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1.
Mol Psychiatry ; 28(10): 4474-4484, 2023 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-37648779

RESUMEN

Mitochondrial dysfunction has been implicated in Parkinson's Disease (PD) progression; however, the mitochondrial factors underlying the development of PD symptoms remain unclear. One candidate is CR6-interacting factor1 (CRIF1), which controls translation and membrane insertion of 13 mitochondrial proteins involved in oxidative phosphorylation. Here, we found that CRIF1 mRNA and protein expression were significantly reduced in postmortem brains of elderly PD patients compared to normal controls. To evaluate the effect of Crif1 deficiency, we produced mice lacking the Crif1 gene in dopaminergic neurons (DAT-CRIF1-KO mice). From 5 weeks of age, DAT-CRIF1-KO mice began to show decreased dopamine production with progressive neuronal degeneration in the nigral area. At ~10 weeks of age, they developed PD-like behavioral deficits, including gait abnormalities, rigidity, and resting tremor. L-DOPA, a medication used to treat PD, ameliorated these defects at an early stage, although it was ineffective in older mice. Taken together, the observation that CRIF1 expression is reduced in human PD brains and deletion of CRIF1 in dopaminergic neurons leads to early-onset PD with stepwise PD progression support the conclusion that CRIF1-mediated mitochondrial function is important for the survival of dopaminergic neurons.


Asunto(s)
Neuronas Dopaminérgicas , Enfermedad de Parkinson , Humanos , Ratones , Animales , Anciano , Neuronas Dopaminérgicas/metabolismo , Enfermedad de Parkinson/genética , Levodopa/farmacología , Dopamina/metabolismo , Encéfalo/metabolismo , Proteínas de Ciclo Celular/genética
2.
Immunity ; 43(1): 80-91, 2015 Jul 21.
Artículo en Inglés | MEDLINE | ID: mdl-26200012

RESUMEN

The orphan nuclear receptor estrogen-related receptor α (ERRα; NR3B1) is a key metabolic regulator, but its function in regulating inflammation remains largely unknown. Here, we demonstrate that ERRα negatively regulates Toll-like receptor (TLR)-induced inflammation by promoting Tnfaip3 transcription and fine-tuning of metabolic reprogramming in macrophages. ERRα-deficient (Esrra(-/-)) mice showed increased susceptibility to endotoxin-induced septic shock, leading to more severe pro-inflammatory responses than control mice. ERRα regulated macrophage inflammatory responses by directly binding the promoter region of Tnfaip3, a deubiquitinating enzyme in TLR signaling. In addition, Esrra(-/-) macrophages showed an increased glycolysis, but impaired mitochondrial respiratory function and biogenesis. Further, ERRα was required for the regulation of NF-κB signaling by controlling p65 acetylation via maintenance of NAD(+) levels and sirtuin 1 activation. These findings unravel a previously unappreciated role for ERRα as a negative regulator of TLR-induced inflammatory responses through inducing Tnfaip3 transcription and controlling the metabolic reprogramming.


Asunto(s)
Cisteína Endopeptidasas/biosíntesis , Inflamación/inmunología , Péptidos y Proteínas de Señalización Intracelular/biosíntesis , Macrófagos/metabolismo , Receptores de Estrógenos/genética , Receptor Toll-Like 4/inmunología , Acetilación , Animales , Calcio/metabolismo , Quinasa de la Proteína Quinasa Dependiente de Calcio-Calmodulina/metabolismo , Células Cultivadas , Cisteína Endopeptidasas/genética , Activación Enzimática/genética , Glucólisis/genética , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Péptidos y Proteínas de Señalización Intracelular/genética , Lipopolisacáridos , Macrófagos/inmunología , Ratones , Ratones Noqueados , Factor 88 de Diferenciación Mieloide/metabolismo , NAD/metabolismo , Fosforilación Oxidativa , Regiones Promotoras Genéticas/genética , Regiones Promotoras Genéticas/inmunología , Choque Séptico/inmunología , Transducción de Señal , Sirtuina 1/metabolismo , Factor 6 Asociado a Receptor de TNF/metabolismo , Factor de Transcripción ReIA/metabolismo , Transcripción Genética/genética , Proteína 3 Inducida por el Factor de Necrosis Tumoral alfa , Ubiquitinación , Receptor Relacionado con Estrógeno ERRalfa
3.
Int J Mol Sci ; 25(3)2024 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-38338992

RESUMEN

Knee osteoarthritis (OA), an age-related degenerative disease characterized by severe pain and disability, is treated using polynucleotides (PNs) and hyaluronic acid (HA). The intra-articular (IA) injection of HA has been studied extensively in both animal models and in humans; however, the efficacy and mechanisms of action remain unclear. In addition, there has been a paucity of research regarding the use of PN alone or in combination with HA in OA. To investigate the effect of the combined injection of PN and HA in vivo, pathological and behavioral changes were assessed in an OA model. Anterior cruciate ligament transection and medial meniscectomy were performed in Sprague-Dawley rats to create the OA animal model. The locomotor activity improved following PNHA injection, while the OARSI grade improved in the medial tibia and femur. In mild OA, TNFα levels decreased histologically in the PN, HA, and PNHA groups but only the PNHA group showed behavioral improvement in terms of distance. In conclusion, PNHA exhibited anti-inflammatory effects during OA progression and improved locomotor activity regardless of the OARSI grade.


