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1.
Immunopharmacol Immunotoxicol ; 46(2): 161-171, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38051589

RESUMO

AIM: One of the serious complications of sepsis is liver damage and liver failure. This study aimed to evaluate the protective and therapeutic potential of melatonin in rats with lipopolysaccharide-induced sepsis. MAIN METHODS: Female Spraque-Dawley rats received single a dose of 7.5 mg/kg lipopolysaccharide in saline to create a 24-h sepsis model. One of the other groups received melatonin at a dose of 10 mg/kg/day beginning 1 week before sepsis induction to the end of the experiment. The melatonin group received the same doses of melatonin for the same duration but not lipopolysaccharide. The vehicle group received the same doses of saline, the vehicle of melatonin, for the same duration. Twenty-four hours after the last injection, the rats were decapitated. By appropriate histochemical, immunohistochemical, biochemical, and molecular techniques, anti-necrotic, anti-apoptotic, anti-necroptotic, anti-inflammatory, and antioxidant effects of melatonin were assessed. KEY FINDINGS: Lipopolysaccharide has disrupted liver functions by inducing oxidative stress, inflammation, necrotic, apoptotic, and necroptotic cell death, thus disrupting liver functions. Melatonin was found to be beneficial in terms of inhibiting the intrinsic pathway of apoptosis and tissue oxidant levels, stimulating tissue antioxidant enzyme levels, and restoring hepatocyte functions. SIGNIFICANCE: Melatonin, at those doses and duration, was found to be hepatoprotective by mainly modulating oxidative status and apoptosis rate, however, failed to significantly reduce histopathological damage. We suggest that longer-term melatonin administration may produce anti-inflammatory and anti-necrotic effects as well.


Assuntos
Melatonina , Sepse , Ratos , Feminino , Animais , Melatonina/farmacologia , Lipopolissacarídeos/toxicidade , Ratos Wistar , Antioxidantes/metabolismo , Estresse Oxidativo , Apoptose , Necrose/tratamento farmacológico , Necrose/metabolismo , Necrose/patologia , Sepse/induzido quimicamente , Sepse/tratamento farmacológico , Sepse/metabolismo , Fígado , Anti-Inflamatórios/farmacologia
2.
J Biochem Mol Toxicol ; 37(7): e23372, 2023 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-37102204

RESUMO

Cis-diamminedichloroplatinum (II) (cisplatin, Cis) is widely employed to treat several types of cancer. It has many important toxic side effects; one of the most important of which is nephrotoxicity. Clemizole hydrochloride (Clem) as the most potent inhibitor of TRPC5 channels was tested in an animal model of Cis-induced nephrotoxicity. Rats were divided into the following groups: control; Cis (8 mg/kg); Cis + 1 mg/kg Clem; Cis + 5 mg/kg Clem; Cis + 10 mg/kg Clem. Kidney injury was detected by histopathological and biochemical analysis. Urine urea nitrogen (UUN), creatinine, urine neutrophil gelatinase-associated lipocalin (NGAL), serum catalase (CAT), and malondialdehyde (MDA) levels were determined by enzyme-linked immunosorbent assay. Total antioxidant status (TAS) and total oxidant status (TOS) were studied using a colorimetric assay. Nephrin, synaptopodin, and Rac family small GTPase 1 (RAC1) expressions were detected by Western blot analysis. Cis was found to induce histopathological alterations, including tubular degeneration, congestion, hemorrhage, hyaline casts, glomerular collapse, and apoptotic cell death. Clem at a dose of 1 and 5 mg/kg attenuated histopathological alterations. UUN, creatinine, and NGAL levels increased in the Cis-administered group, while all doses of Clem decreased in those. CAT and TAS levels decreased, while TOS and oxidative stress index levels increased in the Cis-treated group. A dose of 1 and 5 mg Clem showed antioxidant effects against oxidative stress. Cis induced lipid peroxidation by increasing MDA levels. All doses of Clem reduced MDA levels. Nephrin and synaptopodin expressions were decreased by Cis, and all doses of Clem increased that. All doses of Clem successfully depressed RAC1 expression. Clem showed a highly ameliorating effect on toxicity caused by Cis by blocking TRPC5 calcium channels.


Assuntos
Cisplatino , Insuficiência Renal , Ratos , Animais , Cisplatino/toxicidade , Lipocalina-2/metabolismo , Lipocalina-2/farmacologia , Creatinina , Rim , Insuficiência Renal/induzido quimicamente , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Estresse Oxidativo , Ureia , Canais de Cátion TRPC/metabolismo
3.
Int J Clin Pract ; 75(11): e14810, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34487588

