Biochemical Characterization of HDL/LDL‐like Particles Secreted by Astrocytes

Lipids from the diet or those synthesized in the liver are packaged with proteins into HDL, LDL, and VLDL particles that circulate to places where they are needed. These particles are lipid core vesicles with protein wrapped around the external surface. However, these particles can not cross the blood brain barrier due to their size, so the bulk of neural lipid must be synthesized de novo in the brain. With their extensive processes of dendrites and axons, neurons have a plasma membrane surface several hundred times larger relative to a cell of the same soma size in another tissue and thus have much more lipid to synthesize, manage, and keep healthy. In order to address this biosynthetic need, the brain uses astrocytes to synthesize the bulk of the lipid and then transport it to neurons. Lipid and cholesterol synthesized in astrocytes are packaged into “HDL/LDL‐like” particles along with apolipoproteins and are released to be picked up by adjacent neurons with special receptors that recognize the apolipoproteins on the surface of these particles. Our initial proteomic studies identified ApoE and Apo1A as major components of the astrocyte secretome along with ApoC. Utilizing astrocytes isolated from mice at different ages from 4 to 28 months, we observed that ApoE protein levels secreted from the astrocytes decreased 3.5 fold in the oldest animals. We have hypothesized that the reduction in apolipoprotein production, ApoE in particular, is associated with a decrease in the maintenance of the extensive neuronal plasma membrane which results in enhanced susceptibility of the aged animal for damage to neurons and subsequent degeneration. We have developed an equilibrium sucrose centrifugation protocol to isolate these HDL/LDL‐like particles based on density as well as a more rapid isolation procedure using Cleanascite media to isolate all the lipid containing material and apolipoproteins from the astrocyte conditioned media. We are currently performing quantitative GC‐MS analysis to determine the lipid composition of these particles and if that composition changes with the age of the astrocytes. Western blot analysis is being utilized to determine if the apolipoprotein composition also changes with age of the astrocytes. These studies will clarify the role of lipid turnover/maintenance in the brain as it relates to the progression of neurodegenerative processes. Support or Funding Information McNair Scholars Program, Cowles Endowed Professorship Research Fund, Trinity University