CIRM-Funded Researchers Develop Chimeric “Mighty Mouse” Model to Study Alzheimer’s Disease

Dr. Mathew Blurton-Jones, leader of team that developed the chimeric “Mighty Mouse” model at the University of California, Irvine

In ancient Greek mythology, a Chimera was a creature that was usually depicted as a lion with an additional goat head and a serpent for a tail. Due to the Chimera’s animal hybrid nature, the term “chimeric” came to fruition in the scientific community as a way to describe an organism containing two or more different sets of DNA.

A CIRM-funded study conducted by Dr. Mathew Blurton-Jones and his team at UC Irvine describes a way for human brain immune cells, known as microglia, to grow and function inside mice. Since the mice contain a both human cells and their own mice cells, they are described as being chimeric.

In order to develop this chimeric “mighty mouse” model, Dr. Blurton-Jones and his team generated induced pluripotent stem cells (iPSCs), which have the ability to turn into any kind of cell, from cell samples donated by adult patients. For this study, the researchers converted iPSCs into microglia, a type of immune cell found in the brain, and implanted them into genetically modified mice. After a few months, they found that the implanted cells successfully integrated inside the brains of the mice.

By finding a way to look at human microglia grow and function in real time in an animal model, scientists can further analyze crucial mechanisms contributing to neurological conditions such as Alzheimer’s, Parkinson’s, traumatic brain injury, and stroke.

For this particular study, Dr. Blurton-Jones and his team looked at human microglia in the mouse brain in relation to Alzheimer’s, which could hold clues to better understand and treat the disease. The team did this by introducing amyloid plaques, protein fragments in the brain that accumulate in people with Alzheimer’s, and evaluating how the human microglia responded. They found that the human microglia migrated toward the amyloid plaques and surrounding them, which is what is observed in Alzheimer’s patients.

In a press release, Dr. Blurton-Jones expressed the importance of studying microglia by stating that,

“Microglia are now seen as having a crucial role in the development and progression of Alzheimer’s. The functions of our cells are influenced by which genes are turned on or off. Recent research has identified over 40 different genes with links to Alzheimer’s and the majority of these are switched on in microglia. However, so far we’ve only been able to study human microglia at the end stage of Alzheimer’s in post-mortem tissues or in petri dishes.”

Furthermore, Dr. Blurton-Jones highlighted the importance of looking at human microglia in particular by saying that,

“The human microglia also showed significant genetic differences from the rodent version in their response to the plaques, demonstrating how important it is to study the human form of these cell.”

The full results of this study were published in Cell.

CIRM-funded research is helping unlock the secrets behind “chemo brain”

chemo brain

Every year millions of Americans undergo chemotherapy. The goal of the treatment is to destroy cancer, but along the way more than half of the people treated lose something else. They suffer from something called “chemo brain” which causes problems with thinking and memory. In some cases it can be temporary, lasting a few months. In others it can last years.

Now a CIRM-funded study by researchers at Stanford has found what may be behind chemo brain and identifying potential treatments.

In an article on the Stanford Medicine News Center, senior author Michelle Monje said:

“Cognitive dysfunction after cancer therapy is a real and recognized syndrome. In addition to existing symptomatic therapies — which many patients don’t know about — we are now homing in on potential interventions to promote normalization of the disorders induced by cancer drugs. There’s real hope that we can intervene, induce regeneration and prevent damage in the brain.”

The team first looked at the postmortem brains of children, some of whom had undergone chemotherapy and some who had not. The chemotherapy-treated brains had far fewer oligodendrocyte cells, a kind of cell important in protecting nerve cells in the brain.

Next the team injected methotrexate, a commonly used chemotherapy drug, into mice and then several weeks later compared the brains of those mice to untreated mice. They found that the brains of the treated mice had fewer oligodendrocytes and that the ones they had were in an immature state, suggested the chemo was blocking their development.

The inner changes were also reflected in behavior. The treated mice had slower movement, showed more anxiety, and impaired memory compared to untreated mice; symptoms that persisted for up to six months after the injections.

As if that wasn’t enough, they also found that the chemo affected other cells in the brain, creating a kind of cascade effect that seemed to amplify the impaired memory and other cognitive functions.

However, there is some encouraging news in the study, which is published in the journal Cell. The researchers gave the treated mice a drug to reverse some of the side effects of methotrexate, and that seemed to reduce some of the cognitive problems the mice were having.

Monje says that’s where her research is heading next.

“If we understand the cellular and molecular mechanisms that contribute to cognitive dysfunction after cancer therapy, that will help us develop strategies for effective treatment. It’s an exciting moment.”

 

Stem cell stories that caught our eye: brains, brains and more brains!

