Recent evidences have linked abdominal obesity, insulin resistance, and dyslipidemia to male infertility. Since a defective energy metabolism may play an important role in the impairment of sperm quality, the aim of this study is to investigate the sperm energetic metabolism in rats fed with a high-fat diet, an animal model associated with metabolic syndrome development. Sexually mature male Sprague-Dawley rats were divided into two groups and fed for 4 weeks a standard diet (control group) or a diet enriched in 35% of fat (high fat group). Liver and adipose tissue weight, plasma glucose, insulin, and lipid concentrations were determined. Activities of enzymes involved in sperm energetic metabolism were evaluated by spectrophotometric assays. Sperm mitochondrial respiratory activity was evaluated with a polarographic assay of oxygen consumption. The administration of a high-fat diet caused a significant increase in body weight of rats and provoked hyperglycemia, hyperinsulinemia, and dyslipidemia. In these animals, we also observed a reduction in sperm concentration and motility. The investigation of sperm energetic metabolism in animals fed a high-fat diet revealed an impairment in the activity of pyruvate and lactate dehydrogenase, citrate synthase, and respiratory chain complexes. A parallel reduction in the cellular levels of adenosine triphosphate (ATP) and an increase in oxidative damage were also observed. A defective energy metabolism may play an important role in the impairment of sperm quality in the high-fat diet fed rats.

Krill oil (KO) is a dietary source of n-3 polyunsaturated fatty acids, mainly represented by eicosapentaenoic acid and docosahexaenoic acid bound to phospholipids. The supplementation of a high-fat diet with 2.5% KO efficiently prevented triglyceride and cholesterol accumulation in liver of treated rats. This effect was accompanied by a parallel reduction of the plasma levels of triglycerides and glucose and by the prevention of a plasma insulin increase. The investigation of the molecular mechanisms of KO action in high-fat fed animals revealed a strong decrease in the activities of the mitochondrial citrate carrier and of the cytosolic acetyl-CoA carboxylase and fatty acid synthetase, which are both involved in hepatic de novo lipogenesis. In these animals a significant increase in the activity of carnitine palmitoyl-transferase I and in the levels of carnitine was also observed, suggesting a concomitant stimulation of hepatic fatty acid oxidation. The KO supplemented animals also retained an efficient mitochondrial oxidative phosphorylation, most probably as a consequence of a KO induced arrest of the uncoupling effects of a high-fat diet. Lastly, the KO supplementation prevented an increase in body weight, as well as oxidative damage of lipids and proteins, which is often found in high-fat fed animals.

The mitochondrial tricarboxylate carrier supplies cytosol with the carbon units necessary for hepatic lipogenesis. The activities of cytosolic acetyl-CoA carboxylase and fatty acid synthetase are therefore strictly connected to the function of mitochondrial tricarboxylate carrier. Dietary polyunsaturated fatty acids (PUFA) are potent modulators of hepatic lipogenesis. In rats fed with a diet enriched with a low dose (2.5% by weight) of krill oil (KO), a novel source of dietary n-3 PUFA, a time-dependent decrease in the activities of the mitochondrial tricarboxylate carrier and of the lipogenic enzymes was found. The KO induced inhibition of hepatic lipogenesis was more pronounced than that found in fish oil (FO)-fed rats, at least at short feeding times. The decrease in the activity of the mitochondrial tricarboxylate carrier caused by KO was due to a reduced expression of the protein and this effect is obtained by both transcriptional and post-transcriptional mechanisms. Furthermore, dietary KO reduces hepatic triglycerides and cholesterol more efficiently than FO and KO also lowers plasma triglycerides more efficiently than FO at shorter feeding times, whereas KO and FO similarly decreased plasma cholesterol in the course of feeding treatment. We believe that the present investigation opens up new possibilities regarding the use of dietary KO as a preventive factor for dyslipidaemia.

Non-alcoholic fatty liver disease (NAFLD) is a common clinicopathological condition, encompassing a range of conditions caused by lipid deposition within liver cells. To date, no approved drugs are available for the treatment of NAFLD, despite the fact that it represents a serious and growing clinical problem in the Western world. Identification of the molecular mechanisms leading to NAFLD-related fat accumulation, mitochondrial dysfunction and oxidative balance impairment facilitates the development of specific interventions aimed at preventing the progression of hepatic steatosis. In this review, we focus our attention on the role of dysfunctions in mitochondrial bioenergetics in the pathogenesis of fatty liver. Major data from the literature about the mitochondrial targeting of some antioxidant molecules as a potential treatment for hepatic steatosis are described and critically analysed. There is ample evidence of the positive effects of several classes of antioxidants, such as polyphenols (i.e., resveratrol, quercetin, coumestrol, anthocyanins, epigallocatechin gallate and curcumin), carotenoids (i.e., lycopene, astaxanthin and fucoxanthin) and glucosinolates (i.e., glucoraphanin, sulforaphane, sinigrin and allyl-isothiocyanate), on the reversion of fatty liver. Although the mechanism of action is not yet fully elucidated, in some cases an indirect interaction with mitochondrial metabolism is expected. We believe that such knowledge will eventually translate into the development of novel therapeutic approaches for fatty liver.

After ejaculation, the mammalian male gamete must undergo the capacitation process, which is a prerequisite for egg fertilization. The bioenergetics of sperm capacitation is poorly understood although its fundamental role in sustaining the biochemical and molecular events occurring during gamete activation. Glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) are the two major metabolic pathways producing ATP which is the primary source of energy for spermatozoa. Since recent data suggest that spermatozoa have the ability to use different metabolic substrates, the main aim of this work is to present a broad overview of the current knowledge on the energy-producing metabolic pathways operating inside sperm mitochondria during capacitation in different mammalian species. Metabolism of glucose and of other energetic substrates, such as pyruvate, lactate and citrate, are critically analyzed. Such knowledge, besides its obvious importance for basic science, could eventually translate into the development of novel strategies for treatment of male infertility, for artificial reproduction and sperm selection methods.

