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1.
BMC Microbiol ; 23(1): 257, 2023 09 13.
Artigo em Inglês | MEDLINE | ID: mdl-37704938

RESUMO

BACKGROUND: Enzybiotics are promising alternatives to conventional antibiotics for drug-resistant infections. Exolysins, as a class of enzybiotics, show antibacterial effects against methicillin-resistant Staphylococcus aureus (MRSA). This study evaluated a novel exolysin containing an SH3b domain for its antibacterial activity against MRSA. METHODS: This study designed a chimeric exolysin by fusing the Cell-binding domain (SH3b) from Lysostaphin with the lytic domain (LYZ2) from the gp61 enzyme. Subsequently, LYZ2-SH3b was cloned and expressed in Escherichia coli (E. coli). Finally, the antibacterial effects of LYZ2-SH3b compared with LYZ2 and vancomycin against reference and clinical isolates of MRSA were measured using the disc diffusion method, the minimal inhibitory concentration (MIC), and the minimal bactericidal concentration (MBC) assays. RESULTS: Analysis of bioinformatics showed that LYZ2-SH3b was stable, soluble, and non-allergenic. Protein purification was performed with a 0.8 mg/ml yield for LYZ2-SH3b. The plate lysis assay results indicated that, at the same concentrations, LYZ2-SH3b has a more inhibitory effect than LYZ2. The MICs of LYZ2 were 4 µg/mL (ATCC 43,300) and 8 µg/mL (clinical isolate ST239), whereas, for LYZ2-SH3b, they were 2 µg/mL (ATCC 43,300) and 4 µg/mL (clinical isolate ST239). This suggests a higher efficiency of LYZ2-SH3b compared to LYZ2. Furthermore, the MBCs of LYZ2 were 4 µg/mL (ATCC 43,300) and 8 µg/mL (clinical isolate ST239), whereas, for LYZ2-SH3b, they were 2 µg/mL (ATCC 43,300) and 4 µg/mL (clinical isolate ST239), thus confirming the superior lytic activity of LYZ2-SH3b over LYZ2. CONCLUSIONS: The study suggests that phage endolysins, such as LYZ2-SH3b, may represent a promising new approach to treating MRSA infections, particularly in cases where antibiotic resistance is a concern. But further studies are needed.


Assuntos
Bacteriófagos , Staphylococcus aureus Resistente à Meticilina , Staphylococcus aureus Resistente à Meticilina/genética , Escherichia coli/genética , Antibacterianos/farmacologia , Vancomicina
2.
J Cell Mol Med ; 26(20): 5235-5245, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36098216

RESUMO

Cell therapy and tissue engineering as promising candidates for the liver transplantation dilemma are of special interest. Induced pluripotent stem cells (iPSCs) are one of the best sources in this field, but their differentiation methods to hepatocytes have remained challenging. We transduced human iPSCs (hiPSCs) with miR-122 and off-let-7f (hiPSCsmiR-122 + off-let-7f ) to evaluate how they can differentiate hiPSCs to hepatocyte-like cells (HLCs) without any extrinsic growth factor. Additionally, we studied the effect of Poly ɛ-caprolactone-gelatin-hyaluronic acid (PCL-Gel-HA) nanofibrous scaffold as an extracellular matrix (ECM) simulator on differentiation improvement. Definitive endoderm markers (FOXA2 and SOX17), as well as hepatic markers (AFP, Albumin, CK18, HNF4α) expression, were significantly higher in hiPSCsmiR-122 + off-let-7f derived HLCs (hiPSCs-HLCs) compared to the control group (miR-scramble transduced hiPSCs: hiPSCsscramble ). hiPSCs-HLCs indicated hepatocyte morphological characteristics and positive immunostaining for AFP, Albumin and HNF4α. Albumin and urea secretion were significantly higher in hiPSCs-HLCs than hiPSCsscramble . Comparing these markers in the PCL-Gel-HA group with the tissue culture plate (TCP) group revealed that PCL-Gel-HA could improve differentiation towards HLCs significantly. Regarding our results, these microRNAs can be used to differentiate hiPSCs to the functional hepatocytes for disease modelling, drug screening and cell-based therapy in future studies.