This week we bring you three separate stories about the brain. Two are exciting new advances that use stem cells to understand the brain and the third is plain creepy.

Bioengineering better brains. Lab grown mini-brains got an upgrade thanks to a study published this week in Nature Biotechnology. Mini-brains are tiny 3D organs that harbor similar cell types and structures found in the human brain. They are made from pluripotent stem cells cultured in laboratory bioreactors that allow these cells to mature into brain tissue in the span of a month.

The brain organoid technology was first published back in 2013 by Austrian scientists Jürgen Knoblich and Madeline Lancaster. They used mini-brains to study human brain development and a model a birth defect called microcephaly, which causes abnormally small heads in babies. Mini-brains filled a void for scientists desperate for better, more relevant models of human brain development. But the technology had issues with consistency and produced organoids that varied in size, structure and cell type.

Cross-section of a mini-brain. (Madeline Lancaster/MRC-LMB)

Fast forward four years and the same team of scientists has improved upon their original method by adding a bioengineering technique that will generate more consistent mini-brains. Instead of relying on the stem cells to organize themselves into the proper structures in the brain, the team developed a biological scaffold made of microfilaments that guides the growth and development of stem cells into organoids. They called these “engineered cerebral organoids” or enCORs for short.

In a news feature on IMBA, Jürgen Knoblich explained that enCORs are more reproducible and representative of the brain’s architecture, thus making them more effective models for neurological and neurodevelopmental disorders.

“An important hallmark of the bioengineered organoids is their increased surface to volume ratio. Because of their improved tissue architecture, enCORs can allow for the study of a broader array of neurological diseases where neuronal positioning is thought to be affected, including lissencephaly (smooth brain), epilepsy, and even autism and schizophrenia.”

Salk team finds genetic links between brain’s immune cells and neurological disorders. (Todd Dubnicoff)

Dysfunction of brain cells called microglia have been implicated in a wide range of neurologic disorders like Alzheimer’s, Parkinson’s, Huntington’s, autism and schizophrenia. But a detailed examination of these cells has proved difficult because they don’t grow well in lab dishes. And attempts to grow microglia from stem cells is hampered by the fact that the cell type hasn’t been characterized enough for researchers to know how to distinguish it from related cell types found in the blood.

By performing an extensive analysis of microglia gene activity, Salk Institute scientists have now pinpointed genetic links between these cells and neurological disease. These discoveries also demonstrate the importance of the microglia’s environment within the brain to maintain its identity. The study results were reported in Science.

Microglia are important immune cells in the brain. They are related to macrophages which are white blood cells that roam through the body via the circulatory system and gobble up damaged or dying cells as well as foreign invaders. Microglia also perform those duties in the brain and use their eating function to trim away faulty or damage nerve connections.

To study a direct source of microglia, the team worked with neurosurgeons to obtain small samples of brain tissue from patients undergoing surgery for epilepsy, a tumor or stroke. Microglia were isolated from healthy regions of brain tissue that were incidentally removed along with damaged or diseased brain tissue.

Salk and UC San Diego scientists conducted a vast survey of microglia (pictured here), revealing links to neurodegenerative diseases and psychiatric illnesses. (Image: Nicole Coufal)

A portion of the isolated microglia were immediately processed to take a snap shot of gene activity. The researchers found that hundreds of genes in the microglia had much higher activities compared to those same genes in macrophages. But when the microglia were transferred to petri dishes, gene activity in general dropped. In fact, within six hours of tissue collection, the activity of over 2000 genes in the cells had dropped significantly. This result suggests the microglial rely on signals in the brain to stimulate their gene activity and may explain why they don’t grow well once removed from that environment into lab dishes.

Of the hundreds of genes whose activity were boosted in microglia, the researchers tracked down several that were linked to several neurological disorders. Dr. Nicole Coufal summarized these results and their implications in a Salk press release:

“A really high proportion of genes linked to multiple sclerosis, Parkinson’s and schizophrenia are much more highly expressed in microglia than the rest of the brain. That suggests there’s some kind of link between microglia and the diseases.”

Future studies are needed to explain the exact nature of this link. But with these molecular descriptions of microglia gene activity now in hand, the researchers are in a better position to study microglia’s role in disease.

A stem cell trial to bring back the dead, brain-dead that is. A somewhat creepy stem cell story resurfaced in the news this week. A company called Bioquark in Philadelphia is attempting to bring brain-dead patients back to life by injecting adult stem cells into their spinal cords in combination with other treatments that include protein blend injections, electrical nerve stimulation and laser therapy. The hope is that this combination stem cell therapy will generate new neurons that can reestablish lost connections in the brain and bring it back to life.

