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1.
Nat Rev Mol Cell Biol ; 20(3): 175-193, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30655609

RESUMO

The loss of vital cells within healthy tissues contributes to the development, progression and treatment outcomes of many human disorders, including neurological and infectious diseases as well as environmental and medical toxicities. Conversely, the abnormal survival and accumulation of damaged or superfluous cells drive prominent human pathologies such as cancers and autoimmune diseases. Apoptosis is an evolutionarily conserved cell death pathway that is responsible for the programmed culling of cells during normal eukaryotic development and maintenance of organismal homeostasis. This pathway is controlled by the BCL-2 family of proteins, which contains both pro-apoptotic and pro-survival members that balance the decision between cellular life and death. Recent insights into the dynamic interactions between BCL-2 family proteins and how they control apoptotic cell death in healthy and diseased cells have uncovered novel opportunities for therapeutic intervention. Importantly, the development of both positive and negative small-molecule modulators of apoptosis is now enabling researchers to translate the discoveries that have been made in the laboratory into clinical practice to positively impact human health.


Assuntos
Apoptose/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Animais , Doença , Homeostase , Humanos , Neoplasias/metabolismo , Neoplasias/fisiopatologia , Patologia
3.
Adv Exp Med Biol ; 978: 145-166, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28523545

RESUMO

Anxiety disorders are highly prevalent psychiatric disorders often comorbid with depression and substance abuse. Twin studies have shown that anxiety disorders are moderately heritable. Yet, genome-wide association studies (GWASs) have failed to identify gene(s) significantly associated with diagnosis suggesting a strong role for environmental factors and the epigenome. A number of anxiety disorder subtypes are considered "stress related." A large focus of research has been on the epigenetic and anxiety-like behavioral consequences of stress. Animal models of anxiety-related disorders have provided strong evidence for the role of stress on the epigenetic control of the hypothalamic-pituitary-adrenal (HPA) axis and of stress-responsive brain regions. Neuroepigenetics may continue to explain individual variation in susceptibility to environmental perturbations and consequently anxious behavior. Behavioral and pharmacological interventions aimed at targeting epigenetic marks associated with anxiety may prove fruitful in developing treatments.


Assuntos
Transtornos de Ansiedade/genética , Ansiedade/genética , Epigênese Genética/genética , Adolescente , Corticosteroides/fisiologia , Adulto , Animais , Ansiolíticos/uso terapêutico , Ansiedade/tratamento farmacológico , Ansiedade/etiologia , Ansiedade/psicologia , Transtornos de Ansiedade/tratamento farmacológico , Transtornos de Ansiedade/etiologia , Transtornos de Ansiedade/psicologia , Criança , Modelos Animais de Doenças , Suscetibilidade a Doenças , Doenças em Gêmeos/genética , Feminino , Estudo de Associação Genômica Ampla , Humanos , Sistema Hipotálamo-Hipofisário/fisiopatologia , Padrões de Herança , Plasticidade Neuronal , Sistema Hipófise-Suprarrenal/fisiopatologia , Gravidez , Efeitos Tardios da Exposição Pré-Natal , Estresse Psicológico , Estudos em Gêmeos como Assunto
4.
J Biol Chem ; 289(26): 18258-69, 2014 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-24825908

RESUMO

Collaborator of ARF (CARF) has been shown to directly bind to and regulate p53, a central protein that controls tumor suppression via cellular senescence and apoptosis. However, the cellular functions of CARF and the mechanisms governing its effect on senescence, apoptosis, or proliferation are still unknown. Our previous studies have shown that (i) CARF is up-regulated during replicative and stress-induced senescence, and its exogenous overexpression caused senescence-like growth arrest of cells, and (ii) suppression of CARF induces aneuploidy, DNA damage, and mitotic catastrophe, resulting in apoptosis via the ATR/CHK1 pathway. In the present study, we dissected the cellular role of CARF by investigating the molecular pathways triggered by its overexpression in vitro and in vivo. We found that the dosage of CARF is a critical factor in determining the proliferation potential of cancer cells. Most surprisingly, although a moderate level of CARF overexpression induced senescence, a very high level of CARF resulted in increased cell proliferation. We demonstrate that the level of CARF is crucial for DNA damage and checkpoint response of cells through ATM/CHK1/CHK2, p53, and ERK pathways that in turn determine the proliferative fate of cancer cells toward growth arrest or proproliferative and malignant phenotypes. To the best of our knowledge, this is the first report that demonstrates the capability of a fundamental protein, CARF, in controlling cell proliferation in two opposite directions and hence may play a key role in tumor biology and cancer therapeutics.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Proliferação de Células , Dano ao DNA , Neoplasias/metabolismo , Neoplasias/fisiopatologia , Proteínas de Ligação a RNA/metabolismo , Apoptose , Proteínas Reguladoras de Apoptose/genética , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Linhagem Celular Tumoral , Humanos , Neoplasias/genética , Proteínas de Ligação a RNA/genética , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
5.
Biogerontology ; 16(6): 775-88, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26318578

