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1.
Braz. j. med. biol. res ; 54(2): e9017, 2021. graf
Article in English | LILACS, ColecionaSUS | ID: biblio-1142574

ABSTRACT

The purpose of this study was to investigate the anti-cancer effect of melittin on growth, migration, invasion, and apoptosis of non-small-cell lung cancer (NSCLC) cells. This study also explored the potential anti-cancer mechanism of melittin in NSCLC cells. The results demonstrated that melittin suppressed growth, migration, and invasion, and induced apoptosis of NSCLC cells in vitro. Melittin increased pro-apoptotic caspase-3 and Apaf-1 gene expression. Melittin inhibited tumor growth factor (TGF)-β expression and phosphorylated ERK/total ERK (pERK/tERK) in NSCLC cells. However, TGF-β overexpression (pTGF-β) abolished melittin-decreased TGF-β expression and pERK/tERK in NSCLC cells. Treatment with melittin suppressed tumor growth and prolonged mouse survival during the 120-day observation in vivo. Treatment with melittin increased TUNEL-positive cells and decreased expression levels of TGF-β and ERK in tumor tissue compared to the control group. In conclusion, the findings of this study indicated that melittin inhibited growth, migration, and invasion, and induced apoptosis of NSCLC cells through down-regulation of TGF-β-mediated ERK signaling pathway, suggesting melittin may be a promising anti-cancer agent for NSCLC therapy.


Subject(s)
Animals , Rabbits , Apoptosis , Carcinoma, Non-Small-Cell Lung/genetics , Carcinoma, Non-Small-Cell Lung/drug therapy , MAP Kinase Signaling System , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Melitten/pharmacology , Down-Regulation , Gene Expression Regulation, Neoplastic , Cell Movement , Transforming Growth Factor beta/metabolism , Cell Line, Tumor , Caspase 3 , Apoptotic Protease-Activating Factor 1 , Neoplasm Invasiveness
2.
Article in Chinese | WPRIM | ID: wpr-880101

ABSTRACT

OBJECTIVE@#To investigate the effect of 2-methoxyestradiol (2-ME2) to lymphoma Raji cells and its mechanism.@*METHODS@#Different concentrations of 2-ME2 were used to treat lymphoma Raji cells. CCK8 method was used to detect the effect of 2-ME2 to proliferation of Raji cells. Flow cytometry FITC/PI double labeling method was used to detect early apoptosis of the cells. Western blotting was used to detect the effect of 2-ME2 to the expression of BCL-2, Bax, Caspase-3 and C-myc proteins in Raji cells.@*RESULTS@#2-ME2 significantly inhibited the proliferation of Raji cells. The inhibition rate increased with the increasing of drug concentration, and increased significantly with the prolongation of drug treatment time (r=0.9215). Flow cytometry FITC/PI double staining showed that the apoptotic rate of 2.5 μmol/L 2-ME2 treatment group was (33.79±1.63) %, while the apoptosis rate of the 48 h group was (51.90±2.72) %, and that of the control group was (7.08±0.36) %. After treated with 2.5 μmol/L 2-ME2 for 12 h, the expression of Bax protein was up-regulated, BCL-2 protein was down-regulated, caspase-3 protein expression was up-regulated, and C-myc protein expression was down-regulated, all of them showed a time-dependent relationship.@*CONCLUSION@#2-ME2 shows obvious inhibitory effect on lymphoma Raji cells in a dose- and time-dependent manner. Its mechanism of treatment on lymphoma Raji cells may be related to up-regulation of Bax/BCL-2 ratio and activation of Caspase-3 to induce apoptosis in cancer cells. Down-regulation of C-myc protein expression also participates in the apoptotic process.


Subject(s)
2-Methoxyestradiol , Apoptosis , Caspase 3/metabolism , Cell Line, Tumor , Cell Proliferation , Humans , Lymphoma , Proto-Oncogene Proteins c-bcl-2/metabolism , Up-Regulation , bcl-2-Associated X Protein
3.
Int. j. morphol ; 38(5): 1217-1222, oct. 2020. graf
Article in English | LILACS | ID: biblio-1134428

ABSTRACT

SUMMARY: Repeated stress is a risk factor for memory impairment and neurological abnormalities in both humans and animals. We sought to investigate the extent of (i) brain tissue injury; (ii) nitrosative and oxidative stress in brain tissue homogenates; (iii) apoptotic and survival biomarkers in brain tissue homogenates; and (iv) immobility and climbing abilities, induced over a period of three weeks by chronic unpredictable stress (CUS). Wistar rats were either left untreated (Control group) or exposed to a variety of unpredictable stressors daily before being sacrificed after 3 weeks (model group). Assessment of depression-like behavior was performed and animals were then culled and harvested brain tissues were stained with basic histological staining and examined under light microscopy. In addition, brain tissue homogenates were prepared and assayed for these parameters; inducible nitric oxide synthase (iNOS), malondialdehyde (MDA), superoxide dismutase (SOD), caspase-3, and B-cell lymphoma 2 (Bcl-2). Histology images showed CUS induced profound damage to the cerebral cortex as demonstrated by severe neuronal damage with shrunken cells, disrupted atrophic nuclei, perineuronal vacuolation and swollen glial cells. CUS also significantly (p<0.05) induced iNOS, MDA, and caspase-3, whereas SOD and Bcl-2 brain tissue levels were inhibited by CUS. In addition, data from the depression-like behavior, forced swimming test showed significant (p<0.05) increase in animal immobility and decrease in climbing ability in the model group of rats. Thus, here we demonstrated a reliable rat model of chronic stress-induced brain injury, which can further be used to investigate beneficial drugs or agents used for a period of three weeks to protect against CUS-induced brain damage.


RESUMEN: El estrés crónico es un factor de riesgo para el deterioro de la memoria y las anomalías neurológicas tanto en humanos como en animales. Intentamos investigar el alcance de lesión del tejido cerebral; (ii) estrés nitrosativo y oxidativo en homogeneizados de tejido cerebral; (iii) biomarcadores apoptóticos y de supervivencia en homogeneizados de tejido cerebral; y (iv) inmovilidad y habilidades de escalada, inducidas durante un período de tres semanas por estrés crónico impredecible (ECI). Se dejaron sin tratamiento (grupo control) ratas Wistar, o se expusieron a una variedad de factores estresantes impredecibles diariamente antes de ser sacrificadas después de 3 semanas (grupo modelo). Se realizó una evaluación del comportamiento similar a la depresión y luego se sacrificaron los animales y se tiñeron los tejidos cerebrales con tinción histológica básica y se examinaron con microscopía óptica. Además, se prepararon homogeneizados de tejido cerebral y se analizaron los siguientes parámetros; óxido nítrico sintasa inducible (iNOS), malondialdehído (MDA), superóxido dismutasa (SOD), caspasa- 3 y linfoma de células B 2 (Bcl-2). Las imágenes histológicas mostraron que el CUS indujo un daño profundo en la corteza cerebral como lo demuestra el daño neuronal severo con células encogidas, núcleos atróficos alterados, vacuolación perineuronal y células gliales inflamadas. ECI también indujo significativamente (p <0,05) iNOS, MDA y caspase-3, mientras que los niveles de tejido cerebral SOD y Bcl-2 fueron inhibidos por ECI. Además, los datos del comportamiento similar a la de- presión, la prueba de natación forzada mostró un aumento significativo (p <0,05) en la inmovilidad animal y una disminución en la capacidad de escalada en el grupo modelo de ratas. Por lo tanto, aquí demostramos un modelo confiable de daño cerebral crónico en rata inducido por el estrés, que se puede utilizar para investigar medicamentos o agentes beneficiosos usados durante un período de tres semanas para proteger el daño cerebral inducido por ECI.