Asunto(s)
Ácido Hialurónico , Osteoartritis de la Rodilla , Ratas , Humanos , Animales , Ácido Hialurónico/farmacología , Polinucleótidos/farmacología , Polinucleótidos/uso terapéutico , Ratas Sprague-Dawley , Osteoartritis de la Rodilla/tratamiento farmacológico , Ligamento Cruzado Anterior/cirugía , Inyecciones Intraarticulares
4.
Pharmacol Res ; 184: 106423, 2022 10.
Artículo en Inglés | MEDLINE | ID: mdl-36064078

RESUMEN

BMP2 is clinically used as an ectopic bone inducer and plays a significant role in bone development, formation, and diseases. Chitinase 3-like 1 protein (Chi3L1) is found in the skeletal system. However, Chi3L1-mediated bone metabolism and aging-related bone erosion via BMP2 signaling have not yet been demonstrated. Herein, Chi3L1 increased BMP2-induced osteoblast differentiation in mesenchymal precursor cells and human primary osteoblasts. Chi3L1KO(-/-) showed abnormal bone development, and primary osteoblasts isolated from Chi3L1KO(-/-) exhibited impaired osteoblast differentiation and maturation. Chi3L1 also potentiated BMP2 signaling and RUNX2 expression in primary osteoblasts. Chi3L1 interacted with BMPRIa, which increased the surface expression of BMPRIa and promoted BMP2 signaling to induce osteoblast differentiation. Chi3L1KO(-/-) mice showed bone formation reduced with a decrease in RUNX2 expression in calvarial defects. Chi3L1KO(-/-) mice exhibited aging-related osteoporotic bone loss with decreases in the levels of RUNX2 and OPG, while serum PYD level and osteoclast number increased. Chi3L1 increased OPG via non-canonical BMP2 signaling in osteoblasts, which suppressed osteoclastogenesis in BMMs. Furthermore, ROC analysis showed that serum Chi3L1 level clinically decreased in osteoporosis patients. Our findings demonstrate that Chi3L1 promotes bone formation, suppresses osteoclastogenesis, and prevents aging-related osteoporosis.


Asunto(s)
Quitinasas , Osteoporosis , Animales , Biomarcadores/metabolismo , Diferenciación Celular , Proteína 1 Similar a Quitinasa-3/genética , Proteína 1 Similar a Quitinasa-3/metabolismo , Quitinasas/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Subunidad alfa 1 del Factor de Unión al Sitio Principal/metabolismo , Humanos , Ratones , Osteoblastos/metabolismo , Osteogénesis , Osteoporosis/tratamiento farmacológico , Osteoporosis/metabolismo
5.
J Neurochem ; 156(1): 76-87, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-32639632

RESUMEN

While recent studies strongly suggest that a single, short anesthetic exposure does not affect neurodevelopment, the effects of multiple exposures remain unclear. Unfortunately, studying "multiple exposures" is challenging as it is an extremely heterogeneous descriptor comprising diverse factors. One potentially important, but unrecognized factor is the interval between anesthetic exposures. In order to evaluate the significance of interval, we exposed post-natal day 16, 17 mice to three sevoflurane exposures (2.5%, 1 hr) with short (2 hr) or long (24 hr) intervals. Changes in synaptic transmission, plasticity, protein expression, and behavior were assessed in male and female mice. We discovered that short-interval exposures induced a female-dependent decrease in miniature inhibitory post-synaptic current (mIPSC) frequency 5 days after the last exposure (control: 18.44 ± 2.86 Hz, sevoflurane:14.65 ± 4.54 Hz). Short-interval sevoflurane exposed mice also displayed long-term behavioral deficits at adult age (hypoactivity, anxiety). These behavioral changes were consistent with the sex-dependent changes in inhibitory transmission, as they were more robust in female mice. Although there was no change in learning and memory, short-interval sevoflurane exposures also impaired LTP in a non-sex-dependent manner (control: 171.10 ± 26.90%, sevoflurane: 149.80 ± 26.48 %). Most importantly, we were unable to find long-lasting consequences in mice that received long-interval sevoflurane exposures. Our study provides novel insights regarding the significance of the interval between multiple exposures, and also suggests that the neurotoxic effects of multiple anesthetic exposures may be reduced by simply increasing the interval between each exposure.


