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Appropriate diversification of cellular lineages from multi-potent progenitors is essential for normal development and homeostasis. The specification of erythroid and megakaryocytic lineages represents an especially vital developmental event whose molecular regulation remains incompletely defined. We now demonstrate the role of Rgs18, a GTPase-activating protein and transcriptional target of the repressor Gfi1b, in regulating these processes in mouse and human cells. Gfi1b stringently represses Rgs18 expression in erythroid cells, whereas, during megakaryocytic differentiation, declining Gfi1b levels facilitate a robust induction of Rgs18. Concordantly, alterations in Rgs18 expression produce disparate outcomes by augmenting megakaryocytic and potently suppressing erythroid differentiation and vice versa. These phenotypes reflect the differential impact of Rgs18 on signaling through p38 MAPK family proteins, and ERK1 and ERK2 (also known as MAPK3 and MAPK1, respectively) in the two lineages, which in turn alter the balance between the mutually antagonistic transcription factors Fli1 and Klf1. Overall, these results identify Rgs18 as a new and crucial effector of Gfi1b that regulates downstream signaling and gene expression programs to orchestrate erythro-megakaryocytic lineage choices. This dual role of Rgs18 in reciprocally regulating divergent lineages could exemplify generic mechanisms characteristic of multiple family members in different contexts.
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Linaje de la Célula , Proteínas Proto-Oncogénicas/metabolismo , Proteínas RGS/metabolismo , Proteínas Represoras/metabolismo , Animales , Diferenciación Celular , Línea Celular , Células Eritroides/citología , Células Eritroides/metabolismo , Histona Demetilasas/metabolismo , Factores de Transcripción de Tipo Kruppel/metabolismo , Sistema de Señalización de MAP Quinasas , Megacariocitos/citología , Ratones , Proteína Proto-Oncogénica c-fli-1/metabolismo , Transcripción GenéticaRESUMEN
Lysine-specific demethylase 1 (LSD1) demethylates nucleosomal histone H3 lysine 4 (H3K4) residues in collaboration with the corepressor CoREST/REST corepressor 1 (Rcor1) and regulates cell fates by epigenetically repressing gene targets. The balanced regulation of this demethylase, if any, is however unknown. We now demonstrate the actions of two other Rcor paralogs, Rcor2 and Rcor3, in regulating LSD1 enzymatic activity and biological function in hematopoietic cells. All three Rcor proteins interact with LSD1 and with the erythro-megakaryocytic transcription factor growth factor independence (Gfi)1b; however, whereas Rcor2, like Rcor1, facilitates LSD1-mediated nucleosomal demethylation, Rcor3 competitively inhibits this process. Appending the SANT2 domain of Rcor1 to Rcor3 confers the ability to facilitate LSD1-mediated demethylation on the chimeric Rcor protein. Consistent with their biochemical activities, endogenous Rcor1, Rcor2, and LSD1 promote differentiation, whereas Rcor3 opposes these processes. Recruitment of Rcor3 to cognate gene targets by Gfi1b and LSD1 leads to inhibition of H3K4 demethylation of chromatin and transcriptional derepression of these loci. Remarkably, profound alterations in Rcor1/3 levels during erythroid versus megakaryocytic differentiation potentiate antagonistic outcomes. In mature erythroid cells, a strong upsurge in Rcor3 and a sharp decline in Rcor1 levels counteract LSD1/Rcor1/2-mediated differentiation. In contrast, the opposite changes in Rcor1/3 levels in megakaryocytes favor differentiation and likely maintain homeostasis between these lineages. Overall, our results identify Rcor3 as a natural inhibitor of LSD1 and highlight a dual mechanism of regulating the enzymatic activity and restraining the epigenetic impact of this robust demethylase during hematopoietic differentiation.
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Proteínas Co-Represoras/metabolismo , Hematopoyesis/fisiología , Proteínas del Tejido Nervioso/metabolismo , Oxidorreductasas N-Desmetilantes/metabolismo , Proteínas Represoras/metabolismo , Animales , Diferenciación Celular/fisiología , Proteínas Co-Represoras/genética , Epigénesis Genética/fisiología , Eritrocitos/citología , Eritrocitos/fisiología , Células HEK293 , Histona Demetilasas , Homeostasis/fisiología , Humanos , Megacariocitos/citología , Megacariocitos/fisiología , Ratones , Células Mieloides/citología , Células Mieloides/fisiología , Proteínas del Tejido Nervioso/genética , Oxidorreductasas N-Desmetilantes/genética , Proteínas Represoras/genéticaRESUMEN
Pancreatic cancer (PC) is the fourth leading cause of cancer-related death in United States. Efforts have been made towards the development of the viable solution for its treatment with constrained accomplishment because of its complex biology. It is well established that pancreatic cancer stem cells (CSCs), albeit present in a little count, contribute incredibly to PC initiation, progression, and metastasis. Customary chemo and radiotherapeutic alternatives, however, expands general survival, the related side effects are the significant concern. Amid the most recent decade, our insight about molecular and cellular pathways involved in PC and role of CSCs in its progression has increased enormously. Presently the focus is to target CSCs. The herbal products have gained much consideration recently as they, usually, sensitize CSCs to chemotherapy and target molecular signaling involved in various tumors including PC. Some planned studies have indicated promising results proposing that examinations in this course have a lot to offer for the treatment of PC. Although preclinical studies uncovered the importance of herbal products in attenuating pancreatic carcinoma, limited studies have been conducted to evaluate their role in clinics. The present review provides a new insight to recent advances in pancreatic cancer biology, treatment and current status of herbal products in its anticipation.
