Localization of iodopsin and rod-opsin immunoreactivity in the retina and pineal complex of the river lamprey,Lampetra japonica

Melano-macrophages in the head-kidney, spleen and liver of sea bass and gilthead seabream have been investigated by means of light and electron microscopy, histochemistry and phagocytic assays. The results demonstrate the presence of both free and clustered melano-macrophages (melano-macrophage centres), with similar ultrastructural features. These large cells are PAS-, hemosiderin-and melanin-positive, and contain large, alkaline-and acid phosphatase-positive lysosomes, whose reaction intensity depends on the amount of accumulated pigment. The relationship between the cytochemical features of these lysosomes and the capacity of the melano-macrophages to phagocytose bacteria and latex beads, has been studied. The large melanomacrophage centres have a capsule of flattened cells, whose ultrastructural and cytochemical features are similar to fibroblast-like reticular cells. Melanin is the main accumulated pigment. A subpopulation of head-kidney mononuclear phagocytes engulfs melanin associated with cell debris. The relationship between the origin of the melano-macrophage pigment and the ability of monocytes/macrophages to phagocytise the melanin from melanocytes, is considered. The origin and possible function of melano-macrophage centres are discussed.     

A quantitative and histochemical study on melano-macrophage centres in the spleen of the teleost fish Diplodus annularis L.

The incidence and histochemical features of melano-macrophage centres in the spleen of the teleost fish D. annularis were studied with respect to starved and fed fish. Tissue catabolism following complete starvation appears to induce formation of new centres containing melanin and lipofuscin pigments. However, a change in the amount of melano-macrophages was also recorded in fed fish, suggesting that some other factor(s), internal to fish, may affect the presence of such structures within the spleen of D. annularis .     

Structural characterization of the melano-macrophage centres (MMC) of goldfish Carassius auratus

In the present work we have studied the organization of melano-macrophage centres (MMCs) in the peripheral lymphoid organs, including spleen, pro- and mesonephros, of the goldfish, Carassius auratus, in an attempt to clarify their cellular composition, origins and functional relationships. Histological analysis demonstrated a similar organization in the three organs on the basis of closely packed phagocytic cells containing abundant pigment. The MMCs of Carassius auratus are found throughout the parenchyma of spleen and kidney and show a close association with the vascular system, i.e. splenic ellipsoids, sinusoids of red pulp and renal blood sinuses. They exhibit distinct degree of development from small groups of actively phagocytic macrophages to large, totally or partially encapsulated centres, where effete phagocytic cells are filled by cell debris. Ultrastructural and histochemical data suggest that the main inclusion observed in the MMCs of Carassius auratus is lipofuscin. Haemosiderin occurs in lesser amounts and melanin is almost restricted to kidney MMCs,--mainly mesonephros--. Our results suggest various non-specific physiological roles for the teleost MMCs, including tissue breakdown and erythrocyte catabolism.     

Detection of a Fine Lamellar Gridwork After Degradation of Ocular Melanin Granules by Cultured Peritoneal Macrophages

In the present study we investigated by electron microscopy whether melanin granules derived from choroidal melanocytes and retinal pigment epithelium of cattle could be degraded in the phagolysosomes of cultured murine macrophages. It was found that degradation of ocular melanin is possible by the lysosomes of these macrophages. During degradation of the melanin granules an internal gridwork of fine concentric, highly ordered membranes, 3-4 nm thick, became visible. These membranes may represent remnants of the melanin polymer in the original melanosome or may result from self-assembly of degradation products. Early-stage melanosome-like structures also appeared during digestion of these melanin granules. Melanin granules that seemed to break down into smaller fragments without any visible internal structure were also observed.     