RESUMO

BACKGROUND: Technetium-99m-dimercapto succinic acid (Tc-99m DMSA) scintigraphy is a commonly used imaging modality in children with urological abnormalities. The radiopharmaceuticals, which have the effects of ionising radiation, are used in this method. This study aimed to investigate the impact of the Tc-99m DMSA scan on renal oxidative stress and mononuclear leukocyte (MNL) DNA damage. METHODS: Children, who were followed up by paediatric nephrology at Bezmialem Vakif University and underwent Tc-99m DMSA scintigraphy between April 2015 and January 2016 with the indication of detection of renal scars, were included in this study. The exclusion criteria were nephrolithiasis, history of premature birth and recent urinary tract infection 3 months prior to scintigraphy or antibiotic use in the last 1 month. 3 mL heparinised blood samples were obtained just before, immediately after and 1 week after the scintigraphy. MNL DNA damage, total antioxidant status (TAS) and total oxidant status (TOS) were measured in the blood samples. The oxidative stress index (OSI) was calculated. Spot urine samples were obtained from each patient before and within 3 days after performing the scintigraphy. TAS/Creatinine (TAS/Cr), TOS/Creatinine (TOS/Cr) and N-acetyl-glucosaminidase/creatinine (NAG/Cr) levels were measured in the urine samples. RESULTS: Twenty-seven children were evaluated. The values between TAS, TOS and OSI levels in serum samples at baseline, immediately after and 1 week after the scintigraphy (P = .105, P = .913, and P = .721, respectively) showed no statistically significant difference. The levels of TAS/Cr, TOS/Cr, NAG/Cr ratios and OSI, which were evaluated from urine samples before and within 3 days after the scintigraphy scan were also similar (P = .391, P = .543, P = .819 and P = .179, respectively). The levels of DNA damage only increased following scintigraphy scan and decreased a week later (P < .05). CONCLUSIONS: The effect of Tc-99m DMSA scintigraphy is insufficient to create oxidative damage, but it can cause DNA damage via the direct impact of ionising radiation which can be repaired again in a short time.


Assuntos
Ácido Succínico , Tecnécio , Criança , Dano ao DNA , Humanos , Rim , Estresse Oxidativo , Cintilografia , Compostos Radiofarmacêuticos , Ácido Dimercaptossuccínico Tecnécio Tc 99m
4.
Am J Otolaryngol ; 41(1): 102328, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31732304

RESUMO

OBJECTIVE: This study aimed to investigate the healing effect of metformin on noise induced hearing loss (NIHL) by measuring audiological, biochemical and histological parameters. MATERIALS AND METHODS: 32 rats were divided into four groups (Group 1: Noise, Group 2: Noise + Metformin, Grup 3: Metformin, Grup 4: Control). Broadband noise was applied to Group 1 and Group 2 after basal measurements. Measuring audiological (distortion product otoacoustic emission (DPOAE) and Auditory Brainstem Response (ABR)), biochemical (total antioxidant status (TAS), total oxidant status (TOS), oxidative status index (OSI), DNA damage, IL-1 beta, IL-6, TNF alfa, HSF-1 and COX-2) and histological parameters. RESULTS: Group 2 had significant decreases in ABR thresholds on day 7 and day 14 compared to day 1. DPOAE values of Group 2 on the 7th and 14th days were significantly higher than the post-noise levels. DNA damage, TOS and OSI values of Group 1 were significantly higher than the other groups. The Cox-2 value of Group 1 was higher than all other groups. The HSF-1 value of Group 2 was significantly higher than that of Group 1. In terms of IL-1 Beta, IL-6 and TNF-alpha values, there was no significant difference between groups 2, 3 and 4 and these values were significantly lower than group 1. In histopathological results of our study, no significant difference was found between the groups being exposed to noise and the control group. CONCLUSION: This study showed that early period of Metformin treatment has therapeutic effect on NIHL.


Assuntos
Potenciais Evocados Auditivos do Tronco Encefálico/efeitos dos fármacos , Perda Auditiva Provocada por Ruído/tratamento farmacológico , Metformina/farmacologia , Emissões Otoacústicas Espontâneas/efeitos dos fármacos , Animais , Limiar Auditivo , Biomarcadores/metabolismo , Modelos Animais de Doenças , Feminino , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Ratos , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/metabolismo
5.
Neurol Neurochir Pol ; 54(6): 576-584, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33252137

RESUMO

AIM OF THE STUDY: Among subarachnoid haemorrhage (SAH) patients, delayed cerebral injury (DCI) and infarction are the most important causes of death and major disability. Cerebral vasospasm (cVS) and DCI remain the major cause of death and disability. Thymoquinone (TQ) is the substance most responsible for the biological activity of nigella sativa (NS) and is useful in the treatment of ischaemic and neurodegenerative diseases, oxidative stress, inflammatory events, cardiovascular and neurological diseases. We conducted an experimental study aimed to investigate the preventive and corrective effects of TQ. MATERIALS AND METHODS: 24 Sprague-Dawley rats were randomly divided into three groups. The first was the control group which was a sham surgery group. The second group was the SAH group where the double haemorrage SAH protocol was used to induce vasospasm. The third group was the SAH+TQ group, where cVS was induced by the SAH protocol and the animals received oral 2 cc thymoquinone solution for seven days at a dose of 10 mg/kg, after the induction of SAH. The rats were euthanised seven days after the first procedure. The degree of cerebral vasospasm was evaluated by measuring the basilar artery luminal area and arterial wall thickness. Apoptosis was measured by the western blot method at brainstem neural tissue. Oxidative stress was measured by the Erel Method. Endothelin-1 was measured with ELISA analysis at blood. Statistical analysis was performed. RESULTS: Endothelin-1 values were found to be statistically significantly lower in the control and SAH+TQ groups compared to the SAH group (P < 0.001). Mean lumen area values were significantly higher in the control and SAH+TQ groups than in the SAH group (P < 0.001). In the control and SAH+TQ groups, wall thickness values decreased significantly compared to the SAH group (P < 0.001). OSI values were significantly lower in the control and SAH+TQ groups than in the SAH group (P < 0.001). Apoptosis was significantly lower in the control and SAH+TQ groups than in the SAH group (P < 0.001). CONCLUSION: Our results show that post-SAH TQ inhibits/improves DCI and cVS with positive effects on oxidative stress, apoptosis, ET-1, lumen area, and vessel wall thickness, probably due to its anti-ischaemic, antispasmodic, antioxidant, anti-inflammatory, anti-apoptotic and neuroprotective effects.