Multiple sclerosis (MS) is a chronic inflammatory autoimmune demyelinating disease of the central nervous system. There are four clinical forms of MS, the most common of which is characterized by a relapsing remitting course (RRMS). The etiology of MS is unknown, but many studies suggested that genetic, environmental and infectious agents may contribute to the development of this disease. In experimental autoimmune encephalomyelitis (EAE), the animal model for MS, it has been shown that CD4+ T cells play a key role in MS pathogenesis. In fact, these cells are able to cross the blood-brain barrier and cause axonal damage with neuronal death. T cell activation critically depends on mitochondrial ATP synthesis and reactive oxygen species (ROS) production. Interestingly, lots of studies linked the oxidative damage arising from mitochondrial changes to neurodegenerative disorders, such as MS. Based on these evidences, this work focused on the metabolic reprogramming of CD4+ T cells in MS subjects, being this cell population directly implicated in pathogenesis of disease, paying attention to mitochondrial function and response to oxidative stress. Such aspects, once clarified, may open new opportunities for a therapeutic metabolic modulation of MS disorder.

Mitochondrial metabolite carriers are hydrophobic proteins which catalyze the flux of several charged or hydrophilic substrates across the inner membrane of mitochondria. These proteins, like most mitochondrial proteins, are nuclear encoded and after their synthesis in the cytosol are transported into the inner mitochondrial membrane. Most metabolite carriers, differently from other nuclear encoded mitochondrial proteins, are synthesized without a cleavable presequence and contain several, poorly characterized, internal targeting signals. However, an interesting aspect is the presence of a positively charged N-terminal presequence in a limited number of mitochondrial metabolite carriers. Over the last few years the molecular mechanisms of import of metabolite carrier proteins into mitochondria have been thoroughly investigated. This review summarizes the present knowledge and discusses recent advances on the import and sorting of mitochondrial metabolite carriers.

Biological invasions are deemed as one of the most severe threats to biodiversity and can result in huge economic impacts on natural resources. Many studies have assessed structural changes caused by the introduction of nonindigenous species, but urgent questions remain to be answered about the complex indirect effects of invasive pests on marine biodiversity at the species and ecosystem level. Previous investigations focusing on the biological invasion by Caulerpa racemosa in the Mediterranean indicated that the lipophilic algal metabolite caulerpin accumulates in the tissues of the native edible white sea bream Diplodus sargus feeding on the exotic alga. The level of caulerpin in D. sargus has been thus used as an indicator of the trophic exposure of the fish to the seaweed and related with adverse effects on the fish health. In the present study, we show that the C. racemosa-based diet can also alter muscle fatty acids composition in the white sea bream by reducing the percentage of polyunsaturated fatty acids of the n-3 and n-6 series, such as eicosapentaenoic (C20:5), docosahexaenoic (C22:6) and arachidonic acids (C20:4). This represents both a real threat to the health of fish unable to biosynthesize these essential fatty acids and an impoverishment of an important source of essential fatty acids for human nutrition, which helps to prevent cardiovascular diseases, diabetes, hepatic steatosis, cancer and neurological disorders. On the whole, this work sheds light on an unexplored and critical aspect of biological invasions with implication on the health of both humans and the environment.

L’assunzione di una dieta iperlipidica, caratterizzata da un elevato contenuto in acidi grassi saturi e monoinsaturi e priva di acidi grassi polinsaturi della serie ω-3 ha causato, in modelli animali, obesità, insulino-resistenza, dislipidemie e, parallelamente, una riduzione della qualità spermatica in termini di decremento della concentrazione e della motilità dei gameti maschili. Un’approfondita analisi dei meccanismi molecolari che sono alla base di tale effetto ha evidenziato la presenza di difetti nel metabolismo energetico, riguardanti sia la glicolisi che i processi mitocondriali responsabili della produzione di ATP. Pertanto, l’impiego di nuove strategie terapeutiche, quali la restrizione calorica o l’integrazione con acidi grassi polinsaturi, che regolano la funzione e la biogenesi mitocondriale, potrebbe avere un enorme potenziale preventivo e/o terapeutico anche nel trattamento dell’infertilità maschile.

Le alterazioni del metabolismo lipidico, ampiamente diffuse tra gli individui dei paesi sviluppati, rappresentano la maggiore causa scatenante le patologie cardiovascolari. Un ruolo molto importante nella prevenzione di queste patologie è svolto dall’alimentazione. Differenti trattamenti nutrizionali inducono risposte metaboliche differenti nell’organismo e, in particolare, nel fegato che ne riveste un ruolo centrale. Saggi enzimatici hanno dimostrato l’elevata flessibilità del fegato nella modulazione della biosintesi degli acidi grassi in risposta ai diversi tipi di dieta. Una dieta arricchita in zuccheri semplici stimola moderatamente la lipogenesi epatica, inducendo un lieve e transiente aumento di trigliceridi nel fegato e un significativo e duraturo aumento di trigliceridi nel plasma, suggerendo come l’eccesso di saccarosio sia facilmente convertito in grassi, che vengono distribuiti ai tessuti attraverso il circolo ematico, causando dislipidemie. Al contrario, una dieta arricchita in acidi grassi saturi causa una forte inibizione della lipogenesi, limitatamente alle prime settimane di trattamento, effetto reversibile nel tempo. Questa condizione si associa ad un iniziale aumento dei livelli di trigliceridi e di colesterolo plasmatici che vanno progressivamente a ripristinarsi raggiungendo valori normali, in maniera speculare rispetto a quanto si osserva nel fegato che nel corso delle settimane di trattamento diviene un vero e proprio deposito di lipidi, favorendo così l’insorgenza e l’instaurarsi della condizione di steatosi.

