Shubha Priyamvada

A novel nutrient sensing mechanism underlies substrate-induced regulation of monocarboxylate transporter-1

Monocarboxylate transporter isoform-1 (MCT1) plays an important role in the absorption of short-chain fatty acids (SCFAs) in the colon. Butyrate, a major SCFA, serves as the primary energy source for the colonic mucosa, maintains epithelial integrity, and ameliorates intestinal inflammation. Previous studies have shown substrate (butyrate)-induced upregulation of MCT1 expression and function via transcriptional mechanisms. The present studies provide evidence that short-term MCT1 regulation by substrates could be mediated via a novel nutrient sensing mechanism. Short-term regulation of MCT1 by butyrate was examined in vitro in human intestinal C2BBe1 and rat intestinal IEC-6 cells and ex vivo in rat intestinal mucosa. Effects of pectin feeding on MCT1, in vivo, were determined in rat model. Butyrate treatment (30-120 min) of C2BBe1 cells increased MCT1 function {p-(chloromercuri) benzene sulfonate (PCMBS)-sensitive [(14)C]butyrate uptake} in a pertussis toxin-sensitive manner. The effects were associated with decreased intracellular cAMP levels, increased V(max) of butyrate uptake, and GPR109A-dependent increase in apical membrane MCT1 level. Nicotinic acid, an agonist for the SCFA receptor GPR109A, also increased MCT1 function and decreased intracellular cAMP. Pectin feeding increased apical membrane MCT1 levels and nicotinate-induced transepithelial butyrate flux in rat colon. Our data provide strong evidence for substrate-induced enhancement of MCT1 surface expression and function via a novel nutrient sensing mechanism involving GPR109A as a SCFA sensor.

Gene-environment interactions in heavy metal and pesticide carcinogenesis

Cancer is a complex disease involving a sequence of gene-environment interactions. Lifestyle, genetics, dietary factors, and environmental pollutants can increase the risk of cancer. Gene-environment interactions have been studied by a candidate-gene approach focusing on metabolism, DNA repair, and apoptosis. Here, we review the influence of gene-environment interactions in carcinogenesis, with emphasis on heavy metal and pesticide exposures.

Mechanisms Underlying Dysregulation of Electrolyte Absorption in Inflammatory Bowel Disease-Associated Diarrhea.

Abstract:Inflammatory bowel diseases (IBDs), including Crohns disease and ulcerative colitis, are chronic relapsing inflammatory disorders of the gastrointestinal tract. Chronic inflammation of the intestine affects the normal fluid and electrolyte absorption leading to diarrhea, the hallmark symptom of IBD. The management of IBD-associated diarrhea still remains to be a challenge, and extensive studies over the last 2 decades have focused on investigating the molecular mechanisms underlying IBD-associated diarrhea. These studies have shown that the predominant mechanism of diarrhea in IBD involves impairment of electroneutral NaCl absorption, with very little role if any played by anion secretion. The electroneutral NaCl absorption involves coupled operation of Na+/H+ exchanger 3 (NHE3 or SLC9A3) and Cl−/HCO3− exchanger DRA (Down Regulated in Adenoma, or SLC26A3). Increasing evidence now supports the critical role of a marked decrease in NHE3 and DRA function and/or expression in IBD-associated diarrhea. This review provides a detailed analysis of the current knowledge related to alterations in NHE3 and DRA function and expression in IBD including the mechanisms underlying these observations and highlights the potential of these transporters as important and novel therapeutic targets.

Targeting Parkinson’s - Tyrosine Hydroxylase and Oxidative Stress as Points of Interventions

Parkinsons disease (PD) is characterized by the progressive loss of the dopaminergic neurons leading to decrease in striatal dopamine (DA) levels. In the present review, our focus was on recent advances in the treatment procedures of PD to achieve an increase in deficient tyrosine hydroxylase (TH) activity and/or expression. Stimulation of residual TH activity by the cofactors, 6R-L-erythro-tetrahydrobiopterin (BPH4) or NADH, or by brain transplant of natural TH-containing cells (fetal substantia nigra) or genetically engineered TH-containing cells, has been tried experimentally and clinically lately. As a promising approach to the gene therapy, intrastriatal expression of DAsynthesizing enzymes through transduction with separate adeno-associated virus (AAV) vectors/ marrow stromal cells (MSCs) or nonviral intravenous administration of rat transferrin receptor monoclonal antibody (TfRmAb)-targeted PEGylated immunoliposomes (PILs) has been found to be effective in animal models. Oxidative stress has been identified as one of the intermediary risk factors that could initiate and/or promote degeneration of DA neurons. TH itself is a prime target of oxidative/nitrosative injury. Certain superoxide dismutase and catalase mimetic prevented nitration of TH in cultured dopaminergic neurons. Therefore, development of therapeutic agents that can prevent formation of or specifically remove nitrating agents without interfering with normal neuronal function may protect protein from inactivation and provide means of limiting neuronal injury in PD. Non-pharmacological approaches such as diet therapy or use of active constituents of plants and phytomedicines have also emerged as a new - area of high interest. New treatment strategies for TH dysfunction rectification, a provision for neuroprotection in PD, seem to be on the horizon with many therapies under investigation.

