Now, working with mice, researchers at Johns Hopkins Medicine and the University of Lausanne in Switzerland have identified that missing link — an immune system cell that they say travels from the gut to the brain and attacks cells rather than protect them as it normally does.

The team’s findings are published Jan. 6, 2021, in the journal Science Translational Medicine.

Seen in as many as 12% of infants weighing less than 3.5 pounds at birth, NEC is a rapidly progressing gastrointestinal emergency in which bacteria invade the wall of the colon and cause inflammation that can ultimately destroy healthy tissue at the site. If enough cells become necrotic (die) so that a hole is created in the intestinal wall, bacteria can enter the bloodstream and cause life-threatening sepsis.

In a 2018 mouse study, researchers at Johns Hopkins Medicine and the Fred Hutchinson Cancer Research Center found that animals with NEC make a protein called toll-like receptor 4 (TLR4) that binds to bacteria in the gut and precipitates the intestinal destruction. They also determined that TLR4 simultaneously activates immune cells in the brain known as microglia, leading to white matter loss, brain injury and diminished cognitive function. What wasn’t clear was how the two are connected.

For this latest study, the researchers speculated that CD4+ T lymphocytes — immune system cells also known as helper T cells — might be the link. CD4+ T cells get their “helper” nickname because they help another type of immune cell called a B lymphocyte (or B cell) respond to surface proteins — antigens — on cells infected by foreign invaders such as bacteria or viruses. Activated by the CD4+ T cells, immature B cells become either plasma cells that produce antibodies to mark the infected cells for disposal from the body or memory cells that “remember” the antigen’s biochemistry for a faster response to future invasions.

CD4+ T cells also send out chemical messengers that bring another type of T cell — known as a killer T cell — to the area so that the targeted infected cells can be removed. However, if this activity occurs in the wrong place or at the wrong time, the signals may inadvertently direct the killer T cells to attack healthy cells instead.

“We knew from comparing the brains of infants with NEC with ones from infants who died from other causes that the former had accumulations of CD4+ T cells and showed increased microglial activity,” says study senior author David Hackam, M.D., Ph.D., surgeon-in-chief at Johns Hopkins Children’s Center and professor of surgery at the Johns Hopkins University School of Medicine. “We suspected that these T cells came from the NEC-inflamed regions of the gut and set out to prove it by using neonatal mice as a model of what happens in human infants.”

In the first of a series of experiments, the researchers induced NEC in infant mice and then examined their brains. As expected, the tissues showed a significant increase in CD4+ T cells as well as higher levels of a protein associated with increased microglial activity. In a follow up test, the researchers showed that mice with NEC had a weakened blood-brain barrier — the biological wall that normally prevents bacteria, viruses and other hazardous materials circulating in the bloodstream from reaching the central nervous system. This could, the researchers surmised, explain how CD4+ T cells from the gut could travel to the brain.

Next, the researchers determined that accumulating CD4+ T cells were the cause of the brain injury seen with NEC. They did this first by biologically blocking the movement of the helper T cells into the brain and then in a separate experiment, neutralizing the T cells by binding them to a specially designed antibody. In both cases, microglial activity was subdued and white matter in the brain was preserved.

To further define the role of CD4+ T cells in brain injury, the researchers harvested T cells from the brains of mice with NEC and injected them into the brains of mice bred to lack both T and B lymphocytes. Compared with control mice that did not receive any T cells, the mice that did receive the lymphocytes had higher levels of the chemical signals which attract killer T cells. The researchers also observed activation of the microglia, inflammation of the brain and loss of white matter — all markers of brain injury.

The researchers then sought to better define how the accumulating CD4+ T cells were destroying white matter — actually a fat called myelin that covers and protects neurons in the brain, and facilitates communication between them. To do this, they used organoids, mouse brain cells grown in the laboratory to simulate the entire brain. Brain-derived CD4+ T cells from mice with NEC were added to these laboratory “mini-brains” and then examined for several weeks.

Hackam and his colleagues found that a specific chemical signal from the T cells — a cytokine (inflammatory protein) known as interferon-gamma (IFN-gamma) — increased in the organoids as the amount of myelin decreased. This activity was not seen in the organoids that received CD4+ T cells from mice without NEC.