Evidence for melano-macrophage centres of teleost as evolutionary precursors of germinal centres of higher vertebrates: an immunohistochemical study

The melano-macrophage centres (MMCs) of the haemolymphopoietic organs of teleost fish trap and retain antigens and are closely associated with immunoglobulin-secreting cells. The hypothesis that they are the phylogenetic precursors of the germinal centres of higher vertebrates has been questioned due to their apparent lack of organising cells. In this study the immunoreactivity of MMC cells from spleen and kidney of the teleosts Cyprinus carpio, Odontesthes bonariensis and Solea senegalensis to CNA-42, an antibody usually employed for labelling follicular dendritic cells of higher vertebrates was investigated. Free melano-macrophages and MMCs in the spleens of all three species were labelled by the antibody. This finding adds new evidence to the hypothesis that an evolutionary relationship exists between the MMCs of fish and the germinal centres of many birds and mammals.     

Microstructure of cell wall-associated melanin in the human pathogenic fungus Cryptococcus neoformans

Melanin is a virulence factor for many pathogenic fungal species, including Cryptococcus neoformans. Melanin is deposited in the cell wall, and melanin isolated from this fungus retains the shape of the cells, resulting in hollow spheres called "ghosts". In this study, atomic force, scanning electron, and transmission electron microscopy revealed that melanin ghosts are covered with roughly spherical granular particles approximately 40-130 nm in diameter, and that the melanin is arranged in multiple concentric layers. Nuclear magnetic resonance cryoporometry indicated melanin ghosts contain pores with diameters between 1 and 4 nm, in addition to a small number of pores with diameters near 30 nm. Binding of the antibodies to melanin reduced the apparent measured volume of these pores, suggesting a mechanism for their antifungal effect. We propose a model of cryptococcal melanin structure whereby the melanin granules are held together in layers. This structural model has implications for cell division, cell wall remodeling, and antifungal drug discovery.     

The effect of melanin on iron associated decomposition of hydrogen peroxide

The effects of melanin on the iron-catalyzed decomposition of hydrogen peroxide to hydroxyl radicals and hydroxyl ions have been studied using electron spin resonance, spin trapping and visible light spectrophotometry. Melanin altered these reactions by several different mechanisms and consequently, depending on conditions, can significantly increase or decrease the yield of reactive products, including hydroxyl radicals. For low concentrations of ferrous ions, melanin decreased the yield of hydroxyl radicals due to binding of ferrous ions by melanin; ferrous ions bound to melanin did not decompose H2O2 efficiently. Melanins increased the rate of hydroxyl radical production if the predominant form of iron was ferric, due to the ability of melanin to reduce ferric to ferrous iron. Hydroxyl radical production in the presence of a strong chelator (e.g. EDTA) and melanin was greater than in the presence of a weak chelator (e.g. ADP) and melanin. Melanin also increased the rate of destruction of the DMPO-OH adduct.     

The radioprotective properties of fungal melanin are a function of its chemical composition, stable radical presence and spatial arrangement

Melanized microorganisms are often found in environments with very high background radiation levels such as in nuclear reactor cooling pools and the destroyed reactor in Chernobyl. These findings and the laboratory observations of the resistance of melanized fungi to ionizing radiation suggest a role for this pigment in radioprotection. We hypothesized that the radioprotective properties of melanin in microorganisms result from a combination of physical shielding and quenching of cytotoxic free radicals. We have investigated the radioprotective properties of melanin by subjecting the human pathogenic fungi Cryptococcus neoformans and Histoplasma capsulatum in their melanized and non-melanized forms to sublethal and lethal doses of radiation of up to 8 kGy. The contribution of chemical composition, free radical presence, spatial arrangement, and Compton scattering to the radioprotective properties of melanin was investigated by high-performance liquid chromatography, electron spin resonance, transmission electron microscopy, and autoradiographic techniques. Melanin protected fungi against ionizing radiation and its radioprotective properties were a function of its chemical composition, free radical quenching, and spherical spatial arrangement.     

