Meningitis: Neisseria Meningitidis Disseminates Itself by Sending out 'Scouts'

ScienceDaily (Feb. 17, 2011) — Although, in the majority of cases, the localized presence ofNeisseria meningitidis in the throat has no consequence, it can sometimes lead to meningitis or septicaemia. The seriousness of these two infections is driving researchers from around the world to improve their understanding of the mode of operation of this bacterium, which, once it leaves its favourite location (the throat) becomes extremely dangerous.

The structure of a pilus. Green: Pilin assembly monomers. Red: Visualization of phosphoglycerol at the surface of the pilus.

The Avenir team directed by Guillaume Duménil within Inserm Mixed Research Unit 970, "Paris centre de recherche cardiovasculaire" at the Université Paris Descartes, has recently discovered how this bacterium disseminates, leaving the throat to pass into the bloodstream.

The results of this research have been published in the 11 February 2011 issue of the journal Science.

Neisseria meningitidis is a bacterium specific to man. It is frequently present in the non-pathogenic state in the throat of healthy carriers (5% to 30% of the population).

Its persistence in the organism can however prove dangerous in some cases. The location where the bacteria multiply, the throat, represents a port of entry, from where it disseminates into the bloodstream and may penetrate into the brain. In both these cases, the infection becomes very serious since it results in septicaemia or meningitis. Unless dealt with quickly, the mortality rate linked to these two infections is very high.

Guillaume Duménil and his Inserm research team were, therefore, interested to understand more about this bacterium which, when it passes into the bloodstream, becomes very dangerous. "Certain advances made in the past few years provided the starting point for this work, which is published inScience", he explained. We know, for example, that Neisseria meningitidis are equipped with special structures known as pili. These allow the bacteria to adhere to the cells of the throat and to multiply and form aggregates there. We are closely studying the main protein which makes up the pili; namely, pilin", adds Guillaume Duménil.

The researchers then discovered that the protein underwent various modifications over time. One of these, in particular, has proved more interesting than the others: The addition of a phosphoglycerol. This chemical group once grafted onto the pilin, gives the signal for dissemination.

Bacteria isolated from the colony depart as "scouts"

Following these initial results, the researchers discovered the presence of a gene which enabled the transfer of phosphoglycerol onto the pilin: the gene, pptB. This gene only becomes fully functional when the bacteria is in contact with the cells lining the wall of the throat. The frantic activity of the pptB gene causes the addition of phosphoglycerol to the pilin. This then loses one of its essential properties: its capacity to form aggregates. As a consequence, some of the bacteria detach themselves from the colony and, little by little, are disseminated. This strategy is used by the bacteria in order to colonize other areas of the throat and to cross the cells lining it. "This phenomena could almost be compared with the formation of metastases in cancer", says Guillaume Duménil.

This is the first time that scientists have been able to accurately identify the chain of events which controls the bacteria in the bloodstream. It is a first step. "We now know how the Neisseria meningitidis passes from the throat into the blood. We hope to be able to demonstrate that an identical process is involved when the bacteria passes from the blood into the brain, instigating meningitis", concludes Guillaume Duménil.

Furthermore, if the researchers were able to find molecules which can block this dissemination, they would have both a preventative tool (blocking colonization from the throat and passage into the bloodstream) and a therapeutic tool (limiting colonization from the blood vessels and transmission into the brain).

Although the strategy developed by Neisseria meningitidisensures its multiplication in the throat, and therefore its survival over the course of evolution, it is also responsible for the death of the host organism and hence its own death. Further proof that living in harmony with a host is not easy.

Conventional Wisdom of How Neurons Operate Challenged: Axons Can Work in Reverse

ScienceDaily (Feb. 17, 2011) — Neurons are complicated, but the basic functional concept is that synapses transmit electrical signals to the dendrites and cell body (input), and axons carry signals away (output). In one of many surprise findings, Northwestern University scientists have discovered that axons can operate in reverse: they can send signals to the cell body, too.

Computer-generated image representing connections between neurons.

It also turns out axons can talk to each other. Before sending signals in reverse, axons can perform their own neural computations without any involvement from the cell body or dendrites. This is contrary to typical neuronal communication where an axon of one neuron is in contact with another neuron's dendrite or cell body, not its axon. And, unlike the computations performed in dendrites, the computations occurring in axons are thousands of times slower, potentially creating a means for neurons to compute fast things in dendrites and slow things in axons.

A deeper understanding of how a normal neuron works is critical to scientists who study neurological diseases, such as epilepsy, autism, Alzheimer's disease and schizophrenia.

