engleski

Immunohistochemical changes in spleen during Klebsiella sepsis

Background: The immunological significance of spleen is already known. It is highly organised organ that functions as a repository for immune cells. It has an important role in response to pathogens because of its natural function to collect antigens from the blood. The parenchyma of the spleen is divided into the white pulp and the red pulp. The white pulp is the lymphatic portion of the spleen. It is well known fact that the patients with splenectomy are susceptible to infections with encapsulated bacteria. Klebsiella pneumoniae (K. pneumoniae) is caracterised as heavily encapsulated bacteria. We decided to monitor the difference of spleen morphology and immunohistochemical changes by CD11b, CD4, CD8 T cells, B220, GR-1 markers expression after intraperitoneal challenge with K. pneumoniae, at different time points. Materials: In our experiments we used male, 8 weeks old BALB/c mice purchased form the facility of Medical Faculty in Rijeka. Mice were infected with extremely virulent strain of K. pneumoniae with 50 CFU per mice, which is also a lethal dose. Methods: In different time point mice were scarified and the spleen was aseptically removed and frozen. Spleen sections (5μ m) were stained with haematoxylin and eosin. Immunohistochemical analysis was preformed using primary monoclonal antibodies for different subpopulation of the cell markers (CD11b, GR-1, CD4, CD8 and B220). Results: Haematoxylin and eosin stain showed the reduction of red pulp interstitial areas as well as enlargement of white pulp especially in later stages of the infection. B220 positive cells showed no significant changes in different time points nor comparing with uninfected controls. Other markers tested showed obvious differences between selected time points as well as compared with controls. The changes are considerable regarding the reduction of CD4 positive T cells, especially 48 hours post infection. Even more pronounced changes were noticed in the ratio of infiltrated GR-1 and CD11b positive cells compared to uninfected animals. The greatest changes occur 24 hours post infection for GR-1 and 48 hours after infection for CD11b positive cells. Conclusions: These changes suggest the immune suppression of infected animals at later stages of experimental infection and probable deleterious influence of increased macrophage infiltration and their products.

Immunohistochemical staining; spleen; Klebsiella pneumoniae

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