Structural and functional changes in organs and cells of the immune system in Solanum glaucophyllum intoxicated heifers

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Enzootic calcinosis (EC) is an intoxication of bovine livestock caused by the chronic ingestion of Solanum glaucophyllum (Sg) also known as “Duraznillo blanco” Solanum glaucophyllum is a plant with calcinogenic activity  that contains high levels of glycosideconjugates with 1,25-dihydroxyvitamin D3 or calcitriol Calcitriol has, in addition to its well known functions on mineral metabolism, several effects on growth and differentiation of many cell types and  marked immunomodulatory properties Here we describe the alterations in the morphological and functional characteristics of the organs related to the immune system in Sg-intoxicated cattle. Two - year-old Aberdeen Angus heifers were  dosed with Sg during 15, 30 and 60 days (experimental groups). A group was dose for15 days and sacrificed at 60 days post intoxication (recovery group). Control animals only received tap water during the same period and were sacrificed at 0 and 60 days We ekly control test s were performed to determine EC classical signs. The animals were fed with hay and water ad libitum. Fresh samples as well as frozen sections of subiliac lymph nodes were taken for immunodetection of DC in spots, using rabbit anti-cow monoclonal S -100 protein antibody (DakoCytomation, Carpinteria, CA, USA). The immunohistochemical detection system was a dextran polymer based method (Universal EnVision®System, DakoCytomation), 3-amino-9-ethyl-carbazol was employed as a chromogen while hematoxylin was used for counterstaining. The slides were mounted in aqueous medium Aquamount® (Lerner Laboratories, Pittsburgh, PA, USA). Samples of spleen, thymus and subiliac and prescapular lymph nodes were fixed in buffered formalin paraffin embedded and cut in 5 µm-thick section. The slides were routinely stained for general observations. Morphometric analysis was performed in all processed sections. The lectinhistochemical study, to determinate the carbohydrate expression pattern of tissues, was also performed with 7 biotinylated lectins: PNA, UEA-I, CON-A, SBA, RCA-I, WGA (VectorLab, Burlingame, CA, USA). The reactions were amplified with ABC complex. The 3,3´diaminobenzidine tetrahydrocloride (DAB)(DakoCytomation) was used as a chromogen and hematoxylin used for counterstaining. The intensity of the structures observed was graded using a semiquantitative scale from 0 to 3. The phagocytic activity of the peritoneal macrophages was progressively reduced in the intoxicated animals with time. The main histological findings in lymph nodes were a decrease in the cortex width and a reduction in the lymph nodules and germinal centers (GC) size, the latest presenting lymphocyte and tingible-body macrophages depletion The thymus showed atrophy with reduction of the parenchyma and a relative increase of adipose and connective tissue per unit area. In addition, we also observed a progressive reduction of the cortical region. In the spleen no particular changes were observed in the number of lymphoid follicles nor in their size. Lectinhistochemical studies revealed that normal lymphatic tissues had a great reactivity with PNA PNA bound to T and B dependent areas in spleen and lymph nodes and to the thymic cortex. In Sg-intoxicated animals even though the intensity of staining never decreased in all studied organs, the lectinhistochemical stained area decreased with intoxication time. Recovery group animals showed similar results than those observed in control heifer. We describe some functional and structural modifications of the cell belonging to and associated with the immune system in Sg-intoxicated heifers , that contributeto the negative influence of calcitriol on cellular immune responses. Información suministrada por el agente en SIGEVA