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Neoplasias Pancreáticas , Humanos , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/prevención & control , Neoplasias Pancreáticas/terapiaRESUMEN
Varicella zoster virus (VZV) is a highly contagious human herpes virus responsible for causing chickenpox (varicella) and shingles (herpes zoster). Despite the approval of a highly effective vaccine, Shingrix®, the global incidence of herpes zoster is increasing and the economic burden to the health care system and society are substantial due to significant loss of productivity and health complications, particularly among elderly and immunocompromised individuals. This is primarily because access to the vaccines remains mostly limited to countries within developed economies, such as USA and Canada. Therefore, similarly effective vaccines against VZV that are more accessible to the rest-of-the-world are necessary. In this study, we aimed to evaluate immunogenicity and memory response induced by three mRNA-LNP-based vaccine candidates targeting VZV's surface glycoprotein E (gE). C57BL/6 mice were immunized with each candidate vaccine, and humoral and cellular immune responses were assessed. Our results demonstrate that the mRNA-LNP-based vaccine candidates elicited robust and durable humoral responses specific to the gE antigen. Notably, mice vaccinated with the mRNA-LNP vaccines exhibited significantly higher antigen-specific T-cell cytokine production compared to the group receiving Shingrix®, the current standard of care vaccine. Additionally, mRNA-LNP vaccines induced long-lasting memory response, as evidenced by detection of persistent gE-specific Long-Lived Plasma Cells (LLPCs) and memory T cells four months after final immunization. These findings underscore the potential of our mRNA-LNP-based vaccine candidates in generating potent immune responses against VZV, offering promising prospects for their clinical development as an effective prophylactic vaccine against herpes zoster.
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Previously, we reported that pyrogallol, an anti-psoriatic agent, causes hepatotoxicity in experimental animals and silymarin, an herbal antioxidant, reduces pyrogallol-induced changes [Upadhyay, G., Kumar, A., Singh, M.P., 2007. Effect of silymarin on pyrogallol- and rifampicin-induced hepatotoxicity in mouse. Eur. J. Pharmacol. 565, 190-201.]. The present study was undertaken to assess the effect of resveratrol against pyrogallol-induced changes in hepatic damage markers, xenobiotic metabolizing enzymes and oxidative stress. Swiss albino mice were treated intraperitoneally, daily with pyrogallol (40 mg/kg), for one to four weeks, along with respective controls. In some set of experiments, animals were pre-treated with resveratrol (10 mg/kg), 2 h prior to pyrogallol treatment, along with respective controls. Alanine aminotransaminase, aspartate aminotransaminase and bilirubin were measured in blood plasma and mRNA expression of cytochrome P-450 (CYP) 1A1, CYP1A2, CYP2E1, glutathione-S-transferase (GST)-ya and GST-yc, catalytic activity of CYP1A1, CYP1A2, CYP2E1, GST, glutathione reductase and glutathione peroxidase, lipid peroxidation and reduced glutathione (GSH) level were measured in liver. Resveratrol reduced pyrogallol-mediated increase in alanine aminotransaminase, aspartate aminotransaminase, bilirubin, lipid peroxidation and mRNA expression and catalytic activity of CYP2E1 and CYP1A2. Pyrogallol-mediated decrease in GST-ya and GST-yc expressions, GST, glutathione peroxidase and glutathione reductase activities and GSH content was significantly attenuated in resveratrol co-treated animals. CYP1A1 expression and catalytic activity were not altered significantly in any treated groups. The results demonstrate that resveratrol modulates pyrogallol-induced changes in hepatic toxicity markers, xenobiotic metabolizing enzymes and oxidative stress.
