Acyl-CoA synthetase 4 modulates mitochondrial function in breast cancer cells.

Mitochondria are dynamic organelles that respond to cellular stress through changes in global mass, interconnection, and subcellular location. As mitochondria play an important role in tumor development and progression, alterations in energy metabolism allow tumor cells to survive and spread even in challenging conditions. Alterations in mitochondrial bioenergetics have been recently proposed as a hallmark of cancer, and positive regulation of lipid metabolism constitutes one of the most common metabolic changes observed in tumor cells. Acyl-CoA synthetase 4 (ACSL4) is an enzyme catalyzing the activation of long chain polyunsaturated fatty acids with a strong substrate preference for arachidonic acid (AA). High ACSL4 expression has been related to aggressive cancer phenotypes, including breast cancer, and its overexpression has been shown to positively regulate the mammalian Target of Rapamycin (mTOR) pathway, involved in the regulation of mitochondrial metabolism genes. However, little is known about the role of ACSL4 in the regulation of mitochondrial function and metabolism in cancer cells. In this context, our objective was to study whether mitochondrial function and metabolism, processes usually altered in tumors, are modulated by ACSL4 in breast cancer cells. Using ACSL4 overexpression in MCF-7 cells, we demonstrate that this enzyme can increase the mRNA and protein levels of essential mitochondrial regulatory proteins such as nuclear respiratory factor 1 (NRF-1), voltage-dependent anion channel 1 (VDAC1) and respiratory chain Complex III. Furthermore, respiratory parameters analysis revealed an increase in oxygen consumption rate (OCR) and in spare respiratory capacity (SRC), among others. ACSL4 knockdown in MDA-MB-231 cells led to the decrease in OCR and in SCR, supporting the role of ACSL4 in the regulation of mitochondrial bioenergetics. Moreover, ACSL4 overexpression induced an increase in glycolytic function, in keeping with an increase in mitochondrial respiratory activity. Finally, there was a decrease in mitochondrial mass detected in cells that overexpressed ACSL4, while the knockdown of ACSL4 expression in MDA-MB-231 cells showed the opposite effect. Altogether, these results unveil the role of ACSL4 in mitochondrial function and metabolism and expand the knowledge of ACSL4 participation in pathological processes such as breast cancer.

Supplementation with EPA and DHA omega-3 fatty acids improves peripheral immune cell mitochondrial dysfunction and inflammation in subjects with obesity.

Omega-3 fatty acids (w-3 FA) have anti-inflammatory effects and improve mitochondrial function. Nonetheless, little is known about their effect on mitochondrial bioenergetics of peripheral blood mononuclear cells (PBMCs) in individuals with obesity. Thus, this study aimed to determine the mitochondrial bioenergetics status and cell subset composition of PBMCs during obesity, before and after 1 month supplementation with w-3 FA. We performed a case-control study with twelve women with normal BMI (lean group) and 19 with grade 2 obesity (obese group), followed by a before-after prospective study where twelve subjects with obesity received a 1 month intervention with 5.25 g of w-3 FA (3.5 g eicosapentaenoic (EPA) and 1.75 g docosahexaenoic (DHA) acids), and obtained PBMCs from all participants. Mitochondrial bioenergetic markers, including basal and ATP-production associated respiration, proton leak, and nonmitochondrial respiration, were higher in PBMCs from the obese group vs. the lean group. The bioenergetic health index (BHI), a marker of mitochondrial function, was lower in the obese vs. the lean group. In addition, Th1, Th2, Th17, CD4+ Tregs, CD8+ Tregs, and Bregs, M1 monocytes and pDCreg cells were higher in PBMCs from the obese group vs. the lean group. The w-3 FA intervention improved mitochondrial function, mainly by decreasing nonmitochondrial respiration and increasing the reserve respiratory capacity and BHI. The intervention also reduced circulating pro-inflammatory and anti-inflammatory lymphocyte and monocytes subsets in individuals with obesity. The mitochondrial dysfunction of PBMCs and the higher proportion of peripheral pro-inflammatory and anti-inflammatory immune cells in subjects with obesity, improved with 1 month supplementation with EPA and DHA.

UV-R mitigation strategies in encapsulated embryos of the intertidal gastropod Acanthina monodon: A way to compensate for lack of parental care.

Intracapsular embryonic development in the intertidal zone exposes embryos to various stress sources characteristic of this environment, including UV-R. They require defensive mechanisms to mitigate its adverse effects. The presence of total carotenoids (TC), and mycosporine-like amino acids (MAAs) was studied in adults, in encapsulated embryos, and in the egg capsule walls of the intertidal gastropod Acanthina monodon. Oxygen consumption rates (OCR) were determined in encapsulated and excapsulated embryos exposed to photosynthetically active radiation (PAR) and PAR + UV-A + UV-B to understand if the capsule wall is a protective structure for encapsulated embryos. The results showed the presence of TC in adult pedal and gonad tissues, and in all encapsulated stages. MAAs were not detected. The physical structure of the capsule wall retained most wavelengths, being particularly efficient in the UV-B range. Excapsulated embryos exposed to PAR + UV-A + UV-B radiation increased its OCR compared to encapsulated embryos, indicating the protective character of the capsule wall.