Subject(s)
Animals , Male , Rats , Stress, Psychological/complications , Brain Damage, Chronic/pathology , Superoxide Dismutase/analysis , Behavior, Animal , Brain Injuries/metabolism , Biomarkers , Cerebral Cortex , Chronic Disease , Analysis of Variance , Rats, Wistar , Apoptosis , Oxidative Stress , Nitric Oxide Synthase/analysis , Proto-Oncogene Proteins c-bcl-2 , Depression , Disease Models, Animal , Caspase 3/analysis , Nitrosative Stress , Malondialdehyde/analysis
4.
Int. j. morphol ; 38(3): 523-529, June 2020. graf
Article in English | LILACS | ID: biblio-1098282

ABSTRACT

This study aimed to investigate the morphometric and the pattern of protein and gene expression related to the extrinsic apoptotic pathway in experimental focal cerebral ischemia and the hole of neuroprotection with hypothermia and ketoprofen. For this analysis, 120 rats were randomly divided into 3 groups (20 animals each): control - no surgery (20 animals); sham - simulation of surgery (20 animals); ischemic - focal ischemia for 1 hour, without reperfusion (80 animals) and divided into four subgroups with 20 animals each: ischemic + intraischemic hypothermia; ischemic + previous intravenous ketoprofen, and ischemic + hypothermia and ketoprofen. The infarct volume was measured using morphometric analysis of infarct areas defined by triphenyl tetrazolium chloride and the patterns of expression of the apoptosis genes (Fas, c-Flip, caspase-8 and caspase-3) and the apoptosis protein caspase-3 were evaluated by quantitative real-time PCR and immunohistochemistry, respectively. Hypo expression of genes of extrinsic pathway of apoptosis was observed: Fas receptor, c-Flip and caspase-8 in the ischemics areas. Increases in the gene and protein caspase-3 in the ischemic areas were also observed, and these increases were reduced by hypothermia and ketoprofen, also noted in the morphometric study. The caspases-3 increase suggests that this gene plays an important role in apoptosis, probably culminating in cell death and that the neuroprotective effect of hypothermia and ketoprofen is involved.


Este estudio tuvo como objetivo investigar la morfometría y el patrón de expresión de proteínas y genes relacionados con la vía apoptótica extrínseca en la isquemia cerebral focal experimental y el agujero de neuroprotección con hipotermia y ketoprofeno. Se dividieron aleatoriamente 120 ratas en 3 grupos (20 animales cada uno): control - sin cirugía (20 animales); simulación - simulación de cirugía (20 animales); isquemia isquemia focal durante 1 hora, sin reperfusión (80 animales) y dividida en cuatro subgrupos con 20 animales cada uno: isquemia + hipotermia intraisquémica; isquemia + ketoprofeno intravenoso previo, e isquemia + hipotermia y ketoprofeno. El volumen del infarto se midió utilizando un análisis morfométrico de áreas de infarto definidas por cloruro de trifenil tetrazolio y los patrones de expresión de los genes de apoptosis (Fas, c-Flip, caspase-8 y caspase-3) y la proteína de apoptosis caspase-3 fueron evaluados por PCR cuantitativa en tiempo real e inmunohistoquímica, respectivamente. Se observó hipoexpresión de genes de la vía extrínseca de la apoptosis: receptor Fas, c-Flip y caspasa-8 en las áreas isquémicas. También se observaron aumentos en el gen y la proteína caspasa-3 en las áreas isquémicas y estos aumentos se redujeron por hipotermia y ketoprofeno, también observado por estudio morfométrico. El aumento de caspasas-3 sugiere que este gen tiene un papel importante en la apoptosis, y probable causa de muerte celular, involucrando el efecto neuroprotector de la hipotermia y el ketoprofeno.


Subject(s)
Animals , Rats , Brain Ischemia/genetics , Brain Ischemia/metabolism , Immunohistochemistry , Brain Ischemia/pathology , Brain Ischemia/therapy , Ketoprofen/pharmacology , Apoptosis/genetics , Neuroprotective Agents/pharmacology , Disease Models, Animal , Caspase 3/genetics , Caspase 8/genetics , Real-Time Polymerase Chain Reaction , Hypothermia, Induced
5.
Rev. ADM ; 77(2): 70-79, mar.-abr. 2020. ilus, graf
Article in Spanish | LILACS | ID: biblio-1100338

ABSTRACT

Las enfermedades autoinmunes tienen múltiples manifestaciones en estomatología, entre las más frecuentes se encuentra el liquen plano oral (LPO), se trata de una enfermedad crónica con manifestaciones clínicas en piel y mucosas. Se agrupa en dos formas anatomoclínicas, la de curso evolutivo benigno identificado como típico y la susceptible de transformación maligna, identificada como atípico. Histológicamente, la degeneración vacuolar del estrato basal del epitelio es el signo histomorfológico patognomónico seguido de apoptosis celular. La apoptosis es un evento esencial entre los fenómenos del ciclo celular, sucede con la finalidad de eliminar células dañadas o inútiles. De todas las proteínas implicadas las caspasas son los responsables de la ejecución de este mecanismo, especialmente la caspasa 3 por fragmentar y activar otras caspasas responsables de la proteólisis. El potencial de transformación maligna del LPO podría estar en relación con el fallo de este mecanismo de regulación del ciclo de las células epiteliales agredidas y la persistencia de células dañadas. El presente trabajo de investigación tuvo como objetivo analizar la presencia y proporción de apoptosis en las distintas variantes de LPO con técnicas histológicas de rutina y posterior aplicación de inmunohistoquímica, utilizando como marcador la caspasa 3. Se obtuvieron 20 biopsias de LPO de cinco variedades clínicas nueve variantes típicas (VT): cinco placa, cuatro reticulares y 11 variantes atípicas (VA): dos atróficos, seis erosivos, tres ampollares. El método de evaluación fue semicuantitativo, se consideró en función del porcentaje, se realizó un recuento celular de un total de 100 células en cinco campos de gran aumento considerando las siguientes categorías según ausencia, presencia leve (< 10%), moderada (10 ≤ 25%), intensa (25 ≤ 50%), no valorables. Se encontró una buena correlación de los cambios histológicos y el grado de expresión del marcador utilizado para poner en evidencia la apoptosis, sobre todo con las muestras de LPO de variante atípica. En los casos de las variantes atípicas de liquen observados en comparación con la tinción de rutina (H/E) se observó igualdad o una disminución en algunos casos del número de queratinocitos apoptóticos. En cuanto a las variantes clínicas consideradas «típicas¼ se observó que el recuento de células en apoptosis estaba significativamente elevado. Obtuvimos excelentes resultados con el inmunomarcador caspasa 3, el cual coincide con la literatura en su alta sensibilidad como recurso para cuantificar el número de apoptosis en estas lesiones orales (AU)