Assuntos
Células-Tronco Pluripotentes Induzidas , MicroRNAs , Nanofibras , Albuminas/metabolismo , Caproatos , Regulação para Baixo , Gelatina , Humanos , Ácido Hialurônico/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Lactonas , MicroRNAs/metabolismo , Regulação para Cima , Ureia/metabolismo , alfa-Fetoproteínas/genética , alfa-Fetoproteínas/metabolismo
3.
J Cell Mol Med ; 26(8): 2392-2403, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35224849

RESUMO

This study aimed to investigate if Telmisartan as a novel N-cadherin antagonist, can overcome cell migration of cancer cells. We investigated the mechanism and influence of Docetaxel and Telmisartan (as an analogous to ADH-1, which is a well-known N-cadherin antagonist) on cancer cells. The effect of ADH-1 and Telmisartan on cell attachment in PC3, DU145, MDA-MB-468 cell lines using recombinant human N-cadherin was studied. Cell viability assay was performed to examine the anti-proliferative effects of Telmisartan, ADH-1 and Docetaxel. Migration was examined via wound healing assay, and apoptosis was determined by flow cytometry. The expression of AKT-1 as a downstream gene of N-cadherin signalling pathway was assayed by real-time PCR. Treatment of PC3, MDA-MB-468 and DU145 cells with Telmisartan (0.1 µM) and ADH-1 (40 µM) resulted in 50%, 58% and approximately 20% reduction in cell attachment to N-cadherin coated plate respectively. It shows reduction of cell attachment in PC3 and MDA-MB-468 cell lines appeared to be more sensitive than that of DU145 cells to the Telmisartan and ADH-1 treatments. Telmisartan (0.1 µM) and Docetaxel (0.01 nM) significantly reduced cell migration in PC3 and MDA-MB-468 cell lines compared with the control group. Using Real-time PCR, we found that Telmisartan, Docetaxel and ADH-1 had significant influence on the AKT-1 mRNA level. The results of the current study for the first time suggest that, Telmisartan, exerts anti-proliferation and anti-migration effects by targeting antagonistically N-cadherin. Also, these data suggest that Telmisartan as a less expensive alternative to ADH-1 could potentiate Docetaxel anticancer effects.


Assuntos
Caderinas , Oligopeptídeos , Peptídeos Cíclicos , Proteínas Proto-Oncogênicas c-akt , Telmisartan , Antígenos CD/metabolismo , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Docetaxel/farmacologia , Humanos , Terapia de Alvo Molecular , Oligopeptídeos/farmacologia , Células PC-3 , Peptídeos Cíclicos/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Telmisartan/farmacologia
4.
Biotechnol Appl Biochem ; 69(6): 2592-2598, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34965611

RESUMO

Coronavirus 2019 (COVID-19) is a global concern for public health. Thus, early and accurate diagnosis is a critical step in management of this infectious disease. Currently, RT-PCR is routine diagnosis test for COVID-19, but it has some limitations and false negative results. enzyme-linked immunosorbent assay (ELISA) against SARS-CoV-2 antigens seems to be an appropriate approach for serodiagnosis of COVID-19. In the current study, an ELISA system, using a recombinant nucleocapsid (N) protein, was developed for the detection of IgM and IgG antibodies to SARS-CoV-2. The related protein was expressed, purified, and used in an ELISA system. Sera samples (67) for COVID-19 patients, as well as sera samples from healthy volunteers (112), along with sera samples from non-COVID-19 patients were examined by the ELISA system. The expression and purity of the recombinant N protein were approved by SDS-PAGE and Western blotting. The sensitivity of ELISA system was 91.04 and 92.53% for the detection of IgG and IgM antibodies, respectively. Moreover, the specificity of the developed ELISA system for IgG and IgM were 98.21 and 97.32%, respectively. Our developed ELISA system showed satisfactory sensitivity and specificity for the detection of antiSARS-CoV-2 IgM and IgG antibodies and could be used as a complementary approach for proper diagnosis of COVID-19.


Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Proteínas do Nucleocapsídeo , Imunoglobulina G , COVID-19/diagnóstico , Nucleocapsídeo , Ensaio de Imunoadsorção Enzimática , Sensibilidade e Especificidade , Proteínas Recombinantes , Imunoglobulina M
5.
J Cell Physiol ; 236(5): 3495-3509, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33030247