The findings are published in the February issue of the journalNature Neuroscience.

"We have discovered a number of things fundamental to how neurons work that are contrary to the information you find in neuroscience textbooks," said Nelson Spruston, senior author of the paper and professor of neurobiology and physiology in the Weinberg College of Arts and Sciences. "Signals can travel from the end of the axon toward the cell body, when it typically is the other way around. We were amazed to see this."

He and his colleagues first discovered individual nerve cells can fire off signals even in the absence of electrical stimulations in the cell body or dendrites. It's not always stimulus in, immediate action potential out. (Action potentials are the fundamental electrical signaling elements used by neurons; they are very brief changes in the membrane voltage of the neuron.)

Similar to our working memory when we memorize a telephone number for later use, the nerve cell can store and integrate stimuli over a long period of time, from tens of seconds to minutes. (That's a very long time for neurons.) Then, when the neuron reaches a threshold, it fires off a long series of signals, or action potentials, even in the absence of stimuli. The researchers call this persistent firing, and it all seems to be happening in the axon.

Spruston and his team stimulated a neuron for one to two minutes, providing a stimulus every 10 seconds. The neuron fired during this time but, when the stimulation was stopped, the neuron continued to fire for a minute.

"It's very unusual to think that a neuron could fire continually without stimuli," Spruston said. "This is something new -- that a neuron can integrate information over a long time period, longer than the typical operational speed of neurons, which is milliseconds to a second."

This unique neuronal function might be relevant to normal process, such as memory, but it also could be relevant to disease. The persistent firing of these inhibitory neurons might counteract hyperactive states in the brain, such as preventing the runaway excitation that happens during epileptic seizures.

Spruston credits the discovery of the persistent firing in normal individual neurons to the astute observation of Mark Sheffield, a graduate student in his lab. Sheffield is first author of the paper.

The researchers think that others have seen this persistent firing behavior in neurons but dismissed it as something wrong with the signal recording. When Sheffield saw the firing in the neurons he was studying, he waited until it stopped. Then he stimulated the neuron over a period of time, stopped the stimulation and then watched as the neuron fired later.

"This cellular memory is a novelty," Spruston said. "The neuron is responding to the history of what happened to it in the minute or so before."

Spruston and Sheffield found that the cellular memory is stored in the axon and the action potential is generated farther down the axon than they would have expected. Instead of being near the cell body it occurs toward the end of the axon.

Their studies of individual neurons (from the hippocampus and neocortex of mice) led to experiments with multiple neurons, which resulted in perhaps the biggest surprise of all. The researchers found that one axon can talk to another. They stimulated one neuron, and detected the persistent firing in the other unstimulated neuron. No dendrites or cell bodies were involved in this communication.

"The axons are talking to each other, but it's a complete mystery as to how it works," Spruston said. "The next big question is: how widespread is this behavior? Is this an oddity or does in happen in lots of neurons? We don't think it's rare, so it's important for us to understand under what conditions it occurs and how this happens."

In addition to Spruston and Sheffield, other authors of the paper are Tyler K. Best and William L. Kath, from Northwestern, and Brett D. Mensh, from Harvard Medical School.

Host Genetics Plays Unexpected Role in Dance With Pathogen

ScienceDaily (Feb. 17, 2011) — A new study suggests that differences in the host's genetics can make a big difference in susceptibility bacterial infection. In a study in the February 2011 Infection and Immunity, Virginia L. Miller of the University of North Carolina, Chapel Hill, and her collaborators show that the virulence of a strain of Yersinia pestis, notable for causing bubonic plague, varies drastically among mice strains with different genetic backgrounds. These findings carry major implications for vaccine development, says Miller.

A number of earlier reports dating back 20 years had suggested that removing this bacterium's capsule--an envelope of a loose protein gel surrounding the bacterial cell -- had no effect on its virulence. Then, Miller and her collaborators performed the same experiment, with opposite results.

Searching for an explanation for the conflicting results, the only difference in the experiments that Miller could find was in the strains of mice, and so it occurred to her that their susceptibilities might be different. Her team tested that hypothesis by infecting two different strains of mice with Y. pestis in which the capsule had been removed. In one strain, the bacteria were nearly normally virulent, while in the other, they were relatively impotent.

This research made sense of the earlier experiments, "while highlighting the importance that host genetics can play in the dance between host and pathogen, and how it can influence the phenotype of a potential virulence factor," says Miller.

Moreover, these findings "demonstrate for the first time that the capsule is a Y. pestis virulence factor in a mouse infection model," says James Bliska of Stony Brook University, New York. "It had already been shown that [the capsule] is important for flea transmission, and therefore it was clear why [the capsule] was conserved in Y. pestis."