RESUMO

Intermittent fasting dietary restriction (IF-DR) is recently reported to be an effective intervention to retard age associated disease load and to promote healthy aging. Since sustaining long term caloric restriction regimen is not practically feasible in humans, so use of alternate approach such as late onset short term IF-DR regimen which is reported to trigger similar biological pathways is gaining scientific interest. The current study was designed to investigate the effect of IF-DR regimen implemented for 12 weeks in middle age rats on their motor coordination skills and protein and DNA damage in different brain regions. Further, the effect of IF-DR regimen was also studied on expression of energy regulators, cell survival pathways and synaptic plasticity marker proteins. Our data demonstrate that there was an improvement in motor coordination and learning response with decline in protein oxidative damage and recovery in expression of energy regulating neuropeptides. We further observed significant downregulation in nuclear factor kappa B (NF-κB) and cytochrome c (Cyt c) levels and moderate upregulation of mortalin and synaptophysin expression. The present data may provide an insight on how a modest level of short term IF-DR, imposed in middle age, can slow down or prevent the age-associated impairment of brain functions and promote healthy aging by involving multiple regulatory pathways aimed at maintaining energy homeostasis.


Assuntos
Envelhecimento/fisiologia , Encéfalo/fisiologia , Restrição Calórica/métodos , Jejum/fisiologia , Envelhecimento/patologia , Animais , Encéfalo/anatomia & histologia , Dano ao DNA , Kisspeptinas/metabolismo , Masculino , Modelos Neurológicos , Destreza Motora/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Plasticidade Neuronal , Neuropeptídeo Y/metabolismo , Estresse Oxidativo , Carbonilação Proteica , Ratos , Ratos Wistar , Fatores de Tempo
6.
Exp Cell Res ; 322(2): 324-34, 2014 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-24485912

RESUMO

CARF is an ARF-binding protein that has been shown to regulate the p53-p21-HDM2 pathway. CARF overexpression was shown to cause growth arrest of human cancer cells and premature senescence of normal cells through activation of the p53 pathway. Because replicative senescence involves permanent withdrawal from the cell cycle in response to DNA damage response-mediated signaling, in the present study we investigated the relationship between CARF and the cell cycle and whether it is involved in the DNA damage response. We demonstrate that the half-life of CARF protein is less than 60 min, and that in cycling cells CARF levels are highest in G2 and early prophase. Serially passaged normal human skin and stromal fibroblasts showed upregulation of CARF during replicative senescence. Induction of G1 growth arrest and senescence by a variety of drugs was associated with increase in CARF expression at the transcriptional and translational level and was seen to correlate with increase in DNA damage response and checkpoint proteins, ATM, ATR, CHK1, CHK2, γH2AX, p53 and p21. Induction of growth arrest by oncogenic RAS and shRNA-mediated knockdown of TRF2 in cancer cells also caused upregulation of CARF. We conclude that CARF is associated with DNA damage response and checkpoint signaling pathways.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Pontos de Checagem do Ciclo Celular/fisiologia , Dano ao DNA/fisiologia , Fibroblastos/metabolismo , Proteínas de Ligação a RNA/metabolismo , Pele/metabolismo , Células Estromais/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/genética , Western Blotting , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Senescência Celular/fisiologia , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Imunofluorescência , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Neoplasias/patologia , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Proteínas de Ligação a RNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Pele/citologia , Pele/efeitos dos fármacos , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Proteína 2 de Ligação a Repetições Teloméricas/antagonistas & inibidores , Proteína 2 de Ligação a Repetições Teloméricas/genética
7.
BMC Genomics ; 14 Suppl 5: S6, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24564733