After adding IFN-gamma alone to the organoids, the researchers saw the same increased levels of inflammation and reduction of myelin that they had seen in mice with NEC. When they added an IFN-gamma neutralizing antibody, cytokine production was significantly reduced, inflammation was curtailed and white matter was partially restored.

The researchers concluded that IFN-gamma directs the process leading to NEC-related brain injury. Their finding was confirmed when an examination of brain tissues from mice with NEC revealed higher levels of IFN-gamma than in tissues from mice without the disease.

Next, the researchers investigated whether CD4+ T cells could migrate from the gut to the brain of mice with NEC. To do this, they obtained CD4+ T cells from the intestines of infant mice with and without NEC. Both types of cells were injected into the brains of infant mice in two groups — one set that could produce the protein Rag1 and one that could not. Rag1-deficient mice do not have mature T or B lymphocytes.

The Rag1-deficient mice that received gut-derived helper T cells from mice with NEC showed the same characteristics of brain injury seen in the previous experiments. T cells from both mice with and without NEC did not cause brain injury in mice with Rag1, nor did T cells from mice without NEC in Rag1-deficient mice. This showed that the gut-derived helper T cells from mice with NEC were the only ones that could cause brain injury.

In a second test, gut-derived T cells from mice with and without NEC were injected into the peritoneum — the membrane lining the abdominal cavity — of Rag1-deficient mice. Only the intestinal T cells from mice with NEC led to brain injury.

This finding was confirmed by genetically sequencing the same portions from both the brain-derived and gut-derived T lymphocytes from mice with and without NEC. The sequences of the helper T cells from mice with NEC, on average, were 25% genetically similar while the ones from mice without NEC were only 2% alike.

In a final experiment, the researchers blocked IFN-gamma alone. Doing so provided significant protection against the development of brain injury in mice with severe NEC. This suggests, the researchers say, a therapeutic approach that could benefit premature infants with the condition.

“Our research strongly suggests that helper T cells from intestines inflamed by NEC can migrate to the brain and cause damage,” says Hackam. “The mouse model in our study was previously shown to closely match what occurs in humans, so we believe that this is the likely mechanism by which NEC-related brain injury develops in premature infants.”

Based on these findings, Hackam says measures for preventing this type of brain injury, including therapies to block the action of INF-gamma, may be possible.

Along with Hackam, the Johns Hopkins Medicine researchers on the study team are Qinjie Zhou, Diego Niño, Yukihiro Yamaguchi, Sanxia Wang, William Fulton, Hongpeng Jia, Peng Lu, Thomas Prindle, Meaghan Morris, Chhinder Sodhi and Liam Chen (now at the University of Minnesota). Also on the team is David Pamies from the University of Lausanne.

The study was funded by National Institutes of Health grants RO1DK117186, RO1DK121824, RO1GM078238, RO1AI148446 and R21AI49321.

Hackam, Sodhi and Pamies have patents on NEC treatments that are unrelated to the research in this study.

The new research, which will appear Jan. 6 in the journal Science Advances, is the latest to probe the lives of what may be nature’s hardest working microbes: cyanobacteria.

These single-celled, photosynthetic organisms, also known as “blue-green algae,” can be found in almost any large body of water today. But more than 2 billion years ago, they took on an extra important role in the history of life on Earth: During a period known as the “Great Oxygenation Event,” ancient cyanobacteria produced a sudden, and dramatic, surge in oxygen gas.

“We see this total shift in the chemistry of the oceans and the atmosphere, which changed the evolution of life, as well,” said study lead author Sarah Hurley, a postdoctoral research associate in the departments of Geological Sciences and Biochemistry. “Today, all higher animals need oxygen to survive.”

To date, scientists still don’t know what these foundational microbes might have looked like, where they lived or what triggered their transformation of the globe.

But Hurley and her colleagues think they might have gotten closer to an answer by drawing on studies of naturally-occurring and genetically-engineered cyanobacteria. The team reports that these ancient microbes may have floated freely in an open ocean and resembled a modern form of life called beta-cyanobacteria.

Studying them, the researchers said, offers a window into a time when single-celled organisms ruled the Earth.

“This research gave us the unique opportunity to form and test hypotheses of what the ancient Earth might have looked like, and what these ancient organisms could have been,” said co-author Jeffrey Cameron, an assistant professor of biochemistry.