A novel anti-inflammatory role of GPR120 in intestinal epithelial cells

GPR120 (free fatty acid receptor-4) is a G protein-coupled receptor for medium- and long-chain unsaturated fatty acids, including ω-3 fatty acids. Recent studies have shown GPR120 to play cardinal roles in metabolic disorders via modulation of gut hormone secretion and insulin sensitivity and to exert anti-inflammatory effects in macrophages and adipose tissues. However, information on anti-inflammatory role of GPR120 at the level of intestinal epithelium is very limited. Current studies demonstrated differential levels of GPR120 mRNA and protein along the length of the human, mouse, and rat intestine and delineated distinct anti-inflammatory responses following GPR120 activation in model human intestinal epithelial Caco-2 cells, but not in model mouse intestinal epithelial endocrine cell line STC-1. In Caco-2 cells, GPR120 was internalized, bound to β-arrestin-2, and attenuated NF-κB activation in response to 30-min exposure to the agonists GW9508, TUG-891, or docosahexaenoic acid. These effects were abrogated in response to small interfering RNA silencing of β-arrestin-2. Treatment of STC-1 cells with these agonists did not induce receptor internalization and had no effects on NF-κB activation, although treatment with the agonists GW9508 or TUG-891 for 6 h augmented the synthesis and secretion of the gut hormone glucagon-like peptide-1 in this cell line. Our studies for the first time demonstrated a GPR120-mediated novel anti-inflammatory pathway in specific intestinal epithelial cell types that could be of therapeutic relevance to intestinal inflammatory disorders.

Lactobacillus acidophilus attenuates downregulation of DRA function and expression in inflammatory models

Probiotics, including Lactobacilli, are commensal bacteria that have been used in clinical trials and experimental models for the prevention and treatment of diarrheal disorders. Our previous studies have shown that Lactobacillus acidophilus (LA) and its culture supernatant (CS) stimulated Cl(-)/HCO3 (-) exchange activity, acutely via an increase in the surface levels of downregulated in adenoma (DRA, SLC26A3) and in long-term treatments via increasing its expression involving transcriptional mechanisms. However, the role of LA in modulating DRA activity under inflammatory conditions is not known. Current in vitro studies using human intestinal epithelial Caco-2 cells examined the efficacy of LA or its CS in counteracting the inhibitory effects of interferon-γ (IFN-γ) on Cl(-)/HCO3 (-) exchange activity. Pretreatment of cells with LA or LA-CS for 1 h followed by coincubation with IFN-γ significantly alleviated the inhibitory effects of IFN-γ on Cl(-)/HCO3 (-) exchange activity. In the in vivo model of dextran sulfate sodium-induced experimental colitis (3% in drinking water for 7 days) in C57BL/6J mice, administration of live LA (3 × 10(9) colony-forming units) via oral gavage attenuated colonic inflammation. LA administration also counteracted the colitis-induced decrease in DRA mRNA and protein levels. Efficacy of LA or its secreted soluble factors in alleviating inflammation and inflammation-associated dysregulation of DRA activity could justify their therapeutic potential in inflammatory diarrheal diseases.

LPA stimulates intestinal DRA gene transcription via LPA2 receptor, PI3K/AKT, and c-Fos-dependent pathway

DRA (downregulated in adenoma) or SLC26A3 is the major apical anion exchanger mediating Cl(-) absorption in intestinal epithelial cells. Disturbances in DRA function and expression have been implicated in diarrheal conditions such as congenital chloride diarrhea and inflammatory bowel diseases. Previous studies have shown that DRA is subject to regulation by short-term and transcriptional mechanisms. In this regard, we have recently shown that short-term treatment by lysophosphatidic acid (LPA), an important bioactive phospholipid, stimulates Cl(-)/HCO(3)(-)(OH(-)) exchange activity via an increase in DRA surface levels in human intestinal epithelial cells. However, the long-term effects of LPA on DRA at the level of gene transcription have not been examined. The present studies were aimed at investigating the effects of LPA on DRA function and expression as well as elucidating the mechanisms underlying its transcriptional regulation. Long-term LPA treatment increased the Cl(-)/HCO(3)(-) exchange activity in Caco-2 cells. LPA treatment (50-100 μM) of Caco-2 cells significantly stimulated DRA mRNA levels and DRA promoter activity (-1183/+114). This increase in DRA promoter activity involved the LPA2 receptor and phosphatidylinositol 3-kinase (PI3K)/AKT pathways. Progressive deletions from -1183/+114 to -790/+114 abrogated the stimulatory effects of LPA, indicating that the -1183/-790 promoter region harbors LPA response elements. Utilizing EMSA and mutational studies, our results showed that LPA induced the DRA promoter activity in a c-Fos-dependent manner. LPA also increased the protein expression of c-Fos and c-Jun in Caco-2 cells. Furthermore, overexpression of c-Fos but not c-Jun enhanced the DRA promoter activity. This increase in DRA transcription in response to LPA indicates that LPA may act as an antidiarrheal agent and could be exploited for the treatment of diarrhea associated with inflammatory or infectious diseases of the gut.

