RESUMEN
BACKGROUND: A huge array of function is played by the Wnt/ß-catenin signaling pathway in development by balancing gene expression through the modulation of cell-specific DNA binding downstream effectors such as T-cell factor/lymphoid enhancer factor (TCF/LEF). The ß-catenin/TCF-4 complex is a central regulatory switch for differentiation and proliferation of intestinal cells (both normal and malignant). Thus, in the present study we evaluated each of 60 cases of sporadic adenocarcinoma, alongside adjoining and normal mucosa specimens of colorectum in humans, for mutation and expression analysis of the gene coding for TCF-4 protein. METHODS: DNA sequencing following PCR amplification and SSCP analysis (single strand conformation polymorphism) was employed to detect TCF-4 gene mutations in the case of exon 1. Quantitative real-time (qRT) PCR, immunohistochemistry (IHC), confocal microscopy and western blot analysis were used to detect TCF-4 gene/protein expression. RESULTS: Sequencing analysis confirmed 5/60 patients with a point mutation in exon 1 of the TCF-4 gene in tumor samples. mRNA expression using qRT-PCR showed approximately 83% decreased TCF-4 mRNA expression in tumor tissue and adjoining mucosa compared to normal mucosa. Similarly, a significant decrease in protein expression using IHC showed decreased TCF-4 protein expression in tumor tissue and adjoining mucosa compared to normal mucosa, which also corresponds to some important clinicopathological factors, including disease metastasis and tumor grade. Mutational alterations and downregulation of TCF-4 mRNA and hence decreased expression of TCF-4 protein in tumors suggest its involvement in the pathogenesis of CRC. CONCLUSIONS: A remarkable decrease in TCF-4 mRNA and protein expression was detected in tumorous and adjoining tissues compared to normal mucosa. Hence the alterations in genomic architecture along with downregulation of TCF-4 mRNA and decreased expression of TCF-4 protein in tumors, which is in accordance with clinical features, suggest its involvement in the pathogenesis of CRC. Thus, deregulation and collaboration of TCF-4 with CRC could be a concrete and distinctive feature in the prognosis of the disease at an early stage of development.
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Adenocarcinoma/metabolismo , Neoplasias Colorrectales/metabolismo , Factor de Transcripción 4/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/patología , Adenocarcinoma/secundario , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/mortalidad , Neoplasias Colorrectales/patología , Supervivencia sin Enfermedad , Regulación hacia Abajo , Exones , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Inmunohistoquímica , Masculino , Microscopía Confocal , Persona de Mediana Edad , Clasificación del Tumor , Mutación Puntual , Pronóstico , Factor de Transcripción 4/genética , Proteínas Supresoras de Tumor/genética , beta Catenina/metabolismoRESUMEN
Epigenetic therapy induced by dietary components has become a strong interest in the field of cancer prevention. Olive oil, a potent dietary chemopreventive agent, control colon cancer, however, its role in epigenetic therapy remains unclear. Thus, we aimed to investigate the effect of olive oil in a preclinical model of colon cancer by targeting genetic and epigenetic mechanisms. DMH was used to induce colon cancer in rats; while olive oil was given to separate group of rats along with DMH treatment. Tumor burden and incidence in DMH and DMH + olive oil-treated rats was observed by macroscopic examination and histoarchitectural studies. Potent anti-inflammatory, anti-angiogenic and pro-apoptotic activity of olive oil was explored by gene expression and immunohistochemical studies. The effect of olive oil on epigenetic alterations was examined by detecting promoter methylation with MS-HRM and dysregulation of miRNA by TaqMan MicroRNA Assay. We observed that olive oil administration lowered tumor incidence and inhibited the development of tumors in DMH-treated rats. Olive oil markedly decreased the expression of inflammatory and angiogenic markers and restored the expression of pro-apoptotic markers in DMH-treated rats. Furthermore, the inverse relationship between gene expression and DNA methylation, deviant miRNA pattern and miRNA silencing mediated by aberrant DNA methylation was also seen in DMH-treated rats, which was potentially reversible upon olive oil treatment. Our study concludes that olive oil may play a role in the epigenetic therapy by altering NF-κB and apoptotic pathways via targeting noncoding RNAs and methylation machinery that affecting epigenome to prevent colon carcinogenesis.