RESUMO

BACKGROUND: Coffee contains several compounds that have the potential to influence breast cancer risk and survival. However, epidemiologic data on the relation between coffee compounds and breast cancer survival are sparse and inconsistent. RESULTS: We show that coffee component HHQ has significant apoptotic effect on MDA-MB-231 and MCF-7 cells in vitro, and that ROS generation, change in mitochondrial membrane permeability, upregulation of Bax and Caspase-8 as well as down regulation of PGK1 and PKM2 expression may be important apoptosis-inducing mechanisms. The results suggest that PPARγ ligands may serve as potential therapeutic agents for breast cancer therapy. HHQ was also validated as a ligand for PPARγ by docking procedure. CONCLUSION: This is the first report on the anti-breast cancer (in vitro) activity of HHQ.


Assuntos
Neoplasias da Mama/metabolismo , Café/química , Hidroquinonas/farmacologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , PPAR gama/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Neoplasias da Mama/genética , Caspase 8/metabolismo , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Hidroquinonas/química , Células MCF-7 , Potencial da Membrana Mitocondrial/genética , Modelos Moleculares , Simulação de Acoplamento Molecular , PPAR gama/química , Fosfoglicerato Quinase/metabolismo , Rosiglitazona , Tiazolidinedionas/química , Tiazolidinedionas/farmacologia
8.
Cancer Res ; 83(20): 3442-3461, 2023 10 13.
Artigo em Inglês | MEDLINE | ID: mdl-37470810

RESUMO

Although external beam radiotherapy (xRT) is commonly used to treat central nervous system (CNS) tumors in patients of all ages, young children treated with xRT frequently experience life-altering and dose-limiting neurocognitive impairment (NI) while adults do not. The lack of understanding of mechanisms responsible for these differences has impeded the development of neuroprotective treatments. Using a newly developed mouse model of xRT-induced NI, we found that neurocognitive function is impaired by ionizing radiation in a dose- and age-dependent manner, with the youngest animals being most affected. Histologic analysis revealed xRT-driven neuronal degeneration and cell death in neurogenic brain regions in young animals but not adults. BH3 profiling showed that neural stem and progenitor cells, neurons, and astrocytes in young mice are highly primed for apoptosis, rendering them hypersensitive to genotoxic damage. Analysis of single-cell RNA sequencing data revealed that neural cell vulnerability stems from heightened expression of proapoptotic genes including BAX, which is associated with developmental and mitogenic signaling by MYC. xRT induced apoptosis in primed neural cells by triggering a p53- and PUMA-initiated, proapoptotic feedback loop requiring cleavage of BID and culminating in BAX oligomerization and caspase activation. Notably, loss of BAX protected against apoptosis induced by proapoptotic signaling in vitro and prevented xRT-induced apoptosis in neural cells in vivo as well as neurocognitive sequelae. On the basis of these findings, preventing xRT-induced apoptosis specifically in immature neural cells by blocking BAX, BIM, or BID via direct or upstream mechanisms is expected to ameliorate NI in pediatric patients with CNS tumor. SIGNIFICANCE: Age- and differentiation-dependent apoptotic priming plays a pivotal role in driving radiotherapy-induced neurocognitive impairment and can be targeted for neuroprotection in pediatric patients.


Assuntos
Proteínas Reguladoras de Apoptose , Apoptose , Animais , Criança , Pré-Escolar , Humanos , Camundongos , Apoptose/fisiologia , Proteínas Reguladoras de Apoptose/metabolismo , Proteína X Associada a bcl-2/metabolismo , Morte Celular , Transdução de Sinais , Proteína Supressora de Tumor p53/genética
9.
Sci Adv ; 8(45): eabn6579, 2022 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-36351019

RESUMO

Although major organ toxicities frequently arise in patients treated with cytotoxic or targeted cancer therapies, the mechanisms that drive them are poorly understood. Here, we report that vascular endothelial cells (ECs) are more highly primed for apoptosis than parenchymal cells across many adult tissues. Consequently, ECs readily undergo apoptosis in response to many commonly used anticancer agents including cytotoxic and targeted drugs and are more sensitive to ionizing radiation and BH3 mimetics than parenchymal cells in vivo. Further, using differentiated isogenic human induced pluripotent stem cell models of ECs and vascular smooth muscle cells (VSMCs), we find that these vascular cells exhibit distinct drug toxicity patterns, which are linked to divergent therapy-induced vascular toxicities in patients. Collectively, our results demonstrate that vascular cells are highly sensitive to apoptosis-inducing stress across life span and may represent a "weakest link" vulnerability in multiple tissues for development of toxicities.