RESUMO

Osteoporosis is the most prevalent metabolic bone disease and one of the most important postmenopausal consequences. The aim of this study was to investigate the effects of quercetin (Q) and vitamin E (vitE) on ovariectomy-induced osteoporosis. Animals were ovariectomized and treated with Q (15 mg/kg/day), vitE (60 mg/kg/day), estradiol (10 µg/kg/day), and Q (7.5 mg/kg/day) + vitE (30 mg/kg/day) for 10 weeks by gavage, and osteoporosis markers and messenger RNA (mRNA) expression of autophagy and apoptosis-related genes were analyzed in serum and tibia of rats. Data indicated that ovariectomy resulted in development of osteoporosis as demonstrated by reduction in serum calcium, bone weight, bone volume, trabeculae volume, and the total number of osteocytes and osteoblasts, and increase in the total number of osteoclasts and serum osteocalcin. Total mRNA expressions of LC3, beclin1, and caspase 3 were also increased and bcl2 expression was decreased in the tibia. By reversing these changes, treatment with Q and vitE markedly improved osteoporosis. In conclusion, Q, and to a lesser extent, vitE, prevented osteoporosis by regulating the total number of bone cells, maybe through regulating autophagy and apoptosis.


Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Osteócitos/efeitos dos fármacos , Ovariectomia/efeitos adversos , Quercetina/farmacologia , Animais , Densidade Óssea/efeitos dos fármacos , Feminino , Humanos , Osteoblastos/metabolismo , Osteoclastos/metabolismo , Osteócitos/metabolismo , Osteoporose/genética , Ratos Transgênicos
6.
Nutr Cancer ; 73(10): 2003-2013, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32924610

RESUMO

To assess the effect of sequential treatment with Vitamin C (VC) and Quercetin (Q) on Nrf2-related oxidative stress in PC3 and DU145 cells, viability was measured by MTT assay. Intracellular ROS levels were determined, using 2'-7'-dichlorodihydrofluorescein diacetate fluorescent as a probe. Nrf2 gene expression was investigated by quantitative reverse transcription polymerase chain reaction, and Nrf2 protein levels were defined by western blot analysis. The activity of glutathione peroxidase (GPx), glutathione reductase (GR), nicotinamide adenine dinucleotide phosphate dehydrogenase quinone 1 (NQO1) and hemeoxygenase 1 (HO-1) enzymes were measured. The IC50 values for VC + Q were 263.03-372.1 µM and 144.2-194.1 µM respectively and 200 µM VC + 50 µM Q (dose no.1) and 100 µM VC + 75 µM Q (dose no.2) were selected. Sequential treatment of PC3 cells led to a significant reduction of Nrf2 mRNA expression and protein levels in addition to a significant reduction of GPx, GR and NQO1 enzymatic activity. Although the data was slightly different for DU145 cells after the treatments, in terms of Nrf2 gene expression, we obtained the same results. Our study revealed the significant effects of sequential treatment with VC + Q on Nrf2 suppression in prostate cancer cells.


Assuntos
Fator 2 Relacionado a NF-E2 , Neoplasias da Próstata , Ácido Ascórbico/farmacologia , Quimioprevenção , Humanos , Masculino , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo , Neoplasias da Próstata/tratamento farmacológico , Quercetina/farmacologia , Espécies Reativas de Oxigênio
7.
Mol Biol Rep ; 48(10): 6749-6756, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34424445

RESUMO

BACKGROUND: Metastasis is a major cause of death in Colorectal cancer (CRC) patients, and the Epithelial-mesenchymal transition (EMT) has been known to be a crucial event in cancer metastasis. Downregulated expression of AT-rich interaction domain-containing protein 1A (ARID1A), a bona fide tumor suppressor gene, plays an important role in promoting EMT and CRC metastasis, but the underlying molecular mechanisms remain poorly understood. Here, we evaluated the impact of ARID1A knockdown and overexpression on the expression of EMT­related genes, E-cadherin and ß-catenin, in human CRC cells. METHODS AND RESULTS: The expression levels of ARID1A, E-cadherin and ß-catenin in CRC cell lines were detected via real-time quantitative PCR (qPCR) and western blot. ARID1A overexpression and shRNA-mediated knockdown were performed to indicate the effect of ARID1A expression on E-cadherin and ß-catenin expression in CRC cell lines. The effect of ARID1A knockdown on the migration ability of HCT116 cells was assessed using wound-healing assay. We found that the mRNA and protein expression of adhesive protein E-cadherin was remarkably downregulated in response to shRNA-mediated ARID1A knockdown in HCT116 and HT29 cells. Conversely, overexpression of ARID1A in SW48 cells significantly increased E-cadherin expression. In addition, ARID1A silencing promoted the migration of HCT116 cells. ARID1A knockdown and overexpression did not alter the level of ß-catenin expression. CONCLUSIONS: Our study demonstrates that E-cadherin levels were closely correlated with ARID1A expression. Thus, ARID1A downregulation may promote CRC metastasis through decreasing EMT­related protein E-cadherin and promoting epithelial cell movement. ARID1A could represent a promising candidate therapeutic target for CRC.