Non-consumptive effects of a predatory snail (Acanthina monodon) on a dominant habitat-forming mussel species (Perumytilus purpuratus).

Predators can influence prey through direct consumption as well as through non-consumptive effects (NCEs). NCEs usually occur mediated by behavioral changes in the prey upon detection of predator cues. Such changes may involve reduction of feeding with a variety of physiological consequences. We evaluated NCEs from an intertidal predatory snail (Acanthina monodon) on a dominant habitat-forming mussel species (Perumytilus purpuratus) from the southeastern Pacific coast. We tested whether A. monodon exerts negative NCEs on clearance rate, oxygen consumption rate, biodeposit production, and between-valve gap size in P. purpuratus. We found that waterborne predator cues triggered a decrease in these variables except biodeposit production. However, the organic content of the biodeposits increased in the presence of predator cues. The snail's physical contact with the mussels strengthened the negative NCEs on between-valve gap size. Since P. purpuratus is a dominant filter-feeder and foundation species in rocky intertidal habitats, predator NCEs on this species might indirectly influence ecosystem-level processes and community structure.

Phytomodulatory proteins isolated from Calotropis procera latex promote glycemic control by improving hepatic mitochondrial function in HepG2 cells.

The medicinal uses of Calotropis procera are diverse, yet some of them are based on effects that still lack scientific support. Control of diabetes is one of them. Recently, latex proteins from C. procera latex (LP) have been shown to promote in vivo glycemic control by the inhibition of hepatic glucose production via AMP-activated protein kinase (AMPK). Glycemic control has been attributed to an isolated fraction of LP (CpPII), which is composed of cysteine peptidases (95%) and osmotin (5%) isoforms. Those proteins are extensively characterized in terms of chemistry, biochemistry and structural aspects. Furthermore, we evaluated some aspects of the mitochondrial function and cellular mechanisms involved in CpPII activity. The effect of CpPII on glycemic control was evaluated in fasting mice by glycemic curve and glucose and pyruvate tolerance tests. HepG2 cells was treated with CpPII, and cell viability, oxygen consumption, PPAR activity, production of lactate and reactive oxygen species, mitochondrial density and protein and gene expression were analyzed. CpPII reduced fasting glycemia, improved glucose tolerance and inhibited hepatic glucose production in control animals. Additionally, CpPII increased the consumption of ATP-linked oxygen and mitochondrial uncoupling, reduced lactate concentration, increased protein expression of mitochondrial complexes I, III and V, and activity of peroxisome-proliferator-responsive elements (PPRE), reduced the presence of reactive oxygen species (ROS) and increased mitochondrial density in HepG2 cells by activation of AMPK/PPAR. Our findings strongly support the medicinal use of the plant and suggest that CpPII is a potential therapy for prevention and/or treatment of type-2 diabetes. A common epitope sequence shared among the proteases and osmotin is possibly the responsible for the beneficial effects of CpPII.

Relationship between over-crowding within egg capsules of the marine gastropod Acanthina monodon and prospects for juvenile success.

Encapsulated development with extraembryonic yolk may lead to competition for nutrients within egg capsules. In this research, different degrees of competition among embryos in subtidal egg capsules of Acanthina monodon resulted in considerable differences in hatching size. For newly hatched juveniles, individuals hatching from less crowded egg capsules showed better survival, larger SL, higher rates of oxygen consumption, and higher rates of food consumption. However, by 28 days after hatching, the largest surviving juveniles were the best-performing individuals, regardless of the initial embryo density within the capsules. In summary, more crowded egg capsules resulted in poorer survival. These findings may help to explain the variability seen in juvenile success in some field populations; much of that variation may reflect stressful experiences that the new recruits have had during the early stages of their encapsulated development.

Acetyl-CoA-driven respiration in frozen muscle contributes to the diagnosis of mitochondrial disease.

