In the first evidence of a natural intervention triggering stem cell-based regeneration of an organ or system, a study in the June 5 issue of the Cell Press journal Cell Stem Cell shows that cycles of prolonged fasting not only protect against immune system damage — a major side effect of chemotherapy — but also induce immune system regeneration, shifting stem cells from a dormant state to a state of self-renewal.
In both mice and a Phase 1 human clinical trial, long periods of not eating significantly lowered white blood cell counts. In mice, fasting cycles then “flipped a regenerative switch”: changing the signaling pathways for hematopoietic stem cells, which are responsible for the generation of blood and immune systems, the research showed.
The study has major implications for healthier aging, in which immune system decline contributes to increased susceptibility to disease as we age. By outlining how prolonged fasting cycles — periods of no food for two to four days at a time over the course of six months — kill older and damaged immune cells and generate new ones, the research also has implications for chemotherapy tolerance and for those with a wide range of immune system deficiencies, including autoimmunity disorders.
“We could not predict that prolonged fasting would have such a remarkable effect in promoting stem cell-based regeneration of the hematopoietic system,” said corresponding author Valter Longo, the Edna M. Jones Professor of Gerontology and the Biological Sciences at the USC Davis School of Gerontology, and director of the USC Longevity Institute.
“When you starve, the system tries to save energy, and one of the things it can do to save energy is to recycle a lot of the immune cells that are not needed, especially those that may be damaged,” Longo said. “What we started noticing in both our human work and animal work is that the white blood cell count goes down with prolonged fasting. Then when you re-feed, the blood cells come back. So we started thinking, well, where does it come from?”
Prolonged fasting forces the body to use stores of glucose, fat and ketones, but also breaks down a significant portion of white blood cells. Longo likens the effect to lightening a plane of excess cargo.
During each cycle of fasting, this depletion of white blood cells induces changes that trigger stem cell-based regeneration of new immune system cells. In particular, prolonged fasting reduced the enzyme PKA, an effect previously discovered by the Longo team to extend longevity in simple organisms and which has been linked in other research to the regulation of stem cell self-renewal and pluripotency — that is, the potential for one cell to develop into many different cell types. Prolonged fasting also lowered levels of IGF-1, a growth-factor hormone that Longo and others have linked to aging, tumor progression and cancer risk.
“PKA is the key gene that needs to shut down in order for these stem cells to switch into regenerative mode. It gives the ‘okay’ for stem cells to go ahead and begin proliferating and rebuild the entire system,” explained Longo, noting the potential of clinical applications that mimic the effects of prolonged fasting to rejuvenate the immune system. “And the good news is that the body got rid of the parts of the system that might be damaged or old, the inefficient parts, during the fasting. Now, if you start with a system heavily damaged by chemotherapy or aging, fasting cycles can generate, literally, a new immune system.”
Prolonged fasting also protected against toxicity in a pilot clinical trial in which a small group of patients fasted for a 72-hour period prior to chemotherapy, extending Longo’s influential past research: “While chemotherapy saves lives, it causes significant collateral damage to the immune system. The results of this study suggest that fasting may mitigate some of the harmful effects of chemotherapy,” said co-author Tanya Dorff, assistant professor of clinical medicine at the USC Norris Comprehensive Cancer Center and Hospital. “More clinical studies are needed, and any such dietary intervention should be undertaken only under the guidance of a physician.”
“We are investigating the possibility that these effects are applicable to many different systems and organs, not just the immune system,” said Longo, whose lab is in the process of conducting further research on controlled dietary interventions and stem cell regeneration in both animal and clinical studies.
- Chia-Wei Cheng, Gregor B. Adams, Laura Perin, Min Wei, Xiaoying Zhou, Ben S. Lam, Stefano Da Sacco, Mario Mirisola, David I. Quinn, Tanya B. Dorff, John J. Kopchick, Valter D. Longo. Prolonged Fasting Reduces IGF-1/PKA to Promote Hematopoietic-Stem-Cell-Based Regeneration and Reverse Immunosuppression. Cell Stem Cell, 2014; 14 (6): 810 DOI:1016/j.stem.2014.04.014
A team led by scientists from The Scripps Research Institute (TSRI) and the International AIDS Vaccine Initiative (IAVI) has unveiled a new technique for vaccine design that could be particularly useful against HIV and other fast-changing viruses.
The report, which appears March 28, 2013, in Science Express, the early online edition of the journal Science, offers a step toward solving what has been one of the central problems of modern vaccine design: how to stimulate the immune system to produce the right kind of antibody response to protect against a wide range of viral strains. The researchers demonstrated their new technique by engineering an immunogen (substance that induces immunity) that has promise to reliably initiate an otherwise rare response effective against many types of HIV.
