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Fake news is misinformation or distorted news that is spread over social media with the intent of harming an individual, organization, or government body. Collecting news from an internet platform is simple because it is quick, user-friendly, and constantly updated. However, this information is also subject to personal preferences or interests, which can negatively affect individuals or organizations. As a result, it is critical to detect the spread of fake news by computational methods. Semantic web approaches also plays an important role in detecting false news since they are used to understand the data that is available online in the same manner that humans do. As a result, this research examined various learning models for classifying whether the information is true or false using a fake and real news dataset. Around 40,000 items were analyzed, including approximately 20,000 from each fake and actual news dataset. Ensemble learning models such as support vector machine, logistic regression, CatBoost, XgBoost, multinomial, Naive Bayes, and random forest were used and evaluated using recall, accuracy, false rejection rate (FRR), F1 score, precision, negative predictive value (NPV), false discovery rate (FDR), and Matthews' correlation coefficient. Based on these evaluations, the best learning models were hybridized and computed alongside the passive-aggressive classifier and deep Auto_ViML model. Following computation, it was determined that the deep Auto_ViML model had the highest accuracy, precision, recall, and F1 score of 99%. In contrast, the hybrid learning model had the best value of false rejection rate of 71%. In comparison, the support vector machine was computed in 0.000245 seconds.

Introduction: The novel corona virus (COVID-19), a pandemic, spread to around 198 countries, and has affected millions of people globally. The associated morbidity and mortality challenged the nations in several ways. One such unaddressed area is the mental health impact on the healthcare workers and staff during this pandemic. The review aims at summarizing the evidence of mental health issues of COVID-19 pandemic on the frontline healthcare workers (HCW’s). Materials and methods: A review of the literature of the mental health issues faced by the healthcare workers during the pandemic was done. Literature search was conducted in the following databases: PubMed, Google Scholar, Embase; articles relevant to the subject, in the duration of last 8 months (Jan-2020 to Aug-2020) were reviewed. We have highlighted the most relevant data concerning the disease characteristics, personal and organizational factors that may have contributed to development of psychological changes, distress and mental health symptoms. Results: There is a generalized climate of wariness and uncertainty, particularly amongst the health professionals, which has been provoked by the disease characteristics of the current COVID-19 pandemic and various sociodemographic variables. Other factors included, rapid spread of disease, severity of symptoms, lack of knowledge of the disease, organizational factors and death among healthcare professionals. The mental health problems varied with respect to the sociodemographic variables like, gender, profession, age, place of work, social support and department of work. Evidence suggests that COVID-19 may be an independent risk factor for increase in stress, anxiety, burnout and depression amongst the healthcare professionals, which could have long-term psychological implications. Conclusion: It is of utmost importance to protect and ensure the mental health of the healthcare professionals, to address large scale health crisis. Therefore, planning of future prevention strategies is essential to promote mental well-being. Implementation to bolster response and prevention strategies by training healthcare professionals on crisis management and mental help. Evaluation of mental health status should be done by regular screening of the personnel by the multidisciplinary Psychiatry teams.

The serotonin transporter (SERT) functions to regulate the availability of serotonin (5-HT) in the brain and intestine. An intestine-specific mRNA variant arising from a unique transcription start site and alternative promoter in the SERT gene has been identified (iSERT; spanning exon 1C). A decrease in SERT is implicated in several gut disorders, including inflammatory bowel diseases (IBD). However, little is known about mechanisms regulating the iSERT variant, and a clearer understanding is warranted for targeting SERT for the treatment of gut disorders. The current studies examined the expression of iSERT across different human intestinal regions and investigated its regulation by HNF4α (hepatic nuclear factor-4α), a transcription factor important for diverse cellular functions. iSERT mRNA abundance was highest in the human ileum and Caco-2 cell line. iSERT mRNA expression was downregulated by loss of HNF4α (but not HNF1α, HNF1β, or FOXA1) in Caco-2 cells. Overexpression of HNF4α increased iSERT mRNA concomitant with an increase in SERT protein. Progressive promoter deletion and site-directed mutagenesis revealed that the HNF4α response element spans nucleotides −1,163 to −1150 relative to the translation start site. SERT mRNA levels in the intestine were drastically reduced in the intestine-specific HNF4α-knockout mice relative to HNF4αFL/FL mice. Both HNF4α and SERT mRNA levels were also downregulated in mouse model of ileitis (SAMP) compared with AKR control mice. These results establish the transcriptional regulation of iSERT at the gut-specific internal promoter (hSERTp2) and have identified HNF4α as a critical modulator of basal SERT expression in the intestine.

The ileal apical sodium-dependent bile acid transporter (ASBT) is crucial for the enterohepatic circulation of bile acids. ASBT function is rapidly regulated by several posttranslational modifications. One reversible posttranslational modification is S-acylation, involving the covalent attachment of fatty acids to cysteine residues in proteins. However, whether S-acylation affects ASBT function and membrane expression has not been determined. Using the acyl resin-assisted capture method, we found that the majority of ASBT (∼80%) was S-acylated in ileal brush border membrane vesicles from human organ donors, as well as in HEK293 cells stably transfected with ASBT (2BT cells). Metabolic labeling with alkyne–palmitic acid (100 μm for 15 h) also showed that ASBT is S-acylated in 2BT cells. Incubation with the acyltransferase inhibitor 2-bromopalmitate (25 μm for 15 h) significantly reduced ASBT S-acylation, function, and levels on the plasma membrane. Treatment of 2BT cells with saturated palmitic acid (100 μm for 15 h) increased ASBT function, whereas treatment with unsaturated oleic acid significantly reduced ASBT function. Metabolic labeling with alkyne–oleic acid (100 μm for 15 h) revealed that oleic acid attaches to ASBT, suggesting that unsaturated fatty acids may decrease ASBT's function via a direct covalent interaction with ASBT. We also identified Cys-314 as a potential S-acylation site. In conclusion, these results provide evidence that S-acylation is involved in the modulation of ASBT function. These findings underscore the potential for unsaturated fatty acids to reduce ASBT function, which may be useful in disorders in which bile acid toxicity is implicated.