Autoimmune diseases have multiple manifestations in stomatology, among the most frequent is oral lichen planus (LPO), it is a chronic disease with clinical manifestations in skin and mucous membranes. It is grouped into two anatomoclinic forms, the benign evolutionary course identified as typical and susceptible to malignant transformation, identified as atypical. Histologically, vacuolar degeneration of the basal stratum of the epithelium is the pathognomonic histomorphological sign followed by cellular apoptosis. Apoptosis is an essential event among cell cycle phenomena, it happens in order to eliminate damaged or useless cells. Of all the proteins involved, caspases are responsible for the execution of this mechanism, especially caspase-3 for fragmenting and activating other caspases responsible for proteolysis. The potential for malignant transformation of the LPO could be related to the failure of this mechanism to regulate the cycle of attacked epithelial cells and the persistence of damaged cells. This research work aimed to analyze the presence and proportion of apoptosis in the different variants of LPO with routine histological techniques and subsequent application of immunohistochemistry, using caspase as a marker 3. 20 LPO biopsies from 5 clinical varieties were obtained 9 typical variants (VT): 5 plate, 4 reticular and 11 atypical variants (VA): 2 atrophic, 6 erosive, 3 ampoules. The evaluation method was semi-quantitative considering the percentage, making a cell count of a total of 100 cells, in five large-scale fields considering the following categories according to absence, mild presence (< 10%), moderate (10 ≤ 25%), intense (25 ≤ 50%), not valuable. We found a good correlation of histological changes and the degree of expression of the marker used to highlight apoptosis, especially with the atypical variant LPO samples. In the cases of atypical variants of lichen observed, compared with routine staining (H/E) we find equality or a decrease in some cases of the number of apoptotic keratinocytes. For clinical variants considered «typical¼ it was observed that the cell count in apoptosis was significantly increased. We obtained excellent results with the caspase 3 immunomarker coinciding with the literature of its high sensitivity as a resource to quantify the number of apoptosis in these oral lesions (AU)


Subject(s)
Humans , Male , Female , Biomarkers , Cell Transformation, Neoplastic , Apoptosis , Lichen Planus, Oral/immunology , Caspase 3 , Biopsy , Immunohistochemistry , Statistical Analysis
6.
Korean Circulation Journal ; : 250-263, 2020.
Article in English | WPRIM | ID: wpr-811353

ABSTRACT

BACKGROUND AND OBJECTIVES: To reveal the detail mechanism of miR-484 on myocardial ischemia-reperfusion (MI/R) injury.METHODS: Rats model of MI/R injury was established based on control (Con; sham operate) group, ischemia-reperfusion (I/R) group, miR-484 treatment (miR) group, and I/R-negative control (IR-C) group, followed by pathological and interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-1β expression evaluation. Then the myocardial apoptosis, as well as the expression of miR-484, caspase-3, and caspase-9 in myocardium were examined. Finally, the regulatory relation between miR-484 and SMAD family member 7 (SMAD7) was predicated, followed by verification analysis.RESULTS: Compared with Con group, the expression of miR-484 in I/R and IR-C group was decreased. Compared with I/R and IR-C group, the expression of miR-484 was increased in miR group. Compared with Con group, the expression levels of IL-6, TNF-α, and IL-1β in cardiac myocytes of I/R group and IR-C group were increased. Compared with Con group, the apoptotic index, membrane potential of I/R, and the expression of caspase-3/9 were increased in IR-C group. Compared with the I/R and IR-C groups, the apoptotic index of myocardial cells in the ischemic region was decreased, the membrane potential was increased, and the expression of caspase-3/9 was decreased significantly in the miR group. SMAD7 was the target gene of miR-484.CONCLUSIONS: MiR-484 protected myocardial cells from I/R injury by suppressing caspase-3 and caspase-9 expression during cardiomyocyte apoptosis. MiR-484 reduced the expression of IL-6, TNF-α, and IL-1β in MI/R. MiR-484 might alleviate the decreasing of mitochondrial membrane potential in MI/R cells.


Subject(s)
Animals , Apoptosis , Caspase 3 , Caspase 9 , Humans , Interleukin-6 , Interleukins , Membrane Potential, Mitochondrial , Membrane Potentials , Myocardium , Myocytes, Cardiac , Rats , Reperfusion Injury , Tumor Necrosis Factor-alpha
9.
Int. j. odontostomatol. (Print) ; 13(4): 411-417, dic. 2019. graf
Article in Spanish | LILACS | ID: biblio-1056477

ABSTRACT

RESUMEN: Las patologías pulpares han sido un verdadero reto para la odontología principalmente por su tratamiento. Actualmente, existen numerosos biomateriales en el mercado que reportan tener propiedades inherentes en los tejidos dentarios. Sin embargo, diferentes estudios sobre múltiples líneas celulares expuestas a estos biomateriales demuestran resultados controversiales como biocompatiblidad y citotoxicidad celular. Biodentine, es un cemento endodóntico en base a silicatos cálcico de múltiples aplicaciones, que prestaría propiedades de biocompatibilidad como bioactividad celular, características que le permitirían incluso ser utilizado en contacto directo con la pulpa dental. El objetivo de este estudio es la evaluación in-vitro de Biodentine, sobre cultivos de células de la pulpa dental humana (CCPDH). Se prepararon discos de cemento de Biodentine™ de 2 x 6 mm, los que se expusieron a cultivos de células aisladas de la pulpa dental humana. Luego de 24, 48 y 72 horas de exposición, se realizaron ensayos de viabilidad celular utilizando el método colorimétrico MTT. También se realizaron ensayos de expresión proteica de dos proteínas involucradas en la vía de señalización de la apoptosis celular: Caspasa - 3 clivada y Poli (ADP-Ribosa) Polimerasa, PARP - 1. Existen diferencias estadísticamente significativas (p<0,05) en los ensayos de viabilidad celular entre las células expuestas a Biodentine y el grupo control, como también a medida que aumenta el tiempo de exposición (p<0,05). Por otra parte, también existen diferencias significativas (p<0,05) en la expresión de PARP- 1 en los grupos sometidos a Biodentine. Los resultados obtenidos en este estudio demuestran que Biodentine genera citotoxicidad celular en cultivos celulares de pulpa dental humana, por disminución de la viabilidad celular como por la expresión de proteínas apoptóticas. Es por esto que la utilización de este biomaterial debería ser estudiado y considerarse en cada caso clínico, especialmente como recubridor pulpar directo.


