RESUMEN
The process of spermatogenesis is a complex and delicate process that is still not fully understood. In this study, we examined the role of fatty acid oxidase 3-hydroxy acyl CoA dehydrogenase (HADH) in maintaining normal spermatogenesis in mice. In male mice, ablation of the Hadh gene using CRISPR/Cas9 technology arrested spermatocyte meiosis, increased multinucleated giant germ cells and vacuoles in seminiferous tubules, and accompanied with acrosomal dysplasia. Hadh-/- male mice showed the typical features of oligoasthenoteratozoospermia (OAT), including decreased sperm concentration and motility and increased sperm abnormalities. Next, we explored the molecular events in the testes of the mutant mice. We found fatty acids accumulated in the testis of Hadh-/- mice. And also, inflammatory factors TNF-α, IL-1ß, and IL-6 were significantly increased, apoptosis-related protein Bcl-2 was decreased, and Bax and cleaved-Caspase3 were increased in Hadh-/- male mice testis. After using etanercept, a specific inhibitor of TNF-α, testis injury caused by Hadh knockout was significantly alleviated, the sperm quality and motility were improved, and germ cell apoptosis was reduced. So our study demonstrated that Hadh deletion caused an increase in fatty acids. The accumulated fatty acids further induced testicular inflammation and germ cell apoptosis through the TNF-α/Bcl-2 signaling pathway, finally resulting in OAT in the Hadh-/- mice. Inhibiting TNF-α may be used as a new treatment approach for testicular inflammation and OAT.
Asunto(s)
3-Hidroxiacil-CoA Deshidrogenasa , Astenozoospermia , Infertilidad Masculina , Oligospermia , Animales , Masculino , Ratones , Astenozoospermia/genética , Astenozoospermia/metabolismo , Ácidos Grasos , Infertilidad Masculina/genética , Infertilidad Masculina/metabolismo , Inflamación/genética , Inflamación/metabolismo , Oligospermia/genética , Oligospermia/metabolismo , Semen/metabolismo , Espermatocitos/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , 3-Hidroxiacil-CoA Deshidrogenasa/deficiencia , 3-Hidroxiacil-CoA Deshidrogenasa/genética , 3-Hidroxiacil-CoA Deshidrogenasa/metabolismo , Genes bcl-2/genética , Genes bcl-2/fisiologíaRESUMEN
With the widespread use of plastics, microplastics (MPs) and di(2-ethylhexyl) phthalate (DEHP) have become emerging environmental pollutants. The combined toxicity of MPs and DEHP on the mouse pancreas and the specific mechanism of toxicity remain unclear. To establish in vitro and in vivo models to address these questions, mice were continuously exposed to 200 mg/kg/d DEHP and 10 mg/L MPs for 4 weeks. In vitro, MIN-6 cells were treated with 200 µg/mL MPs and 200 µM DEHP for 24 h. Based on toxicity assessed using CCK8 of the equivalent TU binary mixture, the IC50 of the TU-mix of DEHP and MPs 0.692 < 0.8, indicating a synergistic effect of the two toxicants. Meanwhile, our data revealed that compared to the control group, MPs and DEHP combined treatment increased ROS levels, inhibited the activity, and enhanced the expression of GRP78, and CHOP. Simultaneously, activated CHOP decreased the expression of Bcl-2, and increased the expression of Bax. In conclusion, DEHP and MPs synergistically induce oxidative stress, and activate the GRP78/CHOP/Bcl-2 pathway to induce pancreatic apoptosis in mice. Our finding provides a new direction for the research on the specific mechanism of MPs and DEHP combined toxicity.
Asunto(s)
Dietilhexil Ftalato , Chaperón BiP del Retículo Endoplásmico , Genes bcl-2 , Microplásticos , Estrés Oxidativo , Páncreas , Factor de Transcripción CHOP , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Dietilhexil Ftalato/toxicidad , Chaperón BiP del Retículo Endoplásmico/genética , Chaperón BiP del Retículo Endoplásmico/metabolismo , Genes bcl-2/genética , Genes bcl-2/fisiología , Ratones , Microplásticos/efectos adversos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Páncreas/efectos de los fármacos , Páncreas/metabolismo , Páncreas/patología , Ácidos Ftálicos , Plásticos , Factor de Transcripción CHOP/genética , Factor de Transcripción CHOP/metabolismoRESUMEN
SUMMARY: Statins inhibit cholesterol synthesis, but also have other pleiotropic effects. There are indications that they affect macrophage survival trough the regulation of apoptosis. We analyzed 50 samples of aortic wall, selected based on statins in patients' therapy (n=25, Th-S group) or statin-free therapy (n=25, Th-nonS group). Each group had 5 samples of healthy aortic tissue, 10 samples of mild and 10 samples of severe atherosclerotic changes in aortic wall. Tissue was stained with hematoxylin-eosin and immunohistochemical methods (anti-Bcl-2 antibody). Presence of Bcl2-positive macrophages (Bcl-2+ MP) was determined semiquantitatively, and data were processed in Microsoft Excell and IMB SPSS 23 Statistics. 60 % of patients in the Th-S group had a mild increase of Bcl-2+ MP The use of statins leads to a significantly more frequent increase in Bcl2+ macrophages in the intima of the healthy aortic tissue. Analysis of all aortic samples with pathohistological diagnosis showed that statin therapy was statistically significantly more often leading to a markedly increased presence of Bcl-2+ MP. In the media, all samples of the Th-S group have a mild increase of Bcl-2+ MP, and in adventitia 40 % of patients. The use of statins more often leads to a markedly increased presence of Bcl-2+ MP in aortic tissue with diagnosed mild and severe atherosclerosis. In samples of severe atherosclerosis, statins lead to a markedly increased presence of Bcl-2+ MP in the parts of the plaque towards the intima and towards the media. Statins lead to an increased presence of Bcl-2+ macrophages, prolong their life, both in healthy and atherosclerotic altered aortic tissue. This indicates potentiation of inflammation and damage to the aortic wall, and calls into question the positive effect of statins on the aortic wall with atherosclerosis.
