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1.
Article in Chinese | WPRIM | ID: wpr-940454

ABSTRACT

ObjectiveTo observe the effects of Hedysarum polysaccharides(HPS)on the signaling pathways of B-cell lymphoma 2 (Bcl-2), cysteinyl aspartate-specific protease 3 (Caspase-3), and Bcl-2-associated X protein (Bax) in Schwann cells(SCs)cultured in high glucose,and explore the possible mechanism of HPS against diabetic peripheral neuropathy(DPN). MethodFour SD suckling mice aged 5-7 days were randomly divided into a normal group,a high-glucose group,an HPS + high-glucose group,and an α-lipoic acid(α-LA)+ high-glucose group. SCs were extracted from the sciatic nerve and cultured in a 37 ℃,5% CO2 incubator. After the cells reached 80% confluence,Cell Counting Kit-8(CCK-8)was used to screen the experimental concentrations suitable for high glucose,HPS, and α-LA interventions. Western blot and Real-time polymerase chain reaction (Real-time PCR)were used to detect the protein and mRNA expression of Bcl-2,Bax,and Caspase-3. The apoptosis rate of SCs was detected by flow cytometry using Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI). ResultAs revealed by Western blot and real-time PCR,compared with the normal group,the high-glucose group showed reduced protein and mRNA expression of Bcl-2 and increased protein and mRNA expression of Bax and Caspase-3(P<0.01). Compared with the high-glucose group,the HPS + high-glucose group and the α-LA + high-glucose group showed increased protein and mRNA expression of Bcl-2 and decreased protein and mRNA expression of Bax and Caspase-3(P<0.01). As displayed by the results of flow cytometry using Annexin V/PI, compared with the normal group,the high-glucose group showed increased apoptosis rate;compared with the high-glucose group,the HPS + high-glucose group and the α-LA + high-glucose group showed reduced apoptosis rate(P<0.01). ConclusionHPS can alleviate the apoptotic response of SCs,and its mechanism may be related to the inhibition of the activation of the Bcl-2/Caspase-3 signaling pathway.

2.
International Eye Science ; (12): 1257-1261, 2022.
Article in Chinese | WPRIM | ID: wpr-934994

ABSTRACT

AIM:To investigate the effect of epigallocatechin gallate(EGCG)on the apoptosis of human retinal pigment epithelium(ARPE-19)cells and its mechanism. METHODS:The ARPE-19 cells were cultured in vitro and treated with 0,40,80 and 160 μg/mL EGCG, respectively. At the proposed time of treatment the morphological changes were detected by hoechst 33258 staining. The apoptosis rate was detected by flow cytometry. The expression of apoptosis-related factors B lymphocytoma-2 gene(bcl-2), BCL2-Associated X protein(Bax),caspase-3 and p53 were detected by quantitative RT-PCR and Western blotting.RESULTS: Hoechst 33258 staining showed that the ARPE-19 cells with the increase of EGCG drug concentration, the number of apoptotic cells gradually increased and the apoptotic bodies were observed. Flow cytometry showed that the apoptosis rate increased gradually with the increase of EGCG drug concentration. The apoptosis rates at 40, 80 and 160 μg/mL were 4.95%±0.071%, 11.75%±0.075% and 21.25%±0.919% respectively, which was significantly different compared with the control group(2.8%±1.556%)(P<0.01), presented with a drug concentration-dependent. The results of quantitative PCR and Western blotting showed that EGCG could significantly up-regulate the expression of apoptosis-promoting factors Bax, caspase-3 and the mRNA and protein expression of p53, and down-regulate the apoptosis-inhibiting factor bcl-2, all of these showed concentration-dependent effects.CONCLUSION:EGCG can obviously induce the apoptosis of ARPE-19 cells. The mechanism is related with the inhibition of bcl-2 and increase the expression of Bax, caspase-3 and p53.

3.
Chinese Journal of Biotechnology ; (12): 1025-1038, 2022.
Article in Chinese | WPRIM | ID: wpr-927760

ABSTRACT

In order to explore the antitumor effect and mechanism of different extracts of cultivated Phellinus vaninii fruit body on H22 tumor bearing mice, 150 ICR mice were randomly divided into blank group, model group, CTX group, P. vaninii water extract group, ethanol extract group, petroleum ether extract group and crude polysaccharide group. H22 liver cancer cells were used to establish a solid tumor model and the mice were sacrificed on the 10th day after administration. The spleen and thymus organ index and tumor inhibition rate were calculated, the serum levels of TNF-α, INF-γ, VEGF, and hematoxylin-eosin were detected, and the immunohistochemical staining method was used to observe the pathological changes of tumor tissues, while Western blotting was used to detect the expression of tumor-related proteins. The high-dose petroleum ether extract group showed the best tumor inhibition rate (73.21%), increased serum levels of TNF-α, IFN-γ, and VEGF, as well as significantly promoted tumor necrosis and ablation. The immunohistochemistry of the water extract group showed negative regulation, indicating an insignificant tumor suppression. Western blotting showed the apoptosis genes Caspase-3, Caspase-9 and pathway genes NF-κB and JAK were all highly expressed in each administration group compared with the model group, and their expression levels gradually decreased with increasing doses. In summary, the petroleum ether extract of P. vaninii fruit body showed a significant anti-tumor effect which is presumably mediated through the mitochondrial pathway. The metabolism of drug in the body induces activation of Caspase-3 and Caspase-9 apoptotic proteins by Bax, Bcl-2, and TNF, which further caused nuclear chromatin or DNA to condense or degrade, and subsequently destroy the normal proliferation of tumor cells, thereby inducing their apoptosis and inhibiting tumor growth.