ABSTRACT: Oral pathologies have been a real challenge for dentistry, mainly due to its treatment. Currently, there are numerous biomaterials on the market that may present inherent properties in dental tissues. However, studies on multiple cell lines are based on biocompatible results such as biocompatibility and cellular cytotoxicity. Biodentine is endodontic cement based on calcium silicates of multiple applications, which would provide biocompatibility properties as cellular bioactivity, characteristics that will allow it to be used in direct contact with the dental pulp. The objective of this study is the in vitro evaluation of Biodentine, on cultures of cells of the human dental pulp (HDPC). Biodentine cement disks of 2 x 6 mm were prepared, and HDPC culture plates were introduced. After 24, 48 and 72 hours of exposure, cell viability tests were performed using the MTT colorimetric method. On the other hand, protein expression assays of two proteins involved in the signaling pathway of cell apoptosis Caspase-3 cleaved (cas-3 clv) and PARP-1 are carried out. There are statistically significant differences (p <0,05) in the cell viability tests between Biodentine and control group, as well as the exposure time increases (p <0,05). Otherwise, there are also significant differences (p <0,05) in the expression of PARP-1 in the groups, sometimes a Biodentine. The results in this study that Biodentine generates a cellular cytotoxicity in HDPC cultures, therefore, cell viability as the expression of apoptotic proteins. This is why the use of this biomaterial should be studied for each particular clinical case, especially as a direct pulp capping agent.


Subject(s)
Humans , Apoptosis , Calcium Compounds/chemistry , Caspase 3/analysis , Poly (ADP-Ribose) Polymerase-1 , Stem Cells/physiology , In Vitro Techniques , Cell Survival , Silicates/chemistry , Dental Pulp/anatomy & histology , Dentin/pathology , Antibody-Dependent Cell Cytotoxicity
10.
Arq. gastroenterol ; 56(4): 405-411, Oct.-Dec. 2019. graf
Article in English | LILACS | ID: biblio-1055165

ABSTRACT

ABSTRACT BACKGROUND: Serotonin (5-HT) is present in the epithelial enterochromaffin cells (EC), mast cells of the lamina propria and enteric neurons. The 5-HT is involved in regulating motility, secretion, gut sensation, immune system and inflammation. OBJECTIVE: Evaluate the effects of diabetes and quercetin supplementation on serotoninergic cells and its cell loss by apoptosis in jejunal mucosa of streptozotocin-induced diabetic rats (STZ-rats). METHODS: Twenty-four male Wistar rats were divided into four groups: normoglycemic (C), normoglycemic supplemented with 40 mg/day quercetin (Q), diabetic (D) and diabetic supplemented with 40 mg/day quercetin (DQ). After 120 days, the jejunum was collected and fixated in Zamboni's solution for 18 h. After obtaining cryosections, immunohistochemistry was performed to label 5-HT and caspase-3. Quantification of 5-HT and caspase-3 immunoreactive (IR) cells in the lamina propria, villi and crypts were performed. RESULTS: The diabetic condition displayed an increase of the number of 5-HT-IR cells in villi and crypts, while decreased number of these cells was observed in lamina propria in the jejunum of STZ-rats. In the diabetic animals, an increased density of apoptotic cells in epithelial villi and crypts of the jejunum was observed, whereas a decreased number of caspase-3-IR cells was observed in lamina propria. Possibly, quercetin supplementation slightly suppressed the apoptosis phenomena in the epithelial villi and crypts of the STZ-rats, however the opposite effect was observed on the 5-HT-IR cells of the lamina propria. Quercetin supplementation on healthy animals promoted few changes of serotoninergic function and apoptotic stimuli. CONCLUSION: These results suggest that quercetin supplementation mostly improved the serotonergic function affected by diabetes maybe due to antioxidant and anti-inflammatory properties of quercetin.


RESUMO CONTEXTO: A serotonina (5-HT) está presente nas células epiteliais enterocromafins (CE), nos mastócitos da lâmina própria e nos neurônios entéricos. A 5-HT está envolvida na regulação da motilidade, secreção, nocepção intestinal, sistema imunológico e inflamação. Objetivo: Avaliar os efeitos do diabetes e da suplementação de quercetina sobre a função serotoninérgica e a perda celular por apoptose na mucosa jejunal de ratos diabéticos induzidos por estreptozotocina (ratos STZ). MÉTODOS: Vinte e quatro ratos Wistar machos foram divididos em quatro grupos: normoglicêmico (C), normoglicêmico suplementado com quercetina 40 mg/dia (Q), diabético (D) e diabético suplementado com quercetina 40 mg/dia (DQ). Após 120 dias, o jejuno foi coletado e fixado na solução de Zamboni por 18 horas. Após a obtenção de cortes em criostato, a imuno-histoquímica foi realizada para marcar 5-HT e caspase-3. A quantificação de células imunorreativas (IR) à 5-HT e caspase-3 foram realizadas na lâmina própria, vilosidades e criptas. RESULTADOS: A condição diabética ocasionou um aumento do número de células 5-HT-IR nas vilosidades e criptas, enquanto que na lâmina própria houve uma redução dessas células, no jejuno de ratos STZ. Nos animais diabéticos, foi observada uma densidade aumentada de células apoptóticas no epitélio do jejuno, tanto nas vilosidades quanto nas criptas, por outro lado um número reduzido de células caspase-3-IR foi observado na lâmina própria. Possivelmente, a suplementação de quercetina suprimiu ligeiramente os fenômenos de apoptose no epitélio de vilosidades e criptas do jejuno de ratos STZ, no entanto, o efeito oposto foi observado nas células 5-HT-IR da lâmina própria. A suplementação com quercetina em animais saudáveis promoveu poucas alterações na função serotoninérgica e nos estímulos apoptóticos. CONCLUSÃO: Estes resultados sugerem que a suplementação de quercetina melhorou principalmente a função serotoninérgica afetada pelo diabetes, talvez devido às propriedades antioxidantes e anti-inflamatórias da quercetina.


Subject(s)
Animals , Male , Rats , Quercetin/administration & dosage , Serotonin/metabolism , Apoptosis/drug effects , Dietary Supplements , Diabetes Mellitus, Experimental/drug therapy , Caspase 3/metabolism , Jejunum/pathology , Antioxidants/administration & dosage , Immunohistochemistry , Rats, Wistar , Diabetes Mellitus, Experimental/pathology , Interstitial Cells of Cajal/drug effects , Interstitial Cells of Cajal/pathology , Intestinal Mucosa/drug effects , Jejunum/drug effects
11.
Rev. Assoc. Med. Bras. (1992) ; 65(9): 1193-1200, Sept. 2019. graf
Article in English | LILACS | ID: biblio-1041079

