A potential role of autophagy-mediated vascular senescence in the pathophysiology of HFpEF.

Heart failure with preserved ejection fraction (HFpEF) is one of the most complex and most prevalent cardiometabolic diseases in aging population. Age, obesity, diabetes, and hypertension are the main comorbidities of HFpEF. Microvascular dysfunction and vascular remodeling play a major role in its development. Among the many mechanisms involved in this process, vascular stiffening has been described as one the most prevalent during HFpEF, leading to ventricular-vascular uncoupling and mismatches in aged HFpEF patients. Aged blood vessels display an increased number of senescent endothelial cells (ECs) and vascular smooth muscle cells (VSMCs). This is consistent with the fact that EC and cardiomyocyte cell senescence has been reported during HFpEF. Autophagy plays a major role in VSMCs physiology, regulating phenotypic switch between contractile and synthetic phenotypes. It has also been described that autophagy can regulate arterial stiffening and EC and VSMC senescence. Many studies now support the notion that targeting autophagy would help with the treatment of many cardiovascular and metabolic diseases. In this review, we discuss the mechanisms involved in autophagy-mediated vascular senescence and whether this could be a driver in the development and progression of HFpEF.

Methodology Aspects of Colony Maintain for a Murine Model of Amyotrophic Lateral Sclerosis (ALS) TDP-43 Proteinopathy.

The use of genetically engineered mouse (GEMs) models provides an unprecedented opportunity to study the genetic basis of diseases and gene function, therefore it is paramount to determine reproductive parameters that guarantee proper colony maintenance. We studied the reproductive parameters of mice hemizygous for TDP-43A315T transgene, which are viable, fertile, and express a mutant human TAR DNA binding protein (hTDP-43) cDNA harboring an amino acid substitution associated with familial amyotrophic lateral sclerosis (fALS). TDP43A315T mice were backcrossed to a C57Bl6/J pure background for four consecutive generations. The Tg offspring genotype were then confirmed by PCR assays. Our statistical analysis indicated there were no differences in the sex and number of pups per offspring when hemizygous female and male TDP43A315T mice were backcrossed to C57Bl6/J mice. Interestingly, our results showed significant differences in the number of offspring expressing the transgene when hemizygous TDP43A315T male mice were used as breeders. Therefore, our findings suggest that male TDP43A315T mice transfer the transgene with a greater genetic strengths. Such is an important breeding consideration to ensure the principle of reduction in animal experimentation considering most basic research with models focuses on males and excludes female mice.

Pro-fibrotic effect of oxidized LDL in cardiac myofibroblasts.

Inflammatory signals associated with cardiac diseases trigger trans-differentiation of cardiac fibroblasts to cardiac myofibroblasts. Cardiac myofibroblasts are the main cell type involved in the development of cardiac fibrosis, a diffuse and disproportionate accumulation of collagen in the myocardium. Although the role of the scavenger like-lectin receptor LOX-1 was previously investigated in cardiac fibroblasts and fibrosis, the involvement of the LOX-1 ligand -oxidized low-density lipoprotein (oxLDL)- on cardiac myofibroblast function still remains unexplored. In the present work, we investigated the effect of oxLDL/LOX-1 on fibrotic markers and cardiac myofibroblast function. Our in vitro results showed that oxLDL increased cardiac myofibroblast proliferation, triggered an increase in the synthesis of collagen type I and fibronectin containing extra domain A, and stimulated collagen type I secretion. oxLDL also decreased cardiac myofibroblast migration, collagen gel contraction and cell area, without modifying α-smooth muscle actin protein levels. These effects were dependent on LOX-1, because LOX-1 knockdown abolished oxLDL effects. Collectively these data showed that oxLDL has important modulatory effects on cardiac myofibroblast function.

Enfermedades cardiovasculares y cáncer: ¿dos entidades mutuamente relacionadas? / Cardiovascular diseases and cancer: ¿mutually related entities?