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Antioxidantes/farmacología , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Fármacos Dermatológicos/efectos adversos , Estrés Oxidativo , Pirogalol/efectos adversos , Estilbenos/farmacología , Xenobióticos/metabolismo , Animales , Hidrocarburo de Aril Hidroxilasas/metabolismo , Bilirrubina/sangre , Biomarcadores/metabolismo , Catálisis , Enfermedad Hepática Inducida por Sustancias y Drogas/enzimología , Glutatión/metabolismo , Glutatión Peroxidasa/metabolismo , Glutatión Reductasa/metabolismo , Glutatión Transferasa/metabolismo , Peroxidación de Lípido , Hígado/efectos de los fármacos , Hígado/enzimología , Hígado/metabolismo , Masculino , Ratones , Resveratrol , Transaminasas/sangreRESUMEN
Rifampicin and pyrogallol, besides beneficial effects, elicit hepatotoxicity in experimental animals and humans. The present investigation was undertaken to elucidate the role of drug/toxicant-metabolizing enzymes in rifampicin- and pyrogallol-induced hepatotoxicity and the effect of silymarin, a herbal antioxidant, on rifampicin- and pyrogallol-induced alterations in mouse liver. Male Swiss albino mice were treated intraperitoneally with and without rifampicin (20 mg/kg) and/or pyrogallol (40 mg/kg) for 1, 2, 3 and 4 weeks. In some experiments, animals were treated with silymarin (40 mg/kg), 2 h prior to rifampicin and/or pyrogallol. The differential expression and catalytic activity of cytochrome P-450 (CYP) 1A1, CYP1A2 and CYP2E1, the activity of glutathione-S-transferase, glutathione peroxidase and glutathione reductase, and lipid peroxidation were measured in the liver of control and treated groups. CYP1A1 expression and catalytic activity were not altered following individual or combinational treatment. A significant augmentation in the expression and activity of CYP1A2 and CYP2E1 was observed following pyrogallol and rifampicin+pyrogallol treatment; however, rifampicin exhibited a significant induction of CYP2E1 only. Attenuation of glutathione-S-transferase, glutathione reductase and glutathione peroxidase activities and augmentation of lipid peroxidation were observed following rifampicin and/or pyrogallol treatment and a cumulative effect was seen when the two drugs were administered in combination. Silymarin restored the rifampicin- and/or pyrogallol-induced alterations in the expression and activity of CYP1A2 and CYP2E1, the activity of glutathione-S-transferase, glutathione reductase, and glutathione peroxidase, and lipid peroxidation. The results demonstrate the role of CYP1A2, CYP2E1, glutathione-S-transferase, glutathione reductase and glutathione peroxidase in rifampicin- and pyrogallol-induced hepatotoxicity and provide evidence for the involvement of silymarin in attenuation of drug-induced hepatotoxicity.
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Hígado/efectos de los fármacos , Pirogalol/toxicidad , Rifampin/toxicidad , Silimarina/farmacología , Alanina Transaminasa/sangre , Animales , Western Blotting , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Citocromo P-450 CYP1A2/genética , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/metabolismo , Glutatión/metabolismo , Peroxidación de Lípido , Hígado/metabolismo , Masculino , Ratones , ARN Mensajero/análisisRESUMEN
Benzo(a)pyrene induces cytochrome P-450 1A1 (CYP1A1) expression in rat polymorphonuclear leukocytes (PMNs) that upregulates expression of inducible nitric oxide synthase (iNOS). In the present study, the involvement of secondary signaling molecules in CYP1A1-mediated augmentation of iNOS expression in benzo(a)pyrene-treated rat PMNs was investigated. PMNs were isolated from the peripheral blood of controls and benzo(a)pyrene-treated rats. The expression and/or activity of CYP1A1, iNOS, tumor necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1beta), and intracellular calcium ([Ca(2+)]i) concentrations were measured in control and benzo(a)pyrene-treated rat PMNs with and without alpha-naphthoflavone, aminoguanidine, genistein, pyrrolidine dithiocarbamate (PDTC), felodipine, or SB202190 pre-treatment. A significant elevation in CYP1A1 and [Ca(2+)]i was observed in benzo(a)pyrene-treated rat PMNs, which was significantly restored by alpha-naphthoflavone or genistein. Neither PDTC, SB202190, nor aminoguanidine altered the benzo(a)pyrene-mediated increase in [Ca(2+)]i. Although felodipine reduced the benzo(a)pyrene-mediated increase in [Ca(2+)]i, no significant change was observed in CYP1A1 expression and activity. Benzo(a)pyrene-augmented iNOS expression and activity in PMNs were significantly reverse by felodipine, genistein, or PDTC. Benzo(a)pyrene also induced TNF-alpha and IL-1beta production in PMNs, which was significantly reversed by genistein. The results demonstrated the involvement of [Ca(2+)]i, tyrosine kinase, inflammatory cytokines, and NF-kappaB in CYP1A1-mediated iNOS expression in benzo(a)pyrene-treated rat PMNs.