ABSTRACT

SUMMARY OBJECTIVES This study was conducted to reveal the possible protective effects of ticagrelor and enoxaparin pretreatment against ischemia-reperfusion (IR)-induced injury on the lung tissue of a rat model. METHODS Wistar albino rats were randomly divided into 4 groups as follows: group-1 (control-sham), group-2 (control-saline+IR), group-3 (ticagrelor+IR), group-4 (enoxaparin+IR). Before the ischemic period, saline, ticagrelor, and enoxaparin were administered to the 2nd-4th groups, respectively. In these groups, IR injury was induced by clamping the aorta infrarenally for 2 h, followed by 4 h of reperfusion except group-1. After the rats were euthanized, the lungs were processed for histological examinations. Paraffin sections were stained with Haematoxylin&Eosin (H&E) for light microscopic observation. Apoptosis was evaluated by caspase-3 immunoreactivity. Data were statistically analyzed using the SPSS software. RESULTS In the lung sections stained with H&E, a normal histological structure was observed in group-1, whereas disorganized epithelial cells, hemorrhage, and inflammatory cell infiltration were seen in the alveolar wall in group-2. The histologic structure of the treatment groups was better than that of group-2. Caspase-3(+) apoptotic cells were noticeable in sections of group-2 and were lower in the treatment groups. In group-4, caspase-3 immunostaining was lower than in group-3. In group-2, apoptotic cells were significantly higher than in the other groups (p<0.001). CONCLUSION Based on the histological results, we suggested that both therapies ameliorated the detrimental effects of IR. Caspase-3 immunohistochemistry results also revealed that pre-treatment with enoxaparin gave better results in an IR-induced rat injury model. In further studies, other parameters such as ROS and inflammatory gene expressions should be evaluated for accurate results.


RESUMO OBJETIVOS Este estudo foi realizado para revelar os possíveis efeitos protetores do ticagrelor e do pré-tratamento da enoxaparina no tecido pulmonar contra o modelo de lesão induzida por isquemia-reperfusão (IR). MÉTODOS Ratos albinos Wistar foram randomizados e divididos em quatro grupos: grupo 1 (controle-sham), grupo 2 (controle-salina + IR), grupo 3 (ticagrelor + IR), grupo 4 (enoxaparina + IR). Antes do período isquêmico, salina, ticagrelor e enoxaparina foram administrados nos grupos 2-4, respectivamente. Nesses grupos, a lesão de IR foi induzida pelo clampeamento da aorta na região da infrarrenal por duas horas, seguida por quatro horas de reperfusão, exceto no grupo 1. Após a sacrificação, os pulmões foram processados para exames histológicos. Secções de parafina foram coradas com hematoxilina e eosina (H&E) para observação microscópica de luz. A apoptose foi avaliada pela imunorreatividade da caspase-3. Os dados foram analisados estatisticamente pelo programa SPSS. RESULTADOS Nas secções pulmonares coradas com H&E, estrutura histológica normal foi observada no grupo 1, enquanto células epiteliais desorganizadas, hemorragia e infiltração de células inflamatórias foram observadas na parede alveolar no grupo 2. A estrutura histológica dos grupos de tratamento foi melhor que o grupo 2. Células apoptóticas caspase-3 (+) foram notadas em secções do grupo 2, e essas células foram mais baixas nos grupos de tratamento. No grupo 4, a imunocoloração com caspase-3 foi menor que no grupo 3. No grupo 2, as células apoptóticas foram significativamente maiores que nos outros grupos (p<0,001). CONCLUSÃO Com base nos resultados histológicos, sugerimos que ambas as terapias atenuaram os efeitos prejudiciais da RI. Resultados de imuno-histoquímica com caspase-3 também revelaram que o pré-tratamento com enoxaparina proporcionou melhores resultados no modelo de lesão induzida por IR. Em estudos posteriores, outros parâmetros, como ROS e expressões gênicas inflamatórias, devem ser avaliados quanto a resultados precisos.


Subject(s)
Animals , Male , Aorta, Abdominal/surgery , Reperfusion Injury/prevention & control , Enoxaparin/pharmacology , Protective Agents/pharmacology , Ticagrelor/pharmacology , Lung/drug effects , Reperfusion Injury/pathology , Random Allocation , Rats, Wistar , Apoptosis/drug effects , Disease Models, Animal , Caspase 3/metabolism , Lung Injury/prevention & control , Lung/pathology
12.
Arq. bras. oftalmol ; 82(4): 322-328, July-Aug. 2019. tab, graf
Article in English | LILACS | ID: biblio-1019415

ABSTRACT

ABSTRACT PURPOSE: We examined the effect of intracameral administration of cefuroxime on oxidative stress and endothelial apoptosis in rat corneal tissue. METHODS: In total, 30 rats were divided into 3 groups of 10 rats each (intracameral administration of cefuroxime 0.1 mg/0.01 mL (cefuroxime group); intracameral administration of balanced salt solution 0.01 mL (control group); or absence of intracameral injection (sham group). Corneal endothelial apoptosis was assessed by immunohistochemical analysis using caspase-3 and caspase-8. Total oxidant status, total antioxidant status, oxidative stress index, and paraoxonase and arylesterase levels were examined in corneal endothelial tissue and serum. RESULTS: Paraoxonase levels in the serum were significantly different between the sham and cefuroxime groups (p=0.027). A significant difference was also observed in total oxidant status levels between the cefuroxime and balanced salt solution groups (p=0.023). In addition, there were significant differences in total antioxidant status levels in corneal tissue between the cefuroxime and sham groups (p<0.001) and between the cefuroxime and balanced salt solution groups (p<0.001). Furthermore, significant differences were also observed in oxidative stress index levels between the cefuroxime and balanced salt solution groups (p=0.001) and between the cefuroxime and sham groups (p=0.026). According to the immunohistochemical staining results, a significant association with caspase-3 activity existed between the cefuroxime and balanced salt solution groups (p=0.007), while no significant difference was found with caspase-8 activity (p=0.541). Caspase-3 activity exhibited a significant relationship between the sham and balanced salt solution groups (p=0.018), but no relationship was found with caspase-8 activity (p=0.623). CONCLUSION: Immunohistochemical examination revealed that intracameral cefuroxime increased apoptosis when compared to the sham and balanced salt solution groups. Moreover, intracameral cefuroxime increased oxidative stress in the cornea and simultaneously induced apoptosis.


RESUMO OBJETIVO: Examinamos o efeito da administração intracameral da cefuroxima sobre o estresse oxidativo e a apoptose endotelial no tecido corneano de ratos. MÉTODOS: No total, 30 ratos foram divididos em 3 grupos de 10 ratos cada (administração intracameral de cefuroxima 0,1 mg/0,01 mL (grupo cefuroxima), administração intracameral de solução salina balanceada 0,01 mL (grupo controle) ou ausência de injeção intracameral (grupo sham)). A apoptose endotelial da córnea foi avaliada por análise imuno-histoquimica usando caspase-3 e -8. O status oxidante total, o status antioxidante total, o índice de estresse oxidativo e os níveis de a paraoxonase e arilesterase foram investigados no tecido endotelial da córnea e no soro. RESULTADOS: Os níveis de paraoxonase no soro foram significativamente diferentes entre os grupos sham e cefuroxima (p=0,027). Foi também observada uma diferença significativa nos níveis de estado oxidante total entre os grupos cefuroxima e solução salina balanceada (p=0,023). Além disso, houve diferenças significativas nos níveis de status antioxidante total no tecido da córnea entre os grupos cefuroxima e sham (p<0,001) e entre os grupos cefuroxima e solução salina balanceada (p<0,001). Diferenças significativas também foram observadas nos níveis do índice de estresse oxidativo entre os grupos cefuroxima e solução salina balanceada (p=0,001) e entre os grupos cefuroxima e sham (p=0,026). De acordo com os resultados de coloração imuno-histoquimica, houve associação significativa com a atividade da caspase-3 entre os grupos cefuroxima e solução salina balanceada (p=0,007), enquanto não houve diferença significativa com a atividade da caspase-8 (p=0,541). A atividade da caspase-3 exibiu uma relação significativa entre os grupos sham e solução salina balanceada (p=0,018), mas nenhuma relação foi encontrada com a atividade da caspase-8 (p=0,623). CONCLUSÃO: O exame imuno-histoquímico revelou que a cefuroxima intracameral aumentou a apoptose quando comparada com os grupos sham e solução salina balanceada. Além disso, a cefuroxima intracameral aumentou o estresse oxidativo na córnea e induziu simultaneamente a apoptose.


