Western blot analyses of neuronal lysates and culture medium revealed that human apoE4/4 neurons produce ~35% less full-length intracellular apoE and secrete ~60% less full-length apoE into the medium than human apoE3/3 neurons, both in individual lines (Supplementary Fig. 2a) and as shown by mean values (Fig. 1a–c). Consequently, the ratio of intracellular apoE to secreted apoE was over 20% higher for apoE4 than apoE3, suggesting that apoE4 tends to be retained inside neurons. The ratio of apoE fragments to full-length apoE in neuronal lysates was also significantly higher in apoE4/4 neurons (Fig. 1d,e and Supplementary Fig. 2b). The major apoE fragments were ~12–20 kDa (Fig. 1d), similar in size to those in brains of neuron-specific apoE4 transgenic mice18 and AD patients with an apoE4 genotype19. ApoE4 fragments were undetectable in the culture medium, suggesting that the fragments tend to accumulate inside neurons.