RESUMEN: Las estatinas inhiben la síntesis de colesterol, pero también tienen otros efectos pleiotrópicos. Hay indicios de que afectan la supervivencia de los macrófagos a través de la regulación de la apoptosis.Se analizaron 50 muestras de pared aórtica, seleccionadas en base a estatinas en tratamiento de pacientes (n=25, grupo Th-S) o en tratamiento libre de estatinas (n=25, grupo Th- nonS). Cada grupo tenía 5 muestras de tejido aórtico sano, 10 muestras de cambios ateroscleróticos leves y 10 muestras de cambios ateroscleróticos severos en la pared aórtica. El tejido se tiñó con hematoxilina-eosina y métodos inmunohistoquímicos (anticuerpo anti-Bcl-2). La presencia de macrófagos positivos para Bcl2 (Bcl- 2+ MP) se determinó semicuantitativamente y los datos se procesaron en Microsoft Excell e IMB SPSS 23 Statistics. El 60 % de los pacientes del grupo Th-S tuvo un aumento leve de Bcl-2+ MP. El uso de estatinas conduce a un aumento significativamente más frecuente de macrófagos Bcl2+ en la íntima del tejido aórtico sano. El análisis de todas las muestras aórticas con diagnóstico anatomopatológico mostró que la terapia con estatinas fue significativamente más frecuente desde el punto de vista estadístico, lo que condujo a una presencia marcadamente mayor de Bcl-2+ MP. En los medios, todas las muestras del grupo Th-S tienen un leve aumento de Bcl-2+ MP, y en adventicia en el 40 % de los pacientes. El uso de estatinas con mayor frecuencia conduce a una presencia marcadamente mayor de MP Bcl-2+ en el tejido aórtico con aterosclerosis leve y grave diagnosticada. En muestras de aterosclerosis severa, las estatinas conducen a una presencia aumentada de Bcl-2+ MP en las partes de la placa hacia la íntima y hacia la media. Las estatinas conducen a una mayor presencia de macrófagos Bcl-2+, prolongan su vida, tanto en tejido aórtico sano como aterosclerótico alterado. Esto indica la potenciación de la inflamación y el daño a la pared aórtica y pone en duda el efecto positivo de las estatinas en la pared aórtica con aterosclerosis.
Asunto(s)
Humanos , Inhibidores de Hidroximetilglutaril-CoA Reductasas/administración & dosificación , Aterosclerosis/metabolismo , Aorta/efectos de los fármacos , Factores de Riesgo , Apoptosis/efectos de los fármacos , Medición de Riesgo , Genes bcl-2/fisiología , Aterosclerosis/tratamiento farmacológico , Proteína bcl-X/metabolismo , Placa Aterosclerótica , Macrófagos/efectos de los fármacosRESUMEN
OBJECTIVE: To investigate the expression of miR-195 and its target gene Bcl-2 in intervertebral disc degeneration (IVDD) and its effect on nucleus pulposus (NP) cell apoptosis. METHODS: The expressions of miR-195 and Bcl-2 in NP tissues of IVDD patients were quantified by qRT-PCR and western blotting, respectively. NP cells were divided into blank group, TNF-α group, TNF-α + miR-NC group, TNF-α + siBcl-2 group, and TNF-α + miR-195 inhibitors + siBcl-2 group. Cell proliferation was detected by MTT assay, cell apoptosis evaluated by flow cytometry, and mitochondrial membrane potential (MMP) tested by JC-1 staining. Moreover, the function of miR-195 on IVDD in vivo was investigated using a puncture-induced IVDD rat model. RESULTS: IVDD patients had significantly increased miR-195 expression and decreased Bcl-2 protein expression in NP tissues. The expression of miR-195 was negatively correlated with the expression of Bcl-2 in IVDD patients. Dual-luciferase reporter gene assay indicated that Bcl-2 was a target gene of miR-195. In comparison with blank group, TNF-α group showed decreased cell proliferation and MMP, increased cell apoptosis, upregulated expression of miR-195, Bax, and cleaved caspase 3, and downregulated Bcl-2 protein, while these changes were attenuated by miR-195 inhibitors. Additionally, siBcl-2 can reverse the protective effect of miR-195 inhibitors on TNF-α-induced NP cells. Besides, inhibition of miR-195 alleviated IVDD degeneration and NP cell apoptosis in the rat model. CONCLUSION: MiR-195 was significantly upregulated in NP tissues of IVDD patients, and inhibition of miR-195 could protect human NP cells from TNF-α-induced apoptosis via upregulation of Bcl-2.