Assuntos
Células-Tronco Pluripotentes Induzidas , Neoplasias , Adulto , Humanos , Músculo Liso Vascular/fisiologia , Células Endoteliais , Longevidade , Células-Tronco Pluripotentes Induzidas/fisiologia , Células Cultivadas , Neoplasias/etiologia
10.
Biogerontology ; 11(2): 197-209, 2010 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19609710

RESUMO

Numerous reports implicate increased oxidative stress in the functional and structural changes occurring in the brain and other organs as a part of the normal aging process. Dietary restriction (DR) has long been shown to be life-prolonging intervention in several species. This study was aimed to assess the potential efficacy of late-onset short term DR when initiated in 21 months old male wistar rats for 3 months on the antioxidant defense system and lipid peroxidation, cellular stress response protein HSP 70 and synaptic marker protein synapsin 1 in discrete brain regions such as cortex, hypothalamus, and hippocampus as well as liver, kidney and heart from 24 month old rats. Age-associated decline in activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione, and elevated levels of lipid peroxidation was observed in brain and peripheral organ as well as increased expression of HSP 70 and reduction in synapsin 1 was observed in brain studied. Late-onset short term DR was effective in partially restoring the antioxidant status and in decreasing lipid peroxidation level as well as enhancing the expression of HSP 70 and synapsin 1 in aged rats. Late onset short term DR also prevented age-related neurodegeneration as revealed by Fluoro-Jade B staining in hippocampus and cortex regions of rat brain. Thus our current results suggest that DR initiated even in old age has the potential to improve age related decline in body functions.


Assuntos
Envelhecimento/fisiologia , Encéfalo/metabolismo , Dieta Redutora , Proteínas de Choque Térmico HSP70/metabolismo , Estresse Oxidativo/fisiologia , Sinapsinas/metabolismo , Animais , Catalase/metabolismo , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Rim/metabolismo , Peroxidação de Lipídeos/fisiologia , Fígado/metabolismo , Masculino , Modelos Animais , Miocárdio/metabolismo , Ratos , Ratos Wistar , Superóxido Dismutase/metabolismo
11.
Artigo em Inglês | MEDLINE | ID: mdl-31097976

RESUMO

Skin color/pigmentation is regulated through melanogenesis process in specialized melanin-producing cells, melanocytes, involving multiple signaling pathways. It is highly influenced by intrinsic and extrinsic factors such as oxidative, ultraviolet radiations and other environmental stress conditions. Besides determining the color, it governs response and tolerance of skin to a variety of environmental stresses and pathological conditions including photodamage, hyperpigmentation, and skin cancer. Depigmenting reagents have been deemed useful not only for cosmetics but also for pigmentation-related pathologies. In the present study, we attempted modulation of 1-oleoyl-2-acetyl-glycerol- (OAG-) induced melanogenesis in human melanoma and primary melanocytes. In both cell types, OAG-induced melanogenesis was associated with increase in enhanced expression of melanin, tyrosinase, as well as stress chaperones (mortalin and HSP60) and Reactive Oxygen Species (ROS). Treatment with TXC (trans-4-(Aminomethyl) cyclohexanecarboxylic acid hexadecyl ester hydrochloride) and 5/40 natural compounds resulted in their reduction. The data proposed an important role of mortalin and oxidative stress in skin pigmentation and the use of TXC and natural extracts for modulation of pigmentation pathways in normal and pathological conditions.

12.
Nat Cell Biol ; 20(4): 413-421, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29531308

RESUMO

Ageing of haematopoietic stem cells (HSCs) contributes to deficits in the aged haematopoietic system. HSC decline is driven in part by DNA damage accumulation; yet, how ageing impacts the acute DNA damage response (DDR) of HSCs is poorly understood. We show that old HSCs exhibit diminished ATM activity and attenuated DDR, leading to elevated clonal survival in response to a range of genotoxins that was underwritten by diminished apoptotic priming. Distinct HSC subsets exhibited ageing-dependent and subtype-dependent differences in apoptotic priming and survival in response to DNA damage. The defective DDR of old HSCs was non-cell autonomous, as ATM signalling and clonal survival in response to DNA damage could be restored to levels observed in young HSCs post-transplantated into young recipients. These data indicate that defective DDR and diminished apoptotic priming provide a selective advantage to old HSCs that may contribute to mutation accrual and disease predisposition.