“We’re hoping to test this immunogen soon in mice engineered to produce human antibodies, and eventually in humans,” said team leader William R. Schief, who is an associate professor of immunology and member of the IAVI Neutralizing Antibody Center at TSRI.
Seeking a Better Way
For highly variable viruses such as HIV and influenza, vaccine researchers want to elicit antibodies that protect against most or all viral strains — not just a few strains, as seasonal flu vaccines currently on the market. Vaccine researchers have identified several of these broadly neutralizing antibodies from long-term HIV-positive survivors, harvesting antibody-producing B cells from blood samples and then sifting through them to identify those that produce antibodies capable of neutralizing multiple strains of HIV. Such broadly neutralizing antibodies typically work by blocking crucial functional sites on a virus that are conserved among different strains despite high mutation elsewhere.
However, even with these powerful broadly neutralizing antibodies in hand, scientists need to find a way to elicit their production in the body through a vaccine. “For example, to elicit broadly neutralizing antibodies called VRC01-class antibodies that neutralize 90 percent of known HIV strains, you could try using the HIV envelope protein as your immunogen,” said Schief, “but you run into the problem that the envelope protein doesn’t bind with any detectable affinity to the B cells needed to launch a broadly neutralizing antibody response.”
To reliably initiate that VRC01-class antibody response, Schief and his colleagues therefore sought to develop a new method for designing vaccine immunogens.
From Weak to Strong
Joseph Jardine, a TSRI graduate student in the Schief laboratory, evaluated the genes of VRC01-producing B cells in order to deduce the identities of the less mature B cells — known as germline B cells — from which they originate. Germline B cells are major targets of modern viral vaccines, because it is the initial stimulation of these B cells and their antibodies that leads to a long-term antibody response.
In response to vaccination, germline B cells could, in principle, mature into the desired VRC01-producing B cells — but natural HIV proteins fail to bind or stimulate these germline B cells so they cannot get the process started. The team thus set out to design an artificial immunogen that would be successful at achieving this.
Jardine used a protein modeling software suite called Rosetta to improve the binding of VRC01 germline B cell antibodies to HIV’s envelope protein. “We asked Rosetta to look for mutations on the side of the HIV envelope protein that would help it bind tightly to our germline antibodies,” he said.
Rosetta identified dozens of mutations that could help improve binding to germline antibodies. Jardine then generated libraries that contained all possible combinations of beneficial mutations, resulting in millions of mutants, and screened them using techniques called yeast surface display and FACS. This combination of computational prediction and directed evolution successfully produced a few mutant envelope proteins with high affinity for germline VRC01-class antibodies.
Jardine then focused on making a minimal immunogen — much smaller than HIV envelope — and so continued development using the “engineered outer domain (eOD)” previously developed by Po-Ssu Huang in the Schief lab while Schief was at the University of Washington. Several iterative rounds of design and selection using a panel of germline antibodies produced a final, optimized immunogen — a construct they called eOD-GT6.
A Closer Look
To get a better look at eOD-GT6 and its interaction with germline antibodies, the team turned to the laboratory of Ian A. Wilson, chair of the Department of Integrative Structural and Computational Biology and a member of the IAVI Neutralizing Antibody Center at TSRI.
Jean-Philippe Julien, a senior research associate in the Wilson laboratory, determined the 3D atomic structure of the designed immunogen using X-ray crystallography — and, in an unusual feat, also determined the crystal structure of a germline VRC01 antibody, plus the structure of the immunogen and antibody bound together.
“We wanted to know whether eOD-GT6 looked the way we anticipated and whether it bound to the antibody in the way that we predicted — and in both cases the answer was ‘yes’,” said Julien. “We also were able to identify the key mutations that conferred its reactivity with germline VRC01 antibodies.”
Mimicking a Virus
Vaccine researchers know that such an immunogen typically does better at stimulating an antibody response when it is presented not as a single copy but in a closely spaced cluster of multiple copies, and with only its antibody-binding end exposed. “We wanted it to look like a virus,” said Sergey Menis, a visiting graduate student in the Schief laboratory.
Menis therefore devised a tiny virus-mimicking particle made from 60 copies of an obscure bacterial enzyme and coated it with 60 copies of eOD-GT6. The particle worked well at activating VRC01 germline B cells and even mature B cells in the lab dish, whereas single-copy eOD-GT6 did not.
