Soluble ANPEP Released From Human Astrocytes as a Positive Regulator of Microglial Activation and Neuroinflammation: Brain Renin- Angiotensin System in Astrocyte-Microglia Crosstalk

Abstract

Astrocytes are major supportive glia and immune modu-lators in the brain; they are highly secretory in nature and interact with other cell types via their secreted proteomes. To understand how astrocytes communicate during neu-roinflammation, we profiled the secretome of human as-trocytes following stimulation with proinflammatory factors. A total of 149 proteins were significantly upregu-lated in stimulated astrocytes, and a bioinformatics anal-ysis of the astrocyte secretome revealed that the brain renin-angiotensin system (RAS) is an important mecha-nism of astrocyte communication. We observed that the levels of soluble form of aminopeptidase N (sANPEP), an RAS component that converts angiotensin (Ang) III to Ang IV in a neuroinflammatory milieu, significantly increased in the astrocyte secretome. To elucidate the role of sANPEP and Ang IV in neuroinflammation, we first evaluated the expression of Ang IV receptors in human glial cells because Ang IV mediates biological effects through its receptors. The expression of angiotensin type 1 receptor was considerably upregulated in activated human micro-glial cells but not in human astrocytes. Moreover, inter-leukin-1 beta release from human microglial cells was synergistically increased by cotreatment with sANPEP and its substrate, Ang III, suggesting the proinflammatory ac-tion of Ang IV generated by sANPEP. In a mouse neuro-inflammation model, brain microglial activation and proinflammatory cytokine expression levels were increased by intracerebroventricular injection of sANPEP and attenuated by an enzymatic inhibitor and neutralizing antibody against sANPEP. Collectively, our results indicate that astrocytic sANPEP-induced increase in Ang IV exacerbates neuroinflammation by interacting with microglial proinflammatory receptor angiotensin type 1 receptor, highlighting an important role of indirect cross-talk between astrocytes and microglia through the brain RAS in neuroinflammation.