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

Abstract

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 veterinary medicine, 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 veterinary medicine for the evaluation of metabolic and nutritional status in healthy and diseased dogs.

The Fatty Acid-Based Erythrocyte Membrane Lipidome in Dogs with Chronic Enteropathy

“…Dogs already in dietary management were not excluded from the study. The control group was made of 68 healthy dogs, whose fatty acid-based membrane lipidome profile was recently published [ 31 ].…”

“…Whole blood in EDTA as anticoagulant was used to isolate red blood cells and perform membrane lipidome analysis, as previously described [ 31 ]. Briefly, starting from a 1 mL whole blood sample, the separation of erythrocytes from plasma, followed by washings and centrifugation for the membrane isolation, finally produces a pellet that was re-suspended in pure water to extract lipids using 2:1 chloroform:methanol as the organic phase [ 31 ]. The organic layer was separated and evaporated under vacuum to dryness.…”

“…In Supplementary Information , there are details of the calibration procedure and a figure of a representative GC run ( Supplementary Figures S2 and S3 ) with the 10 fatty acids identified by appropriate standard references, which are commercially available as reference materials at known concentrations. The quantification procedure was performed as already described [ 31 ] and reported in Supplementary Information . The data of the 10 FAs are represented in μg/mL, and each quantity is reported as percentage relative to the cluster as sum of the quantities (100%) that corresponds to >97% of the peaks found in the GC runs (% rel.…”

“…As previously described [ 31 ], a cluster of 10 fatty acids, representative of the main fatty acid moieties present in the cell membrane, was chosen. In particular, the cohort of fatty acids included: palmitic (C16:0) and stearic (C18:0) acids as SFAs; palmitoleic (C16:1), oleic (9c, C18:1), and vaccenic (11c, C18:1) acids as MUFAs; linoleic (LA, C18:2), dihomo-gamma-linolenic (DGLA; C20:3), and arachidonic (AA, C20:4) acids as PUFA ω-6; and eicosapentaenoic (EPA, C20:5) and docosahexaenoic (DHA, C22:6) acids as PUFA ω-3.…”

“…The benchmark of a healthy dog membrane lipidome was reported following the protocol used also for humans, isolating the blood fraction containing erythrocytes [ 31 ], and based on this premise, we were interested in characterizing dogs affected by CE, to investigate the molecular differences from the healthy condition. It can be expected that the quantity and quality of fatty acids in the erythrocyte membrane lipidome may mirror the “gut health” in dogs affected by different forms of CE, allowing us to gain new insights related to the balance among fatty acid types that regulate signaling cascades.…”