Abstract image of a neuron. (Dom Smith/STAT)

You might wonder why the company is trying multiple different treatments simultaneously. In a conversation with STAT news, Bioquark CEO Ira Pastor explained,

“It’s our contention that there’s no single magic bullet for this, so to start with a single magic bullet makes no sense. Hence why we have to take a different approach.”

Bioquark is planning to relaunch a clinical trial testing its combination therapy in Latin America sometime this year. The company previously attempted to launch its first trial in India back in April of 2016, but it never got off the ground because it failed to get clearance from India’s Drug Controller General.

STATnews staff writer Kate Sheridan called the trial “controversial” and raised questions about how it would impact patients and their families.

“How do researchers complete trial paperwork when the person participating is, legally, dead? If the person did regain brain activity, what kind of functional abilities would he or she have? Are families getting their hopes up for an incredibly long-shot cure?”

Scientists also have questions mainly about whether this treatment will actually work or is just a shot in the dark. Adding to the uncertainty is the fact that Bioquark has no preclinical evidence that its combination treatment is effective in animal models. The STAT piece details how the treatments have been tested individually for other conditions such as stroke and coma, but not in brain-dead patients. To further complicate things, there is no consensus on how to define brain death in patients, so patient improvements observed during the trial could be unrelated to the treatment.

STAT asked expert doctors in the field whether Bioquark’s strategy was feasible. Orthopedic surgeon Dr. Ed Cooper said that there’s no way electric stimulation would work, pointing out that the technique requires a functioning brain stem which brain-dead patients don’t have. Pediatric surgeon Dr. Charles Cox, who works on a stem cell treatment for traumatic brain injury and is unrelated to Bioquark, commented, “it’s not the absolute craziest thing I’ve ever heard, but I think the probability of that working is next to zero.”

But Pastor seems immune to the skepticism and naysayers.

“I give us a pretty good chance. I just think it’s a matter of putting it all together and getting the right people and the right minds on it.”

Alzheimer’s and the Inflamed Brain: Their Links Run Deeper than Thought

Given that Alzheimer’s disease (AD) is a brain disorder and the leading cause of dementia, it seems logical to assume that some sort of breakdown in the connections of the brain’s nerve cells is mostly to blame.

But based on an increasing volume of research, it turns out that our immune system is also closely linked in a negative way to the disease. Yes, the very same immune system that fights off infections from the bacteria and viruses we come in contact with everyday.

The brain’s local cleaning crew overwhelmed by beta-amyloid and Alzheimer’s
Like a foot soldier keeping watch over the castle for enemy invaders or internal traitors, immune cells called microglia that reside in the brain constantly survey the brain, and literally gobble up any potentially harmful “enemies”. This is true for the potentially harmful protein clumps called beta-amyloid plaques that form in the Alzheimer’s brain. But studies suggest the microglia’s clearance of beta-amyloid plaques gets overwhelmed in Alzheimer’s, shifting the cells’ activity into primarily an inflammation mode and leading to progressive damage to the brain.

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Microglia (green) surround an amyloid plaque (blue) in the brain of an Alzheimer’s disease mouse model. Image: Samuel Marsh/UC Irvine

Microglia (green) surround an amyloid plaque (blue) in the brain of an Alzheimer’s disease mouse model. UCI researchers have shown that endogenous mouse antibodies (red) associate with microglia in the brains of such mice and boost microglia’s ability to degrade plaques.
Samuel Marsh

Microglia represent the localized, first-responder arm of the immune system also called the innate immune system. A second wave, composed namely of immune cells called B cells and T cells, which targets the “enemy” with much more specificity, is initiated outside the brain. To what extent this so-called adaptive immune system is also linked to Alzheimer’s is less understood.

Now, scientists with the Sue & Bill Gross Stem Cell Research Center at the UC Irvine report in  PNAS that they have some answers.

Knocking out B and T cells makes matters worse
Beginning with a strain of mice that mimics Alzheimer’s symptoms with severe beta-amyloid plaque deposits in their brains, the research team bred the animals to also lack B and T cells. Six months later, the results were dramatic. The level of beta-amyloid plaques in AD mice without B and T cells was twice that of AD mice with an intact immune system. As Mathew Blurton-Jones, the team’s lead and UCI assistant professor (he’s also a CIRM-grantee though we did not fund this research), mentioned in a press release on Tuesday, this finding was unexpected:

blurton-jones

Mathew Blurton-Jones, assistant professor of neurobiology & behavior at UCI. Image: Steve Zylius / UCI

“We were very surprised by the magnitude of this effect. We expected the influence of the deficient immune system on Alzheimer’s pathology to be much more subtle.”