Assuntos
Hemorragia Subaracnóidea , Vasoespasmo Intracraniano , Animais , Artéria Basilar , Benzoquinonas/uso terapêutico , Modelos Animais de Doenças , Humanos , Ratos , Ratos Sprague-Dawley , Hemorragia Subaracnóidea/complicações , Hemorragia Subaracnóidea/tratamento farmacológico , Vasoespasmo Intracraniano/tratamento farmacológico , Vasoespasmo Intracraniano/etiologia , Vasoespasmo Intracraniano/prevenção & controle
6.
Cell Mol Biol (Noisy-le-grand) ; 64(7): 30-35, 2018 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-29974843

RESUMO

We investigated the effects of an aqueous root extract of Cichorium intybus on Bcl-2 and cyclin B1 levels in the brain, kidney and liver volumes and changes of serum total antioxidant status (TAS) and total oxidant status (TOS) levels in ethanol induced damage in rats. The rats were divided into five groups: non-treated controls (C), maltodextrin in tap water treated (MD), 6.4% ethanol in tap water treated (ET), Cichorium intybus + maltodextrin in tap water treated (CI+MD), and Cichorium intybus + 6.4% ethanol in tap water treated (CI+ET). Rats in the CI+MD and CI+ET groups were treated with 200 mg/kg water extract of Cichorium intybus. Chronic ethanol aMDinistration significantly increased cyclin B1 and decreased Bcl-2 levels in the brain and significantly decreased TAS values, increased TOS values of serum and significantly decreased kidney volume in the ET group. There was no significant difference in the liver volume or liver cell count. Our data revealed that ethanol aMDinistration induces an overexpression of cyclin B1 and decreases levels of Bcl-2 in rat brains and induced oxidative stress in the blood. C. intybus treatment possessed a partial amelioration effect on cyclin B1 levels and TAS values.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Cichorium intybus/química , Etanol/administração & dosagem , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Antioxidantes/química , Antioxidantes/farmacologia , Encéfalo/patologia , Doença Hepática Induzida por Substâncias e Drogas/patologia , Ciclina B1/metabolismo , Rim/patologia , Fígado/patologia , Masculino , Extratos Vegetais/química , Raízes de Plantas/química , Polissacarídeos/farmacologia , Ratos , Ratos Wistar , Fatores de Tempo , Proteína de Morte Celular Associada a bcl/metabolismo
7.
Cell Mol Biol (Noisy-le-grand) ; 64(15): 71-77, 2018 Dec 31.
Artigo em Inglês | MEDLINE | ID: mdl-30672439

RESUMO

We investigated the effects of Leontice leontopetalum and Bongardia chrysogonum on apoptosis, gamma-aminobutyric acid (GABAA) receptor positive cell number, cyclin-B1 and bcl-2 levels and oxidative stress in pentylenetetrazol (PTZ) kindling in rats. Kindling was produced by subconvulsant doses of PTZ treatments in rats. Wistar albino rats were divided into 4 groups; Control, PTZ treated (PTZ), PTZ+L. leontopetalum extract treated (PTZ+LLE) and PTZ+B. chrysogonum extract treated (PTZ+BCE) groups. Extracts were given a dose (200 mg/kg) 2h before each PTZ injection. PTZ treatment significantly decreased the glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) activities and bcl-2 levels and increased the total oxidant status (TOS), malondialdehyde (MDA), cyclin B1, oxidative stress index (OSI) and number of neurons that expressed GABAA receptors when compared to the control. LLE and BCE possessed antioxidant activity in the brain and ameliorated PTZ induced oxidative stress, decreased cyclin-B1, increased bcl-2 levels, and kept the GABAA receptor number similar to that of the control despite the PTZ application.


Assuntos
Berberidaceae/química , Epilepsia/induzido quimicamente , Epilepsia/patologia , Excitação Neurológica/efeitos dos fármacos , Neuroproteção/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Ciclina B1/metabolismo , Epilepsia/tratamento farmacológico , Glutationa Peroxidase/metabolismo , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Masculino , Malondialdeído/metabolismo , Pentilenotetrazol , Extratos Vegetais/uso terapêutico , Ratos Wistar , Receptores de GABA/metabolismo , Superóxido Dismutase/metabolismo
8.
Anticancer Drugs ; 28(5): 522-530, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28244942

RESUMO

Gastric cancer (GC) is the most common cause of morbidity and mortality because of cancer. Medicinal plants containing polyphenolic compounds have gained importance in anticancer treatment. In this context, carvacrol is a main component of many plants in the family Lamiaceae that are frequently used in folk medicine and a good candidate to investigate for GC treatment. The present study aimed to explore the cytotoxic, genotoxic, apoptotic, and reactive oxygen species (ROS)-generating effects of carvacrol on gastric adenocarcinoma in vitro. For these purposes, human gastric adenocarcinoma (AGS) cells were used and analyzed after 24 h of exposure to carvacrol with different concentrations. The cytotoxicity, ROS generation, glutathione (GSH) level, and genotoxicity were investigated by the ATP cell viability assay, 2',7'-dichlorodihydrofluorescein-diacetate assay, GSH/GSSG-Glo assay, and comet assay, respectively. Apoptosis induction was detected by acridine orange/ethidium bromide staining and western blotting at below the half-maximal growth inhibitory concentration value. Carvacrol showed cytotoxic, genotoxic, apoptotic, ROS generating, and GSH-reducing effects on AGS cells in a dose-dependent manner. There was a close negative relationship between cell viability and ROS level. Carvacrol inhibited the proliferation of AGS cells, suggesting that it could be a novel and strong anticancer agent against the human gastric adenocarcinoma. These results support the interest of natural diet components in the development of therapeutic products for diseases.


