RESUMEN
Precise control of cellular signaling events during programmed cell death is crucial yet challenging for cancer therapy. The modulation of signal transduction in cancer cells holds promise but is limited by the lack of efficient, biocompatible, and spatiotemporally controllable approaches. Here we report a photodynamic strategy that modulates both apoptotic and pyroptotic cell death by altering caspase-3 protein activity and the associated signaling crosstalk. This strategy employs a mitochondria-targeting, near-infrared activatable probe (termed M-TOP) that functions via a type-I photochemical mechanism. M-TOP is less dependent on oxygen and more effective in treating drug-resistant cancer cells, even under hypoxic conditions. Our study shows that higher doses of M-TOP induce pyroptotic cell death via the caspase-3/gasdermin-E pathway, whereas lower doses lead to apoptosis. This photodynamic method is effective across diverse gasdermin-E-expressing cancer cells. Moreover, the M-TOP mediated shift from apoptotic to pyroptotic modulation can evoke a controlled inflammatory response, leading to a robust yet balanced immune reaction. This effectively inhibits both distal tumor growth and postsurgical tumor recurrence. This work demonstrates the feasibility of modulating intracellular signaling through the rational design of photodynamic anticancer drugs.
Asunto(s)
Gasderminas , Neoplasias , Humanos , Caspasa 3/metabolismo , Apoptosis , Transducción de Señal , Mitocondrias/metabolismo , Neoplasias/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Caspasa 1/metabolismo , Caspasa 1/farmacologíaRESUMEN
Propyl gallate (PG) has been found to exert an inhibitory effect on the growth of different cell types, including lung cancer cells. However, little is known about the cytotoxicological effects of PG specifically on normal primary lung cells. The current study examined the cellular effects and cell death resulting from PG treatment in human pulmonary fibroblast (HPF) cells. DNA flow cytometry results demonstrated that PG (100-1,600 µM) had a significant impact on the cell cycle, leading to G1 phase arrest. Notably, 1,600 µM PG slightly increased the number of sub-G1 cells. Additionally, PG (400-1,600 µM) resulted in the initiation of cell death, a process that coincided with a loss of mitochondrial membrane potential (MMP; ΔΨm). This loss of MMP (ΔΨm) was evaluated using a FACS cytometer. In PG-treated HPF cells, inhibitors targeting pan-caspase, caspase-3, caspase-8, and caspase-9 showed no significant impact on the quantity of annexin V-positive and MMP (ΔΨm) loss cells. The administration of siRNA targeting Bax or caspase-3 demonstrated a significant attenuation of PG-induced cell death in HPF cells. However, the use of siRNAs targeting p53, Bcl-2, or caspase-8 did not exhibit any notable effect on cell death. Furthermore, none of the tested MAPK inhibitors, including MEK, c-Jun N-terminal kinase (JNK), and p38, showed any impact on PG-induced cell death or the loss of MMP (ΔΨm) in HPF cells. In conclusion, PG induces G1 phase arrest of the cell cycle and cell death in HPF cells through apoptosis and/or necrosis. The observed HPF cell death is mediated by the modulation of Bax and caspase-3. These findings offer insights into the cytotoxic and molecular effects of PG on normal HPF cells.
Asunto(s)
Glutatión , Galato de Propilo , Humanos , Galato de Propilo/metabolismo , Galato de Propilo/farmacología , Caspasa 8/metabolismo , Caspasa 8/farmacología , Proteína X Asociada a bcl-2/metabolismo , Proteína X Asociada a bcl-2/farmacología , Caspasa 3/metabolismo , Caspasa 3/farmacología , Glutatión/metabolismo , Glutatión/farmacología , Especies Reactivas de Oxígeno/metabolismo , Proliferación Celular , Muerte Celular , Apoptosis , Pulmón , Fibroblastos/metabolismoRESUMEN
BACKGROUND: The use of nanomaterials in cancer diagnosis and treatment has received considerable interest. Preparation of nanoscale complex molecules could be considered to improve the efficacy and minimize toxicity of the product. This work aimed to biosynthesize BiFe2O4@Ag nanocomposite using the Chlorella vulgaris extract and its cytotoxic effect on colon cancer cell line. METHODS: The physicochemical properties of the bioengineered BiFe2O4 @Ag were investigated by Transmission Electron Microscopy (TEM), Field Emission Scanning Electron Microscopy (FE-SEM), Zeta potential, Dynamic Light Scattering (DLS), Fourier Transform Infrared Spectroscopy (FT-IR), Energy Dispersive X-ray Spectroscopy (EDX), Vibrating-sample Magnetometer (VSM) and X-ray Diffraction Analysis (XRD). The cytotoxic potential of BiFe2O4 @Ag was evaluated by MTT assay against SW480 colon cancer cell line. The expression levels of apoptotic genes including BAX, BCL2 and CASP8 were determined by Real-time PCR. The rate of apoptosis and necrosis of the cancer cells as well as the cell cycle analysis were evaluated by flow cytometry. RESULTS: Physicochemical assays indicated the nanoscale synthesis (10-70 nm) and functionalization of BiFe2O4 nanoparticles by Ag atoms. The VSM analysis revealed the magnetism of BiFe2O4 @Ag nanocomposite. According to the MTT assay, colon cancer cells (SW480) were considerably more sensitive to BiFe2O4 @Ag nanocomposite than normal cells. Apoptotic cell percentage increased from 1.93% to 73.66%, after exposure to the nanocomposite. Cell cycle analysis confirmed an increase in the number of the cells in subG1 and G0/G1 phases among nanocomposite treated cells. Moreover, treating the colon cancer cells with BiFe2O4 @Ag caused an increase in the expression of CASP8, BAX, and BCL2 genes by 3.1, 2.6, and 1.2 folds, respectively. Moreover, activity of Caspase-3 protein increased by 2.4 folds and apoptotic morphological changes appeared which confirms that exposure to the nanocomposite induces extrinsic pathway of apoptosis in colon cancer cells. CONCLUSION: The considerable anticancer potential of the synthesized BiFe2O4 @Ag nanocomposite seems to be related to the induction of oxidative stress which leads to inhibit cell cycle progression and cell proliferation. This study reveals that the BiFe2O4 @Ag is a potent compound to be used in biomedical fields.