Subject(s)
Animals , Apoptosis , Basidiomycota , Mice , Mice, Inbred ICR , Neoplasms/metabolism
4.
Biol. Res ; 552022.
Article in English | LILACS-Express | LILACS | ID: biblio-1383906

ABSTRACT

Abstract Background: Chinese hamster ovary cell line has been used routinely as a bioproduction factory of numerous biopharmaceuticals. So far, various engineering strategies have been recruited to improve the production efficiency of this cell line such as apoptosis engineering. Previously, it is reported that the caspase-7 deficiency in CHO cells reduces the cell proliferation rate. But the effect of this reduction on the CHO cell productivity remained unclear. Hence, in the study at hand the effect of caspase-7 deficiency was assessed on the cell growth, viability and protein expression. In addition, the enzymatic activity of caspase-3 was investigated in the absence of caspase-7. Results: Findings showed that in the absence of caspase-7, both cell growth and cell viability were decreased. Cell cycle analysis illustrated that the CHO knockout (CHO-KO) cells experienced a cell cycle arrest in G2/M phase. This cell cycle arrest resulted in a 1.7-fold increase in the expression of luciferase in CHO-KO cells compared to parenteral cells. Furthermore, in the apoptotic situation the enzymatic activity of caspase-3 in CHO-KO cells was approximately 3 times more than CHO-K1 cells. Conclusions: These findings represented that; however, caspase-7 deficiency reduces the cell proliferation rate but the resulted cell cycle arrest leads to the enhancement of recombinant protein expression. Moreover, increasing in the caspase-3 enzymatic activity compensates the absence of caspase-7 in the caspase cascade of apoptosis.

5.
Int. j. morphol ; 39(1)feb. 2021.
Article in English | LILACS-Express | LILACS | ID: biblio-1385291

ABSTRACT

SUMMARY: We aimed to investigate the possible protective effects of Potentilla fulgens on kidney tissue with ischemia- reperfusion using immunohistochemical methods. Wistar rats were grouped as sham, ischemia, ischemia-reperfusion (I/R) and I/R treated with Potentilla fulgens. Renal vessels of the left rat kidney were clamped for 60 minutes for ischemia, IR group had 6 h of reperfusion. 400 mg/kg Potentilla fulgens were given intraperitoneally 5 days before ischemia+reperfusion procedure. Biochemical analysis (MDA, GSH and MPO) of samples were performed. Kidney tissues were fixed with 10 % neutral formalin and routine paraffin tissue follow-up protocol was applied, stained with routine Hematoxylin and Eosin. ADAMTS-5 and Caspase-3 immunostaining was applied for immunohistochemistry and examined under a light microscope. In the ischemia group, inflammation and congestion in the vessels and increased ADAMTS-5 expression in glomerular cells and tubule cells were observed. In reperfusion, an increase in degenerative glomerular cells, tubule cells and intertubular connective tissue and inflammatory cells ADAMTS-5 expression was observed. In the P. fulgens group, degeneration and inflammation decreased and positive ADAMTS-5 expression was observed. In the ischemia and ischemia reperfusion group, increased apoptotic appearance and Caspase-3 positive expression in glomerular and tubular cells, and negative expression in most cells in the P. fulgens group. Potentilla fulgens are thought to stop apoptotic cell development at a certain stage, which affects the cytokine mechanism and plays an important role in the reduction of inflammatory cells and angiogenic regulation.


RESUMEN: El objetivo de este estudio fue investigar los posibles efectos protectores de Potentilla fulgens en el tejido renal con isquemia-reperfusión utilizando métodos inmunohistoquímicos. Se agruparon ratas Wistar como simulación, isquemia, isquemia-reperfusión (I / R) e I / R tratadas con Potentilla fulgens. Los vasos renales del riñón iz- quierdo de las ratas se fijaron durante 60 min por isquemia, el grupo de IR tuvo 6 h de reperfusión. Se administraron 400 mg / kg de Potentilla fulgens por vía intraperitoneal 5 días antes del procedimiento de isquemia + reperfusión. Se realizaron análisis bioquímicos (MDA, GSH y MPO) de muestras. Los tejidos renales se fijaron con formalina neutra al 10 % y se aplicó el protocolo de seguimiento de tejido de parafina de rutina y teñido con hematoxilina y eosina. Se aplicó inmunotinción de ADAMTS-5 y Caspasa-3 para inmunohistoquímica y se examinó con un microscopio óptico. En el grupo de isquemia, se observó inflamación y congestión en los vasos y el aumento de la expresión de ADAMTS-5 en células glomerulares y células tubulares. En la reperfusión, se observó un aumento en la expresión de ADAMTS-5 de células glomerulares degenerativas, células tubulares y tejido conjuntivo intertubular y células inflamatorias. En el grupo de Potentilla fulgens, la degeneración y la inflamación disminuyeron y se observó expresión positiva de ADAMTS-5. En el grupo de isquemia y reperfusión de isquemia, aumentó la apariencia apoptótica y expresión positiva de Caspasa-3 en células glomerulares y tubulares, y expresión negativa en la mayoría de las células del grupo de Potentilla fulgens. Se cree que Potentilla fulgens detiene el desarrollo de las células apoptóticas en una determinada etapa, lo que afecta el mecanismo de las citocinas y juega un papel importante en la reducción de las células inflamatorias y la regulación angiogénica.