Role of Melanin as a Scavenger of Active Oxygen Species

The protective role of melanin, either synthetic or derived from a metastatic lung melanoma nodule, was studied in terms of its ability to interact with active oxygen species (O₂, H₂O₂, RO, ROO, etc.). Both melanins showed the ability to react with O₂. The superoxide dismutase-like activity corresponds to 21 and 10 U/mg for synthetic and tumor melanin, respectively. The latter value accounts for about 8% of the superoxide dismutase activity of cultured melanoma cells. Neither type of melanin showed catalase-like or glutathione peroxidase-like activity. Both types of melanin reacted with RO and ROO radicals as determined by inhibition of the lipid peroxidation reaction of rat liver homogenates. The spontaneous lipid peroxidation of rat liver homogenate was inhibited up to 90% and 80% by synthetic and tumor melanin with half-maximal effects at 2.5 and 5.5 μg melanin/ml, respectively. The 2,2-azobis-(2 amidino propane) (AAPH)-initiated lipid peroxidation of rat liver homogenate was inhibited up to 3% and 20% by synthetic and tumor melanin, with half maximal effect at 120 and 500 μg melanin/ml, respectively. Both types of melanin were able to protect the in vitro inactivation of glucose oxidase, which occurs in the presence of AAPH-generated radicals.     

Physico-Chemical Evaluation of Rationally Designed Melanins as Novel Nature-Inspired Radioprotectors

Background

Melanin, a high-molecular weight pigment that is ubiquitous in nature, protects melanized microorganisms against high doses of ionizing radiation. However, the physics of melanin interaction with ionizing radiation is unknown.

Methodology/Principal Findings

We rationally designed melanins from either 5-S-cysteinyl-DOPA, L-cysteine/L-DOPA, or L-DOPA with diverse structures as shown by elemental analysis and HPLC. Sulfur-containing melanins had higher predicted attenuation coefficients than non-sulfur-containing melanins. All synthetic melanins displayed strong electron paramagnetic resonance (2.14·10¹⁸, 7.09·10¹⁸, and 9.05·10¹⁷ spins/g, respectively), with sulfur-containing melanins demonstrating more complex spectra and higher numbers of stable free radicals. There was no change in the quality or quantity of the stable free radicals after high-dose (30,000 cGy), high-energy (¹³⁷Cs, 661.6 keV) irradiation, indicating a high degree of radical stability as well as a robust resistance to the ionizing effects of gamma irradiation. The rationally designed melanins protected mammalian cells against ionizing radiation of different energies.

Conclusions/Significance

We propose that due to melanin''s numerous aromatic oligomers containing multiple π-electron system, a generated Compton recoil electron gradually loses energy while passing through the pigment, until its energy is sufficiently low that it can be trapped by stable free radicals present in the pigment. Controlled dissipation of high-energy recoil electrons by melanin prevents secondary ionizations and the generation of damaging free radical species.     

The effects of splenectomy and subsequent starvation on the storage of haemosiderin by the melano-macrophages of rainbow trout Salmo gairdneri Richardson

In clinically normal, cachectic and diseased teleost fish, haemosiderin is observed mostly in the melano-macrophage centres of the spleen and to a much lesser extent in the centres of the kidney and liver. When rainbow trout that had been splenectomised were starved, accumulation of haemosiderin was diverted to the kidney melano-macrophages.     

Interaction of melanin with carbon- and oxygen-centered radicals from methanol and ethanol

The interaction of dopa-melanin (DM) and cysteinyldopa-melanin (CDM) with carbon- and oxygen-centered radicals generated by benzophenone-photosensitized hydrogen abstraction from ethanol, or by pulse radiolysis of aqueous solutions of methanol and ethanol, is reported. Photosensitized formation of carbon-centered radicals and their interaction with melanin was monitored by electron paramagnetic resonance (EPR) spin trapping using DMPO, and via the melanin free radical signal itself. In the pulse radiolysis experiments, the interaction of DM or CDM with hydroxymethyl, hydroxyethyl, and the corresponding methanol peroxyl radical was monitored by recording time-dependent changes of the melanin absorbance at selected wavelengths. The data indicate that both melanins are good scavengers of carbon-centered radicals, with corresponding rate constants in the range of 10⁷ to 10⁸ M⁻¹ s⁻¹. Significantly, compared to DM, CDM is also an exceptionally efficient scavenger of oxygen-centered radicals derived from methanol with corresponding rate constants of 2.7 × 10⁴ and 2 × 10⁶, M⁻¹ s⁻¹ for DM and CDM, respectively. The results are discussed with reference to the potential role of melanin in protecting the integrity of melanosomes by inhibiting peroxidation of lipid components of the organelle membrane.     

