RESUMEN
Stomach cancer is the fifth most common cancer in terms of prevalence and incidence and the fourth leading cause of mortality in men and women worldwide. It is well-established that aberrant DNA methylation in cells can lead to carcinogenesis. The primary objective of our study was to investigate the aberrant DNA methylation status of genes associated with stomach cancer with a particular reference to the ethnic population of Mizoram, North East India. The site-level analysis identified 2883 CpG sites differentially methylated, representing â¼922 genes. Out of which 476 Differentially Methylated Positions (DMPs) were promoter-associated, 452 DMPs were hypermethylated, and 24 were hypomethylated. The region-level analysis identified 462 Differentially Methylated Regions (DMRs) corresponding to â¼320 genes, of which â¼281 genes were hypermethylated and â¼40 genes were hypomethylated. TCGA analysis showed that some of the genes had been previously implicated in other cancers including stomach cancer. Five hypermethylated genes were selected as candidate genes for further investigations and they have shown to be novel and could serve as candidate hypermethylation biomarkers for stomach cancer in this particular ethnic group.
Asunto(s)
Metilación de ADN , Neoplasias Gástricas , Islas de CpG , Epigénesis Genética , Etnicidad , Femenino , Humanos , India , Masculino , Neoplasias Gástricas/genéticaRESUMEN
Mycobacterium tuberculosis-macrophage interactions are key to pathogenesis and clearance of these bacteria. Although interactions between M. tuberculosis-associated lipids and TLRs, non-TLRs, and opsonic receptors have been investigated, interactions of these lipids and infected macrophage lipid repertoire with lipid-sensing nuclear receptors expressed in macrophages have not been addressed. In this study, we report that M. tuberculosis-macrophage lipids can interact with host peroxisome proliferator-activated receptor γ and testicular receptor 4 to ensure survival of the pathogen by modulating macrophage function. These two lipid-sensing nuclear receptors create a foamy niche within macrophage by modulating oxidized low-density lipoprotein receptor CD36, phagolysosomal maturation block by induction of IL-10, and a blunted innate response by alternative polarization of the macrophages, which leads to survival of M. tuberculosis. These results also suggest possible heterologous ligands for peroxisome proliferator-activated receptor γ and testicular receptor 4 and are suggestive of adaptive or coevolution of the host and pathogen. Relative mRNA expression levels of these receptors in PBMCs derived from clinical samples convincingly implicate them in tuberculosis susceptibility. These observations expose a novel paradigm in the pathogenesis of M. tuberculosis amenable for pharmacological modulation.
Asunto(s)
Células Espumosas/inmunología , Células Espumosas/microbiología , Metabolismo de los Lípidos/inmunología , Mycobacterium tuberculosis/crecimiento & desarrollo , Mycobacterium tuberculosis/inmunología , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/metabolismo , PPAR gamma/metabolismo , Animales , Células COS , Línea Celular Tumoral , Chlorocebus aethiops , Células Espumosas/metabolismo , Humanos , LigandosRESUMEN
The phytotherapeutic protein stem bromelain (SBM) is used as an anti-obesity alternative medicine. We show at the cellular level that SBM irreversibly inhibits 3T3-L1 adipocyte differentiation by reducing adipogenic gene expression and induces apoptosis and lipolysis in mature adipocytes. At the molecular level, SBM suppressed adipogenesis by downregulating C/EBPα and PPARγ independent of C/EBPß gene expression. Moreover, mRNA levels of adipocyte fatty acid-binding protein (ap2), fatty acid synthase (FAS), lipoprotein lipase (LPL), CD36, and acetyl-CoA carboxylase (ACC) were also downregulated by SBM. Additionally, SBM reduced adiponectin expression and secretion. SBM's ability to repress PPARγ expression seems to stem from its ability to inhibit Akt and augment the TNFα pathway. The Akt-TSC2-mTORC1 pathway has recently been described for PPARγ expression in adipocytes. In our experiments, TNFα upregulation compromised cell viability of mature adipocytes (via apoptosis) and induced lipolysis. Lipolytic response was evident by downregulation of anti-lipolytic genes perilipin, phosphodiestersae-3B (PDE3B), and GTP binding protein G(i)α(1), as well as sustained expression of hormone sensitive lipase (HSL). These data indicate that SBM, together with all-trans retinoic-acid (atRA), may be a potent modulator of obesity by repressing the PPARγ-regulated adipogenesis pathway at all stages and by augmenting TNFα-induced lipolysis and apoptosis in mature adipocytes.