Assuntos
Antígenos CD/genética , Caderinas/genética , Neoplasias Colorretais/genética , Proteínas de Ligação a DNA/metabolismo , Regulação Neoplásica da Expressão Gênica , Terapia de Alvo Molecular , Fatores de Transcrição/metabolismo , Antígenos CD/metabolismo , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Técnicas de Silenciamento de Genes , Inativação Gênica , Células HEK293 , Humanos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , beta Catenina/metabolismo
8.
J Immunoassay Immunochem ; 41(6): 1010-1020, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-32795213

RESUMO

BACKGROUND: Cystic echinococcosis (CE) is a widespread parasitic disease caused by the larval stage of Echinococcus granulosus. Since current methods for the diagnosis of CE are not efficient enough, rapid, and reliable tests are required for the acceleration of CE diagnosis. The present study aimed to produce recombinant B8/1 and B8/2 antigens of E. granulosus and evaluate their sensitivities and specificities separately and simultaneously for the diagnosis of CE. METHODS: The recombinant B8/1 and B8/2 antigens were produced and used in an ELISA system for the diagnosis of CE. The sera specimens including 30 sera from pathologically confirmed CE patients, 30 from other non-CE patients, and 30 from healthy controls, were evaluated by the ELISA, using AgB8/1 and AgB8/2. RESULTS: The results showed a sensitivity of 93.33%, 90%, and 96.7% for AgB8/1, AgB8/2, and their combination, respectively. The specificities were 91.7%, 93.33%, and 93.33% for AgB8/1, AgB8/2, and their combination, respectively. CONCLUSION: Simultaneous usage of AgB8/1 and AgB8/2 increased the test sensitivity for the diagnosis of CE. Furthermore, the specificity of AgB8/1 and AgB8/2 combination was more than AgB8/1 and equal to AgB8/2 alone. The findings revealed that the simultaneous usage of AgB8/1 and AgB8/2 could be a suitable approach for the diagnosis of CE.


Assuntos
Antígenos de Helmintos/sangue , Equinococose/diagnóstico , Echinococcus granulosus/química , Ensaio de Imunoadsorção Enzimática , Animais , Antígenos de Helmintos/genética , Antígenos de Helmintos/imunologia , Equinococose/sangue , Equinococose/imunologia , Echinococcus granulosus/imunologia , Humanos , Proteínas Recombinantes/sangue , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia
9.
Iran J Med Sci ; 44(4): 315-324, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31439975

RESUMO

BACKGROUND: Endometriosis is a common gynecological disease in which oxidative stress is a potential factor. Caffeine and caffeic acid are present in various foods and beverages with anti-oxidant, anti-inflammatory, and anti-carcinogenic properties. In this study, we aimed to investigate the ameliorative effects of caffeine, caffeic acid, and caffeine+caffeic acid treatments on oxidative stress in ectopic endometrial cells taken from patients and eutopic ones from women without endometriosis. METHODS: In this experimental study, eutopic and ectopic endometrial cells were obtained from biopsies of women free of disease (n=10) and patients with endometriosis (n=10) who referred to Shiraz reference hospitals (2017-2018). Both eutopic and ectopic endometrial cells were divided into four groups: Treated with caffeine, with caffeic acid, with caffeine+caffeic acid, and the control. Also, antioxidant enzyme activities and the levels of glutathione (GSH) and malondialdehyde (MDA) were determined in each group. The data were analyzed using independent sample t test and one-way ANOVA followed by Tukey post-hoc test. RESULTS: Caffeic acid, but not caffeine treatment demonstrated a decrease in MDA level (P<0.001) as well as an increase in GSH level (P<0.001) and antioxidant enzyme activities in ectopic endometrial cells. Also, the treatment of the cells with caffeine+caffeic acid caused similar effects as those ectopic cells treated with caffeic acid. CONCLUSION: According to the findings of the present study, caffeic acid reduced oxidative stress which may alleviate the complications associated with endometriosis. However, more investigations are needed for evaluating the efficiency and safety of caffeic acid.