Assuntos
Adenocarcinoma/tratamento farmacológico , Monoterpenos/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Trifosfato de Adenosina/metabolismo , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ensaio Cometa , Cimenos , Relação Dose-Resposta a Droga , Glutationa/metabolismo , Humanos , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia
9.
Protein J ; 42(5): 596-606, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37634214

RESUMO

Bacteriophage endolysins have been shown to hold great promise as new antibacterial agents for animal and human health in food preservation. In the present study, endolysin from Staphylococcus aureus subsp. aureus ATCC 27692-B1 bacteriophage 52 (LysSA52) was cloned, expressed, and characterized for its antimicrobial properties. Following DNA extraction from bacteriophage 52, a 1446-bp DNA fragment containing the endolysin gene (lysSA52) was obtained by PCR amplification and cloned into pET SUMO expression vector. The positive clone was validated by sequencing and open-reading frame analysis. The LysSA52 sequence shared high homology with staphylococcal phage endolysins of the SA12, SA13, and DSW2 phages and others. The cloned lysSA52 gene encoding 481 amino acids endolysin was expressed in Escherichia coli BL21 with a calculated molecular mass of 66 kDa (LysSA52). This recombinant endolysin LysSA52 exhibited lytic activity against 8 of 10 Gram-positive bacteria via agar spot-on lawn antimicrobial assay, including methicillin-resistant Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus haemolyticus, Streptococcus pneumonia, Streptococcus pyogenes, Enterococcus faecium, Enterococcus faecalis, and Bacillus atrophaeus. In addition, the 0.50 mg/mL, LysSA52 endolysins reduced about 60% of the biofilms of S. aureus and S. epidermidis established on a microtiter plate in 12 h treatment. The data from this study indicate that LysSA52 endolysin could be used as an antibacterial protein to prevent and treat infections caused by staphylococci and several other Gram-positive pathogenic bacteria irrespective of their antibiotic resistance.


Assuntos
Bacteriófagos , Staphylococcus aureus Resistente à Meticilina , Infecções Estafilocócicas , Animais , Humanos , Staphylococcus aureus , Antibacterianos
10.
J Photochem Photobiol B ; 248: 112797, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37862898

RESUMO

Drug resistant and undetectable tumors easily escape treatment leading metastases and/or recurrence of the lethal disease. Therefore, it is vital to diagnose and destroy micro tumors using simple yet novel approaches. Here, we present fluorescence-based detection and light-based destruction of cancer cells that are known to be resistant to standard therapies. We developed a superparamagnetic iron oxide nanoparticle (SPION)-based theranostic agent that is composed of self-quenching light activated photosensitizer (BPD) and EGFR targeting ligand (Anti-EGFR ScFv or GE11 peptide). Photosensitizer (BPD) was immobilized to PEG-PEI modified SPION with acid-labile linker. Prior to stimulation of the theranostic system by light its accumulation within cancer cells is vital since BPD phototoxicity and fluorescence is activated by lysosomal proteolysis. As BPD is cleaved, the system switches from off to on position which triggers imaging and therapy. Targeting, therapeutic and diagnostic features of the theranostic system were evaluated in high and moderate level EGFR expressing pancreatic cancer cell lines. Our results indicate that the system distinguishes high and moderate EGFR expression levels and yields up to 4.3-fold increase in intracellular fluorescence intensity. Amplification of fluorescence signal was as low as 1.3-fold in the moderate or no EGFR expressing cell lines. Anti-EGFR ScFv targeted SPION caused nearly 2-fold higher cell death via apoptosis in high EGFR expressing Panc-1 cell line. The developed system, possessing advanced targeting, enhanced imaging and effective therapeutic features, is a promising candidate for multi-mode detection and destruction of residual drug-resistant cancer cells.


Assuntos
Fármacos Fotossensibilizantes , Medicina de Precisão , Fármacos Fotossensibilizantes/farmacologia , Fármacos Fotossensibilizantes/uso terapêutico , Linhagem Celular Tumoral , Nanopartículas Magnéticas de Óxido de Ferro , Nanomedicina Teranóstica/métodos , Receptores ErbB/metabolismo , Concentração de Íons de Hidrogênio
12.
Mol Biotechnol ; 63(1): 24-39, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33058020

RESUMO

Laccases are polyphenol oxidoreductases used in a number of industrial applications. Due to the increasing demand for these "green catalysis" enzymes, the identification and biochemical characterisation of their novel properties is essential. In our study, cloned Madurella mycetomatis laccase (mmlac) genes were heterologously expressed in the methylotrophic yeast host Pichia pastoris. The high yield of the active recombinant protein in P. pastoris demonstrates the efficiency of a reliably constructed plasmid to express the laccase gene. The optimal biochemical conditions for the successfully expressed MmLac enzyme were identified. Detailed structural properties of the recombinant laccase were determined, and its utility in decolourisation and textile bleaching applications was examined. MmLac demonstrates good activity in an acidic pH range (4.0-6.0); is stable in the presence of cationic metals, organic solvents and under high temperatures (50-60 °C); and is stable for long-term storage at - 20 °C and - 80 °C for up to eight weeks. The structural analysis revealed that the catalytic residues are partially similar to other laccases. MmLac resulted in an increase in whiteness, whilst demonstrating high efficiency and stability and requiring the input of fewer chemicals. The performance of this enzyme makes it worthy of investigation for use in textile biotechnology applications, as well as within environmental and food technologies.