The research is critical for the development of a vaccine against both bubonic and pneumonic plague, also caused by Y. pestis, because considerable effort has been invested in establishing Caf1, an antigen within the capsule, as a protective antigen in vaccines against plague. But all the papers showing that removing the capsule had no effect on virulence had gradually undermined the case for using the capsule antigen in a vaccine, when it had been a major target for vaccine development. But now, "This paper may revive hope that Caf1 in conjunction with other antigens would be a useful component of a multivalent vaccine," says Eric Krukonis of the University of Michigan, Ann Arbor.

Developing such vaccines is important because "Y. pestis is still a major threat to humans, due to endemic pockets of Y. pestis-infected animals and fleas and potential bioterrorism use," Miller and her collaborators note. "A greater understanding of the requirement of the capsule for Y. pestis to cause disease is required. It is particularly important to investigate if natural capsule mutants are able to cause disease and in what contexts, as the current vaccine potentially would not protect against these strains."

Tau-Induced Memory Loss in Alzheimer’s Mice Is Reversible; Study Raises Hopes for the Development of Effective Therapies

ScienceDaily (Feb. 17, 2011) — Amyloid-beta and tau protein deposits in the brain are characteristic features of Alzheimer disease. The effect on the hippocampus, the area of the brain that plays a central role in learning and memory, is particularly severe. However, it appears that the toxic effect of tau protein is largely eliminated when the corresponding tau gene is switched off. Researchers from the Max Planck Research Unit for Structural Molecular Biology at DESY in Hamburg have succeeded in demonstrating that once the gene is deactivated, mice with a human tau gene, which previously presented symptoms of dementia, regain their ability to learn and remember, and that the synapses of the mice also reappear in part. The scientists are now testing active substances to prevent the formation of tau deposits in mice. This may help to reverse memory loss in the early stages of Alzheimer disease -- in part, at least.

To test their capacity to learn, the mice are trained to find an underwater platform which is not visible to them from the edge of a water basin. The swimming path is marked in red. Normal mice learn to find the path after just a few training sessions; they remember it and swim straight to the platform (left) when tested. A mouse with too much aggregated tau protein in its neurons finds it difficult to learn and swims aimlessly around the basin (centre) for extended periods. If the gene for the toxic tau protein in this mouse is switched off for a few weeks using a genetic trick, the mouse is able to learn normally again and quickly finds its way to the platform (right).

Whereas aggregated amyloid-beta protein forms insoluble clumps between the neurons, the tau protein accumulates inside them. Tau protein stabilises the tube-shaped fibers of the cytoskeleton, known as microtubules, which provide the "rails" for cellular transport. In Alzheimer disease, excess phosphate groups cause the tau protein to malfunction and form clumps (the 'neurofibrillary tangles'). As a result, nutrient transport breaks down and the neurons and their synapses die off. This process is accompanied by the initial stage of memory loss.

Together with colleagues from Leuven, Hamburg and Erlangen, Eva and Eckhard Mandelkow's team from the Max Planck Research Unit for Structural Molecular Biology generated regulatable transgenic mice with two different human tau gene variants that can be switched on and off again: one group was given a form of the protein that cannot become entangled (anti-aggregant), and a second was provided with the code for the strongly aggregating protein variant (pro-aggregant). The mice with the first form developed no Alzheimer symptoms; the rodents that were given the pro-aggregant tau developed the disease.

The scientists measured the mice's memory loss with the help of a swimming test: the healthy mice quickly learn how to find a life-saving platform located under the surface of the water in a water basin. In contrast, the transgenic animals, which have the additional pro-aggregant tau gene paddle aimlessly around the basin until they accidentally stumble on the platform; they require over four times more time to do this than their healthy counterparts. However, if the mutated toxic tau gene is switched off again, the mice learn to reach "dry land" with ease just a few weeks later. As a control, the mice with the anti-aggregant form of tau have no defects in learning, just as normal non-transgenic mice.

Surprising tissue results

Tissue tests showed that, as expected, no tau clumps had formed in the brains of the first group of mice expressing anti-aggregant tau. In the second group -- the mice suffering from Alzheimer's -- co-aggregates from human tau and "mouse tau" were formed -- against expectations, because tau protein from mice does not usually aggregate. "Even more astonishingly, weeks after the additional gene had been switched off, the aggregated human tau had dissolved again. However, the 'mouse tau' remained clumped. Despite this, the mice were able to learn and remember again," says Eckhard Mandelkow. More precise tests revealed that new synapses had actually formed in their brains.

