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Antimicrobial peptides (AMPs) are found widespread in nature and possess antimicrobial and immunomodulatory activities. Due to their multifunctional properties, these peptides are a focus of growing body of interest and have been characterized in several fish species. Due to their similarities in amino-acid composition and amphipathic design, it has been suggested that neuropeptides may be directly involved in the innate immune response against pathogen intruders. In this review, we report the molecular characterization of the fish-specific AMP piscidin1, the production of an antibody raised against this peptide and the immunohistochemical identification of this peptide and enkephalins in the neuroepithelial cells (NECs) in the gill of several teleost fish species living in different habitats. In spite of the abundant literature on Piscidin1, the biological role of this peptide in fish visceral organs remains poorly explored, as well as the role of the neuropeptides in neuroimmune interaction in fish. The NECs, by their role as sensors of hypoxia changes in the external environments, in combination with their endocrine nature and secretion of immunomodulatory substances would influence various types of immune cells that contain piscidin, such as mast cells and eosinophils, both showing interaction with the nervous system. The discovery of piscidins in the gill and skin, their diversity and their role in the regulation of immune response will lead to better selection of these immunomodulatory molecules as drug targets to retain antimicrobial barrier function and for aquaculture therapy in the future.

The immune system of a fish has cellular and molecular defense mechanisms that are substantially retained throughout the evolution of vertebrates. The innate immune system provides biological processes, such as phagocytosis and mechanical barriers, to implement an efficient defensive response after exposure to chemical or biological contaminants, pollutants, and contact with parasites, germs, and pathogens. Club cells (CCs) are widespread in the skin of Ostariophysi. After a predator attack or exposure to toxins and parasites, these cells can produce alarming substances. Given their effectiveness against viruses, parasites, and common skin lesions, recent studies have suggested that CCs are a component of the immune system. This study aims to immunohistochemically characterize the CCs for the first time in the skin of zebrafish, using mitogen-activated protein kinase (MAPK) p38, Toll-like receptor (TLR)2, Piscidin1, and inducible nitric oxide synthase (iNOS) peptides involved in the function of all types of vertebrate immune cells. According to our analysis, the intermediate layer of the epidermis exhibited rounded, oval, and elongated CCs, with central acidophilic cytoplasm and a spherical basophilic nucleus, that are positive to the antibodies tested. Our results may confirm that CCs could be involved in the immune function, increasing our knowledge of the immune system of teleosts.

The amphibious teleost Giant mudskipper (Periophthalmodon schlosseri, Pallas 1770) inhabit muddy plains and Asian mangrove forests. It spends more than 90% of its life outside of the water, using its skin, gills, and buccal-pharyngeal cavity mucosa to breathe in oxygen from the surrounding air. All vertebrates have been found to have mast cells (MCs), which are part of the innate immune system. These cells are mostly found in the mucous membranes of the organs that come in contact with the outside environment. According to their morphology, MCs have distinctive cytoplasmic granules that are released during the degranulation process. Additionally, these cells have antimicrobial peptides (AMPs) that fight a variety of infections. Piscidins, hepcidins, defensins, cathelicidins, and histonic peptides are examples of fish AMPs. Confocal microscopy was used in this study to assess Piscidin1 expression in Giant Mudskipper branchial MCs. Our results demonstrated the presence of MCs in the gills is highly positive for Piscidin1. Additionally, colocalized MCs labeled with TLR2/5-HT and Piscidin1/5-HT supported our data. The expression of Piscidin1 in giant mudskipper MCs highlights the involvement of this peptide in the orchestration of teleost immunity, advancing the knowledge of the defense system of this fish.

The fish intestine operates as a complicated interface between the organism and the environment, providing biological and mechanical protections as a result of a viscous layer of mucus released by goblet cells, which serves as a barrier against bacteria, viruses, and other pathogens, and contributes to the functions of the immune system. Therefore, goblet cells have a role in preserving the health of the body by secreting mucus and acting as sentinels. The ancient jawless fish broadgilled hagfish (Eptatretus cirrhatus, Forster, 1801) has a very basic digestive system because it lacks a stomach. By examining the presence, localization, and co-localization of 5-HT, TLR2, iNOS, and Piscidin1, this study intends to provide insight into the potential immune system contributions arranged by the gut goblet cells of broadgilled hagfish. Our results characterize intestinal goblet cells of broadgilled hagfish, for the first time, with the former antibodies, suggesting the hypothesis of conservation of the roles played by these cells also in primitive vertebrates. Moreover, this study deepens the knowledge about the still little-known immune system of hagfish.