Subject(s)
Animals , Male , Cefuroxime/pharmacology , Apoptosis/drug effects , Oxidative Stress/drug effects , Cornea/drug effects , Cornea/metabolism , Anti-Bacterial Agents/pharmacology , Endothelium, Corneal/drug effects , Endothelium, Corneal/metabolism , Endothelium, Corneal/pathology , Immunohistochemistry , Carboxylic Ester Hydrolases/analysis , Reproducibility of Results , Oxidants/blood , Rats, Wistar , Cornea/pathology , Aryldialkylphosphatase/analysis , Caspase 3/analysis , Caspase 8/analysis , Injections, Intraocular
13.
Int. j. morphol ; 37(2): 515-521, June 2019. tab, graf
Article in English | LILACS | ID: biblio-1002253

ABSTRACT

SUMMARY: Reproductive dysfunction is a complication for many diseases and toxins. Its early diagnosis and treatment are immensely important. Here the morphological histoarchitecture changes in early testicular and cauda toxicity before and after treatment with angiotensin receptor blockers were evaluated. Low-grade testicular damage was induced using thioacetamide (TAA, 50 mg/kg/day) intraperitoneally for two weeks in rats. The rats were randomly divided into four groups (n = 8) treated daily orally for three weeks as follows: Normal control (distilled water), TAA (positive control), TAA+candesartan (0.2 mg/kg) and TAA+losartan (7.5 mg/kg). Serum testosterone and testicular malondialdehyde and glutathione were measured. The changes in histoarchitecture of testis and cauda epididymis were evaluated by hematoxylin and eosin for general structure, Masson's trichrome for collagen, periodic acid Schiff for basement membrane, and caspase-3 and proliferating cell nuclear antigen (PCNA) for immunohistochemical analysis. The TAA-rats showed decreases of serum testosterone and testicular glutathione, increases in testicular malondialdehyde, degenerative changes and apoptosis in germ cells, thickening of tubular basal lamina and increases in expression of caspase 3, and decreases in expression of PCNA. The ARBs (candesartan and losartan) significantly reversed these changes with non-significant differences in-between. Treatment with ARBs (candesartan and losartan) significantly reversed TAA-induced low-grade testicular and cauda toxicity in rats. This could be potentially useful for early treatment of male patients with occupational toxicant-induced reproductive dysfunction especially if they are using ARBs for other comorbidities.


RESUMEN: La disfunción reproductiva es una complicación por muchas enfermedades y toxinas. Su diagnóstico y tratamiento tempranos son inmensamente importantes. Aquí se evaluaron los cambios morfológicos en la histoarquitectura en la toxicidad precoz testicular y cauda antes y después del tratamiento con bloqueadores de receptores de angiotensina. Se indujo daño testicular de bajo grado usando tioacetamida (TAA, 50 mg / kg / día) por vía intraperitoneal durante dos semanas en ratas. Las ratas se dividieron aleatoriamente en cuatro grupos (n = 8) tratados diariamente por vía oral durante tres semanas de la siguiente manera: control normal (agua destilada), TAA (control positivo), TAA + candesartan (0,2 mg / kg) y TAA + losartán (7,5 mg / kg). Se midieron la testosterona sérica, el malondialdehído testicular y el glutatión. Los cambios en la histoarquitectura de los testículos y la epidermis de la cauda se evaluaron mediante Hematoxilina y Eosina para determinar la estructura general, con tricrómicro de Masson para el colágeno, ácido periódico de Schiff para la membrana basal y la caspasa-3 y el antígeno nuclear de células proliferantes (PCNA) para análisis inmunohistoquímico. Las ratas TAA mostraron disminución de la testosterona sérica y glutatión testicular, aumentos en el malondialdehído testicular, cambios degenerativos y apoptosis en células germinales, engrosamiento de la lámina basal tubular y aumentos en la expresión de la caspasa 3, y disminución en la expresión de PCNA. Los ARB (candesartán y losartán) revirtieron significativamente estos cambios con diferencias no significativas en el medio. El tratamiento con BRA (candesartán y losartán) revirtió significativamente la toxicidad testicular y cauda inducida por TAA en ratas. Esto podría ser potencialmente útil para el tratamiento temprano de pacientes con disfunción reproductiva inducida por tóxicos ocupacionales, especialmente si están usando BRA para otras comorbilidades.


Subject(s)
Animals , Male , Rats , Testis/drug effects , Thioacetamide/toxicity , Benzimidazoles/pharmacology , Losartan/pharmacology , Angiotensin II Type 1 Receptor Blockers/pharmacology , Testis/pathology , Testosterone/analysis , Tetrazoles/pharmacology , Immunohistochemistry , Rats, Sprague-Dawley , Proliferating Cell Nuclear Antigen/metabolism , Caspase 3/metabolism , Glutathione/analysis , Malondialdehyde/analysis
14.
Laboratory Animal Research ; : 194-201, 2019.
Article in English | WPRIM | ID: wpr-786403

ABSTRACT

TW-37 is a small molecule B cell lymphoma-2 (Bcl-2) homology 3 mimetic with potential anticancer activities. However, the in vivo anti-cancer effect of TW-37 in human oral cancer has not been properly studied yet. Here, we attempted to confirm antitumor activity of TW37 in human oral cancer. TW-37 significantly inhibited cell proliferation and increased the number of dead cells in MC-3 and HSC-3 human oral cancer cell lines. TW-37 enhanced apoptosis of both cell lines evidenced by annexin V/propidium iodide double staining, sub-G1 population analysis and the detection of cleaved poly (ADP-ribose) polymerase and caspase-3. In addition, TW-37 markedly downregulated the expression of Bcl-2 protein, while not affecting Bcl-xL or myeloid cell leukemia-1. In vivo, TW-37 inhibited tumor growth in a nude mice xenograft model without any significant liver and kidney toxicities. Collectively, these data reveal that TW-37 may be a promising small molecule to inhibit human oral cancer.


