Woman side view, memory lapses, forgetting things, degenerative disease. Brain problems. Parkinson and alzheimer desease.
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Investigators at the Keck School of Medicine of USC have published research showing that the APOE4 gene, already widely tied to Alzheimer’s disease (AD) susceptibility, produces molecular changes in the brain’s blood vessels, which are then followed by changes in the synapses between brain cells. The research brings into better focus how the APOE4 gene influences development of the disease, and builds upon prior research showing that the gene has an adverse effect on the blood-brain barrier, the protective layer that protects the brain from toxic substances. But exactly how those changes occurred were not known, until now.

“There was no large-scale comprehensive molecular analysis to understand how the APOE4 gene affects blood vessels in relation to brain functions,” said lead author Berislav Zlokovic, M.D., Ph.D., chair of the Department of Physiology and Neuroscience and director of the Zilkha Neurogenetic Institute at the Keck School of Medicine.

For this study, the results of which are published in the Journal of Experimental Medicine, Keck researchers used knock-in mice whose APOE gene was replaced with humanized APOE gene variants. Studying the brains of these mice at a molecular level, the Keck team showed that APOE4 first caused changes in the cells of the blood-brain barrier that led to problems in synapses, which ultimately caused behavior changes and cognitive dysfunction. This highly detailed molecular analysis of changes in the brain wrought by APOE4 has identified new molecular pathways of AD that are promising therapeutic targets.

“Understanding the order of events—what’s happening first, what’s happening next—is very valuable, because it had not yet been shown how changes in the blood-brain barrier relate to synaptic dysfunction at this molecular level,” Zlokovic said.

A multi-omic approach

In order to get a comprehensive view of what is happening inside individual cells, Zlokovic and his team used both transcriptomic techniques to better understand the activity of RNA molecules, and proteomic studies done in tandem with Marcelo Pablo Coba, Ph.D., associate professor of psychiatry and the behavioral sciences at Keck. The knock-in mice with the APOE4 gene were compared with mice that possessed the APOE3 gene, one that confers a lower risk of AD development.

In the APOE4 mice, the team first detected damage to the endothelial cells that make up the blood-brain barrier and regulate the substances that enter and leave the brain. Among the problems noted was a breakdown of the cytoskeletons which help endothelial cells maintain their shape and structure, as well as irregularities in other molecules that hold cells together and transport cells in and out of the brain. They also observed changes to pericytes, which line the blood-brain barrier to keep it healthy. The pericytes were prone to cell death and damage to the cellular DNA. These changes were a precursor to changes that led to symptoms of the disease.

“We saw this progressive decline in the cells of the blood-brain barrier, at a time when synaptic function was still OK,” Zlokovic said. “But that only lasted for a relatively short period of time.”

Between two to five months after the cellular changes were noted, the researchers observed disruptions in synaptic signaling between brains cells which led to behavior changes in the mice and a degradation in their ability to perform four tasks, one of which was object location recognition, an indication of memory performance.

New drug targets

By identifying a number of different mechanisms of brain damage resulting from the APOE4 variant, the researchers have uncovered promising pathways of AD that might be targeted by therapeutics. One potential approach is to protect the protein networks of the blood brain barrier to help keep it healthier longer.

Zlokovic and his team believe they may have already identified one such substance, a genetically engineered variant of activated protein C, which is already in Phase III clinical trials as a treatment for stroke and has been shown to protect the blood-brain barrier in mice and humans. Another approach suggested by the Keck team is to target dysfunctional TJP1, a gene that controls many of the functions inside endothelial cells.

The study was supported by the National Institutes of Health, the Cure Alzheimer’s Fund, and the Foundation Leducq Transatlantic Network of Excellence for the Study of Perivascular Spaces in Small Vessel Disease. The proteomic and genetic data generated by the work is freely available to the broader research community in the hopes it can generate additional potential treatment approaches.

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