Industrialization has resulted in a massive increase in garbage output, which is frequently discharged or stored in waterways like rivers and seas. Due to their toxicity, durability, bioaccumulation, and biomagnification, heavy metals (such as mercury, cadmium, and lead) have been identified as strong biological poisons. Their presence in the aquatic environment has the potential to affect water quality parameters and aquatic life in general. Teleosts’ histopathology provides a sensitive indicator of pollutant-induced stress, because their organs have a central role in the transformation of different active chemical compounds in the aquatic environment. In particular, the gills, kidneys, and liver are placed at the center of toxicological studies. The purpose of this study is to examine the morphological changes caused by heavy metals in the kidney and gills of Boops boops, with a focus on melanomacrophages centers (MMCs) and rodlet cells (RCs) as environmental biomarkers, using histological and histochemical stainings (hematoxylin/eosin, Van Gieson trichrome, Periodic Acid Schiff reaction, and Alcian Blue/PAS 2.5), and immunoperoxidase methods. Our findings show an increase of MMCs and RCs linked to higher exposure to heavy metals, confirming the role of these aggregates and cells as reliable biomarkers of potential aquatic environmental changes reflected in fish fauna. The cytological study of RCs and MMCs could be important in gaining a better understanding of the complicated immune systems of teleosts.

Dendritic cells (DCs) are antigen-presenting cells (APCs) that regulate the beginning of adaptive immune responses. The mechanisms of tolerance to antigens moving through the digestive tract are known to be regulated by intestinal DCs. Agnatha and Gnathostoma are descendants of a common ancestor. The Ostracoderms gave rise to Cyclostomes, whereas the Placoderms gave rise to Chondrichthyes. Sarcopterygii and Actinopterygii are two evolutionary lines of bony fishes. Brachiopterygii and Neopterygii descend from the Actinopterygii. From Neopterygii, Holostei and Teleostei evolved. Using immunohistochemistry with TLR-2, Langerin/CD207, and MHC II, this study aimed to characterize intestinal DCs, from myxines to teleosts. The findings reveal that DCs are positive for the antibodies tested, highlighting the presence of DCs and DC-like cells phylogenetically from myxines, for the first time, to teleosts. These findings may aid in improving the level of knowledge about the immune system’s evolution and these sentinel cells, which are crucial to the body’s defense.

The aryl hydrocarbon receptor (AHR) mediates a variety of biological responses to ubiquitous dioxin and PCB dioxin-like. AHR together with ARNT, AHRR, represent a novel basic helix-loop-helix/PAS family of transcriptional regulators. Their interplay may affect the xenobiotic response. The aim of this study was to investigate, by histological, immunohistochemical investigations and western-blot analysis, the expression of AHR, ARNT and AHRR in liver of seabrem (Spaurus aurata) after exposure at different time to dioxin-like PCB126 in order to deep the knowledge about their specific role. The findings showed a significant increase of AHR and ARNT expression in juvenile fishes after 12 h than control group. The induction of AHR and ARNT is also significant at 24 and 72 hours compared to the control group. Furthemore, induction of AHRR expression has proved to increase both 12 h but this induction does not seem significant to 24 and 72 hours. The most important data of this work is that the induction of AHRR, when the action of the toxic persistence substances, as dioxin and PCB-126, it is not enough to reduce AHR signaling and thus its hyperactivation leads to toxic effects in seabrem (Spaurus aurata). All this confirms the importance of AHR ligands as new class of drugs that can be directed against severe disease such as cancer.

