New Connection Sprouts Between Alzheimer’s Disease and the Immune System

High-resolution confocal images from the hippocampal CA3 region of Alzheimer’s mouse brain show amyloid-beta plaques (green) and microglia/macrophages (red). Mice with complement C3 deficiency show an altered glial response to plaques.

Just as trimming back the branches of an overgrown plant can encourage healthy growth, a little pruning of the connections in the human brain can be a good thing during brain development. But what happens when this natural process goes wrong later in life? Investigators at Brigham and Women’s Hospital have found new clues from preclinical models to indicate that this “synaptic refinement” may play a role in neurodegenerative disease. Their findings, published in Science Translational Medicine, offer new insights into the interplay between the immune system and the development of Alzheimer’s disease.

The new study looks at the role of complement C3 – a molecule involved in the immune response that is elevated in Alzheimer’s disease. Previous studies have shown that C3 helps to trim back the connections between brain cells – known as synapses – during normal brain development. Synapse loss occurs early in Alzheimer’s disease and is associated with cognitive decline. Researchers have not known whether blocking the “complement cascade” – of which C3 is a central part – could protect against impairment and neurodegeneration at later stages of the disease. In the new study, the team examined the effects of C3 deficiency in a mouse model for Alzheimer’s disease. The team found that mice with the engineered C3 deficiency were protected against age-related loss of synapses and brain cells and had fewer markers of inflammation in the brain.

Interestingly, they also find that in aged mice, the telltale amyloid plaques of Alzheimer’s disease remain – and are even more abundant – but cognitive function improved: mice performed better on a learning and memory task, despite the accumulation of plaque in the brain.

“Amyloid plaque deposition occurs years before memory loss in Alzheimer’s disease, but targeting how the immune system responds to these plaques could be an excellent therapeutic approach,” said corresponding author Cynthia Lemere, PhD, of the Ann Romney Center for Neurologic Diseases at BWH. “We think that in later stages of the disease, it’s not necessarily the plaques but the immune system’s response to them that leads to neurodegeneration.”

C3 has also been implicated in other central nervous system conditions, including stroke and macular degeneration. Although the current study is limited by the differences in the immune system and life span of mice and humans, the team’s findings – and clues from previous studies – suggest that modulating complement signaling may represent a potential therapeutic strategy for combating Alzheimer’s disease.

This work was funded by Fidelity Biosciences Research Initiative (F-PRIME), NIH/National Institute on Aging (R21 AG044713), BrightFocus Foundation Fellowship, and Edward R. and Anne G. Lefler Fellowship.

Paper cited: Shi Q et al. “Complement C3 deficiency protects against neurodegeneration in aged plaque-rich APP/PS1 mice” Science Translational Medicine DOI: 10.1126/scitranslmed.aaf6295


An In-Depth Look at IL-33 and a surprising intersection with PGE2

Interleukin 33 (IL-33) is a member of the IL-1 family and plays a role in various inflammatory diseases. IL-33 is found in cells at barrier sites such as the skin, lung and intestine, but can also appear in macrophages and dendritic cells, contributing to inflammatory conditions such as sepsis, cardiovascular disease and allergy. Despite its role in these disease states, little is known about the mechanisms that govern when, where and how IL-33 becomes active.

In a new study published in The Journal of Biological Chemistry, BWH investigators validate a critical link between PGE2 (an inflammatory lipid mediator that is suppressed by commonly prescribed cyclooxygenase inhibitors) signaling in macrophages with production and amplification of IL-33.

“The amplification mechanisms that we report may play an important role in sepsis, viral infection and other circumstances in which PGE2 and IL-33 play roles in orchestrating innate immunologic states,” said lead author Sachin Kumar Samuchiwal, PhD, of the Division of Rheumatology, Immunology and Allergy at BWH.

The researchers induced IL-33 protein and mRNA expression, identifying contributions from PGE2 by comparing the responses of wild-type cells to those derived from mice lacking the synthase enzyme for lipid mediator or its receptors. In cells without endogenous PGE2, the team found reduced mRNA and protein levels for IL-33. Their findings suggest that PGE2 may be crucial for IL-33 driven disease states, and suggest a mechanism by which cyclooxygenase inhibitors may impact these diseases.

Paper cited: Samuchiwal SK et al. “Endogenous prostaglandin E2 amplifies IL-33 production by macrophages through an E prostanoid (EP)2/EP4-cAMP-EPAC dependent pathway” J. Biol Chem DOI: 10.1074/jbc.M116.769422


I­­­dentifying Notch Binding Sites and Their Relation to Leukemia Cells

A pair of Notch transcription complexes binds to a sequence-paired site. Researchers are learning more about these binding sites and their role in embryonic development and human disease. Image courtesy of the Blacklow lab

The Notch signaling pathway is a highly-conserved mechanism that helps control gene expression in multi-cellular animals. Notch signaling is finely regulated and, when working properly, it is vital for brain development, cell communication and cell differentiation during the development of an embryo.  However, in certain human diseases such as cancer and cardiovascular disease, the Notch pathway goes awry.