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Carcinogénesis/efectos de los fármacos , Neoplasias del Colon/patología , Neoplasias del Colon/prevención & control , Metilación de ADN/efectos de los fármacos , Aceite de Oliva/farmacología , ARN no Traducido/genética , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Biomarcadores de Tumor/genética , Neoplasias del Colon/genética , Metilación de ADN/genética , Epigénesis Genética/efectos de los fármacos , Epigénesis Genética/genética , Expresión Génica/efectos de los fármacos , Expresión Génica/genética , Masculino , MicroARNs/genética , FN-kappa B/genética , Regiones Promotoras Genéticas/efectos de los fármacos , Regiones Promotoras Genéticas/genética , Ratas , Ratas Sprague-DawleyRESUMEN
BACKGROUND & OBJECTIVES: Clostridium difficile is the primary cause of hospital-acquired colitis in patients receiving antibiotics. The pathogenicity of the organism is mainly due to the production of toxins. This study was conducted to investigate the presence of toxigenic C. difficile in the faecal samples of hospitalized patients suspected to have C. difficile infection (CDI) and corroborating the findings with their clinical and demographic data. METHODS: Diarrhoeic samples obtained from 1110 hospitalized patients were cultured for C. difficile and the isolates confirmed by phenotypic and molecular methods. Toxigenicity of the isolates was determined using enzyme-linked immunosorbent assay for toxins A and B. Details of patients included in the study were noted and analyzed. RESULTS: Of the 1110 patients (mean age 39±19.6 yr), 63.9 per cent were males and 36.1 per cent were females. The major antibiotics received by the patients were nitazoxanide (23.9%), penicillins/penicillin combinations (19.0%), quinolones including fluoroquinolones (13.1%), carbapenems (11.5%), glycopeptides (11.0%) and cephalosporins (8.4%). The clinical symptoms predominantly present were watery diarrhoea (56.4%), fever (40.0%) and abdominal pain (35.3%). The underlying diseases were gastrointestinal disorders (52.6%), followed by cancers (13.2%), surgical conditions (8.3%), and hepatic disorders (8.0%). Of the 174 C. difficile isolates, 54.6 per cent were toxigenic. Toxigenic C. difficile was present in all patients with surgical conditions, 65.2 per cent with cancers and 57.1 per cent with gastrointestinal disorders. INTERPRETATION & CONCLUSIONS: C. difficile was found to be an important cause of gastrointestinal infections in hospitalized patients with underlying diseases and on antibiotics. Clinical conditions of the patients correlating with toxigenic culture can be an important tool for establishing CDI diagnosis.
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Toxinas Bacterianas/aislamiento & purificación , Clostridioides difficile/aislamiento & purificación , Infecciones por Clostridium/microbiología , Diarrea/microbiología , Adolescente , Adulto , Anciano , Antibacterianos/uso terapéutico , Técnicas de Cultivo de Célula/métodos , Niño , Preescolar , Clostridioides difficile/química , Clostridioides difficile/patogenicidad , Infecciones por Clostridium/tratamiento farmacológico , Infecciones por Clostridium/epidemiología , Infecciones por Clostridium/patología , Diarrea/tratamiento farmacológico , Diarrea/epidemiología , Diarrea/patología , Heces/microbiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Centros de Atención Terciaria , Adulto JovenRESUMEN
ß-catenin (CTNNB1), an oncogene/onco-protein and an adhesion molecule is a key effector in colorectal cancer (CRC). Its activation, and subsequent up-regulation of Wnt-signaling, is an important event in the development of certain human cancers including CRC. Mutations in the ß-catenin gene in the region of serine-threonine glycogen kinase (GSK)-3ß phosphorylation target sites have been identified in colorectal cancer in humans. In the current study, we investigated 60 sporadic colorectal adenocarcinomas along with adjoining and normal mucosa cases in humans for ß-catenin mutations. Thirteen of sixty colorectal tumors from humans had point mutations with a frequency of 21.66% at codons 24, 26, 27, 32, 34, 35, 41, 42,43, 46, 49, 54, 55, or 67 sites which are mutated in colorectal cancer and some of these sites in other cancers. Thus, there appears to be a key involvement of ß-catenin activation in human colorectal carcinogenesis. mRNA expression analysis using q-Real Time PCR showed 21.5-fold up-regulation of ß-catenin mRNA in tumor tissue compared to normal and adjoining mucosa. Protein expression analysis using immunohistochemistry, confocal microscopy, and Western blot confirmed aberrant accumulation of ß-catenin protein along the nucleus and cytoplasm following mutation. The observed mutations and up-regulation of mRNA in tumors, and the increased expression of ß-catenin protein in CRC suggest that these alterations are early and prognostic events in sporadic colorectal carcinogenesis in humans. © 2015 Wiley Periodicals, Inc.