Asunto(s)
Antineoplásicos , Chlorella vulgaris , Neoplasias del Colon , Nanopartículas del Metal , Nanocompuestos , Humanos , Chlorella vulgaris/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Espectroscopía Infrarroja por Transformada de Fourier , Apoptosis , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Antineoplásicos/farmacología , Antineoplásicos/química , Nanocompuestos/química , Nanopartículas del Metal/química , Caspasa 8/metabolismo , Caspasa 8/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
BACKGROUND: Pyroptosis plays an important role in the pathological process of ischemic stroke (IS). However, the exact mechanism of pyroptosis remains unclear. This paper aims to reveal the key molecular markers associated with pyroptosis in IS. METHODS: We used random forest learning, gene set variation analysis, and Pearson correlation analysis to screen for biomarkers associated with pyroptosis in IS. Middle cerebral artery occlusion/reperfusion (MCAO/R) and oxygen and glucose deprivation/reoxygenation (OGD/R) models were constructed in vitro and in vivo. Cells were transfected with an Annexin A3 silencing (si-ANXA3) plasmid to observe the effects of ANXA3 on OGD/R + lipopolysaccharides (LPS)-induced pyroptosis. qRTâPCR and western blotting were used to detect the expression of potential biomarkers and pyroptotic pathways. RESULTS: Samples from a total of 170 IS patients and 109 healthy individuals were obtained from 5 gene expression omnibus databases. Thirty important genes were analyzed by random forest learning from the differentially expressed genes. Then, we investigated the relationship between the above genes and the pyroptosis score, obtaining three potential biomarkers (ANXA3, ANKRD22, ADM). ANXA3 and ADM were upregulated in the MCAO/R model, and the fold difference in ANXA3 expression was greater. Pyroptosis-related factors (NLRP3, NLRC4, AIM2, GSDMD-N, caspase-8, pro-caspase-1, cleaved caspase-1, IL-1ß, and IL-18) were upregulated in the MCAO/R model. Silencing ANXA3 alleviated the expression of pyroptosis-related factors (NLRC4, AIM2, GSDMD-N, caspase-8, pro-caspase-1, cleaved caspase-1, and IL-18) induced by OGD/R + LPS or MCAO/R. CONCLUSION: This study identified ANXA3 as a possible pyroptosis-related gene marker in IS through bioinformatics and experiments. ANXA3 could inhibit pyroptosis through the NLRC4/AIM2 axis.
Asunto(s)
Accidente Cerebrovascular Isquémico , Daño por Reperfusión , Humanos , Piroptosis/genética , Interleucina-18/metabolismo , Interleucina-18/farmacología , Caspasa 1/metabolismo , Caspasa 1/farmacología , Caspasa 8/metabolismo , Caspasa 8/farmacología , Accidente Cerebrovascular Isquémico/genética , Lipopolisacáridos/farmacología , Biomarcadores , Daño por Reperfusión/genética , Daño por Reperfusión/metabolismo , Anexina A3/genética , Anexina A3/metabolismo , Anexina A3/farmacologíaRESUMEN
Dietary high-fructose may cause metabolic disturbances; however, its effect on the reproductive system is little understood. The insulin signaling pathway is critical in testicular development, maintenance of microcirculation and spermatogenesis. Therefore, in this study, we aimed to investigate the impact of dietary high-fructose on insulin signaling pathway as well as macrophage and apoptotic markers in testicular tissue of rats. Fructose was administered to male Wistar rats as a 20% solution in drinking water for fifteen-week. Gene expression of ir-ß, irs-1, irs-2, pi3k, akt, mtor, and enos in the testicular samples was determined by real-time PCR. Protein expression of IR, IRS-1, IRS-2, PI3K, Akt, phospho-Akt (p-Akt), mTOR, eNOS, phospho-eNOS (p-eNOS), and GLUT5 was established by analysis of Western Blot. Testicular expression of occludin, CD163, CD68, caspase-8, and caspase-3 was analyzed by using immunohistochemical assay. Testicular level of fructose was measured by colorimetric method. Dietary high-fructose decreased mRNA expressions of irs-1, irs-2, pi3k, and mtor in the testicular tissue of rats. Also, this dietary intervention impaired protein expressions of IR, IRS-1, IRS-2, PI3K, p-Akt, mTOR, eNOS, and p-eNOS as well as p-Akt/Akt and p-eNOS/eNOS ratios in the testis of rats. However, a high-fructose diet increased the expression of CD163, CD68, caspase-8 and caspase-3, but decreased that of occludin, in the testicular tissue of rats. The high-fructose consumption in rats suppresses testicular insulin signaling but activates macrophages-related factors and apoptotic markers. These changes induced by dietary fructose could be related to male reproductive dysfunction.
