First person – Juan Garrido-Maraver

First Person is a series of interviews with the first authors of a selection of papers published in Biology Open, helping early-career researchers promote themselves alongside their papers. Juan Garrido-Maraver is first author on 'Forcing contacts between mitochondria and the endoplasmic reticulum extends lifespan in a Drosophila model of Alzheimer's disease', published in BiO. Juan conducted the research described in this article while a postdoctoral scientist in L. Miguel Martins's lab at the MRC Toxicology Unit, University of Cambridge, Leicester, UK. He is now a postdoctoral scientist in the lab of Alvaro A. Tavares at the Centre for Biomedical Research (CBMR), University of Algarve, Faro, Portugal, investigating molecular mechanisms linked to human diseases from a therapeutic point of view.info:eu-repo/semantics/publishedVersio

Differential pathophysiology in MELAS syndrome

Programa de Doctorado en Biotecnología y Tecnología QuímicaMELAS (mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes) is a mitochondrial disorder caused mainly by the m.3243A>G mutation in mitochondrial DNA. In this thesis, we report on how the severity of pathophysiological alterations is differently expressed in fibroblasts derived from patients with MELAS disease. We evaluated mitophagy activation and mitochondrial biogenesis which are the main mechanisms regulating the degradation and genesis of mitochondrial mass in transmitochondrial cybrids and fibroblasts derived form MELAS patients. Our results suggest a critical balance between mitophagy and mitochondrial biogenesis which leads to the expression of different degrees of pathological severity among MELAS fibroblast cell lines according to their heteroplasmy load and the activation of AMP-activated protein kinase (AMPK). AMPK-activators such as 5-aminoimidazole-4-carboxamide 1-ß-D-ribofuranoside (AICAR) or coenzyme Q10 (CoQ) increased peroxisome proliferator-activated receptor alpha (PGC-1¿) nuclear translocation, mitochondrial biogenesis, antioxidant enzyme system response, autophagic flux and ultimately improved pathophysiological alterations in MELAS fibroblasts with the most severe phenotype. Our findings support the hypothesis that mitochondrial biogenesis, increased antioxidant response and autophagy clearance serve as compensatory mechanisms in response to mitophagic degradation of dysfunctional mitochondria and point out that AMPK is an important player in this balance. These results are particularly important since currently no efficient treatments are available for this chronic progressive disorder. Furthermore, in this thesis we propose the evaluation of the effectiveness of putative beneficial pharmacological agents in the treatment of MELAS by using cellular models such as transmitochondrial cybrids and fibroblasts with high mutational load. According to our results, supplementation with riboflavin or coenzyme Q10 effectively reversed the pathologic alterations in MELAS cybrid and fibroblast cell models. Our results indicate that cell models manifesting severe pathophysiological alterations and high heteroplasmy load have great potential as a screening and validation assays of novel drug candidates for MELAS treatment and presumably also for other diseases with mitochondrial impairmentUniversidad Pablo de Olavide. Departamento de Fisiología, Anatomía y Biología Celula

AMPK Phosphorylation Modulates Pain by Activation of NLRP3 Inflammasome

Impairment in adenosine monophosphate-activated protein kinase (AMPK) activity and NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome activation are associated with several metabolic and inflammatory diseases. In this study, we investigated the role of AMPK/NLRP3 inflammasome axis in the molecular mechanism underlying pain perception. Results: Impairment in AMPK activation induced by compound C or sunitinib, two AMPK inhibitors, provoked hyperalgesia in mice ( p < 0.001) associated with marked NLRP3 inflammasome protein activation and increased serum levels of interleukin-1b (IL-1b) (24.56 – 0.82 pg/ml) and IL-18 (23.83 – 1.882 pg/ml) compared with vehicle groups (IL-1b: 8.15 – 0.44; IL-18:4.92 – 0.4). This effect was rescued by increasing AMPK phosphorylation via metformin treatment ( p < 0.001), caloric restriction diet ( p < 0.001), or NLRP3 inflammasome genetic inactivation using NLRP3 knockout (nlrp3-/ - ) mice ( p < 0.001). Deficient AMPK activation and overactivation of NLRP3 inflammasome axis were also observed in blood cells from patients with fibromyalgia (FM), a prevalent human chronic pain disease. In addition, metformin treatment (200 mg/daily), which increased AMPK activation, restored all biochemical alterations examined by us in blood cells and significantly improved clinical symptoms, such as, pain, fatigue, depression, disturbed sleep, and tender points, in patients with FM. Innovation and Conclusions: These data suggest that AMPK/NLRP3 inflammasome axis participates in chronic pain and that NLRP3 inflammasome inhibition by AMPK modulation may be a novel therapeutic target to fight against chronic pain and inflammatory diseases as FM. Antioxid. Redox Signal. 24, 157–170.Junta de Andalucía CTS11