Asunto(s)
Anestésicos por Inhalación/toxicidad , Conducta Animal/efectos de los fármacos , Plasticidad Neuronal/efectos de los fármacos , Sevoflurano/toxicidad , Transmisión Sináptica/efectos de los fármacos , Anestésicos por Inhalación/administración & dosificación , Animales , Animales Recién Nacidos , Encéfalo/efectos de los fármacos , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Sevoflurano/administración & dosificación , Caracteres Sexuales
6.
Int J Mol Sci ; 22(7)2021 Mar 26.
Artículo en Inglés | MEDLINE | ID: mdl-33810547

RESUMEN

The hypothalamic regulation of appetite governs whole-body energy balance. Satiety is regulated by endocrine factors including leptin, and impaired leptin signaling is associated with obesity. Despite the anorectic effect of leptin through the regulation of the hypothalamic feeding circuit, a distinct downstream mediator of leptin signaling in neuron remains unclear. Angiopoietin-like growth factor (AGF) is a peripheral activator of energy expenditure and antagonizes obesity. However, the regulation of AGF expression in brain and localization to mediate anorectic signaling is unknown. Here, we demonstrated that AGF is expressed in proopiomelanocortin (POMC)-expressing neurons located in the arcuate nucleus (ARC) of the hypothalamus. Unlike other brain regions, hypothalamic AGF expression is stimulated by leptin-induced signal transducers and activators of transcription 3 (STAT3) phosphorylation. In addition, leptin treatment to hypothalamic N1 cells significantly enhanced the promoter activity of AGF. This induction was abolished by the pretreatment of ruxolitinib, a leptin signaling inhibitor. These results indicate that hypothalamic AGF expression is induced by leptin and colocalized to POMC neurons.


Asunto(s)
Proteínas Similares a la Angiopoyetina/genética , Proteínas Similares a la Angiopoyetina/metabolismo , Hipotálamo/metabolismo , Leptina/metabolismo , Transducción de Señal , Proteína 6 similar a la Angiopoyetina , Animales , Núcleo Arqueado del Hipotálamo/metabolismo , Encéfalo/metabolismo , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/metabolismo , Fosforilación , Proopiomelanocortina/metabolismo , Factor de Transcripción STAT3/metabolismo
7.
Lab Invest ; 99(9): 1389-1399, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-31043679

RESUMEN

High-mobility group box 1 (HMGB1) is actively secreted from inflammatory cells and acts via a non-cell-autonomous mechanism to play an important role in mediating cell proliferation and migration. The HMGB1-RAGE (receptor for advanced glycation end products) axis upregulates tyrosine hydroxylase (TH) expression in response to extracellular insults in dopaminergic neurons in vitro, but little is known about HMGB1 in modulation of dopaminergic neurons in vivo. Here, using immunohistochemistry, we show that HMGB1 and RAGE expression are higher in the nigral area of MPTP (methyl-4-phenyl-1,2,3,6-tetrahydropyridine)-treated mice, a toxin-induced Parkinsonian mouse model, compared with saline-treated controls. HMGB1 was predominantly localized to astrocytes and may affect neighboring dopaminergic neurons in the MPTP mouse model, owing to co-localization of RAGE in these TH-positive cells. In addition, MPTP induced a decrease in TH expression, an effect that was potentiated by inhibition of c-Jun N-terminal kinase (JNK) or RAGE. Moreover, stereotaxic injection of recombinant HMGB1 attenuated the MPTP-induced reduction of TH in a Parkinsonian mouse model. Collectively, our results suggest that an increase of HMGB1, released from astrocytes, upregulates TH expression in an acute MPTP-induced Parkinsonian mouse model, thereby maintaining dopaminergic neuronal functions.


Asunto(s)
Astrocitos/metabolismo , Proteína HMGB1/metabolismo , Trastornos Parkinsonianos/metabolismo , Tirosina 3-Monooxigenasa/metabolismo , 1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/efectos adversos , Animales , Masculino , Ratones , Ratones Endogámicos C57BL , Trastornos Parkinsonianos/inducido químicamente , Receptor para Productos Finales de Glicación Avanzada/metabolismo
9.
Int J Mol Sci ; 20(14)2019 Jul 11.
Artículo en Inglés | MEDLINE | ID: mdl-31336718

RESUMEN

Current therapeutics for Parkinson's disease (PD) are only effective in providing relief of symptoms such as rigidity, tremors and bradykinesia, and do not exert disease-modifying effects by directly modulating mitochondrial function. Here, we investigated auraptene (AUR) as a potent therapeutic reagent that specifically protects neurotoxin-induced reduction of mitochondrial respiration and inhibits reactive oxygen species (ROS) generation. Further, we explored the mechanism and potency of AUR in protecting dopaminergic neurons. Treatment with AUR significantly increased the viability of substantia nigra (SN)-derived SN4741 embryonic dopaminergic neuronal cells and reduced rotenone-induced mitochondrial ROS production. By inducing antioxidant enzymes AUR treatment also increased oxygen consumption rate. These results indicate that AUR exerts a protective effect against rotenone-induced mitochondrial oxidative damage. We further assessed AUR effects in vivo, investigating tyrosine hydroxylase (TH) expression in the striatum and substantia nigra of MPTP-induced PD model mice and behavioral changes after injection of AUR. AUR treatment improved movement, consistent with the observed increase in the number of dopaminergic neurons in the substantia nigra. These results demonstrate that AUR targets dual pathogenic mechanisms, enhancing mitochondrial respiration and attenuating ROS production, suggesting that the preventative potential of this natural compound could lead to improvement in PD-related neurobiological changes.