Asunto(s)
Insulina , Proteínas Proto-Oncogénicas c-akt , Ratas , Masculino , Animales , Insulina/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Fructosa/farmacología , Ratas Wistar , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Testículo/metabolismo , Ocludina/metabolismo , Ocludina/farmacología , Transducción de Señal , Fosfatidilinositol 3-Quinasas/metabolismo , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
During differentiation, neutrophils undergo a spontaneous pro-inflammatory program that is hypothesized here to be under caspase-8 control. In mice, intraperitoneal administration of the caspase-8 inhibitor z-IETD-fmk is sufficient to unleash the production of pro-inflammatory cytokines and neutrophil influx in the absence of cell death. These effects are due to selective inhibition of caspase-8 and require tonic interferon-ß (IFN-ß) production and RIPK3 but not MLKL, the essential downstream executioner of necroptotic cell death. In vitro, stimulation with z-IETD-fmk is sufficient to induce significant cytokine production in murine neutrophils but not in macrophages. Therapeutic administration of z-IETD-fmk improves clinical outcome in models of lethal bacterial peritonitis and pneumonia by augmenting cytokine release, neutrophil influx, and bacterial clearance. Moreover, the inhibitor protects mice against high-dose endotoxin shock. Collectively, our data unveil a RIPK3- and IFN-ß-dependent pathway that is constitutively activated in neutrophils and can be harnessed therapeutically using caspase-8 inhibition.
Asunto(s)
Apoptosis , Infecciones Bacterianas , Animales , Ratones , Infecciones Bacterianas/tratamiento farmacológico , Caspasa 8/metabolismo , Caspasa 8/farmacología , Citocinas/metabolismo , Activación NeutrófilaRESUMEN
Particles of various types of plastics, including polystyrene nanoparticles (PS-NPs), have been determined in human blood, placenta, and lungs. These findings suggest a potential detrimental effect of PS-NPs on bloodstream cells. The purpose of this study was to assess the mechanism underlying PS-NPs-induced apoptosis in human peripheral blood mononuclear cells (PBMCs). Non-functionalized PS-NPs of three diameters: 29 nm, 44 nm, and 72 nm were studied used in this research. PBMCs were isolated from human leukocyte-platelet buffy coat and treated with PS-NPs at concentrations ranging from 0.001 to 200 µg/mL for 24 h. Apoptotic mechanism of action was evaluated by determining the level of cytosolic calcium ions, as well as mitochondrial transmembrane potential, and ATP levels. Furthermore, detection of caspase-8, -9, and -3 activation, as well as mTOR level was conducted. The presence of apoptotic PBMCs was confirmed by the method of double staining of the cells with propidium iodide and FITC-conjugated Annexin V. We found that all tested NPs increased calcium ion and depleted mitochondrial transmembrane potential levels. The tested NPs also activated caspase-9 and caspase-3, and the smallest NPs of 29 nm of diameter also activated caspase-8. The results clearly showed that apoptotic changes and an increase of mTOR level depended on the size of the tested NPs, while the smallest particles caused the greatest alterations. PS-NPs of 26 nm of diameter activated the extrinsic pathway (increased caspase-8 activity), as well as intrinsic (mitochondrial) pathway (increased caspase-9 activity, raised calcium ion level, and decreased transmembrane mitochondrial potential) of apoptosis. All PS-NPs increased mTOR level at the concentrations smaller than those that induced apoptosis and its level returned to control value when the process of apoptosis escalated.
Asunto(s)
Leucocitos Mononucleares , Nanopartículas , Humanos , Poliestirenos/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Caspasa 9/metabolismo , Calcio/metabolismo , Apoptosis , Nanopartículas/toxicidad , Potencial de la Membrana Mitocondrial , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Fluoride compounds are abundant and widely distributed in the environment at various concentrations, which can seriously injure the human body. In this study, we aim to evaluate the effects of excessive fluoride exposure on the liver, kidney, and heart tissues of healthy female Xenopus laevis by administering NaF (0, 100, and 200 mg/L) in drinking water for 90 days. The expression level of procaspase-8, cleaved-caspase-8, and procaspase-3 proteins were determined by Western blot. Compared with the control group, the group exposed to NaF exhibited expression levels of procaspase-8, cleaved-caspase-8, and procaspase-3 proteins that were considerably upregulated at a concentration of 200 mg/L in the liver and kidney. The cleaved-caspase-8 protein expression in the group exposed to a high concentration of NaF was lower than that in the control group in heart. Histopathological results by hematoxylin and eosin staining showed that excessive NaF exposure caused necrosis of hepatocytes and vacuolization degeneration. Granular degeneration and necrosis in renal tubular epithelial cells were also observed. Moreover, hypertrophy of myocardial cells, atrophy of myocardial fibers and disorder of myocardial fibers were detected. These results demonstrated that NaF-induced apoptosis and the mediated death receptor pathway activation ultimately damaged the liver and kidney tissues. This finding offers a fresh perspective on the effects of F-induced apoptosis in X. laevis.