Subject(s)
Animals , Apoptosis , Caspase 3 , Cell Line , Cell Proliferation , Heterografts , Humans , Kidney , Liver , Mice , Mice, Nude , Mouth Neoplasms , Myeloid Cells
15.
Laboratory Animal Research ; : 114-123, 2019.
Article in English | WPRIM | ID: wpr-786396

ABSTRACT

In our efforts to understand the systemic features of tumors, the importance of animal models is increasing due to the recent growth in the development of immunotherapy and targeted therapies. This has resulted in increased attention towards tumor animal models using C57BL/6N, which are mainly used in immunological studies. In this study, the C57BL/6NKorl stock and two other commercial stocks (C57BL/6NA and C57BL/N6B) are evaluated by comparing the occurrence of tumors using the syngeneic model; furthermore, we compare the response to anti-cancer drugs in the syngeneic model by evaluating survival, growth of tumors, proliferation and molecular biology analysis. In the syngeneic model using LLC (Lewis lung carcinoma) cells, the survival of mice and growth of the tumor showed a better response in the C57BL/6NKorl stock, and was dependent on the cell concentration of the dosing tumor, as compared to the other C57BL/6N stocks. However, the Ki-67 staining showed only little difference in cell proliferation within the tumor tissue each mouse stocks. Comparing the sensitivity to anti-cancer drug by examining changes in growth, volume and weight revealed that cisplatin treatment for tumor-bearing C57BL/6NKorl was more dependent on concentration. The Ki-67 staining, however, showed no difference among the C57BL/6N stocks after cisplatin treatment. The expressions of p27 and p53 tumor suppressor proteins, caspase-3 and Bax showed dose-dependent increase after exposure to cisplatin, whereas the expression of Bcl-2 was reduced in a dose-dependent manner. Furthermore, the expressions of MMP-2 and VEGF involved in metastasis, as well as inflammatory genes IL-1β, IL-6 and IL-10, showed dose-dependent decrease in tumor tissue after cisplatin exposure. Differences observed among the C57BL/6N stocks were not significant. Taken together, our studies reveal that C57BL/6NKorl has the potential of being a useful biological resource established in Korea, as it does not differ from the two commercially available C57BL/6N stocks when considering response to tumor generation and sensitivity to anti-cancer drugs using the syngeneic tumor model.


Subject(s)
Animals , Caspase 3 , Cell Proliferation , Cisplatin , Immunotherapy , Interleukin-10 , Interleukin-6 , Korea , Lung , Mice , Models, Animal , Molecular Biology , Neoplasm Metastasis , Tumor Suppressor Protein p53 , Vascular Endothelial Growth Factor A
16.
Article in English | WPRIM | ID: wpr-785644

ABSTRACT

OBJECTIVE: This study aimed to examine the effect of vitrification on apoptosis and survival in human preantral follicles after thawing.METHODS: This experimental study was conducted at an acute tertiary care hospital from March 2012 to April 2013. Ovaries were sliced into 5×5×1-mm pieces and divided into the following three groups: preantral follicle isolation, ovarian tissue vitrification-warming followed by follicle isolation, and immunohistochemistry of fresh ovarian tissue. For statistical analyses, the Student t-test, chi-square test, Kruskal-Wallis test, and Kaplan-Meier survival analysis were used.RESULTS: A total of 161 preantral follicles (70% secondary) were collected from ovarian cortex tissue of six women between 30 and 37 years of age who underwent oophorectomy due to cervical cancer or breast cancer. There were no significant differences in the follicular morphology of fresh preantral follicles and vitrified follicles after thawing. The mean Fas ligand (FasL) mRNA expression level was 0.43±0.20 (relative to β-actin) in fresh preantral follicles versus 0.51±0.20 in vitrified follicles (p=0.22). The mean caspase-3 mRNA expression level in fresh preantral follicles was 0.56±0.49 vs. 0.27±0.21 in vitrified follicles (p=0.233). One vitrified-thawed secondary follicle grew and developed to an antral follicle within 6 days of culture.CONCLUSION: Vitrification did not affect preantral follicle morphology or mRNA expression of the apoptosis markers FasL and caspase-3. Further studies are required to establish whether vitrification affects the outcomes of in vitro culture and the maturation of preantral follicles.


Subject(s)
Apoptosis , Breast Neoplasms , Caspase 3 , Fas Ligand Protein , Female , Humans , Immunohistochemistry , In Vitro Techniques , Ovariectomy , Ovary , RNA, Messenger , Tertiary Healthcare , Uterine Cervical Neoplasms , Vitrification
17.
Asian Spine Journal ; : 875-889, 2019.
Article in English | WPRIM | ID: wpr-785500

ABSTRACT

STUDY DESIGN: Development of an in vitro model for assessing the anti-inflammatory efficacies of naringin (Nar) and naringenin (NG).PURPOSE: To evaluate the efficacy of natural flavonoids as therapeutic drugs against anti-inflammatory processes in the nucleus pulposus (NP) cells using in-vitro and in-silico methods.OVERVIEW OF LITERATURE: Intervertebral disc (IVD) disease is a common cause of low back pain. Chronic inflammation and degeneration play a significant role in its etiopathology. Thus, a better understanding of anti-inflammatory agents and their role in IVD degeneration and pro-inflammatory cytokines expression is necessary for pain management and regeneration in IVD.METHODS: We performed primary cell culture of NP cells; immunocytochemistry; gene expression studies of cytokines, metalloproteases, extracellular proteins, and apoptotic markers using quantitative polymerase chain reaction and reverse transcription-polymerase chain reaction (RT-PCR); cytotoxicity assay (MTT); and molecular docking studies using AutoDock 4.2 software (Molecular Graphics Laboratory, La Jolla, CA, USA) to confirm the binding mode of proteins and synthesized complexes. We calculated the mean±standard deviation values and performed analysis of variance and t-test using SPSS ver. 17.0 (SPSS, Inc., Chicago, IL, USA).RESULTS: Molecular docking showed that both Nar and NG bind to the selected genes of interest. Semi-quantitative RT-PCR analysis reveals differential gene expression of collagen (COL)9A1, COL9A2, COL9A3, COL11A2, COMT (catechol-O-methyltransferase), and THBS2 (thrombospondin 2); up regulation of ACAN (aggrecan), COL1A1, COL11A1, interleukin (IL)6, IL10, IL18R1, IL18RAP, metalloprotease (MMP)2, MMP3, MMP9, ADAMTS5 (a disintegrin and metalloproteinase with thrombospondin motifs 5), IGF1R (insulin-like growth factor type 1 receptor), SPARC (secreted protein acidic and cysteine rich), PARK2 (parkin), VDR (vitamin D receptor), and BCL2 (B-cell lymphoma 2); down regulation of IL1A, CASP3 (caspase 3), and nine genes with predetermined concentrations of Nar and NG.CONCLUSIONS: The present study evaluated the anti-inflammatory and regenerative efficiencies of Nar and NG in degenerated human NP cells. Altered gene expressions of cytokines, metalloproteases, extracellular proteins, apoptotic genes were dose responsive. The molecular docking (in silico) studies showed effective binding of these native ligands (Nar and NG) with genes identified as potent inhibitors of inflammation. Thus, these natural flavonoids could serve as anti-inflammatory agents in the treatment of low back pain and sciatica.


