NEK10 kinase ablation affects mitochondrial morphology, function and protein phosphorylation status.

BACKGROUND: NEK10, a serine/threonine/tyrosine kinase belonging to the NEK (NIMA-related kinases) family, has been associated with diverse cellular processes. However, no specific target pathways have been identified. Our previous work knocking down NEK10 in HeLa cells suggested a functional association with mitochondria, as we observed altered mitochondrial morphology, mitochondrial oxygen consumption, mtDNA integrity, and reactive oxygen species levels. METHODS: To better understand this association, we studied human HAP1 cells fully knockout for NEK10 and confirmed that NEK10 has an important role in mitochondrial homeostasis. We performed the study of mitochondrial respiration, mitochondrial morphology, mitochondrial mass, and mtDNA analysis. Additionally, we showed proteome and phosphoproteome data of crude mitochondrial fraction of Parental and NEK10 KO cells using liquid chromatography-mass spectrometry (LC-MS/MS). RESULTS: In the absence of NEK10 several mitochondrial functions were disturbed. Moreover, proteome and phosphoproteome analyses of mitochondrial fractions showed that NEK10 alters the threonine phosphorylation status of several mitochondrial/endoplasmic reticulum components, including HSP60, NDUFB4, and TOM20. These changes impacted the steady-state levels of a larger group of proteins, preferentially involving respiratory complexes and autophagy pathways. CONCLUSION: We concluded that NEK10 plays a key role in mitochondrial function, possibly by modulating the phosphorylation status of mitochondrial proteins.

Influence of Mitochondrial NAD(P) + Transhydrogenase (NNT) on Hypothalamic Inflammation and Metabolic Dysfunction Induced by a High-Fat Diet in Mice.

The mitochondrial protein NAD(P)+ transhydrogenase (NNT) has been implicated in the metabolic derangements observed in obesity. Mice with the C57BL/6J genetic background bear a spontaneous mutation in the Nnt gene and are known to exhibit increased susceptibility to diet-induced metabolic disorders. Most of the studies on NNT in the context of diet-induced obesity have compared C57BL/6J mice with other mouse strains, where differences in genetic background can serve as confounding factors. Moreover, these studies have predominantly employed a high-fat diet (HFD) consisting of approximately 60% of calories from fat, which may not accurately mimic real-world fat-rich diets. In this study, we sought to examine the role of NNT in diet-induced hypothalamic inflammation and metabolic syndrome by using a congenic mice model lacking NNT, along with a HFD providing approximately 45% of calories from fat. Our findings indicate that mice lacking NNT were more protected from HFD-induced weight gain but presented a worse performance on glucose tolerance test, albeit not in insulin tolerance test. Interestingly, the brown adipose tissue of HFD-fed Nnt +/+ mice presented a greater mass and a higher whole-tissue ex-vivo oxygen consumption rate. Also, HFD increased the expression of the inflammatory markers Il1ß, Tlr4 and Iba1 in the hypothalamus of Nnt -/- mice. In conclusion, our study highlights the importance of NNT in the context of diet-induced obesity and metabolic syndrome, indicating its contribution to mitigate hypothalamic inflammation and suggesting its role in the brown adipose tissue increased mass.

Gait, Quality of Life, and Knee Function in Advanced Knee Osteoarthritis: A Single-Center, Prospective, Observational Study.

Background/Objectives: Treatment of advanced knee osteoarthritis with total knee arthroplasty typically results in an improvement in function, gait, and quality of life, which tend to be affected by the condition. It is, however, necessary to determine the baseline factors that could influence the patients' postoperative outcome. Methods: This is a single-center prospective observational study of patients with advanced knee osteoarthritis (Kellgren-Lawrence grade 3 or 4) treated with total knee arthroplasty. Gait parameters were recorded at baseline and at various postoperative time points using a wireless device. Progression of function was assessed using the Knee Society Score questionnaire and quality of life by means of the EQ-5D and Knee Injury and Osteoarthritis Outcome Score questionnaires. Progression of gait and quality of life was analyzed in all patients, distinguishing between those where baseline velocity was < 1 m/s and those where it was ≥1 m/s. The potential correlation between baseline and postoperative parameters was also evaluated. Results: All 119 patients showed a significant improvement in their gait, function, and quality of life parameters at one year from the procedure (p < 0.05). No statistically significant differences were found in any of the postoperative subscales, regardless of baseline velocity (< o ≥ 1 m/s) or between any of the baseline or postoperative parameters (r < 0.29). Conclusions: Baseline gait parameters in patients with advanced knee osteoarthritis do not appear to bear a statistically significant relationship with function or quality of life outcomes following total knee arthroplasty. Such parameters exhibit a significant improvement one year after surgery.