Asunto(s)
Apoptosis , Fluoruros , Animales , Femenino , Humanos , Fluoruros/metabolismo , Fluoruros/farmacología , Xenopus laevis/metabolismo , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Riñón/metabolismo , Hígado/metabolismo , Transducción de Señal , NecrosisRESUMEN
Paracetamol is the most predominantly used antipyretic and analgesic drug. As paracetamol is metabolised mostly in the liver, both deliberate and unintentional overdoses of paracetamol are reported to provoke severe hepatotoxicity, including liver failure. Caesalpinia bonducella seed is well known for its medicinal and therapeutic properties. However, there is no report on its potential protective effects against paracetamol-instigated hepatotoxicity. Therefore, we studied the protective effects of aqueous seed extract of Caesalpinia bonducella (ASECB) on paracetamol-instigated hepatotoxicity in rats. Thirty female albino rats were divided into five groups: control, paracetamol-intoxicated, ASECB + paracetamol, silymarin + paracetamol, and ASECB alone. The rats were assessed for liver enzyme markers (alkaline phosphatase, alanine aminotransferase, aspartate aminotransferase, gamma-glutamyl transpeptidase), antioxidant activity (superoxide dismutase, catalase, reduced glutathione, glutathione peroxidase), lipid peroxidation (malondialdehyde), histopathological, cytokine levels (pro-inflammatory cytokines TNF-α and IL-6, and anti-inflammatory cytokine IL-10), and protein expression (pro-apoptotic markers caspase 3 and caspase 8 and anti-apoptotic marker Bcl-2) after the 8-day study period. Repercussions of paracetamol intoxication induced upregulation of liver enzyme markers, antioxidant depletion, malondialdehyde production, decreased expression of Bcl-2 and IL-10, and overexpression of apoptotic and pro-inflammatory mediators, which were attenuated by pre-treatment with ASECB. ASECB markedly mitigated paracetamol-instigated liver injury by suppressing caspase-8/3 signalling and inflammatory infiltration in liver tissue by significantly reducing TNF-α and IL-6. In conclusion, ASECB pre-treatment exerts potent liver protection against paracetamol-instigated hepatotoxicity evidenced by mitigation of oxidative stress, lipid peroxidation, inflammation, and apoptosis.
Asunto(s)
Caesalpinia , Enfermedad Hepática Inducida por Sustancias y Drogas , Ratas , Femenino , Animales , Acetaminofén/toxicidad , Acetaminofén/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Caesalpinia/metabolismo , Interleucina-6/metabolismo , Interleucina-10/metabolismo , Caspasa 3/metabolismo , Antioxidantes/farmacología , Estrés Oxidativo , Hígado/metabolismo , Citocinas/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/tratamiento farmacológico , Enfermedad Hepática Inducida por Sustancias y Drogas/prevención & control , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Malondialdehído/metabolismoRESUMEN
Degenerative cervical myelopathy (DCM) is a severe condition of the spinal cord caused by chronic compression. However, no studies to date have examined the effects of zonisamide (ZNS) on DCM via the Fas/FasL-mediated pathway. The aim of this study was to investigate the effects of ZNS on a DCM rat model and to explore the potential mechanisms. First, 40 adult Sprague-Dawley rats were used to establish the DCM rat model and were individually divided into four groups: the Sham group, DCM model group (DCM), ZNS group (DCM model rats treated with ZNS, 30 mg/kg/day), and ZNS + CD95 group (DCM model rats treated with ZNS and CD95). Histopathology injury and cell apoptosis, Fas and Fas ligand (FasL) expression and Fas/FasL relative protein levels were detected by hematoxylin and eosin staining, TUNEL assay, and immunofluorescence and western blotting, respectively. The results of our study demonstrated that ZNS could promote motor recovery while reversing histopathological injury and cell apoptosis in DCM rats. Moreover, Iba-1, Fas and FasL expression in DCM rats was decreased, accompanied by a decrease in cleaved caspase-3/caspase-3, cleaved caspase-8/caspase-8, cleaved caspase-9/caspase-9, cleaved caspase-10/caspase-10 and B-cell lymphoma-2 (Bcl-2)/Bcl-2 associated X (Bax) levels. All these results revealed that ZNS attenuates DCM injury in a rat model via the regulation of Fas and FasL signaling. Our study indicated that ZNS had beneficial effects on DCM and thus provided a novel theoretical approach for subsequent academic and clinical research on DCM injury.
