Technological and Biotechnological Processes To Enhance the Bioavailability of Dietary (Poly)phenols in Humans.

The health effects of (poly)phenols (PPs) depend upon their bioavailability that, in general, is very low and shows a high interindividual variability. The low bioavailability of PPs is mainly attributed to their low absorption in the upper gastrointestinal tract as a result of their low water solubility, their presence in foods as polymers or in glycosylated forms, and their tight bond to food matrices. Although many studies have investigated how technological and biotechnological processes affect the phenolic composition of fruits and vegetables, limited information exists regarding their effects on PP bioavailability in humans. In the present review, the effect of food processing (mechanical, thermal, and non-thermal treatments), oral-delivery nanoformulations, enzymatic hydrolysis, fermentation, co-administration with probiotics, and generation of postbiotics in PP bioavailability have been overviewed, focusing in the evidence provided in humans.

Clinical, pathological and genetic spectrum in 89 cases of mitochondrial progressive external ophthalmoplegia.

BACKGROUND: Mitochondrial progressive external ophthalmoplegia (PEO) encompasses a broad spectrum of clinical and genetic disorders. We describe the phenotypic subtypes of PEO and its correlation with molecular defects and propose a diagnostic algorithm. METHODS: Retrospective analysis of the clinical, pathological and genetic features of 89 cases. RESULTS: Three main phenotypes were found: 'pure PEO' (42%), consisting of isolated palpebral ptosis with ophthalmoparesis; Kearns-Sayre syndrome (10%); and 'PEO plus', which associates extraocular symptoms, distinguishing the following subtypes: : myopathic (33%), bulbar (12%) and others (3%). Muscle biopsy was the most accurate test, showing mitochondrial changes in 95%. Genetic diagnosis was achieved in 96% of the patients. Single large-scale mitochondrial DNA (mtDNA) deletion was the most frequent finding (63%), followed by multiple mtDNA deletions (26%) due to mutations in TWNK (n=8), POLG (n=7), TK2 (n=6) or RRM2B (n=2) genes, and point mtDNA mutations (7%). Three new likely pathogenic mutations were identified in the TWNK and MT-TN genes. CONCLUSIONS: Phenotype-genotype correlations cannot be brought in mitochondrial PEO. Muscle biopsy should be the first step in the diagnostic flow of PEO when mitochondrial aetiology is suspected since it also enables the study of mtDNA rearrangements. If no mtDNA deletions are identified, whole mtDNA sequencing should be performed.

Different mitochondrial genetic defects exhibit the same protein signature of metabolism in skeletal muscle of PEO and MELAS patients: A role for oxidative stress.

A major challenge in mitochondrial diseases (MDs) is the identification of biomarkers that could inform of the mechanisms involved in the phenotypic expression of genetic defects. Herein, we have investigated the protein signature of metabolism and of the antioxidant response in muscle biopsies of clinically and genetically diagnosed patients with Progressive External Ophthalmoplegia due to single large-scale (PEO-sD) or multiple (PEO-mD) deletions of mtDNA and Mitochondrial Encephalopathy Lactic Acidosis and Stroke-like episode (MELAS) syndrome, and healthy donors. A high-throughput immunoassay technique that quantitates the expression of relevant proteins of glycolysis, glycogenolysis, pentose phosphate pathway, oxidative phosphorylation, pyruvate and fatty acid oxidation, tricarboxylic acid cycle and the antioxidant response in two large independent and retrospectively collected cohorts of PEO-sD, PEO-mD and MELAS patients revealed that despite the heterogeneity of the genetic alterations, the three MDs showed the same metabolic signatures in both cohorts of patients, which were highly divergent from those of healthy individuals. Linear Discriminant Analysis and Support Vector Machine classifier provided a minimum of four biomarkers to discriminate healthy from pathological samples. Regardless of the induction of a large number of enzymes involved in ameliorating oxidative stress, the down-regulation of mitochondrial superoxide dismutase (SOD2) and catalase expression favored the accumulation of oxidative damage in patients' proteins. Down-regulation of SOD2 and catalase expression in MD patients is not due to relevant changes in the availability of their mRNAs, suggesting that oxidative stress regulates the expression of the two enzymes post-transcriptionally. We suggest that SOD2 and catalase could provide specific targets to improve the detoxification of reactive oxygen species that affects muscle proteins in these patients.

Beyond cervical lipomas: myoclonus, gait disorder and multisystem involvement leading to mitochondrial disease.