The recognition and elimination of invading pathogens are vital for host survival. Macrophages play a central role in host protection and cells functionally reminiscent of vertebrate macrophages are present in all multicellular organisms. A pattern responsible for bacterial recognition found on the surface of macrophages is CD14. These cells possess a repertoire of antimicrobial molecules stored in their granules and lysosomes. Polarization states observed in mammalian macrophages termed M1 and M2 also likely exist in fish macrophages. Markers for macrophage subtypes are slowly but definitively emerging in fish species. In the present study cell markers such as CD14, acetylcholine, alpha 7 acetylcholine nicotinic receptor (nAChR) subtype, the inducible nitric oxidase synthase (iNOS), and the antimicrobial peptide piscidin 1 are reported for the first time in the intestinal macrophages of both catfish Heteropneustes fossilis (Bloch, 1794) and the African bonytongue Heterotis niloticus (Cuvier, 1829) along the anterior and the posterior axis and the concentric muscle layers. Many antimicrobial effector responses of vertebrate macrophages including respiratory burst and NO induction are similar across the diverse animal taxa. Antibodies against calbindin coupled with ones to VAChT and tubulin revealed the localization of myenteric and submucosal plexuses, which are made up of enteric neurons, glial cells, and nerves near macrophages. Current studies allow for the elucidation of multiple roles of macrophages in disease models providing an insight into their in vivo function in fish.

Gill and air sac of the indian catfish Heteropneustes fossilis harbour a nerve network comprising an innervated system of neuroepithelial endocrine cells; the latter cells are found especially in the gill. A series of antibodies was used for the immunohistochemical detection of neurotransmitters of the neural non-adrenergic, non-cholinergic (NANC) systems such as the sensory neuropeptides (enkephalins), the inhibitory neuropeptide VIP and neuronal nitric oxide synthase (nNOS) responsible for nitric oxide (NO) production which is an inhibitory NANC neurotransmitter. NADPH-diaphorase (NADPH-d) histochemistry was used as marker of nNOS although it is not a specific indicator of constitutively-expressed NOS in gill and air sac tissues. A tyrosine hydroxylase antibody was used to investigate adrenergic innervation. Nitrergic and VIP-positive sensory innervation was found to be shared by gill and air sac. Immunohistochemistry revealed the presence of enkephalins, VIP, NOS and NADPH-d in nerves associated with branchial and air sac vasculature, and in the neuroendocrine cell systems of the gill. Adrenergic nerve fibers were found in some parts of the air sac vasculature. The origin of the nerve fibers remains unclear despite previous findings showing the presence of both NADPH-d and nNOS in the sensory system of the glossopharyngeal and vagus nerves including the branchial structure. Scarce faintly stained nNOS-positive neurons were located in the gill but were never detected in the air sac. These findings lead to the conclusion that a postganglionic innervation of the airways is absent. Mucous goblet cells in the gill were found to express nNOS and those located in the non-respiratory interlamellar areas of the air sac were densely innervated by nNOS-positive and VIP-positive nerve fibers. Our immunohistochemical studies demonstrate that most arteries of the gill and air sac share a NANC (basically nitrergic) innervation which strongly suggests that they are homologous structures.

The innervation of the respiratory tract of amphibians is still poorly understood. Therefore, the respiratory tracts of the frogs Rana esculenta and Discoglossus pictus have been investigated in order to describe non-adrenergic non-cholinergic (NANC) and adrenergic innervation, and the localization of neuromediators that are possibly involved. Immunohistochemical staining of many bioactive substances was found in neuroepithelial cells of the buccopharynx, larynx, lung septa, nerves and neurons throughout the airway system. The findings indicate the occurrence of vasoactive intestinal peptide (VIP)-immunopositive nerve fibers in fibromuscular septa and the vasculature, nitrergic innervation of the large pulmonary veins showing a plexus of nNOS-immunopositive nerve fibers that also innervate the lung wall and the localization of neuronal nitric oxide synthase (nNOS) in neurons in the lung wall. In addition, laryngeal blood vessels and small arteries in the wall of septa that form capillary networks are supplied by enkephalin-immunopositive nerve terminals. We conclude that the airway system of the two frog species studied is innervated by a parasympathetic NANC system. Adrenergic innervation was also found that was immunostained for tyrosine hydroxylase. Adrenergic fibers were mainly present in muscles in septal edges, arteries present in septa and the wall of the lung. It is suggested that nNOS-positive and leu-enkephalin-positive neurons mediate vasodilation via the release of NO, but the nature of the NANC innervation remains obscure. Despite the many pharmacological studies of the lungs of amphibians, the physiological role of pulmonary autonomic innervation remains poorly understood.