Asunto(s)
Apoptosis , Enfermedades de la Médula Espinal , Ratas , Animales , Proteína Ligando Fas/metabolismo , Ratas Sprague-Dawley , Caspasa 3/metabolismo , Caspasa 9/metabolismo , Caspasa 9/farmacología , Zonisamida/farmacología , Caspasa 10/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Inflamación/tratamiento farmacológico , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
Cisplatin is widely used as an anti-neoplastic agent but is limited by its nephrotoxicity. The use of mesenchymal stem cells (MSCs) for the management of acute kidney injury (AKI) represents a new era in treatment but effective homing of administered cells is needed. This study aimed to investigate the effect of bone marrow-derived mesenchymal stem cells (BM-MSCs) on cisplatin-induced AKI in rats after directed migration by electric field (EF). Forty-eight adult male albino rats were equally classified into four groups: control, cisplatin-treated, cisplatin plus BM-MSCs, and cisplatin plus BM-MSCs exposed to EF. Serum levels of IL-10 and TNF-α were measured by ELISA. Quantitative real-time PCR analysis for gene expression of Bcl2, Bax, caspase-3, and caspase-8 was measured. Hematoxylin and eosin (H&E) staining, periodic acid Schiff staining, and immunohistochemical analysis were also done. MSC-treated groups showed improvement of kidney function; increased serum levels of IL-10 and decreased levels of TNF-α; and increased mRNA expression of Bcl2 and decreased expression of Bax, caspase-3, and caspase-8 proteins comparable to the cisplatin-injured group. EF application increased MSCs homing with significant decrease in serum urea level and caspase-3 gene expression together with significant increase in Bcl2 expression than occurred in the MSCs group. Restoration of normal kidney histomorphology with significant decrease in immunohistochemical expression of caspase-3 protein was observed in the BM-MSCs plus EF group compared to the BM-MSCs group. EF stimulation enhanced the MSCs homing and improved their therapeutic potential on acute cisplatin nephrotoxicity.
Asunto(s)
Lesión Renal Aguda , Células Madre Mesenquimatosas , Humanos , Masculino , Lesión Renal Aguda/inducido químicamente , Lesión Renal Aguda/terapia , Proteína X Asociada a bcl-2/metabolismo , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Cisplatino/toxicidad , Interleucina-10/genética , Interleucina-10/metabolismo , Células Madre Mesenquimatosas/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Animales , RatasRESUMEN
Photodynamic therapy (PDT) is a light-based anti-neoplastic therapeutic approach. Growing evidence indicates that combining conventional anti-cancer therapies with PDT can be a promising approach to treat malignancies. Herein, we aimed to investigate anti-cancer effects of the combination treatment of zinc phthalocyanine (ZnPc)-PDT with tamoxifen (TA) on MDA-MB-231 cells (as a triple-negative breast cancer (TNBC) cell line). For this purpose, we investigated the cytotoxicity of TA and ZnPc-PDT on MDA-MB-231 cells performing the MTT assay. The effect of TA and ZnPc-PDT on the apoptosis of MDA-MB-231 cells was studied using Annexin V/PI and DAPI staining. The wound-healing assay, and colony formation assay were performed to study the effect of TA and ZnPc-PDT on the migration, and clonogenicity of MDA-MB-231 cells, respectively. The qRT-PCR was done to study the gene expression of caspase-8, caspase-9, caspase-3, ZEB1, ROCK1, SNAIL1, CD133, CD44, SOX2, and ABCG2 (ATP-binding cassette sub-family G member 2). Based on our results, monotherapies with TA and ZnPc-PDT can remarkably increase cell cytotoxicity effects, stimulate apoptosis via downregulating Bcl-2 and upregulating caspase-3 and caspase-9, inhibit migration via downregulating SNAIL1 and ZEB1, and suppress clonogenicity via downregulating SOX2 and CD44 in MDA-MB-231 cells. Besides, these monotherapies can downregulate the expression of ABCG2 in MDA-MB-231 cells. Nevertheless, the combination treatment can potentiate the above-mentioned anti-cancer effects compared to monotherapy with TA. Of interest, the combined treatment of TA with ZnPc-PDT can synergically increase cell cytotoxicity effects on MDA-MB-231 cells. In fact, synergistic effects were estimated by calculation of Combination Index (CI); that synergistic outcomes were observed in all groups. Also, this combination treatment can significantly upregulate the caspase-8 gene expression and downregulate ROCK1 and CD133 gene expression in MDA-MB-231 cells. Overall, our results show that ZnPc-PDT can more sensitize the MDA-MB-231 cells to TA treatment. Based on our knowledge and experiment, the synergistic effects of ZnPc-PDT and TA deserve further evaluation in cancer research.
Asunto(s)
Fotoquimioterapia , Neoplasias de la Mama Triple Negativas , Humanos , Fármacos Fotosensibilizantes/uso terapéutico , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Caspasa 3 , Caspasa 9/farmacología , Caspasa 8/farmacología , Caspasa 8/uso terapéutico , Fotoquimioterapia/métodos , Tamoxifeno/farmacología , Tamoxifeno/uso terapéutico , Línea Celular Tumoral , Indoles , Apoptosis , Quinasas Asociadas a rho/farmacología , Quinasas Asociadas a rho/uso terapéuticoRESUMEN
OBJECTIVES: Excessive inflammatory responses and apoptosis are critical pathologies that contribute to sepsis-induced acute kidney injury (SI-AKI). Annexin A1 (ANXA1), a member of the calcium-dependent phospholipid-binding protein family, protects against SI-AKI through its anti-inflammatory and antiapoptotic effects, but the underlying mechanisms are still largely unknown. METHODS: In vivo, SI-AKI mouse models were established via caecal ligation and puncture (CLP) and were then treated with the Ac2-26 peptide of ANXA1 (ANXA1 (Ac2-26)), WRW4 (Fpr2 antagonist) or both. In vitro, HK-2 cells were induced by lipopolysaccharide (LPS) and then treated with ANXA1 (Ac2-26), Fpr2-siRNA or both. RESULTS: In the present study, we found that the expression levels of ANXA1 were decreased, and the expression levels of TNF-α, IL-1ß, IL-6, cleaved caspase-3, cleaved caspase-8 and Bax were significantly increased, accompanied by marked kidney tissue apoptosis in vivo. Moreover, we observed that ANXA1 (Ac2-26) significantly reduced the levels of TNF-α, IL-1ß and IL-6 and cleaved caspase-3, cleaved caspase-8, FADD and Bax and inhibited apoptosis in kidney tissue and HK-2 cells, accompanied by pathological damage to kidney tissue. Seven-day survival, kidney function and cell viability were significantly improved in vivo and in vitro, respectively. Furthermore, the administration of ANXA1 (Ac2-26) inhibited the CLP- or LPS-induced phosphorylation of PI3K and AKT and downregulated the level of NF-κB in vivo and in vitro. Moreover, our data demonstrate that blocking the Fpr2 receptor by the administration of WRW4 or Fpr2-siRNA reversed the abovementioned regulatory role of ANXA1, accompanied by enhanced phosphorylation of PI3K and AKT and upregulation of the level of NF-κB in vivo and in vitro. CONCLUSIONS: Taken together, this study provides evidence that the protective effect of ANXA1 (Ac2-26) on SI-AKI largely depends on the negative regulation of inflammation and apoptosis via the Fpr2 receptor.