Introduction: The diagnosis of parotid tuberculosis requires a high degree of clinical suspicion, so as to avoid unnecessary surgery, and fine-needle aspiration cytology is an easy and reliable investigation for the same. Salivary glands are suitable for unguided fine needle aspiration cytology (FNAC) because of their superficial location. Methods: The present study involved a total of 42 cases of salivary gland swellings in which fine needle aspiration cytology was performed in the department of pathology of our hospital from December 2011 to August 2019. Results: The prevalence of granulomatous inflammation among salivary gland swellings was 38.1 % (n=16). Anti-tubercular treatment was given in 5 patients, all of whom showed improvement after anti-tubercular treatment. Therefore, 5 out of the total of 16 cases were a case of tuberculous parotitis (31.2 %). Conclusion: Although tuberculous parotitis is a rare entity, it should be always kept in mind as a differential during the evaluation of solitary parotid mass lesion. Fine needle aspiration cytology of all mass lesions of salivary glands must be done before doing excision of salivary gland so as to avoid unnecessary surgery.

Coronaviruses have led to three major outbreaks to date-Severe Acute Respiratory Syndrome (SARS; 2002), Middle East Respiratory Syndrome (MERS; 2012) and the ongoing pandemic, Coronavirus Disease (COVID-19; 2019). Coronavirus infections are usually mild in children. However, a few children with MERS had presented with a severe phenotype in the acute phase resulting in progressive pneumonic changes with increasing oxygen dependency and acute respiratory distress requiring ventilatory support. A subset of children with a history of SARS-CoV-2 infection develops a multisystem hyper-inflammatory phenotype known as Multisystem Inflammatory Syndrome in Children (MIS-C). This syndrome occurs 4-6 weeks after infection with SARS-CoV-2 and has been reported more often from areas with high community transmission. Children with MIS-C present with high fever and often have involvement of cardiovascular, gastrointestinal and hematologic systems leading to multiorgan failure. This is accompanied by elevation of pro-inflammatory cytokines such as IL-6 and IL-10. MIS-C has several similarities with Kawasaki disease (KD) considering children with both conditions present with fever, rash, conjunctival injection, mucosal symptoms and swelling of hands and feet. For reasons that are still not clear, both KD and MIS-C were not reported during the SARS-CoV and MERS-CoV outbreaks. As SARS-CoV-2 differs from SARS-CoV by 19.5% and MERS by 50% in terms of sequence identity, differences in genomic and proteomic profiles may explain the varied disease immunopathology and host responses. Left untreated, MIS-C may lead to severe abdominal pain, ventricular dysfunction and shock. Immunological investigations reveal reduced numbers of follicular B cells, increased numbers of terminally differentiated CD4+T lymphocytes, and decreased IL-17A. There is still ambiguity about the clinical and immunologic risk factors that predispose some children to development of MIS-C while sparing others. Host-pathogen interactions in SARS, MERS and COVID-19 are likely to play a crucial role in the clinical phenotypes that manifest. This narrative review focuses on the immunological basis for development of MIS-C syndrome in the ongoing SARS-CoV-2 pandemic. To the best of our knowledge, these aspects have not been reviewed before.

Niemann-Pick C1 Like-1 (NPC1L1) mediates the uptake of micellar cholesterol by intestinal epithelial cells and is the molecular target of the cholesterol-lowering drug ezetimibe (EZE). The detailed mechanisms responsible for intracellular shuttling of micellar cholesterol are not fully understood due to the lack of a suitable NPC1L1 substrate that can be traced by fluorescence imaging and biochemical methods. 27-Alkyne cholesterol has been previously shown to serve as a substrate for different cellular processes similar to native cholesterol. However, it is not known whether alkyne cholesterol is absorbed via an NPC1L1-dependent pathway. We aimed to determine whether alkyne cholesterol is a substrate for NPC1L1 in intestinal cells. Human intestinal epithelial Caco2 cells were incubated with micelles containing alkyne cholesterol in the presence or absence of EZE. Small intestinal closed loops in C57BL/6J mice were injected with micelles containing alkyne cholesterol with or without EZE. Alkyne cholesterol esterification in Caco2 cells was significantly inhibited by EZE and by inhibitor of clathrin-mediated endocytosis Pitstop 2. The esterification was similarly reduced by inhibitors of the acyl-CoA cholesterol acyltransferase (ACAT). Alkyne cholesterol efficiently labeled the apical membrane of Caco2 cells and the amount retained on the membrane was significantly increased by EZE as judged by accessibility to exogenous cholesterol oxidase. In mouse small intestine, the presence of EZE reduced total alkyne cholesterol uptake by ∼75%. These data show that alkyne cholesterol acts as a substrate for NPC1L1 and may serve as a nonradioactive tracer to measure cholesterol absorption in both in vitro and in vivo models.