The scientists concluded from this that mutated or pathological tau can alter healthy tau. It appears that pro-aggregant tau can act similar to a crystal nucleus -- once it has started to clump up, it drags neighboring "healthy" tau into the clumps as well. This is what makes the process so toxic to the neurons. "The really important discovery here, however, is that the progression of Alzheimer's disease can be reversed in principle -- at least at an early stage of the illness before too many neurons have been destroyed," explains Eva Mandelkow who, together with her husband, will be awarded the Potamkin Prize 2011 for Alzheimer's disease research, which is sponsored by the American Academy of Neurology.

The aggregation of tau proteins, however, cannot simply be switched off in humans the way it can in the transgenic mice. Nevertheless, special substances exist that could dissolve the tau aggregates. By screening 200,000 substances, the Hamburg researchers have already identified several classes of active substances that could re-convert the tau aggregates into soluble tau. These are now being tested on animals.

Group of Enzymes Could Have a Positive Impact on Health, from Cholesterol to Osteoporosis

ScienceDaily (Feb. 17, 2011) — Recent studies conducted at the Institut de recherches cliniques de Montréal (IRCM) on a group of PCSK enzymes could have a positive impact on health, from cholesterol to osteoporosis. A team led by Dr. Nabil G. Seidah, Director of the Biochemical Neuroendocrinology research unit, has published six articles in scientific journals over the past four months, all shedding light on novel functions of certain PCSK enzymes.

PCSK enzymes belong to the proprotein convertase family, responsible for the conversion of an inactive protein into its active state. The latest projects led by Dr. Seidah and his team focused on five of the nine PCSK enzymes, which are implicated in diseases such as cardiovascular and neuroendocrine disorders, cancer, and viral infections.

PCSK9 could help lower bad cholesterol levels

A member of the proprotein convertase family, PCSK9 plays a key role in the regulation of cholesterol. It is involved in causing familial hypercholesterolemia, a genetic disorder characterized by high total cholesterol levels in the blood, specifically very high levels of LDL (low-density lipoprotein) or bad cholesterol, which can lead to the early onset of cardiovascular diseases. PCSK9 is thus a target for the treatment of dyslipidemia, which results from an abnormal concentration of lipids (fat) in the blood. It is believed that inhibition of PCSK9 function could lower LDL-cholesterol levels, and such treatments are currently undergoing early phase clinical trials.

"Members of my team, led by Dr. Annik Prat and Dr. Anna Roubtsova, discovered that PCSK9 also regulates fatty acids," explains Dr. Seidah. "They investigated its role in the metabolism of body fat, and found that PCSK9 is pivotal in regulating cholesterol and fat metabolism: it maintains high circulating cholesterol levels, but it also limits fat generation."

In another study, the researchers uncovered further data on PCSK9's mechanism of action and functional structure. Their data showed that a part of PCSK9 inhibits its own function, and that acidic pH levels affect the degradation of bad cholesterol receptors.

In a third study, the team discovered that two other PCSK enzymes, Furin (PCSK3) and PC5/6 (PCSK5), reduce the level of active PCSK9. "They were able to demonstrate how enzymes of the PCSK family communicate with one another and found that liver-derived Furin cuts PCSK9 and inactivates it," adds Dr. Seidah. "This provided genetic evidence for the mechanism behind the functioning of a mutant gene found in some human hypercholesterolemic patients."

An important enzyme for osteoporosis, especially for women after menopause

The eighth member of the family (PCSK8), known as SKI-1/S1P, is critical in the intracellular pathway leading to the synthesis of cholesterol and fatty acids. Researchers discovered a new function of the enzyme in the regulation of bone formation. They demonstrated that mineralization -- the process by which the body uses minerals to build bone structure -- was blocked by inhibitors of SKI-1/S1P.

"The team found that SKI-1/S1P also regulates the activation of a membrane-bound transcription factor required for bone formation, which could have an impact on osteoporosis," says Dr. Seidah. "This could be especially important for women after menopause, as they are most likely to develop bone disease."

A better understanding of an enzyme implicated in various cancers

PC7 (or PCSK7) is the most ancient and highly-conserved basic amino acid-specific member of the proprotein convertase family. The team of researchers, led by Estelle Rousselet, shed light on how PC7 functions within cells, and was able to define its intracellular trafficking pathways. "We identified a mechanism for PC7's unique ability to activate the precursor of epidermal growth factor, which is involved in various cancers, tumour growth, and the maintenance of stem cells," concludes Dr. Seidah.