10.
Nutr Cancer ; 70(5): 770-775, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-29781726

RESUMO

Some types of cancers show a strong relationship with diabetes and play a central role in mortality in the patient population suffering from diabetes mellitus. In this study, HepG2 cells have been used to investigate the toxic effects of hyperglycemia and/or quercetin (Q) on mammalian target of rapamycin (m-TOR) and nuclear factor erythroid 2-related factor 2 (Nrf-2) expression as central molecules involved in cancer. HepG2 cells were cultured with different concentrations of glucose (5.5, 30, and 50 mM) and/or Q (25 µM) for 48 and 72 h. Effects of glucose and/or Q on m-TOR and Nrf-2 expression were assayed by quantitative real-time PCR (qRT-PCR). qRT-PCR results revealed that 30 and 50 mM of glucose increased m-TOR expression at 48 h, although after 72 h, only 30 mM had an increasing effect. At 50 mM, glucose-induced Nrf-2 gene expression after both 48 and 72 h. The results also showed that 25 µM of Q reduced m-TOR and Nrf-2 expression at both 30 and 50 mM after 48 and 72 h incubation. Q has potential effects on reducing oxidative stress caused by hyperglycemia and during diabetes may be able to modulate some carcinogenic signaling pathways.


Assuntos
Glucose/farmacologia , Fator 2 Relacionado a NF-E2/genética , Quercetina/farmacologia , Serina-Treonina Quinases TOR/genética , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Glucose/administração & dosagem , Células Hep G2 , Humanos , Hiperglicemia/metabolismo , Quercetina/administração & dosagem
11.
Mol Biol Rep ; 45(3): 245-252, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29411210

RESUMO

Human Wharton's jelly mesenchymal stem cells (hWJSCs) are multipotent stem cells that could be aggregated into 3D spherules. ITGA4 and ITGA5 genes encode α4 and α5 subunits of integrins, respectively. In this study, we analyzed expression levels of ITGA4 and ITGA5 gene mRNAs in undifferentiated and 3D spherules forming hWJSCs in order to determine their expression pattern for possible future treatment of cancer cells in a co-culture fashion. For the purpose of obtaining hWJSCs, umbilical cords were collected from patients with caesarian section at full term delivery. The cells were then characterized according to cell surface markers using flow cytometry. Furthermore pluripotency of the obtained cells was verified. Subsequently the cells were aggregated in 3D spherules using hanging drop cultures. Expression levels of ITGA4 and ITGA5 gene mRNAs were determined by RT-PCR and Real time PCR, both in the initial undifferentiated cells and those aggregated in the spherules. The obtained hWJSCs demonstrated pluripotency, differentiating to adipogenic and osteogenic cells. They also expressed mesenchymal stem cell surface markers. Following the aggregation of these cells and formation of 3D spherules, mRNA expression levels of both genes were significantly reduced (P < 0.05) compared with the initial undifferentiated state. The results of this study demonstrated that aggregation of hWJSCs into spherules alters their expression of ITGA4 and ITGA5. The implications of such an alteration would require further research.


Assuntos
Integrina alfa4/genética , Integrina alfa5/genética , Células-Tronco Mesenquimais/fisiologia , Adipócitos/citologia , Diferenciação Celular/fisiologia , Células Cultivadas , Técnicas de Cocultura , Meios de Cultivo Condicionados , Regulação para Baixo , Citometria de Fluxo , Humanos , Integrina alfa4/biossíntese , Integrina alfa5/biossíntese , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Osteogênese/fisiologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Cordão Umbilical/citologia
12.
Iran J Med Sci ; 43(5): 523-532, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30214105

RESUMO

BACKGROUND: Differentiating oligodendrocyte precursor cells (OPCs) into oligodendrocytes could be improved by inhibiting signaling pathways such as Wnt and Notch. 9-cis-retinoic acid (9-cis-RA) and 1,25-dihydroxyvitamin D3 (1,25[OH]2D3) can ameliorate oligodendrogenesis. We investigated whether they could increase oligodendrogenesis by inhibiting the Wnt and Notch signaling pathways. METHODS: Cortical neural stem cells were isolated from 14-day-old rat embryos and cultured using the neurosphere assay. The cells were treated in 4 different conditions for 1 week: the negative control group received only the basic fibroblast growth factor, the positive control group received only T3 without growth factors, the RA group was treated with 9-cis-RA, and the Vit D3 group was treated with 1,25(OH)2D3. The effects of 9-cis-RA and 1,25(OH)2D3 on the level of the myelin basic protein (MBP) and the gene expression of the SOX10, MBP gene, HES5, and LRP6 were studied using flow cytometry and real-time PCR. The data were analyzed using one-way ANOVA with GraphPad Prism. A P value less than 0.05 was considered significant. RESULTS: The mRNA expressions of the SOX10, MBP, and MBP gene were significantly increased in the treated groups compared with the negative control group; the increase was similar in the 9-cis-RA group and the positive control group. Furthermore, 9-cis-RA significantly decreased the expression of the HES5 gene, a Notch signaling pathway transcription factor, and 1,25(OH)2D3 significantly reduced the expression of the LRP6 gene, a Wnt signaling pathway co-receptor. CONCLUSION: It seems that 9-cis-RA and 1,25(OH)2D3 are good candidates to improve the differentiation of OPCs into oligodendrocytes.