Non-alcoholic fatty liver disease (NAFLD) is a major health problem associated with obesity and other features of the metabolic syndrome including insulin resistance and dyslipidemia. The accumulation of lipids in hepatocytes causes liver damage and triggers inflammation, fibrosis, and cirrhosis. Beside fatty acids and triglycerides, evidence showed an increased accumulation of free cholesterol in the liver with subsequent toxic effects contributing to liver damage. The maintenance of cholesterol homeostasis in the body requires a balance between several pathways responsible for cholesterol synthesis, transport and conversion into bile acids. Intestinal absorption is also one of the major determinants of cholesterol homeostasis. The nature of changes in cholesterol homeostasis associated with NAFLD has been a subject of extensive investigations. In this article, we will attempt to provide a brief overview of the current knowledge about the disturbances in cholesterol metabolism associated with NAFLD and discuss how certain molecular targets of these pathways could be exploited for the treatment of this multifactorial disease.

The intestinal reclamation of bile acids is crucial for the maintenance of their enterohepatic circulation. The majority of bile acids are actively absorbed via specific transport proteins that are highly expressed in the distal ileum. The uptake of bile acids by intestinal epithelial cells modulates the activation of cytosolic and membrane receptors such as the farnesoid X receptor (FXR) and G protein-coupled bile acid receptor 1 (GPBAR1), which has a profound effect on hepatic synthesis of bile acids as well as glucose and lipid metabolism. Extensive research has focused on delineating the processes of bile acid absorption and determining the contribution of dysregulated ileal signaling in the development of intestinal and hepatic disorders. For example, a decrease in the levels of the bile acid-induced ileal hormone FGF15/19 is implicated in bile acid-induced diarrhea (BAD). Conversely, the increase in bile acid absorption with subsequent overload of bile acids could be involved in the pathophysiology of liver and metabolic disorders such as fatty liver diseases and type 2 diabetes mellitus. This review article will attempt to provide a comprehensive overview of the mechanisms involved in the intestinal handling of bile acids, the pathological implications of disrupted intestinal bile acid homeostasis, and the potential therapeutic targets for the treatment of bile acid-related disorders. Published 2020. Compr Physiol 10:21-56, 2020.

Intestinal Niemann-Pick C1 Like 1 (NPC1L1) protein plays a key role in cholesterol absorption. A decrease in NPC1L1 expression has been implicated in lowering plasma cholesterol and mitigating the risk for coronary heart disease. Little is known about the mechanisms responsible for NPC1L1 protein degradation that upon activation may lead to a reduction in NPC1L1 protein levels in intestinal epithelial cells (IECs). In current studies, the human intestinal Caco-2 and HuTu-80 cell lines expressing NPC1L1-hemagglutinin fusion protein were used to investigate the mechanisms of NPC1L1 protein degradation. Incubation with the proteasome inhibitors MG-132 and lactacystin (10 μM, 24 h) significantly increased NPC1L1 protein levels in IECs. Also, the inhibition of the lysosomal pathway with bafilomycin A1 (80 nM, 24 h) resulted in a significant increase in NPC1L1 protein levels. Immunoprecipitation studies showed that NPC1L1 protein is both a poly- and monoubiquinated polypeptide and that the inhibition of the proteasomal pathway remarkably increased the level of the polyubiquinated NPC1L1. The surface expression of NPC1L1 was increased by the inhibition of both proteasomal and lysosomal pathways. Furthermore, the pharmacological inhibition of mitogen-activated protein kinase pathway (PD-98059, 15 μM, 24 h) and siRNA silencing of ERK1/2 resulted in a significant decrease in NPC1L1 protein levels in IECs. In conclusion, our results showed that basal level of intestinal cholesterol transporter NPC1L1 protein is modulated by both ubiquitin proteasome- and lysosome-dependent degradation as well as by ERK1/2-dependent pathway. The modulation of these pathways may provide novel clues for therapeutic intervention to inhibit cholesterol absorption and lower plasma cholesterol.

A thermodynamically and kinetically simple picture of protein folding envisages only two states, native (N) and unfolded (U), separated by a single activation free energy barrier, and interconverting by cooperative two-state transitions. The folding/unfolding transitions of many proteins occur, however, in multiple discrete steps associated with the formation of intermediates, which is indicative of reduced cooperativity. Furthermore, much advancement in experimental and computational approaches has demonstrated entirely non-cooperative (gradual) transitions via a continuum of states and a multitude of small energetic barriers between the N and U states of some proteins. These findings have been instrumental towards providing a structural rationale for cooperative versus noncooperative transitions, based on the coupling between interaction networks in proteins. The cooperativity inherent in a folding/unfolding reaction appears to be context dependent, and can be tuned via experimental conditions which change the stabilities of N and U. The evolution of cooperativity in protein folding transitions is linked closely to the evolution of function as well as the aggregation propensity of the protein. A large activation energy barrier in a fully cooperative transition can provide the kinetic control required to prevent the accumulation of partially unfolded forms, which may promote aggregation. Nevertheless, increasing evidence for barrier-less "downhill" folding, as well as for continuous "uphill" unfolding transitions, indicate that gradual non-cooperative processes may be ubiquitous features on the free energy landscape of protein folding.