Role of MOB4 in cell proliferation and neurogenesis

Signaling pathways that integrate a large set of inputs (both extra- and intracellular) to control cell proliferation are essential during both development and adult stages to guarantee organism homeostasis. Mobs are small adaptor proteins that participate in several of these signaling pathways. Here, we review recent advances unravelling Mob4 cellular functions, a highly conserved non-catalytic protein, that plays a diversity of roles in cell proliferation, sperm cell differentiation and is simultaneously involved in synapse formation and neural development. In addition, the gene is often overexpressed in a large diversity of tumors and is linked to poor clinical outcomes. Nevertheless, Mob4 molecular functions remain poorly defined, although it integrates the core structure of STRIPAK, a kinase/phosphatase protein complex, that can act upstream of the Hippo pathway. In this review we focus on the recent findings of Mob4 functions, that have begun to clarify its critical role on cell proliferation and the development of tissues and individuals.Grant number ALG-01-0145-FEDER-030014info:eu-repo/semantics/publishedVersio

Enhancing folic acid metabolism suppresses defects associated with loss of Drosophila mitofusin.

Mutations in the mitochondrial GTPase mitofusin 2 (MFN2) cause Charcot-Marie-Tooth disease type 2 (CMT2A), a form of peripheral neuropathy that compromises axonal function. Mitofusins promote mitochondrial fusion and regulate mitochondrial dynamics. They are also reported to be involved in forming contacts between mitochondria and the endoplasmic reticulum. The fruit fly, Drosophila melanogaster, is a powerful tool to model human neurodegenerative diseases, including CMT2A. Here, we have downregulated the expression of the Drosophila mitofusin (dMfn RNAi) in adult flies and showed that this activates mitochondrial retrograde signalling and is associated with an upregulation of genes involved in folic acid (FA) metabolism. Additionally, we demonstrated that pharmacological and genetic interventions designed to increase the FA metabolism pathway suppresses the phenotype of the dMfn RNAi flies. We conclude that strategies to increase FA metabolism may ameliorate diseases, such as peripheral neuropathies, that are associated with loss of mitochondrial function. A video abstract for this article is available at  https://youtu.be/fs1G-QRo6xI .Medical Research Counci

Pharmacological Chaperones and Coenzyme Q10 Treatment Improves Mutant β-Glucocerebrosidase Activity and Mitochondrial Function in Neuronopathic Forms of Gaucher Disease

Gaucher disease (GD) is caused by mutations in the GBA1 gene, which encodes lysosomal β-glucocerebrosidase. Homozygosity for the L444P mutation in GBA1 is associated with high risk of neurological manifestations which are not improved by enzyme replacement therapy. Alternatively, pharmacological chaperones (PCs) capable of restoring the correct folding and trafficking of the mutant enzyme represent promising alternative therapies.Here, we report on how the L444P mutation affects mitochondrial function in primary fibroblast derived from GD patients. Mitochondrial dysfunction was associated with reduced mitochondrial membrane potential, increased reactive oxygen species (ROS), mitophagy activation and impaired autophagic flux.Both abnormalities, mitochondrial dysfunction and deficient β-glucocerebrosidase activity, were partially restored by supplementation with coenzyme Q10 (CoQ) or a L-idonojirimycin derivative, N-[N’-(4-adamantan-1-ylcarboxamidobutyl)thiocarbamoyl]-1,6-anhydro-L-idonojirimycin (NAdBT-AIJ), and more markedly by the combination of both treatments. These data suggest that targeting both mitochondria function by CoQ and protein misfolding by PCs can be promising therapies in neurological forms of GD.España, Ministerio de Sanidad FIS PI13/00129España, Ministerio de Economía y Competitividad SAF2013-44021-R and CTQ2010-15848España, Junta de Andalucía CTS-5725 and FQM-146

Critical role of AMP-activated protein kinase in the balance between mitophagy and mitochondrial biogenesis in MELAS disease

MELAS syndrome is a mitochondrial disorder that is caused mainly by the m.3243A > G mutation in mitochondrial DNA. Here, we report on how the severity of pathophysiological alterations is differently expressed in fibroblasts derived from patients with MELAS disease. We evaluated mitophagy activation and mitochondrial biogenesis which are the main mechanisms regulating the degradation and genesis of mitochondrial mass in MELAS fibroblasts and transmitochondrial cybrids. Our results suggest a critical balance between mitophagy and mitochondrial biogenesis which leads to the expression of different degrees of pathological severity among MELAS fibroblast cell lines according to their heteroplasmy load and the activation of AMP-activated protein kinase (AMPK). AMPK-activators such as 5-aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR) or coenzyme Q10 (CoQ) increased peroxisome proliferator-activated receptor alpha (PGC-1α) nuclear translocation, mitochondrial biogenesis, antioxidant enzyme system response, autophagic flux and improved pathophysiological alterations in MELAS fibroblasts with the most severe phenotype. Our findings support the hypothesis that mitochondrial biogenesis, increased antioxidant response and autophagy clearance serve as compensatory mechanisms in response to mitophagic degradation of dysfunctional mitochondria and point out that AMPK is an important player in this balance.This work was supported by FIS PI13/00129 grant, Ministerio de Sanidad, Spain and Fondo Europeo de Desarrollo Regional (FEDER-Unión Europea), Proyecto de Investigación de Excelencia de la Junta de AndalucíaCTS-5725, and by AEPMI (Asociación de Enfermos de Patología Mitocondrial).Peer Reviewe