Asunto(s)
Respiración de la Célula/efectos de los fármacos , Cumarinas/farmacología , Depuradores de Radicales Libres/farmacología , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Animales , Conducta Animal/efectos de los fármacos , Biomarcadores , Cumarinas/química , Modelos Animales de Enfermedad , Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/metabolismo , Depuradores de Radicales Libres/química , Expresión Génica , Ratones , Modelos Biológicos , Oxidación-Reducción/efectos de los fármacos , Enfermedad de Parkinson/tratamiento farmacológico , Enfermedad de Parkinson/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Tirosina 3-Monooxigenasa/genética , Tirosina 3-Monooxigenasa/metabolismo
10.
Biochem Biophys Res Commun ; 493(1): 358-364, 2017 11 04.
Artículo en Inglés | MEDLINE | ID: mdl-28887039

RESUMEN

The derangement of tyrosine hydroxylase (TH) activity reduces dopamine synthesis and is implicated in the pathogenesis of Parkinson's disease. However, the extracellular modulator and intracellular regulatory mechanisms of TH have yet to be identified. Recently, high-mobility group box 1 (HMGB1) was reported to be actively secreted from glial cells and is regarded as a mediator of dopaminergic neuronal loss. However, the mechanism for how HMGB1 affects TH expression, particularly through the receptor for advanced glycation endproducts (RAGE), has not yet been investigated. We found that recombinant HMGB1 (rHMGB1) upregulates TH mRNA expression via simultaneous activation of JNK phosphorylation, and this induction of TH expression is blocked by inhibitors of RAGE and JNK. To investigate how TH expression levels change through the HMGB1-RAGE axis as a result of MPP+ toxicity, we co-treated SN4741 dopaminergic cells with MPP+ and rHMGB1. rHMGB1 blocked the reduction of TH mRNA following MPP+ treatment without altering cell survival rates. Our results suggest that HMGB1 upregulates TH expression to maintain dopaminergic neuronal function via activating RAGE, which is dependent on JNK phosphorylation.


Asunto(s)
Neuronas Dopaminérgicas/fisiología , Proteína HMGB1/metabolismo , MAP Quinasa Quinasa 4/metabolismo , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Transducción de Señal/fisiología , Tirosina 3-Monooxigenasa/metabolismo , Animales , Línea Celular , Fosforilación , Ratas , Regulación hacia Arriba/fisiología
11.
Anesthesiology ; 126(2): 288-299, 2017 02.
Artículo en Inglés | MEDLINE | ID: mdl-27922840

RESUMEN

BACKGROUND: Anesthesia during the synaptogenic period induces dendritic spine formation, which may affect neurodevelopment. The authors, therefore, evaluated whether changes in synaptic transmission after dendritic spine formation induced by sevoflurane were associated with long-term behavioral changes. The effects of sevoflurane on mitochondrial function were also assessed to further understand the mechanism behind spinogenesis. METHODS: Postnatal day 16 to 17 mice were exposed to sevoflurane (2.5% for 2 h), and synaptic transmission was measured in the medial prefrontal cortex 6 h or 5 days later. The expression of postsynaptic proteins and mitochondrial function were measured after anesthesia. Long-term behavioral changes were assessed in adult mice. RESULTS: Sevoflurane increased the expression of excitatory postsynaptic proteins in male and female mice (n = 3 to 5 per group). Sevoflurane exposure in male mice transiently increased miniature excitatory postsynaptic current frequency (control: 8.53 ± 2.87; sevoflurane: 11.09 ± 2.58) but decreased miniature inhibitory postsynaptic current frequency (control: 10.18 ± 4.66; sevoflurane: 6.88 ± 2.15). Unexpectedly, sevoflurane increased miniature inhibitory postsynaptic current frequency (control: 1.81 ± 1.11; sevoflurane: 3.56 ± 1.74) in female mice (neurons, n = 10 to 21 per group). Sevoflurane also increased mitochondrial respiration in male mice (n = 5 to 8 per group). However, such changes from anesthesia during the critical period did not induce long-term behavioral consequences. Values are presented as mean ± SD. CONCLUSIONS: Sevoflurane exposure during the critical period induces mitochondrial hyperactivity and transient imbalance of excitatory/inhibitory synaptic transmission, without long-lasting behavioral consequences. Further studies are needed to confirm sexual differences and to define the role of mitochondrial activity during anesthesia-induced spine formation.


Asunto(s)
Anestésicos por Inhalación/farmacología , Conducta Animal/efectos de los fármacos , Éteres Metílicos/farmacología , Mitocondrias/efectos de los fármacos , Transmisión Sináptica/efectos de los fármacos , Animales , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Sevoflurano , Factores Sexuales
12.
PLoS Genet ; 9(3): e1003356, 2013.
Artículo en Inglés | MEDLINE | ID: mdl-23516375

RESUMEN

Impaired mitochondrial oxidative phosphorylation (OXPHOS) has been proposed as an etiological mechanism underlying insulin resistance. However, the initiating organ of OXPHOS dysfunction during the development of systemic insulin resistance has yet to be identified. To determine whether adipose OXPHOS deficiency plays an etiological role in systemic insulin resistance, the metabolic phenotype of mice with OXPHOS-deficient adipose tissue was examined. Crif1 is a protein required for the intramitochondrial production of mtDNA-encoded OXPHOS subunits; therefore, Crif1 haploinsufficient deficiency in mice results in a mild, but specific, failure of OXPHOS capacity in vivo. Although adipose-specific Crif1-haploinsufficient mice showed normal growth and development, they became insulin-resistant. Crif1-silenced adipocytes showed higher expression of chemokines, the expression of which is dependent upon stress kinases and antioxidant. Accordingly, examination of adipose tissue from Crif1-haploinsufficient mice revealed increased secretion of MCP1 and TNFα, as well as marked infiltration by macrophages. These findings indicate that the OXPHOS status of adipose tissue determines its metabolic and inflammatory responses, and may cause systemic inflammation and insulin resistance.