The citrate carrier (CIC) is an integral protein of the inner mitochondrial membrane that has a fundamental role in hepatic intermediary metabolism. Its primary function is to catalyze the transport of citrate from mitochondria, where this molecule is formed, to cytosol, where this molecule is used for fatty acid and cholesterol synthesis. Therefore, mitochondrial CIC acts upstream of cytosolic lipogenic reactions and its regulation is particularly important in view of the modulation of hepatic lipogenesis. Whereas a great deal of data are currently available on the dietary modulation of cytosolic lipogenic enzymes, little is known about the nutritional regulation of the CIC transport activity. In this review we describe the differential effects of distinct fatty acids present in the diet on the activity of the mitochondrial CIC. In particular, polyunsaturated fatty acids were powerful modulators of the activity of mitochondrial CIC by influencing its expression through transcriptional and post-transcriptional mechanisms. On the contrary, saturated and monounsaturated fatty acids did not influence at all the mitochondrial CIC activity. Moreover, variations in the CIC activity were connected to similar alterations in the metabolic pathways to which the transported citrate is channelled. Therefore, CIC may be considered as a sensor for changes occurring inside the hepatocyte and may represent an important target for the regulation of hepatic lipogenesis. The crucial role of this protein is reinforced by the recent discovery of its involvement in other cellular processes, such as glucose-stimulated insulin secretion, inflammation, tumorigenesis, genome stability and sperm metabolism.

Recently, obesity has been linked to male infertility. In animal models the administration of a high-fat diet caused a reduction in sperm quality, by impairing gamete energy metabolism. The aim of this study was to investigate a possible effect of dietary fatty acids supplementation in the modulation of sperm energy metabolism and, in turn, in the improvement of sperm quality in rats fed a high-fat diet. Sexually mature male Sprague–Dawley rats were divided into four groups and fed for 4 weeks a standard diet (con- trol group), a high-fat diet (enriched in 35% of fat and 15% sucrose), a high-fat diet supplemented with 2.5% olive oil (a source of monounsaturated fatty acids) or a high-fat diet supplemented with 2.5% krill oil (a source of n-3 polyunsaturated fatty acids). Liver and adipose tissue weight, plasma glucose, insulin and lipid concentrations were determined. Activities of enzymes involved in sperm energetic metabolism were evaluated by spectrophotometric assays. Sperm mitochondrial respiratory efficiency was also assayed. The obtained results suggest that olive oil partially counteracts the negative effects of a high-fat diet on sperm quality, by increasing gamete motility, by reducing oxidative stress and slightly improving mitochondrial respiration efficiency. On the other hand, krill oil determines an increase in sperm concentration and motility, an increase in the activities of lactate dehydrogenase, Krebs cycle enzymes and respiratory chain complexes; a parallel increase in the cellular levels of ATP and a reduction in oxidative damage were also observed. These results suggest that dietary fatty acids are able to positively influence sperm quality and function.

L’assunzione di una dieta iperlipidica, caratterizzata da un elevato contenuto in acidi grassi saturi e monoinsaturi e priva di acidi grassi polinsaturi della serie ω-3, causa obesità e steatosi epatica. Un’approfondita analisi dei meccanismi molecolari che sono alla base di tale effetto ha evidenziato, oltre all’inibizione dell’ossidazione degli acidi grassi epatici, l’esistenza di una forte compromissione del metabolismo energetico mitocondriale. Pertanto, l’impiego di nuove strategie terapeutiche, quali la restrizione calorica o l’integrazione con acidi grassi polinsaturi della serie ω-3, che regolano la funzione e la biogenesi mitocondriale, possono avere un enorme potenziale terapeutico nel trattamento dell’obesità e dell’accumulo di lipidi nel fegato.

PURPOSE: Hepatic fatty acid synthesis is influenced by several nutritional and hormonal factors. In this study, we have investigated the effects of distinct experimental diets enriched in carbohydrate or in fat on hepatic lipogenesis. METHODS: Male Wistar rats were divided into four groups and fed distinct experimental diets enriched in carbohydrates (70% w/w) or in fat (20% and 35% w/w). Activity and expression of the mitochondrial citrate carrier and of the cytosolic enzymes acetyl-CoA carboxylase and fatty acid synthetase were analyzed through the study with assessments at 0, 1, 2, 4 and 6 weeks. Liver lipids and plasma levels of lipids, glucose and insulin were assayed in parallel. RESULTS: Whereas the high carbohydrate diet moderately stimulated hepatic lipogenesis, a strong inhibition of this anabolic pathway was found in animals fed high fat diets. This inhibition was time and concentration-dependent. Moreover, whereas the high carbohydrate diet induced an increase in plasma triglycerides, the high fat diets determined an accumulation of triglycerides in liver. An increase in the plasmatic levels of glucose and insulin was observed in all cases. CONCLUSIONS: The excess of sucrose in the diet is converted into fat, which is distributed by bloodstream in the organism in the form of circulating triglycerides. On the other hand, a high amount of dietary fat caused a strong inhibition of lipogenesis and a concomitant increase in the level of hepatic lipids, thereby highlighting, in these conditions, the role of liver as a reservoir of exogenous fat.

