The lipidomic analysis in the veterinary field: Study of the red blood cell membrane lipid profile in physiological and pathological conditions

The role of lipids in health and disease in humans has been recognized for many decades and over the last fifteen years there has been an intense effort to develop suitable methodologies to discover, identify, and quantitatively monitor lipids in biological systems. That led to a dramatical increase on the use lipidomics in search of new possible biomarkers or to better understand pathological mechanisms. The cell membrane lipidomics is based on homeostatic balance of the fatty acid metabolic pathways and can give us important information on the composition activity and the cause of oxidative stress in the cellular membranes. Erythrocyte membrane lipidomic analysis offers some new perspectives in nutraceuticals and personalized health interventions in order to help therapies at molecular level, to maintain the best homeostasis possible in a pathological condition. Several studies have shown modification of the phospholipid content of cell membranes and point to alterations of the enzymic activity on the fatty acid metabolic pathways in diabetic patients. Using a protocol widely tested in human profiling, in the present study erythrocyte membrane lipidome was examined in healthy and diseased dogs. In particular, a cluster composed of 10 cis fatty acids, present in membrane glycerophospholipids and representative of structural and functional properties of cell membrane, was chosen and quantitatively analyzed. This cluster consists of: 2 saturated fatty acids (SFA: palmitic and stearic acids); 3 monounsaturated fatty acids (MUFA, palmitoleic, oleic and cis-vaccenic acids); 3 polyunsaturated fatty acids omega-6 (PUFA, linoleic, dihomo-gamma linolenic, arachidonic acids); 2 polyunsaturated fatty acids omega-3 (PUFA, eicosapentaenoic and docosahexaenoic acids). For the purpose of this study, blood samples were collected from 68 clinically healthy dogs, including 30 males (6 neutered) and 38 females (12 sterilized), weighting from 2.6 to 43 kg, aged from 2 to 156 months (median 41). The interval values and distribution for each fatty acid of the cluster were determined, providing the first panel describing the healthy dog lipidomic membrane profile. We used these interval values to evaluate correlations between FAME types, families and lipid indexes with the dog characteristics and the results showed an interesting correlation to bodyweight increase. More specific, with the increase of the bodyweight, an increase in palmitic acid and in the total levels of SFA, while a decrease of the omega-6 and the total PUFA was observed. The same intervals when then used to evaluate diseased dogs: ι) 49 dogs affected with chronic enteropathy (CE), consisting of 17 females (7 spayed) and 32 males (1 neutered), with a median age of 47 months (range 4–144). Interestingly, the CE dogs had higher values of palmitic acid and lower levels of stearic acid compared to healthy dogs, with an overall reduction of total SFA. In addition, the CE dogs showed reduced content of LA and increased DGLA when compared to healthy dogs. ii) 12 dogs affected with diabetes mellitus (DM), consisting of 7 females (5 spayed) and 5 males (1 neutered), with a median age of 131 months (range 94–181). The results obtained showed increased levels of palmitoleic and oleic acids as well as the total MUFA content on DM dogs when compared to the healthy dogs. These fatty acids cannot derive from the enzymatic transformation of palmitic acid by the delta-9 desaturase and their increased levels indicate an accelerated Δ9 desaturase activity on the DM dogs. The activation of this enzymatic transformation is known to be related to the insulin response and the carbohydrate management, which is connected to lipid biosynthesis through the pyruvate and acetyl-CoA pathways. In the current study, we provided the first panel of erythrocyte membrane fatty acids in healthy dogs choosing a cohort representative for the main structural and functional roles of these hydrophobic molecules in the cell membrane compartment. and demonstrated a first utility of the interval values established for healthy dogs to evaluate the condition of these two pathological conditions. The results obtained from this study indicate that the erythrocyte membrane lipidome of animal samples may be successfully applied in veterinary medicine and provide important of the cell membrane status in physiological and pathological conditions and a better understanding to how membranes are influenced by dietary habits

Fatty acid-based lipidomics and membrane remodeling induced by apoE3 and apoE4 in human neuroblastoma cells

Apolipoprotein E (apoE) is a major lipid carrier of the lipoprotein transport system that plays critical roles in various pathologies. Human apoE has three common isoforms, the apoE4 being associated with Alzheimer's disease. This is the first study in the literature investigating the effects of apoE (apoE3 and apoE4 isoforms) on membrane fatty acid profile in neuroblastoma SK-N-SH cells. Fatty acid analyses were carried out by gas chromatography of the corresponding methyl esters (FAME). We observed the occurrence of membrane fatty acid remodeling in the presence of each of the two apoE isoforms. ApoE3 increased the membrane level of stearic acid and dihomo-gamma-linolenic acid (DGLA), whereas apoE4 had opposite effects. Both apoE3 and apoE4 increased saturated and monounsaturated fatty acids (SFA and MUFA), omega-6/omega-3 ratio and decreased total polyunsaturated fatty acid (PUFA) amount, but with various intensities. Moreover, both apoE isoforms decreased membrane homeostasis indexes such as PUFA balance, unsaturation index and peroxidation index. Our results highlight membrane property changes connected to the apoE isoforms suggesting membrane lipidomics to be inserted in further model studies of apolipoproteins in health and disease

Effects of Aging and Disease Conditions in Brain of Tumor-Bearing Mice: Evaluation of Purine DNA Damages and Fatty Acid Pool Changes