Linking Alzheimer’s Disease and Type 2 Diabetes Mellitus via Aberrant Insulin Signaling and Inflammation

Alzheimers disease (AD) and type 2 diabetes mellitus (T2DM) are two progressive and devastating health disorders afflicting millions of people worldwide. The probability and incidence of both have increased considerably in recent years consequent to increased longevity and population growth. Progressively more links are being continuously found between inflammation and central nervous system disorders like AD, Parkinsons disease, Huntingtons disease, motor neuron disease, multiple sclerosis, stroke, traumatic brain injury and even cancers of the nervous tissue. The depth of the relationship depends on the timing and extent of anti- or pro-inflammatory gene expression. Inflammation has also been implicated in T2DM. Misfolding and fibrillization (of tissue specific and/or non-specific proteins) are features common to both AD and T2DM and are induced by as well as contribute to inflammation and stress (oxidative/ glycation). This review appraises the roles of inflammation and abnormalities in the insulin signaling system as important shared features of T2DM and AD. The capacity of anti-cholinesterases in reducing the level of certain common inflammatory markers in particular if they may provide therapeutic potential to mitigate awry mechanisms leading to AD.

Probiotic Bifidobacterium species stimulate human SLC26A3 gene function and expression in intestinal epithelial cells.

SLC26A3, or downregulated in adenoma (DRA), plays a major role in mediating Cl(-) absorption in the mammalian intestine. Disturbances in DRA function and expression have been implicated in intestinal disorders such as congenital Cl(-) diarrhea and gut inflammation. We previously showed that an increase in DRA function and expression by Lactobacillus acidophilus and its culture supernatant (CS) might underlie antidiarrheal effects of this probiotic strain. However, the effects of Bifidobacterium species, important inhabitants of the human colon, on intestinal Cl(-)/HCO3 (-) exchange activity are not known. Our current results demonstrate that CS derived from Bifidobacterium breve, Bifidobacterium infantis, and Bifidobacterium bifidum increased anion exchange activity in Caco-2 cells (∼1.8- to 2.4-fold). Consistent with the increase in DRA function, CS also increased the protein, as well as the mRNA, level of DRA (but not putative anion transporter 1). CS of all three Bifidobacterium sp. increased DRA promoter activity (-1,183/+114 bp) in Caco-2 cells (1.5- to 1.8-fold). Furthermore, the increase in DRA mRNA expression by CS of B. breve and B. infantis was blocked in the presence of the transcription inhibitor actinomycin D (5 μM) and the ERK1/2 MAPK pathway inhibitor U0126 (10 μM). Administration of live B. breve, B. infantis, and B. bifidum by oral gavage to mice for 24 h increased DRA mRNA and protein levels in the colon. These data demonstrate an upregulation of DRA via activation of the ERK1/2 pathway that may underlie potential antidiarrheal effects of Bifidobacterium sp.

Lactobacillus acidophilus Counteracts Inhibition of NHE3 and DRA Expression and Alleviates Diarrheal Phenotype in Mice Infected With Citrobacter rodentium.

Impaired absorption of electrolytes is a hallmark of diarrhea associated with inflammation or enteric infections. Intestinal epithelial luminal membrane NHE3 (Na+/H+ exchanger 3) and DRA (Down-Regulated in Adenoma; Cl-/HCO3- exchanger) play key roles in mediating electroneutral NaCl absorption. We have previously shown decreased NHE3 and DRA function in response to short-term infection with enteropathogenic E coli (EPEC), a diarrheal pathogen. Recent studies have also shown substantial downregulation of DRA expression in a diarrheal model of infection with Citrobacter rodentium, the mouse counterpart of EPEC. Since our previous studies showed that the probiotic Lactobacillus acidophilus (LA) increased DRA and NHE3 function and expression and conferred protective effects in experimental colitis, we sought to evaluate the efficacy of LA in counteracting NHE3 and DRA inhibition and ameliorating diarrhea in a model of C rodentium infection. FVB/N mice challenged with C rodentium [1 × 109 colony-forming units (CFU)] with or without administration of live LA (3 × 109 CFU) were assessed for NHE3 and DRA mRNA and protein expression, mRNA levels of carbonic anhydrase, diarrheal phenotype (assessed by colonic weight-to-length ratio), myeloperoxidase activity, and proinflammatory cytokines. LA counteracted C rodentium-induced inhibition of colonic DRA, NHE3, and carbonic anhydrase I and IV expression and attenuated diarrheal phenotype and MPO activity. Furthermore, LA completely blocked C rodentium induction of IL-1β, IFN-γ, and CXCL1 mRNA and C rodentium-induced STAT3 phosphorylation. In conclusion, our data provide mechanistic insights into antidiarrheal effects of LA in a model of infectious diarrhea and colitis.