Asunto(s)
Lesión Renal Aguda , Anexina A1 , Sepsis , Ratones , Animales , FN-kappa B/metabolismo , Caspasa 3/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Lipopolisacáridos/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Interleucina-6/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Anexina A1/farmacología , Anexina A1/uso terapéutico , Anexina A1/genética , Proteína X Asociada a bcl-2/metabolismo , Proteína X Asociada a bcl-2/farmacología , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Lesión Renal Aguda/tratamiento farmacológico , Lesión Renal Aguda/etiología , Lesión Renal Aguda/metabolismo , Apoptosis , Sepsis/complicaciones , Sepsis/tratamiento farmacológico , Sepsis/metabolismo , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/farmacología , Fosfatidilinositol 3-Quinasas/metabolismoRESUMEN
Glioblastoma multiforme (GBM) is the most aggressive type of glioma, displaying atypical glycosylation pattern that may modulate signaling pathways involved in tumorigenesis. Lectins are glycan binding proteins with antitumor properties. The present study was designed to evaluate the antitumor capacity of the Dioclea reflexa lectin (DrfL) on glioma cell cultures. Our results demonstrated that DrfL induced morphological changes and cytotoxic effects in glioma cell cultures of C6, U-87MG and GBM1 cell lines. The action of DrfL was dependent upon interaction with glycans, and required a carbohydrate recognition domain (CRD), and the cytotoxic effect was apparently selective for tumor cells, not altering viability and morphology of primary astrocytes. DrfL inhibited tumor cell migration, adhesion, proliferation and survival, and these effects were accompanied by activation of p38MAPK and JNK (p46/54), along with inhibition of Akt and ERK1/2. DrfL also upregulated pro-apoptotic (BNIP3 and PUMA) and autophagic proteins (Atg5 and LC3 cleavage) in GBM cells. Noteworthy, inhibition of autophagy and caspase-8 were both able to attenuate cell death in GBM cells treated with DrfL. Our results indicate that DrfL cytotoxicity against GBM involves modulation of cell pathways, including MAPKs and Akt, which are associated with autophagy and caspase-8 dependent cell death.
Asunto(s)
Antineoplásicos , Muerte Celular Autofágica , Dioclea , Glioma , Humanos , Dioclea/química , Caspasa 8/metabolismo , Caspasa 8/farmacología , Caspasa 8/uso terapéutico , Lectinas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas Proto-Oncogénicas c-akt/farmacología , Proteínas Proto-Oncogénicas c-akt/uso terapéutico , Línea Celular Tumoral , Glioma/tratamiento farmacológico , Glioma/metabolismo , Glioma/patología , Movimiento Celular , Autofagia , Antineoplásicos/farmacología , Proliferación Celular , ApoptosisRESUMEN
BACKGROUND: The purpose of the present research was to explore the therapeutic impact of raw lacquer extract from Toxicodendron vernicifluum on colorectal cancer cells and to investigate the outcome of raw lacquer extract and ONC201 co-treatment on the activity of colorectal cancer cells. METHODS: The cells of HCT116 were treated with raw lacquer extract, ONC201, or co-treatment. Subsequently, MTT, trypan blue staining, colony formation, annexin V/propidium iodide staining, wound healing, and transwell assays were performed to assess the effects of raw lacquer extract, ONC201 and the synthesis effect of co-treatment on cell activity, survival, proliferation, apoptosis, migration, and invasion in HCT116 cells. Western blotting and immunostaining assay were also performed to detect the expressions of tumor necrosis factor-related apoptosis-inducing ligand, death receptor-5, cleaved caspase-8, p-mTOR/mTOR, and p-S6K/S6K in cells. RESULTS: The results showed that ONC201 and raw lacquer extract had effective anti-cancer effects on HCT116 cells. ONC201 and raw lacquer extract treatment on colorectal cancer cells inhibited cell viability and growth, as well as induced cell apoptosis and cell death of HCT116. The migration and invasion of HCT116 cells were also inhibited. Significantly, raw lacquer extract and ONC201 cotreatment further enhanced the anti-colorectal cancer cell activity in HCT116 cells. Western blotting and immunostaining assay showed that raw lacquer extract in combination with ONC201 induced tumor necrosis factor-related apoptosis-inducing ligand/death receptor-5 expression activation, inhibited the expression of cleaved caspase-8/procaspase-8, and reduced the expression of p-mTOR/mTOR and p-S6K/S6K. CONCLUSION: These results indicated that raw lacquer extract in combination with ONC201 enhanced the inhibitory effects on colorectal cancer cell activity.