Il varicocele è una patologia andrologica caratterizzata da ectasia del plesso venoso pampiniforme, complesso di vene testicolari che drena il sangue refluo dai testicoli. E’ causato, in primo luogo, da ipertermia scrotale e dal reflusso di prodotti tossici per la struttura testicolare, come catecolamine e ormoni steroidei provenienti dai surreni. Il varicocele risulta essere una delle principali cause di infertilità maschile: circa il 20% dei soggetti affetti da tale patologia, infatti, manifesta infertilità. Le lesioni a carico del testicolo, causate dal varicocele, si riflettono in alterazioni della qualità spermatica e possono indurre diminuzione del numero degli spermatozoi, riduzione nella loro motilità e/o alterazioni nella loro morfologia. Evidenze sperimentali suggeriscono che lo stress ossidativo è una condizione fondamentale ai fini della compromissione della fertilità nei soggetti con varicocele. D’altro canto, sono state messe in luce interessanti correlazioni tra elevati livelli di specie reattive dell’ossigeno (ROS), danni indotti da stress ossidativo come perossidazione lipidica e frammentazione del DNA spermatico, e riduzione della qualità dei gameti maschili. Recentemente, è stato anche proposto che lo stress ossidativo è in grado di compromettere la funzionalità mitocondriale determinando un “disaccoppiamento” tra il trasporto degli elettroni attraverso la catena respiratoria e la sintesi di ATP. Pertanto, il ruolo che difetti dell’efficienza respiratoria mitocondriale esercitano nella riduzione della qualità spermatica dei soggetti affetti da varicocele merita di essere approfondito.

The role of mitochondria in sperm motility was the subject of several investigations. However, different views on this topic emerged among scientists. In particular, very little is known on the mechanisms of energy production occurring during human sperm capacitation and related processes. In this study, we have investigated the mitochondrial respiratory efficiency in human sperm samples from normozoospermic subjects before and after swim-up selection and incubation under capacitating condition. Sperm cells, selected by swim-up treatment, were incubated up to 24 h and then demembranated by hypotonic swelling at selected times. The oxygen uptake rate was measured in both basal and swim-up selected samples by a polarographic assay. Mitochondria of swim-up selected cells showed an impressive oxygen consumption rate, which was about 20 times higher than that measured in basal samples. The high mitochondrial respiratory efficiency remained stable up to 24 h after the swimup treatment. The respiration control ratio, the substrate specificity and the inhibitor sensitivity in the swim-up selected samples were similar to those of basal samples thereby suggesting that the physiology of mitochondria was preserved after the swim-up treatment. Furthermore, the remarkably high mitochondrial respiration in swim-up selected samples allowed the oxygraphic analysis of just 200 000 sperm cells. Sperm selection and incubation under capacitating condition are therefore associated with a high activity of the mitochondrial respiratory chain. The sperm oxygen consumption rate could be useful to exclude mitochondria malfunctioning in male infertility.

In this study the authors evaluated the prevalence of Gestational Diabetes Mellitus (GDM) and the incidence in determining the occurrence thereof, given by the following risk factors: age, family history for Diabetes Mellitus (DM), Body Mass Index (BMI), smoking, diet, macrosomia in previous pregnancies and physical activity. To this end, they investigated a cohort of 61 pregnant women, who had at least one risk factor, in a total period of nine months, during which women checked in the Clinic of Endocrinology and Diabetology of the Hospital “F. Ferrari” in Casarano (Lecce) Italy, and were subjected to an anamnestic investigation and then to a diagnostic test (Oral Glucose Tolerance Test, OGTT 75 g). The anamnesis has allowed to identify the type of risk factors that are present in the population, while the OGTT has allowed to diagnose gestational diabetes. Later, an analysis of the cross-collected data was performed in order to evaluate the percentage incidence of the disease, in relation to risk factors observed. The authors’ results confirmed a prevalence of GDM of 26.23% and the existence of a close association between risk factors and GDM. In particular, the risk factors, macrosomia, BMI ≥ 25 kg/m2, a hypercaloric diet, light physical activity, and age ≥ 35 years, seem to have more influence than smoking and family history for DM, inducing the onset of the illness.

In recent years, several studies focused their attention on the role of dietary fats in the pathogenesis of hepatic steatosis. It has been demonstrated that a high-fat diet is able to induce hyperglycemia, hyperinsulinemia, obesity and non-alcoholic fatty liver disease. On the other hand krill oil, a novel dietary supplement of n-3 PUFAs, has the ability to improve lipid and glucose metabolism, exerting possible protective effects against hepatic steatosis. In this study we have investigated the effects of krill oil on mitochondrial energetic metabolism in animals fed a high-fat diet. To this end, male Sprague-Dawley rats were divided into three groups and fed for 4 weeks with a standard diet (control group), a diet with 35% fat (HF group) or a high-fat diet supplemented with 2.5% krill oil (HF+KO group). The obtained results suggest that krill oil promotes the burning of fat excess introduced by the high-fat diet. This effect is obtained by stimulating mitochondrial metabolic pathways such as fatty acid oxidation, Krebs cycle and respiratory chain complexes activity. Modulation of the expression of carrier proteins involved in mitochondrial uncoupling was also observed. Overall, krill oil counteracts the negative effects of a high-fat diet on mitochondrial energetic metabolism.

Risulta quanto mai evidente la necessità di ulteriori studi che permettano di rivelare gli aspetti ancora poco chiari dell’assembly del complesso III, nonché evidenziare ulteriori ruoli delle proteine chaperone coinvolte durante il suo processo di biogenesi. La specificità dei tessuti maggiormente colpiti e il sovraccarico di ferro, tipici delle malattie associate a BCS1L, sollevano infatti numerosi interrogativi circa il ruolo di questa proteina, suggerendo possibili funzioni addizionali nei mitocondri, oltre a quella di fattore di assemblaggio del complesso III. Inoltre, nella maggior parte degli individui con carenze del complesso del citocromo bc1 deve essere ancora identificato il difetto molecolare, permettendo di ipotizzare l’esistenza di addizionali fattori di assemblaggio ancora da scoprire3. Allo stato attuale non esistono cure risolutive per il trattamento delle malattie mitocondriali, ma solo terapie che permettono di migliorare i sintomi, in particolare l’intolleranza allo sforzo e l’affaticamento. La loro efficacia varia, comunque, da paziente a paziente e dipende dal deficit enzimatico, nonché dalla gravità dei sintomi. Una comprensione più profonda dei meccanismi molecolari alla base dei disordini mitocondriali potrebbe quindi porre le basi per lo sviluppo di adeguati trattamenti terapeutici.