6.
J. vasc. bras ; 20: e20210040, 2021. tab, graf
Article in Portuguese | LILACS | ID: biblio-1279382

ABSTRACT

Resumo Contexto Estudos demonstraram, por análise histológica e Dopplerfluxométrica, a interferência da isquemia renal unilateral, realizada em algumas cirurgias, sobre o rim contralateral, identificando o fenômeno de kidney-kidney crosstalk. Objetivos Identificar o efeito da isquemia de duas estratégias de oclusão da vasculatura renal esquerda sobre o rim contralateral através do volume de células renais positivas para Caspase 3. Métodos Suínos foram divididos em 2 grupos: A (n = 8), artéria renal esquerda clampeada, e AV (n = 8), artéria e veia renais esquerdas clampeadas. Foi realizado o estudo imuno-histoquímico (anti-Caspase 3), com o material de biópsias coletadas do rim isquêmico e contralateral em 0, 30, 60 e 90 minutos de isquemia, e análise morfométrica, sendo que a média representou o volume de área de Caspase 3 positiva (%). Resultados A análise morfométrica do rim contralateral nos tempos 30, 60 e 90 minutos de isquemia mostrou que a média da área marcada por Caspase 3 foi estatisticamente superior à média do rim isquêmico nos dois grupos: artéria renal clampeada (A) e artéria e veia renais clampeadas (AV). Comparando o rim isquêmico e contralateral nos dois tipos de clampeamento, não houve diferença estatisticamente significante da área marcada por Caspase 3. Conclusões No modelo experimental de isquemia renal unilateral, o rim não isquêmico apresentou dano celular, demonstrado pela expressão da Caspase 3 de forma aguda em decorrência da isquemia contralateral. O tipo de clampeamento do hilo não parece ter influência sobre o volume de área marcada por Caspase 3.


Abstract Background Studies have demonstrated with histological analysis and Doppler flow measurement analysis that unilateral renal ischemia, which is performed in some surgeries, interfered with the contralateral kidney, identifying the phenomenon of kidney-kidney crosstalk. Objectives To identify the effects on the ischemic and contralateral kidney of renal ischemia induced by two types of clamping technique by analyzing the volume of kidney cells positive for Caspase 3. Methods Sixteen pigs were divided into 2 groups, as follows: A (n = 8) - clamping of left renal artery only and AV (n = 8) - clamping of left renal artery and vein. Immunohistochemical analyses (anti Caspase 3) were conducted with biopsy specimens collected from the ischemic and contralateral kidney at 0, 30, 60, and 90 minutes of ischemia and morphometric analysis was performed, taking the mean to represent the volume of the Caspase 3 positive area (%). Results Morphometric analysis of specimens collected at 30, 60, and 90 minutes of ischemia showed that the mean area marked for Caspase 3 was statistically larger in the contralateral kidney than the ischemic kidney in both groups: clamped renal artery (A) and clamped renal artery and vein (AV). Comparing the ischemic and contralateral kidney, there was no statistically significant difference in the area marked for Caspase 3 between the two types of clamping. Conclusions In the experimental model of unilateral renal ischemia, the non-ischemic kidney exhibited cell damage, demonstrated by Caspase 3 expression. The type of hilum clamping does not appear to influence the area marked for Caspase 3.


Subject(s)
Animals , Renal Circulation , Ischemia , Swine , Intervention Studies , Apoptosis , Constriction , Caspase 3
7.
Braz. dent. sci ; 24(4, suppl 1): 1-13, 2021. tab, ilus
Article in English | LILACS, BBO | ID: biblio-1349352

