The ratio of lentic to lotic habitat features strongly affects macroinvertebrate metrics used in southern Europe for ecological status classification

Abstract Biological quality in rivers based on benthic macroinvertebrates is typically assessed by comparison with expected reference conditions, which represent relatively undisturbed situations. Commonly, reference conditions are set in agreement with river typologies to handle major ecological differences and limit biological variability. Although natural hydrological variation can be highly influential, site-specific tuning of reference conditions is rare in Mediterranean countries. River flow and local hydraulics change continuously over time, shaping the occurrence of lentic and lotic habitat features. Thus, biological reference conditions might require site-specific adjustment based on the ratio of lentic to lotic habitats assessed at the time of sampling. This would help reducing systematic bias in ecological assessments, interpreting benthic invertebrate responses to pressures, and diminishing the amount of unexplained biological variability. In this study, the response to the lentic-lotic character of river reaches was assessed for nineteen macroinvertebrate metrics and indices commonly used for the classification of ecological status in South European rivers. The study sites, with a prevalent temporary character, were located in Sardinia, southwestern Italy. Most metrics were significantly related to the lentic-lotic habitat conditions, both in pool and riffle mesohabitats, and their response curves were either parabolic or linearly decreasing at increasing lentic conditions. Taxonomic richness, score-based metrics, ovoviviparous taxa and multi-metric indices related well to the lentic-lotic conditions, while abundance metrics correlated less. The potential impact on ecological status classification was tested for the method formally used in Italy, which had a major role in comparing and inter-calibrating European assessment methods for the Water Framework Directive. After adjusting for bias due to the ratio of lentic to lotic habitat features, quality classification shifted towards better ecological status for ≈ 23% samples. This highlighted the impact of ignoring lentic-lotic information when defining reference conditions for assessing ecological status, varying from difficulties in understanding the biological response to pressures, to largely biased ecological status classification. The observed response of macroinvertebrate metrics to lentic-lotic conditions should be a key consideration for realistic ecological status assessment and could further be a valuable input for evaluating the effects of human-induced hydrological alteration and for assessing environmental flows

Inhibition of NAPDH Oxidase 2 (NOX2) Prevents Oxidative Stress and Mitochondrial Abnormalities Caused by Saturated Fat in Cardiomyocytes

Obesity and high saturated fat intake increase the risk of heart failure and arrhythmias. The molecular mechanisms are poorly understood. We hypothesized that physiologic levels of saturated fat could increase mitochondrial reactive oxygen species (ROS) in cardiomyocytes, leading to abnormalities of calcium homeostasis and mitochondrial function. We investigated the effect of saturated fat on mitochondrial function and calcium homeostasis in isolated ventricular myocytes. The saturated fatty acid palmitate causes a decrease in mitochondrial respiration in cardiomyocytes. Palmitate, but not the monounsaturated fatty acid oleate, causes an increase in both total cellular ROS and mitochondrial ROS. Palmitate depolarizes the mitochondrial inner membrane and causes mitochondrial calcium overload by increasing sarcoplasmic reticulum calcium leak. Inhibitors of PKC or NOX2 prevent mitochondrial dysfunction and the increase in ROS, demonstrating that PKC-NOX2 activation is also required for amplification of palmitate induced-ROS. Cardiomyocytes from mice with genetic deletion of NOX2 do not have palmitate-induced ROS or mitochondrial dysfunction. We conclude that palmitate induces mitochondrial ROS that is amplified by NOX2, causing greater mitochondrial ROS generation and partial depolarization of the mitochondrial inner membrane. The abnormal sarcoplasmic reticulum calcium leak caused by palmitate could promote arrhythmia and heart failure. NOX2 inhibition is a potential therapy for heart disease caused by diabetes or obesity

Coenzyme Q deficiency causes impairment of the sulfide oxidation pathway

Coenzyme Q (CoQ) is an electron acceptor for sulfide‐quinone reductase (SQR), the first enzyme of the hydrogen sulfide oxidation pathway. Here, we show that lack of CoQ in human skin fibroblasts causes impairment of hydrogen sulfide oxidation, proportional to the residual levels of CoQ. Biochemical and molecular abnormalities are rescued by CoQ supplementation in vitro and recapitulated by pharmacological inhibition of CoQ biosynthesis in skin fibroblasts and ADCK3 depletion in HeLa cells. Kidneys of Pdss2kd/kd mice, which only have ~15% residual CoQ concentrations and are clinically affected, showed (i) reduced protein levels of SQR and downstream enzymes, (ii) accumulation of hydrogen sulfides, and (iii) glutathione depletion. These abnormalities were not present in brain, which maintains ~30% residual CoQ and is clinically unaffected. In Pdss2kd/kd mice, we also observed low levels of plasma and urine thiosulfate and increased blood C4‐C6 acylcarnitines. We propose that impairment of the sulfide oxidation pathway induced by decreased levels of CoQ causes accumulation of sulfides and consequent inhibition of short‐chain acyl‐CoA dehydrogenase and glutathione depletion, which contributes to increased oxidative stress and kidney failure

The clinical heterogeneity of coenzyme Q10 deficiency results from genotypic differences in the Coq9 gene