Human fibroblasts from sporadic Alzheimer's disease (AD) patients show mitochondrial alterations and lysosome dysfunction.

Mitophagy is a mechanism that maintains mitochondrial integrity and homeostasis and is thought to promote longevity and reduce the risk of age-related neurodegenerative diseases, including Alzheimer's disease (AD). Here, we investigate the abundance of mitochondrial reactive oxygen species (ROS), mitochondrial function, and mitophagy in primary fibroblasts from patients with sporadic AD (sAD) and normal healthy controls. The results show increased levels of mitochondrial ROS, changes in mitochondrial morphology, altered bioenergetic properties, and defects in autophagy, mitophagy, and lysosome-mediated degradation pathways in sAD fibroblasts relative to control fibroblasts. Interestingly, lysosome abundance and the staining of lysosomal markers remained high, while the capacity of lysosome-dependent degradation was lower in sAD fibroblasts than in controls fibroblasts. Nicotinamide riboside supplementation decreased mitochondrial ROS, while capacity for lysosomal degradation remained unchanged in sAD fibroblasts relative to healthy control fibroblasts. These findings provide insight into molecular mechanisms involving the dysregulation of lysosome and autophagy/mitophagy pathways that may contribute significantly to clinical signs and pathological features of sAD.

Aging-dependent mitochondrial bioenergetic impairment in the skeletal muscle of NNT-deficient mice.

Overall health relies on features of skeletal muscle that generally decline with age, partly due to mechanisms associated with mitochondrial redox imbalance and bioenergetic dysfunction. Previously, aged mice genetically devoid of the mitochondrial NAD(P)+ transhydrogenase (NNT, encoded by the nicotinamide nucleotide transhydrogenase gene), an enzyme involved in mitochondrial NADPH supply, were shown to exhibit deficits in locomotor behavior. Here, by using young, middle-aged, and older NNT-deficient (Nnt-/-) mice and age-matched controls (Nnt+/+), we aimed to investigate how muscle bioenergetic function and motor performance are affected by NNT expression and aging. Mice were subjected to the wire-hang test to assess locomotor performance, while mitochondrial bioenergetics was evaluated in fiber bundles from the soleus, vastus lateralis and plantaris muscles. An age-related decrease in the average wire-hang score was observed in middle-aged and older Nnt-/- mice compared to age-matched controls. Although respiratory rates in the soleus, vastus lateralis and plantaris muscles did not significantly differ between the genotypes in young mice, the rates of oxygen consumption did decrease in the soleus and vastus lateralis muscles of middle-aged and older Nnt-/- mice. Notably, the soleus, which exhibited the highest NNT expression level, was the muscle most affected by aging, and NNT loss. Additionally, histology of the soleus fibers revealed increased numbers of centralized nuclei in older Nnt-/- mice, indicating abnormal morphology. In summary, our findings suggest that NNT expression deficiency causes locomotor impairments and muscle dysfunction during aging in mice.

Lack of NAD(P)+ transhydrogenase activity in patients with primary adrenal insufficiency due to NNT variants.

BACKGROUND: Pathogenic variants in the nicotinamide nucleotide transhydrogenase gene (NNT) are a rare cause of primary adrenal insufficiency (PAI), as well as functional impairment of the gonads. OBJECTIVE: Despite the description of different homozygous and compound heterozygous NNT variants in PAI patients, the extent to which the function and expression of the mature protein are compromised remains to be clarified. DESIGN: The activity and expression of mitochondrial NAD(P)+ transhydrogenase (NNT) were analyzed in blood samples obtained from patients diagnosed with PAI due to genetically confirmed variants of the NNT gene (n = 5), heterozygous carriers as their parents (n = 8), and healthy controls (n = 26). METHODS: NNT activity was assessed by a reverse reaction assay standardized for digitonin-permeabilized peripheral blood mononuclear cells (PBMCs). The enzymatic assay was validated in PBMC samples from a mouse model of NNT absence. Additionally, the PBMC samples were evaluated for NNT expression by western blotting and reverse transcription quantitative polymerase chain reaction and for mitochondrial oxygen consumption. RESULTS: NNT activity was undetectable (<4% of that of healthy controls) in PBMC samples from patients, independent of the pathogenic genetic variant. In patients' parents, NNT activity was approximately half that of the healthy controls. Mature NNT protein expression was lower in patients than in the control groups, while mRNA levels varied widely among genotypes. Moreover, pathogenic NNT variants did not impair mitochondrial bioenergetic function in PBMCs. CONCLUSIONS: The manifestation of PAI in NNT-mutated patients is associated with a complete lack of NNT activity. Evaluation of NNT activity can be useful to characterize disease-causing NNT variants.