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Benzo(a)pireno/toxicidad , Citocromo P-450 CYP1A1/fisiología , Neutrófilos/efectos de los fármacos , Neutrófilos/enzimología , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Transducción de Señal/fisiología , Animales , Calcio/metabolismo , Catálisis , Citocromo P-450 CYP1A1/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Interleucina-1beta/análisis , Interleucina-1beta/biosíntesis , Masculino , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Nitritos/metabolismo , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Ratas , Ratas Wistar , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Necrosis Tumoral alfa/análisis , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Understanding the production and differentiation of megakaryocytes from progenitors is crucial for realizing the biology and functions of these vital cells. Previous gene ablation studies demonstrated the essential role of the transcriptional repressor Gfi1b (growth factor independence 1b) in the generation of both erythroid and megakaryocytic cells. However, our recent work has demonstrated the down-regulation of this factor during megakaryocytic differentiation. In this study we identify two new gene targets of Gfi1b, the cytoskeletal proteins Kindlin3 and Talin1, and demonstrate the inverse expression and functions of these cytoskeletal targets relative to Gfi1b, during megakaryocytic differentiation. Both kindlin3 and talin1 promoters exhibit dose dependent Gfi1b and LSD1 (lysine specific demethylase 1; a Gfi1b cofactor) enrichment in megakaryocytes and repression in non-hematopoietic cells. Accordingly the expression of these genes is elevated in gfi1b mutant and LSD1 inhibited hematopoietic cells, while during megakaryocytic differentiation, declining Gfi1b levels fostered the reciprocal upregulation of these cytoskeletal factors. Concordantly, manipulation of Kindlin3 and Talin1 expression demonstrated positive correlation with megakaryocytic differentiation with over-expression stimulating, and inhibition diminishing, this process. Co-operativity between these factors and integrins in promoting differentiation was further underscored by physical interactions between them and integrinß3/CD61 and by stimulation of differentiation by the Talin1 head domain, which is necessary and sufficient for integrin activation. Therefore this study demonstrates the significance of Gfi1b regulated Kindlin3-Talin1 expression in driving megakaryocytic differentiation and highlights the contribution of cytoskeletal agents in the developmental progression of these platelet progenitors.
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Diferenciación Celular/genética , Megacariocitos/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genética , Talina/metabolismo , Animales , Línea Celular , Inmunoprecipitación de Cromatina , Humanos , Integrina beta3/metabolismo , Megacariocitos/citología , RatonesRESUMEN
The liver is the center for drug and xenobiotic metabolism, which is influenced most with medication/xenobiotic-mediated toxic activity. Drug-induced hepatotoxicity is common and its actual frequency is hard to determine due to underreporting, difficulties in detection or diagnosis, and incomplete observation of exposure. The death rate is high, up to about 10% for drug-induced liver damage. Endorsed medications represented >50% of instances of intense liver failure in a study from the Acute Liver Failure Study Group of the patients admitted in 17 US healing facilities. Albeit different studies are accessible uncovering the mechanistic aspects of medication prompted hepatotoxicity, we are in the dilemma about the virtual story. The expanding prevalence and effectiveness of Ayurveda and natural products in the treatment of various disorders led the investigators to look into their potential in countering drug-induced liver toxicity. Several natural products have been reported to date to mitigate the drug-induced toxicity. The dietary nature and less adverse reactions of the natural products provide them an extra edge over other candidates of supplementary medication. In this paper, we have discussed the mechanism involved in drug-induced liver toxicity and the potential of herbal antioxidants as supplementary medication.
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Neurodegenerative diseases are still a challenge for researchers and clinicians due to its complexity. Traditional medicines usually do not provide sufficient protection against these diseases due to drug resistance and relapse. The discovery of the therapeutic potential of stem cells offers new opportunities for the treatment of incurable neurological diseases. Based on their biological properties, stem cells can differentiate into specific tissue type and maintain the cellular tissue/organ homeostasis in physiological and pathological conditions. Recently, it has been demonstrated that somatic cells of patients can be reprogrammed to a pluripotent state from which neural lineage cells can be derived. Potential strategies such as cell replacement therapy and gene transfer to the diseased or injured brain have opened a new line of therapeutic approach for a broad spectrum of human neurological diseases. Thus, stem cell replacement therapy for central and peripheral nervous system disorders aims at repopulating the affected neural tissue with new neurons. However, the limiting factors that have hampered the development of this promising therapeutic approach are the lack of suitable cell types for cell replacement therapy in patients suffering from neurological disorders. In this review, we have discussed the recent advances in stem cell replacement therapy with particular emphasis to neurological disorders.
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Enfermedades Neurodegenerativas/terapia , Trasplante de Células Madre , Animales , HumanosRESUMEN
Matrix metalloproteinases (MMPs), a large family of calcium-dependent zinc-containing endopeptidases, are involved in the tissue remodeling and degradation of the extracellular matrix. MMPs are widely distributed in the brain and regulate various processes including microglial activation, inflammation, dopaminergic apoptosis, blood-brain barrier disruption, and modulation of α-synuclein pathology. High expression of MMPs is well documented in various neurological disorders including Parkinson's disease (PD), Alzheimer's disease (AD), Japanese encephalitis (JE), and Glaucoma. Although potentially critical, the role of MMPs in neuronal disorders is under-investigated. The present review summarizes the role of MMPs in neurodegeneration with a particular emphasis on PD, AD, JE, and Glaucoma.