Assuntos
Biotecnologia/métodos , Lacase/química , Lacase/genética , Madurella/genética , Saccharomycetales/metabolismo , Clareadores/química , Catálise , Clonagem Molecular , Estabilidade Enzimática , Expressão Gênica , Concentração de Íons de Hidrogênio , Cinética , Lacase/isolamento & purificação , Madurella/enzimologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Temperatura
13.
Protein Pept Lett ; 28(2): 195-204, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-32729412

RESUMO

BACKGROUND: Aside from its pervasiveness, whereby it affects as much as 20% of the world's population, depression continues to be one of the most crucial psychiatric problems due to the loss of power it causes by disrupting daily life functioning, containing economic consequences, and having a high suicidal tendency. Major depression (MD) is a systemic and multifactorial disorder involving complex interactions between genetic predisposition and disturbances of various molecular pathways. OBJECTIVES: In our current study, we aimed to identify the proteins obtained from serum samples that change during depression with the MD model. METHODS: The MD model was applied through the forced swim test in rats. 14 Winstar Albino male rats were divided into two equal groups as follows: depression and control groups. Serum samples were separated by chromatographic methods and then compared with two-dimensional (2D) electrophoresis. RESULTS: A total of 9 potential diagnostic protein sequences were identified, which were distinguished with computer software. During the last phase of the study, the Matrix-Assisted Laser Desorption/ Ionization - Time of Flight (MALDI-TOF) analysis, the previous expression sequences identified among the groups were determined and classified. By comparing protein expressions, it was concluded that 9 different points could be used together as a potential biomarker. CONCLUSION: Results can help us identify a new diagnostic system that can be used to diagnose MD.


Assuntos
Biomarcadores/metabolismo , Transtorno Depressivo Maior/diagnóstico , Eletroforese em Gel Bidimensional/métodos , Proteínas/metabolismo , Proteoma/análise , Proteoma/metabolismo , Natação , Animais , Biomarcadores/análise , Transtorno Depressivo Maior/metabolismo , Masculino , Ratos , Ratos Wistar , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
14.
Protein Pept Lett ; 28(7): 817-830, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33413052

RESUMO

BACKGROUND: Polygalacturonases are a group of enzymes under pectinolytic enzymes related to enzymes that hydrolyse pectic substances. Polygalacturonases have been used in various industrial applications such as fruit juice clarification, retting of plant fibers, wastewater treatment drinks fermentation, and oil extraction. OBJECTIVES: The study was evaluated at the heterologous expression, purification, biochemical characterization, computational modeling, and performance in apple juice clarification of a new exo-polygalacturonase from Sporothrix schenckii 1099-18 (SsExo-PG) in Pichia pastoris. METHODS: Recombinant DNA technology was used in this study. Two different pPIC9K plasmids were constructed with native signal sequence-ssexo-pg and alpha signal sequence-ssexo-pg separately. Protein expression and purification performed after plasmids transformed into the Pichia pastoris. Biochemical and structural analyses were performed by using pure SsExo-PG. RESULTS: The purification of SsExo-PG was achieved using a Ni-NTA chromatography system. The enzyme was found to have a molecular mass of approximately 52 kDa. SsExo-PG presented as stable at a wide range of temperature and pH values, and to be more storage stable than other commercial pectinolytic enzyme mixtures. Structural analysis revealed that the catalytic residues of SsExo- PG are somewhat similar to other Exo-PGs. The KM and kcat values for the degradation of polygalacturonic acid (PGA) by the purified enzyme were found to be 0.5868 µM and 179 s-1, respectively. Cu2+ was found to enhance SsExo-PG activity while Ag2+ and Fe2+ almost completely inhibited enzyme activity. The enzyme reduced turbidity up to 80% thus enhanced the clarification of apple juice. SsExo-PG showed promising performance when compared with other commercial pectinolytic enzyme mixtures. CONCLUSION: The clarification potential of SsExo-PG was revealed by comparing it with commercial pectinolytic enzymes. The following parameters of the process of apple juice clarification processes showed that SsExo-PG is highly stable and has a novel performance.