ABSTRACT

Objective: Head and neck cancer is the sixth leading cancer by incidence worldwide and eighth by death. Recent reports revealed that, not only radiotherapy but also chemotherapy may induce xerostomia. The aim of this study was to compare the possible regenerative effect of BMSCs through systemic and local injections. Material and Methods: 52 male Albino rats were randomly divided into 4 groups: Group 1: 10 rats received 0.5 ml of PBS by injection. Group 2: 14 rats received an intraperitoneal injection of 5-FU drug. Group 3: 14 rats were injected the same dose of 5-FU then received an intraglandular transplantation of BMSCs suspended in 0.5 ml of PBS at day 1 after 5-FU administration. Group 4: 14 rats were injected the same dose of 5-FU then received an intravenous injection of BMSCs suspended in 0.5 ml of PBS via the tail vein at day 1 after 5-FU administration. Results: Histological examination showed that group 2 showed features of severe degenerative changes which increased over time. Group 3 showed increasing amelioration in the ductal structure overtime. Group 4 also showed regenerated ductal elements however concerning apoptotic changes, immunohistochemistry results revealed improvement in both group 3 and 4 over group 2 with no statistical difference between groups 3 and 4. Conclusion: Histological and immunohistochemical features in group 3 and group 4 revealed similar amelioration in regenerative potentials. On the other hand, regenerative features of both experimental groups were statistically significant as compared independently to group 1 (AU)


Objetivo: O carcinoma de cabeça e pescoço é o sexto câncer de maior incidência no mundo sendo a oitava causa de morte por cancer. Relatos recentes revelaram que não apenas a radioterapia, mas também a quimioterapia podem induzir xerostomia. O objetivo desse estudo foi comparar a possivel ação regenerative de BMSCs através de injeção local e sistêmica. Material e Métodos: 52 ratos Albino foram aleatoriamente alocados em 4 grupos: Grupo 1: 10 ratos que receberam 0.5 ml de injeção de PBS. Grupo 2: 14 ratos que receberam injeção intraperitoneal da droga 5-FU. Grupo 3: 14 ratos que foram injetados com a mesma dose de 5-FU e receberam transplante intraglandular de BMSCs ressuspendidas em 0.5mL de PBS no dia 1 após a administração do 5-FU. Grupo 4: 14 ratos que foram injetados com a mesma dose de 5-FU e receberam injeção intravenosa de BMSCs ressuspendidas em 0.5mL de PBS via veia caudal 1 dia após a administração de 5-FU. Resultados: O exame histológico demonstrou que o grupo 2 apresentou alterações degenerativas severas que se agravaram com o tempo. O Grupo 3 mostrou melhora da estrutura ductal ao longo do experimento. Group 4 também mostrou elementos ductais regenerados. Referente a alterações apoptóticas,análise imunohistoquimica mostrou melhora nos grupos 3 e 4 comparados ao grupo 2, sendo que os grupos 3 e 4 foram estatisticamente semelhantes. Conclusão: Análises histológicas e imunohistoquímicas mostram que os grupos 3 e 4 apresentam melhora no potencial regenerativo Por outro lado, os resultados observados para os dois grupos foi estatisticamente semlhante quando comparados independentemente ao grupo 1 (AU)


Subject(s)
Animals , Rats , Salivary Glands , Mesenchymal Stem Cells , Squamous Cell Carcinoma of Head and Neck
8.
Article in Chinese | WPRIM | ID: wpr-906124

ABSTRACT

Objective:To reveal the effective components, targets and possible mechanisms of Qinggan Huayu granules in the treatment of non-alcoholic fatty liver disease (NAFLD) and liver cancer based on network pharmacology and experimental verification, and to provide a basis for its rational interpretation of treating different diseases with same method for NAFLD and liver cancer. Method:Based on databases of traditional Chinese medicine and disease, the network pharmacology was used to screen main active compounds and potential targets of Qinggan Huayu granules for NAFLD and liver cancer. STRING 11.0 was used to analyze the interaction between potential targets. The core targets were selected from the interaction targets by cytoHubba plug-in. The gene ontology (GO) function and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed on the target by Metascape database. At the same time, <italic>in vitro</italic> experiments were conducted to validate the effect of kaempferol, one of the main active ingredients of Qinggan Huayu granules, on hepatocellular carcinoma cell model and NAFLD cell model. Result:A total of 43 potential targets of Qinggan Huayu granules for for NAFLD and liver cancer were screened, corresponding to 136 active ingredients in 8 herbal medicines. Through enrichment analysis of potential targets, there were 20 biological processes, 13 molecular functions, 9 cellular components and 15 signaling pathways. Qinggan Huayu granules regulated biological behaviors of tumors related to liver cancer and NAFLD (such as apoptosis inhibition and oxidative stress) mainly through kaempferol, quercetin, luteolin and other active ingredients for Caspase-3 (CASP3), tumor protein p53 (TP53), vascular endothelial growth factor A (VEGFA) and other hub genes. <italic>In vitro</italic> experiments revealed that kaempferol could inhibit cell proliferation in a dose-dependent manner in hepatocellular carcinoma cell model. And kaempferol could modulate the levels of malondialdehyde (MDA) and glutathione peroxidase (GPx), which were the molecular markers of oxidative stress of NAFLD cell model. Kaempfero also regulated the expression level of CASP3 in hepatocellular carcinoma cell model and NAFLD cell model. Conclusion:The main mechanism of Qinggan Huayu granules in treating liver cancer and NAFLD with concept of treating different diseases with same method is related to systematic synergy effect of multiple compounds (represented by quercetin, luteolin and kaempferol), multiple targets (represented by VEGFA, TP53 and CASP3) and multiple signaling pathways (represented by oxidative stress and cell apoptosis).