Scrota1 melanins in bats (Chiroptera): description, distribution and function

Several species of bats contain pigment granules within the scrotal skin, tunica vaginalis, or tunica albuginea surrounding the testis and/or epididymis. Seventy-two species of bats, representing 49 genera were examined for the presence of such pigmentation. Histological, chemical and spectrophotometric tests were performed and confirmed the pigment as melanin. Melanin was found only in the families Pteropidae, Megadermatidae, Myzopodidae and Vespertilionidae. A strong correlation exists between scrotal pigmentation and roosting in locations where the bats are exposed to solar radiation. Melanin pigmentation in the scrotal region appears to be an adaption protecting male germinal tissue from the harmful effects of ultraviolet radiation. In one species, Laoiu frons , melanin deposited within the scrotal skin appears to have a social/reproductive communication function.     

Melanin both causes and prevents oxidative base damage in DNA: quantification by anti-thymine glycol antibody.

The present study employs immunological methods to measure modified bases in DNA. A polyclonal antibody specific for thymine glycol was used to quantify the level of thymine glycol in calf thymus DNA gamma-irradiated in solutions containing varying concentrations of DOPA-eumelanin. Melanin decreased the number of thymine glycols produced by 200 Gy at low melanin concentrations. At high melanin concentrations, the number of thymine glycols increased. Thymine glycol was also produced in unirradiated DNA-eumelanin mixtures. DOPA-eumelanin was found to produce single-strand breaks in supercoiled phi X174 RF DNA. The breaks were measured by conversion of form I to form II as detected by agarose gel electrophoresis. The level of damage produced by melanin could be modulated by agents known either to stabilize or to scavenge active oxygen species. These studies demonstrate that melanin can both scavenge and generate active free radicals.     

The effect of melanins on oxidation of lecithin in liposomal membranes

Lecithin peroxidation in liposomal membranes induced by UV light was studied in the presence of natural eye melanin and synthetic melanins prepared from various precursors. It was shown that melanins inhibited lecithin photooxidation, and that the extent of this effect strongly depended on the type and concentration of melanin. Comparative study indicated that melanin obtained from adrenolutin was the most effective antioxidant. The ability to inhibit lipid peroxidation depends both on the concentration of paramagnetic centers in the melanin polymer and the accessibility of these centers for free radicals formed during irradiation of liposomes.     

Ligula intestinalis (L.) (Cestoda: Pseudophyllidea): plerocercoid-induced changes in the spleen and pronephros of roach, Rutilus rutilus (L.), and gudgeon, Gobio gobio (L.)

The effects of the plerocercoid of Ligula intestinalis (L.) (Cestoda: Pseudophyllidea) on the major lymphoid organs, the spleen and pronephros, of roach, Rutilus rutilus (L.), and gudgeon, Gobio gobio (L.), are described. The spleen of ligulosed roach showed a significant decrease in weight. Differential cell counts suggested this was due to a reduction in erythrocytes, despite significant increases in macrophages and vacuolated granulocytes. The spleen of gudgeon, which consisted almost entirely of erythrocytes, showed a slight reduction in weight in ligulosed fish. In both roach and gudgeon this decrease was independent of parasite burden. Differential cell counts of the pronephros from ligulosed roach revealed a significant decrease in neutrophils and increase in vacuolated granulocytes. In the pronephros of gudgeon, however, cell counts were unaffected by ligulosis. Ultrastructural observations included an apparent disintegration of vacuolated granulocytes and increased pinocytic activity in specialized endothelial cells in the spleens of ligulosed roach. Also, melano-macrophage centres and melano-macrophages increased in the spleen of ligulosed roach and gudgeon, respectively. The marked changes in spleen weight and differential cell counts in ligulosed roach and lack of such changes in ligulosed gudgeon correlate with the differential response to the parasite by these two fish species.     