Asunto(s)
Apoptosis/genética , Genes bcl-2/fisiología , Degeneración del Disco Intervertebral/genética , MicroARNs/metabolismo , Núcleo Pulposo/citología , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Humanos , RatasRESUMEN
BACKGROUND: When redox balance is lost in the brain, oxidative stress can cause serious damage that leads to neuronal loss, in congruence with neurodegenerative diseases. Aucubin (AU) is an iridoid glycoside and that is one of the active constituents of Eucommia ulmoides, has many pharmacological effects such as anti-inflammation, anti-liver fibrosis, and anti-atherosclerosis. PURPOSE: The present study aimed to evaluate the inhibitory effects of AU on cell oxidative stress against hydrogen peroxide (H2O2)-induced injury in SH-SY5Y cells in vitro. METHODS: SH-SY5Y cells were simultaneously treated with AU and H2O2 for 24 h. Cell viability was measured by CCK-8. Additionally, mitochondrial membrane depolarization, reactive oxygen species (ROS) generation, and cell apoptosis were measured by flow cytometry. RESULTS: The results showed that AU can significantly increase the H2O2-induced cell viability and the mitochondrial membrane potential, decrease the ROS generation, malondialdehyde (MDA), and increase glutathione (GSH) contents and the superoxide dismutase (SOD) activity. We also found that H2O2 stimulated the production of nitric oxide (NO), which could be reduced by treatment with AU through inhibiting the inducible nitric oxide synthase (iNOS) protein expression. In H2O2-induced SH-SY5Y cells, the levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-1ß (IL-1ß) content and cell apoptosis were significantly reduced by AU treatment through nuclear factor E2-related factor 2/hemo oxygenase-1 (Nrf2/HO-1) activation, inhibiting the expression of p-NF-κB/NF-κB and down-regulating MAPK and Bcl-2/Bax pathways. CONCLUSION: These results indicate that AU can reduce inflammation and oxidative stress through the NF-κB, Nrf2/HO-1, and MAPK pathways.
Asunto(s)
Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Hemo-Oxigenasa 1/metabolismo , Peróxido de Hidrógeno/toxicidad , Glucósidos Iridoides/farmacología , Factor 2 Relacionado con NF-E2/metabolismo , Fármacos Neuroprotectores/farmacología , Apoptosis/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Genes bcl-2/genética , Genes bcl-2/fisiología , Hemo-Oxigenasa 1/genética , Humanos , Quinasas de Proteína Quinasa Activadas por Mitógenos/genética , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Factor 2 Relacionado con NF-E2/genética , FN-kappa B/metabolismo , Neuroblastoma , Oxidación-Reducción , Estrés Oxidativo/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismoRESUMEN
The molecular mechanisms underlying the capability of pituitary tumours to avoid unregulated cell proliferation are still not well understood. However, the NF-κB transcription factor, which is able to modulate not only cellular senescence but also tumour progression, has emerged as a targeted candidate. This work was focused on the NF-κB role in cellular senescence during the progression of experimental pituitary tumours. Also, the contribution of the signalling pathways in senescence-associated NF-κB activation and the senescence-associated secretory phenotype (SASP) and pro-survival-NF-κB target genes transcription were analysed. A robust NF-κB activation was seen at E20-E40 of tumour development accompanied by a marked SA-ß-Gal co-reactivity in the tumour pituitary parenchyma. The induction of TNFα and IL1-ß as specific SASP-related NF-κB target genes as well as Bcl-2 and Bcl-xl pro-survival genes was shown to be accompanied by increases in the p-p38 MAPK protein levels, starting at the E20 stage and strengthening from 40 to 60 days of tumour growth. It is noteworthy that p-JNK displayed a similar pattern of activation during pituitary tumour development, while p-AKT and p-ERK1/2 were downregulated. By employing a pharmacological strategy to abrogate NF-κB activity, we demonstrated a marked reduction in SA-ß-Gal activity and a slight decrease in Ki67 immunopositive cells after NF-κB blockade. These results suggest a central role for NF-κB in the regulation of the cellular senescence programme, leading to the strikingly benign intrinsic nature of pituitary adenomas.
Asunto(s)
Senescencia Celular/genética , FN-kappa B/fisiología , Neoplasias Hipofisarias/genética , Transducción de Señal/genética , Animales , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Genes bcl-2/fisiología , Hipoxantina Fosforribosiltransferasa/metabolismo , Interleucina-1beta/metabolismo , Masculino , Ratas , Ratas Wistar , Factor de Necrosis Tumoral alfa/metabolismo , Proteína bcl-X/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
This study aimed to investigate the effect of miR-206 on apoptosis and autophagy of cardiomyocytes in ischaemia/reperfusion (I/R) injured rats treated with sevoflurane post-conditioning (SP) through the AMPK/Nampt signalling pathway. Rat models of myocardial I/R injury were established. The combination of SP, miR-206 inhibitor, AMPK activator AICAR and inhibitor Compound C was induced in rats, and their effects on I/R injury were determined with detection of malondialdehyde (MDA), hydroxyproline (HYP) and superoxide dismutase (SOD) levels in cardiomyocytes, autophagy, and apoptosis. We also conducted experiments to determine p62, Beclin1, Bax and Bcl-2 expression and the mRNA and expression pattern of AMPK/Nampt signalling pathway-related genes. Myocardial I/R injured rats revealed decreased SOD activity and elevated MDA content, autophagy and apoptosis. With the combined performance of SP, miR-206 inhibitor and AMPK activator AICAR, the rats presented higher SOD level and lower MDA and HYP levels, suppressed autophagy and apoptosis. Meanwhile, miR-206 inhibition also contributed to elevated expression of Nampt and the extent of AMPK phosphorylation, increased Bcl-2/Bax ratio, but degraded expression of p62 and Beclin1. Collectively, inhibition of miR-206 could activate the AMPK/Nampt signalling pathway, thus protecting against myocardial I/R injury on the basis of SP.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , MicroARNs/genética , Daño por Reperfusión Miocárdica/genética , Daño por Reperfusión Miocárdica/patología , Nicotinamida Fosforribosiltransferasa/metabolismo , Animales , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Beclina-1/biosíntesis , Genes bcl-2/fisiología , Miocitos Cardíacos/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Péptidos/metabolismo , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Sevoflurano/farmacología , Proteína X Asociada a bcl-2/biosíntesisRESUMEN
