Immunohistochemical and immunological detection of ghrelin and leptin in rainbow trout Oncorhynchus mykiss and murray cod Maccullochella peelii peelii as affected by different dietary fatty acids

In this study, we report ghrelin and leptin immunoreactive (ir) cells distribution in the gastrointestinal tract and blood ghrelin and leptin levels in rainbow trout (Oncorhynchus mykiss) and Murray cod (Maccullochella peelii peelii) fed diets with different fatty acid compositions. Juvenile rainbow trout and Murray cod were fed five iso-energetic experimental diets containing fish oil (FO) or one of the following vegetable oils (VO): olive oil (OO), sunflower oil (SO), linseed oil (LO), and palm oil (PO); as the added dietary lipid source. The presence and distribution of both ghrelin and leptin ir cells in the gastrointestinal tract were affected by the inclusion of VO. Ghrelin ir cells were found in the gastric glands of rainbow trout and in the mioenteric plexuses of the stomach of Murray cod fed FO. Ghrelin ir cells were localized in the mucosa of the intestine of rainbow trout and Murray cod fed VO. Leptin ir cells were more abundant in the epithelial lining of the mucosa folds and in the glands of the stomach of rainbow trout fed VO. Leptin immunoreactivity was detected in the gastric mioenteric plexus of Murray cod fed FO. No differences were found both in ghrelin and leptin levels in blood plasma or in the growth rates of rainbow trout and Murray cod fed the different experimental diets. These observations suggest that dietary fatty acids play a role in the peripheral feeding regulation.     

The use of fractal dimension and lacunarity in the characterization of mast cell degranulation in rainbow trout (Onchorhynchus mykiss)

Fractal analysis is a reliable method for describing, summarizing object complexity and heterogeneity and has been widely used in biology and medicine to deal with scale, size and shape management problems. The aim of present survey was to use fractal analysis as a complexity measure to characterize mast cells (MCs) degranulation in a rainbow trout ex vivo model (isolated organ bath). Compound 48/80, a condensation product of N‐methyl‐p‐methoxyphenethylamine with formaldehyde, was adopted as MCs degranulation agent in trout intestinal strips. Fractal dimension (D), as a measure of complexity, ‘roughness’ and lacunarity (λ), as a measure of rotational and translational invariance, heterogeneity, in other words, of the texture, were compared in MCs images taken from intestinal strips before and after compound 48/80 addition to evaluate if and how they were affected by degranulation. Such measures were also adopted to evaluate their discrimination efficacy between compound 48/80 degranulated group and not degranulated group and the results were compared with previously reported data obtained with conventional texture analysis (image histogram, run‐length matrix, co‐occurrence matrix, autoregressive model, wavelet transform) on the same experimental material. Outlines, skeletons and original greyscale images were fractal analysed to evaluate possible significant differences in the measures values according to the analysed feature. In particular, and considering outline and skeleton as analysed features, fractal dimensions from compound 48/80 treated intestinal strips were significantly higher than the corresponding untreated ones (paired t and Wilcoxon test, p < 0.05), whereas corresponding lacunarity values were significantly lower (paired Wilcoxon test, p < 0.05) but only for outline as analysed feature. Outlines roughness increase is consistent with an increased granular mediators interface, favourable for their biological action; while lacunarity (image heterogeneity) reduction is consistent with the biological informative content decrease, due to granule content depletion. In spite of the significant differences in fractal dimension and lacunarity values registered according to the analysed feature (greyscale obtained values were, on average, lower than those obtained from outlines and skeletons; General Linear Model, p < 0.01), the discrimination power between not degranulated and degranulated MCs was, on average, the same and fully comparable with previously performed texture analysis on the same experimental material (outline and skeleton misclassification error, 20% [two false negative cases]; greyscale misclassification error, 30% [two false negative cases and one false positive case]). Fractal analysis proved to be a reliable and objective method for the characterization of MCs degranulation.