The bile acid transporter ASBT is a glycoprotein responsible for active absorption of bile acids. Inhibiting ASBT function and bile acid absorption is an attractive approach to lower plasma cholesterol and improve glucose imbalance in diabetic patients. Deglycosylation of ASBT was shown to decrease its function. However, the exact roles of N-glycosylation of ASBT, and how it affects its function, is not known. Current studies investigated the roles of N-glycosylation in ASBT protein stability and protection against proteases utilizing HEK-293 cells stably transfected with ASBT-V5 fusion protein. ASBT-V5 protein was detected as two bands with molecular mass of ∼41 and ∼35 kDa. Inhibition of glycosylation by tunicamycin significantly decreased ASBT activity and shifted ASBT bands to ∼30 kDa, representing a deglycosylated protein. Treatment of total cellular lysates with PNGase F or Endo H glycosidases showed that the upper 41-kDa band represents a fully mature N-acetylglucosamine-rich glycoprotein and the lower 35-kDa band represents a mannose-rich core glycoprotein. Studies with the glycosylation deficient ASBT mutant (N10Q) showed that the N-glycosylation is not essential for ASBT targeting to plasma membrane. However, mature glycosylation significantly increased the half-life and protected ASBT protein from digestion with trypsin. Incubating the cells with high glucose (25 mM) for 48 h increased mature glycosylated ASBT along with an increase in its function. These results unravel novel roles for N-glycosylation of ASBT and suggest that high levels of glucose alter the composition of the glycan and may contribute to the increase in ASBT function in diabetes mellitus.

Na+/H+ exchanger-3 (NHE3) is crucial for intestinal Na+ absorption, and its reduction has been implicated in infectious and inflammatory bowel diseases (IBD)-associated diarrhea. Epigenetic mechanisms such as DNA methylation are involved in the pathophysiology of IBD. Whether changes in DNA methylation are involved in modulating intestinal NHE3 gene expression is not known. Caco-2 and HuTu 80 cells were used as models of human intestinal epithelial cells. Normal C57/BL6, wild-type, or growth arrest and DNA damage-inducible 45b (GADD45b) knockout (KO) mice were used as in vivo models. NHE3 gene DNA methylation levels were assessed by MBDCap (MethyMiner) assays. Results demonstrated that in vitro methylation of NHE3 promoter construct (p-1509/+127) cloned into a cytosine guanine dinucleotide-free lucia vector decreased the promoter activity in Caco-2 cells. DNA methyltransferase inhibitor 5-azacytidine (10 μM, 24 h) caused a significant decrease in DNA methylation of the NHE3 gene and concomitantly increased NHE3 expression in Caco-2 cells. Similarly, 5-azacytidine treatment increased NHE3 mRNA levels in HuTu 80 cells. 5-Azacytidine treatment for 3 wk (10 mg/kg body wt ip, 3 times/wk) also resulted in an increase in NHE3 expression in the mouse ileum and colon. Small-interfering RNA knockdown of GADD45b (protein involved in DNA demethylation) in Caco-2 cells decreased NHE3 mRNA expression. Furthermore, there was a significant decrease in NHE3 mRNA and protein expression in the ileum and colon of GADD45b KO mice. Our findings demonstrate that NHE3 gene expression is regulated by changes in its DNA methylation. NEW & NOTEWORTHY Our studies for the first time demonstrate that Na+/H+ exchanger-3 gene expression is regulated by an epigenetic mechanism involving DNA methylation.

Nonalcoholic fatty liver disease (NAFLD) is characterized by lipid accumulation in the liver that may progress to hepatic fibrosis and nonalcoholic steatohepatitis (NASH). Mechanisms underlying NAFLD and NASH are not yet fully understood. Dietary cholesterol was recently shown to be a risk factor for the development of NASH, suggesting a role for intestinal handling of cholesterol. One important regulator of cholesterol homeostasis is the sterol response element-binding protein-2 (SREBP-2) transcription factor. We tested the hypothesis that the overactivation of intestinal SREBP-2 increases the susceptibility to diet-induced NASH. A transgenic mouse model with intestine-specific overexpression of active SREBP-2 (ISR2 mice) driven by villin promoter was used. ISR2 mice and their wild-type littermates were fed a regular chow diet or a high-fat, high-cholesterol (HFHC) diet (15% fat, 1% cholesterol) for 15 wk. Results showed that HFHC feeding to ISR2 mice caused hepatic inflammation with increased levels of proinflammatory cytokines. Histological examination demonstrated extensive fibrosis after a HFHC diet associated with a perivascular as well as pericellular collagen deposits in ISR2 mice compared with wild-type littermates. The severe hepatic inflammation and advanced fibrosis in ISR2 mice was not associated with a difference in lipid accumulation in ISR2 mice compared with wild type littermates after HFHC feeding. These data indicate that overactivation of intestinal SREBP2 promotes diet-induced hepatic inflammation with features of human NASH resulting in rapid severe fibrosis and provide a novel link between regulatory processes of intestinal cholesterol and progression of fatty liver. NEW & NOTEWORTHY The current study highlights the role of overactivation of intestinal SREBP-2 transcription factor in the progression of hepatic fibrosis associated with diet-induced NASH. Mice with intestine-specific overexpression of SREBP-2 demonstrated more inflammation and severe fibrosis in the liver in response to 15 wk of being fed a high-cholesterol, high-fat diet as compared with their wild-type littermates. These data demonstrate a novel link between intestinal regulatory processes of cholesterol metabolism and the pathogenesis of fatty liver diseases.