RATIONALE: Intracranial solitary fibrous tumor (ISFT) is a rare spindle cell tumor derived from dendritic mesenchymal cells expressing CD34 antigens, which are widely distributed in human connective tissues. PATIENT CONCERNS: In two case reports, we describe a 61-year-old woman and a 42-year-old man who present with intracranial malignant SFTs. Computed tomography or magnetic resonance imaging of head revealed that the largest size is about 3.3â×â3.0âcm in left occipital part and 4.0â×â3.0âcm in right skull base. DIAGNOSIS: Postoperative pathological results demonstrated that all of two cases are SFT. Case one: Immunohistochemical examination demonstrated a strong immunoreaction for cluster of differentiation (CD)34, B-cell lymphoma 2 (Bcl-2) and Vimentin (Vim). Case two: The tumor was distinctively positive for Bcl-2, but not for CD34 and Vim. INTERVENTIONS: One of the two patients recurred 6 years after the first tumor resection. After the recurrence, two gamma knife treatments were given, and another operation was performed about five years later. In one case, only tumor resection was performed. OUTCOMES: Case one: The postoperative neurological status was substantially improved and regular follow-up examinations for 6 months postsurgery have shown that the patient is currently disease-free. Case two: The patient achieved a good outcome, with no epilepsy or other neurological symptoms experienced on a regular 6-month follow-up. The patient is currently disease free. LESSONS: Imaging findings can be used to assist the diagnosis. The diagnostic method is pathology, and total surgical resection is the most effective treatment. The main treatment methods were total resection, supplemented by radiotherapy and chemotherapy if necessary.
Asunto(s)
Neoplasias Encefálicas/diagnóstico , Tumores Fibrosos Solitarios/diagnóstico , Adulto , Antígenos CD34/biosíntesis , Biomarcadores de Tumor , Neoplasias Encefálicas/diagnóstico por imagen , Neoplasias Encefálicas/inmunología , Femenino , Genes bcl-2/fisiología , Humanos , Masculino , Persona de Mediana Edad , Tumores Fibrosos Solitarios/diagnóstico por imagen , Tumores Fibrosos Solitarios/inmunología , Vimentina/biosíntesisRESUMEN
BACKGROUND: Subarachnoid hemorrhage (SAH) results in many brain dysfunctions and the neuroprotective function of puerarin after brain damage has been demonstrated in several studies. But whether puerarin can reduce brain nerve damage after SAH is not clear.In this study, we hypothesized that puerarin had the neuroprotective effect after SAH, and this protection could be mediated by bothBcl-2/Bax/Cleaved caspase-3 and SIRT3/SOD2 apoptotic signaling pathways. METHODS: First, we used neurological score, brain water content and so on to detect the neurological deficits after SAH. Then apoptosis neuron rate was detected by TUNEL staining. Western blot was carried out to explore the alteration of Blc-2, Bax, cleaved caspase-3 and Sirt3.Also, ROS acitivity and As-lysine level of SOD2 should be detected with assays. RESULTS: We demonstrate that puerarin attenuated the neurological deficits, effectively relieves cerebral edema, and reduce BBB disruption in SAH mice.And we revealed that a reduced rate of apoptosis neuron has been found out in puerarin treatment after SAH. In addition, obviously higher ratio of Blc-2/Bax and decreased expression of cleaved caspase-3 in puerarin-treated SAH micecomparing with vehicle-treated SAH animals had been found. Furthermore, puerarin effectively reversed these alterations in expression and inhibits ROSproduction induced by SAH. Also, puerarin can increase SOD activation after SAH and protect the expression of Sirt3 after SAH. CONCLUSIONS: In coclusion, puerarin can provide potential neuroprotection from the SAH damages, and can be act as a novel therapy for SAH.
Asunto(s)
Caspasa 3/metabolismo , Genes bcl-2/fisiología , Isoflavonas/uso terapéutico , Sirtuina 3/metabolismo , Hemorragia Subaracnoidea/metabolismo , Superóxido Dismutasa/metabolismo , Proteína X Asociada a bcl-2/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Genes bcl-2/efectos de los fármacos , Isoflavonas/farmacología , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedades del Sistema Nervioso/metabolismo , Enfermedades del Sistema Nervioso/prevención & control , Hemorragia Subaracnoidea/tratamiento farmacológico , Vasodilatadores/farmacología , Vasodilatadores/uso terapéuticoRESUMEN
BACKGROUND: Glioma is the most prevalent malignant tumor in human central nervous systems. Recently, the development of resistance to radiotherapy in glioma patients markedly vitiates the therapy outcome. MiR-153-3p has been reported to be closely correlated with tumor progression, but its effect and molecular mechanism underlying radioresistance remains unclear in glioma. METHODS: The expression of miR-153-3p was determined in radioresistant glioma clinical specimens as well as glioma cell lines exposed to irradiation (IR) using quantitative real-time PCR. Cell viability, proliferation and apoptosis were then evaluated by MTT assay, colony formation assay, Flow cytometry analysis and caspase-3 activity assay in glioma cells (U87 and U251). Tumor forming was evaluated by nude mice model in vivo. TUNEL staining was used to detect cell apoptosis in nude mice model. The target genes of miR-153-3p were predicted and validated using integrated bioinformatics analysis and a luciferase reporter assay. RESULTS: Here, we found that miR-153-3p was down-regulated in radioresistant glioma clinical specimens as well as glioma cell lines (U87 and U251) exposed to IR. Enhanced expression of miR-153-3p promoted the radiosensitivity, promoted apoptosis and elevated caspase-3 activity in glioma cells in vitro, as well as the radiosensitivity in U251 cell mouse xenografs in vivo. Mechanically, B cell lymphoma-2 gene (BCL2) was identified as the direct and functional target of miR-153-3p. Moreover, restoration of BCL2 expression reversed miR-153-3p-induced increase of radiosensitivity, apoptosis and caspase-3 activity in U251 cells in vitro. In addition, clinical data indicated that the expression of miR-153-3p was significantly negatively associated with BCL2 in radioresistance of glioma samples. CONCLUSIONS: Our findings suggest that miR-153-3p is a potential target to enhance the effect of radiosensitivity on glioma cells, thus representing a new potential therapeutic target for glioma.