Asunto(s)
Neoplasias Colorrectales , Toxicodendron , Humanos , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Toxicodendron/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Caspasa 8/uso terapéutico , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/uso terapéutico , Laca , Ligandos , Serina-Treonina Quinasas TOR/metabolismo , Apoptosis , Factores de Necrosis Tumoral/metabolismo , Factores de Necrosis Tumoral/farmacología , Factores de Necrosis Tumoral/uso terapéutico , Proliferación CelularRESUMEN
Photodynamic therapy (PDT) is a new therapy for treating cancer with less toxicity, high selectivity, good cooperativity, and repetitive usability. However, keloid treatment by PDT is mainly focused on clinical appearance, and few studies have been conducted on the mechanisms of PDT. In this study, key factors of the classical mitochondrial apoptosis signaling pathway were measured to assess the effect of a new PDT photosensitizer (p1). A specific inhibitor of caspase-8 (Z-IETD-FMK) was also used to verify the possible mechanisms. Twelve samples were obtained from 12 patients (six with keloids and six without) selected randomly from the Department of Plastic Surgery at Peking Union Medical College Hospital from January to December 2020. After cell culture, fibroblasts were divided into 13 groups. The morphology of fibroblasts in each group was observed by microscopy. Cell activity was measured by cell counting kit-8, and cell apoptotic morphology was observed by TUNEL staining. The reactive oxygen species (ROS) relative value was measured by a ROS test kit. The expression levels of key mitochondrial factors (caspase-3, caspase-8, cytochrome-c, Bax, and Bcl-2) were assessed by western blot, and mRNA expression of caspase-3 and caspase-8 was measured by RT-qPCR. We showed that p1 had a satisfactory proapoptotic effect on keloid fibroblasts by increasing the expression of ROS, caspase-3, caspase-8, and cytochrome-c, and decreasing the Bcl-2/Bax ratio; however, this effect was partially inhibited by Z-IETD-FMK, indicating that caspase-8 may be one of the p1's targets to achieve the proapoptotic effect.
Asunto(s)
Queloide , Fotoquimioterapia , Humanos , Fármacos Fotosensibilizantes/farmacología , Fármacos Fotosensibilizantes/metabolismo , Fármacos Fotosensibilizantes/uso terapéutico , Caspasa 3/metabolismo , Caspasa 3/farmacología , Caspasa 3/uso terapéutico , Queloide/tratamiento farmacológico , Queloide/patología , Especies Reactivas de Oxígeno/metabolismo , Especies Reactivas de Oxígeno/farmacología , Especies Reactivas de Oxígeno/uso terapéutico , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismo , Proteína X Asociada a bcl-2/farmacología , Caspasa 8/metabolismo , Caspasa 8/farmacología , Caspasa 8/uso terapéutico , Apoptosis , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/uso terapéutico , Fibroblastos/patología , Citocromos/metabolismo , Citocromos/farmacología , Citocromos/uso terapéuticoRESUMEN
The activation of receptor-interacting protein kinase 1 (RIPK1) by death-inducing signaling complex (DISC) formation is essential for triggering the necroptotic mode of cell death under apoptosis-deficient conditions. Thus, targeting the induction of necroptosis by modulating RIPK1 activity could be an effective strategy to bypass apoptosis resistance in certain types of cancer. In this study, we screened a series of arborinane triterpenoids purified from Rubia philippinesis and identified rubiarbonol B (Ru-B) as a potent caspase-8 activator that induces DISC-mediated apoptosis in multiple types of cancer cells. However, in RIPK3-expressing human colorectal cancer (CRC) cells, the pharmacological or genetic inhibition of caspase-8 shifted the mode of cell death by Ru-B from apoptosis to necroptosis though upregulation of RIPK1 phosphorylation. Conversely, Ru-B-induced cell death was almost completely abrogated by RIPK1 deficiency. The enhanced RIPK1 phosphorylation and necroptosis triggered by Ru-B treatment occurred independently of tumor necrosis factor receptor signaling and was mediated by the production of reactive oxygen species via NADPH oxidase 1 in CRC cells. Thus, we propose Ru-B as a novel anticancer agent that activates RIPK1-dependent cell death via ROS production, and suggest its potential as a novel necroptosis-targeting compound in apoptosis-resistant CRC.