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BACKGROUND AND AIM: Genetic factors regulating alcohol metabolism could predispose in developing alcoholic pancreatitis (ACP). Studies revealed that alcohol could be metabolized by both ways, oxidative and non-oxidative. The main oxidative pathway includes alcohol dehydrogenase (ADH), aldehyde dehydrogenase (ALDH), and cytochrome P450 enzyme. We investigated the association of polymorphisms in these enzymes with the alcoholic pancreatitis in the north Indian population. METHOD: Patients with alcoholic pancreatitis (ACP; n = 72), tropical calcific pancreatitis (TCP; n = 75), alcoholic controls (AC; n = 40), and healthy controls (HC; n = 100) were included in the study. Blood samples were collected from the subjects in EDTA coated vials. DNA was extracted and genotyping for ADH3, ALDH2, and CYP2E1 was done by PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism). The products were analyzed by gel electrophoresis. RESULT: The frequency distribution of ADH3(*)1/(*)1 genotype was significantly higher in ACP group (59.7%) compared with TCP (38.7%), HC (42%), and AC (37.5%) and was found to be associated with increased risk of alcoholic pancreatitis. There was no statistically significant difference between the frequency distribution of ADH3(*)1/(*)1, ADH3(*)1/(*)2, and ADH3(*)2/(*)2 genotypes between TCP and HC or healthy alcoholics. ALDH2 gene was monomorphic in our population, and the frequencies for CYP2E1 intron 6 Dra I polymorphism were comparable in all the four groups. CONCLUSION: This study shows that carriers of ADH3(*)1/(*)1 individuals consuming alcohol are at higher risk for alcoholic pancreatitis than those with other genotypes such as ADH3(*)1/(*)2 and ADH3(*)2/(*)2.
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The purpose of the study was to examine the molecular mechanisms by which rottlerin inhibited growth of human pancreatic tumors in Balb C nude mice, and pancreatic cancer cells isolated from Kras(G12D) mice. AsPC-1 cells were injected subcutaneously into Balb c nude mice, and tumor-bearing mice were treated with rottlerin. Cell proliferation and apoptosis were measured by Ki67 and TUNEL staining, respectively. The expression of components of Akt, Notch, and Sonic Hedgehog (Shh) pathways were measured by the immunohistochemistry, Western blot analysis, and/or q-RT-PCR. The effects of rottlerin on pancreatic cancer cells isolated from Kras(G12D) mice were also examined. Rottlerin-treated mice showed a significant inhibition in tumor growth which was associated with suppression of cell proliferation, activation of capase-3 and cleavage of PARP. Rottlerin inhibited the expression of Bcl-2, cyclin D1, CDK2 and CDK6, and induced the expression of Bax in tumor tissues compared to untreated control. Rottlerin inhibited the markers of angiogenesis (Cox-2, VEGF, VEGFR, and IL-8), and metastasis (MMP-2 and MMP-9), thus blocking production of tumorigenic mediators in tumor microenvironment. Rottlerin also inhibited epithelial-mesenchymal transition by up-regulating E-cadherin and inhibiting the expression of Slug and Snail. Furthermore, rottlerin treatment of xenografted tumors or pancreatic cancer cells isolated from Kras(G12D) mice showed a significant inhibition in Akt, Shh and Notch pathways compared to control groups. These data suggest that rottlerin can inhibit pancreatic cancer growth by suppressing multiple signaling pathways which are constitutively active in pancreatic cancer. Taken together, our data show that the rottlerin induces apoptosis and inhibits pancreatic cancer growth by targeting Akt, Notch and Shh signaling pathways, and provide a new therapeutic approach with translational potential for humans.