Asunto(s)
Genes bcl-2/fisiología , Glioma/genética , MicroARNs/fisiología , MicroARNs/efectos de la radiación , Tolerancia a Radiación/genética , Adulto , Análisis de Varianza , Western Blotting , Caspasa 3/análisis , Línea Celular Tumoral , Proliferación Celular/efectos de la radiación , Supervivencia Celular/efectos de la radiación , Regulación hacia Abajo , Femenino , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica , Marcación de Gen/métodos , Genes bcl-2/efectos de la radiación , Glioma/radioterapia , Humanos , Etiquetado Corte-Fin in Situ , Masculino , MicroARNs/análisis , Persona de Mediana Edad , Reacción en Cadena en Tiempo Real de la Polimerasa , Factores de TiempoRESUMEN
OBJECTIVE: The aim of the study was to investigate the role of microRNA-21 (miR-21) in cardiomyocyte apoptosis and to determine a possible mechanism. METHODS: H9c2 embryonic rat heart-derived cells were used in the study. Cell viability was determined using the 3-(4.5-dimethyl-2-thiazolyl)-2,5- diphenyl-2-H-tetrazolium bromide (MTT) assay, and flow cytometry was used to evaluate cell apoptosis. Reverse transcription-polymerase chain reaction and western blot assays were used to detect mRNA and protein expression of the apoptosis-related proteins and miR-21. ELISA was used to detect reactive oxygen species (ROS). RESULTS: Palmitate exposure greatly reduced miR-21 expression in cardiomyocytes. Apoptosis increased when miR-21 was inhibited with or without palmitate exposure. Consistently, reduced apoptosis was observed when miR-21 was overexpressed in cardiomyocytes. Caspase-3 activity was reduced after palmitate exposure. Bcl-2 protein expression was increased in H9c2 cells when transfected with the miR-21 mimic. MiR-21 overexpression alone did not induce ROS or DNA fragmentation; however, in conjunction with palmitate exposure, miR-21 mimic reduced ROS and DNA fragmentation. Moreover, palmitate administration overcame the antioxidant effect of 3 mM N-acetylcysteine to significantly inhibit apoptosis, DNA fragmentation, and caspase-3 activity. The exposure to palmitate greatly reduced p65 and p-p38 expression in the nucleus. A p38 inhibitor had no effect on the expression of Bcl-2 and cleaved caspase-3 in H9c2 cells alone; however, when combined with exposure to palmitate the p38 inhibitor induced Bcl-2 expression and inhibited caspase-3 activity. The p38 inhibitor by itself did not induce apoptosis, ROS production, or DNA fragmentation in H9c2 cells, but when palmitate was included with the p38 inhibitor, apoptosis, ROS production, and DNA fragmentation were reduced. CONCLUSION: miR-21 protects cardiomyocytes from apoptosis that is induced by palmitate through the caspase-3/NF-κB signal pathways.
Asunto(s)
Apoptosis/fisiología , Caspasa 3/metabolismo , MicroARNs/fisiología , Miocitos Cardíacos/fisiología , FN-kappa B/metabolismo , Palmitatos/metabolismo , Acetilcisteína/farmacología , Animales , Antagomirs/metabolismo , Antagomirs/farmacología , Apoptosis/efectos de los fármacos , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Fragmentación del ADN/efectos de los fármacos , Regulación de la Expresión Génica , Genes bcl-2/fisiología , MicroARNs/antagonistas & inhibidores , MicroARNs/genética , MicroARNs/metabolismo , Palmitatos/farmacología , Ratas , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Factor de Transcripción ReIA/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
The current study examined the ameliorative effects of nano-elemental selenium (Nano-Se) against chromium-VI (K2Cr2O7)-induced apoptosis in chickens. The expression of apoptosis-related genes was evaluated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot. A total of 60, one-day-old broiler chickens allotted to six equal groups, i.e., control group (standard diet), Cr(VI)-exposed group (K2Cr2O7 via drinking water), Nano-Se group (Nano-Se at 0.5 mg/kg via diet), protection group (K2Cr2O7 + Nano-Se), cure group (K2Cr2O7 for initial 2 weeks and then Nano-Se), and prevention group (opposite to the cure group) and were detected by the activities of pro-apoptosis (Bax, Caspase-3) and anti-apoptosis (Bcl-2) genes expression at day 35 of the experiment. Intense apoptosis was observed in liver tissues of chickens exposed to K2Cr2O7. The Nano-Se supplementation caused a significant decrease (P < 0.01) in the mRNA expression levels of Bax and Caspase-3 genes, while significantly elevated (P < 0.05) mRNA expression level of Bcl-2 gene was observed in Nano-Se experimental groups as compare to control and Cr(VI)-exposed group. The results quantified by the RT-qPCR were further confirmed by the western blot analysis. Altogether, these results suggest anti-apoptotic effects of Nano-Se in the chicken liver, which is interesting for further study. The present findings suggested that Nano-Se has protective effects against K2Cr2O7-induced apoptosis in broilers liver and can serve a key role as a protective agent against apoptosis.