Glatiramer acetate (GA; Copaxone) is a random copolymer of glutamic acid, lysine, alanine, and tyrosine used for the treatment of patients with multiple sclerosis (MS). Its mechanism of action has not been already fully elucidated, but it seems that GA has an immune-modulatory effect and neuro-protective properties. Lymphocyte mitochondrial dysfunction underlines the onset of several autoimmune disorders. In MS first diagnosis patients, CD4+, the main T cell subset involved in the pathogenesis of MS, undergo a metabolic reprogramming that consist in the up-regulation of glycolysis and in the down-regulation of oxidative phosphorylation. Currently, no works exist about CD4+ T cell metabolism in response to GA treatment. In order to provide novel insight into the potential use of GA in MS treatment, blood samples were collected from 20 healthy controls (HCs) and from 20 RR MS patients prior and every six months during the 12 months of GA administration. GA treated patients' CD4+ T cells were compared with those from HCs analysing their mitochondrial activity through polarographic and enzymatic methods in association with their antioxidant status, through the analysis of SOD, GPx and CAT activities. Altogether, our findings suggest that GA is able to reduce CD4+ T lymphocytes' dysfunctions by increasing mitochondrial activity and their response to oxidative stress.

Lo studio del metabolismo energetico degli spermatozoi, nell’ambito del quale i mitocondri rivestono un ruolo fondamentale, rappresenta un nuovo ed intrigante approccio per la comprensione dei meccanismi biochimici alla base della fertilità maschile e, al tempo stesso, per il trattamento di patologie andrologiche. Due aspetti in particolare, la compartimentalizzazione e la versatilità metabolica, fanno, dei gameti maschili, delle cellule con la peculiare capacità di modulare il proprio metabolismo e attivare diverse vie metaboliche, a seconda della disponibilità dei substrati e delle condizioni chimico-fisiche presenti nell’ambiente in cui si trovano. Tale versatilità è fondamentale per garantire il successo della fertilizzazione. La glicolisi e la fosforilazione ossidativa (OXPHOS) rappresentano i due principali processi metabolici dai quali viene prodotta ATP, che è la fonte energetica maggiormente utilizzata dagli spermatozoi, al fine di supportare e garantire diverse funzioni cellulari. Oltre al metabolismo del glucosio, anche quello di altri substrati energetici, come piruvato, lattato, acidi grassi e citrato, fornisce intermedi e prodotti metabolici necessari per sostenere la fisiologia di uno spermatozoo. La comprensione dei processi biochimici rappresenta senza dubbio una nuova sfida per caratterizzare aspetti fisiologici e patologici dei gameti maschili.

The red pigment caulerpin, a secondary metabolite from the marine invasive green algae Caulerpa cylindracea can be accumulated and transferred along the trophic chain, with detrimental consequences on biodiversity and ecosystem functioning. Despite increasing research efforts to understand how caulerpin modifies fish physiology, little is known on the effects of algal metabolites on mammalian cells. Here we report for the first time the mitochondrial targeting activity of both caulerpin, and its closely related derivative caulerpinic acid, by using as experimental model rat liver mitochondria, a system in which bioenergetics mechanisms are not altered. Mitochondrial function was tested by polarographic and spectrophotometric methods. Both compounds were found to selectively inhibit respiratory complex II activity, while complexes I, III, and IV remained functional. These results led us to hypothesize that both algal metabolites could be used as antitumor agents in cell lines with defects in mitochondrial complex I. Ovarian cancer cisplatin-resistant cells are a good example of cell lines with a defective complex I function on which these molecules seem to have a toxic effect on proliferation. This provided novel insight toward the potential use of metabolites from invasive Caulerpa species for the treatment of human ovarian carcinoma cisplatin-resistant cells.

Oxygen consumption rate (OCR) is a significant parameter helpful to determine in vitro respiratory efficiency of living cells. Oxygen is an excellent oxidant and its electrocatalytic reduction on a noble metal allows accurately detecting it. By means of microfabrication technologies, handy, low-cost, and disposable chip can be attained, minimizing working volumes and improving sensitivity and response time. In this respect, here is presented a microoxygraph device (MOD), based on Clark’s electrode principle, displaying many advantageous features in comparison to other systems. This lab-on-chip platform is composed of a three-microelectrode detector equipped with a microgrooved electrochemical cell, sealed with a polymeric reaction chamber. Au working/counter electrodes and Ag/AgCl reference electrode were fabricated on a glass slide. A microchannel was realized by photoresist lift-off technique and a polydimethylsiloxane (PDMS) nanoporous film was integrated as oxygen permeable membrane (OPM) between the probe and the microreaction chamber. Electrochemical measurements showed good reproducibility and average response time, assessed by periodic injection and suction of a reducing agent. OCR measurements on 3T3 cells, subjected, in real time, to chemical stress on the respiratory chain, were able to show that this chip allows performing consistent metabolic analysis.