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Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Análisis Mutacional de ADN/métodos , Regulación hacia Arriba , beta Catenina/genética , beta Catenina/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Núcleo Celular/metabolismo , Neoplasias Colorrectales/metabolismo , Citoplasma/metabolismo , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Mutación Puntual , Pronóstico , Análisis de Supervivencia , Adulto JovenRESUMEN
BACKGROUND: Lactase activity declines during childhood in the majority of human populations leading to adult-type hypolactasia (AtH). C/T-13910 and G/A-22018 single nucleotide polymorphisms (SNPs) have been suggested to be associated with AtH in different human populations. Coeliac disease (CD) is an autoimmune condition characterized by damage to intestinal cells leading to ultimate deterioration. AIM: This study investigated the association between coeliac disease (CD) and SNPs leading to AtH in children from North India. SUBJECTS AND METHODS: Intestinal biopsies and saliva samples were obtained from 52 children with CD diagnosis and 102 control subjects. Biopsies were assayed for disaccharidase activities and samples were genotyped for given SNPs. RESULTS: Prevalence of C/C and G/G genotypes of AtH was almost equal in the CD and control group. The CD group had low lactase activity compared to the control group, irrespective of genotype at C/T -13910 and G/A -22018 SNPs (p < 0.05). For the control group, lactase activity was high in children with C/T + G/A genotypes compared to C/C + G/G (p < 0.05). CONCLUSION: There appears to be no significant correlation between C/T -13910 or G/A -22018 SNPs of AtH and CD. Children with C/C or G/G genotype of AtH may not be at greater risk of CD.
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Enfermedad Celíaca/genética , Lactasa/deficiencia , Lactasa/genética , Intolerancia a la Lactosa/genética , Enfermedad Celíaca/epidemiología , Enfermedad Celíaca/inmunología , Niño , Preescolar , Estudios de Cohortes , Genotipo , Humanos , India/epidemiología , Polimorfismo de Nucleótido SimpleRESUMEN
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 < 0.05) was observed. Normal expression of hMLH1 and hMSH2 was seen in all of the unmethylated tumors (100%). Nuclear staining and promoter methylation of hMLH1 and hMSH2 did not significantly influence survival. hMLH1 methylation was common and was significantly correlated with loss of hMLH1 protein expression. In contrast, hMSH2 methylation was infrequent. These findings suggest that the inactivation of MMR gene expression probably via hypermethylation may lead to inactivation of their functions which finally leads to tumor aggressiveness and the immunostaining of hMLH1 protein can be used as a prognostic factor for determining the grade of the tumor.
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Proteínas Adaptadoras Transductoras de Señales/genética , Neoplasias Colorrectales/genética , Metilación de ADN , Proteína 2 Homóloga a MutS/genética , Proteínas Nucleares/genética , Regiones Promotoras Genéticas , Proteínas Adaptadoras Transductoras de Señales/análisis , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Colorrectales/mortalidad , Femenino , Humanos , Inmunohistoquímica , India , Masculino , Persona de Mediana Edad , Homólogo 1 de la Proteína MutL , Proteína 2 Homóloga a MutS/análisis , Proteínas Nucleares/análisis , Estudios ProspectivosRESUMEN
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.