Resumen: Las enfermedades cardiovasculares y el cáncer son enfermedades crónicas transmisibles culturalmente, y las dos causas principales de mortalidad en el mundo. Además del gran impacto sobre la mortalidad y morbilidad, estas enfermedades han mostrado un alto grado de relación entre ellas debido, entre otras razones, a que comparten factores de riesgo y mecanismos biológicos. La alta incidencia de enfermedad cardiovascular en pacientes con cáncer es un fenómeno conocido que ha orientado el desarrollo del campo interdisciplinario de la cardio-oncología. Sin embargo, en la última década han surgido evidencias que muestran el papel que desempeñan las enfermedades cardiovasculares en el desarrollo de cáncer. Un estudio reciente publicado por Meijers y cols, en agosto de 2018 en Circulation, mostró que la insuficiencia cardiaca post-infarto del miocardio contribuye significativamente al desarrollo del cáncer de colón, apoyando lo obtenido en estudios epidemiológicos anteriores. Este estudio también sugiere que el crecimiento tumoral podría producirse por factores secretados por el corazón insuficiente abriendo un amplio grupo de posibilidades de investigación en lo que sería un nuevo campo de la medicina cuyo propósito sería el desarrollo de nuevas estrategias para el seguimiento y tratamiento del cáncer en pacientes con enfermedades cardiovasculares. El presente artículo revisa los factores de riesgo, y mecanismos celulares y moleculares, que son comunes en las enfermedades cardiovasculares y el cáncer, la contribución del trabajo de Meijers y cols hacia un mayor entendimiento de la interrelación entre estas patologías y las perspectivas futuras con respecto a los nuevos hallazgos.

Abstracts: Cardiovascular diseases and cancer are culturally transmitted chronic diseases and the two main causes of death globally. In addition to their high morbidity and mortality, these diseases are closely related, due to their common risk factors and biological mechanisms. The high incidence of cardiovascular diseases in cancer patients is widely known phenomenon, which has oriented the development of the interdisciplinary field of cardio-oncology Nonetheless, there is emerging evidence in the last decade suggesting a potential role for cardiovascular diseases in the onset of cancer. A recent publication by Meijers et al in the scientific cardiovascular journal Circulation showed that heart failure significantly contributes to tumor growth, confirming previous epidemiological findings suggesting this hypothesis. Moreover, this study indicates that tumor growth may be stimulated by the secretion of factors from the failing heart, opening a wide spectrum of research areas in what may be suggested as a new field in medicine that would seek to develop new strategies to treat and prevent cancer in patients with cardiovascular diseases. This article will review shared risk factor and common cellular and molecular pathways in cardiovascular diseases and cancer, the contribution of Meijers et al to a better understanding of the connection of these diseases and future perspectives in light of the new evidence.

A Culturally Adapted Intervention for Mexican-Origin Parents of Adolescents: The Need to Overtly Address Culture and Discrimination in Evidence-Based Practice.

Latino/a populations in the United States are negatively impacted by widespread mental health disparities. Although the dissemination of culturally relevant parent training (PT) programs constitutes an alternative to address this problem, there is a limited number of efficacious culturally adapted PT prevention interventions for low-income Latino/a immigrant families with adolescents. The current manuscript describes the level of acceptability of a version of the GenerationPMTO® intervention adapted for Latino/a immigrant families, with an explicit focus on immigration-related challenges, discrimination, and promotion of biculturalism. Qualitative reports were provided by 39 immigrant parents who successfully completed the prevention parenting program. The majority of these parents self-identified as Mexican-origin. According to qualitative findings, participants reported overall high satisfaction with immigration and culture-specific components. Parents also expressed high satisfaction with the core GenerationPMTO parenting components and provided specific recommendations for improving the intervention. Current findings indicate the need to adhere to the core components that account for the effectiveness of PT interventions. Equally important is to thoroughly adapt PT interventions according to the cultural values and experiences that are relevant to target populations, as well as to overtly address experiences of discrimination that negatively impact underserved Mexican-origin immigrant families. Due to the exploratory nature of this study, the efficacy and effectiveness of the adapted prevention intervention remains to be established in empirical research.

Las poblaciones latinas de los Estados Unidos están afectadas negativamente por desigualdades generalizadas en el área de salud mental. Aunque la difusión de programas de capacitación para padres culturalmente pertinentes constituye una alternativa para abordar este problema, existe un número limitado de capacitaciones para padres con intervenciones preventivas adaptadas culturalmente que son eficaces para familias inmigrantes latinas con adolescentes de bajos recursos. El presente manuscrito describe el nivel de aceptabilidad de una versión de la intervención GenerationPMTO® adaptada para familias inmigrantes latinas, que hace hincapié específicamente en dificultades relacionadas con la inmigración, la discriminación y la promoción del biculturalismo. Treinta y nueve padres inmigrantes que completaron satisfactoriamente el programa preventivo sobre crianza proporcionaron informes cualitativos. La mayoría de estos padres se autoidentificaron como de origen mexicano. De acuerdo con los resultados cualitativos, los participantes informaron una alta satisfacción en general con la inmigración y los componentes específicos de la cultura. Los padres también expresaron una alta satisfacción con los componentes principales relativos a la crianza de GenerationPMTO y ofrecieron recomendaciones específicas para mejorar la intervención. Los resultados actuales indican la necesidad de cumplir con los componentes principales que justifican la eficacia real de las intervenciones de capacitación para padres. Es igualmente importante adaptar totalmente las intervenciones de capacitación para padres de acuerdo con los valores culturales y las experiencias que son relevantes para las poblaciones objetivo, así como abordar expresamente las experiencias de discriminación que afectan negativamente a las familias inmigrantes de bajos recursos de origen mexicano. Debido a la índole exploratoria de este estudio, la eficacia real y la eficacia teórica de la intervención de prevención adaptada quedan por demostrarse en la investigación empírica.