Construcción y validación de un cuestionario para evaluar burnout académico en estudiantes de ciencias de la salud / Construction and validation of a questionnaire to assess academic burnout in health science students / Construção e validação de um questionário para avaliar o burnout acadêmico em estudantes da área da saúde

Objetivo: el burnout (BA) académico puede presentarse en el contexto universitario, se caracteriza por un estado de agotamiento vital que afecta la salud física y mental de los estudiantes, lo que genera una disminución en el rendimiento académico. Este problema aparece con mayor frecuencia en programas universitarios con altas exigencias académicas. El objetivo de este trabajo fue construir y evaluar la estructura factorial de un cuestionario de BA académico en estudiantes que cursaban los programas de medicina, enfermería y psicología. Metodología: los participantes fueron 710 estudiantes de ciencias de la salud (hombres 40.8 % y mujeres 59.2 %), de 16 a 33 años (M = 20.42 años, DT = 3 años). Se evaluó la validez de constructo mediante análisis factorial exploratorio (AFE) y análisis factorial confirmatorio (AFC); además, se calculó la consistencia interna por medio del estadístico alfa de Cronbach. Resultados: el cuestionario burnout académico (CBA-24) quedó conformado por 24 reactivos y una estructura factorial de cuatro dimensiones (agotamiento emocional, cinismo hacia el estudio, cinismo hacia las personas y realización personal). Con la prueba se evaluó el nivel de malestar emocional ante las demandas del entorno académico. Los índices de ajuste alcanzaron valores altos en el modelo propuesto, por lo tanto, el modelo de cuatro factores alcanzó los criterios para considerar que el ajuste es adecuado en todos los índices y mostró una estructura multidimensional. Dichos índices se agruparon de acuerdo con la taxonomía propuesta. Conclusiones: el cuestionario permitió identificar de manera ecológica el constructo de BA ajustado a las demandas de los contextos universitarios.

Objective: Academic burnout (AB) can occur in the university context and is characterized by a state of vital exhaustion that affects the physical and mental health of students, leading to a decrease in academic performance. This problem is more commonly observed in college programs with high academic demands. The aim of this study was to construct and evaluate the factorial structure of a questionnaire for assessing AB in students enrolled in medicine, nursing, and psychology programs. Methodology: The participants consisted of 710 health science students (40.8% male and 59.2% female) aged between 16 and 33 years (M = 20.42 years, SD = 3 years). Construct validity was assessed using exploratory factor analysis (EFA) and confirmatory factor analysis (CFA). Additionally, internal consistency was calculated using Cronbach's alpha statistic. Results: The academic burnout questionnaire (ABQ-24) consisted of 24 items and a four-factor factorial structure (emotional exhaustion, cynicism towards studying, cynicism towards people, and personal accomplishment). The questionnaire assessed the level of emotional distress experienced in response to academic demands. The fit indices achieved high values in the proposed structure, indicating that the four-factor model met the criteria for adequate fit across all indices and exhibited a multidimensional structure. These indices were grouped according to the proposed taxonomy. Conclusions: The questionnaire provided an ecologically valid means of identifying the construct of AB, adapted to the demands of university contexts.