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Adenocarcinoma/genética , Proteína de la Poliposis Adenomatosa del Colon/genética , Neoplasias Colorrectales/genética , Mutación/genética , Proteínas Proto-Oncogénicas/genética , Proteína p53 Supresora de Tumor/genética , Proteínas ras/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidad , Proteína de la Poliposis Adenomatosa del Colon/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Colon/metabolismo , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/mortalidad , Femenino , Estudios de Seguimiento , Humanos , Técnicas para Inmunoenzimas , India , Metástasis Linfática , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Reacción en Cadena de la Polimerasa , Polimorfismo Conformacional Retorcido-Simple , Pronóstico , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas p21(ras) , Recto/metabolismo , Tasa de Supervivencia , Proteína p53 Supresora de Tumor/metabolismo , Adulto Joven , Proteínas ras/metabolismoRESUMEN
The kinetics of Na+ activation of brush border sucrase (sucrose D-glucosidase E.C. 3.2.1.48) has been studied in mice intestine. At pH 5.0, 50 mM Na+ ions stimulated sucrase activity by 84%. At pH 7.2, enzyme stimulation was reduced to 16%, whereas, atpH 8.5, 10-100 mMNa+ ions produced 18-45% inhibition of enzyme activity. Kinetic studies revealed that at pH 5.0, the enzyme activation by Na+ ions was V-type, which changed to K-type atpH 7.2, whereas at alkaline pH (8.5), Na+ ions inhibited the enzyme activity non-competitively. Using the non-compulsory model of Na+ ion stimulation of brush border sucrase [Mahmood & Alvarado, Arch Bioch Biophys, 168 (1975) 585] various kinetic constants involved in activation of sucrase by Na ions were determined. It is apparent that Na+ stimulation of brush border sucrase is pH dependent, which is similar to that described for rat, rabbit and other mammalian species and conform to identical mechanisms, at least with reference to the affinity type effects, as observed in mice intestine.
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Mucosa Intestinal/metabolismo , Microvellosidades/enzimología , Sodio/metabolismo , Sacarasa/metabolismo , Animales , Activación Enzimática , Intestinos/enzimología , Cinética , RatonesRESUMEN
Intestinal brush border sucrase-isomaltase (sucrose D-glucosidase E.C. 3.2.1.48, E.C. 3.2.1.10) exhibits pH-dependent stimulatory or inhibitory effects in response to Na+ ions. However, whether the enzyme undergoes conformational modulations as a function of pH and in the presence of alkali metal ions is not known. In this paper, we investigated the structural and functional relationship of purified murine sucrase in response to pH and Na+ ions using UV-CD fluorescence and spectroscopic studies. Kinetic studies revealed that at pH 5.0, the enzyme activation by Na+ ions was V-type, which changed to K-type at pH 7.2, whereas at alkaline pH (8.5), Na+ ions inhibited the enzyme activity and inhibition was uncompetitive in nature, affecting both the Km and Vmax components. Far UV-CD spectra of protein at pH 7.2 in the absence and presence of Na+ were almost overlapping, suggesting that secondary structure of protein was not affected upon addition of the salt. However, near UV-CD spectra indicated marked alterations in the tertiary structure of protein in presence of 50 mM Na+ ions. Increase in pH from 7.2 to 8.5 resulted in a marked rise in fluorescence intensity and red shift in lambda max due to tryptophan residues in the enzyme molecule. These findings suggested that alterations in enzyme activity as a function of pH and Na+ ions was associated with ionization of key amino acid residues together with structural modifications in the enzyme conformation around neutral or alkaline pH.