9.
Article in Chinese | WPRIM | ID: wpr-906052

ABSTRACT

Objective:To investigate the effect of<italic> Stemona tuberosa</italic> alkaloids on the apoptosis of human hepatoma SMMC-7721 cells and the expression of apoptosis-related proteins including B lymphocytoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), and cleaved cysteinyl aspartate-specific protease-3 (cleaved Caspase-3). Method:SMMC-7721 cells were routinely cultured, passaged, and treated with various concentrations (50, 75, 112, 167, and 250 mg·L<sup>-1</sup>) of <italic>S. tuberosa </italic>alkaloids, while those in the blank control group were only treated with 10% fetal bovine serum. The cell proliferation was determined by tetrazolium bromide (MTT) colorimetry and colony assay and the cell apoptosis by Hoechst 33258 staining. The protein expression levels of Bcl-2, Bax, and cleaved Caspase-3 were detected by Western blot. Result:<italic>S. tuberosa</italic> alkaloids inhibited the proliferation of SMMC-7721 cells, and the inhibition rate was significantly increased in comparison with that in the blank control group (<italic>P</italic><0.01), with the half maximal inhibitory concentrations (IC<sub>50</sub>) at 24 h, 48 h, and 72 h being (173.36±8.75), (112.14±16.50), and (96.41±2.60)mg·L<sup>-1</sup>, respectively. The cell colony-inhibitory activity was significantly increased in a dose-dependent manner (<italic>P</italic><0.01). Compared with the blank control group, <italic>S. tuberosa</italic> alkaloids promoted the apoptosis of SMMC-7721 cells, manifested as increased number of apoptotic cells and elevated apoptotic rate (<italic>P</italic><0.01). The typical morphological changes such as brightly blue-fluorescent condensed nuclei, cytoplasmic shrinking, and karyopyknosis were found under the upright fluorescence microscope. As revealed by comparison with the blank control group, the expression of Bcl-2 was significantly down-regulated (<italic>P</italic><0.01), while the protein expression levels of pro-apoptotic protein Bax and cleaved Caspase-3 in the 75, 112, 167, and 250 mg·L<sup>-1</sup> <italic>S. tuberosa</italic> alkaloids groups were significantly up-regulated (<italic>P</italic><0.01). Conclusion:<italic>S. tuberosa </italic>alkaloids inhibit the proliferation of SMMC-7721 cells and promote their apoptosis possibly by inhibiting Bcl-2 protein expression and promoting Bax and cleaved Caspase-3 protein expression.

10.
China Pharmacy ; (12): 1565-1571, 2021.
Article in Chinese | WPRIM | ID: wpr-881456

ABSTRACT

OBJECTIVE:To study the effects of ferulic acid on t he proliferation ,invasion and apoptosis of HepG 2 hepatocelluar carcinoma cells. METHODS :CCK-8 assay was used to screen the concentration of ferulic acid. Western blot assay was adopted to screen the optimal concentration of interleukin 6(IL-6)to induce HepG 2 cell model with high expression of phosphorylated signal transduction protein and activator 3(p-STAT3)protein. HepG 2 cell were divided into blank control group , model group ,ferulic acid group (0.5 mmol/L)and positive control group (p-STAT3 inhibitor C 188-9,10 μmol/L). Except for blank control group ,model group treated with IL- 6,while administration groups were treated with IL- 6 and relevant drugs. Cell survival rate ,invasion and apoptosis rate in early and late stage were detected by CCK- 8 assay,Transwell assay and Annexin V-FITC/PI double staining ,respectively. Western blot assay was used to detect the expression of p-STAT 3,caspase-3,ZBP-89 and vimentin proteins in each group. On the basis of the PDB protein database ,using 1BG1,a highly similar crystal structure of STAT3,as docking template ,using the region around Tyr 705 as the putative binding pocket ,the docking analysis of ferulic acid with STAT 3 protein was carried out. RESULTS :It is selected to use 0.5 mmol/L ferulic acid intervention for 48 h as the follow-up experimental condition ;50 ng/mL IL- 6 was selected as the modeling condition. Compared with blank control group ,the number of cell invasion ,p-STAT3/STAT3 ratio and protein expression of vimentin were increased significantly in model group (P<0.05 or P<0.01),while late apoptosis rate and protein expression 20 of caspase- 3 were decreased significantly (P<0.05 or P< 0.01). Compared with model group ,cell survival rate ,the number of cell invasion ,p-STAT3/STAT3 ratio and protein expression of vimentin were d ecreased significantly in ferulic acid group and positive control group (P<0.05 or P<0.01);early apoptotic rate (except for ferulic acid group ),late apoptotic rate,the protein expression of caspase- 3 and ZBP- 89(except for positive control group )were increased significantly (P<0.05 or P<0.01). The results of molecular docking showed that the carboxylic groups of ferulic acid could interact with 1.9 Å hydrogen bond of Asn 581 and 2.0 Å hydrogen bond of Lys 591,with binding energy of -4.4 kcal/mol. CONCLUSIONS :Ferulic acid may inhibit the activity of p-STAT 3 by directly binding to the phosphorylation site of STAT 3;it may up-regulate the protein expression of caspase- 3 via STAT 3 dependent pathway ,or up-regulate the protein expression of ZBP- 89 via STAT 3 independent pathway and then down-regulate the protein expression of vimentin ,so as to inhibit the proliferation ,invasion and apoptosis of HepG 2 cells.