Autophagy protects against neural cell death induced by piperidine alkaloids present in Prosopis juliflora (Mesquite).

Prosopis juliflora is a shrub that has been used to feed animals and humans. However, a synergistic action of piperidine alkaloids has been suggested to be responsible for neurotoxic damage observed in animals. We investigated the involvement of programmed cell death (PCD) and autophagy on the mechanism of cell death induced by a total extract (TAE) of alkaloids and fraction (F32) from P. juliflora leaves composed majoritary of juliprosopine in a model of neuron/glial cell co-culture. We saw that TAE (30 µg/mL) and F32 (7.5 µg/mL) induced reduction in ATP levels and changes in mitochondrial membrane potential at 12 h exposure. Moreover, TAE and F32 induced caspase-9 activation, nuclear condensation and neuronal death at 16 h exposure. After 4 h, they induced autophagy characterized by decreases of P62 protein level, increase of LC3II expression and increase in number of GFP-LC3 cells. Interestingly, we demonstrated that inhibition of autophagy by bafilomycin and vinblastine increased the cell death induced by TAE and autophagy induced by serum deprivation and rapamycin reduced cell death induced by F32 at 24 h. These results indicate that the mechanism neural cell death induced by these alkaloids involves PCD via caspase-9 activation and autophagy, which seems to be an important protective mechanism.

Autophagy protects against neural cell death induced by piperidine alkaloids present in Prosopis juliflora (Mesquite)

ABSTRACT Prosopis juliflora is a shrub that has been used to feed animals and humans. However, a synergistic action of piperidine alkaloids has been suggested to be responsible for neurotoxic damage observed in animals. We investigated the involvement of programmed cell death (PCD) and autophagy on the mechanism of cell death induced by a total extract (TAE) of alkaloids and fraction (F32) from P. juliflora leaves composed majoritary of juliprosopine in a model of neuron/glial cell co-culture. We saw that TAE (30 µg/mL) and F32 (7.5 µg/mL) induced reduction in ATP levels and changes in mitochondrial membrane potential at 12 h exposure. Moreover, TAE and F32 induced caspase-9 activation, nuclear condensation and neuronal death at 16 h exposure. After 4 h, they induced autophagy characterized by decreases of P62 protein level, increase of LC3II expression and increase in number of GFP-LC3 cells. Interestingly, we demonstrated that inhibition of autophagy by bafilomycin and vinblastine increased the cell death induced by TAE and autophagy induced by serum deprivation and rapamycin reduced cell death induced by F32 at 24 h. These results indicate that the mechanism neural cell death induced by these alkaloids involves PCD via caspase-9 activation and autophagy, which seems to be an important protective mechanism.

Autofagia en el sistema cardiovascular: pasado, presente y futuro / Cardiovascular autophagy: past, present and future

La enfermedad cardiovascular se mantiene como la principal causa de morbimortalidad a nivel mundial a pesar de los avances científicos y tecnológicos recientes, por esto existe la necesidad de búsqueda de nuevas dianas terapéuticas. La autofagia es un mecanismo de degradación de proteínas y organelos disfuncionales que ocurre en vacuolas especializadas de doble membrana denominadas autofagosomas y que requiere la participación de los lisosomas. Este proceso permite el auto abastecimiento celular de energía a través del reciclaje de diversos substratos energéticos. Se activa en respuesta a diversas formas de estrés, principalmente debido a la ausencia de nutrientes y su presencia ha sido caracterizada en todos los tipos celulares que componen el sistema cardiovascular. Existe una ventana de actividad de autofagia óptima la que se relaciona con la mantención de la homeostasis cardiovascular y su desregulación participa en la patogénesis de diversas patologías cardiovasculares. En este artículo se revisa el curso temporal que llevó el descubrimiento de la autofagia, la contribución al área del Dr. Ohsumi, reciente Premio Nobel de Medicina, los principales conceptos, mecanismos celulares y moleculares de la formación del auto-fagosoma, nodos de regulación y sintetizamos su participación en la homeostasis del corazón y en la patogénesis de las enfermedades cardiovasculares y sus perspectivas futuras.