Assuntos
Apoptose , Senescência Celular , Dano ao DNA , Reparo do DNA , Células-Tronco Hematopoéticas/enzimologia , Fatores Etários , Animais , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Células Cultivadas , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/patologia , Masculino , Camundongos Endogâmicos C57BL , Transdução de Sinais , Fatores de Tempo
13.
Mol Neurobiol ; 54(4): 3050-3061, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-27037574

RESUMO

Modern lifestyle and sustained stress of professional commitments in the current societal set up often disrupts the normal sleep cycle and duration which is known to lead to cognitive impairments. In the present study, we report whether leaf extract of Withania somnifera (Ashwagandha) has potential neuroprotective role in acute stress of sleep deprivation. Experiments were performed on three groups of adult Wistar rats: group 1 (vehicle treated-undisturbed sleep [VUD]), group 2 (vehicle treated-sleep deprived [VSD]), and group 3 (ASH-WEX treated-sleep deprived [WSD]). Groups 1 and 2 received single oral feeding of vehicle and group 3 received ASH-WEX orally (140 mg/kg or 1 ml/250 g of body weight) for 15 consecutive days. Immediately after this regimen, animals from group 1 were allowed undisturbed sleep (between 6 a.m. and 6 p.m.), whereas rats of groups 2 and 3 were deprived of sleep during this period. We observed that WSD rats showed significant improvement in their performance in behavioral tests as compared to VSD group. At the molecular level, VSD rats showed acute change in the expression of proteins involved in synaptic plasticity, cell survival, and apoptosis in the hippocampus region of brain, which was suppressed by ASH-WEX treatment thus indicating decreased cellular stress and apoptosis in WSD group. This data suggest that Ashwagandha may be a potential agent to suppress the acute effects of sleep loss on learning and memory impairments and may emerge as a novel supplement to control SD-induced cognitive impairments.


Assuntos
Fármacos Neuroprotetores/uso terapêutico , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Privação do Sono/tratamento farmacológico , Withania/química , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Sobrevivência Celular/efeitos dos fármacos , Feminino , Homeostase/efeitos dos fármacos , Imuno-Histoquímica , Memória/efeitos dos fármacos , Modelos Biológicos , Atividade Motora/efeitos dos fármacos , Proteínas do Tecido Nervoso/metabolismo , Plasticidade Neuronal/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Extratos Vegetais/farmacologia , Ratos Wistar , Privação do Sono/patologia , Privação do Sono/fisiopatologia , Estresse Fisiológico/efeitos dos fármacos
14.
J Gerontol A Biol Sci Med Sci ; 70(6): 701-13, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24747666

RESUMO

The heat shock 70 family protein, mortalin, has pancytoplasmic distribution pattern in normal and perinuclear in cancer human cells. Cancer cells when induced to senesce by either chemicals or stress showed shift in mortalin staining pattern from perinuclear to pancytoplasmic type. Using such shift in mortalin staining as a reporter, we screened human shRNA library and identified nine senescence-inducing siRNA candidates. An independent Comparative Genomic Hybridization analysis of 35 breast cancer cell lines revealed that five (NBS1, BRCA1, TIN2, MRE11A, and KPNA2) of the nine genes located on chromosome regions identified as the gain of locus in more than 80% cell lines. By gene-specific PCR, these five genes were found to be frequently amplified in cancer cell lines. Bioinformatics revealed that the identified targets were connected to MRN (MRE11-RAD50-NBS1) complex, the DNA damage-sensing complex. We demonstrate that the identified shRNAs triggered DNA damage response and induced the expression of tumor suppressor protein p16(INK4A) causing growth arrest of cancer cells. Furthermore, cells showed decreased migration, mediated by decrease in matrix metalloproteases. Taken together, we demonstrate that the MRN complex is a potential target of cancer cell proliferation and migration, and staining pattern of mortalin could serve as an assay to identify senescence-inducing/anticancer reagents.