11.
Article in English | WPRIM | ID: wpr-879951

ABSTRACT

:To investigate the effect of transient receptor potential melastatin 2 (TRPM2) inhibitor A10 on oxygen glucose deprivation/reperfusion (OGD/R) injury in SH-SY5Y cells.:Human neuroblastoma SH-SY5Y cells were subject to OGD/R injury,and then were divided into blank control group,model control group and A10 group randomly. The cell survival rate was detected by cell counting kit 8 (CCK-8); the level of cellular reactive oxygen species (ROS) was detected by reactive oxygen detection kit; the mitochondrial membrane potential was detected by tetramethylrhodamine (TMRM) method; the number of apoptotic cells was detected by TUNEL apoptosis assay kit; the protein expression level of cleaved caspase 3 was detected by Western blot.:Compared with 3,20,30,50, has lower cytotoxicity and better inhibition effect on channel activity. Compared with the model control group,ROS level was reduced,the mitochondrial membrane potential was improved,the number of apoptosis cells was reduced ,and the expression of cleaved caspase 3 was significantly reduced in the A10 group(all <0.05). : A10 can alleviate cell damage after OGD/R by inhibiting TRPM2 channel function,reducing extracellular calcium influx,reducing cell ROS levels,stabilizing mitochondrial membrane potential levels,and reducing apoptosis.


Subject(s)
Apoptosis , Benzeneacetamides , Cell Survival , Glucose , Humans , Oxygen/metabolism , Piperidones , Reactive Oxygen Species/metabolism , Reperfusion , TRPM Cation Channels
12.
Article in English | WPRIM | ID: wpr-922105

ABSTRACT

OBJECTIVE@#To explore the antitumor effects of ethanol extract from Ventilago leiocarpa Benth (EEVLB) on sarcoma 180 (S180) tumor-bearing mice and the potential mechanism.@*METHODS@#Sixty mice were randomly assigned to 6 groups according to a random number table: normal group, model group, 5-fluorouracil (5-FU) group (0.02 g·kg@*RESULTS@#EEVLB with different concentrations achieved inhibition of tumor growth in vivo, wherein the high-dose group showed the most significant reduction in tumor weight and increased apoptosis of tumor cells (P<0.05). In addition, both net weight gain and spleen index of mice showed uptrend in EEVLB treatment groups (P<0.05). Besides, serum levels of IL-2 and IL-6, percentages of CD3@*CONCLUSIONS@#EEVLB exhibits promising antitumor activity in vivo. This effect might be due to activation of apoptotic signaling pathway, increase of cytokine levels and enhancement of immune function in tumor-bearing mice.


Subject(s)
Animals , Cell Line, Tumor , Ethanol , Mice , Plant Extracts/therapeutic use , Rhamnaceae , Sarcoma 180/drug therapy
13.
Article in Chinese | WPRIM | ID: wpr-877377

ABSTRACT

@#目的:探讨成纤维细胞生长因子 13(fibroblast growth factor 13,FGF13)对非小细胞肺癌 A549 细胞活性氧(reactive oxygen species,ROS)的生成和凋亡的影响及其调控机制。方法:WB 法检测 FGF13 在人正常肺上皮细胞 BEAS-2B 和肺癌 A549、H460 细胞中的本底表达量。采用 FGF13 过表达载体转染 BEAS-2B 和 A549 细胞;设计两组靶向 FGF13 的 shRNA 序 列,构建慢病毒干扰载体,包装病毒后侵染 A549 细胞,采用 qPCR 和 WB 法检测干扰效果,DCFH-DA 探针结合荧光酶标仪分 析敲减 FGF13 对 A549 细胞内 ROS 水平的影响,MitoSOX 与 WB 法检测对线粒体 ROS 水平及烟酰胺腺嘌呤二核苷酸磷酸氧 化酶 4(nicotinamide adenine dinucleotide phosphate oxidase 4,NOX4)蛋白表达量的影响,Annexin V-FITC-PI 双染法检测对细胞 凋亡和 Caspase-3 及 Cleaved Caspase-3 蛋白表达的影响。结果:与 BEAS-2B 细胞相比,FGF13 蛋白在两种肺癌细胞中均高表 达(均 P<0.05)。成功构建 FGF13 过表达、低表达的 A549 细胞系。过表达 FGF13 后,BEAS-2B 和 A549 细胞内 ROS 水平显著 降低(P<0.05);敲减 FGF13 表达后,A549 细胞内 ROS 水平显著升高(P<0.05);然而过表达及干扰 FGF13 对 A549 细胞内线粒 体 ROS 水平无显著影响,但 NOX4 蛋白表达量显著下调(P<0.05)及显著上调(P<0.05)。FGF13 干扰后 A549 细胞凋亡率显著 升高(P<0.01),Caspase-3 及 Cleaved Caspase-3 蛋白表达量显著上调(P<0.05)。结论:FGF13 可能通过 NOX 家族途径调控 ROS 的生成,并通过 ROS/Caspase-3 通路调控 A549 细胞凋亡。