Asunto(s)
Apoptosis/efectos de los fármacos , Pollos/metabolismo , Cromo/toxicidad , Hígado/efectos de los fármacos , Dicromato de Potasio/toxicidad , Sustancias Protectoras/farmacología , Selenio/farmacología , Alimentación Animal/análisis , Animales , Proteínas Aviares/genética , Proteínas Aviares/metabolismo , Caspasa 3/genética , Caspasa 3/metabolismo , Pollos/genética , Dieta/veterinaria , Suplementos Dietéticos/análisis , Expresión Génica , Genes bcl-2/genética , Genes bcl-2/fisiología , Hígado/fisiopatología , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismoRESUMEN
Several studies have revealed that certain naturally occurring medicinal plants inhibit the growth of various cancers. The present study was conducted to evaluate cytotoxicity and apoptotic induction potential of Myristica fragrans Houtt mace extract. The cytotoxic activity of the Myristica fragrans Houtt mace acetone extract was assayed by MTT assay on human oral epidermal carcinoma KB cell lines. KB cells were incubated with different concentration of mace extract ranging from 25 to 125 µg/mL for 24hrs. The apoptotic induction potential was also studied by the analysis of Bcl-2 protein and gene expression in mace extract incubated KB cell lines using western blotting technique and real-time polymerase chain reaction. The mace extract exhibited cytotoxicity and anticancer effect against KB cell lines and it also suppressed the growth of cancer cells, therefore growth inhibitory effect was noted in extract treated cell lines. The apoptotic potential of mace extract was accompanied by reduced gene expression of Bcl-2 compared to the untreated KB cells. The mace extract shows the cytotoxic activity and induced the apoptosis through the modulation of its target genes Bcl-2 in the KB cell lines, suggesting the potential of mace as a candidate for oral cancer chemoprevention. This can be further investigated in vivo for its anticancer potential.
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Extractos Vegetales/análisis , Células KB , Myristica/anatomía & histología , Citotoxinas/análisis , Plantas Medicinales/clasificación , Preparaciones Farmacéuticas , Apoptosis , Genes bcl-2/fisiologíaRESUMEN
BACKGROUND: Glioma is the most prevalent malignant tumor in human central nervous systems. Recently, the development of resistance to radiotherapy in glioma patients markedly vitiates the therapy outcome. MiR-153-3p has been reported to be closely correlated with tumor progression, but its effect and molecular mechanism underlying radioresistance remains unclear in glioma. METHODS: The expression of miR-153-3p was determined in radioresistant glioma clinical specimens as well as glioma cell lines exposed to irradiation (IR) using quantitative real-time PCR. Cell viability, proliferation and apoptosis were then evaluated by MTT assay, colony formation assay, Flow cytometry analysis and caspase-3 activity assay in glioma cells (U87 and U251). Tumor forming was evaluated by nude mice model in vivo. TUNEL staining was used to detect cell apoptosis in nude mice model. The target genes of miR-153-3p were predicted and validated using integrated bioinformatics analysis and a luciferase reporter assay. RESULTS: Here, we found that miR-153-3p was down-regulated in radioresistant glioma clinical specimens as well as glioma cell lines (U87 and U251) exposed to IR. Enhanced expression of miR-153-3p promoted the radiosensitivity, promoted apoptosis and elevated caspase-3 activity in glioma cells in vitro, as well as the radiosensitivity in U251 cell mouse xenografs in vivo. Mechanically, B cell lymphoma-2 gene (BCL2) was identified as the direct and functional target of miR-153-3p. Moreover, restoration of BCL2 expression reversed miR-153-3p-induced increase of radiosensitivity, apoptosis and caspase-3 activity in U251 cells in vitro. In addition, clinical data indicated that the expression of miR-153-3p was significantly negatively associated with BCL2 in radioresistance of glioma samples. CONCLUSIONS: Our findings suggest that miR-153-3p is a potential target to enhance the effect of radiosensitivity on glioma cells, thus representing a new potential therapeutic target for glioma.
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Humanos , Masculino , Femenino , Adulto , Persona de Mediana Edad , Tolerancia a Radiación/genética , Genes bcl-2/fisiología , MicroARNs/efectos de la radiación , MicroARNs/fisiología , Glioma/genética , Factores de Tiempo , Regulación hacia Abajo , Regulación Neoplásica de la Expresión Génica , Supervivencia Celular/efectos de la radiación , Western Blotting , Análisis de Varianza , Marcación de Gen/métodos , Genes bcl-2/efectos de la radiación , Etiquetado Corte-Fin in Situ , MicroARNs/análisis , Línea Celular Tumoral , Proliferación Celular/efectos de la radiación , Caspasa 3/análisis , Reacción en Cadena en Tiempo Real de la Polimerasa , Citometría de Flujo , Glioma/radioterapiaRESUMEN
The present study aimed to investigate how bats protect their brain in a hypothermic situation. Formosan leaf-nosed bats (Hipposideros terasensis) were used in this study and treated under three conditions: room temperature (25±1°C), low temperature (4±1°C), and hibernation. The reactive oxygen species (ROS) levels in the blood and apoptosis-related proteins in the brain tissue were assessed and then compared among those bats under three conditions. Our results showed that the blood ROS levels of bats treated under conditions of low temperature and hibernation were significantly reduced compared with bats treated under the condition of room temperature. Both immunohistochemistry and immunoblotting expressions of hypoxia, inflammation, and apoptosis-related proteins in the brain tissue of bats treated under the condition of hibernation were significantly reduced compared with those bats treated under conditions of room temperature and low temperature. Thus, we suggested that bats can protect the brain in cold environment by reducing blood ROS levels and decreasing expressions of hypoxia, inflammation, and apoptosis-related proteins in the brain. Possible protection mechanisms involved in hypothermic adaptations need to be further clarified.