Madelung's disease (benign symmetric lipomatosis) is a rare syndrome in which there are multiple lipomas around the neck, upper limbs and trunk in the context of chronic alcoholism. We report on a female patient with lipomas and slightly progressive myoclonus, neuropathy, myopathy, ataxia and respiratory systemic involvement (labelled in the past as Madelung's disease). Multisystem involvement and family history of lipomas led to the development of mitochondrial genetic tests, which can assess two concurrent mitochondrial mutations: the m.8344A>G mutation in MT-TK gene, related MERRF (myoclonic epilepsy with ragged-red fibre) phenotype and m.14484T>C mutation in the MT-ND6 gene responsible for Leber hereditary optic neuropathy phenotype.

Secondary coenzyme Q10 deficiencies in oxidative phosphorylation (OXPHOS) and non-OXPHOS disorders.

We evaluated the coenzyme Q10 (CoQ) levels in patients who were diagnosed with mitochondrial oxidative phosphorylation (OXPHOS) and non-OXPHOS disorders (n=72). Data from the 72 cases in this study revealed that 44.4% of patients showed low CoQ concentrations in either their skeletal muscle or skin fibroblasts. Our findings suggest that secondary CoQ deficiency is a common finding in OXPHOS and non-OXPHOS disorders. We hypothesize that cases of CoQ deficiency associated with OXPHOS defects could be an adaptive mechanism to maintain a balanced OXPHOS, although the mechanisms explaining these deficiencies and the pathophysiological role of secondary CoQ deficiency deserves further investigation.

Bulk autophagy, but not mitophagy, is increased in cellular model of mitochondrial disease.

Oxidative phosphorylation system (OXPHOS) deficiencies are rare diseases but constitute the most frequent inborn errors of metabolism. We analyzed the autophagy route in 11 skin fibroblast cultures derived from patients with well characterized and distinct OXPHOS defects. Mitochondrial membrane potential determination revealed a tendency to decrease in 5 patients' cells but reached statistical significance only in 2 of them. The remaining cells showed either no change or a slight increase in this parameter. Colocalization analysis of mitochondria and autophagosomes failed to show evidence of increased selective elimination of mitochondria but revealed more intense autophagosome staining in patients' fibroblasts compared with controls. Despite the absence of increased mitophagy, Parkin recruitment to mitochondria was detected in both controls' and patients' cells and was slightly higher in cells harboring complex I defects. Western blot analysis of the autophagosome marker LC3B, confirmed significantly higher levels of the protein bound to autophagosomes, LC3B-II, in patients' cells, suggesting an increased bulk autophagy in OXPHOS defective fibroblasts. Inhibition of lysosomal proteases caused significant accumulation of LC3B-II in control cells, whereas in patients' cells this phenomenon was less pronounced. Electron microscopy studies showed higher content of late autophagic vacuoles and lysosomes in OXPHOS defective cells, accompanied by higher levels of the lysosomal marker LAMP-1. Our findings suggest that in OXPHOS deficient fibroblasts autophagic flux could be partially hampered leading to an accumulation of autophagic vacuoles and lysosomes.

A proposed molecular diagnostic flowchart for myophosphorylase deficiency (McArdle disease) in blood samples from Spanish patients.

McArdle disease is a metabolic myopathy due to molecular defects in the myophosphorylase gene (PYGM), usually diagnosed in muscle biopsy. The aims of this study were to characterize genetically a large series of patients and to establish a protocol of molecular diagnosis on blood samples. We studied 55 Spanish unrelated patients with McArdle disease. Screening for the three more frequent mutations in the PYGM gene in the Spanish population (c.148C>T, p.R50X; c.613G>A, p.G205S; and c.2392T>C, p.W798R) were performed with polymerase chain-reaction and restriction fragment length polymorphism (PCR-RFLP) methods. To identify other mutant alleles, the coding region of PYGM gene was sequenced. The p.R50X mutation was observed in 38 patients, the p.G205S substitution in eight, and the p.W798R change in nine. Nine novel mutations, five missense (c.247A>T, p.I83F; c.521G>A, p.G174D; c.1094C>T, p.A365V; c.1468C>T, p.R490W; and c.1730A>G, p.Q577R), one nonsense mutation (c.2352C>A, p.C784X), three frameshift (c.402del, p.N134KfsX161; c.212_218dup, p.Q73HfsX7; c.1470dup, p.R491AfsX7), and nine previously reported mutations were found. In addition, we also updated the molecular data of 95 unrelated patients with McArdle disease studied thus far in our center. Of these patients, 56 were either homozygous or compound heterozygous for the p.R50X, p.G205S, or p.W798R mutation. By including in the molecular diagnosis protocol sequencing of the exons 1, 14, 17 and 18 of the PYGM gene, 16 further patients were characterized, and therefore we were able to detect the molecular defect in 72 out of 95 patients. A proposed molecular diagnosis protocol of the disease based on blood DNA would avoid muscle biopsy in 75.8% [95% confidence interval (95% CI): 62.1%-78.6%] of patients with McArdle disease.