Cardiovascular disease continues to be the leading cause of morbi-mortality worldwide despite the recent scientific and technological advances. Therefore, more research is needed to discover novel therapeutic targets. Autophagy mediates the removal of dysfunctional proteins and organelles. This process takes place in double-membrane vesicles, named autophagosomes, which later fuse with lysosomes. The mechanism allows self-renewal energy repletion through diverse energy substrate recycling. Diverse forms of cellular stress, mainly nutrient deprivation, activate this process. Autophagy has been widely characterized within the cells of the cardiovascular system. There is a window of optimal autophagy activity implicated in maintaining cardiovascular homeostasis and its dysregulation participates in the pathogenesis of different cardiovascular diseases. In this article, we review the time course of auto-phagy discovery, the Nobel Prize winner Dr. Ohsumi contribution, main concepts, mechanisms involved in autophagosome formation and its regulatory no-des. Additionally, we summarized the role of auto-phagy in cardiovascular homeostasis and pathogenesis and future perspectives.

Glutathione transferase-M2-2 secreted from glioblastoma cell protects SH-SY5Y cells from aminochrome neurotoxicity.

U373MG cells are able to take up aminochrome that induces glutathione transferase M2-2 (GSTM2) expression in a concentration-dependent manner where 100 µM aminochrome increases GSTM2 expression by 2.1-fold (P < 0.001) at 3 h. The uptake of (3)H-aminochrome into U373MG cells was significantly reduced in the presence of 2 µM nomifensine (P < 0.001) 100 µM imipramine (P < 0.001) and 50 mM dopamine (P < 0.001). Interestingly, U373MG cells excrete GSTM2 into the conditioned medium and the excretion was significantly increased (2.7-fold; P < 0.001) when the cells were pretreated with 50 µM aminochrome for 3 h. The U373MG-conditioned medium containing GSTM2 protects SH-SY5Y cells incubated with 10 µM aminochrome. The significant protection provided by U373MG-conditioned medium in SH-SY5Y cells incubated with aminochrome was dependent on GSTM2 internalization into SH-SY5Y cells as evidenced by (i) uptake of (14)C-GSTM2 released from U373MG cells into SH-SY5Y cells, a process inhibited by anti-GSTM2 antiserum; (ii) lack of protection of U373MG-conditioned medium in the presence of anti-GSTM2 antiserum on SH-SY5Y cells treated with aminochrome; and (iii) lack of protection of conditioned medium from U373MGsiGST6 that expresses an siRNA directed against GSTM2 on SH-SY5Y cells treated with aminochrome. In conclusion, our results demonstrated that U373MG cells protect SH-SY5Y cells against aminochrome neurotoxicity by releasing GSTM2 into the conditioned medium and subsequent internalization of GSTM2 into SH-SY5Y cells. These results suggest a new mechanism of protection of dopaminergic neurons mediated by astrocytes by releasing GSTM2 into the intersynaptic space and subsequent internalization into dopaminergic neuron in order to protect these cells against aminochrome neurotoxicity.

Glutathione transferase mu 2 protects glioblastoma cells against aminochrome toxicity by preventing autophagy and lysosome dysfunction.

U373MG cells constitutively express glutathione S-transferase mu 2 (GSTM2) and exhibit (3)H-dopamine uptake, which is inhibited by 2 µM of nomifensine and 15 µM of estradiol. We generated a stable cell line (U373MGsiGST6) expressing an siRNA against GSTM2 that resulted in low GSTM2 expression (26% of wild-type U373MG cells). A significant increase in cell death was observed when U373MGsiGST6 cells were incubated with 50 µM purified aminochrome (18-fold increase) compared with wild-type cells. The incubation of U373MGsiGST6 cells with 75 µM aminochrome resulted in the formation of autophagic vacuoles containing undigested cellular components, as determined using transmission electron microscopy. A significant increase in autophagosomes was determined by measuring endogenous LC3-II, a significant decrease in cell death was observed in the presence of bafilomycin A 1, and a significant increase in cell death was observed in the presence of trehalose. A significant increase in LAMP2 immunostaining was observed, a significant decrease in bright red fluorescence of lysosomes with acridine orange was observed, and bafilomycin A 1 pretreatment reduced the loss of lysosome acidity. A significant increase in cell death was observed in the presence of lysosomal protease inhibitors. Aggregation of TUBA/α-tubulin (tubulin, α) and SQSTM1 protein accumulation were also observed. Moreover, a significant increase in the number of lipids droplets was observed compared with U373MG cells with normal expression of GSTM2. These results support the notion that GSTM2 is a protective enzyme against aminochrome toxicity in astrocytes and that aminochrome cell death in U373MGsiGST6 cells involves autophagic-lysosomal dysfunction.