Intestinal NPC1L1 transporter is essential for cholesterol absorption and the maintenance of cholesterol homeostasis in the body. NPC1L1 is differentially expressed along the gastrointestinal tract with very low levels in the colon as compared with the small intestine. This study was undertaken to examine whether DNA methylation was responsible for segment-specific expression of NPC1L1. Treatment of mice with 5-azacytidine (i.p.) resulted in a significant dose-dependent increase in NPC1L1 mRNA expression in the colon. The lack of expression of NPC1L1 in the normal colon was associated with high levels of methylation in the area flanking the 3-kb fragment upstream of the initiation site of the mouse NPC1L1 gene in mouse colon as analyzed by EpiTYPER® MassARRAY®. The high level of methylation in the colon was observed in specific CpG dinucleotides and was significantly decreased in response to 5-azacytidine. Similar to mouse NPC1L1, 5-azacytidine treatment also increased the level of human NPC1L1 mRNA expression in the intestinal HuTu-80 cell line in a dose- and time-dependent manner. Silencing the expression of DNA methyltransferase DNMT1, -2, -3A, and -3B alone by siRNA did not affect NPC1L1 expression in HuTu-80 cells. However, the simultaneous attenuation of DNMT1 and -3B expression caused a significant increase in NPC1L1 mRNA expression as compared with control. Also, in vitro methylation of the human NPC1L1 promoter significantly decreased NPC1L1 promoter activity in human intestinal Caco2 cells. In conclusion, our data demonstrated for the first time that DNA methylation in the promoter region of the NPC1L1 gene appears to be a major mechanism underlying differential expression of NPC1L1 along the length of the gastrointestinal tract.

The incidence of colorectal cancer (CRC) is increasing rapidly in Asian countries during the past few decades, but no comprehensive analysis has been done to find out the exact cause of this disease. In this study, we investigated the frequencies of mutations and expression pattern of K-ras, APC (adenomatosis polyposis coli) and p53 in tumor, adjoining and distant normal mucosa and to correlate these alterations with patients clinicopathological parameters as well as with the survival. Polymerase chain reaction (PCR)-restriction digestion was used to detect mutations in K-ras and PCR-SSCP (Single Strand Conformation Polymorphism) followed by DNA sequencing was used to detect mutations in APC and p53 genes. Immunohistochemistry was used to detect the expression pattern of K-ras, APC and p53 proteins. The frequencies of mutations of K-ras, APC and p53 in 30 tumor tissues samples were 26.7 %, 46.7 % and 20 %, respectively. Only 3.3 % of tumors contained mutations in all the three genes. The most common combination of mutation was APC and p53 whereas mutation in both p53 and K-ras were extremely rare. There was no association between the mutations and expression pattern of K-ras, APC and p53 (p>0.05). In Indians, the frequency of alterations of K-ras and APC is similar as in Westerns, whereas the frequency of p53 mutation is slightly lower. The lack of multiple mutations in tumor specimens suggests that these genetic alterations might have independent influences on CRC development and there could be multiple alternative genetic pathways to CRC in our present study cohort.

The serotonin (5-HT) transporter (SERT) facilitates clearance of extracellular 5-HT by its uptake and internalization. Decreased expression of SERT and consequent high 5-HT levels have been implicated in various diarrheal disorders. Thus, appropriate regulation of SERT is critical for maintenance of 5-HT homeostasis in health and disease. Previous studies demonstrated that SERT is regulated via posttranslational and transcriptional mechanisms. However, the role of epigenetic mechanisms in SERT regulation is not known. Current studies investigated the effects of histone deacetylase (HDAC) inhibition on SERT expression and delineated the mechanisms. Treatment of Caco-2 cells with the pan-HDAC inhibitors butyrate (5 mM) and trichostatin (TSA, 1 μM) decreased SERT mRNA and protein levels. Butyrate- or TSA-induced decrease in SERT was associated with decreased activity of human SERT (hSERT) promoter 1 (upstream of exon 1a), but not hSERT promoter 2 (upstream of exon 2). Butyrate + TSA did not show an additive effect on SERT expression, indicating that mechanisms involving histone hyperacetylation may be involved. Chromatin immunoprecipitation assays demonstrated enrichment of the hSERT promoter 1 (flanking nt −250/+2) with tetra-acetylated histone H3 or H4, which was increased (∼3-fold) by butyrate. Interestingly, specific inhibition of HDAC2 (but not HDAC1) utilizing small interfering RNA decreased SERT mRNA and protein levels. The decrease in SERT expression by HDAC inhibition was recapitulated in an in vivo model . SERT mRNA levels were decreased in the ileum and colon of mice fed pectin (increased availability of butyrate) compared with controls fed a fiber-free diet (∼50–60%). Our results identify a novel role of HDAC2 as a regulator of SERT gene expression in intestinal epithelial cells.