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Acetofenonas/farmacología , Inhibidores de la Angiogénesis/farmacología , Benzopiranos/farmacología , Proliferación Celular/efectos de los fármacos , Genes ras , Neoplasias Pancreáticas/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Carga Tumoral/efectos de los fármacos , Proteínas Angiogénicas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/metabolismo , Línea Celular Tumoral , Transición Epitelial-Mesenquimal/efectos de los fármacos , Proteínas Hedgehog/antagonistas & inhibidores , Proteínas Hedgehog/metabolismo , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Ratones Transgénicos , Neovascularización Patológica , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores Notch/antagonistas & inhibidores , Receptores Notch/metabolismo , Transducción de Señal/efectos de los fármacos , Factores de Tiempo , Microambiente Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Pancreatic cancer is a deadly disease, and therefore effective treatment and/or prevention strategies are urgently needed. The objectives of this study were to examine the molecular mechanisms by which embelin inhibited human pancreatic cancer cell growth in vitro, and xenografts in Balb C nude mice, and pancreatic cancer cell growth isolated from KrasG12D transgenic mice. XTT assays were performed to measure cell viability. AsPC-1 cells were injected subcutaneously into Balb c nude mice and treated with embelin. Cell proliferation and apoptosis were measured by Ki67 and TUNEL staining, respectively. The expression of Akt, and Sonic Hedgehog (Shh) and their target gene products were measured by the immunohistochemistry, and Western blot analysis. The effects of embelin on pancreatic cancer cells isolated from 10-months old KrasG12D mice were also examined. Embelin inhibited cell viability in pancreatic cancer AsPC-1, PANC-1, MIA PaCa-2 and Hs 766T cell lines, and these inhibitory effects were blocked either by constitutively active Akt or Shh protein. Embelin-treated mice showed significant inhibition in tumor growth which was associated with reduced expression of markers of cell proliferation (Ki67, PCNA and Bcl-2) and cell cycle (cyclin D1, CDK2, and CDK6), and induction of apoptosis (activation of caspase-3 and cleavage of PARP, and increased expression of Bax). In addition, embelin inhibited the expression of markers of angiogenesis (COX-2, VEGF, VEGFR, and IL-8), and metastasis (MMP-2 and MMP-9) in tumor tissues. Antitumor activity of embelin was associated with inhibition of Akt and Shh pathways in xenografts, and pancreatic cancer cells isolated from KrasG12D mice. Furthermore, embelin also inhibited epithelial-to-mesenchymal transition (EMT) by up-regulating E-cadherin and inhibiting the expression of Snail, Slug, and ZEB1. These data suggest that embelin can inhibit pancreatic cancer growth, angiogenesis and metastasis by suppressing Akt and Shh pathways, and can be developed for the treatment and/or prevention of pancreatic cancer.
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Benzoquinonas/farmacología , Proteínas Hedgehog/metabolismo , Neoplasias Pancreáticas/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Transducción de Señal/efectos de los fármacos , Proteínas ras/metabolismo , Animales , Línea Celular Tumoral , Proteínas Hedgehog/genética , Xenoinjertos , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Mutantes , Ratones Desnudos , Metástasis de la Neoplasia , Trasplante de Neoplasias , Neovascularización Patológica/tratamiento farmacológico , Neovascularización Patológica/genética , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patología , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Transducción de Señal/genética , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas ras/genéticaRESUMEN
Gfi1b (growth factor independence 1b) is a zinc finger transcription factor essential for development of the erythroid and megakaryocytic lineages. To elucidate the mechanism underlying Gfi1b function, potential downstream transcriptional targets were identified by chromatin immunoprecipitation and expression profiling approaches. The combination of these approaches revealed the oncogene meis1, which encodes a homeobox protein, as a direct and prominent target of Gfi1b. Examination of the meis1 promoter sequence revealed multiple Gfi1/1b consensus binding motifs. Distinct regions of the promoter were occupied by Gfi1b and its cofactors LSD1 and CoREST/Rcor1, in erythroid cells but not in the closely related megakaryocyte lineage. Accordingly, Meis1 was significantly upregulated in LSD1 inhibited erythroid cells, but not in megakaryocytes. This lineage specific upregulation in Meis1 expression was accompanied by a parallel increase in di-methyl histone3 lysine4 levels in the Meis1 promoter in LSD1 inhibited, erythroid cells. Meis1 was also substantially upregulated in gfi1b-/- fetal liver cells along with its transcriptional partners Pbx1 and several Hox messages. Elevated Meis1 message levels persisted in gfi1b mutant fetal liver cells differentiated along the erythroid lineage, relative to wild type. However, cells differentiated along the megakaryocytic lineage, exhibited no difference in Meis1 levels between controls and mutants. Transfection experiments further demonstrated specific repression of meis1 promoter driven reporters by wild type Gfi1b but neither by a SNAG domain mutant nor by a DNA binding deficient one, thus confirming direct functional regulation of this promoter by the Gfi1b transcriptional complex. Overall, our results demonstrate direct yet differential regulation of meis1 transcription by Gfi1b in distinct hematopoietic lineages thus revealing it to be a common, albeit lineage specific, target of both Gfi1b and its paralog Gfi1.