Objetivo: o burnoutacadêmico (BA) pode ocorrer no contexto universitário, é caracteri-zado por um estado de esgotamento vital que afeta a saúdefísica e mentaldos alunos, o que gera uma diminuição no desempenho acadêmico. Esse problema aparece com mais frequência em programas universitários com altas demandas acadêmicas. O objetivo deste trabalho foi construir e avaliar a estrutura fatorial de um questionário acadêmico de graduação em estudantes de medicina, enfermagem e psicologia. Metodologia: Os participantes foram 710 estudantes de ciências da saúde (40,8% ho-mens e 59,2 % mulheres), de 16 a 33 anos (M = 20,42 anos, DT = 3 anos). A validade de construto foi avaliada por meio de análise fatorial exploratória (EFA) e análise fatorial confirmatória (AFC); além disso, a consistência interna foi calculada usando a estatística alfa de Cronbach. Resultados: o questionário de burnoutacadêmico (CBA-24) foi composto por 24 itens e uma estrutura fatorial de quatro dimensões (exaustão emocional, cinismo em relação ao estudo, cinismo em relação às pessoas e realização pessoal). Com o teste, avaliou-se o nível de desconforto emocional diante das demandas do ambiente acadêmico. Os índices de ajuste atingiram valores altos no modelo proposto, portanto, o modelo de quatro fatores atendeu aos critérios para considerar que o ajuste é adequado em todos os índices e apresentou uma estrutura multidimensional. Esses índices foram agrupados de acordo com a taxonomia proposta.Conclusões:o questionário permitiu identificar de forma ecológica o construto BA ajustado às demandas dos contextos universitários

Low Correlation between Gait and Quality of Life in Advanced Knee Osteoarthritis.

Advanced knee osteoarthritis patients' gait usually undergoes alterations leading to decreased mobility and lower functional performance, which can result in a worsening of their quality of life (QoL). While several authors have reported a moderate correlation between gait parameters and QoL assessed by generic questionnaires, the literature is scarce. This study aimed to explore the relationship between gait and QoL parameters assessed by a generic and a disease-specific questionnaire in patients with advanced knee osteoarthritis. In this single-centre, prospective, observational study, 129 patients with advanced knee osteoarthritis scheduled for elective total knee replacement were selected. The patients' gait was evaluated by means of a validated wireless device while they walked 30 m at a comfortable speed. Patient function was also analysed using the Knee Society Score (KSS). QoL was measured with the EQ-5D and the Knee Injury and Osteoarthritis Outcome Score (KOOS) questionnaires. Patients showed a mean walking speed of 0.95 ± 0.19 m/s, a mean cadence of 105.6 ± 9.9 steps/min, and a mean stride length of 1.25 ± 0.17 m on both legs. They presented poor knee status (KSS < 60) and poor QoL, with an EQ-5D of 0.44 ± 0.24 and a total KOOS of 29.77 ± 13.99. Positive low correlations (r <0.5, p <0.5) were found only between the speed, propulsion and stride length of both legs, and the overall and ADLs subscale scores of the total KOOS questionnaire. In conclusion, several gait parameters have a significant low correlation with the QoL of patients with advanced knee osteoarthritis, as assessed by an osteoarthritis-specific questionnaire.

Calidad de vida relacionada con edad e ingreso económico en adultos mayores diabéticos. Chillán, Chile, 2019

Fundamento padecer diabetes puede influenciar la percepción del propio individuo y cambiar su rutina diaria, afectando así su calidad de vida; partiendo de ello, la profundización en el tema reviste gran importancia Objetivo determinar la relación entre calidad de vida, edad e ingreso económico en adultos mayores con diabetes mellitus tipo 2. Métodos se realizó un estudio descriptivo, de corte transversal, en 33 adultos mayores con diabetes mellitus tipo 2, seleccionados mediante un muestreo no probabilístico por conveniencia, en la comunidad de las Mariposas (Chillán, Chile), en el año 2019. Para el análisis de la calidad de vida fue aplicado el instrumento Diabetes 39. Se utilizaron las pruebas estadísticas de Kolmogorov-Smirnov y Rho de Spearman. Resultados existió predominio del sexo femenino, así como de personas casadas. La edad tuvo un valor medio de 71,45 años. Con respecto al ingreso económico, un 66,7 % de los participantes refirió imposibilidad para costear los gastos del mes. Entre las dimensiones de la calidad de vida analizadas, las más afectadas fueron: severidad de la diabetes (media de 55,05), energía/movilidad (media de 44,1), y calidad de vida (media de 43,9). La edad y el ingreso económico no mostraron una correlación significativa con ninguna de las dimensiones estudiadas. Conclusiones la calidad de vida de los adultos mayores con diabetes mellitus tipo 2 se ve afectada, sin que exista una relación significativa con la edad e ingresos económicos.

