Crosstalk between Bradykinin and Sphingosine-1-Phosphate Exacerbate Inflammation and Oxidative Stress in Microglia

Abstract

Background: Neurodegenerative diseases (NDs) are a set of disorders that involve the death of neurons in the brain. These disorders are described as progressive and irreversible due to the brain’s limited regenerative capacity and thus causing debilitating consequences to the lives of ND patients. The underlying pathogenesis of NDs is still elusive to this date. Therefore, there is no cure for NDs, and therapies are restricted to symptom relief treatments that can only slow down the progression of these disorders. Bradykinin (BK) is an active peptide in the kallikrein-kinin system found to be involved in mechanisms that are prevalent in both diabetes, inflammation, and stroke in diabetic patients that involves the microglia. Sphingosine-1-phosphate (S1P) is a bioactive sphingolipid that is involved in inflammation through the modulation of microglia activation. However, there is limited knowledge on the involvement of BK and S1P in inflammatory pathways when studied in conjunction, given that both molecules activate G-protein coupled receptors. Aims: The aim of this study was to explore the action of BK and S1P in brain resident macrophages known as microglia. We hypothesize that BK and S1P exacerbate inflammation and oxidative stress through crosstalk signaling cascades. Methods: For the purpose of this study, immortalized murine microglial cells (N9) were stimulated with 0.1 μM BK and/or 1 μM S1P to obtain baseline data regarding inflammatory and oxidative stress markers. Inhibitors of Bradykinin-2-Receptor (B2R) and of Sphingosine Kinase (SK) were utilized to explore the possibility of crosstalk among BK and S1P. The gene expression of inflammatory mediators, oxidative stress markers, BK receptors, and S1P related markers were measured through qPCR to assess for changes in their mRNA levels. Results: Our results show that inflammatory and oxidative stress markers were significantly elevated when N9 cells were stimulated with BK. Moreover, BK was able to increase mRNA levels of Sphingosine Kinases (SK1 and SK2) and S1P receptors, implicating a possibility of crosstalk between BK and S1P. Conclusion: BK stimulated the expression of various inflammatory and oxidative markers in microglial cells. Although BK was also capable of inducing the expression of SK1 and SK2 and S1P receptors, further studies are needed to establish a mechanistic pathway through which BK is inducing this expression.