13.
Nutr Cancer ; 69(6): 881-891, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28742385

RESUMO

OBJECTIVE: Prevention by antioxidant agents including vitamin C (VC) and quercetin (QU), which are nontoxic, cost effective, and physiologically bioavailable, is a promising approach in breast cancer handling. The aim of this work is to investigate the influence of VC+QU on cytotoxicity profile of doxorubicin (DOX) plus paclitaxel (PAC) in breast cancer cells. METHODS: The effect of each drug on its own or in combination was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Combination indexes were calculated using the Chou-Talalay method. Apoptosis and cell cycle analysis was investigated by flow cytometric assays. RESULTS: Combination treatment with VC+QU plus drugs diminished IC50 value 2.28-7.7 and 10.5-66.6 fold in comparison with the drugs and PAC treatment alone, respectively, in all breast cancer cells and induced apoptosis at the early stages more than the treatment with the drugs alone (P<0.01). A marked reduction in Go/G1 and S phases was reported after combination therapy in MDA-MB 231 and MDA-MB 468 cells. MCF-7 cells demonstrated lower fractions of cells in S phase with no significant changes in G2/M phase (P < 0.01). The same treatment produced a significant increase in S and G2/M phases in A549 cells (P < 0.001). CONCLUSION: Our results emphasized the importance of VC+QU in combination with the drugs to produce a synergistic antitumor effect in breast cancer cells.


Assuntos
Antineoplásicos/farmacologia , Ácido Ascórbico/farmacologia , Neoplasias da Mama/tratamento farmacológico , Quercetina/farmacologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Doxorrubicina/farmacologia , Feminino , Fase G2 , Humanos , Concentração Inibidora 50 , Células MCF-7 , Paclitaxel
15.
Iran J Med Sci ; 42(3): 275-283, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28533576

RESUMO

BACKGROUND: Discriminating latent tuberculosis infection (LTBI) from active TBI may be challenging. The objective of this study was to produce the recombinant L-alanine dehydrogenase (AlaDH) antigen and evaluate individuals with LTBI, those with active TBI, and uninfected individuals by enzyme-linked immunospot assay (ELISPOT) in order to distinguish LTBI from active TBI. METHODS: This exploratory study was performed in the Iranian city of Shiraz from 2014 to 2015. The study population (N=99) was divided into 3 groups: individuals with newly diagnosed active TBI (n=33), their household contacts (n=33), and controls (n=33). AlaDH was produced through PCR and cloning methods. The diagnostic characteristics of AlaDH vs. ESAT-6/CFP-10 were evaluated in responses to interferon-γ (IFN-γ) and interleukin-2 (IL-2) with ELISPOT. Differences between the groups were assessed with the Kruskal-Wallis and Mann-Whitney tests for nonparametric data analysis. The statistical analyses were performed with SPSS, version 16. RESULTS: IFN-γ responses to both ESAT-6/CFP-10 (P=0.81) and AlaDH (P=0.18) revealed that there were no significant differences between the individuals with LTBI and those with active TBI. The same results were determined for IL-2 responses to ESAT-6/CFP-10 between the 2 groups, while significantly higher IL-2 responses to AlaDH were observed in LTBI than in active TBI. According to the ROC curve analysis, a cutoff value of 275 SFC showed sensitivity of 75.8% and specificity of 78.8% for distinguishing LTBI from active TBI by IL-2 responses to AlaDH. CONCLUSION: The current study suggests that it may be possible to discriminate LTBI from active TBI by IL-2 responses to AlaDH.

