RESUMO
Mesenchymal stem cells (MSC) are potentially a good material for transplantation in many diseases, including neurodegenerative diseases. The main problem with using them is the low percentage of surviving cells after the transplant procedure and the naturally poor ability of MSC to spontaneously differentiate into certain types of cells, which results in their poor integration with the host cells. The aim and the novelty of this work consists in the synergistic overexpression of two genes, BCL2 and BDNF, using lentiviral vectors. According to our hypothesis, the overexpression of the BCL2 gene is aimed at increasing the resistance of cells to stressors and toxic factors. In turn, the overexpression of the BDNF gene is suspected to direct the MSC into the neural differentiation pathway. As a result, it was shown that the overexpression of both genes and the overproduction of proteins is permanent and persists for at least 60 days. The synergistically transduced MSC were significantly more resistant to the action of staurosporine; 12 days after transduction, the synergistically transduced MSC had a six-times greater survival rate. The overexpression of the Bcl-2 and BDNF proteins was sufficient to stimulate a significant overexpression of the CHAT gene, and under specific conditions, the TH, TPH1, and SYP genes were also overexpressed. Modified MSC are able to differentiate into cholinergic and dopaminergic neurons, and the release of acetylcholine and dopamine may indicate their functionality.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/biossíntese , Diferenciação Celular , Neurônios Dopaminérgicos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Fator Neurotrófico Derivado do Encéfalo/genética , Neurônios Dopaminérgicos/citologia , Humanos , Lentivirus , Células-Tronco Mesenquimais/citologia , Proteínas Proto-Oncogênicas c-bcl-2/genética , Transdução GenéticaRESUMO
PURPOSE: The blood-brain barrier limits the application of idarubicin in the therapy of glioblastoma multiforme. Biodegradable, intracranial wafers with prolonged release may increase therapy efficiency. METHODS: Blank wafers, wafers containing 5% w/w and 10% w/w of idarubicin were formulated by solution casting from poly(L-lactide-co-glycolide) and poly(glycolide-co-ε-caprolactone). The following methods were used: NMR, GPC, DSC, FTIR, AFM, UV-VIS, and a viability and proliferation assay for idarubicin action (U87MG cell line). RESULTS: Wafers showed a surface with numerous immersions and hills. A lack of interactions between idarubicin and the copolymers was observed. The substance was entrapped in the matrix and released in two phases for all wafers with the appropriate bolus and maintenance dose. The burst effect was observed for all wafers, however, the biggest bolus for poly(L-lactide-co-glycolide) wafers containing 5% w/w of idarubicin was noted. The stable and steady degradation of poly(glycolide-co-ε-caprolactone) wafers containing 5% w/w of idarubicin ensures the most optimal release profile and high inhibition of proliferation. CONCLUSIONS: Copolymer wafers with idarubicin are an interesting proposition with great potential for the local treatment of glioblastoma multiforme. The release rate and dose may be regulated by the amount and kind of wafers for various effects.
Assuntos
Portadores de Fármacos/síntese química , Glioblastoma/tratamento farmacológico , Idarubicina/uso terapêutico , Polímeros/síntese química , Linhagem Celular Tumoral , Sobrevivência Celular , Liberação Controlada de Fármacos , Humanos , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Tecnologia Farmacêutica/métodosRESUMO
Static magnetic field (SMF) is widely used in industry, in consumer devices and diagnostic medical equipment, hence the widespread exposure to SMF in the natural environment and in people occupationally exposed to it. In environment and in some workplaces, there is a risk of exposure also to various chemicals. Environmental factors can affect the cellular processes which can be the cause of the development of various pathological conditions. Therefore, the aim of this study was to assess the effect of SMF on the expression of the apoptosis-related genes in human fibroblast cultures that had been co-treated with fluoride ions. The control and NaF-treated cells were subjected to the influence of SMF with a moderate induction. The flow-cytometric analysis showed that the fluoride ions reduced the number of viable cells and induced early apoptosis. However, exposure to the SMF reduced the number of dead cells that had been treated with fluoride ions. Moreover, specific genes that were involved in apoptosis exhibited a differential expression in the NaF-treated cells and exposure to the SMF yielded a modulation of their transcriptional activity. Our results suggest some beneficial properties of using a moderate-intensity static magnetic field to reduce the adverse effects of fluoride.