Mitochondrial dysfunction and mitophagy activation in blood mononuclear cells of fibromyalgia patients: implications in the pathogenesis of the disease

This is an open access article distributed under the terms of the Creative Commons Attribution License.[Introduction]: Fibromyalgia is a chronic pain syndrome with unknown etiology. Recent studies have shown some evidence demonstrating that oxidative stress may have a role in the pathophysiology of fibromyalgia. However, it is still not clear whether oxidative stress is the cause or the effect of the abnormalities documented in fibromyalgia. Furthermore, the role of mitochondria in the redox imbalance reported in fibromyalgia also is controversial. We undertook this study to investigate the role of mitochondrial dysfunction, oxidative stress, and mitophagy in fibromyalgia. [Methods]: We studied 20 patients (2 male, 18 female patients) from the database of the Sevillian Fibromyalgia Association and 10 healthy controls. We evaluated mitochondrial function in blood mononuclear cells from fibromyalgia patients measuring, coenzyme Q10 levels with high-performance liquid chromatography (HPLC), and mitochondrial membrane potential with flow cytometry. Oxidative stress was determined by measuring mitochondrial superoxide production with MitoSOX™ and lipid peroxidation in blood mononuclear cells and plasma from fibromyalgia patients. Autophagy activation was evaluated by quantifying the fluorescence intensity of LysoTracker™ Red staining of blood mononuclear cells. Mitophagy was confirmed by measuring citrate synthase activity and electron microscopy examination of blood mononuclear cells. [Results]: We found reduced levels of coenzyme Q10, decreased mitochondrial membrane potential, increased levels of mitochondrial superoxide in blood mononuclear cells, and increased levels of lipid peroxidation in both blood mononuclear cells and plasma from fibromyalgia patients. Mitochondrial dysfunction was also associated with increased expression of autophagic genes and the elimination of dysfunctional mitochondria with mitophagy. [Conclusions]: These findings may support the role of oxidative stress and mitophagy in the pathophysiology of fibromyalgia.This work was supported by grants FIS PI080500 and FIS EC08/00076, Ministerio de Sanidad, Spain. The authors dedicate this manuscript to FM patients and AFIBROSE (Asociación de Fibromialgia de Sevilla) for their unconditional help.Peer Reviewe

Coenzyme Q10 therapy

For a number of years, coenzyme Q10 (CoQ10) was known for its key role in mitochondrial bioenergetics; later studies demonstrated its presence in other subcellular fractions and in blood plasma, and extensively investigated its antioxidant role. These 2 functions constitute the basis for supporting the clinical use of CoQ10. Also, at the inner mitochondrial membrane level, CoQ10 is recognized as an obligatory cofactor for the function of uncoupling proteins and a modulator of the mitochondrial transition pore. Furthermore, recent data indicate that CoQ 10 affects the expression of genes involved in human cell signaling, metabolism and transport, and some of the effects of CoQ10 supplementation may be due to this property. CoQ10 deficiencies are due to autosomal recessive mutations, mitochondrial diseases, aging-related oxidative stress and carcinogenesis processes, and also statin treatment. Many neurodegenerative disorders, diabetes, cancer, and muscular and cardiovascular diseases have been associated with low CoQ10 levels as well as different ataxias and encephalomyopathies. CoQ10 treatment does not cause serious adverse effects in humans and new formulations have been developed that increase CoQ10 absorption and tissue distribution. Oral administration of CoQ10 is a frequent antioxidant strategy in many diseases that may provide a significant symptomatic benefit.This work was supported by grants (FIS PI10/00543, FIS EC08/00076) from the Ministerio de Sanidad, Spain, and Fondo Europeo de Desarrollo Regional (FEDER-Unión Europea); Servicio Andaluz de Salud-Junta de Andalucía (SAS 111242); Proyecto de Investigación de Excelencia de la Junta de Andalucía (CTS-5725); and by AEPMI (Asociación de Enfermos de Patología Mitocondrial), FEEL (Fundación Española de Enfermedades Lisosomales) and ALBA Andalucía (Federación Andaluza de Fibromialgia y Fatiga Crónica).Peer Reviewe