16.
Toxicol Mech Methods ; 26(8): 595-600, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27552315

RESUMO

Cigarette smoke is a complex mixture of toxic chemicals, including nicotine, carbon monoxide, and several recognized carcinogens and mutagens. Nicotine has a direct disturbing influence on steroid hormones (estrogen and progesterone), which are essential components of the female reproductive system, but the effect of nicotine on the hormone receptors is not yet clear. The aim of this study was to elucidate the effect of nicotine on the expression of estrogen receptor (ER), progesterone receptor (PR), and vascular endothelial growth factor (VEGF) in endometrial stromal cells. Expression levels of PR, ER, and VEGF in human endometrial stromal primary cells treated with nicotine (0, 10-11, 10-8, and 10-6 µM) for 24 h were measured by quantitative real-time PCR. MTT assay demonstrated that nicotine decreased cell viability in a dose-dependent manner. Real-time PCR data showed that despite decrease in ER expression in the nicotine-treated groups compared with the control, nicotine exerted an increased inhibitory effect on PR expression compared to that on ER expression. VEGF mRNA expression in nicotine-treated endometrial stromal cells was increased. The results from this study provide novel evidence for inhibitory effects of nicotine on steroid hormones receptor expression in human primary endometrial cells. Also, our data suggest that nicotine might have angiogenesis effects on these cells.


Assuntos
Endométrio/efeitos dos fármacos , Receptor alfa de Estrogênio/genética , Expressão Gênica/efeitos dos fármacos , Nicotina/toxicidade , Receptores de Progesterona/genética , Fator A de Crescimento do Endotélio Vascular/genética , Adulto , Técnicas de Cultura de Células , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , Endométrio/citologia , Endométrio/metabolismo , Feminino , Humanos , Reação em Cadeia da Polimerase em Tempo Real , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Adulto Jovem
17.
Iran J Med Sci ; 41(4): 296-304, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27365551

RESUMO

BACKGROUND: Macrophage immune responses are affected by the secretory proteins of Mycobacterium tuberculosis (Mtb). This study aimed to examine the immune responses of macrophages to Mtb secretory antigens, namely ESAT-6, CFP-10, and ESAT-6/CFP-10. METHODS: THP-1 cells (a human monocytic cell line) were cultured and differentiated to macrophages by phorbol 12-myristate 13-acetate. The cytotoxicity of the recombinant Mtb proteins was assessed using the MTT assay. Two important immune responses of macrophages, namely NO and ROS production, were measured in response to the ESAT-6, CFP-10, and ESAT-6/CFP-10 antigens. The data were analyzed using one-way ANOVA with SPSS, version 16, and considered significant at P<0.05. RESULTS: The results showed that the ESAT-6, CFP-10, and ESAT-6/CFP-10 proteins markedly reduced macrophage immune response. The treatment of the THP-1-differentiated cells with ESAT-6, CFP-10, and ESAT-6/CFP-10 reduced NO and ROS production. The treated THP-1-differentiated cells exhibited less inducible NO synthase activity than did the untreated cells. No toxic effect on macrophage viability was observed for the applied proteins at the different concentrations. CONCLUSION: It seems that the decline in macrophage immune response is due to the suppression of NO and ROS production pathways without any effect on cell viability.

18.
Toxicol Mech Methods ; 25(2): 98-104, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25418342

RESUMO

BACKGROUND AND OBJECTIVE: Cyclosporine A (CsA) is an immunosuppressive agent administered to transplant patients. A well-known reported oral side effect of CsA consumption is gingival overgrowth (GO). Changes in the expression of integrins occurring in the gingiva following CsA treatment have been reported but these reports are mainly concerned with the connective tissue of the gingiva. In this study we targeted the alterations in the oral epithelium using KB cells, an oral epithelial cell line. METHODS: Cultured oral epithelial cells were treated with increasing concentrations of CsA (0.1, 1 and 10 µg/mL) and the molecular changes involving antioxidant enzymes [glutathione peroxidase (GPx) and glutathione reductase (GR)] and the level of reactive oxygen species (ROS) were measured. Quantitative real-time PCR was used to assess the expression of selected integrins (α2, α5 and ß1). RESULTS: At CsA concentration above 0.1 µg/mL GPx demonstrated an increase in activity while GR activity and the level of reduced glutathione were diminished (p < 0.05). α5 and ß1 integrin were downregulated at all treatment concentrations of CsA while α2 integrin presented this effect at concentrations above 1 µg/mL (p < 0.05). CONCLUSION: The results suggest a possible role for oxidative stress and the altered expression of integrins in the pathology of CsA-induced gingival overgrowth.