Assuntos
Apoptose/efeitos dos fármacos , Poluentes Ambientais/toxicidade , Fibroblastos/efeitos dos fármacos , Campos Magnéticos , Fluoreto de Sódio/toxicidade , Proteínas Reguladoras de Apoptose/genética , Linhagem Celular , Fibroblastos/patologia , Expressão Gênica/efeitos dos fármacos , HumanosRESUMO
AIM: Many data showed a role of inflammation and dysfunction of immune system as important factors in the risk of schizophrenia. The TNFR2 receptor is a molecule that adapts to both areas. Tumor necrosis factor receptor 2 (TNFR2) is a receptor for the TNF-α cytokine which is a strong candidate gene for schizophrenia. The serum level of TNFR2 was significantly increased in schizophrenia and associated with more severe symptoms of schizophrenia. METHODS: We examined the association of the three single nucleotide polymorphisms (rs3397, rs1061622, and rs1061624) in TNFR2 gene with a predisposition to and psychopathology of paranoid schizophrenia in Caucasian population. The psychopathology was measured by a five-factor model of the PANSS scale. We also assessed a haplotype analysis with the -308G/A of TNF-α gene. RESULTS: Our case-control study (401 patients and 657 controls) revealed that the genetic variants of rs3397, rs1061622, and rs1061624 in the TNFR2 gene are associated with a higher risk of developing schizophrenia and more severe course in men. However, the genotypes with polymorphic allele for rs3397 SNP are protective for women. The rs1061624 SNP might modulate the appearance of the disease in relatives of people with schizophrenia. The CTGG haplotype build with tested SNPs of TNFR2 and SNP -308G/A of TNF-α has an association with a risk of schizophrenia in Caucasian population depending on sex. Our finding is especially true for the paranoid subtypes of schizophrenia.
Assuntos
Receptores Tipo II do Fator de Necrose Tumoral/genética , Esquizofrenia/genética , Fator de Necrose Tumoral alfa/genética , Adulto , Alelos , Estudos de Casos e Controles , Feminino , Predisposição Genética para Doença/genética , Genótipo , Haplótipos/genética , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genéticaRESUMO
The microglial hypothesis of schizophrenia suggests that its neuropathology is closely associated with neuroinflammation manifested, inter alia, by an increased expression of cytokines. However, clinical investigations imply that schizophrenia is a heterogeneous disease and in some groups of patients the activated inflammatory process does not contribute to the disease-associated impairment of brain function. Clinical studies revealed also an equivocal impact of antipsychotics on peripheral and CSF cytokines, whereas experimental research performed on the stimulated glia cultures showed their inhibitory effect on pro-inflammatory cytokine levels. In the present study, the effect of chlorpromazine, haloperidol and risperidone (0.5, 5 or 10µM) on production of pro-inflammatory cytokines IL-1ß and TNF-α and anti-inflammatory IL-10 was investigated in the unstimulated and lipopolysaccharide-stimulated primary rat mixed glial cell cultures. In the unstimulated cultures, haloperidol at all applied concentrations, risperidone at 5, 10µM and chlorpromazine at 10µM increased IL-10 levels in the culture supernatants without a significant influence on IL-1ß or TNF-α levels, and all drugs applied at 10µM induced a robust increase in IL-10 mRNA expression. Under strong inflammatory activation, haloperidol and risperidone at all concentrations reduced production of both pro-inflammatory cytokines, without adverse effects on IL-10 expression when used at 10µM. Chlorpromazine at all concentrations diminished the production of three cytokines and did not induce anti-inflammatory effect. These results suggest that dependently on glia activation antipsychotics via different mechanisms may induce anti-inflammatory effect and that this activity is not common for all drugs under conditions of strong glia activation.
