such as sphingolipid metabolism in males, FAHFA in females, and PE metabolism in both sexes, were suggested. This study provides new insight into the sex-dependent lipid change associated with alcohol-induced gut-microbiota dysfunction and its potential health impacts.

Low-dose thyroxine attenuates autism-associated adverse effects of fetal alcohol in male offspring's social behavior and hippocampal gene expression.

BACKGROUND: Fetal alcohol spectrum disorder (FASD) is characterized by neurodevelopmental anomalies manifesting in cognitive and behavioral deficits in the offspring with diverse severities. Social behavior is affected in FASD, and these deficits overlap with those of autism spectrum disorder (ASD). Identifying some of the molecular characteristics related to ASD in an animal model of FASD could ultimately provide details on the underlying molecular mechanisms of both disorders that could lead to novel treatments. METHODS: Pregnant Sprague-Dawley rats received the following diets: control (C; ad libitum standard laboratory chow), nutritional control pair-fed (PF), ethanol (EtOH), or an EtOH diet supplemented with 0.3, 1.5, or 7.5 mg thyroxine (T4)/l in the diet. Social behavior and memory were tested in the adult offspring. Plasma total T4, free T3 (fT3), and thyroid-stimulating hormone (TSH) levels were measured. Hippocampal expression of Gabrb3, Ube3a, Nr2b, Rasgrf1, and Dio3 were measured by RT-qPCR and protein levels of Mecp2 and Slc25a12 by Western blotting. RESULTS: Adult male offspring of EtOH dams showed elevated fT3 and low TSH levels. Adult male, but not female, offspring of EtOH dams exhibited social behavior and memory deficits. Expression of autism candidates, Gabrb3, Ube3a, Mecp2, and Slc25a12, was significantly increased in the hippocampus of male offspring of EtOH dams. Hippocampal Nr2b and Dio3 were also increased, while Rasgrf1 was decreased in the same population. Peripheral thyroid function, social behavioral deficits, and altered expression of the above genes were normalized by simultaneous administration of 0.3 mg/l T4 in the EtOH diet. CONCLUSIONS: Our data suggest that social interaction deficits of FASD share molecular mechanism with ASD by showing altered hippocampal expression of several ASD candidate genes. Social interaction deficits as well as the gene expression changes in the offspring of EtOH-consuming dams can be reversed by low dose of thyroid hormone supplementation to the mothers.