Asunto(s)
Apoptosis , Necroptosis , Humanos , Especies Reactivas de Oxígeno/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Muerte Celular , Necrosis , Proteína Serina-Treonina Quinasas de Interacción con Receptores/genética , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , NADPH Oxidasa 1/metabolismo , NADPH Oxidasa 1/farmacologíaRESUMEN
Recent reports have suggested that abnormal miR-29c expression in hippocampus have been implicated in the pathophysiology of some neurodegenerative and neuropsychiatric diseases. However, the underlying effect of miR-29c in regulating hippocampal neuronal function is not clear. In this study, HT22 cells were infected with lentivirus containing miR-29c or miR-29c sponge. Cell counting kit-8 (CCK8) and lactate dehydrogenase (LDH) assay kit were applied to evaluate cell viability and toxicity before and after TNF-α administration. Reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP) were measured with fluorescent probes. Hoechst 33258 staining and TUNEL assay were used to evaluate cell apoptosis. The expression of key mRNA/proteins (TNFR1, Bcl-2, Bax, TRADD, FADD, caspase-3, -8 and -9) in the apoptosis pathway was detected by PCR or WB. In addition, the protein expression of microtubule-associated protein-2 (MAP-2), nerve growth-associated protein 43 (GAP-43) and synapsin-1 (SYN-1) was detected by WB. As a result, we found that miR-29c overexpression could improve cell viability, attenuate LDH release, reduce ROS production and inhibit MMP depolarization in TNF-α-treated HT22 cells. Furthermore, miR-29c overexpression was found to decrease apoptotic rate, along with decreased expression of Bax, cleaved caspase-3, cleaved caspase-9, and increased expression of Bcl-2 in TNF-α-treated HT22 cells. However, miR-29c sponge exhibited an opposite effects. In addition, in TNF-α-treated HT22 cells, miR-29c overexpression could decrease the expressions of TNFR1, TRADD, FADD and cleaved caspase-8. However, in HT22 cells transfected with miR-29c sponge, TNF-α-induced the expressions of TNFR1, TRADD, FADD and cleaved caspase-8 was significantly exacerbated. At last, TNF-α-induced the decreased expression of MAP-2, GAP-43 and SYN-1 was reversed by miR-29c but exacerbated by miR-29c sponge. Overall, our study demonstrated that miR-29c protects against TNF-α-induced HT22 cells injury through alleviating ROS production and reduce neuronal apoptosis. Therefore, miR-29c might be a potential therapeutic agent for TNF-α accumulation and toxicity-related brain diseases.
Asunto(s)
MicroARNs , Factor de Necrosis Tumoral alfa , Ratones , Animales , Especies Reactivas de Oxígeno/metabolismo , Caspasa 3/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Factor de Necrosis Tumoral alfa/metabolismo , Caspasa 8/metabolismo , Caspasa 8/farmacología , Receptores Tipo I de Factores de Necrosis Tumoral , Proteína X Asociada a bcl-2/metabolismo , Proteína GAP-43/metabolismo , Línea Celular , Apoptosis , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , MicroARNs/metabolismo , Hipocampo/metabolismoRESUMEN
Intestinal microecology was closely related to immune regulation, but the related mechanism was still unclear. This study aimed to reveal how microorganisms improved immune response via casepase-3 and Bak of FAS/CD95 pathway. Bifidobacterium animalis F1-7 inhibited the melanoma B16-F10 cells in vitro effectively; had a potent anticancer effect of lung cancer mice; effectively improved the spleen immune index and CD3+ (75.8%) and CD8+ (19.8%) expression level; strengthened the phagocytosis of macrophages; inhibited the overexpression of inflammatory factors IL-6 (319.10 ± 2.46 pg/mL), IL-8 (383.05 ± 9.87 pg/mL), and TNF-α (2003.40 ± 11.42 pg/mL); and promoted the expression of anti-inflammatory factor IL-10 (406.00 ± 3.59 pg/mL). This process was achieved by promoting caspase-8/3 and BH3-interacting domain death agonist (Bid), Bak genes, and protein expression. This study confirmed the B. animalis F1-7 could act as an effective activator to regulate immune response by promoting the expression of caspase-8/3, Bid and Bak genes, and proteins and by activating the FAS/CD95 pathway. Our study provided a data support for the application of potentially beneficial microorganisms of B. animalis F1-7 as an effective activator to improve immunity.
Asunto(s)
Apoptosis , Bifidobacterium animalis , Ratones , Animales , Caspasa 8/genética , Caspasa 8/metabolismo , Caspasa 8/farmacología , Transducción de Señal/fisiología , Receptor fas/genética , Receptor fas/metabolismo , InmunidadRESUMEN
Cellular c-FLIP prevents apoptosis mediated by death receptor through inhibiting activation of caspase-8. Therefore, when c-FLIP is downregulated or eliminated, caspase-8 activation is promoted, and death receptor ligand-induced apoptosis is activated. It was reported that triptolide (TPL) sensitized tumor cells to TNF-α-induced apoptosis by blocking TNF-α-induced activation of NF-κB and transcription of c-IAP1 and c-IAP2. However, the effect of TPL on basal c-FLIP expression was not understood. In this study, we found that the combination of TNF-α and TPL accelerated apoptosis in human hepatocellular carcinoma cells and TNF-α-induced elevated as well as basal level of FLIPS protein were downregulated by TPL. Additionally, we demonstrated that the basal level of FLIPS in Huh7 cells was continuously downregulated following the incubation of TPL and downregulated more when dosage of TPL for treatment was increased. Subsequently, we showed that TPL reduced FLIPS level in a transcription- and degradation-independent mechanism. Our findings suggest that TPL induces loss of FLIPS at the post-transcriptional level independently of proteasome-mediated pathway, an additional mechanism of TPL sensitizing cancer cells to TNF-α-induced apoptosis.