Assuntos
Antipsicóticos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Citocinas/metabolismo , Neuroglia/efeitos dos fármacos , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Clorpromazina/farmacologia , Meios de Cultura/química , Citocinas/genética , Haloperidol/farmacologia , Técnicas In Vitro , Inflamação , Interleucina-10/genética , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Lipopolissacarídeos/farmacologia , Neuroglia/fisiologia , Ratos , Risperidona/farmacologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The present study was designed to evaluate, for the first time, the potential sex differences in BDNF and VEGF systems under normal conditions and in response to IL-1ß given ip. Peripheral overproduction of this cytokine mediates the pathophysiology of various acute neuroinflammatory states. Until now, the effect of IL-1ß on VEGF expression in rat brain structures and its serum level has not been examined. In male and female rats, the BDNF and VEGF mRNA expression, and BDNF level were evaluated in the amygdala, hippocampus, hypothalamus and pituitary gland. The VEGF levels were determined in the pituitary. Serum BDNF and VEGF levels were also measured. The pituitary BDNF mRNA, and BDNF and VEGF levels were higher in females than in male rats whereas in males, the BDNF levels were higher in the other brain structures. The serum BDNF concentration was similar in both groups but VEGF levels were enhanced in females. Following IL-1ß (50µg/kg ip.) administration, a higher serum IL-1ß level was detected in females than in males. In male rats, IL-1ß decreased BDNF mRNA in all the brain structures, except for the pituitary, and VEGF mRNA in the amygdala. In opposite, IL-1ß challenge in females increased the pituitary VEGF mRNA and serum BDNF and VEGF levels. These results suggest that in females BDNF and VEGF may play a more important role in the pituitary function. In males, amygdala trophic system seems to be especially sensitive to the enhanced peripheral IL-1ß production. Our findings point to the need to consider sex-related differences to be able to draw reliable conclusions about changes in BDNF and VEGF levels during inflammation.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/sangue , Encéfalo/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-1beta/farmacologia , Caracteres Sexuais , Fator A de Crescimento do Endotélio Vascular/sangue , Animais , Feminino , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Schizophrenia is a devastating mental disorder with undetermined aetiology. Previous research has suggested that dysregulation of proinflammatory cytokines and their receptors plays a role in developing schizophrenia. We examined the association of the three single nucleotide polymorphisms (SNPs; rs4149576, rs4149577, and rs1860545) in the tumor necrosis factor receptor 1 (TNFR1) gene with the development and psychopathology of paranoid schizophrenia in the Polish Caucasian sample consisting of 388 patients and 657 control subjects. The psychopathology was assessed using a five-factor model of the Positive and Negative Syndrome Scale (PANSS). SNPs were genotyped using the TaqMan 5'-exonuclease allelic discrimination assay. The SNPs tested were not associated with a predisposition to paranoid schizophrenia in either the entire sample or after stratification according to gender. However, rs4149577 and rs1860545 SNPs were associated with the intensity of the PANSS excitement symptoms in men, which may contribute to the risk of violent behavior. Polymorphisms in the TNFR1 gene may have an impact on the symptomatology of schizophrenia in men.
Assuntos
Polimorfismo de Nucleotídeo Único , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Esquizofrenia Paranoide/genética , Adolescente , Adulto , Idade de Início , Idoso , Estudos de Casos e Controles , Feminino , Frequência do Gene , Estudos de Associação Genética , Predisposição Genética para Doença , Humanos , Masculino , Pessoa de Meia-Idade , Polônia , Esquizofrenia Paranoide/psicologia , Fatores Sexuais , Tentativa de Suicídio , Violência , População Branca , Adulto JovemRESUMO
Adipose tissue is a promising source of mesenchymal stem cells. Their potential to differentiate and regenerate other types of tissues may be affected by several factors. This may be due to in vitro cell-culture conditions, especially the supplementation with antibiotics. The aim of our study was to evaluate the effects of a penicillin-streptomycin mixture (PS), amphotericin B (AmB), a complex of AmB with copper (II) ions (AmB-Cu2+) and various combinations of these antibiotics on the proliferation and differentiation of adipose-derived stem cells in vitro. Normal human adipose-derived stem cells (ADSC, Lonza) were routinely maintained in a Dulbecco's Modified Eagle Medium (DMEM) that was either supplemented with selected antibiotics or without antibiotics. The ADSC that were used for the experiment were at the second passage. The effect of antibiotics on proliferation was analyzed using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and sulforhodamine-B (SRB) tests. Differentiation was evaluated based on Alizarin Red staining, Oil Red O staining and determination of the expression of ADSC, osteoblast and adipocyte markers by real-time RT-qPCR. The obtained results indicate that the influence of antibiotics on adipose-derived stem cells depends on the duration of exposure and on the combination of applied compounds. We show that antibiotics alter the proliferation of cells and also promote natural osteogenesis, and adipogenesis, and that this effect is also noticeable in stimulated osteogenesis.