14.
Article in Chinese | WPRIM | ID: wpr-876058

ABSTRACT

@#[摘 要] 细胞焦亡(pyroptosis)是新近发现的一种依赖炎性半胱天冬酶(caspase)的促炎程序性细胞死亡方式(regulated cell death,RCD),主要通过gasdermin家族成员发生剪切活化,引起细胞膜穿孔和细胞内容物释放的过程。Gasdermin E(GSDME)是gasdermin家族的主要成员之一,近来研究发现在几种常见的肿瘤中常因其启动子甲基化而低表达,进而增强了肿瘤的增殖和转移能力;分析其结构和功能显示,GSDME可被激活态的caspase-3切割并形成具有造孔活性的GSDME-N末端结构域,从而诱导肿瘤细胞发生焦亡,介导肿瘤细胞死亡。本文就GSDME介导焦亡的机制及其与肿瘤发生发展的关系作一综述,以期提出新的临床肿瘤诊疗方向。

15.
Article in Chinese | WPRIM | ID: wpr-906359

ABSTRACT

Objective:To explore the mechanism of Bushen Huoxue prescription in regulating the related factors in phosphatidylinositol 3-kinase (PI3K)/ protein kinase B (Akt)/ mammalian target of rapamycin (mTOR) pathway and improving ovarian reserve function of rats with diminished ovarian reserve (DOR). Method:Sixty DOR model rats were duplicated by Ataya method (intraperitoneal injection of cyclophosphamide) and then randomized into the model group, estradiol valerate (0.000 9 g·kg<sup>-1</sup>) group, and high- (33 g·kg<sup>-1</sup>), middle- (16.5 g·kg<sup>-1</sup>) and low-dose (8.25 g·kg<sup>-1</sup>) Bushen Huoxue prescription groups, with 12 rats in each group. Another 12 healthy rats were classified into the blank control group. The rats in each group were given the corresponding drugs by gavage, while those in the blank control group and model group received the same volume of normal saline, once per day, for 14 successive days. After treatment, the ovarian tissue was stained with hematoxylin-eosin (HE) for observing the changes in quantities of primary follicles, mature follicles, and total follicles under a light microscope, followed by the detection of vascular endothelial growth factor (VEGF) expression in the ovarian tissue by immunohistochemistry. The protein expression levels of PI3K, Akt, mTOR, and cysteine-dependent aspartate-directed protease-3 (Caspase-3) in the ovarian tissue were assayed by Western blot, whereas the mRNA expression levels of PI3K, Akt, and mTOR were measured by Real-time polymerase chain reaction (Real-time PCR). Result:As revealed by comparison with the blank control group, the quantities of mature follicles and total follicles in the ovarian tissue of model group were significantly reduced (<italic>P</italic><0.05, <italic>P</italic><0.01). The protein expression levels of VEGF and Caspase-3 in the ovarian tissue were increased (<italic>P</italic><0.05), while the protein and mRNA expression levels of PI3K, Akt, and mTOR were decreased (<italic>P</italic><0.05, <italic>P</italic><0.01). Compared with the model group, Bushen Huoxue prescription at the high and middle doses elevated the quantities of mature follicles and total follicles to varying degrees (<italic>P</italic><0.05, <italic>P</italic><0.01), and VEGF increased most significantly in the middle-dose Bushen Huoxue prescription group (<italic>P</italic><0.05). Caspase-3 in the low-, middle-, and high-dose Bushen Huoxue prescription groups and the western medicine group declined. The protein and mRNA expression levels of PI3K, Akt, and mTOR were up-regulated in the middle- and high-dose Bushen Huoxue prescription groups (<italic>P</italic><0.05, <italic>P</italic><0.01), and the levels in the middle-dose Bushen Huoxue prescription group were closer to those in the blank control group. Conclusion:Bushen Huoxue prescription effectively improves the ovarian reserve function of rats with DOR and increases the number of follicles possibly by up-regulating VEGF expression in the ovarian tissue, activating the PI3K/Akt/mTOR signaling pathway, and regulating the content of Caspase-3.