Assuntos
Movimento Celular , Senescência Celular , Dano ao DNA , Proteínas de Choque Térmico HSP70/metabolismo , Neoplasias/patologia , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Proteínas de Ligação a DNA/genética , Biblioteca Gênica , Humanos , Proteína Homóloga a MRE11 , Neoplasias/genética , Proteínas Nucleares , RNA de Cadeia Dupla , Proteínas de Ligação a Telômeros , Ubiquitina-Proteína Ligases , alfa Carioferinas
15.
PLoS One ; 10(3): e0120554, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25789768

RESUMO

BACKGROUND: Ashwagandha, a traditional Indian herb, has been known for its variety of therapeutic activities. We earlier demonstrated anticancer activities in the alcoholic and water extracts of the leaves that were mediated by activation of tumor suppressor functions and oxidative stress in cancer cells. Low doses of these extracts were shown to possess neuroprotective activities in vitro and in vivo assays. METHODOLOGY/PRINCIPAL FINDINGS: We used cultured glioblastoma and neuroblastoma cells to examine the effect of extracts (alcoholic and water) as well as their bioactive components for neuroprotective activities against oxidative stress. Various biochemical and imaging assays on the marker proteins of glial and neuronal cells were performed along with their survival profiles in control, stressed and recovered conditions. We found that the extracts and one of the purified components, withanone, when used at a low dose, protected the glial and neuronal cells from oxidative as well as glutamate insult, and induced their differentiation per se. Furthermore, the combinations of extracts and active component were highly potent endorsing the therapeutic merit of the combinational approach. CONCLUSION: Ashwagandha leaf derived bioactive compounds have neuroprotective potential and may serve as supplement for brain health.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ácido Glutâmico/toxicidade , Humanos , Peróxido de Hidrogênio/toxicidade , Microscopia de Fluorescência , Fármacos Neuroprotetores/química , Extratos Vegetais/química , Folhas de Planta/química , Folhas de Planta/metabolismo , Ratos , Withania/química , Withania/metabolismo
16.
PLoS One ; 8(10): e77189, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24130852

RESUMO

BACKGROUND: Cancer is a leading cause of death accounting for 15-20% of global mortality. Although advancements in diagnostic and therapeutic technologies have improved cancer survival statistics, 75% of the world population live in underdeveloped regions and have poor access to the advanced medical remedies. Natural therapies hence become an alternative choice of treatment. Ashwagandha, a tropical herb used in Indian Ayurvedic medicine, has a long history of its health promoting and therapeutic effects. In the present study, we have investigated an anticancer activity in the water extract of Ashwagandha leaves (ASH-WEX). METHODOLOGY/PRINCIPAL FINDINGS: Anticancer activity in the water extract of Ashwagandha leaves (ASH-WEX) was detected by in vitro and in vivo assays. Bioactivity-based size fractionation and NMR analysis were performed to identify the active anticancer component(s). Mechanism of anticancer activity in the extract and its purified component was investigated by biochemical assays. We report that the ASH-WEX is cytotoxic to cancer cells selectively, and causes tumor suppression in vivo. Its active anticancer component was identified as triethylene glycol (TEG). Molecular analysis revealed activation of tumor suppressor proteins p53 and pRB by ASH-WEX and TEG in cancer cells. In contrast to the hypophosphorylation of pRB, decrease in cyclin B1 and increase in cyclin D1 in ASH-WEX and TEG-treated cancer cells (undergoing growth arrest), normal cells showed increase in pRB phosphorylation and cyclin B1, and decrease in cyclin D1 (signifying their cell cycle progression). We also found that the MMP-3 and MMP-9 that regulate metastasis were down regulated in ASH-WEX and TEG-treated cancer cells; normal cells remained unaffected. CONCLUSION: We provide the first molecular evidence that the ASH-WEX and TEG have selective cancer cell growth arrest activity and hence may offer natural and economic resources for anticancer medicine.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Folhas de Planta/química , Água/química , Withania/química , Animais , Antineoplásicos/análise , Antineoplásicos/isolamento & purificação , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Camundongos , Extratos Vegetais/análise , Extratos Vegetais/isolamento & purificação , Solventes/química , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Age (Dordr) ; 34(4): 917-33, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21861096