Assuntos
Anfotericina B/farmacologia , Antibacterianos/farmacologia , Diferenciação Celular , Proliferação de Células , Células-Tronco Mesenquimais/efeitos dos fármacos , Penicilinas/farmacologia , Estreptomicina/farmacologia , Adipócitos/citologia , Tecido Adiposo/citologia , Células Cultivadas , Humanos , Células-Tronco Mesenquimais/citologia , Osteoblastos/citologiaRESUMO
Abstract: AKT, a serine/threonine protein kinase and mammalian target of rapamycin (mTOR) plays a critical role in the proliferation and resistance to apoptosis that are essential to the development and progression of colon cancer. Therefore, AKT/mTOR signaling pathway has been recognized as an attractive target for anticancer therapy. Inositol hexaphosphate (InsP6), a natural occurring phytochemical, has been shown to have both preventive and therapeutic effects against various cancers, however, its exact molecular mechanisms of action are not fully understood. The aim of the in vitro study was to investigate the anticancer activity of InsP6 on colon cancer with the focus on inhibiting the AKT1 kinase and p70S6K1 as mTOR effector, in relation to proliferation and apoptosis of cells. The colon cancer Caco-2 cells were cultured using standard techniques and exposed to InsP6 at different concentrations (1 mM, 2.5 mM and 5 mM). Cellular proliferative activity was monitored by 5-bromo-2'-deoxyuridine (BrdU) incorporation into cellular DNA. Flow cytometric analysis was performed for cell cycle progression and apoptosis studies. Real-time RT-qPCR was used to validate mRNA levels of CDNK1A, CDNK1B, CASP3, CASP9, AKT1 and S6K1 genes. The concentration of p21 protein as well as the activities of caspase 3, AKT1 and p70S6K1 were determined by the ELISA method. The results revealed that IP6 inhibited proliferation and stimulated apoptosis of colon cancer cells. This effect was mediated by an increase in the expression of genes encoding p21, p27, caspase 3, caspase 9 as well a decrease in transcription of AKT1 and S6K1. InsP6 suppressed phosphorylation of AKT1 and p70S6K1, downstream effector of mTOR. Based on these studies it may be concluded that InsP6 can reduce proliferation and induce apoptosis through inhibition of the AKT/mTOR pathway and mTOR effector followed by modulation of the expression and activity of several key components of these pathways in colon cancer cells.
Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Ácido Fítico/farmacologia , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Células CACO-2 , Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Humanos , Ácido Fítico/química , Ácido Fítico/uso terapêutico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismoRESUMO
INTRODUCTION: Epigenetic mechanisms regulate the transcription of genes, which can affect the differentiation of MSCs. The aim of the current work is to determine how the histone deacetylase inhibitors TSA and VPA affect the expression of neuronal lineage genes in a culture of rat MSCs (rMSCs). MATERIALS AND METHODS: We analyzed the expression of early neuron marker gene (Tubb3), mature neuron markers genes (Vacht, Th, Htr2a) and the oligodendrocyte progenitor marker gene (GalC). Moreover, changes in the gene expression after three different periods of exposure to TSA and VPA were investigated for the first time. RESULTS: After six days of exposition to TSA and VPA, the expression of Tubb3 and GalC decreased, while the expression of Th increased. The highest increase of VAChT expression was observed after three days of TSA and VPA treatment. A decrease in Htr2a gene expression was observed after TSA treatment and an increase was observed after VPA treatment. We also observed that TSA and VPA inhibited cell proliferation and the formation of neurospheres in the rMSCs culture. DISCUSSION: The central findings of our study are that TSA and VPA affect the expression of neuronal lineage genes in an rMSCs culture. After exposure to TSA or VPA, the expression of early neuronal gene decreases but equally the expression of mature neuron genes increases. After TSA and VPA treatment ER of the oligodendrocyte progenitor marker decreased. TSA and VPA inhibit cell proliferation and the formation of neurospheres in rMSCs culture.