One-electron reduction of 6-hydroxydopamine quinone is essential in 6-hydroxydopamine neurotoxicity.

6-Hydroxydamine has widely been used as neurotoxin in preclinical studies related on the neurodegenerative process of dopaminergic neurons in Parkinson's disease based on its ability to be neurotoxic as a consequence of free radical formation during its auto-oxidation to topaminequinone. We report that 50-µM 6-hydroxydopamine is not neurotoxic in RCSN-3 cells derived from substantia nigra incubated during 24 h contrasting with a significant sixfold increase in cell death (16 ± 2 %; P < 0.001) was observed in RCSN-3NQ7 cells expressing a siRNA against DT-diaphorase that silence the enzyme expression. To observe a significant cell death in RCSN-3 cells induced by 6-hydroxydopamine (24 ± 1 %; P < 0.01), we have to increase the concentration to 250 µm while a 45 ± 2 % cell death (P < 0.001) was observed at this concentration in RCSN-3NQ7 cells. The cell death induced by 6-hydroxydopamine in RCSN-3NQ7 cells was accompanied with a (i) significant increase in oxygen consumption (P < 0.01), (ii) depletion of reduced glutathione and (iii) a significant decrease in ATP level (P < 0.05) in comparison with RCSN-3 cells. In conclusion, our results suggest that one-electron reduction of 6-hydroxydopamine quinone seems to be the main reaction responsible for 6-hydroxydopamine neurotoxic effects in dopaminergic neurons and DT-diaphorase seems to play an important neuroprotective role by preventing one-electron reduction of topaminequinone.

Protective effects of nicotine against aminochrome-induced toxicity in substantia nigra derived cells: implications for Parkinson's disease.

Parkinson's disease is a debilitating progressive neurodegenerative disorder that results from the loss of or damage to dopaminergic cells containing neuromelanin in the substantia nigra (SN). The underlying neurodegenerative mechanism(s), however, remain elusive. Aminochrome, the precursor of neuromelanin is an endogenous substance capable of inducing selective neurotoxicity to dopaminergic neurons in SN. Nicotine, on the other hand, may offer protective effects against dopaminergic cell damage induced by various neurotoxins including MPTP and salsolinol. In this study, we sought to determine whether nicotine may also protect against aminochrome-induced toxicity in SN derived RCSN-3 cells. Exposure of RCSN-3 cells to a combination of aminochrome (50 µM) and dicoumarol (50 µM) for 48 h induced approximately 70 % cell death. Pretreatment with nicotine, dose-dependently blocked this toxicity. The effects of nicotine in turn were dose-dependently blocked by mecamylamine, a non-selective nicotinic receptor antagonist. These results suggest involvement of nicotinic receptors in protective effects of nicotine against aminochrome-induced toxicity and provide further evidence for possible therapeutic effects of nicotine or nicotinic agonists in Parkinson's disease.

The dopamine metabolite aminochrome inhibits mitochondrial complex I and modifies the expression of iron transporters DMT1 and FPN1.

Hallmarks of idiopathic and some forms of familial Parkinson's disease are mitochondrial dysfunction, iron accumulation and oxidative stress in dopaminergic neurons of the substantia nigra. There seems to be a causal link between these three conditions, since mitochondrial dysfunction can give rise to increased electron leak and reactive oxygen species production. In turn, recent evidence indicates that diminished activity of mitochondrial complex I results in decreased Fe­S cluster synthesis and anomalous activation of Iron Regulatory Protein 1. Thus, mitochondrial dysfunction could be a founding event in the process that leads to neuronal death. Here, we present evidence showing that at low micromolar concentrations, the dopamine metabolite aminochrome inhibits complex I and ATP production in SH-SY5Y neuroblastoma cells differentiated into a dopaminergic phenotype. This effect is apparently direct, since it is replicated in isolated mitochondria. Additionally, overnight treatment with aminochrome increased the expression of the iron import transporter divalent metal transporter 1 and decreased the expression of the iron export transporter ferroportin 1. In accordance with these findings, cells treated with aminochrome presented increased iron uptake. These results suggest that aminochrome is an endogenous toxin that inhibits by oxidative modifications mitochondrial complex I and modifies the levels of iron transporters in a way that leads to iron accumulation.