Asunto(s)
Tejido Adiposo , Proteínas de Ciclo Celular , Inflamación , Resistencia a la Insulina/genética , Obesidad , Adipocitos/citología , Adipocitos/metabolismo , Tejido Adiposo/citología , Tejido Adiposo/metabolismo , Animales , Proteínas de Ciclo Celular/deficiencia , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Haploinsuficiencia , Inflamación/metabolismo , Inflamación/patología , Insulina/genética , Insulina/metabolismo , Macrófagos/citología , Macrófagos/metabolismo , Ratones , Mitocondrias/metabolismo , Obesidad/metabolismo , Obesidad/patología , Fosforilación Oxidativa
13.
Diabetologia ; 58(4): 771-80, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25660120

RESUMEN

AIM/HYPOTHESIS: Although mitochondrial oxidative phosphorylation (OxPhos) dysfunction is believed to be responsible for beta cell dysfunction in insulin resistance and mitochondrial diabetes, the mechanisms underlying progressive beta cell failure caused by defective mitochondrial OxPhos are largely unknown. METHODS: We examined the in vivo phenotypes of beta cell dysfunction in beta cell-specific Crif1 (also known as Gadd45gip1)-deficient mice. CR6-interacting factor-1 (CRIF1) is a mitochondrial protein essential for the synthesis and formation of the OxPhos complex in the inner mitochondrial membrane. RESULTS: Crif1(beta-/-) mice exhibited impaired glucose tolerance with defective insulin secretion as early as 4 weeks of age without defects in islet structure. At 11 weeks of age, Crif1(beta-/-) mice displayed characteristic ultrastructural mitochondrial abnormalities as well as severe glucose intolerance. Furthermore, islet area and insulin content was decreased by approximately 50% compared with wild-type mice. Treatment with the glucoregulatory drug exenatide, a glucagon-like peptide-1 (GLP-1) agonist, was not sufficient to preserve beta cell function in Crif1(beta-/-) mice. CONCLUSIONS/INTERPRETATION: Our results indicate that mitochondrial OxPhos dysfunction triggers progressive beta cell failure that is not halted by treatment with a GLP-1 agonist. The Crif1(beta-/-) mouse is a useful model for the study of beta cell failure caused by mitochondrial OxPhos dysfunction.


Asunto(s)
Proteínas de Ciclo Celular/deficiencia , Diabetes Mellitus/metabolismo , Células Secretoras de Insulina/metabolismo , Mitocondrias/metabolismo , Fosforilación Oxidativa , Factores de Edad , Animales , Autofagia , Glucemia/metabolismo , Proteínas de Ciclo Celular/genética , Línea Celular , Diabetes Mellitus/tratamiento farmacológico , Diabetes Mellitus/genética , Diabetes Mellitus/patología , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Exenatida , Genotipo , Receptor del Péptido 1 Similar al Glucagón/agonistas , Receptor del Péptido 1 Similar al Glucagón/metabolismo , Hipoglucemiantes/farmacología , Incretinas/farmacología , Insulina/sangre , Células Secretoras de Insulina/efectos de los fármacos , Células Secretoras de Insulina/ultraestructura , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias/efectos de los fármacos , Mitocondrias/ultraestructura , Péptidos/farmacología , Fenotipo , Factores de Tiempo , Ponzoñas/farmacología
14.
Carcinogenesis ; 36(11): 1407-18, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26354775

RESUMEN

Abnormal accumulation of defective mitochondria is the hallmark of oncocytes, which are frequently observed in thyroid Hürthle cell lesions. Autophagy is an essential cellular catabolic mechanism for the degradation of dysfunctional organelles and has been implicated in several human diseases. It is yet unknown how autophagic turnover of defective mitochondria in Hürthle cell tumors is regulated. We characterized the expression patterns of molecular markers including Beclin1, LC3, PINK1 and Parkin, which are required for autophagy or mitophagy, in human oncocytic lesions of the thyroid. To undertake mechanistic studies, we investigated autophagy and mitophagy using XTC.UC1 cells, the only in vitro model of Hürthle cell tumors. Beclin1 and LC3 were highly expressed in oncocytes of Hürthle cell tumors. XTC.UC1 showed autophagic responses to starvation and rapamycin treatment, whereas they displayed ineffective activation of mitophagy, which is triggered by the coordinated action of PINK1 and Parkin in response to CCCP. This resulted in a decreased turnover of abnormal mitochondria. The mechanisms underlying defective mitophagy and mitochondrial turnover were investigated by genetic analysis of the PARK2 gene in XTC.UC1 and Hürthle cell tumor tissues. XTC.UC1 and several tumors harbored the V380L mutation, resulting in dysfunctional autoubiquitination and decreased E3 ligase activity. Consistently, oncocytes in Hürthle cell tumors displayed comparable expression of PINK1 but decreased Parkin expression in comparison to normal thyrocytes. The introduction of wild-type Parkin sensitized XTC.UC1 to death induced by CCCP. This study provides a possible etiological basis for oncocytic formation in heterogeneous Hürthle cell tumors through insufficient mitophagy leading to ineffective turnover of aberrant mitochondria caused by dysfunctional Parkin-mediated pathways of mitochondria quality control.