Assuntos
Sucos de Frutas e Vegetais/análise , Proteínas Fúngicas/química , Malus/química , Pectinas/química , Poligalacturonase/química , Sporothrix/química , Cátions Bivalentes , Clonagem Molecular , Cobre/química , Estabilidade Enzimática , Tecnologia de Alimentos/métodos , Proteínas Fúngicas/isolamento & purificação , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Ferro/química , Cinética , Peso Molecular , Pichia/genética , Pichia/metabolismo , Poligalacturonase/isolamento & purificação , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Prata/química , Sporothrix/enzimologia , Temperatura
15.
Appl Biochem Biotechnol ; 193(2): 363-376, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32974869

RESUMO

The NAD+-dependent formate dehydrogenase (FDH; EC 1.2.1.2) from Candida boidinii (CboFDH) has been extensively used in NAD(H)-dependent industrial biocatalysis as well as in the production of renewable fuels and chemicals from carbon dioxide. In the present work, the effect of amino acid residues Phe285, Gln287, and His311 on structural stability was investigated by site-directed mutagenesis. The wild-type and mutant enzymes (Gln287Glu, His311Gln, and Phe285Thr/His311Gln) were cloned and expressed in Escherichia coli. Circular dichroism (CD) spectroscopy was used to determine the effect of each mutation on thermostability. The results showed the decisive roles of Phe285, Gln287, and His311 on enhancing the enzyme's thermostability. The melting temperatures for the wild-type and the mutant enzymes Gln287Glu, His311Gln, and Phe285Thr/His311Gln were 64, 70, 77, and 73 °C, respectively. The effects of pH and temperature on catalytic activity of the wild-type and mutant enzymes were also investigated. Interestingly, the mutant enzyme His311Gln exhibits a large shift of pH optimum at the basic pH range (1 pH unit) and substantial increase of the optimum temperature (25 °C). The present work supports the multifunctional role of the conserved residues Phe285, Gln287, and His311 and further underlines their pivotal roles as targets in protein engineering studies.


Assuntos
Formiato Desidrogenases/química , Proteínas Fúngicas/química , Saccharomycetales/enzimologia , Substituição de Aminoácidos , Aminoácidos , Estabilidade Enzimática , Formiato Desidrogenases/genética , Proteínas Fúngicas/genética , Mutação de Sentido Incorreto , Domínios Proteicos , Saccharomycetales/genética
16.
Asian Pac J Cancer Prev ; 21(9): 2531-2537, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32986349

RESUMO

Cornelian Cherry (Cornus Mas L) has widespread use due to its anti-inflammatory, anti-carcinogenic and anti-oxidant properties. In this study, the effects of Cornus Mas L (C. mas L) in different dosages on the biochemical values of mice organs were investigated in the Ehrlich Ascites tumor model, which originated from mice breast adenocarcinoma and developed in Balb/C mice. In our study, 32 Balb/C type male mice were used. Ehrlich Ascites Tumor (EAT) cells (1x106 EAT cell) from the stock animal were injected into all the mice in an intraperitoneal way. Experimental groups were given 100 and 200mg/kg C. mas L extract intraperitoneally for 9 days. The weights of the animals were recorded every day and were sacrificed on the 9th day. To estimate tumor proliferation of the lung, brain, kidney, liver, and testis, antioxidant parameters were recorded including, the reduced glutathione (GSH), lipid peroxidation, glutathione S-transferase (GST), superoxide dismutase (SOD) and catalase (CAT). Treatments of different doses of C. mas L. meaningfully (p < 0.05) modulated the lung, brain, kidney, liver and testis tissues antioxidant parameters as compared to the control. Our study showed the anti-tumor effect of C. mas L. in assisted tumor development with EAT cells, conceivably moderated by the enhancement of oxidative stress due to numerous mechanisms.
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Assuntos
Antioxidantes/farmacologia , Neoplasias Encefálicas/tratamento farmacológico , Carcinoma de Ehrlich/tratamento farmacológico , Cornus/química , Neoplasias Renais/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Testiculares/tratamento farmacológico , Animais , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Carcinoma de Ehrlich/metabolismo , Carcinoma de Ehrlich/patologia , Neoplasias Renais/metabolismo , Neoplasias Renais/patologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Estresse Oxidativo , Fitoterapia , Extratos Vegetais/farmacologia , Neoplasias Testiculares/metabolismo , Neoplasias Testiculares/patologia
17.
Brain Res Bull ; 154: 68-80, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31715313

RESUMO

Traumatic brain injury (TBI) is one of the important reason of morbidity and mortality. While the primary injury due to mechanical impact is unavoidable, the secondary injury which is formed as a result of primary injury and thought to occur due to neuroinflammation in the forefront can be prevented and by this way mortality and morbidity can be reduced. High mobility group box-1 (HMGB1) is a protein that triggers the neuroinflammatory process by being released from the nucleus of necrotic tissues after primary injury. The aim of this study is to investigate the effects of HMGB1 on its receptors TLR4 and RAGE, cerebral edema, blood-brain barrier, oxidative stress and apoptosis causing secondary damage in an experimental traumatic brain injury model. Weighing between 280-320 g, 10 to 12 weeks-old, a total of 30 adult male Sprague-Dawley rats were used for the experiments. The rats were randomly assigned to 3 groups: 1) Control, 2) TBI and 3) TBI + ethyl pyruvate group (n = 10 per group). Right parietal cortical contusion was made by using a weight-dropping TBI method. Brain samples were harvested from pericontusional area at 24 h after TBI. HMGB1, TLR4, RAGE, occludin, claudin-5, ZO-1 levels are investigated by western blot analyses and immunohistochemistry examinations. HMGB-1, TLR4 and RAGE expressions increased after TBI. Major tight junction proteins in the blood-brain barrier: occludin, claudin-5 and ZO-1 expressions decreased after TBI. Brain edema increased after TBI. Also, proapoptotic bax and active caspase 3 expressions increased, antiapoptotic bcl-2 levels decreased after TBI. Total oxidant status and oxidative stress increased, total antioxidant status decreased after TBI. HMGB-1 protein plays a key role in the pathophysiology of traumatic brain injury.