Take a breath

You can still make the case that cyanobacteria rule the planet. Hurley noted that these organisms currently produce about a quarter of the oxygen that comes from the world’s oceans.

One secret to their success may lie in carboxysomes — or tiny, protein-lined compartments that float inside all living cyanobacteria. These pockets are critical to the lives of these organisms, allowing them to concentrate molecules of carbon dioxide within their cells.

“Being able to concentrate carbon allows cyanobacteria to live at what are, in the context of Earth’s history, really low carbon dioxide concentrations,” Hurley said.

Before the Great Oxidation Event, it was a different story. Carbon dioxide levels in the atmosphere may have been as much as 100 times what they are today, and oxygen was almost nonexistent. For that reason, many scientists long assumed that ancient microorganisms didn’t need carboxysomes for concentrating carbon dioxide.

“Cyanobacteria have persisted in some form over two billion years of Earth’s history,” she said. “They could have been really different than today’s cyanobacteria.”

To find out how similar they were, the researchers cultured jars filled with bright-green cyanobacteria under conditions resembling those on Earth 2 billion years ago.

Hurley explained that different types of cyanobacteria prefer to digest different forms, or “isotopes,” of carbon atoms. As a result, when they grow, die and decompose, the organisms leave behind varying chemical signatures in ancient sedimentary rocks.

“We think that cyanobacteria were around billions of years ago,” she said. “Now, we can get at what they were doing and where they were living at that time because we have a record of their metabolism.”

Resurrecting zombie microbes

In particular, the team studied two different types of cyanobacteria. They included beta-cyanobacteria, which are common in the oceans today. But the researchers also added a new twist to the study. They attempted to bring an ancient cyanobacterium back from the dead. Hurley and her colleagues used genetic engineering to design a special type of microorganism that didn’t have any carboxysomes. Think of it like a zombie cyanobacterium.

“We had the ability to do what was essentially a physiological resurrection in the lab,” said Boswell Wing, a study coauthor and associate professor of geological sciences.

But when the researchers studied the metabolism of their cultures, they found something surprising: Their zombie cyanobacterium didn’t seem to produce a chemical signature that aligned with the carbon isotope signatures that scientists had previously seen in the rock record. In fact, the best fit for those ancient signals were likely beta-cyanobacteria — still very much alive today.

The team, in other words, appears to have stumbled on a living fossil that was hiding in plain sight. And, they said, it’s clear that cyanobacteria living around the time of the Great Oxygenation Event did have a structure akin to a carboxysome. This structure may have helped cells to protect themselves from growing concentrations of oxygen in the air.

“That modern organisms could resemble these ancient cyanobacteria — that was really counterintuitive,” Wing said.

Scientists, they note, now have a much better idea of what ancient cyanobacteria looked like and where they lived. And that means that they can begin running experiments to dig deeper into what life was like in the 2 billion-year-old ocean.

“Here is hard evidence from the geological record and a model organism that can shed new light on life on ancient Earth,” Cameron said.

Other coauthors on the new paper included CU Boulder undergraduate student Claire Jasper and graduate student Nicholas Hill.

Medina, an assistant professor of biomedical engineering, led the team who published its results Jan. 4 in Nature Biomedical Engineering.

“One of the best protective mechanisms we have to prevent infection are beneficial bacteria that inhabit our bodies, known as commensals,” Medina said. “For example, we often avoid food poisoning because our guts are already populated by helpful bacteria. There’s no room for the pathogen to take hold and colonize. If you wipe out the good bacteria, opportunistic pathogens can take advantage and cause infections.”

Antibiotics can knock out an infection, but they can also kill off good bacteria, creating an opportunity for a potentially fatal secondary infection. Repeated exposure to antibiotics can also breed bacteria resistant to drugs. The potential for secondary infection and drug-resistant bacteria holds true for infections elsewhere in the body, too, according to Medina.

Led by biomedical engineering doctoral student Andrew W. Simonson, first author on the paper, the team set out to develop a peptide that could eradicate the pathogen that causes tuberculosis (TB), one of the top 10 causes of death worldwide, without harming surrounding good bacteria.

“There are great control strategies and treatments in place for tuberculosis, making it largely preventable and treatable, but drug-resistant TB is an emerging threat that is on track to becoming a serious global health problem,” Medina said. “It’s a scary prospect.”