Background suffering from diabetes can influence the perception of the individual himself and change his daily routine, thus affecting his quality of life; Based on this, the deepening of the subject is of great importance Objective to determine the relationship between quality of life, age and income in older adults with type 2 diabetes mellitus. Methods a descriptive, cross-sectional study was carried out in 33 older adults with type 2 diabetes mellitus, selected by non-probabilistic convenience sampling, in the Las Mariposas community (Chillán, Chile), in 2019. The Diabetes 39 instrument was applied for quality of life analysis. The Kolmogorov-Smirnov and Spearman's Rho statistical tests were used. Results the female sex was the predominant, as well as married people. Age had a mean value of 71.45 years. In the economic income, 66.7% of the participants reported the impossibility of paying the expenses of the month. Among the life's quality dimensions analyzed, the most affected were: diabetes severity (mean 55.05), energy/mobility (mean 44.1), and quality of life (mean 43.9). Age and income did not show a significant correlation with any of the dimensions studied. Conclusions the older adults' quality of life with type 2 diabetes mellitus is affected, without a significant relationship with age and economic income.

Neither Chia Flour nor Whey Protein Supplementation Further Improves Body Composition or Strength Gains after a Resistance Training Program in Young Subjects with a Habitual High Daily Protein Intake.

The aim of this study was to compare the potential additional effect of chia flour, whey protein, and a placebo juice to resistance training on fat-free mass (FFM) and strength gains in untrained young men. Eighteen healthy, untrained young men underwent an 8-week whole-body resistance training program, comprising three sessions per week. Subjects were randomized into three groups that after each training session consumed: (1) 30 g whey protein concentrate containing 23 g protein (WG), (2) 50 g chia flour containing 20 g protein (CG), or (3) a placebo not containing protein (PG). Strength tests (lower- and upper-limb one repetition maximum (1 RM) tests) and body composition analyses (dual-energy X-ray absorptiometry; DXA) were performed before (PRE) and after (POST) the intervention. Resistance training increased FFM and the 1 RM for each of the strength tests similarly in the three groups. FFM increased by 2.3% in WG (p = 0.04), by 3.6% in CG (p = 0.004), and by 3.0% in PG (p = 0.002)., and 1 RM increased in the different strength tests in the three groups (p < 0.05) with no difference between PG, CG, and WG. In conclusion, neither chia flour nor whey protein supplementation elicited an enhanced effect on FFM and strength gains after an 8-week resistance training program in healthy, untrained young men consuming a habitual high protein mixed diet (>1.2 g/kg/day).

Clinical profile of oral mucosa lesions in elderly patients: a 20-year retrospective study / Perfil clínico das lesões da mucosa oral em pacientes idosos: um estudo retrospectivo de 20 anos

Background: Life expectancy has considerably increased resulting in population aging. Studies evaluating the outcomes of aging in oral health are scarce. Objective: Evaluate retrospectively the profile of elderly patients from a public Oral Medicine Center during a period of 20 years. Methods:A qualitative and quantitative retrospective observational study was conducted analyzing medical records from an oral medicine service from January 1994 to December 2014. Results were reported as mean ± standard deviation for quantitative variables and percentages for categorical variables. The Chi-square test and T-student test was applied with significance level of 5%. Results: 2,690 medical records were retrieved, comprising of 61% women and 39% men with an average ageof 68.8 ± 6.79 years. Xerostomia was significantly associated, hypoglycemic usage (p<0.0001), anticoagulantusage (p<0.0001), psychotropic usage (p<0.0001) and analgesics and anti-inflammatory usage (p<0.0001). Forcandidiasis, an association with age, xerostomía (p<0.0001), and use of complete dentures was found(p<0.0001). For oral squamous cell carcinoma and oral leukoplakia the tabacco (p<0.0001) and alcohol consumption (p<0.0001) were significant associated. Conclusion:The elderly population was comprised mostly by women that use a large of drugs which wereassociated with xerostomia development. In addition, tabaco and alcohol consumption were associated withoral leukoplakia and OSCC being these two diseases more frequently in men. Dental care services should aimto prevent and treat these complications as way to improve the elderly's quality of life.