Mitochondria of spermatozoa significantly differ, structurally and functionally, from the corresponding orga- nelles of somatic cells. Nevertheless, sperm mitochondria, as well as the somatic ones, are the location of the ox- idative phosphorylation (OXPHOS) process, which is necessary for the production of metabolic energy in the form of adenosine triphosphate (ATP). In addition to their basic role in oxidative energy generation, mitochon- dria are also a source of reactive oxygen species (ROS), which, at low concentrations, play a physiological role in many sperm processes. It is commonly accepted that a proper functionality of mitochondria is necessary for a high quality of sperm cells; this last parameter, in its turn, is a pre-requisite for a high sperm fertilizing ability. Evaluation of mitochondrial respiratory efficiency represents therefore a valuable test that could integrate routine semen analysis.

I mitocondri presenti negli spermatozoi rivestono un ruolo critico nel determinare la qualità del liquido seminale e la funzionalità degli stessi gameti. Pertanto, l’analisi della funzionalità mitocondriale è un’indagine a cui ricorrere nella valutazione della qualità di una popolazione spermatica e, quindi, nella diagnosi dell’infertilità maschile. Un metodo particolarmente promettente per determinare la funzionalità mitocondriale degli spermatozoi si basa sulla determinazione del consumo di ossigeno in campioni di spermatozoi permeabilizzati in mezzo ipotonico e sul calcolo di un parametro, l’indice di controllo respiratorio, o ICR, in grado di dare indicazioni sull’efficienza respiratoria degli organelli. Tale analisi, ora solo emergente, potrebbe in realtà diventare un’indagine di routine, da eseguire nei centri di procreazione medicalmente assistita e nei laboratori di fisiopatologia seminale, per valutare la capacità degli spermatozoi a fecondare la cellula uovo.

OBJECTIVE To correlate sperm mitochondrial respiratory efficiency with variations in sperm motility and with sperm morphologic anomalies. METHODS Sperm mitochondrial respiratory activity was evaluated with a polarographic assay of oxygen consumption carried out in hypotonically-treated sperm cells. A possible relationship among sperm mitochondrial respiratory efficiency, sperm motility, and morphologic anomalies was investigated. RESULTS Mitochondrial respiratory efficiency was positively correlated with sperm motility and negatively correlated with the percentage of immotile spermatozoa. Moreover, midpiece defects impaired mitochondrial functionality. CONCLUSION The results reported in this study support a strong and positive correlation between mitochondrial respiratory efficiency and sperm progressive motility. In addition and in strict agreement with these findings, morphologic alterations of the sperm midpiece, where mitochondria are found, are associated with lower mitochondrial functionality and lower sperm motility. These results confirm the important role played by mitochondria in supplying metabolic energy for sperm motility and provide more insights into the biochemical basis of mitochondria-based asthenozoospermia. On the other hand, the findings reported here open up new and interesting avenues of investigation in the case of semen samples from asthenozoospermic subjects, for which a more detailed investigation can be carried out at the level of single componente of the mitochondrial respiratory chain.

Non-alcoholic fatty liver disease (NAFLD) describes a range of conditions caused by fat deposition within liver cells. Liver fat content reflects the equilibrium between several metabolic pathways involved in triglyceride synthesis and disposal, such as lipolysis in adipose tissue and de novo lipogenesis, triglyceride esterification, fatty acid oxidation and very-low-density lipoprotein synthesis/secretion in hepatic tissue. In particular, it has been demonstrated that the hepatic de novo lipogenesis plays a significant role in NAFLD pathogenesis. It is widely known that the fatty acid composition of the diet influences hepatic lipogenesis along with other metabolic pathways. Therefore, dietary fat may not only be involved in the pathogenesis of hepatic steatosis, but may also prevent and/or reverse hepatic fat accumulation. In this review major literature data about the role of some dietary fats as a potential cause of hepatic fat accumulation or as a potential treatment for NAFLD are described. Moreover, biochemical mechanisms responsible for the increase or the decrease in the hepatic lipid content are critically analyzed. It is noteworthy that both quantitative and qualitative aspects of dietary fat influence triglyceride deposition in liver. A high-fat diet or the dietary administration of conjugated linoleic acids induced hepatic steatosis. On the contrary, supplementation of the diet with krill oil or pine nut oil helped in the prevention and/or in the treatment of steatotic liver. Quite interesting is the “case” of olive oil, since several studies have often provided different and⁄∕or conflicting results in animal models.

Obesity in men of reproductive age is increasing worldwide, impacting negatively on reproductive potential, sperm function, and assisted reproduction outcomes. Changes in modern eating behaviors are needed to invert the negative correlation between lifestyle and sperm quality. It is known from the literature that fatty acids are able to modulate lipid metabolism by complex mechanisms involving a sequence of interdependent and cross-regulated molecular events. Although a great deal of data is currently available on the dietary modulation of lipid metabolism, little is known about the nutritional regulation of sperm energetic metabolism. In this review, major data from the literature about the role of some dietary fats as modulators of sperm function and quality are described. Moreover, biochemical mechanisms responsible for an impairment or improvement in sperm bioenergetics are critically analyzed. It is noteworthy that both quantitative and qualitative aspects of dietary fats influence sperm function. The administration of a high-fat diet, low in polyunsaturated fatty acids, caused a significant increase in body weight hyperglycemia, hyperinsulinemia, and dyslipidemia, along with a significant reduction in sperm quality. In contrast, unsaturated fatty acids are able to positively influence sperm quality and function. The understanding of the biochemical mechanisms underlying the development of male infertility related to obesity led us to depict a possible framework for the molecular action of fatty acids on sperm quality during dysmetabolic conditions.