Asunto(s)
Mitofagia , Neoplasias de la Tiroides/enzimología , Ubiquitina-Proteína Ligasas/genética , Adenoma Oxifílico , Adulto , Anciano , Proteínas Reguladoras de la Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Autofagia , Beclina-1 , Línea Celular Tumoral , Análisis Mutacional de ADN , Femenino , Expresión Génica , Regulación Neoplásica de la Expresión Génica , Estudios de Asociación Genética , Células HEK293 , Humanos , Masculino , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Persona de Mediana Edad , Mutación Missense , Consumo de Oxígeno , Estudios Retrospectivos , Glándula Tiroides/metabolismo , Glándula Tiroides/patología , Neoplasias de la Tiroides/genética , Ubiquitina-Proteína Ligasas/metabolismo , Adulto Joven
15.
Biochem Biophys Res Commun ; 457(1): 95-100, 2015 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-25545062

RESUMEN

Omega-3 polyunsaturated fatty acid levels are reduced in the substantia nigra area in Parkinson's disease patients and animal models, implicating docosahexaenoic acid (DHA) as a potential treatment for preventing Parkinson's disease and suggesting the need for investigations into how DHA might protect against neurotoxin-induced dopaminergic neuron loss. The herbicide paraquat (PQ) induces dopaminergic neuron loss through the excessive production of reactive oxygen species (ROS). We found that treatment of dopaminergic SN4741 cells with PQ reduced cell viability in a dose-dependent manner, but pretreatment with DHA ameliorated the toxic effect of PQ. To determine the toxic mechanism of PQ, we measured intracellular ROS content in different organelles with specific dyes. As expected, all types of ROS were increased by PQ treatment, but DHA pretreatment selectively decreased cytosolic hydrogen peroxide content. Furthermore, DHA treatment-induced increases in glutathione reductase and glutamate cysteine ligase modifier subunit (GCLm) mRNA expression were positively correlated with glutathione (GSH) content. Consistent with this increase in GCLm mRNA levels, Western blot analysis revealed that DHA pretreatment increased nuclear factor-erythroid 2 related factor 2 (Nrf2) protein levels. These findings indicate that DHA prevents PQ-induced neuronal cell loss by enhancing Nrf2-regulated GSH homeostasis.


Asunto(s)
Ácidos Docosahexaenoicos/farmacología , Neuronas Dopaminérgicas/metabolismo , Glutatión/metabolismo , Homeostasis/efectos de los fármacos , Paraquat/toxicidad , Especies Reactivas de Oxígeno/metabolismo , Animales , Muerte Celular/efectos de los fármacos , Línea Celular , Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/enzimología , Glutatión Reductasa/metabolismo , Ratones , Factor 2 Relacionado con NF-E2/metabolismo , Fármacos Neuroprotectores/farmacología
16.
Liver Int ; 35(4): 1341-53, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25443620

RESUMEN

BACKGROUND & AIMS: Nonalcoholic steatohepatitis (NASH) is associated with cirrhosis and hepatocellular carcinoma. Reactive oxygen species (ROS) and reactive nitrogen species (RNS) play key roles in the development of the disease. However, the therapeutic target of NASH has not been fully defined and new treatments are needed. We investigated the protective effects of the antioxidant indole-derived NecroX-7 in a NASH mouse model using leptin-deficient ob/ob and methionine- and choline-deficient (MCD) diet-fed ob/ob mice. METHODS: Six-week-old male mice were divided into three groups: ob/+ mice, ob/ob mice treated with vehicle and ob/ob mice treated daily with NecroX-7 (20 mg/kg) for 4 weeks. To study the effects of NecroX-7 in a fibrosis model, NASH was induced by feeding ob/ob mice an MCD diet. The effects of NecroX-7 on NASH progression were evaluated using biochemical, histological and molecular markers. RESULTS: NecroX-7-treated ob/ob mice had a marked decrease in serum aspartate aminotransferase and alanine transaminase compared with vehicle-treated controls. Interestingly, hepatic steatosis and lipid peroxidation were significantly improved by NecroX-7 treatment. NecroX-7 inhibited tert-butylhydroperoxide- and H2 O2 -induced mitochondrial ROS/RNS in primary hepatocytes and attenuated mitochondrial dysfunction in vitro and in vivo. Furthermore, NecroX-7-treated mice exhibited fewer infiltrating macrophages and reduced hepatic tumour necrosis factor-alpha expression. Hepatic fibrosis in MCD-fed ob/ob mice was significantly decreased by NecroX-7 treatment. CONCLUSIONS: NecroX-7 treatment improved hepatic steatosis and fibrosis in murine NASH models. These effects occurred through the suppression of whole-cell ROS/RNS and inflammatory responses and suggest that NecroX-7 has a potential therapeutic benefit in steatohepatitis.


