Apoptosis of gut-associated lymphoid tissue in rainbow trout Oncorhynchus mykiss after incubation with Candida albicans and bacterial lipopolysaccharide.

Until now a few studies have been carried out on the gut lymphoid system in fish despite its protective role in the host. Here, we have evaluated the effects of Candida albicans (Ca) and lipopolysaccaride (LPS) on the pyloric and terminal segments of gut in the rainbow trout Oncorhynchus mykiss. In particular, data show that both Ca and LPS are able to cause apoptosis of intestinal lymphoid cells as detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) procedure. These findings suggest a further modality of gut response in fish to environmental antigens.

Antigenically activated avian erythrocytes release cytokine-like factors: a conserved phylogenetic function discovered in fish.

Fish erythrocytes are endowed with the ability to produce cytokine like factors when stimulated with Candida albicans (Ca). To evaluate whether similar activities are still conserved in bird erythrocytes (BE), a morphological, cytochemical and immunological evaluation was conducted on peripheral cells in chickens (Gallus gallus). BE form rosettes with monocytes (Mo)-macrophages (MØ), and Mo-MØ according to cytochemical analysis to maintain phagocytic functions across the evolution. Finally, Ca-activated BE release in the supernatants cytokine like-factors that enhance Mo-MØ phagocytosis (interferon-gamma-like activity) and inhibit Mo-MØ migration in agarose (migration inhibitory factor activity). In conclusion, bird erythrocytes, as nonimmune cells, are able to participate in the immune response contributing to the host defence.

Do fish thrombocytes play an immunological role? Their cytoenzymatic profiles and function during an accidental piscine candidiasis in aquarium.

Fish (F) thrombocytes (THRs) from healthy trouts were studied in terms of cytoenzyme expression. FTHRs were positive to acid periodic of shiff (PAS) and acid phosphatase (ac. phos.) without tartaric acid (-TA) stainings, as well to alkaline phosphatase. However, when compared with autologous macrophages (M psi's), they were negative to naphthol cloroacetate esterase (AS-D), alpha-naphthyl acetate esterase (Anae), peroxidase (perox) and control ac. phos. with tartaric acid (+TA) stainings, thus indicating a lack of typical lysosomial enzymes. This evidence supports the notion that FTHRs are not true digesting cells. Quite interestingly, trouts and human M psi's were positive for PAS, AS-D, Anae, and perox stainings, thus confirming that cellular cytochemistries are maintained across evolution as their phagocytic functions. Additionally, blood films from trouts, accidentally infected with Candida albicans in aquarium, were morphologically analyzed. Actually, FTHRs interact with erythrocytes, potentiating the formation of rosettes around a central Mpsi. Polymorph nuclear cells and lymphocytes are present in these cellular aggregates, thus suggesting that FTHRs may represent a link between innate and adaptive immunity.

Lymphoreticular system in fish: erythrocyte-mediated immunomodulation of macrophages contributes to the formation of melanomacrophage centers.

In fish, melanomacrophage centers (MMC) represent the equivalent of germinal centers of birds and mammals. They consist of aggregates of macrophages filled with fragments of cells, mainly erythrocytes, and various pigments such as hemosiderin, lipofuscin, and melanin. The kidney, the spleen, and the liver contain many MMC either moving freely or encapsulated in the context of the lymphoreticular tissue. Here, we have demonstrated for the first time that MMC exist in the kidney, spleen, and liver of the rainbow trout Salmo gairdneri Richardson by using traditional stainings and cytoenzymatic methods: periodic acid-shiff, alpha-naphthyl acetate esterase, and peroxidase. Finally, in the view of the already described peripheral interaction between fish erythrocytes (FE) and macrophages (MO), we postulate that these cellular aggregates (rosettes) travel to distant organs, thus contributing to the formation of MMC in the kidney, spleen, and liver.

Maturation of fish erythrocytes coincides with changes in their morphology, enhanced ability to interact with Candida albicans and release of cytokine-like factors active upon autologous macrophages.

Erythrocytes from the rainbow trout Salmo gairdneri Richardson (Salmo g.R.) were classified into immature and mature populations, respectively, by measuring longitudinal diameters. More elongated fish erythrocytes (FE), classified as mature cells, were those interacting with Candida albicans (CA) in a higher frequency in terms of either binding to the fungus or its intracellular engulfment. At the same time, in the rosetting phenomenon more elongated mature FE surrounded macrophages (Mø) phagocytosing CA. Finally, FE activated by CA released in the supernatants cytokine-like factors able to modulate Mø functions. In particular, these active supernatants were analyzed for their capacity to inhibit Mø migration Macrophage Inhibition Factor (MIF) activity and enhance Mø phagocytosis. Both activities were detected in supernatants from CA stimulated FE but not in control supernatants. MIF activity could play a role in the accumulation of Mø in the context of functional rosettes, while the factor enhancing Mø phagocytosis could promote clearance of CA in a more efficacious way.

Fish immunology. II. Morphologycal and cytochemical characterization and phagocytic activities of head kidney macrophages from rainbow trout (Salmo gairdneri Richardson).