Asunto(s)
Células Eritroides/metabolismo , Proteínas de Homeodominio/genética , Megacariocitos/metabolismo , Proteínas de Neoplasias/genética , Proteínas del Tejido Nervioso/genética , Oxidorreductasas N-Desmetilantes/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Represoras/genética , Transcripción Genética , Animales , Secuencia de Bases , Sitios de Unión , Diferenciación Celular , Línea Celular Tumoral , Proteínas Co-Represoras , Células Eritroides/citología , Feto , Regulación de la Expresión Génica , Histona Demetilasas , Proteínas de Homeodominio/metabolismo , Humanos , Megacariocitos/citología , Ratones , Datos de Secuencia Molecular , Proteína 1 del Sitio de Integración Viral Ecotrópica Mieloide , Proteínas de Neoplasias/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Oxidorreductasas N-Desmetilantes/metabolismo , Regiones Promotoras Genéticas , Unión Proteica , Estructura Terciaria de Proteína , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Represoras/metabolismo , Transducción de SeñalRESUMEN
The study aimed to investigate the effects of cypermethrin on biochemical, histopathological, and motor behavioral indices of the nigrostriatal dopaminergic system in adult rats treated with or without cypermethrin (1/10 adult dose) during postnatal days 5-19. Spontaneous locomotor activity (SLA) and rotarod tests were performed to assess motor behavior. Levels of dopamine, 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in the striatum, and tyrosine hydroxylase (TH) immunoreactivity and 4',6-diamidino-2-phenylindole (DAPI)/Fluoro-Jade B staining in the substantia nigra were measured to assess dopaminergic neurodegeneration. Postnatal treated animals did not exhibit significant changes in any measured parameters. The significant reduction in the time of stay on rotarod, spontaneous locomotor activity, dopamine, 3,4-dihydroxyphenylacetic acid, and tyrosine hydroxylase immunoreactivity while an increase in homovanillic acid level and Fluoro-Jade B-positive cells were observed in cypermethrin treated adult rats. These changes were more pronounced in the animals treated with cypermethrin during postnatal days followed by adulthood compared with adulthood alone. The results obtained thus demonstrate that exposure to cypermethrin during adulthood induces dopaminergic neurodegeneration in rats and postnatal exposure enhances the susceptibility of animals to dopaminergic neurodegeneration if rechallenged during adulthood.
Asunto(s)
Cuerpo Estriado/fisiopatología , Neuronas Dopaminérgicas/metabolismo , Insecticidas/toxicidad , Enfermedades Neurodegenerativas/inducido químicamente , Enfermedades Neurodegenerativas/fisiopatología , Piretrinas/toxicidad , Sustancia Negra/fisiopatología , Animales , Cuerpo Estriado/efectos de los fármacos , Neuronas Dopaminérgicas/efectos de los fármacos , Exposición a Riesgos Ambientales , Masculino , Vías Nerviosas/efectos de los fármacos , Vías Nerviosas/fisiopatología , Enfermedades Neurodegenerativas/patología , Ratas , Ratas Wistar , Sustancia Negra/efectos de los fármacosRESUMEN
NAD(P)H:quinone oxidoreductase 1 (NQO1) and cytochrome P450 1A2 (CYP1A2) are involved in the metabolism of estrogens. Genetic polymorphisms in these genes may lead to interindividual variation in breast cancer susceptibility. This study was undertaken to investigate the association of NQO1 exon 6 proline187serine (C609T) and CYP1A2 exon 2 phenylalanine21leucine (C63G) polymorphisms with breast cancer susceptibility in North Indian women. Polymorphisms were analyzed by polymerase chain reaction amplification of the desired segment of NQO1 and CYP1A2 genes followed by restriction fragment length polymorphism. NQO1 mRNA expression was analyzed by semiquantitative reverse transcription-polymerase chain reaction and its enzyme activity was estimated spectrofluorophotometrically. Odds ratios for NQO1 C609T heterozygous and homozygous variants were 0.66 (95% confidence interval: 0.39-1.13; p-value: 0.141) and 1.07 (95% confidence interval: 0.46-2.46; p-value: 0.976). All cases and controls were monomorphic for the CYP1A2 exon 2 phenylalanine21leucine (C63G) genotype. NQO1 mRNA expression and its catalytic activity among wild-type genotype, homozygous variant, and heterozygous variant were not significantly altered, except for catalytic activity of the NQO1 homozygous variant, which was observed extremely low. The results of the study suggest that NQO1 exon 6 proline187serine (C609T) and CYP1A2 exon 2 phenylalanine21leucine (C63G) polymorphisms do not play a significant role in breast cancer susceptibility in North Indian women.