RESUMO

Lifelong dietary restriction (DR) is known to have many potential beneficial effects on brain function as well as delaying the onset of neurological diseases. In the present investigation, the effect of late-onset short-term intermittent fasting dietary restriction (IF-DR) regimen was studied on motor coordination and cognitive ability of ageing male rats. These animals were further used to estimate protein carbonyl content and mitochondrial complex I-IV activity in different regions of brain and peripheral organs, and the degree of age-related impairment and reversion by late-onset short-term IF-DR was compared with their levels in 3-month-old young rats. The results of improvement in motor coordination by rotarod test and cognitive skills by Morris water maze in IF-DR rats were found to be positively correlated with the decline in the oxidative molecular damage to proteins and enhanced mitochondrial complex IV activity in different regions of ageing brain as well as peripheral organs. The work was further extended to study the expression of synaptic plasticity-related proteins, such as synaptophysin, calcineurin and CaM kinase II to explore the molecular basis of IF-DR regimen to improve cognitive function. These results suggest that even late-onset short-term IF-DR regimen have the potential to retard age-associated detrimental effects, such as cognitive and motor performance as well as oxidative molecular damage to proteins.


Assuntos
Comportamento Animal , Encéfalo/patologia , Restrição Calórica/métodos , Cognição/fisiologia , Jejum , Estresse Oxidativo/fisiologia , Envelhecimento/metabolismo , Envelhecimento/fisiologia , Análise de Variância , Animais , Western Blotting , Encéfalo/metabolismo , Modelos Animais de Doenças , Imuno-Histoquímica , Masculino , Aprendizagem em Labirinto , Destreza Motora , Oxirredução , Carbonilação Proteica , Distribuição Aleatória , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real/métodos , Valores de Referência , Fatores de Tempo
18.
PLoS One ; 7(1): e30890, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22303466

RESUMO

Cancer is largely marked by genetic instability. Specific inhibition of individual proteins or signalling pathways that regulate genetic stability during cell division thus hold a great potential for cancer therapy. The Aurora A kinase is a Ser/Thr kinase that plays a critical role during mitosis and cytokinesis and is found upregulated in several cancer types. It is functionally regulated by its interactions with TPX2, a candidate oncogene. Aurora A inhibitors have been proposed as anticancer drugs that work by blocking its ATP binding site. This site is common to other kinases and hence these inhibitors lack specificity for Aurora A inhibition in particular, thus advocating the need of some alternative inhibition route. Previously, we identified TPX2 as a cellular target for withanone that selectively kill cancer cells. By computational approach, we found here that withanone binds to TPX2-Aurora A complex. In experiment, withanone treatment to cancer cells indeed resulted in dissociation of TPX2-Aurora A complex and disruption of mitotic spindle apparatus proposing this as a mechanism of the anticancer activity of withanone. From docking analysis, non-formation/disruption of the active TPX2-Aurora A association complex could be discerned. Our MD simulation results suggesting the thermodynamic and structural stability of TPX2-Aurora A in complex with withanone further substantiates the binding. We report a computational rationale of the ability of naturally occurring withanone to alter the kinase signalling pathway in an ATP-independent manner and experimental evidence in which withanone cause inactivation of the TPX2-Aurora A complex. The study demonstrated that TPX2-Aurora A complex is a target of withanone, a potential natural anticancer drug.


Assuntos
Antineoplásicos/farmacologia , Proteínas de Ciclo Celular/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Simulação de Dinâmica Molecular , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Triterpenos/farmacologia , Withania/química , Antineoplásicos/química , Aurora Quinases , Bioensaio , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/genética , Linhagem Celular Tumoral , Análise por Conglomerados , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Proteínas Associadas aos Microtúbulos/química , Proteínas Associadas aos Microtúbulos/genética , Proteínas Nucleares/química , Proteínas Nucleares/genética , Extratos Vegetais , Proteínas Serina-Treonina Quinases/química , Proteínas Serina-Treonina Quinases/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reprodutibilidade dos Testes , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Triterpenos/química , Vitanolídeos
19.
PLoS One ; 7(9): e44419, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22973447