Assuntos
Células da Medula Óssea/metabolismo , Expressão Gênica/efeitos dos fármacos , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Neurais/efeitos dos fármacos , Tubulina (Proteína)/metabolismo , Ácido Valproico/farmacocinética , Animais , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Inibidores de Histona Desacetilases/farmacologia , Células-Tronco Mesenquimais/citologia , Neurônios/metabolismo , RatosRESUMO
Nowadays, it is assumed that therapeutic efficacy of antidepressants depends, at least partly, on their anti-inflammatory properties. The present study investigated for the first time the effect of 21-day oral administration of desipramine on the lipopolysaccharide (LPS)-stimulated IL-1ß concentration in the olfactory bulb, hypothalamus, frontal cortex, hippocampus and plasma of rats, and on the LPS-induced IL-1ß mRNA level in the olfactory bulb. Desipramine (15mg/kg/day) reduced significantly the LPS (250 µg/kg i.p.)-induced IL-1ß concentration in the olfactory bulb, hypothalamus and in plasma, and diminished the LPS effect on IL-1ß mRNA in the olfactory bulb. Plasma concentration of desipramine was comparable to its therapeutic range. By using the α1/α2-adrenoceptor antagonist prazosin and the unspecific ß-adrenoceptor antagonist propranolol given prior to LPS, we found that the effect of desipramine on LPS-induced IL-1ß production was partially mediated by both adrenoceptors in the olfactory bulb and plasma, and that ß-adrenoceptors contributed also to its effect on the stimulated IL-1ß concentration in the hypothalamus. The effect of LPS on the cerebral IL-1ß levels was, in part, mediated by ß-adrenoceptors and, in a region-specific manner, by α1/α2-adrenoceptors. The findings provide evidence for central and peripheral anti-inflammatory activity of desipramine and confirm the impact of the noradrenergic system on IL-1ß production induced by an immunostimulatory challenge.
Assuntos
Antidepressivos Tricíclicos/administração & dosagem , Encéfalo/efeitos dos fármacos , Desipramina/administração & dosagem , Interleucina-1beta/sangue , Interleucina-1beta/metabolismo , Lipopolissacarídeos/imunologia , Administração Oral , Animais , Antidepressivos Tricíclicos/sangue , Anti-Hipertensivos/administração & dosagem , Encéfalo/imunologia , Encéfalo/metabolismo , Desipramina/sangue , Esquema de Medicação , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Interleucina-1beta/biossíntese , Interleucina-1beta/genética , Bulbo Olfatório/efeitos dos fármacos , Bulbo Olfatório/metabolismo , Prazosina/administração & dosagem , Propranolol/administração & dosagem , RatosRESUMO
Glioblastoma multiforme (GBM) is the most common primary brain malignancy, having a very poor prognosis and is characterized by extensive brain invasion as well as resistance to the therapy. The phosphoinositide 3-kinase (PI3K)/Akt/PTEN signaling pathway is deregulated in GBM. Besides, florid vascularization and aberrantly elevated vascular endothelial growth factor (VEGF) occur very often. The present study was designed to examine the inhibitory effect of AKT3, PI3KCA, and VEGFR2 small interfering RNAs (siRNAs) on GBM cell invasiveness. T98G cells were transfected with AKT3, PI3KCA, and/or VEGFR2 siRNAs. VEGFR2 protein-positive cells were identified by flow cytometry using specific monoclonal anti-VEGFR2 antibodies. Alterations in messenger RNA (mRNA) expression of VEGF, VEGFR2, matrix metalloproteinases (MMPs) (MMP-2, MMP-9, MMP-13, MMP-14), tissue inhibitors of metalloproteinases (TIMPs) (TIMP-1, TIMP-3), c-Fos, c-Jun, hypoxia-inducible factor-1α (HIF-1α), ObRa, and cathepsin D genes were analyzed by qRT-PCR. Cells treated with specific siRNA were also analyzed for invasion using the Matrigel invasion assay. We have found significantly lower mRNA levels of MMPs, cathepsin D, VEGF, VEGFR2, HIF-1α, and c-Fos/c-Jun ratio, as well as significantly higher mRNA level of TIMPs in AKT3 and PI3KCA siRNA transfected cells compared to untransfected cells, while significantly lower mRNA levels of MMPs (MMP-2, MMP-9, MMP-14) and TIMP-1, as well as significantly higher mRNA level of TIMP-3, were shown only in cells transfected with VEGFR2 siRNA. The positive correlation between MMP-13 and ObRa mRNA copy number has been found. Summarizing, transfection of T98G cells with AKT3, PI3KCA, or VEGFR2 siRNAs leads to a significant reduction in cell invasiveness. The siRNA-induced AKT3, PI3KCA, and VEGFR2 mRNA knockdown may offer a novel therapeutic strategy to reduce the invasiveness of GBM cells.