Asunto(s)
Neoplasias de la Mama/genética , Citocromo P-450 CYP1A2/genética , Predisposición Genética a la Enfermedad , NAD(P)H Deshidrogenasa (Quinona)/genética , Polimorfismo de Nucleótido Simple/genética , Población Blanca/genética , Adulto , Neoplasias de la Mama/patología , Citocromo P-450 CYP1A2/metabolismo , Femenino , Genotipo , Humanos , Inactivación Metabólica , India , Persona de Mediana Edad , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Xenobióticos/metabolismoRESUMEN
The study aimed to identify the differentially expressed nigrostriatal proteins in cypermethrin-induced neurodegeneration and to investigate the role of microglial activation therein. Proteomic approaches were used to identify the differentially expressed proteins. Microglial activation, tyrosine hydroxylase immunoreactivity (TH-IR), dopamine content, and neurobehavioral changes were measured according to the standard procedures. The expressions of α-internexin intermediate filament (α-IIF), ATP synthase D chain (ATP-SD), heat shock protein (Hsp)-70, truncated connexin-47, Hsp-60, mitogen-activated protein kinase-activated kinase-5, nicotinamide adenine dinucleotide dehydrogenase 24k chain precursor, platelet-activating factor acetyl hydrolase 1b-α2 (PAF-AH 1b-α2), and synaptosomal-associated protein-25 (SNAP-25) were altered in the substantia nigra and nicotinamide adenine dinucleotide- specific isocitrate dehydrogenase, phosphatidylethanolamine-binding protein-1, prohibitin, protein disulfide isomerase-endoplasmic reticulum 60 protease, stathmin, and ubiquitin-conjugating enzyme in the striatum along with motor impairment, decreased dopamine and TH-IR, and increased microglial activation after cypermethrin exposure. Minocycline restored α-IIF, ATP-SD chain, truncated connexin-47, Hsp-60, PAF-AH 1b-α2, stathmin and SNAP-25 expressions, motor impairment, dopamine, TH-IR, and microglial activation. The results suggest that cypermethrin produces microglial activation-dependent and -independent changes in the expression patterns of the nigrostriatal proteins leading to dopaminergic neurodegeneration.
Asunto(s)
Neuronas Dopaminérgicas/metabolismo , Degeneración Nerviosa/patología , Proteómica/métodos , Piretrinas/toxicidad , Animales , Western Blotting , Cromatografía Liquida , Cuerpo Estriado/metabolismo , Dopamina/análisis , Relación Dosis-Respuesta a Droga , Expresión Génica , Proteínas de Filamentos Intermediarios/genética , Proteínas de Filamentos Intermediarios/metabolismo , Masculino , Espectrometría de Masas , Microglía/metabolismo , Degeneración Nerviosa/inducido químicamente , Fenotipo , Ratas , Ratas Wistar , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
Pyrogallol, a potent anti-psoriatic drug, produces toxicity due to its ability to generate free radicals, besides its beneficial effects. Oxidative stress is implicated in pyrogallol-mediated toxicity in general and hepatotoxicity in particular. Naturally occurring antioxidants including, resveratrol and silymarin have been proposed as potential supplements to counteract pyrogallol-mediated toxicity, without reducing its efficacy. Due to increase in the popularity of natural antioxidants in combating pyrogallol-mediated toxicity, a literature-based survey was performed to assess their role in experimental studies and possible implications in real life situations. Although preclinical studies revealed the boons of naturally occurring antioxidants in attenuating/abolishing the undesirable effects of pyrogallol exposure, limited studies have been conducted to evaluate their role in clinics. In this review, an update on the recent development in assessing the potential of natural antioxidants in pyrogallol-mediated toxicity in preclinical interventions, triumphs and pitfalls of such investigations, their translational challenges and future possibilities are discussed.
Asunto(s)
Antioxidantes/uso terapéutico , Pirogalol/toxicidad , Animales , Encefalopatías/tratamiento farmacológico , Enfermedades Cardiovasculares/tratamiento farmacológico , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Evaluación Preclínica de Medicamentos , Medicamentos Herbarios Chinos/farmacología , Enfermedades Gastrointestinales/tratamiento farmacológico , Humanos , Ratones , Estrés Oxidativo , RatasRESUMEN
In this study, the involvement of various molecular events in pyrogallol-mediated hepatotoxicity was deciphered by differential mRNA transcription profiles of control and pyrogallol treated mice liver. The modulatory effects of silymarin on pyrogallol-induced differentially expressed transcripts were also looked into. Swiss albino mice were treated with or without pyrogallol. In some set of experiments, mice were also treated with silymarin 2 h prior to pyrogallol. Total RNA was isolated from liver and polyadenylated RNA was reverse-transcribed into Cye 3 or Cye 5 labelled cDNA. Equal amounts of labelled cDNA from two different groups were mixed and hybridized with mouse 15 k array. The hybridized arrays were scanned, analyzed and the expression level of each transcript was calculated. The differential expression was validated by quantitative real time polymerase chain reaction. Comparative transcription pattern showed an alteration in the expression of 183 transcripts (150 up-regulated and 33 down-regulated) associated with oxidative stress, cell cycle, cytoskeletal network, cell-cell adhesion, extra-cellular matrix, inflammation, apoptosis, cell-signaling and intermediary metabolism in pyrogallol-exposed liver and silymarin pre-treatment modulated the expression of many of these transcripts. Results obtained thus suggest that pyrogallol induces multiple molecular events leading to hepatotoxicity and silymarin effectively counteracts pyrogallol-mediated alterations.