Curcumin, the major phenolic compound in the spice turmeric, exhibits numerous biological effects, including lowering plasma cholesterol and preventing diet-induced hypercholesterolemia. The mechanisms underlying the hypocholesterolemic effect of curcumin are not fully understood. In this regard, intestinal Niemann-Pick C1-like 1 (NPC1L1) cholesterol transporter, the molecular target of intestinal cholesterol absorption inhibitor ezetimibe, plays an essential role in the maintenance of cholesterol homeostasis. The current studies were designed to investigate the effect of curcumin on NPC1L1 function, expression, and promoter activity in intestinal Caco-2 monolayers. NPC1L1 function was evaluated by the measurement of ezetimibe-sensitive [3H]cholesterol esterification. Relative abundance of NPC1L1 mRNA and protein was evaluated by real-time PCR and Western blotting, respectively. Luciferase assays were used to measure NPC1L1 promoter activity. Our results showed that curcumin significantly inhibited ezetimibe-sensitive cholesterol esterification in a dose-dependent manner with a maximum decrease (by 52% compared with control) occurring at 50 μM concentration. Curcumin treatment of Caco-2 monolayers also significantly decreased NPC1L1 mRNA and protein expression. Similarly, the promoter activity of the NPC1L1 gene was inhibited significantly (55%) by 50 μM curcumin. The decrease in NPC1L1 promoter activity by curcumin was associated with a reduction in the expression and the DNA-binding activity of the sterol response element-binding protein 2 (SREBP2) transcription factor. Furthermore, the overexpression of active SREBP2 protected NPC1L1 from the inhibitory effect of curcumin. Our studies demonstrate that curcumin directly modulates intestinal NPC1L1 expression via transcriptional regulation and the involvement of SREBP2 transcription factor.

The role of Information Technology (IT) is indispensable in making the Indian economy globalised. Globalization has turned the whole world into a single market. Marketers have optimally utilized IT particularly internet and mobile applications to make their business global. The present paper is a descriptive paper analyzing the emerging trends and issues in IT particularly ecommerce sector in India in the wake of globalised economy. It tries to explore the challenges in the growth and development of this emerging sector and also suggest the ways to manage these challenges El papel de Information Technology (IT) – la tecnología de la información (TI) en español – es indispensable a la hora de hacer global la economía india. La globalización ha convertido todo el mundo en un mercado único. Los comerciantes han usado de forma óptima TI, en concreto internet y aplicaciones del móvil, para hacer sus negocios globales. Este artículo es un análisis descriptivo de las tendencias y problemas emergentes en TI, particularmente en el sector del comercio electrónico en India siguiendo los pasos de la economía universal. Este intenta explorar los retos en el crecimiento y desarrollo de este sector emergente, y también sugiere maneras para manejar estos.

In this study, we tested the hypothesis that the presence of dietary DHA in the sn-1 position of phosphatidylcholine (PC) results in greater incorporation of absorbed DHA into the phospholipids of plasma lipoproteins, compared to its presence in triacylglycerol (TG) or in the sn-2 position of PC, because of the positional specificities of the pancreatic enzymes. Micelle containing the DHA either in lyso PC (LPC) form (surrogate for the dietary sn-1 DHA PC) or as free fatty acid (surrogate for dietary DHA TG and sn-2 DHA PC) were infused intraduodenally into lymph fistula rats, and the concentration of DHA in PC and TG of lymph chylomicrons and HDL was determined by GC and LC/MS. The total amount of DHA absorbed was comparable with the two preparations. However the ratio of DHA in PC/TG in lymph was much higher (0.4) after LPC-DHA infusion, compared to free DHA infusion (0.1). Also, more DHA appeared in HDL fraction of lymph after LPC-DHA infusion, and more of the HDL DHA was in PC than in TG. Similar resu...

Non-Alcoholic Steatohepatitis (NASH) is characterized by steatosis with varying degrees of inflammation and fibrosis in the liver. The pathophysiology of NASH is not fully understood due to the lack of appropriate animal models mimicking the disease in humans. Recent studies showed that high cholesterol high fat (HCHF) diet induced NASH in mice but only after prolonged 30 weeks of feeding. We have recently shown that in transgenic mice [ISR2, with intestine-specific overexpression of active Sterol Response Binding Element Protein 2 (SREBP2)], intestinal cholesterol synthesis was significantly increased resulting in hypercholesterolemia. Since high cholesterol has been indicated to be a risk factor for NASH, we hypothesized that ISR2 mice will develop a more severe diet-induced NASH. Our data showed that HCHF feeding to ISR2 mice resulted in accelerated (15 wks versus 30 wks) and more severe NASH compared to wild type animals. HCHF feeding (15 wks) to ISR2 mice led to a significant increase in the hepatic ...

The Onto-Tools suite is composed of an annotation database and six seamlessly integrated, web-accessible data mining tools: Onto-Express, Onto-Compare, Onto-Design, Onto-Translate, Onto-Miner and Pathway-Express. The Onto-Tools database has been expanded to include various types of data from 12 new databases. Our database now integrates different types of genomic data from 19 sequence, gene, protein and annotation databases. Additionally, our database is also expanded to include complete Gene Ontology (GO) annotations. Using the enhanced database and GO annotations, Onto-Express now allows functional profiling for 24 organisms and supports 17 different types of input IDs. Onto-Translate is also enhanced to fully utilize the capabilities of the new Onto-Tools database with an ultimate goal of providing the users with a non-redundant and complete mapping from any type of identification system to any other type. Currently, Onto-Translate allows arbitrary mappings between 29 types of IDs. Pathway-Express is a new tool that helps the users find the most interesting pathways for their input list of genes. Onto-Tools are freely available at http://vortex.cs.wayne.edu/Projects.html.