The consequences of aging and disease conditions in tissues involve reactive oxygen species (ROS) and related molecular alterations of different cellular compartments. We compared a murine model of immunodeficient (SCID) xenografted young (4 weeks old) and old (17 weeks old) mice with corresponding controls without tumor implantation and carried out a compositional evaluation of brain tissue for changes in parallel DNA and lipids compartments. DNA damage was measured by four purine 5′,8-cyclo-2′-deoxynucleosides, 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxo-dG), and 8-oxo-7,8-dihydro-2′-deoxyadenosine (8-oxo-dA). In brain lipids, the twelve most representative fatty acid levels, which were mostly obtained from the transformation of glycerophospholipids, were followed up during the aging and disease progressions. The progressive DNA damage due to age and tumoral conditions was confirmed by raised levels of 5′S-cdG and 5′S-cdA. In the brain, the remodeling involved a diminution of palmitic acid accompanied by an increase in arachidonic acid, along both age and tumor progressions, causing increases in the unsaturation index, the peroxidation index, and total TFA as indicators of increased oxidative and free radical reactivity. Our results contribute to the ongoing debate on the central role of DNA and genome instability in the aging process, and on the need for a holistic vision, which implies choosing the best biomarkers for such monitoring. Furthermore, our data highlight brain tissue for its lipid remodeling response and inflammatory signaling, which seem to prevail over the effects of DNA damage

The Lipidomics of Spermatozoa and Red Blood Cells Membrane Profile of Martina Franca Donkey: Preliminary Evaluation

: Fatty acid-based lipidomic analysis has been widely used to evaluate health status in human medicine as well as in the veterinary field. In equine species, there has been a developing interest in fertility and sperm quality. Fatty acids, being the principal components of the membranes, play an active role in the regulation of the metabolic activities, and their role on spermiogenesis seems to be of great importance for the resulting quality of the sperm and, thus, fertility. With the application of widely used lipidomic techniques, the aim of this study was to evaluate: (a) the fatty acid content of the spermatozoa's membranes of 26 healthy male Martina Franca donkeys and its possible correlation with sperm parameters, and (b) the evaluation of the composition of the red blood cells' membrane. PUFA omega-6 are the principal components (40.38%) of the total PUFA content (47.79%) in both types of cells; however, DPA is the predominant one on the spermatozoa's membrane (27.57%) but is not present in the erythrocyte's membrane. Spermatozoa's motility (%) is positively correlated with stearic acid and EPA, and progressive motility (%), with oleic acid. These findings offer information on the composition of both types of cells' membranes in healthy male MF donkeys and reflect the metabolic transformations of the spermatozoa's membrane during the maturation period, providing a better perception of the role of fatty acids in sperm parameters and fertility

The Lipidomics of Spermatozoa and Red Blood Cells Membrane Profile of Martina Franca Donkey: Preliminary Evaluation

Fatty acid-based lipidomic analysis has been widely used to evaluate health status in human medicine as well as in the veterinary field. In equine species, there has been a developing interest in fertility and sperm quality. Fatty acids, being the principal components of the membranes, play an active role in the regulation of the metabolic activities, and their role on spermiogenesis seems to be of great importance for the resulting quality of the sperm and, thus, fertility. With the application of widely used lipidomic techniques, the aim of this study was to evaluate: (a) the fatty acid content of the spermatozoa’s membranes of 26 healthy male Martina Franca donkeys and its possible correlation with sperm parameters, and (b) the evaluation of the composition of the red blood cells’ membrane. PUFA omega-6 are the principal components (40.38%) of the total PUFA content (47.79%) in both types of cells; however, DPA is the predominant one on the spermatozoa’s membrane (27.57%) but is not present in the erythrocyte’s membrane. Spermatozoa’s motility (%) is positively correlated with stearic acid and EPA, and progressive motility (%), with oleic acid. These findings offer information on the composition of both types of cells’ membranes in healthy male MF donkeys and reflect the metabolic transformations of the spermatozoa’s membrane during the maturation period, providing a better perception of the role of fatty acids in sperm parameters and fertility

The Erythrocyte Membrane Lipidomeof Healthy Dogs: Creating a Benchmark of Fatty Acid Distribution and Interval Values

Molecular-based approaches are rapidly developing in medicine for the evaluation of physiological and pathological conditions and discovery of new biomarkers in prevention and therapy. Fatty acid diversity and roles in health and disease in humans are topical subjects of lipidomics. In particular, membrane fatty acid-based lipidomics provides molecular data of relevance in the study of human chronic diseases, connecting metabolic, and nutritional aspects to health conditions. In veterinarymedicine,membrane lipidomics, and fatty acid profiles have not been developed yet in nutritional approaches to health and in disease conditions. Using a protocol widely tested in human profiling, in the present study erythrocyte membrane lipidome was examined in 68 clinically healthy dogs, with different ages, sex, and sizes. In particular, a cluster composed of 10 fatty acids, present in membrane glycerophospholipids and representative of structural and functional properties of cell membrane, was chosen, and quantitatively analyzed. The interval values and distribution for each fatty acid of the cluster were determined, providing the first panel describing the healthy dog lipidomic membrane profile, with interesting correlation to bodyweight increases. This molecular information can be advantageously developed as benchmark in veterinarymedicine for the evaluation of metabolic and nutritional status in healthy and diseased dogs