Subject(s)
Anti-Inflammatory Agents , Caspase 3 , Collagen , Cysteine , Cytokines , Down-Regulation , Flavonoids , Gene Expression , Humans , Immunohistochemistry , In Vitro Techniques , Inflammation , Interleukin-10 , Interleukins , Intervertebral Disc , Intervertebral Disc Degeneration , Ligands , Low Back Pain , Lymphoma , Metalloproteases , Models, Molecular , Pain Management , Polymerase Chain Reaction , Primary Cell Culture , Regeneration , Sciatica , Thrombospondins , Up-Regulation
18.
Article in English | WPRIM | ID: wpr-787536

ABSTRACT

BACKGROUND/OBJECTIVES: Although anaplastic thyroid carcinoma (ATC) is rare, it is one of the deadliest forms of thyroid cancer. The fatality rate for ATC is high, and the survival rate at one year after diagnosis is <20%. The present study aimed to investigate the anti-tumor activities of paclitaxel, radiation, and tyrosine kinase inhibitor (TKI) combined therapy in anaplastic thyroid cancer cells both in vitro and in vivo and explore its effects on apoptotic cell death pathways.MATERIALS #SPCHAR_X0026; METHODS: ATC cell line was exposed to TKI, lenvatinib in the presence or absence of paclitaxel with radiation, and cell viability was determined by MTT assay. Effects of the combined treatment on cell cycle and intracellular signaling pathways were assessed by flow cytometry and western blot analysis. The ATC cell line xenograft model was used to examine the anti-tumor activity in vivo.RESULTS: Our data revealed that the combined administration of paclitaxel, TKI, and radiation decreased cell viability in ATC cells, and also significantly increased apoptotic cell death in these cells, as demonstrated by the cleavage of caspase-3 and DNA fragmentation. This combination therapy reduced anti-apoptotic factor levels in ATC cells, while significantly decreasing tumor volume and increasing survival in ATC xenografts.CONCLUSION: These results indicate that administering the combination of paclitaxel, TKI, and radiation therapy may exert significant anticancer effects in preclinical models, potentially suggesting a new clinical approach for treating patients with ATC.


Subject(s)
Blotting, Western , Caspase 3 , Cell Cycle , Cell Death , Cell Line , Cell Survival , Diagnosis , DNA Fragmentation , Flow Cytometry , Heterografts , Humans , In Vitro Techniques , Paclitaxel , Protein-Tyrosine Kinases , Survival Rate , Thyroid Carcinoma, Anaplastic , Thyroid Neoplasms , Tumor Burden
19.
Article in Chinese | WPRIM | ID: wpr-772118

ABSTRACT

OBJECTIVE@#To investigate the effects of millimeter wave (MMW) exposure on apoptosis of human melanoma A375 cells and explore the mechanisms.@*METHODS@#Through electromagnetic field calculation we simulated MMW exposure in cells and calculated the specific absorption rate (SAR). The optimal irradiation parameters were determined according to the uniformity and intensity of the SAR. A375 cells were then exposed to MMV for 15, 30, 60, or 90 min, with or without pretreatment with the caspase-3 inhibitor AC-DEVD-fmk (10 μmol/L) for 1 h at 90 min before the exposure. CCK-8 assay was used to assess the changes in the viability and Annexin-V/ PI staining was used to detect the apoptosis of the cells following the exposures; Western blotting was used to detect the expression of caspase-3 in the cells.@*RESULTS@#The results of electromagnetic field calculation showed that for optimal MMV exposure, the incident field needed to be perpendicular to the bottom of the plastic Petri dish with the antenna placed below the dish. CCk-8 assay showed that MMW exposure significantly inhibited the cell viability in a time-dependent manner ( < 0.05); exposures for 15, 30, 60, and 90 min all resulted in significantly increased apoptosis of the cells ( < 0.05). The cells with MMW exposure showed significantly increased expression of caspase-3. The inhibitory effect of MMW on the cell viability was antagonized significantly by pretreatment of the cells with AC-DEVD-fmk ( < 0.05), which increased the cell viability rate from (36.7±0.09)% to (59.8±0.06)% ( < 0.05).@*CONCLUSIONS@#35.2 GHz millimeter wave irradiation induces apoptosis in A375 cells by activating the caspase-3 protein.


Subject(s)
Apoptosis , Caspase 3 , Metabolism , Caspase Inhibitors , Pharmacology , Cell Line, Tumor , Cell Survival , Electromagnetic Fields , Enzyme Activation , Humans , Magnetic Field Therapy , Melanoma , Pathology , Therapeutics , Time Factors
20.
Article in Chinese | WPRIM | ID: wpr-774366

ABSTRACT

OBJECTIVE@#To investigate the mechanism of Paris forrestii (Takht.) H. Li (PCT3)-suppressing the proliferation of HL-60, K562, KG-1 and HT-93 cells.@*METHODS@#cute myeloid leukemia cell lines such as HL-60, K562, KG-1 and HT-93 were treated with Paris forrestii (Takht.) H. Li (PCT3) for 24, 48, and 72 h, and MTT assay was employed to determine the cells proliferation. Meanwhile, the apoptosis of K562, HL-60, KG-1 and HT-93 cells were detected by flow cytometry after PCT3 (Control, 4 μg/ml, 8 μg/ml) treated for 24 h and the Western blot was performed to detect the expression of PARP,Caspase-3, MCL-1, BAX, BCL-2, P53, and P27. GAPDH was used as an internal loading control.@*RESULTS@#MTT assay showed that Paris forrestii (Takht.) H. Li (PCT3) significantly inhibited the proliferation of HL-60, K562, KG-1 and HT-93 cells in concentration and time-dependent manners. Compared with the control group, the leukemia cell viabilities were significantly suppressed (r =0.9436; r =0.8623; r =0.9922; r =0.8918). Paris forrestii (Takht.) H. Li (PCT3) induced apoptosis of leukemia cells in a concentration dependent manner, compared with the control group (P<0.05 or P<0.01). Western blot revealed that PARP, a major enzyme in DNA damage repair, and Caspase-3 another one of the major executive apoptotic enzymes were cleaved in cell lines examined, and this cleavage was concentration dependent. Anti-apoptotic proteins such as MCL-1 and BCL-2 were down regulated by Paris forrestii (Takht.) H. Li (PCT3), and Pro-apoptotic protein BAX was upregulated. And the protein of tumor suppressor gene P53 and its downstream signaling protein P27 increased.@*CONCLUSION@#Paris forrestii (Takht.) H. Li (PCT3) can inhibit the proliferation of leukemia cells by activating endogenous apoptosis pathway, and provide a potential new drug selection for clinical treatment of AML leukemia.


Subject(s)
Apoptosis , Caspase 3 , Cell Line, Tumor , Cell Proliferation , Humans , Leukemia, Myeloid, Acute , Melanthiaceae , Proto-Oncogene Proteins c-bcl-2
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