In Salmo gairdneri Richardson trouts, a comparison was made between macrophages (MO) derived from head kidney and peripheral blood monocytes. Morphologically and cytochemically no differences were observed between these two types of mononucleated cells. On the other hand, in parallel studies the ability of trout erythrocytes to engulf Candida albicans (CA) was evaluated and compared to the MO phagocytosis. In erythrocytes, engulfment is preceded by binding to CA and cell membrane invagination, while this was not the case for MO. Finally, MO stimulated with lipopolysaccharides (LPS) did not modify their phagocytic capacities, thus suggesting a lack of LPS receptors or a tolerance to LPS.

Fish immunology. I. Binding and engulfment of Candida albicans by erythrocytes of rainbow trout (Salmo gairdneri Richardson).

The role of fish erythrocytes (FE) as phagocytic cells has poorly been investigated, until now. Here, we have focussed our attention on the interplay between rainbow trout (Salmo gairdneri Richardson) erythrocytes and Candida albicans (CA). At the same time, the intervention of autologous head kidney macrophages (MO) in the CA processing by FE has been studied. Data show that CA particles bind to FE, which, in turn, are able to engulf but not kill them. In the presence of MO, a decrease of FE with bound CA occurs and, in some microscopic images, FE form rosettes with MO. Phagocytosis of CA is higher in rosetting MO than in non-rosetting ones. According to our findings, it appears that FE represent a reservoir of engulfed CA and rosetting is an efficacious phenomenon of presentation of pathogens to MO, where an effective clearance of them can take place.

The rainbow trout (Salmo gairdneri Richardson) granular leukocytes and mast-cells system versus the human one. Cytomorphometrical and cytochemical characters in these two species philogenetically poles apart.

In man, circulating granular leukocytes constitute a cellular system and are able to migrate in the tissues to take part in the immune reactions. This study was to characterize the granular leukocytes and the eventual existence of mast-cells of a low vertebrate, rainbow trout (Salmo gairdneri R.) in order to compare it with man. The blood smears and tissues sections have been tested whit panoptic methods, cytochemistry and toluidine blue. White blood cell count, leukocytes formula and cytomorphometric characterization was performed using an image analyser. Scanning of tissues sections in order to identify mast-cells has been also performed. In this model the granular leukocytes are all neutrophilic like; no eosinophilic, no basophilic no tissue mast-cells, basically existing in allergic and anaphylactic reactions, were found.

Corrosion cast of the vascularization of Mugil Cephalus gills.

The spatial organization of the microcirculation in gills of Mugil Cephalus, was examined by scanning electron microscopic analysis of corrosion cast prepared by intravascular injection of methyl methacrylate. The afferent branchial artery originates from the ventral aorta and gives rise to afferent filamental artery. From the medio-lateral wall of the afferent filamental artery, afferent lamellar arterioles originate which supply one or more lamellae. The lamellar efferent arterioles, which drain the blood coming from the lamellae into the efferent filamental arteries, continue with the efferent branchial arteries and then the dorsal aorta. The techniques used so permitted to evaluate the structure and the interrelationships of the vascular pathways, explaining the regulation and the distribution of the blood flow in the gills.

GABA concentrations in different cerebral areas of bass (dicentrarchus labrax).

In these investigations GABA concentrations in different cerebral areas (cortex, diencephalon, optic lobes and mesencephalon) of bass (Dicentrarchus labrax) were evaluated. According to the results obtained, the GABA, like other chemical mediators (noradrenaline, dopamine, serotonin), investigated in different animal species, shows a variable distribution in the CNS of bass. The highest levels of GABA were observed in the cerebral cortex and the lowest ones in the mesencephalon. Mean values were observed in the diencephalon and the optic lobes. The study of the GABA-ergic system of the CNS of fish, could provide a valid contribution to the comprehension of several biochemical and physiopathological mechanisms responsible for the behavioural activity and/or the appearance of neurotoxic phenomena.

[Comparative anatomo-microscopic observations on the esophagus of teleosts with different feeding habits]. / Osservazioni anatomo-microscopiche comparate sull'esofago di teleostei a diverso regime alimentare.

The authors describe the micro-anatomy of the oesophagus of different diet fish: herbivorous (Boops salpa), omnivorous (Liza aurata) and carnivorous fish (Blennius gattorugine and Gaidropsarus mediterraneus). In all the observed species the oesophagus is formed by a tunica mucosa, tunica muscularis and tunica serosa. The tunica mucosa of the herbivorous fish arises to form thin plicae which protrude into the lumen and are covered with squamous stratified epithelium. The goblet cells do not arrange among the covering cells but take place laterally and on the basis of the plicae. The tunica muscularis is constituted by an external circular layer and an inner longitudinal layer of striated muscle. The oesophagus of omnivorous fish shows thick plicae covered with a higher epithelium than that of the herbivorous fish. Among the epithelial cells a lot of taste buds have been found. The tunica muscularis is like that of the herbivorous fish. In the carnivorous fish Blennius the tunica mucosa presents thin plicae like those of the herbivorous fish but they are covered with a thicker epithelium. The tunica muscularis is formed by an external circular layer and by an inner longitudinal layer which constitutes two diametrically opposed bundles placed dorsally and ventrally. In the other carnivorous fish Gaidropsarus, the tunica mucosa shows very thick plicae covered with a quite high squamous epithelium among which there are many goblet cells. In this fish too numerous taste buds have been found. The tunica muscularis is arranged in the same manner observed in the other carnivorous fish.