Le alterazioni del metabolismo lipidico, ampiamente diffuse tra gli individui dei paesi sviluppati, rappresentano la maggiore causa scatenante le patologie cardiovascolari. Un ruolo molto importante nella prevenzione di queste patologie è svolto dagli acidi grassi EPA e DHA, contenuti negli oli di pesce, dal momento che questi acidi grassi sono in grado di svolgere diversi effetti benefici sull’organismo: riducono i livelli di trigliceridi, prevengono le aritmie, riducono l’infiammazione e abbassano la pressione sanguigna, svolgendo un’azione preventiva nei confronti delle patologie cardiovascolari. Recentemente, è stata introdotta sul mercato una nuova fonte di EPA e DHA: si tratta di un olio estratto da invertebrati marini, appartenenti alla specie Euphasia superba e denominato “olio di krill”. Numerosi sono gli integratori e le pillole a base di questo olio immessi di recente nel mercato internazionale e pubblicizzati in virtù delle proprietà benefiche possedute da EPA e DHA e del potenziale antiossidante delle molecole in esso contenute. Grazie alle sue potenzialità ipolipidemizzanti, maggiori rispetto agli oli di pesce, l’olio di krill potrebbe essere utilizzato nella cura e/o prevenzione di dislipidemie e obesità, come nuova e preziosa fonte di acidi grassi della serie ω-3.

OBJECTIVE: To correlate the level of oxidative stress in serum and seminal fluid and the level of sperm DNA fragmentation with sperm mitochondrial respiratory efficiency. METHODS: Sperm mitochondrial respiratory activity was evaluated with a polarographic assay of oxygen consumption carried out in hypotonically-treated sperm cells. A possible relationship between sperm mitochondrial respiratory efficiency, the level of oxidative stress and the level of sperm DNA fragmentation was investigated. RESULTS: Sperm motility was positively correlated with mitochondrial respiration but was negatively correlated with oxidative stress and DNA fragmentation. Interestingly, sperm mitochondrial respiratory activity was negatively affected by oxidative stress and DNA fragmentation. CONCLUSIONS: Our data indicate that sperm mitochondria respiration is decreased in subjects with high levels of ROS by an uncoupling between electron transport and ATP synthesis. This reduction in mitochondrial functionality may be one of the reasons responsible for the decrease in spermatozoa motility.

La Sclerosi Multipla (SM) è una malattia infiammatoria cronica demielinizzante, a carattere autoimmune, che colpisce il Sistema Nervoso Centrale (SNC). Nonostante siano stati effettuati una serie di studi allo scopo di chiarire le cause della patologia, alcuni meccanismi patogenetici restano ancora oscuri e l’eziologia rimane sconosciuta. Il modello murino della malattia, l'encefalopatia autoimmune, ha tuttavia permesso di ricondurre il processo di demielinizzazione all'azione dei linfociti T CD4+ che, una volta attivati, sarebbero in grado di penetrare attraverso la barriera ematoencefalica e di causare danno assonale con conseguente morte di cellule neuronali. L’attivazione delle cellule T è accompagnata da uno shift metabolico che prevede il passaggio da un metabolismo ossidativo energeticamente efficiente ad un programma prettamente glicolitico; nelle malattie autoimmuni i dati in letteratura concernenti il fenotipo metabolico dei linfociti T sono piuttosto esigui e, tuttora, non vi sono lavori condotti a tal riguardo nella SM.

Sempre più numerose sono le evidenze sperimentali che suggeriscono come la sindrome metabolica possa rappresentare un fattore di rischio per infertilità maschile. I meccanismi patogenetici alla base dell’interconnessione tra le due patologie potrebbero trovare le loro cause nella lipomatosi scrotale, associata ad una condizione di obesità, in un aumento della produzione di ROS, come conseguenza delle alterazioni del metabolismo lipidico o in uno squilibrio dell’asse neuro-endocrino, proprio dei soggetti diabetici. Le conseguenze di questi fenomeni si riflettono in una riduzione della qualità del liquido seminale: decremento della concentrazione e della motilità e alterazioni della morfologia degli spermatozoi. Tuttavia, questi sono prevalentemente dei segni clinici di subfertilità, piuttosto che una vera diagnosi. Studi futuri saranno, quindi, necessari alla comprensione dei meccanismi molecolari che sono alla base delle alterazioni della funzionalità spermatica indotte da sindrome metabolica. Un’analisi approfondita di questo tipo potrebbe risultare utile alla definizione di misure preventive mediante le quali limitare il rischio di insorgenza di infertilità in presenza di disfunzioni metaboliche.

Sperm cells progressive motility is the most important parameter involved in the fertilization process. Sperm middle piece contains mitochondria, which play a critical role in energy production and whose proper operation ensures the reproductive success. Notably, sperm progressive motility is strictly related to mitochondrial membrane potential (MMP) and consequently to mitochondrial functionality. Although previous studies presented an evaluation of mitochondrial function through MMP assessment in entire sperm cells samples, a quantitative approach at single-cell level could provide more insights in the analysis of semen quality. Here we combine laser scanning confocal microscopy and functional fluorescent staining of mitochondrial membrane to assess MMP distribution among isolated spermatozoa. We found that the sperm fluorescence value increases as a function of growing progressive motility and that such fluorescence is influenced by MMP disruptors, potentially allowing for the discrimination of different quality classes of sperm cells in heterogeneous populations.