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Mucosa Intestinal/enzimología , Microvellosidades/enzimología , Sodio/química , Complejo Sacarasa-Isomaltasa/química , Animales , Cationes Monovalentes , Dicroismo Circular , Concentración de Iones de Hidrógeno , Ratones , Ratones Endogámicos BALB C , Estructura Terciaria de Proteína , Sacarasa/químicaRESUMEN
BACKGROUND: Clostridium difficile is an important cause of infectious colitis among hospitalized patients across the globe. The pathogenic potential of C. difficile in producing significant morbidity and mortality is mainly due to production of toxins A and B. The outbreaks of C. difficile infection (CDI) are due to changes in the genetic sequences of the organism. There is hardly any molecular study reported on the prevalent types of C. difficile strains in India. Toxinotyping and sequencing of locally circulating C. difficile isolates from patients presenting to our tertiary care center of North India were done. MATERIALS AND METHODS: C. difficile strains (n = 174) isolated from 1,110 fecal samples from patients with suspected CDI were subjected to toxinotyping and partial sequencing of tcdA and tcdB genes. Comparison of nucleotide sequences with reference C. difficile 630 strain using BLAST was made and translated into corresponding amino acid sequences by ExPASy. RESULTS AND DISCUSSION: Of 174 C. difficile isolates, 121 were toxigenic, belonging to toxinotype 0 (n = 76) and VIII (n = 45). Partial sequencing of toxin genes using bioinformatics approaches revealed changes in toxin A sequences of five (50%) C. difficile isolates, but the translated nucleotide sequences showed substitution in only three of them. No variation was seen in the toxin B nucleotide sequences. Interstrain variations were found in the clinical C. difficile isolates in our region. CONCLUSION: PCR amplified toxigenic genes followed by sequencing can help to identify genetic changes and pathogenicity of varied collection of C. difficile isolates.
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Radiation enteritis is one of the most feared complications of abdominal and pelvic regions. Thus, radiation to abdominal or pelvic malignancies unavoidably injures the intestine. Because of rapid cell turnover, the intestine is highly sensitive to radiation injury, which is the limiting factor in the permissible dosage of irradiation. Bowel injuries such as fistulas, strictures, and chronic malabsorption are potentially life-threatening complications and have an impact on patient quality of life. The incidence of radiation enteritis is increasing because of the current trend of combined chemotherapy and radiation. The consequences of radiation damage to the intestine may result in considerable morbidity and even mortality. The observed effects of ionizing radiation are mediated mainly by oxygen-free radicals that are generated by its action on water and are involved in several steps of signal transduction cascade, leading to apoptosis. The oxyradicals also induce DNA strand breaks and protein oxidation. An important line of defense against free radical damage is the presence of antioxidants. Therefore, administration of antioxidants may ameliorate the radiation-induced damage to the intestine.
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BACKGROUND: Postnatal development of rat intestine is associated with a decline in brush-border lactase activity. This phenomenon is similar to the adulthood hypolactasia in humans. However, the mechanism underlying this process is not understood. METHODS: The effect of luminal proteases from adult rat intestine on the intestinal lactase activity in animals aged 7, 14, 21 and 30 days was studied in in vitro experiments. Lactase levels were estimated using enzyme assays and Western blot analysis. RESULTS: Incubation of purified brush borders with increasing concentrations of luminal proteases reduced the lactase activity in intestine of 7-day-old rats, but not in that of adult animals. Western blot analysis revealed low signal of the 220-kDa lactase protein in 7-day-old animals, but not that of older weaned animals. CONCLUSIONS: Our findings suggest that luminal proteases may be responsible for the maturational decline in intestinal lactase activity.
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Endopeptidasas/farmacología , Intestinos/efectos de los fármacos , Lactasa/efectos de los fármacos , Animales , Animales Recién Nacidos , Western Blotting , Mucosa Intestinal/metabolismo , Intestinos/ultraestructura , Lactasa/metabolismo , Masculino , Microvellosidades/efectos de los fármacos , Microvellosidades/metabolismo , Ratas , Ratas WistarRESUMEN
Lactase-phlorizin hydrolase (LPH), a membrane-bound glycoprotein present in the luminal surface of enterocytes in the intestine is responsible for lactose intolerance, a phenomenon prevalent in humans worldwide. In the rodent intestine, the post-natal development of the LPH follows a specific pattern, such that the enzyme levels are high in the peri-natal period, but declines considerably upon maturation. The observed maturational decline in the LPH activity is very similar to adult-type hypolactasia observed in humans. Majority of the studies have been carried out using animal models or cell lines and a number of hypotheses have been put forward to explain the maturational decline of lactase activity such as: (a) decreased amount of lactase protein, (b) defect in post-translational modification of precursor lactase to the mature enzyme, and (c) synthesis of an inactive, high molecular weight lactase with altered glycosylation, however, the precise underlying mechanism of adult-type hypolactasia remains undefined. The present review describes the recent developments in understanding the regulation of lactase expression and the possible mechanism of adult-type hypolactasia, as a cause of lactose intolerance.