Assuntos
Ciclosporina/toxicidade , Células Epiteliais/efeitos dos fármacos , Crescimento Excessivo da Gengiva/induzido quimicamente , Imunossupressores/toxicidade , Integrinas/metabolismo , Mucosa Bucal/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Linhagem Celular , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Crescimento Excessivo da Gengiva/metabolismo , Crescimento Excessivo da Gengiva/patologia , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Humanos , Integrina alfa2/metabolismo , Integrina alfa5/metabolismo , Integrina beta1/metabolismo , Integrinas/genética , Mucosa Bucal/metabolismo , Mucosa Bucal/patologia , Espécies Reativas de Oxigênio/metabolismo
19.
Int J Vitam Nutr Res ; 84(1-2): 55-64, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25835236

RESUMO

This study was designed to assess oxidative damage and cell apoptosis in the uterus of rats with streptozotocin (STZ)-induced diabetes. The role of vitamin E (VE) and/or folic acid (FA) in the protection from such damage was also evaluated. The treatments were performed for 4 weeks on six groups of rats: 1) normal control 2) diabetic control 3) diabetic rats receiving olive oil as a vehicle (without VE) 4) diabetic rats treated with VE (200 mg/kg) in olive oil 5) diabetic rats treated with FA (25 mg/kg) and 6) diabetic rats treated with VE+FA (200 and 25 mg/kg, respectively). We measured the malondialdehyde level (MDA), glutathione content (GSH) and the activity of GSH peroxidase (GPx), GSH reductase (GR) and catalase. Changes in caspase-3 activity were quantified in uterine tissue to assess the rate of apoptosis. In the rat uterine tissues, MDA content and caspase-3 activity were significantly elevated, while GPx, GR and CAT activities and the GSH level were significantly decreased in the diabetic control compared with those in normal rats (p<0.05). The combination of the vitamins (VE+FA) restored uterine GSH content and enzymatic activities of GPx, GR and CAT and reduced the MDA level (p<0.05). A prominent reduction in apoptosis of uterine cells was detected in diabetic rats treated with two vitamins (p<0.05). Overall, VE alone, not FA, produced results similar to those of the VE+FA combination. Thus, in the uterine tissue of diabetic rats, diabetes complications (that are caused by oxidative damage and apoptosis induction) can be prevented by the systemic administration of VE and FA.


Assuntos
Apoptose/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Ácido Fólico/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Útero/metabolismo , Vitamina E/administração & dosagem , Animais , Caspase 3/análise , Catalase/análise , Complicações do Diabetes/prevenção & controle , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Quimioterapia Combinada , Feminino , Glutationa/análise , Glutationa Peroxidase/análise , Glutationa Redutase/análise , Malondialdeído/análise , Ratos , Ratos Sprague-Dawley , Útero/química , Útero/patologia
20.
Toxicol Mech Methods ; 24(6): 412-6, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24845846

RESUMO

OBJECTIVES: Propranolol, a beta-adrenergic blocker, is used in the treatment of a large number of cardiovascular diseases such as hypertension and arrhythmias. Propranolol, in combination with furosemide, is used to treat hypertensive disorders although their side effect profile is not very obvious. In present study, the effects of the drugs furosemide and propranolol were in corporately investigated both on glutathione homeostasis and their antioxidant effect on ACHN cells. METHODS: The cytoxicities and antioxidant effects of these two clinically important drugs on human kidney cell lines were evaluated using MTT following by the determination of glutathione reductase (GR) and glutathione peroxidase (GPx) activities and measuring the level of reduced glutathione (GSH). RESULTS: Propranolol induced a significant cytotoxic effect at 100 µM, while furosemide was cytotoxic at doses of 250 and 1000 µg/ml. A slight increase in GPx and GR activities and GSH level was observed with propranolol and furosemide treatment alone, while the two drugs together caused a significant increase in GPx and GR activities (35% and 42%, respectively) and GSH content (35%) in ACHN cell lysates (p < 0.05). CONCLUSIONS: Our results demonstrate that although high doses of furosemide and propranolol are cytotoxic, co-administration of low doses may improve the antioxidant defense in patients undergoing treatment with these two important drugs.


Assuntos
Furosemida/toxicidade , Glutationa/metabolismo , Homeostase/efeitos dos fármacos , Rim/citologia , Rim/efeitos dos fármacos , Propranolol/toxicidade , Antagonistas Adrenérgicos beta/administração & dosagem , Antagonistas Adrenérgicos beta/toxicidade , Linhagem Celular , Diuréticos/administração & dosagem , Diuréticos/toxicidade , Quimioterapia Combinada , Fibroblastos/efeitos dos fármacos , Furosemida/administração & dosagem , Furosemida/química , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Glutationa Peroxidase/genética , Glutationa Peroxidase/metabolismo , Glutationa Redutase/genética , Glutationa Redutase/metabolismo , Humanos , Estrutura Molecular , Propranolol/administração & dosagem , Propranolol/química
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