BACKGROUND: Freezing human biopsies is common in clinical practice for storage. However, this technique disrupts mitochondrial membranes, hampering further analyses of respiratory function. To contribute to laboratorial diagnosis of mitochondrial diseases, this study sought to develop a respirometry approach using O2k (Oroboros Ins.) to measure the whole electron transport chain (ETC) activity in homogenates of frozen skeletal muscle biopsies. PATIENTS AND METHODS: We enrolled 16 patients submitted to muscle biopsy in the process of routine diagnostic investigation: four with mitochondrial disease and severe mitochondrial dysfunction; seven with exercise intolerance and multiple deletions of mitochondrial DNA, presenting mild to moderate mitochondrial dysfunction; five without mitochondrial disease, as controls. Whole homogenates of muscle fragments were prepared using grinder-type equipment. O2 consumption rates were normalized using citrate synthase activity. RESULTS: Transmission electron microscopy confirmed mitochondrial membrane discontinuation, indicating increased permeability of mitochondrial membranes in homogenates from frozen biopsies. O2 consumption rates in the presence of acetyl-CoA lead to maximum respiratory rates sensitive to rotenone, malonate and antimycin. This protocol of acetyl-CoA-driven respiration (ACoAR), applied in whole homogenates of frozen muscle, was sensitive enough to identify ETC abnormality, even in patients with mild to moderate mitochondrial dysfunction. We demonstrated adequate repeatability of ACoAR and found significant correlation between O2 consumption rates and enzyme activity assays of individual ETC complexes. CONCLUSIONS: We present preliminary data on a simple, low cost and reliable procedure to measure respiratory function in whole homogenates of frozen skeletal muscle biopsies, contributing to diagnosis of mitochondrial diseases in humans.

Feeding mode influence on treatment performance of unsaturated and partially saturated vertical flow constructed wetland.

This study aimed to evaluate the influence of two different feeding modes on wastewater treatment performance and oxygen consumption rate (OCR) of the unsaturated (UVF wetland) and partially saturated (SVF wetland) vertical flow constructed wetlands operated in parallel under subtropical climate for four years. Each wetland had a superficial area of 7.5 m2 and was planted with Typha domingensis. Both units have a filter media depth of 0.75 m, composed by sand (effective diameter of 0.29 mm and uniformity of 4). UVF wetland operated typically unsaturated, while SVF wetland had the bottom part saturated (57% of total depth). Two feeding modes were evaluated for both wetlands. The feeding mode was operated within the limits recommended by the German standard, with a hydraulic loading rate (HLR) of 75 mm d-1 and specific pulse volume (SPV) of 19 L m-2 for both wetlands and a specific hydraulic loading rate (SHLR) of 8 and 9 L m-2 min-1 for UVF and SVF wetlands, respectively. Meanwhile, the second feeding mode was applied for both wetlands, being an HLR of 103 mm d-1, 26 L m-2 of SPV, and 4 L m-2 min-1 of SHLR. The load removal efficiency of SVF wetland was higher than the UVF wetland for all parameters and feeding modes. No statistical difference was identified for OCR values between wetlands and feeding mode. The results showed that operating UVF and SVF wetlands with a SHLR around 4 L m-2 min-1 and SPV equal of 26 L m-2 is preferable. This fact could represent a significant reduction in inlet pumping power requirements and also less superficial area requirements.

Respiratory analysis of coupled mitochondria in cryopreserved liver biopsies.

The aim of this work was to develop a cryopreservation method of small liver biopsies for in situ mitochondrial function assessment. Herein we describe a detailed protocol for tissue collection, cryopreservation, high-resolution respirometry using complex I and II substrates, calculation and interpretation of respiratory parameters. Liver biopsies from cow and rat were sequentially frozen in a medium containing dimethylsulfoxide as cryoprotectant and stored for up to 3 months at -80 °C. Oxygen consumption rate studies of fresh and cryopreserved samples revealed that most respiratory parameters remained unchanged. Additionally, outer mitochondrial membrane integrity was assessed adding cytochrome c, proving that our cryopreservation method does not harm mitochondrial structure. In sum, we present a reliable way to cryopreserve small liver biopsies without affecting mitochondrial function. Our protocol will enable the transport and storage of samples, extending and facilitating mitochondrial function analysis of liver biopsies.

Volcanic ash in the water column: Physiological impact on the suspension-feeding bivalve Mytilus chilensis.

Ashes settling into the sea from volcanic explosions expose suspension-feeding species to reduced seston quality. Adults and juveniles of the mussel Mytilus chilensis were exposed for 15days to the phytoplankton Isochrysis galbana together with various concentrations of ashes. We then quantified impact on survival and physiology. Although no individuals died during the experiment, by the end of the study clearance rates and oxygen consumption rates had decreased substantially, and tissue weight of mussels exposed to the highest ash concentrations declined substantially. Gills showed no physical damage, but did show abundant mucus secretion in response to ash particles. Moreover, as the relative proportions of microalgae to ash in the diet decreased, individuals showed increasing preferential ingestion of microalgal particles. Increased ash content in the diet altered physiological rates and activated distinct particle selection with a high production of pseudofeces and high energy costs, with potential long-term consequences.