Primary coenzyme Q10 (CoQ10) deficiency is due to mutations in genes involved in CoQ biosynthesis. The disease has been associated with five major phenotypes, but a genotype–phenotype correlation is unclear. Here, we compare two mouse models with a genetic modification in Coq9 gene (Coq9Q95X and Coq9R239X), and their responses to 2,4‐dihydroxybenzoic acid (2,4‐diHB). Coq9R239X mice manifest severe widespread CoQ deficiency associated with fatal encephalomyopathy and respond to 2,4‐diHB increasing CoQ levels. In contrast, Coq9Q95X mice exhibit mild CoQ deficiency manifesting with reduction in CI+III activity and mitochondrial respiration in skeletal muscle, and late‐onset mild mitochondrial myopathy, which does not respond to 2,4‐diHB. We show that these differences are due to the levels of COQ biosynthetic proteins, suggesting that the presence of a truncated version of COQ9 protein in Coq9R239X mice destabilizes the CoQ multiprotein complex. Our study points out the importance of the multiprotein complex for CoQ biosynthesis in mammals, which may provide new insights to understand the genotype–phenotype heterogeneity associated with human CoQ deficiency and may have a potential impact on the treatment of this mitochondrial disorder.This work was supported by grants from the Marie Curie International Reintegration Grant Programme (COQMITMEL-266691 to LCL) within the Seventh European Community Framework Programme, from Ministerio de Economía y Competitividad, Spain (SAF2009-08315 and SAF2013-47761-R to LCL), from the Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucía (P10-CTS-6133 to LCL), and from the ‘CEIBioTic’ (20F12/1 to LCL). MLS is a predoctral fellow from the Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucía. LCL is supported by the ‘Ramón y Cajal’ National Programme, Ministerio de Economía y Competitividad, Spain (RYC-2011-07643). MAT is supported by a predoctoral grant from the University of Granada. EJC is supported by the Research Program of the University of Granada. CMQ is supported by NICHD Grants 5K23 HDO65871-05 and P01 HD080642-01, and by a MDA grant. The proteomic analysis was performed in the CSIC/UAB Proteomics Facility of IIBB-CSIC that belongs to ProteoRed, PRB2-ISCIII, supported by Grant PT13/0001

ANO10 mutations cause ataxia and coenzyme Q₁₀ deficiency.

Inherited ataxias are heterogeneous disorders affecting both children and adults, with over 40 different causative genes, making molecular genetic diagnosis challenging. Although recent advances in next-generation sequencing have significantly improved mutation detection, few treatments exist for patients with inherited ataxia. In two patients with adult-onset cerebellar ataxia and coenzyme Q10 (CoQ10) deficiency in muscle, whole exome sequencing revealed mutations in ANO10, which encodes anoctamin 10, a member of a family of putative calcium-activated chloride channels, and the causative gene for autosomal recessive spinocerebellar ataxia-10 (SCAR10). Both patients presented with slowly progressive ataxia and dysarthria leading to severe disability in the sixth decade. Epilepsy and learning difficulties were also present in one patient, while retinal degeneration and cataract were present in the other. The detection of mutations in ANO10 in our patients indicate that ANO10 defects cause secondary low CoQ10 and SCAR10 patients may benefit from CoQ10 supplementation

Three-dimensional analysis of mitochondrial crista ultrastructure in a Leigh Syndrome patient by in situ cryo-electron tomography

Mitochondrial diseases produce profound neurological dysfunction via mutations affecting mitochondrial energy production, including the relatively common Leigh Syndrome (LS). We recently described an LS case caused by a pathogenic mutation in USMG5, encoding a small supernumerary subunit of mitochondrial ATP synthase. This protein is integral for ATP synthase dimerization, and patient fibroblasts revealed an almost total loss of ATP synthase dimers. Here, we utilize in situ cryo-electron tomography (cryo-ET) in a clinical case-control study of mitochondrial disease to directly study mitochondria within cultured fibroblasts from an LS patient and a healthy human control subject. Through tomographic analysis of patient and control mitochondria, we find that loss of ATP synthase dimerization caused by the pathogenic mutation causes profound disturbances of mitochondrial crista ultrastructure. Overall, this work supports the crucial role of ATP synthase in regulating crista architecture in the context of human disease

Inhibition of NADPH oxidase 2 (NOX2) prevents sepsis-induced cardiomyopathy by improving calcium handling and mitochondrial function

Cardiomyopathy frequently complicates sepsis and is associated with increased mortality. Increased cardiac oxidative stress and mitochondrial dysfunction have been observed during sepsis, but the mechanisms responsible for these abnormalities have not been determined. We hypothesized that NADPH oxidase 2 (NOX2) activation could be responsible for sepsis-induced oxidative stress and cardiomyopathy. Treatment of isolated adult mouse cardiomyocytes with low concentrations of the endotoxin lipopolysaccharide (LPS) increased total cellular reactive oxygen species (ROS) and mitochondrial superoxide. Elevated mitochondrial superoxide was accompanied by depolarization of the mitochondrial inner membrane potential, an indication of mitochondrial dysfunction, and mitochondrial calcium overload. NOX2 inhibition decreased LPS-induced superoxide and prevented mitochondrial dysfunction. Further, cardiomyocytes from mice with genetic ablation of NOX2 did not have LPS-induced superoxide or mitochondrial dysfunction. LPS decreased contractility and calcium transient amplitude in isolated cardiomyocytes, and these abnormalities were prevented by inhibition of NOX2. LPS decreased systolic function in mice, measured by echocardiography. NOX2 inhibition was cardioprotective in 2 mouse models of sepsis, preserving systolic function after LPS injection or cecal ligation and puncture (CLP). These data show that inhibition of NOX2 decreases oxidative stress, preserves intracellular calcium handling and mitochondrial function, and alleviates sepsis-induced systolic dysfunction in vivo. Thus, NOX2 is a potential target for pharmacotherapy of sepsis-induced cardiomyopathy