Proteomic Identification of Novel Plasma Kallikrein Substrates in the Astrocyte Secretome

Abstract

Plasma kallikrein (PK) is activated during hemorrhage and has been implicated in cerebral vascular permeability and edema. To further characterize the potential effects of PK on the brain that may follow cerebral vascular injury, we have utilized a proteomics approach to search for novel PK substrates in the astrocyte secretome. Extracellular proteins released by astrocytes are critical mediators of cerebral homeostasis, including roles in synapse function and vascular integrity. We identified 1,108 proteins in astrocyte condition medium and 295 of these were annotated as secreted proteins. The total abundance of nine proteins was changed after treatment with PK. Characterization of the secreted proteins revealed low molecular weight fragments for 59 proteins in conditioned media exposed to PK that were not observed in untreated controls. The most striking finding from this study was the appearance of fragmentation of 26 extracellular matrix-associated proteins including collagen isoforms 1-6 and11, nidogen-1 and -2, lysyl oxidase-like protein 1, and matrix metalloproteinase 19 in the presence of PK. We also demonstrated that PK induced the fragmentation of non-matrix proteins, including apolipoprotein E. This report further characterizes the astrocyte secretome and identifies novel potential targets of PK-induced proteolysis that may contribute to its effects on the brain following vascular injury.