Assuntos
Lesões Encefálicas Traumáticas/metabolismo , Proteína HMGB1/metabolismo , Animais , Apoptose/fisiologia , Barreira Hematoencefálica/metabolismo , Encéfalo/metabolismo , Edema Encefálico/etiologia , Edema Encefálico/metabolismo , Lesões Encefálicas/complicações , Lesões Encefálicas Traumáticas/fisiopatologia , Claudina-5/metabolismo , Modelos Animais de Doenças , Domínios HMG-Box/fisiologia , Proteína HMGB1/fisiologia , Proteínas de Grupo de Alta Mobilidade/metabolismo , Masculino , Ocludina/metabolismo , Estresse Oxidativo/fisiologia , Piruvatos/farmacologia , Ratos , Ratos Sprague-Dawley , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Receptor 4 Toll-Like/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
18.
J Neurosurg Sci ; 63(6): 714-722, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26439453

RESUMO

BACKGROUND: Despite optimal medical therapy the mortality rate approaches 50% in MCA infarctions. Although recent studies have been showed life-saving effect of hemicraniectomy; there are a few data available in regard to neuroprotection effect of decompressive craniectomy (DC). We induced a malignant cerebral ischemia model by intraluminal permanent middle cerebral artery occlusion (MCAo) in male rats for defining the neuroprotective effects of early DC on brain-blood barrier (BBB) molecular changes, infarct size and cerebral edema. METHODS: A total of 48 male Spraque-Dawley rats were allocated to 4 groups; sham (N.=9), control (N.=9), experiment 1 (N.=15), experiment 2 (N.=15). DC was performed by creating a bone flap, after MCAo at 4 and 24 hours. After 28 hours of survival, all animals were sacrificed. Infarction volumes were calculated from TTC (2,3,5-triphenyl-2H-tetrazolium chloride)-stained brain sections. In all groups, cerebral edema was quantified as a change in the percentage of brain water content. Western Blot was used to analyze the expression of tight junction protein claudin-5 and occludin. RESULTS: Brain water content was calculated 75.18±0.75% in the early DC group and 77.76±0.71% in the late DC group. No significant difference was found between experiment groups (P=0.178). In the early DC group; occludin and claudin-5 were significantly expressed at higher levels compared to late DC group (occluding, P=0.013; claudin-5, P=0.034). At early DC group (73.38±23.11 mm3) the final infarct volumes were significantly smaller than in the late DC group (377.18±39.23 mm3) (P=0.013). CONCLUSIONS: The study results supported the neuroprotective effects of early DC in malignant MCA infarcts.


Assuntos
Barreira Hematoencefálica/patologia , Edema Encefálico/cirurgia , Infarto da Artéria Cerebral Média/cirurgia , Fármacos Neuroprotetores/farmacologia , Animais , Encéfalo/cirurgia , Isquemia Encefálica/cirurgia , Craniectomia Descompressiva/métodos , Masculino , Ratos
19.
World Neurosurg ; 128: e570-e581, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31054338

RESUMO

BACKGROUND: Patients with brain metastasis from melanoma have a dismal prognosis with poor survival time. Gamma Knife (GK) is an effective treatment to control brain metastasis from melanoma. Thymoquinone (TQ) has emerged as a potential therapeutic option due to its antiproliferative effects on various cancers. The purpose of the study was to assess the effect of GK on B16-F10 melanoma cells in vitro and intracerebral melanoma in vivo, and its synergistic effect in combination with TQ. METHODS: The effects of GK and combination treatment of GK and TQ were studied on B16-F10 melanoma cells by evaluating cytotoxicity with an adenosine triphosphate assay, apoptosis by acridine orange staining, and genotoxicity by comet assay. Western blot analysis was performed to investigate the expression of STAT3, p-STAT3 (Tyr705), JAK2, p-JAK2, caspase-3, Bax, Bcl-2, survivin, and ß-actin. Expression of inflammatory cytokines was assessed by enzyme-linked immunosorbent assay. GK alone and in combination with TQ was assessed in an established intracerebral melanoma tumor in mice. RESULTS: The effects of GK on cytotoxicity, genotoxicity, and apoptosis were enhanced by TQ in B16-F10 melanoma cells. GK induced apoptosis through inhibition of p-STAT3 expression, which in turn regulated pro- and antiapoptotic proteins such as caspase-3, Bax, Bcl-2, and survivin. Adding TQ to GK irradiation further enhanced this apoptotic effect of GK irradiation. GK was shown to reduce the levels of tumor-related inflammatory cytokines in B16-F10 melanoma cells. This effect was more pronounced when TQ was added to GK irradiation. GK with 15 Gy increased the survival of mice with intracerebral melanoma compared with untreated mice. However, despite the additive effect of TQ in addition to GK irradiation on B16-F10 melanoma cells in vitro, TQ did not add any significant survival benefit to GK treatment in mice with intracerebral melanoma. CONCLUSIONS: Our findings suggest that TQ would be a potential therapeutic agent in addition to GK to enhance the antitumor effect of irradiation. Further studies are required to support our findings.