Defence mechanisms in fish

The lympho-reticular tissues in the plaice were investigated for their phagocytic properties on colloidal carbon after its intraperitoneal injection. Fish were killed at intervals ranging from 10 min to 25 days after injection. Although peritoneal macrophages constituted a large population of phagocytic cells, most of the carbon apparently gained access to the circulation as free particles and phagocytosis was performed predominantly by the ellipsoids of the spleen, the network of reticulo-endothelial (RE) cells throughout the haemopoietic tissue of the kidney, and by the RE cells occupying intermuscular spaces in the atrium of the heart. The cardiac macrophages rapidly emigrated from the organ while the carbon containing macrophages in the kidney and spleen formed aggregates in the lymphoid areas, either within or outwith pre-existing aggregates of melano-macrophages.
The possible significance of phagocyte aggregations, including melano-macrophages, in association with lymphoid elements in the kidney and spleen is discussed in the context of immune mechanisms.     

The role of free radicals in asbestos-induced diseases.

Asbestos exposure causes pulmonary fibrosis and malignant neoplasms by mechanisms that remain uncertain. In this review, we explore the evidence supporting the hypothesis that free radicals and other reactive oxygen species (ROS) are an important mechanism by which asbestos mediates tissue damage. There appears to be at least two principal mechanisms by which asbestos can induce ROS production; one operates in cell-free systems and the other involves mediation by phagocytic cells. Asbestos and other synthetic mineral fibers can generate free radicals in cell-free systems containing atmospheric oxygen. In particular, the hydroxyl radical often appears to be involved, and the iron content of the fibers has an important role in the generation of this reactive radical. However, asbestos also appears to catalyze electron transfer reactions that do not require iron. Iron chelators either inhibit or augment asbestos-catalyzed generation of the hydroxyl radical and/or pathological changes, depending on the chelator and the nature of the asbestos sample used. The second principal mechanism for asbestos-induced ROS generation involves the activation of phagocytic cells. A variety of mineral fibers have been shown to augment the release of reactive oxygen intermediates from phagocytic cells such as neutrophils and alveolar macrophages. The molecular mechanisms involved are unclear but may involve incomplete phagocytosis with subsequent oxidant release, stimulation of the phospholipase C pathway, and/or IgG-fragment receptor activation. Reactive oxygen species are important mediators of asbestos-induced toxicity to a number of pulmonary cells including alveolar macrophages, epithelial cells, mesothelial cells, and endothelial cells. Reactive oxygen species may contribute to the well-known synergistic effects of asbestos and cigarette smoke on the lung, and the reasons for this synergy are discussed. We conclude that there is strong evidence supporting the premise that reactive oxygen species and/or free radicals contribute to asbestos-induced and cigarette smoke/asbestos-induced lung injury and that strategies aimed at reducing the oxidant stress on pulmonary cells may attenuate the deleterious effects of asbestos.     

Melanin: structure, biosynthesis, biological functions

Melanins are the group of natural black pigments. The structure of melanin macromolecules is irregular network arising from phenolic precursors in consequence of enzymatic and autooxidation. Melanin is stable polyradical, contain some semiquinone radicals and accumulate the exogene radicals and other active oxygen species, heavy metals, electrophyl toxic compounds. Some this properties determine the antioxidant, antitoxic, antiradiation and antitumour activity of melanins. On the base of natural melanins is possible creation of some effective prophylactic and curative preparations.     

Effects of sublethal concentrations of potassium dichromate on the occurrence of splenic melano-macrophage centres in juvenile plaice, Pleuronectes platessa, L.

Plaice were exposed to 0.5 and 2.0 ppm potassium dichromate (K₂Cr₂O₇) concentrations for 27 days. In both water concentrations chromium was accumulated up to a level of 400 ppb in the carcass of the fish. At the same time the number of splenic melano-macrophage centres (MMC) tripled, whereas their average size was reduced to one third. The percentage area of MMC in the splenic tissue remained unchanged. Histological alterations following exposure to K₂Cr₂O₇ included increases both in melanin content of the MMC and in the number of melanin-containing cells within blood vessels; also, the splenic ellipsoids became less clearly demarcated. Splenic melano-macrophages show a decreased ability to aggregate to form large MMC under the influence of K₂Cr₂O₇, which leads to the assumption that the turnover rate of macrophage settlement in, and macrophage elimination from, the spleen is increased.