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Regulación Enzimológica de la Expresión Génica , Intolerancia a la Lactosa/etiología , Lactosa/metabolismo , Animales , Glicosilación , Humanos , Lactasa/biosíntesis , Lactasa-Florizina Hidrolasa/química , Lactosa/genética , Polimorfismo GenéticoRESUMEN
BACKGROUND: Doxycycline (DOX) exhibits anti-inflammatory, anti-tumor, and pro-apoptotic activity and is being tested in clinical trials as a chemotherapeutic agent for several cancers, including colon cancer. MATERIALS & METHODS: In the current study, the chemotherapeutic activity of doxycycline was tested in a rat model of colon carcinogenesis, induced by colon specific cancer promoter, 1,2, dimethylhydrazine (DMH) as well as study the effect of DOX-alone on a separate group of rats. RESULTS: Doxycycline administration in DMH-treated rats (DMH-DOX) unexpectedly increased tumor multiplicity, stimulated progression of colonic tumor growth from adenomas to carcinomas and revealed metastasis in small intestine as determined by macroscopic and histopathological analysis. DOX-alone treatment showed markedly enhanced chronic inflammation and reactive hyperplasia, which was dependent upon the dose of doxycycline administered. Moreover, immunohistochemical analysis revealed evidence of inflammation and anti-apoptotic action of DOX by deregulation of various biomarkers. CONCLUSION: These results suggest that doxycycline caused chronic inflammation in colon, small intestine injury, enhanced the efficacy of DMH in tumor progression and provided a mechanistic link between doxycycline-induced chronic inflammation and tumorigenesis. Ongoing studies thus may need to focus on the molecular mechanisms of doxycycline action, which lead to its inflammatory and tumorigenic effects.
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Carcinogénesis/inducido químicamente , Doxiciclina/efectos adversos , Inflamación/patología , Metástasis de la Neoplasia/patología , 1,2-Dimetilhidrazina , Animales , Peso Corporal/efectos de los fármacos , Carcinogénesis/patología , Caspasa 3/metabolismo , Caspasa 9/metabolismo , Proliferación Celular/efectos de los fármacos , Enfermedad Crónica , Neoplasias del Colon/inducido químicamente , Neoplasias del Colon/patología , Citocromos c/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Inmunohistoquímica , Intestino Delgado/efectos de los fármacos , Intestino Delgado/patología , Masculino , Metaloproteinasa 9 de la Matriz/metabolismo , FN-kappa B/metabolismo , Ratas Sprague-Dawley , Proteína p53 Supresora de Tumor/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
mRNA levels encoding lactase were detected by Northern blot analysis using two different probes in developing rat intestine. Probe I and probe II corresponding to second half of prolactase gene showed a 6.8 kb mRNA transcript in 7, 14, 21 and 30 day old rat intestine. There was no change in quantity of lactase mRNA detected using probe II, but hybridization with probe I showed a progressive decrease in mRNA transcript encoding lactase with age. At day 7 and 14 of postnatal development, the lactase mRNA was quite high, but it reduced upon weaning. The in vitro translation products of RNA detected by Western blot analysis using brush border lactase antibodies showed several isoforms of lactase antigen with molecular weight ranging from 100-220 kDa. Analysed at days 7 and 30 of postnatal development, lactase isoforms of molecular weight 130 kDa and 220 kDa were similar to those found in purified brush border membranes. The translation of RNA to 220 kDa lactase protein was high in 7 and 14 day old pups, but it was markedly reduced in 30 day old animals. These findings support the contention that translation of mRNA to lactase is impaired in weaned animals, which may also be responsible for the maturational decline in lactase activity in adult rat intestine.