To determine the etiological factors of human colorectal cancer (CRC) we assessed the frequency and prognostic significance of hMLH1 and hMSH2 genes in conjunction with hMLH1 and hMSH2 protein expression in 30 Indian CRC patients. The protein expression and promoter methylation of hMLH1 and hMSH2; Mismatch Repair genes (MMR) were analyzed by immunohistochemistry and methylation-specific PCR (MSP), respectively. A loss of hMLH1 expression was recognized in 4(13.3 %) and loss of hMSH2 expression was recognized in 2(6.6 %) of 30 CRC cases whereas 50 % tumors showed reduced expression of hMLH1 and 33.3 % showed reduced expression of hMSH2 protein. One tumor showed a loss of both hMLH1 and hMSH2 expression. Normal nuclear staining pattern of hMLH1 and hMSH2 was observed in almost all the adjoining and normal mucosa. Promoter hypermethylation of the hMLH1 gene was detected in 15 of 30 CRC cases (50 %) and of hMSH2 gene was only in 3 of 30 CRC cases (10 %). No promoter methylation of hMLH1 and hMSH2 genes was observed in adjoining and normal mucosa. Combination of methylation of hMLH1 and hMSH2 gene was observed in two tumors (6.6 %). A significant correlation between histological grade of the tumor, methylation and expression of hMLH1 gene (p

Sterol Response Element Binding Protein 2 (SREBP2) transcription factor is a master regulator of cholesterol homeostasis. Treatment with statins, inhibitors of cholesterol synthesis, activates intestinal SREBP2, which may hinder their cholesterol-lowering effects. Overactivation of SREBP2 in mouse liver was shown to have no effect on plasma cholesterol. However, the influence of activating intestinal SREBP2 on plasma cholesterol is not known. We have generated a novel transgenic mouse model with intestine specific overexpression of active SREBP2 (ISR2) driven by villin promoter. ISR2 mice showed overexpression of active SREBP2 specifically in the intestine. Microarray analysis of jejunal RNA from ISR2 mice showed a significant increase in genes involved in fatty acid and cholesterol synthesis. Cholesterol and triglyceride (TG) in jejunum and liver (mg/g protein) were significantly increased in ISR2 vs wild type mice. Serum Cholesterol was significantly increased in VLDL and LDL fractions whereas the level of serum triglycerides was decreased in ISR2 vs wild type mice. In conclusion, activation of intestinal SREBP2 alone seems to be sufficient to increase plasma cholesterol, highlighting the essential role of intestine in maintaining cholesterol homeostasis in the body.

The expression of intestinal Niemann-Pick C1-like 1 (NPC1L1) cholesterol transporter has been shown to be elevated in patients with diseases associated with hypercholesterolemia such as diabetes mellitus. High levels of glucose were shown to directly increase the expression of NPC1L1 in intestinal epithelial cells, but the underlying mechanisms are not fully defined. The present studies were, therefore, undertaken to examine the transcriptional regulation of NPC1L1 expression in human intestinal Caco2 cells in response to glucose. Removal of glucose from the culture medium of Caco2 cells for 24 h significantly decreased the NPC1L1 mRNA, protein expression, as well as the promoter activity. Glucose replenishment significantly increased the promoter activity of NPC1L1 in a dose-dependent manner compared with control cells. Exposure of Caco2 cells to nonmetabolizable form of glucose, 3- O-methyl-d-glucopyranose (OMG) had no effect on NPC1L1 promoter activity, indicating that the observed effects are dependent on glucose metabolism. Furthermore, glucose-mediated increase in promoter activity was abrogated in the presence of okadaic acid, suggesting the involvement of protein phosphatases. Glucose effects on several deletion constructs of NPC1L1 promoter demonstrated that cis elements mediating the effects of glucose are located in the region between −291 and +56 of NPC1L1 promoter. Consistent with the effects of glucose removal on NPC1L1 expression in Caco2 cells, 24-h fasting resulted in a significant decrease in the relative expression of NPC1L1 in mouse jejunum. In conclusion, glucose appears to directly modulate NPC1L1 expression via transcriptional mechanisms and the involvement of phosphatase-dependent pathways.

To investigate p16 gene methylation and its expression in 30 patients with sporadic colorectal adenocarcinoma in a North Indian population.Methylation specific polymerase chain reaction was used to detect p16 gene methylation and immunohistochemistry was used to study the p16 expression in 30 sporadic colorectal tumors as well as adjoining and normal tissue specimens.Aberrant promoter methylation of p16 gene was detected in 12 (40%) tumor specimens, whereas no promoter methylation was observed in adjoining and normal tissue. Immunohistochemistry showed expression of p16 protein in 26 (86.6%) colorectal tumors whereas complete loss of expression was seen in 4 (13.3%) and reduced expression was observed in 12 (40%) tumors. In the adjoining mucosa, expression of p16 was in 11 (36.6%) whereas no clear positivity for p16 protein was seen in normal tissue. There was a significant difference in the expression of p16 protein in tumor tissue and adjoining mucosa (P0.001). The methylation of the p16 gene had a significant effect on the expression of p16 protein (P = 0.021). There was a significant association of methylation of p16 gene with the tumor size (P = 0.015) and of the loss/reduced expression of p16 protein with the proximal site of the tumor (P = 0.047). Promoter methylation and expression of p16 had no relation with the survival of the patients (P0.05).Our study demonstrated that promoter hypermethylation of the p16 gene results in loss/reduced expression of p16 protein and this loss/reduced expression may contribute to tumor enlargement.