La principale caratteristica della patogenesi della NAFLD è l’accumulo di trigliceridi nel fegato, i cui livelli riflettono un equilibrio tra i vari processi metabolici coinvolti nella loro sintesi e smaltimento. In particolare, l’aumento dell’uptake di acidi grassi liberi, in seguito allo stimolo della lipolisi nel tessuto adiposo, e l’induzione della lipogenesi nel fegato contribuiscono primariamente all’insorgenza del fegato steatotico. Recentemente, numerosi studi condotti su modelli animali hanno messo in evidenza il ruolo svolto da alcuni grassi alimentari nella modulazione della lipogenesi epatica, in considerazione dell’importanza che questa via metabolica riveste nella patogenesi della NAFLD. Mentre SFA e MUFA non hanno alcuna significativa influenza sulla via biosintetica degli acidi grassi, l’assunzione di PUFA della serie ω-3 e della serie ω-6, riducendo fortemente l’attività e l’espressione di proteine coinvolte nella lipogenesi, ostacola l’insorgenza della patologia. Pertanto, è possibile modulare il metabolismo lipidico e, in particolare la lipogenesi epatica, intervenendo sull’alimentazione e ricorrendo a particolari combinazioni di acidi grassi. Ciò evidenzia come i grassi alimentari possano essere utilizzati nella prevenzione e nel trattamento di patologie, spesso associate ad abitudini e stili di vita errati, di cui la steatosi epatica ne è un esempio. Alla luce di ciò, i risultati promettenti ottenuti dalla somministrazione di nuovi supplementi dietetici in modelli animali, come l’olio di krill e l’olio di pino coreano, suggeriscono che sarebbe estremamente interessante approfondire gli studi sull’uomo. Ciò potrebbe avere importanti risvolti nella prevenzione e/o nel trattamento della NAFLD, anche in virtù del fatto che, attualmente, non esistono ancora trattamenti farmacologici accertati tramite i quali intervenire su questa patologia.

In Saccharomyces cerevisiae the mature cytochrome bc1 complex exists as an obligate homo-dimer in which each monomer consists of ten distinct protein subunits inserted into or bound to the inner mitochondrial membrane. Among them, the Rieske iron-sulphur protein (Rip1), besides its catalytic role in electron transfer, may be implicated in the bc1 complex dimerization. Indeed, Rip1 has the globular domain containing the catalytic centre in one monomer while the transmembrane helix interacts with the adjacent monomer. In addition, the lack of Rip1 leads to the accumulation of an immature bc1 intermediate, only loosely associated with cytochrome c oxidase. In this study we have investigated the biogenesis of the yeast cytochrome bc1 complex using epitope tagged proteins to purify native assembly intermediates. We showed that the dimerization process is an early event during bc1 complex biogenesis and that the presence of Rip1, differently from previous proposals, is not essential for this process. We also investigated the multi-step model of bc1 assembly thereby lending further support to the existence of bona fide subcomplexes during bc1 maturation in the inner mitochondrial membrane. Finally, a new model of cytochrome bc1 complex assembly, in which distinct intermediates sequentially interact during bc1 maturation, has been proposed.

Mitochondria of spermatozoa are different from the corresponding organelles of somatic cells, in both their morphology and biochemistry. The biochemical differences are essentially related to the existence of specific enzyme isoforms, which are characterized by peculiar kinetic and regulatory properties. As mitochondrial energy metabolism is a key factor supporting several sperm functions, these organelles host critical metabolic pathways during germ cell development and fertilization. Furthermore, spermatozoa can use different substrates, and therefore activate different metabolic pathways, depending on the available substrates and the physico-chemical conditions in which they operate. This versatility is critical to ensure fertilization success. However, the most valuable aspect of mitochondria function in all types of cells is the production of chemical energy in the form of ATP which can be used, in the case of spermatozoa, for sustaining sperm motility. The latter, on the other hand, represents one of the major determinants of male fertility. Accordingly, the presence of structural and functional alterations in mitochondria from asthenozoospermic subjects confirms the important role played by these organelles in energy maintenance of sperm motility. The present study gives an overview of the current knowledge on the energy-producing metabolic pathways operating inside human sperm mitochondria and critically analyse the differences with respect to somatic mitochondria. Such a comparison has also been carried out between the functional characteristics of human sperm mitochondria and those of other mammalian species. A deeper understanding of mitochondrial energy metabolism could open up new avenues of investigation in bioenergetics of human sperm mitochondria, both in physiological and pathological conditions.

OBJECTIVE: To evaluate the effect of varicocele on oxidative stress, sperm mitochondrial respiratory efficiency, sperm morphology and semen parameters. METHODS: 20 patients with varicocele and 20 normozoospermic subjects without varicocele (control group) were recruited from a medical center for reproductive biology. The levels of serum reactive oxygen metabolites and seminal lipid peroxides were assessed for both control and varicocele subjects. Sperm DNA fragmentation was measured by sperm chromatin dispersion test. Mitochondrial respiratory activity was evaluated with a polarographic assay of oxygen consumption carried out in hypotonically-treated sperm cells. RESULTS: In this study, varicocele patients were compared with men without varicoceles. Oxidative stress was observed in the serum and seminal fluid of varicocele patients. These patients showed an increase of 59% (P < 0.05) in serum reactive oxygen metabolites and a threefold increase in the level of sperm lipid peroxides. A parallel and significant increase (a twofold increase; P < 0.05) in the degree of sperm DNA fragmentation was also observed. Varicocele patients showed a 27% decrease (P < 0.05) in mitochondrial respiratory activity in comparison to the control group. A 32% increase (P < 0.05) in sperm midpiece defects and a 41% decrease (P < 0.05) in sperm concentration and motility were also observed. CONCLUSIONS: Men with varicocele have increased markers of oxidative stress and decreased mitochondrial respiratory activity. These results correlated with abnormalities in semen parameters. For morphology these correlated with midpiece defects.

The present invention refers to a micro culture device (5) to be inserted in a chamber (4) of an oxygraph (7). The micro culture device is divided into an upper microchamber (2) and a lower microchamber (3) by a semi-permeable nanoporous membrane (1), suitable for acting as adhering support for cells. The upper microchamber (2) presents at its upper side either an inlet (2.1) coaxially aligned with an oxygraph plunger precision bore (6), or a semi-permeable membrane. The lower microchamber (3) presents at its bottom side either an inlet (3.1) coaxially aligned with an oxygraph piston bore (8) or a semi-permeable membrane (3.2).