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Animales Recién Nacidos/fisiología , Mucosa Intestinal/metabolismo , Lactasa/metabolismo , Biosíntesis de Proteínas/fisiología , ARN Mensajero/metabolismo , Ratas/fisiología , Factores de Edad , Animales , Animales Recién Nacidos/genética , Northern Blotting , Western Blotting , Cartilla de ADN , Lactasa/genética , Ratas/genética , Ratas WistarRESUMEN
BACKGROUND AND OBJECTIVES: Ethanol exposure during gestation induces marked aberrations in growth and development of offsprings collectively known as foetal alcohol syndrome (FAS);. However, its effects on the postnatal development of alcohol dehydrogenase (ADH) are not adequately investigated. Therefore, ADH activity in liver and intestine of rats exposed to ethanol during gestation was studied in relation to postnatal development. METHODS: Pregnant female rats beginning at day 1 of gestation were fed 1 ml of 30 per cent ethanol daily during the entire gestation period. ADH activity was determined in liver and intestine postnatally at day 4, 8, 14, 20 and 30. DNA and RNA contents and intestinal histology were also examined. RESULTS: During the first two weeks of postnatal life, there was no difference in ADH levels of rat liver and intestine in control and prenatally ethanol exposed pups but ADH levels were significantly reduced at 3-4 wk in ethanol fed group compared to control. A similar decrease in DNA and RNA contents of intestine and changes in tissue morphology were observed in ethanol exposed pups during postnatal development. INTERPRETATION AND CONCLUSION: The findings of our study suggested that prenatal ethanol exposure modified ADH activity in liver and intestine during postnatal development. This could affect ethanol metabolism under these conditions.
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Envejecimiento/metabolismo , Alcohol Deshidrogenasa/metabolismo , Animales Recién Nacidos , Etanol/farmacología , Intestinos/enzimología , Hígado/enzimología , Efectos Tardíos de la Exposición Prenatal , Animales , Animales Recién Nacidos/crecimiento & desarrollo , Animales Recién Nacidos/metabolismo , Femenino , Intestinos/efectos de los fármacos , Intestinos/patología , Hígado/efectos de los fármacos , Hígado/patología , Embarazo , Ratas , Ratas WistarRESUMEN
Clostridium difficile infection (CDI) leads to considerable morbidity and mortality among hospitalized patients. Faecal specimens from 1110 hospitalized patients suspected for CDI were cultured for isolation of C. difficile and characterization of virulence genes. PCR was carried out for toxigenic genes tcdA, tcdB, cdtA and cdtB and PCR-RFLP for fliC and slpA genes. Of 174 (15.7%) C. difficile isolates, 121 (69.5%) were toxigenic, amongst which 68 (56.2%) also had both tcdA and tcdB genes. The remaining 53 (43.8%) of the isolates also had at least one of the toxin genes. Binary toxin genes (cdtA and cdtB) with only one of the two components were present in 16 (9.2%) of the 174 isolates. The other virulence genes - fliC and slpA - were present in 100% of the isolates. The most frequent PCR-RFLP type of fliC gene was type I (n = 101), followed by type VII (n = 49) and type III (n = 24). The slpA gene presented with three combinations of patterns. Characterization of virulence genes in C. difficile isolates is of extreme importance for epidemiological surveillance and control of outbreaks owing to the capacity of this bacterium to adapt to new environmental circumstances, leading to the emergence of new epidemic strains.