those patients. Total and surface expression of NHE3 were reduced in rat ileum treated with rapamycin, along with suppressed S6 protein phophorylation and increased ratio of LCII/I, confirming inhibition of mTOR and subsequent activation of autophagy by rapamycin. We further demonstrated that genetic inactivation of autophagy in mouse intestinal epithelium resulted in a marked accumulation of NHE3 and attenuated the rapamycin-induced down-regulation of NHE3, demonstrating a critical role for autophagy in maintaining NHE3 homeostasis. However, in addition to causing autophagic down-regulation of NHE3, we also demonstrate that rapamycin has a second mode of action: at high doses which mimics the sharp rise in serum rapamycin in non-infectious diarrhea, it causes acute inhibition of NHE3 surface expression in mouse ileum and NHE3 transporter activity in fibroblast PS120 cells, lacking endogenous NHE3, when stably transfected with human NHE3. Together, our data implicate two pathophysiological mechanisms of NHE3 down-regulation in the profound non-infectious diarrhea induced by rapamycin: 1) increased autophagic turnover of NHE3 by chronic exposure to rapamycin, and 2) reduced surface expression of NHE3 by acute exposure to a high concentration (spike) of rapamycin. Our findings not only lead to new insights into NHE3 regulation, but also provide new strategies to manage diarrhea in organ transplantations.

regulating CIE; however, its role in asymmetric CCH-stimulated signal transduction is not well understood. PURPOSE: Since PLD generates phosphatidic acid (PA) that regulates endocytosis, we (1) investigated whether CCH activates BB PLD, (2) identified which PLD isoform is necessary for CCH-inhibition of NHE3 activity, and (3) determined whether PA or PA-derived diacylglycerol (DAG) is required for this effect. METHODS: 12 day postconfluent Caco-2/BBe cells grown on filters were transiently infected with a 3HA-NHE3 adenovirus construct and studied 48 hours later. NHE3 activity was measured by fluorimetry using the pH-sensitive dye, BCECF, in Caco-2/BBe cells treated with vehicle, isoform specific PLD inhibitors (EVJ = PLD1; JWJ = PLD2, 5WO = dual), or propranolol (to inhibit conversion of PA to DAG) in the presence or absence of 10μM CCH to elevate [Ca]i. PLD activity was determined in total cell lysates from Caco-2/BBe cells treated with vehicle or CCH in the presence and absence of 10nM (blocks PLD1) or 50nM (blocks PLD2) FIPI. RESULTS: CCH induced rapid (1min) elevation of PLD activity in Caco-2/BBe cells, an effect which was blocked by BB, but not BLM, pretreatment with FIPI. Pretreatment of Caco-2/BBe cells with apical 10μM 5WO abolished CCH-mediated inhibition of NHE3 activity suggesting that BB PLD1 and/or PLD2 are necessary for NHE3 inhibition by CCH. This result was confirmed in studies in which cells were pretreated with both EVJ and JWJ as exposure to each inhibitor alone did not prevent CCH-inhibition of NHE3 activity. Propranolol treatment also prevented CCH-inhibition of NHE3 activity suggesting that PA-derived DAG is required for this effect. CONCLUSIONS: In summary, these data suggest that both PLD1 and PLD2 are activated by CCH signaling and are both necessary to inhibit NHE3 activity, which involves production of PA and DAG. Elevated [Ca]i-mediated signal transduction from the BLMM3 cholingergic receptor activates both BB PLD1 and PLD2 as part of an asymmetric signaling pathway that stimulates production of PA-derived DAG that results in the inhibition of BB NHE3 activity.

Apical Sodium-Dependent Bile Acid Transporter (ASBT) is responsible for active absorption of bile acids mainly in the ileum and is a central modulator of body cholesterol homeostasis. Inhibition of intestinal bile acid absorption reduces high levels of plasma cholesterol associated with diseases such as diabetes mellitus. Recent clinical trials have demonstrated that blockers of bile acid absorption improve lipid and cholesterol profiles in diabetic patients. Thus, specific inhibitors of ASBT may prove to be more effective therapeutic targets to lower plasma cholesterol. It is therefore, critical to delineate molecular and cellular mechanisms involved in ASBT regulation to develop better therapeuticmodalities for treatment of hypercholesterolemia. Recent studies have reported that 3'-untranslated region (3'-UTR) of ASBT is critical for regulation of ASBT expression. However, the role of micro RNAs (miRNAs) in the regulation of ASBT expression is not known. Using in silico computational target prediction, we initially identified the putative binding site for miR-625 in the 3'UTR of human ASBT mRNA (255bp-251bp downstream of the stop codon) with a context score percentile of 99. Luciferase assays were performed in human intestinal HUTU-80 and Caco2 cells as well as in kidney HEK293 cells. Mimic of miR-625 was transiently transfected in the cells along with a dual luciferase reporter construct (pMIR-GLO vector) containing human ASBT 3'-UTR sequence. The 3'-UTR of ASBT constructs demonstrated a significant decrease in relative luciferase activity as compared to empty vector in HUTU-80 (42.47 ± 1.5%, P