Asunto(s)
Adipocitos/efectos de los fármacos , Adipogénesis/efectos de los fármacos , Apoptosis/efectos de los fármacos , Bromelaínas/farmacología , Lipólisis/efectos de los fármacos , Células 3T3-L1 , Adipocitos/metabolismo , Adipocitos/fisiología , Adipogénesis/genética , Adipogénesis/fisiología , Animales , Apoptosis/genética , Apoptosis/fisiología , Proteína beta Potenciadora de Unión a CCAAT/genética , Proteína beta Potenciadora de Unión a CCAAT/metabolismo , Proteína delta de Unión al Potenciador CCAAT/genética , Proteína delta de Unión al Potenciador CCAAT/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/genética , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Regulación hacia Abajo/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Hipolipemiantes/farmacología , Lipólisis/genética , Lipólisis/fisiología , Ratones , PPAR gamma/genética , PPAR gamma/metabolismo , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genéticaRESUMEN
Stem bromelain (SBM), a therapeutic protein, is rapidly absorbed across the gut epithelium. Because SBM encounters an alkaline pH at its principal site of absorption, we investigated the alkaline-induced denaturation of SBM. From pH 7 to 10, the protein's secondary structure remained the same, although a slight loss of tertiary structure was observed. Above pH 10, there was a significant and irreversible loss of secondary and tertiary structure. At pH 10, SBM showed enhanced tryptophan fluorescence, however, the number of accessible tryptophans remained the same. The thermodynamics of temperature transition at pH 7 and 10 were strikingly different, with the former showing a two-phase transition endotherm, and the latter a broad non-two-state transition. At pH 10, SBM showed a significant increase in 8-anilino-1-naphthalene-sulfonate binding relative to the native state, suggestive of a specific molten globule (SMG) state. These studies suggest a distinct conformational rearrangement in SBM, at the protein's isoelectric point.
Asunto(s)
Bromelaínas/química , Proteínas de Plantas/química , Ananas/enzimología , Bromelaínas/farmacocinética , Dicroismo Circular , Guanidina , Calor , Humanos , Concentración de Iones de Hidrógeno , Técnicas In Vitro , Absorción Intestinal , Punto Isoeléctrico , Fitoterapia , Proteínas de Plantas/farmacocinética , Conformación Proteica , Desnaturalización Proteica , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Espectrometría de Fluorescencia , Triptófano/químicaRESUMEN
Stem bromelain is a proteolytic phytoprotein with a variety of therapeutic effects. Understanding its structural properties could provide insight into the mechanisms underlying its clinical utility. Stem bromelain was evaluated for its conformational and folding properties at the pH conditions it encounters when administered orally. It exists as a partially folded intermediate at pH 2.0. The conformational changes to this intermediate state were evaluated using fluorinated alcohols known to induce changes similar to those seen in vivo. Studies using circular dichroism, fluorescence emission spectroscopy, binding of the hydrophobic dye 1-anilino-8-naphthalene sulfonic acid and mass spectrometry indicate that treatment with 10-30% hexafluoroisopropanol induces the partially folded intermediate to adopt much of the native protein's secondary structure, but only a rudimentary tertiary structure, characteristic of the molten globule state. Addition of slightly higher concentrations of hexafluoroisopropanol caused transformation from an alpha-helix to a beta-sheet and induced formation of a compact nonnative structure. This nonnative form was more inhibitory of cell survival than either the native or the partially folded intermediate forms, as measured by enhanced suppression of proliferative cues (e.g., extracellular-signal-regulated kinase) and initiation of apoptotic events. The nonnative form also showed better antitumorigenic properties, as evaluated using an induced two-stage mouse skin papilloma model. In contrast, the nonnative state showed only a fraction of the proteolytic activity of the native form. This study demonstrates that hexafluoroisopropanol can induce a conformational change in stem bromelain to a form with potentially useful therapeutic properties different from those of the native protein.