Assuntos
Neoplasias Encefálicas/enzimologia , Glioblastoma/metabolismo , Proteínas Nucleares/genética , Proteínas Proto-Oncogênicas c-akt/genética , Fatores de Transcrição/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/genética , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Movimento Celular , Técnicas de Silenciamento de Genes , Glioblastoma/patologia , Humanos , Metaloproteinases da Matriz Secretadas/genética , Metaloproteinases da Matriz Secretadas/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/genética , Inibidor Tecidual de Metaloproteinase-3/genética , Inibidor Tecidual de Metaloproteinase-3/metabolismo , Fatores de Transcrição/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Dermatan sulfate (DS) is a glycosaminoglycan characterized by having a variable structure and wide distribution in animal tissues. We previously demonstrated that some structural variants of DS were able to rapidly induce moderate necroptosis in luminal breast cancer cells when used at a high concentration. We have now investigated the mechanisms underlying the DS-mediated activation of the necroptotic executor MLKL using immunofluorescence, Western blotting and pharmacological inhibition. The two main processes, by which DS influences the phosphorylation of MLKL, are the activation of NFκB, which demonstrates a suppressive impact, and the induction of oxidative stress, which has a stimulatory effect. Moreover, the triggering of the redox imbalance by DS occurs via the modulatory influence of this glycosaminoglycan on the rearrangement of the actin cytoskeleton, requiring alterations in the activity of small Rho GTP-ase Rac1. All of these processes that were elicited by DS in luminal breast cancer cells showed a dependence on the structure of this glycan and the type of cancer cells. Furthermore, our results suggest that a major mechanism that is involved in the stimulation of necroptosis in luminal breast cancer cells by high doses of DS is mediated via the effect of this glycan on the activity of adhesion molecules.
Assuntos
Neoplasias da Mama , Dermatan Sulfato , NF-kappa B , Necroptose , Estresse Oxidativo , Proteínas Quinases , Humanos , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Neoplasias da Mama/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , NF-kappa B/metabolismo , Necroptose/efeitos dos fármacos , Feminino , Dermatan Sulfato/metabolismo , Dermatan Sulfato/farmacologia , Proteínas Quinases/metabolismo , Linhagem Celular Tumoral , Transdução de Sinais/efeitos dos fármacos , Proteínas rac1 de Ligação ao GTP/metabolismo , Fosforilação/efeitos dos fármacos , Células MCF-7 , Proteínas rac de Ligação ao GTP/metabolismoRESUMO
The brain-derived neurotrophic factor (BDNF) is one of the candidate genes for schizophrenia. There is evidence that val66met polymorphism may be involved in the pathophysiology of schizophrenia. The authors genotyped val66met (rs6265) polymorphism of the BDNF gene in 208 inpatients with paranoid schizophrenia and 254 control subjects in a Polish population. There was no association between val66met polymorphism and development of paranoid schizophrenia in either men or women. However, an association was found between this polymorphism and age at onset and psychopathology of paranoid schizophrenia. Men with the val/met genotype had an earlier age at onset, and the val/val genotype predisposed to more severe symptoms, particularly on the General Psychopathology Scale of the Positive and Negative Symptoms Scale (PANSS-G). The analysis of PANSS single items has shown that patients with the val/met genotype had higher scores on a hallucinatory behavior item than those with other genotypes.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/genética , Predisposição Genética para Doença , Metionina/genética , Polimorfismo de Nucleotídeo Único/genética , Esquizofrenia Paranoide/genética , Valina/genética , Adulto , Idade de Início , Análise de Variância , Análise Mutacional de DNA , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Polônia/epidemiologia , Escalas de Graduação Psiquiátrica , Fatores SexuaisRESUMO
Numerous reports have brought attention to the potential role of cytokines in schizophrenia. The aim of the study was to determine whether polymorphisms of IL-2, IL-6, and TNFα genes are risk factors for development of paranoid schizophrenia in a Polish population. Promoter polymorphisms of IL-6 (rs1800795), TNFα (rs1800629), and IL-2 (rs2069762) genes in patients (N=115) and controls (N=135) were genotyped by PCR-RFLP and AS-PCR methods, respectively. Genotype TT and allele T for IL-2 polymorphism, and genotype AA and allele A for TNFα polymorphism were found to be significantly associated with paranoid schizophrenia. Similarly, haplotypes CTA and GTA increased the risk (4.4 times and 5.9 times, respectively) of schizophrenia. To reveal associations between Positive and Negative Symptom Scale subscales and age at onset of schizophrenia, the authors used a novel method called Grade Correspondence Analysis. This analysis revealed that patients with early age at onset have higher scores on the Negative and General subscales of PANSS, and, in that group of patients, haplotype CTA was the most represented. As far as is known, this analysis was used for the first time with reference to genetic data.
