The spleen of the African catfish (Clarias gariepinus) was surrounded by connective tissue capsule consisted mainly of collagen fibers. The white pulp was mainly composed of the melanomacrophage center, lymphocytes and surrounding arterial vessels. The red pulp contained sinusoids, capillaries and splenic cords of erythrocytes, in addition to macrophages and lymphocytes (Fig. A+B) and ellipsoid capillary branches.
Using transmission electron microscope, Langerhans-like cells were observed in between the cells of spleen of catfish, they were irregular to ovoid in shape (Fig. C). Their nuclei were ovoid in shape and sometimes lobulated. No desmosomes could be seen between Langerhans-like cells and neighbouring cells (Fig. C). The Langerhans-like cell cytoplasm was electron lucent and characterized by the presence of numerous Birbeck-like granules in several shapes, varying from oval to round, rod to racket shaped granules (Fig. D). In addition to identifying the rough endoplasmic reticulum, free ribosomes and lysosomes within the Langerhans-like cells (Fig. E), three types of Birbeck-like granules were identified. The first Birbeck-like granule type had heterogenic content, the second type identified were cored granules and the third type were racket shaped granules, some of these latter racket shaped granules were enlarged with granular content (Fig. F).
Immunohistochemical staining showed that both the S100 (Fig. G+H) and CD1a (Fig. I-K) antibodies were expressed in the nucleus and cytoplasm of the Langerhans-like cells, especially around blood vessels in the red pulp tissue.
The histological and electron microscope studies undertaken in this investigation showed many morphological features not described previously in the African catfish spleen. It has been previously reported that the spleen of the ray-finned fish (Barbus pectoralis) has a one layer thick capsule comprising of an epithelium of squamous to cuboidal cells with some small round secretory cells, our study has shown that the connective tissue capsule was mainly comprised from collagen fibers in the catfish. It has been also shown that the thin fibrous capsule showed little evidence of contractile ability, our study confirmed that no muscle cells could be recognized in catfish spleen, adding to the theory that contractility is unlikely.
The white pulp within the spleen in our catfish showed a nodular arrangement with melanomacrophage centers, lymphocytes and surrounding arterial vessels. While in the ray-finned fish a lymphatic nodule was described, like that of mammals with germinal nodes, this was absent in the present catfish study. The melanomacrophage centers have been described as ‘groupings of cells containing pigments that are generally found inside the endothelial reticulum of the matrix of hematopoietic cells in teleost fish’ but that have also been shown to be present in the brain, gonads and gills. Our study confirms the existence of these cells within the spleen of catfish and this opens up theories as to why they are present.
Several studies have suggested that these melanomacrophage centers function to destroy, detoxify or recycle endogenous and exogenous substances, which includes material produced by erythrocytes and other cells undergoing metabolic activities. Also it has been shown that melanomacrophage centers respond to infectious agents and other bodies, thus contributing towards immune reactions.
In our study, the African catfish spleen also contained red pulp which contained sinusoids, capillaries and splenic cords of erythrocytes, in addition to macrophages and lymphocytes. This has been observed in other species such as the ray-finned fish whereby the red pulp fills the majority of the spleen and itself contained red blood cell filled sinusoids with trabeculae and lymphatic tissue diffused throughout the red pulp. In addition, also ellipsoid capillary branches, unlike mammals, were found. The ellipsoids are terminations of arterioles running through a sheath of reticular fibers, reticular cells and macrophages and have narrow lumens. It has been shown that these vessels trap blood born substances and filter plasma and therefore play an important role in immunity.
In this study, Langerhans-like cells were observed by transmission electron microscope among the cells of spleen of catfish (Clarias gariepinus), they were irregular to ovoid in shape and no cytoplasmic processes could be observed. This finding was in agreement with published works which also showed that Langerhans-like cells (dendritic cells) did not possess dendritic processes. In contrast, it has been shown that Langerhans-like cells in all Actinopterygii species (ray-finned fish) demonstrated adherent-like junctions and were in close contact with neighbouring cells. These junctions were not observed in the spleen of catfish in the present study. The previous study in ray-finned fish added that, cytoplasmic multivesicular bodies existed within the cells and that the plasma membrane was associated with large aggregates of exosomes, indicating that they could be released/exported from the cell. These characteristics were also absent in catfish in the present study, as the cytoplasmic contents consisted of rough endoplasmic reticulum, free ribosomes and lysosomes, suggesting that there were no secretory functions in Langerhans-like cells in the catfish spleen.
Despite Langerhans cells being relatively specific to epidermal tissues and some mucosal lined organs, these cells were present in healthy spleen/kidney tissues and diseased gills in salmonids. Birbeck-like granules have been observed previously in salmonids within cells resembling Langerhans cells and the earliest vertebrate with an adaptive immune response are fish and therefore would require dendritic cells for antigen presentation. Our study showed that, the cytoplasm of Langerhans-like cells in the catfish spleen was electron lucent and characterized by the presence of numerous Birbeck-like granules exhibiting several shapes, varying from oval, round, rod to racket shaped granules. Previous work in the salmonids has shown that the Birbeck-like granules had a vacuolated portion and a rod section giving an overall racket shape but that fine ultrastructural differences were noted between the species overall. Three types of Birbeck-like like granules were identified in our study, the first type had heterogenic content, the second type were cored granules, the third type were racket shaped granules. Some of the racket shaped granules were enlarged having granular contents. Previous studies have also indicated that Birbeck granules are only found in Langerhans cells and not in other dendritic cells.
Previous studies have shown that Langerhans cells in the skin were positive for both CD1a and S100, dendritic cells can also be positive for both proteins but that interdigitating cells are S100 positive and CD1a negative. Although the morphological observations and presence of Birbeck-like granules indicated that the cells present in the spleen were Langerhans cells, our immunohistochemical observations showing positive staining for CD1a and S100 antibodies further confirmed the presence of Langerhans-like cells in the spleen of African cat fish and showed the locations of the cells throughout the organ. There were also similarities between the ultrastructure of the salmonids and catfish but that some granules were morphologically different between the two species.