ApoE ranks as the strongest genetic risk factor for late-onset Alzheimer’s, but scientists are still unclear about how the lipoprotein affects the brain. A study in the September 28 Journal of Neuroscience describes a unique mouse model that could help clarify. Scientists led by Courtney Lane-Donovan and Joachim Herz at the University of Texas Southwestern Medical Center, Dallas, created a mouse that expresses no ApoE in the brain but a normal amount in the periphery. Synapses faltered and died away in their brains. Surprisingly, these animals dodged memory deficits seen in complete ApoE knockouts. The authors interpret this to mean that while brain ApoE protects synaptic health in the central nervous system, plasma ApoE helps preserve learning and memory. The authors suggest peripheral ApoE may prevent breakdown of the blood-brain barrier or maintain lipid homeostasis that is essential to brain function.
“This is an elegant approach,” said Daniel Michaelson of Tel Aviv University in Israel. “It’s exciting because it suggests that future treatments targeting ApoE in the blood can help the brain.”
Some studies hint that reducing the ApoE may protect against AD. In mice overexpressing human APP, replacing endogenous ApoE with just one copy of human ApoE3 or ApoE4 reduces accumulation of the Aβ peptide (Dec 2011 news on Kim et al., 2011). Similarly, injecting an anti-ApoE antibody into the peritoneum of an AD mouse model reins in amyloid deposition and improves learning and memory (see May 2014 news on Liao et al., 2014). On the other hand, knocking out the ApoE gene altogether in mice degrades synapses and leads to learning impairments (Masliah et al., 1995). Scientists are not clear what causes these learning problems. Plasma lipid levels shoot through the roof in the ApoE knockouts, causing dyslipidemia and atherosclerosis that may affect the brain. If researchers could restore normal ApoE expression in the periphery of these mice, they might be able to separate out the effects of brain and peripheral ApoE on the central nervous system.
Herz and colleagues got a rare opportunity to do just that with a transgenic mouse they inadvertently created. While replacing mouse ApoE with the human version, the transgene randomly integrated into a spot in the genome where the regulatory elements for expression in the liver were active, but those for production in astrocytes were silent. Those are the cells in the periphery and blood, respectively, that produce ApoE. The authors realized what had happened when they analyzed plasma and brain extracts. The mice, which still made murine ApoE, produced human ApoE only in the plasma. By crossing this mouse with a complete ApoE knockout, they aimed to create a model that expressed human ApoE only in the plasma, serving essentially as a brain ApoE knockout (bEKO).
Lane-Donovan first used ApoE immunohistochemistry to confirm the genetic cross worked. While the antibody bound to astrocytes and in the interstitial space between brain cells in ApoE3 targeted replacement mice, none bound in the brain of the ApoE KO mice or to astrocytes in the bEKO mice. The antibody did detect a trace of ApoE in the interstitial space in the bEKO animals. The authors were uncertain why, but think ApoE in the blood may have crossed into the brain through leaks in the blood-brain barrier. To see if lack of astrocytic ApoE affected synapses, the researchers probed sections from the neocortices and hippocampi of seven- to eight-month-old mice with an anti-synaptophysin antibody. Compared with the ApoE3 controls, both bEKO and ApoE KO mice bound much less antibody in the neocortex (see image at left). In the hippocampus, synaptophysin levels were similar among all the mice.
Despite normal synaptophysin in the hippocampus, electrophysiological experiments indicated synaptic plasticity and synapse strength took a hit there in bEKO mice, just as they did in ApoE knockouts. However, in hippocampal slices from ApoE KO mice, the AMPA/NMDA ratio ran lower than in controls, whereas it was normal in the bEKO animals. Maintaining that ratio might explain why the bEKO mice remembered normally, the authors suggested. In the Morris water maze, the bEKO mice spent as much time as controls in the target quadrant in a probe trial, while ApoE KO wasted more time in other quadrants. Interestingly, when the authors parsed the data by gender, they found that the female ApoE KO mice drove the behavioral differences. All males performed equally well, regardless of ApoE status. Michaelson found this gender effect interesting and said that it paved the way for a wider study on how peripheral ApoE affects distinct cognitive parameters such as short- and long-term memory, which are linked to different brain areas and mechanisms.
Taken together, the results suggest that brain and peripheral ApoE have distinct effects on the CNS, the authors wrote. Brain ApoE is important for synapse development, while ApoE in the blood holds further sway over cognitive function, they conclude.
How could peripheral ApoE do that? By maintaining lipid homeostasis, the lipoprotein may help normalize blood flow to the brain, shore up the blood-brain barrier to keep out neurotoxic factors, or corral free fatty acids, which can impair cognitive function in mice (Heverin et al., 2015), the authors suggested (see image below). They could not exclude the possibility that some ApoE might cross into the brain through a leaky blood-brain barrier in the bEKO mice. This could explain the small amount of ApoE detected in the interstitial space, and could account for the memory advantage over complete knockouts. Gregory Cole, University of California, Los Angeles, suggested that restoring plasma ApoE could also benefit the brain by reducing inflammatory signaling in the circulating myeloid cells that then cross the blood-brain barrier.
ApoE’s Potential Roles. Without astrocytic ApoE, fewer synapses mature (top). Without plasma ApoE (bottom right), the blood-brain barrier may leak and toxins could enter the brain. [Courtesy of Lane-Donovan, et al. The Journal of Neuroscience 2016.]
Notably, this study says nothing about the role of the different isoforms of ApoE, Herz told Alzforum. While ApoE2 protects people from AD and ApoE4 puts them at risk for the disease, scientists still debate whether reducing or increasing ApoE4 in the brain would lower disease risk. “This study gives a more informed basis on which to consider such strategies,” said Herz. “Completely abolishing ApoE expression in the brain may not be the best approach, but reducing it, which has been shown to lead to reduced plaque accumulation in mice, may be useful to mitigate pathology.” Whether this will stave off cognitive impairment is still unclear, he said.
“This paper highlights the potential importance of high plasma lipids and how this can negatively impact brain function,” wrote David Holtzman, Washington University School of Medicine in St. Louis, to Alzforum (see full comment below). It would be interesting to assess other synaptic markers as well as the structure of synapses in these mice to see if they are altered, he said.—Gwyneth Dickey Zakaib
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