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Encéfalo/metabolismo , Quirópteros/fisiología , Frío , Hibernación/fisiología , Neuroprotección/fisiología , Animales , Apoptosis/fisiología , Encéfalo/citología , Calpaína/metabolismo , Caspasa 12/metabolismo , Caspasa 3/metabolismo , Ecolocación , Genes bcl-2/fisiología , Hipoxia/metabolismo , Hipoxia/patología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Inmunohistoquímica , Inflamación/metabolismo , Inflamación/patología , Especies Reactivas de Oxígeno/sangre , Factor de Necrosis Tumoral alfa/metabolismo , Proteína X Asociada a bcl-2/metabolismoRESUMEN
BACKGROUND: Apoptosis plays a fundamental role in appropriate tissue development and function. Although expression of Bcl-2 has been reported during tooth and submandibular gland (SMG) development, the physiological role Bcl-2 plays during these processes has not been addressed. This study was performed to evaluate the impact of Bcl-2 expression on the formation and properties of tooth hard tissue, and saliva production. METHODS: Twenty-four mice (12 males and 12 females) were divided into three groups of eight (n=8): group A (Bcl-2 +/+), group B (Bcl-2 +/-), and group C (Bcl-2 -/-) and subjected to micro-CT analyses. The mineral content of first molars was analyzed by X-Ray diffraction (XRD) and scanning electron microscopy (SEM) color dot map. The surface microhardness was determined by Vickers test on labial surfaces of incisors. Saliva was collected from different groups of mice after subcutaneous injection of pilocarpine. RESULTS: Samples from Bcl-2 -/- mice showed significantly smaller micro-CT values, lower and poor crystallinity of hydroxyapatite (HA), and lowest surface micro hardness. SMG from Bcl-2 -/- mice showed remarkable reduction in size, consistent with reduced saliva accumulation. CONCLUSIONS: The absence of Bcl-2 expression in SMG did not affect the expression of other Bcl-2 family members. Thus, Bcl-2 expression influence on the formation and properties of tooth hard tissue, and saliva accumulation. SIGNIFICANCE: Bcl-2 expression has a significant impact on the mineralogical content of enamel crystals of tooth structure. Lack of Bcl-2 expression led to impaired production of enamel ACP crystals.
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Genes bcl-2/fisiología , Saliva/metabolismo , Diente/metabolismo , Animales , Esmalte Dental/metabolismo , Femenino , Dureza/fisiología , Masculino , Ratones , Microscopía Electrónica de Rastreo/métodos , Propiedades de Superficie , TemperaturaRESUMEN
Acupuncture has been widely used as a treatment for stroke in China for a long time. Recently, studies have demonstrated that electroacupuncture (EA) can accelerate intracerebral hemorrhage (ICH)-induced angiogenesis in rats. In the present study, we investigated the effect of EA on neurobehavioral function and brain injury in ICH rats. ICH was induced by stereotactic injection of collagenase type I and heparin into the right caudate putamen. Adult ICH rats were randomly divided into the following three groups: model control group (MC), EA at non-acupoint points group (non-acupoint EA) and EA at Baihui and Dazhui acupoints group (EA). The neurobehavioral deficits of ICH rats were assessed by modified neurological severity score (mNSS) and gait analysis. The hemorrhage volume and glucose metabolism of hemorrhagic foci were detected by PET/CT. The expression levels of MBP, NSE and S100-B proteins in serum were tested by ELISA. The histopathological features were examined by haematoxylin-eosin (H&E) staining. Apoptosis-associated proteins in the perihematomal region were observed by immunohistochemistry. EA treatment significantly promoted the recovery of neurobehavioral function in ICH rats. Hemorrhage volume reduced in EA group at day 14 when compared with MC and non-acupoint EA groups. ELISA showed that the levels of MBP, NSE and S100-B in serum were all down-regulated by EA treatment. The brain tissue of ICH rat in the EA group was more intact and compact than that in the MC and non-acupoint groups. In the perihematomal regions, the expression of Bcl-2 protein increased and expressions of Caspase-3 and Bax proteins decreased in the EA group vs MC and non-acupoint EA groups. Our data suggest that EA treatment can improve neurobehavioral function and brain injury, which were likely connected with the absorption of hematoma and regulation of apoptosis-related proteins.
