A. Cianciotta

FISH IMMUNOLOGY. I. BINDING AND ENGULFMENT OF CANDIDA ALBICANS BY ERYTHROCYTES OF RAINBOW TROUT (SALMO GAIRDNERI RICHARDSON)

ABSTRACT 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 (MØ) 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 MØ, a decrease of FE with bound CA occurs and, in some microscopic images, FE form rosettes with MØ. Phagocytosis of CA is higher in rosetting MØ 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 MØ, where an effective clearance of them can take place.

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 (MØs), they were negative to naphthol cloroacetate esterase (AS-D), α-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 MØs 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 MØ. Polymorph nuclear cells and lymphocytes are present in these cellular aggregates, thus suggesting that FTHRs may represent a link between innate and adaptive immunity.

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.

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, α-naphthyl acetate esterase, and peroxidase. Finally, in the view of the already described peripheral interaction between fish erythrocytes (FE) and macrophages (MØ), we postulate that these cellular aggregates (rosettes) travel to distant organs, thus contributing to the formation of MMC in the kidney, spleen, and liver.

Expression of proto-oncogene C-kit and correlation with morphological evaluations in canine cutaneous mast cell tumors.

Canine cutaneous mast cell tumor (MCT) is very common disease in dogs, this is more aggressive than in other species. The biologic behavior of MCT is highly variable and a more accurate prognosis for these tumors needs to performed. The proto-oncogene c-kit is known to play a critical role in development and function of mast cells (MC). The aim of this study was to evaluate the expression of immunohistochemical pattern of c-kit in MCTs and to correlate these results with MC density (MCD) and intratumoral microvessel density (MVD). Our results confirm that a more aggressive biologic behavior of canine MCT is associated with the increased c-kit expression, further suggesting a new role for c-kit, as a useful marker, in diagnostic pathology and in tumor progression.

In Vitro Elicitation of Intestinal Immune Responses in Teleost Fish: Evidence for a Type IV Hypersensitivity Reaction in Rainbow Trout

In fish the gut immune system has been the subject of few investigations until now. Here, we provide novel morphological and immunological data on the gut isolated from rainbow trout Salmo gairdneri. The pyloric (P) and terminal (T) segments of trout gut, when morphologically examined, evidenced lymphocytes and macrophages (MØ) loosely dispersed in the intestinal mucosa and in the lamina propria in the absence of typical Peyers patches-like structures. Furthermore, incubation of P and T sections with Candida albicans (Ca) and functional analysis of supernatants generated some interesting results. In fact, active supernatants, when compared with controls, exhibited cytokine-like activities attributable to the presence of interferon (IFN)-γ and migration inhibiting factor (MIF), respectively. In particular, IFN-γ-like activity gave rise to an enhancement of Ca phagocytosis by MØ, whereas MIF inhibited MØ migration in agarose. Taken together, these in vitro data suggest that the gut-associated lymphoreticular tissue in fish possesses the appropriate armamentarium to mount a type IV hypersensitivity response when challenged by microbial antigens.

First observation of an Hashimoto thyroiditis-like disease in horses from Eastern Europe: histopathological and immunological findings.

Among 622 slaughtered horses from eastern Europe, 156 thyroid glands were selected on the basis of macroscopic alterations (e.g., determination of volume and weight). In the 80% of these thyroids, microscopic alterations consistent with a diagnosis of Hashimoto thyroiditis-like disease were found. In particular, a subverted architecture of the thyroid gland with colloid rarefaction, lymphocytic infiltration, and fibrosis was noted. The confirmation of the histopathological diagnosis of an equine Hashimoto thyroiditis-like disease was provided by the increased serum concentration of thyroglobulin, of antithyroglobulin, and of antithyroid peroxidase autoantibodies. Despite evidence consistent with an Hashimoto thyroiditis-like disease in eastern European horses, the etiopathogenesis of this autoimmune disorder deserves further investigation. In this respect, in some horses histopathological alterations in the pituitary gland may suggest an as-yet-unidentified disorder within the hypothalamus-pituitary adrenal axis associated with Hashimoto thyroiditis.

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.

Modifications of Serum and Cellular Parameters in Trotters After a Race. Macrophage Migration Inhibitory Activity Reduction and Serum β-Glucan Elevation

Trotters are exposed to a chronic prolonged stress, such as daily training and frequent races during their active lifespans. There is evidence that trotters undergo very often lethal lung infections after a race, and therefore, is likely that modifications of certain physiologic cellular parameters could account for the increased susceptibility to microbial diseases. Here, we demonstrate that in 7 trotters after a race either serum values (e.g., glycaemia, triglycerides, transaminases, gamma-glutamyltransferase, cholinesterase, amylase, alkaline phosphatase, total proteins, serum albumin, sodium, blood urea nitrogen, lactic dehydrogenase, creatine phosphokinase, and creatinine) or hematological parameters (red blood cell count, hemoglobin, lymphocyte and monocyte count) were increased. At the same time, in the same animals after a race, macrophage migration inhibitory factor activity was depressed, thus indicating an impaired T-lymphocyte response. Finally, increased levels of circulating β-glucans in some horses, after a race, may suggest a reduced clearance of fungal cell wall components. Taken together, these findings indicate a condition of multiple organ dysfunction, such as the liver, the kidney, the pancreas, and skeletal muscles, as well as a reduced cell-mediated immune response in trotters, after a race.

A phylogenetic comparison between acute monocytic leukemia cells and monocytes-macrophages in lower vertebrates

In humans, monocytes and macrophages (MØ) play a central role in immune regulation, tissue maintenance and pathogen control. In lower vertebrates, a few studies have been conducted on MØ like cells. In acute monocytic leukemia monocytic cells, as immature cells restrained in one of the phases of their ontogenesis, would offer the opportunity to rebuild an archaic condition helpful to understand the phylogenesis. Therefore, aim of this work was to characterize in the Rainbow trout (Salmo Gairdneri Richardson) MØ and compare them with acute leukemia monocytic cells. In the trout, MØs morphology is similar to that of mammals. In particular, MØ possess an irregular embryoshaped nucleus occupying 2/3 of the cell, while the peripheral cytoplasmic profile is irregular with extroflexed plasmalemma and pseudopods. A morphological transition towards MØ is featured by a wavy hyaline classical membrane and an irregular and extroflexed surface. Some aspects of erythrophagocytosis represented a finding of great interest indicating that the hemocatheretic function could take place directly in circulation. This condition, also observed in human acute monocytic leukemia, suggests that the information to the erythrophagocytosis is restrained under physiological conditions. Non-specific esterases, which are positive in human MØ smear and MØ from human lymph node tissue, were also positive in the teleost studied but with a dysomogeneous pattern. Consequently non-specific esterase system is phylogenetically conserved. A lack of immune-reactivity with the anti-CD68 monoclonal antibody (MoAb) on smear and trout tissue sections was observed. On the contrary, strong positivity was detected on human lymph node sections. In trout, the presence of MØ and circulating MØ like cells exhibiting an erythrocatheretic function in the circulation would indicate a primordial function that has later been replaced by the liver and the spleen.