Introdução: A expectativa de vida aumentou consideravelmente, resultando no envelhecimento da população. Estudos avaliando os desfechos do envelhecimento na saúde bucal são escassos. Objetivo: Avaliar retrospectivamente o perfil dos pacientes idosos de um Centro de Medicina Oral público durante um período de 20 anos. Materiais e métodos: Estudo observacional retrospectivo qualitativo e quantitativo, analisando os prontuários de um serviço de medicina bucal no período de janeiro de 1994 a dezembro de 2014. Os resultados foram expressos em média ± desvio padrão para variáveis quantitativas e percentuais para variáveis categóricas. Aplicou-se o teste Qui-quadrado e o teste T-student com nível de significância de 5%. Resultados: Foram recuperados 2.690 prontuários, sendo 61% mulheres e 39% homens com idade média de 68,8 ± 6,79 anos. Xerostomia foi significativamente associada, uso de hipoglicemiantes (p<0,0001), uso de anticoagulantes (p<0,0001), uso de psicotrópicos (p<0,0001) e uso de analgésicos e anti-inflamatórios (p<0,0001). Para candidíase, foi encontrada associação com idade, xerostomia (p<0,0001)e uso de prótese total (p<0,0001). Para carcinoma espinocelular oral e leucoplasia oral, o uso de tabaco (p<0,0001) e consumo de álcool (p<0,0001) estiveram associados significativamente. Conclusão: A população idosa foi composta em sua maioria por mulheres que fazem uso de grande quantidade de medicamentos associados ao desenvolvimento de xerostomia. Além disso, o consumo de tabaco e álcool foram associados com leucoplasia oral e OSCC sendo essas duas doenças mais frequentes em homens. Os serviços odontológicos devem ter como objetivo prevenir e tratar essas complicações como forma de melhorar a qualidade de vida dos idosos.

Periodontal treatment of patients with hereditary hemorrhagic telangiectasia.

Hereditary hemorrhagic telangiectasia (HHT) is a rare condition in which fragile vascular walls lead to increased risks of bleeding, cerebral abscesses, arteriovenous malformations, anemia, and thrombosis. To date, no protocol has been established for optimizing the clinical outcomes of periodontal treatment in patients with this condition. The aim of this case report is to describe a safe clinical approach to periodontal treatment in a patient with HHT. A 39-year-old woman had a history of multiple macules on the oral mucosa, and a diagnosis of HHT was made based on the Curaçao diagnostic criteria (epistaxis, telangiectases, visceral lesions, and family history). Evaluation of the patient's periodontal clinical parameters and radiographs led to a diagnosis of generalized periodontitis, stage IV, grade C. The patient underwent nonsurgical periodontal therapy consisting of supragingival and subgingival scaling and root planing under a careful and specific protocol that included antibiotic prophylaxis before each session. Two months after therapy, the periodontal reevaluation showed improvement in the clinical parameters at most sites. Sites with remaining periodontal pockets were re-treated according to the same protocol, including the antibiotic prophylaxis. The patient was enrolled in a periodontal maintenance program, and her HHT was routinely monitored by her physician. Periodontal treatment may promote secondary complications in patients with HHT if appropriate systemic care is not provided, and the periodontal treatment plan should be designed individually for each patient. Establishing the correct HHT diagnosis and coordinating care with the patient's physician are essential to safe, effective treatment.

JMJD3 intrinsically disordered region links the 3D-genome structure to TGFß-dependent transcription activation.

Enhancers are key regulatory elements that govern gene expression programs in response to developmental signals. However, how multiple enhancers arrange in the 3D-space to control the activation of a specific promoter remains unclear. To address this question, we exploited our previously characterized TGFß-response model, the neural stem cells, focusing on a ~374 kb locus where enhancers abound. Our 4C-seq experiments reveal that the TGFß pathway drives the assembly of an enhancer-cluster and precise gene activation. We discover that the TGFß pathway coactivator JMJD3 is essential to maintain these structures. Using live-cell imaging techniques, we demonstrate that an intrinsically disordered region contained in JMJD3 is involved in the formation of phase-separated biomolecular condensates, which are found in the enhancer-cluster. Overall, in this work we uncover novel functions for the coactivator JMJD3, and we shed light on the relationships between the 3D-conformation of the chromatin and the TGFß-driven response during mammalian neurogenesis.

Aggravation of hepatic lipidosis in red-footed tortoise Chelonoidis carbonaria with age is associated with alterations in liver mitochondria.