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Hemorragia Cerebral/rehabilitación , Hemorragia Cerebral/terapia , Electroacupuntura/métodos , Puntos de Acupuntura , Terapia por Acupuntura/métodos , Animales , Encéfalo/metabolismo , Lesiones Encefálicas/metabolismo , Caspasa 3/metabolismo , Hemorragia Cerebral/metabolismo , Modelos Animales de Enfermedad , Marcha/efectos de los fármacos , Marcha/fisiología , Genes bcl-2/fisiología , Masculino , Fármacos Neuroprotectores/metabolismo , Tomografía Computarizada por Tomografía de Emisión de Positrones , Ratas , Ratas Sprague-Dawley , Proteína X Asociada a bcl-2/metabolismoRESUMEN
BACKGROUND Laryngeal cancer is a malignant head and neck tumor with high morbidity and high mortality in humans. Recently, treatments with Chinese medicines and their extracts have gradually received great attention, and studies suggest that Boschniakia rossica polysaccharide (BRP) has potential anti-tumor activity. Therefore, this study investigating the role of BRP in inducing apoptosis in human laryngeal carcinoma cells. MATERIAL AND METHODS The BRP was extracted with organic solvent and HR column. We treated Hep2 laryngeal carcinoma cells with different concentrations of BRP, then assessed cell growth inhibition rate by flow cytometry and apoptosis index by TUNEL staining. The protein expression of p53, Bcl-2, Bax, and caspase-3 were analyzed by Western blot. RESULTS Flow cytometry results showed that BRP inhibited Hep2 cell proliferation in a dose-dependent manner (p<0.05), and TUNEL staining indicated that BRP significantly increased Hep2 apoptosis index (p<0.05). Western blot results showed that the expression levels of p53 and activation of caspase-3 in Hep2 cells were significantly up-regulated (p<0.05), while the expression of Bcl-2 was significantly down-regulated (p<0.05). CONCLUSIONS BRP might induce cell apoptosis by regulating the expression level of cell apoptosis-associated proteins, suggesting strong anti-laryngeal cancer activity.
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Neoplasias Laríngeas/tratamiento farmacológico , Orobanchaceae/toxicidad , Polisacáridos/uso terapéutico , Apoptosis/fisiología , Caspasa 3/efectos de los fármacos , Caspasa 3/metabolismo , Caspasa 3/fisiología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Genes bcl-2/efectos de los fármacos , Genes bcl-2/fisiología , Humanos , Neoplasias Laríngeas/metabolismo , Neoplasias Laríngeas/patología , Laringe/patología , Medicina Tradicional China , Orobanchaceae/metabolismo , Proteína p53 Supresora de Tumor/efectos de los fármacos , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
AIMS: Acute myeloid leukemia (AML) is an immunophenotypically heterogenous malignant disease. The early immature CD34+ AML cell subpopulation is frequently impervious to intensive chemotherapy, making them largely responsible for relapse of AML. CD34+ AML cells have higher level of Bcl-2 protein expression than the CD34- subpopulation. As such, development of drugs that specifically target the Bcl-2 may have the potential to eliminate immature CD34+ AML progenitor cells and provide therapeutic benefit. In this work, we made an attempt to investigate the cytotoxic effect of a novel Bcl-2 family inhibitor, ABT-737, on CD34+ AML cell lines (KG1a and Kasumi-1) as well as CD34+ primary AML cells. METHODS: Primary human CD34+ cells were isolated from bone marrow mononuclear cells using CD34 MicroBead kit. The growth inhibitory effect was measured by cell counting kit-8. Apoptosis was analyzed by annexin V/PI assays. Protein expression was determined by Western blotting analysis. RESULTS: Inhibition of Bcl-2 by ABT-737 effectively inhibited growth and induced apoptosis in CD34+ AML cell lines and CD34+ primary AML cells without affecting CD34+ normal hematopoietic cells. Furthermore, Western blot analysis showed that ABT-737 induced apoptosis associated with caspase-3 activation and poly ADP-ribose polymerase (PARP) degradation. Finally, ABT-737 synergistically enhanced the cytotoxic effect of cytarabine and daunorubicin in CD34+ AML cells. CONCLUSION: Taken together, these findings indicate that ABT-737 may offer as a promising molecular targeting agent in CD34+ AML.
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Apoptosis/efectos de los fármacos , Compuestos de Bifenilo/metabolismo , Genes bcl-2/fisiología , Leucemia Mieloide Aguda/metabolismo , Nitrofenoles/metabolismo , Sulfonamidas/metabolismo , Adulto , Sinergismo Farmacológico , Femenino , Humanos , Leucemia Mieloide Aguda/patología , Masculino , Persona de Mediana Edad , Piperazinas/metabolismo , Adulto JovenRESUMEN
White syndrome has been described as one of the most prolific diseases on the Great Barrier Reef. Previously, apoptotic cell death has been described as the mechanism driving the characteristic rapid tissue loss associated with this disease, but the molecular mechanisms controlling apoptotic cell death in coral disease have yet to be investigated. In situ methods were used to study the expression patterns of 2 distinct regulators of apoptosis in Acropora hyacinthus tissues undergoing white syndrome and apoptotic cell death. Apoptotic genes within the Bcl-2 family were not localized in apparently healthy coral tissues. However, a Bcl-2 family member (bax-like) was found to localize to cells and tissues affected by white syndrome and those with morphological evidence for apoptosis. A potential up-regulation of pro-apoptotic or bax-like gene expression in tissues with apoptotic cell death adjacent to disease lesions is consistent with apoptosis being the primary cause of rapid tissue loss in coral affected by white syndrome. Pro-apoptotic (bax-like) expression in desmocytes and the basal tissue layer, the calicodermis, distant from the disease lesion suggests that apoptosis may also underlie the sloughing of healthy tissues associated with the characteristic, rapid spread of tissue loss, evident of this disease. This study also shows that in situ hybridisation is an effective tool for studying gene expression in adult corals, and wider application of these methods should allow a better understanding of many aspects of coral biology and disease pathology.