To help control and fine-tune which genes are transcribed, Notch transcription complexes (NTCs) attach to certain binding areas on target genes. They bind to either monomeric sites (which require a single NTC to bind), or sequence-paired sites (SPSs, which bind with NTC-dimers). While monomeric sites have previously been studied and identified, SPSs are less understood.

In a new study published in Science Signaling, work led by researchers at BWH and Harvard Medical School identified new information regarding SPSs that shed light on their function and how to identify them. Previous studies suggest that SPS-regulated genes may have stronger and longer responses to Notch compared to monomeric-regulated genes, but understanding SPS responses is difficult due to their structural diversity and variation in response depending on the cell-environment. The researchers developed a method to identify SPSs within genome-wide data sets using a chemical test called a FRET assay. They found that SPSs mainly exist in long-range enhancers on DNA and they are present in about a third of Notch target genes. The experiment also showed that there is diversity in SPS DNA sequences.

The researchers also looked at the prevalence of SPSs in T-cell acute lymphoblastic leukemia (T-ALL) cells. Previous research by Jon Aster, MD, PhD, of BWH Department of Pathology and a corresponding author of the study, found that in certain forms of leukemia, such as T-ALL, mutations in the Notch receptors increase Notch signaling and drive growth of the cancerous cells. In this study, the authors discovered that 15-20 percent of the binding sites in T-ALL cells are SPSs, highlighting the role SPSs play in these kinds of cells.

“These observations greatly expand the known role of SPSs in mammalian gene regulation and reveal the presence of diverse SPS architectures that contribute to the regulation of Notch target genes,” said Aster, a co-corresponding author of the study. “There is still much to be discovered about SPSs, but the ability to identify them in the genome is a significant stepping stone toward understanding how they function and the exact role they play in regulating gene expression in both healthy and diseased cells.”

“This work advances fundamental understanding about how Notch proteins act to stimulate expression of genes that control cell identity and cell proliferation in both normal and disease states,” said co-corresponding author Stephen Blacklow, MD, PhD.

Funding for this work was made possible by the National Institutes of Health and the Leukemia and Lymphoma Society.

Paper cited: Severson, E et al. “Genome-wide identification and characterization of Notch transcription complex–binding sequence-paired sites in leukemia cells.”  Sci. Signal. DOI: 10.1126/scisignal.aag1598


A Virus Visualized

B-cells, shown here, are targeted by the Epstein-Barr virus for degradation.

The Epstein-Barr virus, most commonly known for causing mononucleosis but also a contributor to several forms of cancer, has many mysterious properties. The virus is an expert at replication –more than 95 percent of adults worldwide carry the virus, which is easily transmitted and can persist in a human host decades after a person is initially infected. However, for most of its life cycle, the

Epstein-Barr virus is in a “latent” state, hibernating in a host’s B-cells. Little has been known about its replication phase, in part due to technical constraints. Using a newly developed technique, BWH investigators have, for the first time, visualized how the EBV replication cycle remodels the cell. Their results are published in Cell Reports.

The team used FACS (fluorescence-activated cell sorting) to sort for cells, triggered the lytic cycle (through which cells infected with EBV are destroyed) and then used sensitive mass spectrometry to quantify profiles of more than 8,000 cellular proteins and 69 EBV proteins.

There is significant interest in characterizing the EBV replication cycle because it is required for persistent host colonization, for spread to the next host and has an important connection to cancer. EBV is associated with 200,000 cancers worldwide annually – the virus’s genome can be detected in EBV-associated cancers, and there are indications that drugs that target the EBV replication cycle may be useful in killing cancer cells.

There are 2 EBV strains (type 1 and II), and the researchers studied lytic replication by both.  They then cross-compared EBV replication effects on the host cell proteome with those of a related human herpesvirus, cytomegalovirus (CMV) and with a key immune-modulator of Kaposi sarcoma associated herpesvirus (KSHV). This cross-comparison highlights unique and shared aspects of the replication cycles of these divergent, medically-relevant human herpesviruses.

“Our study serves as the first resource of the EBV lytic cycle effects on the host proteome, and on the infected cell plasma membrane,” said corresponding author Benjamin Gewurz, Division of Infectious Diseases at BWH. “The data generates hypotheses for future studies, and will be of interest to immunologists and those in the infectious disease field.”

Paper cited: Ersing I et al. “A Temporal Proteomic Map of Epstein-Barr Virus Lytic Replication in B Cells” Cell Reports DOI:10.1016/j.celrep.2017.04.062