To develop a pathogen-specific antibacterial against TB, the researchers looked to the pathogen itself. The TB pathogen is wrapped in a thick envelope that is difficult to penetrate, especially compared to other bacteria.

“The envelope has pores, though — channels through which the pathogen takes in nutrients and metabolites,” Medina said. “We asked if we could mimic these channels to design antibacterials that would create holes in the bacterial envelope, and ultimately kill the pathogen.”

The researchers made a peptide that seems to disrupt the protective outer coating of the pathogen, making the TB bacteria susceptible to antibiotics and die, but it does not interact with the good bacteria. Medina said they are currently studying the exact mechanism by which the peptide attacks the TB pathogen, but they suspect it has something to do with a fatty acid that lives on the pathogen’s surface.

“There aren’t many biochemical differences between the targeted pathogen and good bacteria, except for this surface lipid,” Medina said. “We think the interaction of our peptide with this fatty acid is one of the things driving this preferential interaction.”

He also pointed to the bacteria’s thin carbohydrate region. In other types of bacteria, the carbohydrates form a thick defensive barrier that appears to insulate the bacteria against the peptide.

“It’s not entirely clear why this works, but it’s working on TB,” Medina said. “There’s a 10-fold improvement on the effectiveness of antibiotics on this pathogen relative to the other good bacteria.”

Next, the researchers plan to investigate how to administer the peptide to treat TB in a full model system. Peptides tend to break down when injected, Medina said, so his team is working to develop an aerosol that would allow a person to inhale the peptides directly to the infected lung tissue.

“Once we understand why this peptide targets TB, and how to administer the peptide as a viable therapeutic, we can use this platform to design antibacterials toward other lung pathogens,” Medina said.

The National Institutes of Health supported this work.

Story Source:

Materials provided by Penn State. Original written by Ashley J. WennersHerron. Note: Content may be edited for style and length.

The research sought to understand how P. gingivalis colonizes the mouth. The pathogen is unable to produce its own growth molecules until it achieves a large population in the oral microbiome (the community of microorganisms that live on and inside the body).

The answer: It borrows growth molecules from V. parvula, a common yet harmless bacteria in the mouth whose growth is not population dependent.

In a healthy mouth, P. gingivalis makes up a miniscule amount of the bacteria in the oral microbiome and cannot replicate. But if dental plaque is allowed to grow unchecked due to poor oral hygiene, V. parvula will multiply and eventually produce enough growth molecules to also spur the reproduction of P. gingivalis.

More than 47% of adults 30 and older have some form of periodontitis (also known as gum disease), according to the Centers for Disease Control and Prevention. Understanding the relationship between P. gingivalis and V. parvula will help researchers create targeted therapies for periodontitis, says Patricia Diaz, DDS, PhD, lead investigator on the study and Professor of Empire Innovation in the UB School of Dental Medicine.

“Having worked with P. gingivalis for nearly two decades, we knew it needed a large population size to grow, but the specific processes that drive this phenomenon were not completely understood,” says Diaz, also director of the UB Microbiome Center. “Successfully targeting the accessory pathogen V. parvula should prevent P. gingivalis from expanding within the oral microbial community to pathogenic levels.”

The study, which was published on Dec. 28 in the ISME Journal, tested the effects of growth molecules exuded by microorganisms in the mouth on P. gingivalis, including molecules from five species of bacteria that are prevalent in gingivitis, a condition that precedes periodontitis.

Of the bacteria examined, only growth molecules secreted by V. parvula enabled the replication of P. gingivalis, regardless of the strain of either microbe. When V. parvula was removed from the microbiome, growth of P. gingivalis halted. However, the mere presence of any V. parvula was not enough to stimulate P. gingivalis, as the pathogen was only incited by a large population of V. parvula.

Data suggest that the relationship is one-directional as V. parvula received no obvious benefit from sharing its growth molecules, says Diaz.

“P. gingivalis and V. parvula interact at many levels, but the beneficiary is P. gingivalis,” says Diaz, noting that V. parvula also produces heme, which is the preferred iron source for P. gingivalis.

“This relationship that allows growth of P. gingivalis was not only confirmed in a preclinical model of periodontitis, but also, in the presence of V. parvula, P. gingivalis could amplify periodontal bone loss, which is the hallmark of the disease,” says George Hajishengallis, DDS, PhD, co-investigator on the study and Thomas W. Evans Centennial Professor in the University of Pennsylvania School of Dental Medicine.