16.
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
17.
J Biosci ; 2020 Feb; : 1-18
Article | IMSEAR | ID: sea-214321

ABSTRACT

Quinacrine (QC), an FDA-approved anti-malarial drug, has shown to have anticancer activities. Due to its‘shotgun’ nature, QC has become an inevitable candidate for combination chemotherapy. There is lack of studyof the molecular interplay between colorectal cancer (CRC) microenvironment and its metastasis. In this study,we focused on the differential anti-cancerous effect of QC on two different human cancer cell lines, HCT 116and INT 407. Results suggest that cytotoxicity increased in both the cell lines with an increase in QCconcentration. The expression patterns of small-GTPases and caspases were altered significantly in QC-treatedcells compared to non-treated cells. HSP70 and p53 showed comparable differences in the expression pattern.The wound-healing assay showed an increase in the denuded zone, with an increase in the concentration ofQC. The formation of apoptotic nuclei increased with a rise in the concentration of QC in both the cell lines.The decrease and increase in caspase 9 and caspase 3 expression respectively were studied, confirmingapoptosis by the extrinsic pathway

18.
Article | IMSEAR | ID: sea-200777

ABSTRACT

Background : Acute promyelocytic leukemia (APL) affects both kids and adults, however it is more prevalent in younger population. Although APL has a favorable prognostic, patients that relapse often do not respond positively to additional chemotherapy. Therefore, there is a need to further identify ways to overcome these challenges. Hypothesis: In this study, we examined antileukemic effects of xanthohumol (XN), a prenylated flav onoid derived from hops ( Humulus lupulus L ), on human promyelocytic HL - 60 cells. Materials and Methods : HL - 60 cells were exposed to different concentrations of XN (?M) for 24 h. Cell viability, cell morphology, chromatin condensation, cPARP - 1 level, and caspase - 3 activation, and the expression of p21 WAF1/Cip1 were analyzed. Results : XN reduced HL - 60 cell viability in a dose - dependent manner. XN induced a dose - dependent morphological changes including cell shrinkage and b lebbing , and significantly increased the number of cells with condensed chromatin. XN significantly increased the level of cPARP - 1, active caspase - 3, and the expression of p21WAF/CIP mRNA. Conclusion : These data indicate that XN induces HL - 60 cell death by regula ting cell cycle progression and apoptosis. This study suggests that XN may have antileukemic preventive effects.

19.
Acta cir. bras ; 35(3): e202000304, 2020. tab, graf
Article in English | LILACS | ID: biblio-1130623

ABSTRACT

Abstract Purpose To investigate the protective effect of rosmarinic acid (RA) in ovarian ischemia/reperfusion injury using biochemical, histopathological, and immunohistochemical methods. Methods Wistar female rats (n = 32) were randomly divided into four groups: control, ischemia, ischemia-reperfusion, and ischemia-reperfusion with RA. Rosmarinic acid was given at a dose of 50 mg/kg by oral gavage three hours after reperfusion. Malondialdehyde (MDA) levels and glutathione peroxidase (GSH-Px) activities were determined in the ovary tissue homogenates for each rat. Results In the ischemia-reperfusion with RA group, the epithelial cells are regularly regulated at the periphery, and the degenerative changes in preantral and antral follicle cells are reduced. Follicle cells and cells in the corpus luteum showed a decrease in vascular endothelial growth factor (VEGF) expression, while VEGF demonstrated a positive reaction in vascular endothelial cells and stromal cells. The TNF-α expression due to the decreased degenerative effect and inflammation was positive in the macrophage cells. The expression of caspase-3 as an apoptosis change was negative in antral follicle cells and granular cells around the antral follicle. Conclusion Different doses of RA may be useful in preventing ischemic damage after vascularization, inflammation, and apoptotic development after ischemia/reperfusion.

20.
Article in Chinese | WPRIM | ID: wpr-849635

ABSTRACT

Objective To investigate the effect of abdominal paracentesis drainage (APD) on pancreatic cell apoptosis in severe acute pancreatitis. Methods Male adult SD rats were randomized into the sham operation (SO) group, SAP group, and APD group, with 18 rats in each group. In the SAP group, 5% sodium sulfonate was pumped into the retrograde pancreatic bile duct to prepare the SAP model. On this basis, a gastric tube was introduced into the right lower abdomen for drainage, namely the APD group. Blood from the abdominal aorta and pancreatic tissues were collected at 6, 12, and 24 h time points in each group. The changes of serum amylase, inflammatory factor, and endotoxin were detected by ELISA. The HE staining was used to evaluate the pancreatic tissue injury. The apoptosis of pancreatic tissue was detected by TUNEL. Western blot and immunohistochemistry were used to detect the expression of apoptosis-related proteins and PI3K/AKT signaling pathway. Results Pancreatic tissue necrosis and edema were significantly lower in the APD group than in the SAP group, and the pathological score was decreased (P<0.05). Serum amylase, TNF-α, IL-1β, IL-6, and endotoxin levels in the APD group were significantly lower than those in the SAP group (P<0.05). The number of pancreatic cell apoptosis in the APD group was significantly higher than that in the SAP group (P<0.05), and the expression levels of pancreatic apoptotic proteins cleaved-caspase-3 and Bax were significantly increased in the APD group, while the expression levels of anti-apoptotic protein Bcl-2 were significantly decreased (P<0.05). Compared with the SAP group, the expression levels of PI3K/AKT signaling pathway key molecules p-PI3K, p-AKT, and NF-kB p65 were significantly decreased in the APD group (P<0.05). Conclusions Our data indicate that APD attenuates the severity of SAP by enhancing cell apoptosis via suppressing PI3K/AKT signaling pathway. This study provides a new theoretical basis for the treatment of severe acute pancreatitis with APD technology.

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