Assuntos
Apoptose/efeitos dos fármacos , Benzoquinonas/farmacologia , Neoplasias Encefálicas/terapia , Dano ao DNA/efeitos dos fármacos , Melanoma Experimental/terapia , Radiocirurgia/métodos , Fator de Transcrição STAT3/efeitos dos fármacos , Actinas/efeitos dos fármacos , Actinas/metabolismo , Actinas/efeitos da radiação , Animais , Apoptose/efeitos da radiação , Western Blotting , Neoplasias Encefálicas/secundário , Caspase 3/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 3/efeitos da radiação , Linhagem Celular Tumoral , Terapia Combinada , Dano ao DNA/efeitos da radiação , Técnicas In Vitro , Janus Quinase 2/efeitos dos fármacos , Janus Quinase 2/metabolismo , Janus Quinase 2/efeitos da radiação , Melanoma Experimental/secundário , Camundongos , Fosfoproteínas/efeitos dos fármacos , Fosfoproteínas/metabolismo , Fosfoproteínas/efeitos da radiação , Proteínas Proto-Oncogênicas c-bcl-2/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/efeitos da radiação , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT3/efeitos da radiação , Survivina/efeitos dos fármacos , Survivina/metabolismo , Survivina/efeitos da radiação , Proteína X Associada a bcl-2/efeitos dos fármacos , Proteína X Associada a bcl-2/metabolismo , Proteína X Associada a bcl-2/efeitos da radiação
20.
Nefrologia (Engl Ed) ; 39(4): 411-423, 2019.
Artigo em Inglês, Espanhol | MEDLINE | ID: mdl-30712966

RESUMO

BACKGROUND: Ischemia-reperfusion injury causes various severe morphological and functional changes in diabetic patients. To date, numerous antidiabetic and antioxidant agents have been used for treatment of the disease-related changes. OBJECTIVES: We aimed to examine effective therapeutic doses or doses of berberine against renal ischemia/reperfusion injury (IRI) in a streptozotocin (STZ)-induced diabetic rat model by histopathological and biochemical analysis. METHODS: Thirty male Sprague Dawley rats were treated with STZ injection for the development of diabetes, and divided into the following groups: STZ-induced diabetic group (STZ); IRI-induced diabetic group (STZ+IRI); 50mg/kg berberine (BRB) treated diabetic group after inducing IRI (STZ+IRI+BRB1); 100mg/kg BRB treated diabetic group after IRI (STZ+IRI+BRB2); 150mg/kg BRB treated diabetic group after IRI (STZ+IRI+BRB3). Bilateral renal ischemia model was applied for 45min, then reperfusion was allowed for 14 days in STZ-induced diabetic rats. Renal injury was detected histopathologically. Blood urea nitrogen (BUN), creatinine and lactate dehydrogenase (LDH) levels were measured in serum using the ELISA method. Total antioxidant status (TAS) and total oxidant status (TOS) of renal tissue was studied by spectrophotometric assay. Oxidative stress index (OSI) was calculated as TOS-to-TAS ratio. Tumor necrosis factor alpha (TNF-α), C-reactive protein (CRP), Na+/K+-ATPase (sodium pump), and Ca2+-ATPase (calcium ATPase) enzyme levels were measured in tissues using the ELISA method. Anti-apoptotic Bax and pro-apoptotic Bcl-2 protein levels were detected by Western blot analysis. All data were evaluated statistically. RESULTS: The highest histopathological score was detected in the STZ+IRI group compared to the other group. BRB administration at the doses of 100mg/kg and 150mg/kg markedly improved renal injury. BUN and creatinine levels significantly increased in the STZ+IRI group compared to the STZ group (p<0.001). 100mg/kg and 150mg/kg BRB administration significantly decreased those levels (p<0.01). The highest TOS and the lowest TAS levels were detected in the STZ+IRI group (p<0.001). IRI markedly aggravated inflammation via increasing levels of TNF-α and CRP (<0.001), and caused apoptosis via inducing Bcl-2 protein, and suppressing Bax protein (p<0.001). BRB administration at the doses of 100mg/kg and 150mg/kg showed anti-oxidant, anti-inflammatory and anti-apoptotic effects (p<0.01). The LDH enzyme, was used as a necrosis marker, was higher in the STZ+IRI group than other groups. BRB administration at all of the doses, resulted in the decline of LDH enzyme level (p<0.001). Ca2+-ATPase and Na+/K+-ATPase enzyme activities decreased in the STZ+IRI group compared to the STZ group (p<0.001), while BRB administration at the doses of 100mg/kg and 150mg/kg significantly increased those of enzyme activities, respectively (p<0.05). CONCLUSION: Ischemia with diabetes caused severe histopathological and biochemical damage in renal tissue. The high doses of berberine markedly improved histopathological findings, regulated kidney function via decreasing BUN and creatinine levels, and rearranged intercellular ion concentration via increasing Na+/K+-ATPase and Ca2+- ATPase levels. Berberine showed anti-oxidant, anti-apoptotic, and anti-inflammatory effects. According to these data, we suggest that berberine at the doses of 100 and 150mg may be used as a potential therapeutic agent to prevent renal ischemic injury.


Assuntos
Berberina/administração & dosagem , Angiopatias Diabéticas/tratamento farmacológico , Rim/irrigação sanguínea , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Diabetes Mellitus Experimental , Relação Dose-Resposta a Droga , Masculino , Ratos , Ratos Sprague-Dawley
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