Assuntos
Predisposição Genética para Doença/genética , Interleucina-2/genética , Interleucina-6/genética , Polimorfismo Genético/genética , Esquizofrenia Paranoide/genética , Fator de Necrose Tumoral alfa/genética , Adulto , Análise de Variância , Feminino , Frequência do Gene , Estudos de Associação Genética , Genótipo , Humanos , Estimativa de Kaplan-Meier , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Polônia , Escalas de Graduação Psiquiátrica , Esquizofrenia Paranoide/epidemiologiaRESUMO
BACKGROUND: Glioblastoma multiforme (GBM) is a heterogeneous and highly vascularized brain tumor that avoids apoptosis due to P-glycoprotein (P-gp) mediated multidrug resistance. Therefore, the development of new therapeutic strategies that induce apoptosis and inhibit proliferation is urgently warranted. OBJECTIVES: We examined the efficacy of the combination of baicalin (BAI) and knockdown of miR-148a gene in human glioblastoma T98G and U87MG cell lines. METHODS: T98G and U87MG cells were transfected with miR148a siRNA. The influence of miR- 148a siRNA in combination with BAI on T98G and U87MG cell viability, proliferation, apoptosis, and autophagy was evaluated as well. Alterations in the mRNA expression of autophagy-related genes were analyzed using RT-qPCR. RESULTS: The transfection of T98G and U87MG cells with miR148a specific siRNA and exposition on baicalin led to a significant reduction in cell viability and proliferation, the accumulation of sub G1-phase cells and a reduced population of cells in the S and G2/M phases (only in U87MG cell line), increased population of cells in the S phase in T98G cell line and apoptosis or necrosis induction and induction of autophagy for both cell lines. CONCLUSION: The siRNA-induced miR-148a mRNA knockdown in combination with baicalin may offer a novel therapeutic strategy to more effectively control the growth of human GBM cells. Thus, knockdown of this gene in combination with baicalin inhibits proliferation (cell cycle arrest in the S phase in T98G but not in U87MG cells), induces apoptosis, and regulates autophagy in T98G and U87MG cells. However, further studies are urgently needed to confirm a positive phenomenon for the treatment of GBM.
Assuntos
Neoplasias Encefálicas , Glioblastoma , MicroRNAs , Humanos , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Sobrevivência Celular , Linhagem Celular Tumoral , Apoptose , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Autofagia , Proliferação de Células , MicroRNAs/genética , MicroRNAs/uso terapêutico , RNA Interferente PequenoRESUMO
Changes in immunological system are one of dysfunctions reported in schizophrenia. Some changes based on an imbalance between Th1 and Th2 cytokines results from cytokine gene polymorphisms. Interleukin-4 gene (IL4) is considered as a potential candidate gene in schizophrenia association studies. The aim of the current case-control study was to examine whether the -590C/T (rs2243250) and -33C/T (rs2070874) IL4 gene polymorphisms are implicated in paranoid schizophrenia development in the Polish population. Genotyping of polymorphisms was performed by using PCR-RFLP technique. The genotypes and alleles distribution of both SNPs were analysed in patients (n = 182) and healthy individuals constituted the control group (n = 215). The connection between some clinical variables and studied polymorphisms has been examined as well. We did not revealed any association between the -590C/T and -33C/T polymorphisms and paranoid schizophrenia. In case of both SNPs the homozygous TT genotype was extremely rare. Both polymorphic sites of the IL4 gene were found to be in a very strong linkage disequilibrium. However we did not identify a haplotype predispose to paranoid schizophrenia. No associations were also observed between the clinical course and psychopathology of the disease and the genotypes of both analysed polymorphisms. Our results suggest that the polymorphisms -590C/T in IL4 gene promoter region and -33C/T in the 5'-UTR are not involved in the pathophysiology of paranoid schizophrenia in Polish residents.