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Clostridioides difficile/genética , Clostridioides difficile/aislamiento & purificación , Infección Hospitalaria/microbiología , Enterocolitis Seudomembranosa/microbiología , Heces/microbiología , Adolescente , Adulto , Anciano , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Niño , Preescolar , Clostridioides difficile/clasificación , Femenino , Humanos , India , Masculino , Persona de Mediana Edad , Polimorfismo de Longitud del Fragmento de Restricción , Prevalencia , Estudios Prospectivos , Centros de Atención Terciaria/estadística & datos numéricos , Adulto JovenRESUMEN
BACKGROUND & OBJECTIVES: Infection by Salmonella Typhimurium is one of the leading causes of intestinal dysfunction, however the underlying mechanism of this effect is largely unknown. Hence the effect of enterotoxin secreted by Salmonella Typhimurium-(S-LT) was studied on D-glucose absorption and brush border enzymes in rabbit ileum. mRNA levels encoding these proteins were also analysed. METHODS: Adult male New Zealand white rabbits were used. The polymyxine B extract of enterotoxin obtained from Salmonella Typhimurium was tested for the presence of enterotoxicity by rabbit ileal loop test. D-glucose uptake by ileal tissue was measured by the tissue accumulation method. Intestinal brush border membranes were isolated and the effect of S-LT on various brush border enzymes studied. RESULTS: S-LT significantly inhibited (P < 0.01) the absorption of Na+ dependent D-glucose uptake but had no effect on Na+ independent sugar uptake in rabbit ileum. The activities of brush border sucrase (72% P < 0.001) and lactase (47% P < 0.01) and alkaline phosphatase (43% P < 0.01) were also significantly reduced in infected animals as compared to the controls. Northern blot analysis revealed that mRNA levels encoding Na+ glucose co-transporter (SGLT1), brush border lactase and sucrase activities were unaffected in Salmonella infected rabbit ileal loops. INTERPRETATION & CONCLUSION: The findings suggest that the intestinal dysfunctions observed in Salmonella infection are unrelated to mRNA expression encoding Na+ glucose co-transporter and brush border enzyme proteins in rabbit ileum.
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Toxinas Bacterianas/toxicidad , Endotoxinas/toxicidad , Íleon/efectos de los fármacos , Íleon/fisiopatología , Salmonella typhimurium/patogenicidad , Animales , Transporte Biológico Activo/efectos de los fármacos , Expresión Génica/efectos de los fármacos , Glucosa/metabolismo , Absorción Intestinal/efectos de los fármacos , Masculino , Glicoproteínas de Membrana/genética , Microvellosidades/efectos de los fármacos , Microvellosidades/enzimología , Proteínas de Transporte de Monosacáridos/genética , Conejos , Salmonelosis Animal/genética , Salmonelosis Animal/fisiopatología , Transportador 1 de Sodio-GlucosaRESUMEN
Consumption of dietary oil, viz., corn, fish, coconut, or olive, induced the secretion of surfactant-like particles (SLP) in rat intestine. These lipoprotein particles differ in (i) levels of alkaline phosphatase activity, (ii) lipid composition, and (iii) FA composition in response to feeding of different oils. The secreted particles had similar buoyancy (1.07-1.08 g/mL) and cholesterol/phospholipid molar ratios (0.61-0.72) except that feeding coconut oil to rats produced SLP with a low (0.18) cholesterol/phospholipid molar ratio compared to control animals. It is concluded from these observations that feeding different oils induces the secretion of lipoprotein particles in rat intestine with different chemical compositions.
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Grasas de la Dieta/farmacología , Intestino Delgado/efectos de los fármacos , Intestino Delgado/metabolismo , Tensoactivos/química , Tensoactivos/metabolismo , Fosfatasa Alcalina/sangre , Fosfatasa Alcalina/metabolismo , Animales , Colesterol/análisis , Ácidos Grasos/análisis , Masculino , Aceites/farmacología , Fosfolípidos/análisis , Fosfolípidos/sangre , Ratas , Ratas Wistar , Triglicéridos/sangreRESUMEN
The activities of lactase, sucrase, alkaline phosphatase (AP) and y-glutamyl transpeptidase (gamma-GTP) were studied in the intestinal brush border membranes of pups born to rat mothers exposed to ethanol (1 ml of 30% ethanol daily during gestation) at different days of postnatal development. The activities of lactase (at day 4-20) and sucrase (at day 20-30) were considerably reduced in response to prenatal exposure to ethanol, while AP (at day 4-30) and gamma-GTP activities were significantly enhanced (p < 0.05) at day 4, 8, 14 and 20, but there was no significant difference by day 30 of postnatal development. The observed changes in enzyme activities were corroborated by Western blot analysis of lactase, sucrase and AP. Kinetic studies revealed a change in Vmax without affecting apparent Km of enzymes under these conditions. The present findings suggest that in utero ethanol exposure to rats is embryotoxic and affects the postnatal development of various brush border enzymes, which persist long after the ethanol was withdrawn prior to birth.