Nitric Oxide Synthase Expression in Naturally Infected Sheep Brain with Listeria monocytogenes and Relationship with Cell Death

Mehmet Önder Karayiğit


Background: Nitric oxide is synthesized from L-arginine and catalyzed by a family of NOS. There are three different NOS isoforms: neuronal (nNOS), inducible (iNOS) and endothelial (eNOS). Nitric oxide is an important apoptosis regulator in mammalian system that can induce and prevent apoptosis depending on levels of NO production and environmental conditions of the cell. NOS expression and its relationship with apoptosis has not been well elucidated in listerial meningoencephalitis in sheep. The aim of this study was to investigate eNOS and iNOS expressions in the brain of sheep with natural listeriosis and to compare them with apoptosis which is shaped in the region.

Materials, Methods & Results: In the study, formalin fixed and paraffin embedded brainstem tissue from 25 sheep naturally infected with LM were used from archives. Five μm-thick section was taken from each block. Histopathologically, sections were stained with H&E. Five normal sheep brain tissues were used as control. At the end of the study, Histopathologically in brainstem tissue infected with LM, multifocal microabscesses in different sizes mixed with neutrophils and macrophages were detected and perivascular mononuclear cell infiltration and meningitis characterized by mononuclear cell infiltration were found. All sections were also immunohistochemically stained with LM, eNOS and iNOS antibodies. In addition, TUNEL method was used to determine apoptosis in brain tissues. As a result of immunostaining, listeria immunoreactivity was observed in microabscesses. The Listeria antigens were detected mainly in the cytoplasm of the neutrophils and macrophages and located extracellulary in microabscesses. Both eNOS and iNOS immunoreactivity were observed in very few neurons and glial cells in normal control sheep. Neurons and glial cells in brain tissues of infected animals stained with eNOS and iNOS. But, eNOS and iNOS expressions in listeriosis animals more higher than in control and the this difference was statistically significant (P < 0.05). At the same time, TUNEL immunopositivity was observed mainly in the nuclei of neuron and glial cells and this findings was found to be significantly increased compared to the control group (P < 0.05).

Discussion: The pathogenesis of listerial meningoencephalitis depends on many factors and is still a subject of research. NO is a pluripotent regulator of diverse cellular functions. NOS can trigger cellular damage. However, there are studies suggesting that it can prevent oxidative stress. eNOS and iNOS expressions may vary depending on disease and animal species. Increased levels of NO have been reported to induce apoptosis in many diseases. However, there are studies indicating that NO is able to prevent apoptosis according to the isoform in which it is synthesized. eNOS and iNOS have not been reported together expressions and with their relationship apoptosis in sheep brain with listerial meningoencephalitis. This is the first study of eNOS and iNOS expression and and its relation to cell death in sheep brain infected with LM. Our findings are consistent with previous studies suggesting that NO expression is effective on the pathogenesis of some disease in the central nervous system. However, there are studies that do not match the findings of the present study. This suggests that the role of NO synthesis in the pathogenesis of diseases may vary according to the amount of expression, type of disease and tissue, or animal species. In the present study showed that the expression of both eNOS and iNOS with increased TUNEL positive cells was statistically significant in the listeriosis compared the control brain tissue. These results suggests that eNOS and iNOS can be expressed by diverse brain cells in the pathogenesis of listeriosis in sheep. In addition this, synthesis of NO can induce the apoptosis in sheep brain with listerial meningoencephalitis.

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