Differently polarized macrophages affect the viability and growth of NSPCs by regulating the expression of PACAP

Objective To explore the influence of differently polarized macrophages, M1 or M2, to viability and growth of NSPCs and its possible mechanism, especially the role of pituitary adenylate cyclase-activating polypeptide (PACAP) in it.

Method: Spinal cord-derived NSPCs were co-cultured with M1 or M2 through a transwell system. Growth of NSPCs in both groups was observed through an inverted microscope within 3days. NSPCs viability of each group, represented as intracellular ATP levels, was measured using the Cellular ATP Kit HTS following co-culture for 24h. PACAP levels in the co-cultured NSPCs was alleviated with immunofluorescence and Western blot analysis.

Results: Morphologically NSPCs demonstrated a long spindle shape with short sprout on 3rd day when cultured together with M2. NSPCs cultured with M1 showed a small circle or oval shape with no obvious sprout. Expression of PACAP was observed in NSPCs co-cultured with M2 through immunofluorescence. In contrast, NSPCs did not demonstrate PACAP staining in the presence of M1 or cultured alone. PACAP in the NSPCs co-cultured with M2 was upregulated significantly compared with that co-cultured with M1 according to Western blot method. The relative ATP level of NSPCs co-cultured with M1 was markedly decreased while that with M2 was elevated significantly. That trend could be relieved by exogenous PACAP or PACAP 6-38. Viability change of NSPCs by M1/M2 correlated with apoptosis.

Conclusion: Differently polarized macrophages could affect the growth and viability of NSPCs by regulating the expression of PACAP.