The occurrence of hepatic lipidosis is commonly reported in different reptilian species, especially in animals under captivity. Liver accumulation of fat is associated with disorders, better described in mammals as non-alcoholic fatty liver diseases (NAFLD), ranging from simple steatosis, to non-alcoholic steatohepatitis (NASH), and to more severe lesions of cirrhosis and hepatocellular carcinoma. Mitochondria play a central role in NAFLD pathogenesis, therefore in this study we characterized livers of ad libitum fed captive red-footed tortoise Chelonoidis carbonaria through histological and mitochondrial function evaluations of juvenile and adult individuals. Livers from adult tortoises exhibited higher levels of lipids, melanomacrophages centers and melanin than juveniles. The observed high score levels of histopathological alterations in adult tortoises, such as microvesicular steatosis, inflammation and fibrosis, indicated the progression to a NASH condition. Mitochondrial oxygen consumption at different respiratory states and with different substrates was 30 to 58% lower in adult when compared to juvenile tortoises. Despite citrate synthase activity was also lower in adults, cardiolipin content was similar to juveniles, indicating that mitochondrial mass was unaffected by age. Mitochondrial Ca2+ retention capacity was reduced by 70% in adult tortoises. Overall, we found that aggravation of NAFLD in ad libitum fed captive tortoises is associated with compromised mitochondrial function, indicating a critical role of the organelle in liver disease progression in reptiles.

Dichloroacetate reactivates pyruvate-supported peroxide removal by liver mitochondria and prevents NAFLD aggravation in NAD(P)+ transhydrogenase-null mice consuming a high-fat diet.

The mechanisms by which a high-fat diet (HFD) promotes non-alcoholic fatty liver disease (NAFLD) appear to involve liver mitochondrial dysfunction and redox imbalance. The functional loss of the enzyme NAD(P)+ transhydrogenase, a main source of mitochondrial NADPH, results in impaired mitochondrial peroxide removal, pyruvate dehydrogenase inhibition by phosphorylation, and progression of NAFLD in HFD-fed mice. The present study aimed to investigate whether pharmacological reactivation of pyruvate dehydrogenase by dichloroacetate attenuates the mitochondrial redox dysfunction and the development of NAFLD in NAD(P)+ transhydrogenase-null (Nnt-/-) mice fed an HFD (60% of total calories from fat). For this purpose, Nnt-/- mice and their congenic controls (Nnt+/+) were fed chow or an HFD for 20 weeks and received sodium dichloroacetate or NaCl in the final 12 weeks via drinking water. The results showed that HFD reduced the ability of isolated liver mitochondria from Nnt-/- mice to remove peroxide, which was prevented by the dichloroacetate treatment. HFD-fed mice of both Nnt genotypes exhibited increased body and liver mass, as well as a higher content of hepatic triglycerides, but dichloroacetate treatment attenuated these abnormalities only in Nnt-/- mice. Notably, dichloroacetate treatment decreased liver pyruvate dehydrogenase phosphorylation levels and prevented the aggravation of NAFLD in HFD-fed Nnt-/- mice. Conversely, dichloroacetate treatment elicited moderate hepatocyte ballooning in chow-fed mice, suggesting potentially toxic effects. We conclude that the protection against HFD-induced NAFLD by dichloroacetate is associated with its role in reactivating pyruvate dehydrogenase and reestablishing the pyruvate-supported liver mitochondrial capacity to handle peroxide in Nnt-/- mice.

A dataset describing glycolytic inhibitors overcoming the underestimation of maximal mitochondrial oxygen consumption rate in oligomycin-treated cells.

Determination of oxygen consumption is one of the most valuable methodologies to evaluate mitochondrial (dys)function. Previous studies demonstrated that a widely used protocol, consisting of adding the ATP synthase inhibitor oligomycin before mitochondrial respiratory uncoupling by sequential addition of a protonophore (e.g., carbonyl cyanide 3-chlorophenyl hydrazone [CCCP]), may lead to underestimation of maximal oxygen consumption rate (OCRmax) and spare respiratory capacity (SRC) parameters in highly glycolytic tumor cell lines. In this dataset, we report the effects of the glycolytic inhibitors 2-deoxy-D-glucose, iodoacetic acid, and lonidamine on overcoming the underestimation of OCRmax and SRC in oligomycin-treated cells. We propose a protocol in which 2-deoxy-D-glucose is added after oligomycin and just before the sequential addition of CCCP to avoid underestimation of OCRmax and SRC parameters in A549, C2C12, and T98G cells. The oxygen consumption rates were determined in intact suspended cell lines using a high-resolution oxygraph device. The data can be used in several fields of research that require characterization of mitochondrial respiratory parameters in intact cells.