Asunto(s)
Antioxidantes/farmacología , Inflamación/tratamiento farmacológico , Hígado/efectos de los fármacos , Mitocondrias Hepáticas/efectos de los fármacos , Enfermedad del Hígado Graso no Alcohólico/tratamiento farmacológico , Compuestos Orgánicos/farmacología , Estrés Oxidativo/efectos de los fármacos , Especies de Nitrógeno Reactivo/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Animales , Citoprotección , Metabolismo Energético/efectos de los fármacos , Células Hep G2 , Humanos , Inflamación/metabolismo , Inflamación/patología , Mediadores de Inflamación/metabolismo , Peroxidación de Lípido/efectos de los fármacos , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática Experimental/tratamiento farmacológico , Cirrosis Hepática Experimental/metabolismo , Cirrosis Hepática Experimental/patología , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones Obesos , Mitocondrias Hepáticas/metabolismo , Mitocondrias Hepáticas/patología , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Transducción de Señal/efectos de los fármacos , Factores de Tiempo
17.
BMC Cancer ; 14: 36, 2014 Jan 22.
Artículo en Inglés | MEDLINE | ID: mdl-24447339

RESUMEN

BACKGROUND: Combination therapy is key to improving cancer treatment efficacy. Phorbol 12-myristate 13-acetate (PMA), a well-known PKC activator, increases the cytotoxicity of several anticancer drugs. Apicularen A induces cytotoxicity in tumor cells through disrupting microtubule networks by tubulin down-regulation. In this study, we examined whether PMA increases apicularen A-induced cytotoxicity in HeLa cells. METHODS: Cell viability was examined by thiazolyl blue tetrazolium (MTT) assays. To investigate apoptotic potential of apicularen A, DNA fragmentation assays were performed followed by extracting genomic DNA, and caspase-3 activity assays were performed by fluorescence assays using fluorogenic substrate. The cell cycle distribution induced by combination with PMA and apicularen A was examined by flow cytometry after staining with propidium iodide (PI). The expression levels of target proteins were measured by Western blotting analysis using specific antibodies, and α-tubulin mRNA levels were assessed by reverse transcription polymerase chain reaction (RT-PCR). To examine the effect of combination of PMA and apicularen A on the microtubule architecture, α-tubulin protein and nuclei were visualized by immunofluorescence staining using an anti-α-tubulin antibody and PI, respectively. RESULTS: We found that apicularen A induced caspase-dependent apoptosis in HeLa cells. PMA synergistically increased cytotoxicity and apoptotic sub-G1 population induced by apicularen A. These effects were completely blocked by the PKC inhibitors Ro31-8220 and Go6983, while caspase inhibition by Z-VAD-fmk did not prevent cytotoxicity. RNA interference using siRNA against PKCα, but not PKCß and PKCγ, inhibited cytotoxicity induced by combination PMA and apicularen A. PMA increased the apicularen A-induced disruption of microtubule networks by further decreasing α- and ß-tubulin protein levels in a PKC-dependent manner. CONCLUSIONS: These results suggest that the synergy between PMA and apicularen A is involved by PKCα activation and microtubule disruption, and that may inform the development of novel approaches to treat cancer.


Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Microtúbulos/efectos de los fármacos , Neoplasias del Cuello Uterino/metabolismo , Apoptosis/efectos de los fármacos , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Caspasa 3/metabolismo , Puntos de Control del Ciclo Celular , Supervivencia Celular/efectos de los fármacos , Sinergismo Farmacológico , Femenino , Células HeLa , Humanos , Microtúbulos/genética , Microtúbulos/metabolismo , Microtúbulos/patología , Proteína Quinasa C-alfa/antagonistas & inhibidores , Proteína Quinasa C-alfa/genética , Proteína Quinasa C-alfa/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Interferencia de ARN , Acetato de Tetradecanoilforbol/farmacología , Factores de Tiempo , Transfección , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/farmacología , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patología
18.
BMC Cancer ; 14: 481, 2014 Jul 03.
Artículo en Inglés | MEDLINE | ID: mdl-24993937

RESUMEN

BACKGROUND: The role of omega-3 polyunsaturated fatty acids (ω3-PUFAs) in cancer prevention has been demonstrated; however, the exact molecular mechanisms underlying the anticancer activity of ω3-PUFAs are not fully understood. Here, we investigated the relationship between the anticancer action of a specific ω3-PUFA docosahexaenoic acid (DHA), and the conventional mitogen-activated protein kinases (MAPKs) including extracellular signal-regulated kinase (ERK), c-JUN N-terminal kinase (JNK) and p38 whose dysregulation has been implicated in human cancers. METHODS: MTT assays were carried out to determine cell viability of cancer cell lines (PA-1, H1299, D54MG and SiHa) from different origins. Apoptosis was confirmed by TUNEL staining, DNA fragmentation analysis and caspase activity assays. Activities of the conventional MAPKs were monitored by their phosphorylation levels using immunoblotting and immunocytochemistry analysis. Reactive oxygen species (ROS) production was measured by flow cytometry and microscopy using fluorescent probes for general ROS and mitochondrial superoxide. RESULTS: DHA treatment decreased cell viability and induced apoptotic cell death in all four studied cell lines. DHA-induced apoptosis was coupled to the activation of the conventional MAPKs, and knockdown of ERK/JNK/p38 by small interfering RNAs reduced the apoptosis induced by DHA, indicating that the pro-apoptotic effect of DHA is mediated by MAPKs activation. Further study revealed that the DHA-induced MAPKs activation and apoptosis was associated with mitochondrial ROS overproduction and malfunction, and that ROS inhibition remarkably reversed these effects of DHA. CONCLUSION: Together, these results indicate that DHA-induced MAPKs activation is dependent on its capacity to provoke mitochondrial ROS generation, and accounts for its cytotoxic effect in human cancer cells.