RESUMO

BACKGROUND AND PURPOSE: Withanolides are naturally occurring chemical compounds. They are secondary metabolites produced via oxidation of steroids and structurally consist of a steroid-backbone bound to a lactone or its derivatives. They are known to protect plants against herbivores and have medicinal value including anti-inflammation, anti-cancer, adaptogenic and anti-oxidant effects. Withaferin A (Wi-A) and Withanone (Wi-N) are two structurally similar withanolides isolated from Withania somnifera, also known as Ashwagandha in Indian Ayurvedic medicine. Ashwagandha alcoholic leaf extract (i-Extract), rich in Wi-N, was shown to kill cancer cells selectively. Furthermore, the two closely related purified phytochemicals, Wi-A and Wi-N, showed differential activity in normal and cancer human cells in vitro and in vivo. We had earlier identified several genes involved in cytotoxicity of i-Extract in human cancer cells by loss-of-function assays using either siRNA or randomized ribozyme library. METHODOLOGY/PRINCIPAL FINDINGS: In the present study, we have employed bioinformatics tools on four genes, i.e., mortalin, p53, p21 and Nrf2, identified by loss-of-function screenings. We examined the docking efficacy of Wi-N and Wi-A to each of the four targets and found that the two closely related phytochemicals have differential binding properties to the selected cellular targets that can potentially instigate differential molecular effects. We validated these findings by undertaking parallel experiments on specific gene responses to either Wi-N or Wi-A in human normal and cancer cells. We demonstrate that Wi-A that binds strongly to the selected targets acts as a strong cytotoxic agent both for normal and cancer cells. Wi-N, on the other hand, has a weak binding to the targets; it showed milder cytotoxicity towards cancer cells and was safe for normal cells. The present molecular docking analyses and experimental evidence revealed important insights to the use of Wi-A and Wi-N for cancer treatment and development of new anti-cancer phytochemical cocktails.


Assuntos
Modelos Moleculares , Extratos Vegetais/metabolismo , Folhas de Planta/química , Triterpenos/metabolismo , Withania/química , Vitanolídeos/metabolismo , Western Blotting , Morte Celular/efeitos dos fármacos , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Biologia Computacional , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Humanos , Imunoprecipitação , Ayurveda , Estrutura Molecular , Fator 2 Relacionado a NF-E2/metabolismo , Sais de Tetrazólio , Tiazóis , Triterpenos/química , Triterpenos/farmacologia , Proteína Supressora de Tumor p53/metabolismo , Vitanolídeos/química , Vitanolídeos/farmacologia , beta-Galactosidase
20.
PLoS One ; 6(11): e27265, 2011.
Artigo em Inglês | MEDLINE | ID: mdl-22096544

RESUMO

BACKGROUND: Scopolamine is a well-known cholinergic antagonist that causes amnesia in human and animal models. Scopolamine-induced amnesia in rodent models has been widely used to understand the molecular, biochemical, behavioral changes, and to delineate therapeutic targets of memory impairment. Although this has been linked to the decrease in central cholinergic neuronal activity following the blockade of muscarinic receptors, the underlying molecular and cellular mechanism(s) particularly the effect on neuroplasticity remains elusive. In the present study, we have investigated (i) the effects of scopolamine on the molecules involved in neuronal and glial plasticity both in vivo and in vitro and (ii) their recovery by alcoholic extract of Ashwagandha leaves (i-Extract). METHODOLOGY/PRINCIPAL FINDINGS: As a drug model, scopolamine hydrobromide was administered intraperitoneally to mice and its effect on the brain function was determined by molecular analyses. The results showed that the scopolamine caused downregulation of the expression of BDNF and GFAP in dose and time dependent manner, and these effects were markedly attenuated in response to i-Extract treatment. Similar to our observations in animal model system, we found that the scopolamine induced cytotoxicity in IMR32 neuronal and C6 glioma cells. It was associated with downregulation of neuronal cell markers NF-H, MAP2, PSD-95, GAP-43 and glial cell marker GFAP and with upregulation of DNA damage--γH2AX and oxidative stress--ROS markers. Furthermore, these molecules showed recovery when cells were treated with i-Extract or its purified component, withanone. CONCLUSION: Our study suggested that besides cholinergic blockade, scopolamine-induced memory loss may be associated with oxidative stress and Ashwagandha i-Extract, and withanone may serve as potential preventive and therapeutic agents for neurodegenerative disorders and hence warrant further molecular analyses.


Assuntos
Encéfalo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Folhas de Planta/química , Escopolamina/toxicidade , Triterpenos/farmacologia , Withania/química , Animais , Encéfalo/metabolismo , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Dano ao DNA/efeitos dos fármacos , Proteína Glial Fibrilar Ácida , Camundongos , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Vitanolídeos
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