Enhanced resistance to Ca2+-induced mitochondrial permeability transition in the long-lived red-footed tortoise Chelonoidis carbonaria.

The interaction between supraphysiological cytosolic Ca2+ levels and mitochondrial redox imbalance mediates the mitochondrial permeability transition (MPT). The MPT is involved in cell death, diseases and aging. This study compared the liver mitochondrial Ca2+ retention capacity and oxygen consumption in the long-lived red-footed tortoise (Chelonoidis carbonaria) with those in the rat as a reference standard. Mitochondrial Ca2+ retention capacity, a quantitative measure of MPT sensitivity, was remarkably higher in tortoises than in rats. This difference was minimized in the presence of the MPT inhibitors ADP and cyclosporine A. However, the Ca2+ retention capacities of tortoise and rat liver mitochondria were similar when both MPT inhibitors were present simultaneously. NADH-linked phosphorylating respiration rates of tortoise liver mitochondria represented only 30% of the maximal electron transport system capacity, indicating a limitation imposed by the phosphorylation system. These results suggested underlying differences in putative MPT structural components [e.g. ATP synthase, adenine nucleotide translocase (ANT) and cyclophilin D] between tortoises and rats. Indeed, in tortoise mitochondria, titrations of inhibitors of the oxidative phosphorylation components revealed a higher limitation of ANT. Furthermore, cyclophilin D activity was approximately 70% lower in tortoises than in rats. Investigation of critical properties of mitochondrial redox control that affect MPT demonstrated that tortoise and rat liver mitochondria exhibited similar rates of H2O2 release and glutathione redox status. Overall, our findings suggest that constraints imposed by ANT and cyclophilin D, putative components or regulators of the MPT pore, are associated with the enhanced resistance to Ca2+-induced MPT in tortoises.

Feeding effects on liver mitochondrial bioenergetics of Boa constrictor (Serpentes: Boidae).

Snakes are interesting examples of taxa that can overcome energy metabolism challenges, as many species can endure long periods without feeding, and their eventual meals are of reasonably large sizes, thus exhibiting dual extreme adaptations. Consequently, metabolic rate increases considerably to attend to the energetic demand of digestion, absorption and protein synthesis. These animals should be adapted to transition from these two opposite states of energy fairly quickly, and therefore we investigated mitochondrial function plasticity in these states. Herein, we compared liver mitochondrial bioenergetics of the boid snake Boa constrictor during fasting and after meal intake. We fasted the snakes for 60 days, and then we fed a subgroup with 30% of their body size and evaluated their maximum postprandial response. We measured liver respiration rates from permeabilized tissue and isolated mitochondria. From isolated mitochondria, we also measured Ca2+ retention capacity and redox status. Mitochondrial respiration rates were maximized after feeding, reaching an approximately 60% increase from fasting levels when energized with complex I-linked substrates. Interestingly, fasting and fed snakes exhibited similar respiratory control ratios and citrate synthase activity. Furthermore, we found no differences in Ca2+ retention capacity, indicating no increase in susceptibility to mitochondrial permeability transition, and no changes in mitochondrial redox state, although fed animals exhibited increases in the release of H2O2. Thus, we conclude that liver mitochondria from B. constrictor snakes increase respiration rates during the postprandial period and quickly improve the bioenergetic capacity without compromising redox balance.

The histone demethylase PHF8 regulates astrocyte differentiation and function.

Epigenetic factors have been shown to play a crucial role in X-linked intellectual disability (XLID). Here, we investigate the contribution of the XLID-associated histone demethylase PHF8 to astrocyte differentiation and function. Using genome-wide analyses and biochemical assays in mouse astrocytic cultures, we reveal a regulatory crosstalk between PHF8 and the Notch signaling pathway that balances the expression of the master astrocytic gene Nfia. Moreover, PHF8 regulates key synaptic genes in astrocytes by maintaining low levels of H4K20me3. Accordingly, astrocytic-PHF8 depletion has a striking effect on neuronal synapse formation and maturation in vitro. These data reveal that PHF8 is crucial in astrocyte development to maintain chromatin homeostasis and limit heterochromatin formation at synaptogenic genes. Our studies provide insights into the involvement of epigenetics in intellectual disability.