Asunto(s)
Apoptosis/efectos de los fármacos , Ácidos Docosahexaenoicos/farmacología , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Neoplasias/enzimología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Humanos , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Neoplasias/metabolismo , Especies Reactivas de Oxígeno/metabolismo
19.
J Biol Chem ; 287(50): 41875-87, 2012 Dec 07.
Artículo en Inglés | MEDLINE | ID: mdl-23074219

RESUMEN

Cyclic AMP (cAMP) induces steroidogenic enzyme gene expression and stimulates testosterone production in Leydig cells. Phosphoenolpyruvate carboxykinase (PEPCK) is expressed in Leydig cells, but its role has not been defined. In this study, we found that PEPCK and glucose-6-phosphatase (Glc-6-Pase) are increased significantly following cAMP treatment of mouse Leydig cells. Moreover, cAMP treatment increased recruitment of the cAMP-response element-binding transcription factor and decreased recruitment of the corepressor DAX-1 on the pepck promoter. Furthermore, cAMP induced an increase in ATP that correlated with a decrease in phospho-AMP-activated protein kinase (AMPK). In contrast, knockdown or inhibition of PEPCK decreased ATP and increased phospho-AMPK. Treatment with an AMPK activator or overexpression of the constitutively active form of AMPK inhibited cAMP-induced steroidogenic enzyme promoter activities and gene expression. Liver receptor homolog-1 (LRH-1) was involved in cAMP-induced steroidogenic enzyme gene expression but was inhibited by AMPK activation in Leydig cells. Additionally, inhibition or knockdown of PEPCK and Glc-6-Pase decreased cAMP-mediated induction of steroidogenic enzyme gene expression and steroidogenesis. Finally, pubertal mouse (8-week-old) testes and human chorionic gonadotropin-induced prepubertal mouse testes showed increased PEPCK and Glc-6-Pase gene expression. Taken together, these results suggest that induction of PEPCK and Glc-6-Pase by cAMP plays an important role in Leydig cell steroidogenesis.


Asunto(s)
Glucosa-6-Fosfatasa/biosíntesis , Células Intersticiales del Testículo/enzimología , Fosfoenolpiruvato Carboxiquinasa (ATP)/biosíntesis , Proteínas Quinasas Activadas por AMP/genética , Proteínas Quinasas Activadas por AMP/metabolismo , Animales , AMP Cíclico/genética , AMP Cíclico/metabolismo , Receptor Nuclear Huérfano DAX-1/genética , Receptor Nuclear Huérfano DAX-1/metabolismo , Regulación Enzimológica de la Expresión Génica/fisiología , Glucosa-6-Fosfatasa/genética , Células HeLa , Humanos , Células Intersticiales del Testículo/citología , Masculino , Ratones , Fosfoenolpiruvato Carboxiquinasa (ATP)/genética , Regiones Promotoras Genéticas/fisiología , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo
20.
Biochem Biophys Res Commun ; 434(3): 634-40, 2013 May 10.
Artículo en Inglés | MEDLINE | ID: mdl-23583412

RESUMEN

Apicularen A is a novel antitumor agent and strongly induces death in tumor cells. In this study, we synthesized apicularen A acetate, an acetyl derivative of apicularen A, and investigated its antitumor effect and mechanism in HM7 colon cancer cells. Apicularen A acetate induced apoptotic cell death and caspase-3 activation; however, the pan-caspase inhibitor Z-VAD-fmk could not prevent this cell death. Apicularen A acetate induced the loss of mitochondrial membrane potential and the translocation of apoptosis-inducing factor (AIF) from mitochondria. In addition, apicularen A acetate significantly decreased tubulin mRNA and protein levels and induced disruption of microtubule networks. Taken together, these results indicate that the mechanism of apicularen A acetate involves caspase-independent apoptotic cell death and disruption of microtubule architecture.


Asunto(s)
Factor Inductor de la Apoptosis/metabolismo , Apoptosis/efectos de los fármacos , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Neoplasias del Colon/patología , Regulación hacia Abajo/efectos de los fármacos , Microtúbulos/efectos de los fármacos , Tubulina (Proteína)/metabolismo , Western Blotting , Línea Celular Tumoral , Neoplasias del Colon/metabolismo , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Microtúbulos/metabolismo , Transporte de Proteínas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
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