So what’s going on here? Why does knocking out the adaptive immune system in AD mice lead to even more beta-amyloid plaques? One hint came from a careful analysis of microglial cells in the brains of the AD mice lacking B and T cells. These cells showed a weakened “eating” activity compared to microglia in AD mice with an intact immune system. This result suggests that without B and T cells, the beta-amyloid plaques are not cleared away by microglial as well.

Additional experiments pointed to the importance of B cells on microglial function. First author Samuel March explained the result in the press release:

“We found that in Alzheimer’s mice with intact immune systems, antibodies – which are made by B-cells – accumulated in the brain and associated with microglia. This, in turn, helped increase the clearance of beta-amyloid.”

Restoring beta-amyloid clean up with a blood stem cell transplant
So, in other words, eliminating B cells eliminates the recruitment of antibodies in the brain which in turn impairs the microglia’s ability to clear away beta-amyloid. To prove this point, the team transplanted healthy blood stem cells into the AD mice lacking B and T cells. The stem cells are capable of restoring all the cell types of the immune system. Sure enough, four months after the transplantation, antibodies were present in the brain near microglia and were associated with a nearly 50% reduction in beta-amyloid plaques.

All together, this data points to Alzheimer’s as a disease of an aging immune system, an idea that Blurton-Jones plans to tackle next:

“We know that the immune system changes with age and becomes less capable of making T- and B-cells. So whether aging of the immune system in humans might contribute to the development of Alzheimer’s is the next big question we want to ask.”

 

CIRM Creativity Student Hanan Sinada’s ‘Extraordinary’ Journey as a Budding Scientist

This summer we’re sponsoring high school interns in stem cell labs throughout California as part of our annual Creativity Program. We asked those students to share their experiences through blog posts and videos.

Today, we hear from Hanan Sinada, who has been busy at the Gladstone Institutes in San Francisco.

Extraordinary. That is the word I would use to describe my time here at Gladstone. This summer I have been an intern at the Gladstone Institute of Neurology, studying microglia. The brain has two main types of cells. Those cells are neurons and glial cells. Glia makes ninety percent of the cells in your brain. Although the word “glia” is derived from the Greek word meaning “glue”, glia cells are more like the support system that surround the neurons in the brain. Many people have not heard of glial cells because they are the dark matter of the brain and not involved in synaptic transition. However, glial cells have many significant functions in the central nervous system (CNS). Their main functions are to supply oxygen and nutrients to the neurons, hold neurons in place, destroy infectious agents, eliminate dead cells, and provide insulation (myelin) to neurons.

Hanan Sinada with her mentor, Gladstone Postdoctoral Researcher Dr. Grietje Krabbe

Hanan Sinada with her mentor, Gladstone Postdoctoral Researcher Dr. Grietje Krabbe

There are three main types of glial cells: microglia, astrocytes, and oligodendrocytes. In my research we focus specifically on microglial cells. Microglia only make up 10-15 percent of the total glia population. Microglia serve as the central nervous system’s macrophages. One function of microglia is to act as antigen presenting cells. Two other roles of the microglia are phagocytosis and cytotoxicity. In cytotoxicity, microglia release cytotoxic substances such as Nitric Oxide (NO) or hydrogen peroxide (H2O2), to damage neurons that have been infected. This leads to cell death. Microglia’s main function is to maintain homeostasis. As a result, microglia are constantly scavenging for apoptotic cells, infectious agents, or any foreign material. Microglia are the main orchestrators of the inflammatory response in the central nervous system (CNS). When an injury occurs in the spinal cord or the brain, microglia release cytokines that cause inflammation in that given area.

In my research we look closely at microglia because they are related to many neurodegenerative diseases, such as Alzheimer’s disease and Parkinson’s disease. My lab started to question about what would happen if we annihilated all the microglia in the brain. Would it decrease the possibility of avoiding the development of those diseases? So we gave wild type mice a drug that depleted all the microglia in the brain, and surprisingly enough the microglia repopulated the brain rapidly after a couple of days. By doing immunohistochemistry and using certain markers, I was able to find where this new microglia-like cell was coming from. From previous studies we already know that this new microglia is not from the periphery. Monocytes cannot cross the blood brain barrier to replace the microglia. We believe that this new microglia is coming from progenitor cell (a type of stem cell). However, we do not know which cell type is giving rise to this new microglia population.

Before starting my internship I did not know that it was going to be the most amazing and interesting learning experience I have ever had in my life. Although every now and then I would have a science crisis, such as having to change antibody because a certain staining would not work, I am so happy and lucky to be doing this cutting edge research. Not only did I learn so much but I am proud to say that I have contributed to the future of science.