Assuntos
Predisposição Genética para Doença , Interleucina-4/genética , Polimorfismo de Nucleotídeo Único , Esquizofrenia Paranoide/genética , População Branca/genética , Adulto , Idoso , Alelos , Análise de Variância , Estudos de Casos e Controles , Feminino , Genótipo , Humanos , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Polônia , Regiões Promotoras Genéticas , Adulto JovemRESUMO
Neuropeptide S (NPS), which is a peptide that is involved in the regulation of the stress response, seems to be relevant to the mechanism of action of antidepressants that have anxiolytic properties. However, to date, there have been no reports regarding the effect of long-term treatment with escitalopram or venlafaxine on the NPS system under stress conditions. This study aimed to investigate the effects of the above-mentioned antidepressants on the NPS system in adult male Wistar rats that were exposed to neonatal maternal separation (MS). Animals were exposed to MS for 360 min. on postnatal days (PNDs) 2-15. MS causes long-lasting behavioral, endocrine and neurochemical consequences that mimic anxiety- and depression-related features. MS and non-stressed rats were given escitalopram or venlafaxine (10mg/kg) IP from PND 69 to 89. The NPS system was analyzed in the brainstem, hypothalamus, amygdala and anterior olfactory nucleus using quantitative RT-PCR and immunohistochemical methods. The NPS system was vulnerable to MS in the brainstem and amygdala. In the brainstem, escitalopram down-regulated NPS and NPS mRNA in the MS rats and induced a tendency to reduce the number of NPS-positive cells in the peri-locus coeruleus. In the MS rats, venlafaxine insignificantly decreased the NPSR mRNA levels in the amygdala and a number of NPSR cells in the basolateral amygdala, and increased the NPS mRNA levels in the hypothalamus. Our data show that the studied antidepressants affect the NPS system differently and preliminarily suggest that the NPS system might partially mediate the pharmacological effects that are induced by these drugs.
RESUMO
Dermatan sulfate (DS) is widespread in the extracellular matrix (ECM) of animal tissues. This glycosaminoglycan is characterized by a variable structure, which is reflected in the heterogeneity of its sulfation pattern. The sulfate groups are responsible for the binding properties of DS, which determine an interaction profile of this glycan. However, the detailed role of DS in biological processes such as the neoplasm is still poorly understood. The aim of the study was to assess the effects of the structural variants of DS on breast cancer cells. We found that DS isoforms from normal and fibrotic fascia as well as from intestinal mucosa were able to quickly induce oxidative stress in the cytoplasm and affect the mitochondrial function in luminal breast cancer cells. Moreover, the variants caused the necroptosis of the cells most likely via the first of these mechanisms. This death was responsible for a reduction in the viability and number of breast cancer cells. However, the dynamics and intensity of all of the DS variants-triggered effects were strongly dependent on the cell type and the structure of these molecules. The most pronounced activity was demonstrated by those variants that shared structural features with the DS from the tumor niche.
Assuntos
Neoplasias da Mama/patologia , Dermatan Sulfato/farmacologia , Necroptose , Animais , Contagem de Células , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Feminino , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Necroptose/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacosRESUMO
Schizophrenia (SCZ) is a severe psychiatric disorder that has a significant genetic component. HSPB1 (HSP27) is known for its neuroprotective functions under stress conditions and appears to play an important role during the development of the central nervous system, which is in agreement with the neurodevelopmental hypothesis of SCZ. The aim of the present case-control study was to investigate whether HSPB1 variants contribute to the risk and clinical features (age of onset, symptoms, and suicidal behavior) of SCZ in a Polish population. To the best of our knowledge, this is the first study that investigated the association between the HSPB1 polymorphisms and SCZ. Three SNPs of HSPB1 (rs2868370, rs2868371, and rs7459185) were genotyped in a total of 1082 (403 patients and 679 controls) unrelated subjects using TaqMan assays. The results showed that the genotypes, alleles, and haplotypes of the three SNPs were not significantly different between the schizophrenic patients and healthy controls either in the overall analysis or in the gender-stratified analysis (all p > 0.05). However, we did find a significant effect of the rs2868371 genotype on the age of onset, negative symptoms, and disorganized symptoms in the five-factor model of PANSS (all p < 0.01). Post hoc comparisons showed that carriers of the rs2868371 G/G genotype had significantly higher negative and disorganized factor scores than those with the C/G and C/C genotypes, respectively. Further investigations with other larger independent samples are required to confirm our findings and to better explore the effect of the HSPB1 polymorphisms on the risk and symptomatology of SCZ.