“It is not clear whether the growth-promoting cues produced by P. gingivalis and V. parvula are chemically identical,” says Diaz. “Far more work is needed to uncover the identity of these molecules.”

Additional investigators include Anilei Hoare, PhD, assistant professor, University of Chile; Hui Wang, PhD, postdoctoral researcher, University of Pennsylvania; Archana Meethil, resident, University of Connecticut; Loreto Abusleme, PhD, assistant professor, University of Chile; Bo-Young Hong, PhD, associate research scientist, Jackson Laboratory for Genomic Medicine; Niki Moutsopoulos, DDS, PhD, senior investigator, National Institute of Dental and Craniofacial Research; and Philip Marsh, PhD, professor, University of Leeds.

The research was funded by the National Institute of Dental and Craniofacial Research of the National Institutes of Health.

But how much pressure can they stand? The 2014-2016 North Pacific marine heatwave, known as the Blob, led to a harmful algal bloom of unprecedented scale. It necessitated substantial delays in the opening of the 2015-16 U.S. West Coast Dungeness crab fishery. The fishery is vital to West Coast communities. It produces around 26 percent of all annual fishing revenue and supports more than 30 percent of all commercial fishing vessels.

Understanding Impacts from Climate Shocks

Previous studies have documented the devastating economic impacts from the 2015-16 event on Dungeness crab fishermen. Members of affected coastal communities attest that these socioeconomic effects rippled through associated industries and coastal communities. But can changes in fishing practices in response to this significant climate shock be quantified?

“We wanted to examine the extent to which the Dungeness crab fishery delays affected participation in other fisheries, and the duration of those changes,” said Mary Fisher, a doctoral student at the University of Washington. Fisher did the work as a National Science Foundation Graduate Research Internship Program Fellow at NOAA’s Northwest Fisheries Science Center.

Fisher and her colleagues at NOAA Fisheries, University of Washington, and Oregon State University studied the impacts on more than 2,500 vessels across seven California fishing communities. The researchers wanted to see how a climate-related shock (like the heatwave and associated harmful algal bloom) can impact communities’ use of ocean resources.

The researchers used 10 years of fishery landings data to map resource use networks for seven California port groups where Dungeness crab is an important revenue source. The networks visualize the portfolio of fisheries that a community harvests and how vessels move between fisheries. The network visualization is similar to a food web, except that the connections represent harvest by fishing vessels rather than predator-prey relationships.

Resilient Fishing Communities

Researchers found that 71 percent of California Dungeness crab fishing vessels temporarily left the industry and stopped fishing altogether during the delays. The two other strategies that fishermen used to cope with the disruption were:

• Participating in other fisheries that were not affected by the harmful algal bloom.
• Moving out of delayed areas to fish in other more favorable locations.

These strategies significantly changed communities’ resource use patterns, but some communities were more affected by the climate shock than others. Researchers could predict which communities would be least sensitive to the shock by looking at their resource use patterns before the 2015-16 season.

These communities, located in central California, were less dependent on Dungeness crab and had shorter fishery delays. In these communities, fishermen had access to more open fisheries during the winter months and were more flexible in how they fished previously. These characteristics provided more options, buffering fishermen from the disruption to the Dungeness crab fishing season.

No matter how central Dungeness crab was to a community’s fisheries portfolio, the researchers didn’t observe any significant, lasting changes in how vessels participated in fisheries after the closures lifted. This suggests fishing communities may mostly return to “normal” fishing practices relatively quickly after short-term disturbances like climate shocks, provided those changes don’t pile up one after the other.

Access to Other Fisheries and Ports Important

The results highlight the importance of nearshore groundfish fisheries like sablefish, rockfish, and lingcod as alternatives to the Dungeness crab fishery. “Most of the crab vessels that stayed out on the water were using pot or hook-and-line gear to fish groundfish,” says Fisher. “I think we can expect access to these fisheries to continue to be important during crab season delays.”

Fishermen with larger vessels also landed their catch at different ports, a strategy less common among smaller vessels. This was possible in part because regulations only restricted fishing in delayed areas at the district level. In 2018, the California Department of Fish and Wildlife amended these rules, called the Fair Start Provisions. The amendments provide more protection to fishermen at a finer scale than districts in the event of future delays.