“Essentially it’s a self-assembling nanoparticle that presents the immunogen in a properly oriented way,” Menis said. “We’re hoping that this approach can be used not just for an HIV vaccine but for many other vaccines, too.”
The next step for the eOD-GT6 immunogen project, said Schief, is to test its ability to stimulate an antibody response in lab animals that are themselves engineered to produce human germline antibodies. The difficulty with testing immunogens that target human germline antibodies is that animals typically used for vaccine testing cannot make those same antibodies. So the team is collaborating with other researchers who are engineering mice to produce human germline antibodies. After that, he hopes to learn how to drive the response, from the activation of the germline B cells all the way to the production of mature, broadly neutralizing VRC01-class antibodies, using a series of designed immunogens.
Schief also hopes they will be able to test their germline-targeting approach in humans sooner rather than later, noting “it will be really important to find out if this works in a human being.”
The first authors of the paper, “Rational HIV immunogen design to target specific germline B cell receptors,” were Jardine, Julien and Menis. Co-authors were Takayuki Ota and Devin Sok of the Nemazee and Burton laboratories at TSRI, respectively; Travis Nieusma of the Ward laboratory at TSRI; John Mathison of the Ulevitch laboratory at TSRI; Oleksandr Kalyuzhniy and Skye MacPherson, researchers in the Schief laboratory from IAVI and TSRI, respectively; Po-Ssu Huang and David Baker of the University of Washington, Seattle; Andrew McGuire and Leonidas Stamatatos of the Seattle Biomedical Research Institute; and TSRI principal investigators Andrew B. Ward, David Nemazee, Ian A. Wilson, and Dennis R. Burton, who is also head of the IAVI Neutralizing Center at TSRI.
Joseph Jardine, Jean-Philippe Julien, Sergey Menis, Takayuki Ota, Oleksandr Kalyuzhniy, Andrew McGuire, Devin Sok, Po-Ssu Huang, Skye MacPherson, Meaghan Jones, Travis Nieusma, John Mathison, David Baker, Andrew B. Ward, Dennis R. Burton, Leonidas Stamatatos, David Nemazee, Ian A. Wilson, and William R. Schief. Rational HIV Immunogen Design to Target Specific Germline B Cell Receptors. Science, 28 March 2013 DOI: 10.1126/science.1234150
How Deadly Marburg Virus Silences Immune System: Breakthrough Findings Point to Targets for Drugs and Vaccines
Scientists at The Scripps Research Institute have determined the structure of a critical protein from the Marburg virus, a close cousin of Ebola virus. These viruses cause similar diseases and are some of the deadliest pathogens on the planet, each killing up to 90 percent of those infected.
The Marburg virus VP35 protein (beige) surrounds the virus’s double-stranded RNA (blue), masking it from immune system detection. (Credit: Image by Christina Corbaci, The Scripps Research Institute) (up)
Described in the Sept. 13, 2012 publication of the journal PLoS Pathogens, the new research reveals how a key protein component of the Marburg virus, called VP35, blocks the human immune system, allowing the virus to grow unchecked. The structure provides a major step forward in understanding how the deadly virus works, and may be useful in the development of potential treatments for those infected.
“The immune system is designed to recognize certain hallmarks of virus infection,” said Erica Ollmann Saphire, the Scripps Research scientist who led the effort. “When these are sensed, an immediate antiviral defense is launched. However, the Marburg and Ebola viruses mask the evidence of their own infection. By doing so, the viruses are able to replicate rapidly and overwhelm the patient’s ability to launch an effective defense.”
Ebola virus outbreaks have occurred in the last month in both Uganda and the Democratic Republic of the Congo, while Marburg virus broke out in Angola in 2005 to 2006 and again in Uganda in 2007. The Angolan Marburg virus outbreak began in a pediatric ward and killed 88 percent of those it infected. The virus has since been imported into the United States (Colorado) and the Netherlands by tourists who had visited Africa.
There is currently no cure for Marburg hemorrhagic fever. The virus is spread when people come into contact with the bodily fluids of a person or animal who is already infected. The best treatment consists of administering fluids and taking protective measures to ensure containment, like isolating the patient and washing sheets with bleach.
Most people, however, die within two weeks of exposure from a combination of dehydration, massive bleeding, and shock. A smaller number of people have stronger and immediate immune responses against the virus and survive.
A New Roadmap for Defense
The breakthrough described in the PLoS Pathogens article explains a key reason why the viruses are so deadly and provides the necessary templates to develop drugs to treat the infection.
The study’s lead author, Research Associate Shridhar Bale, explains that a key signature of Marburg virus infection is the double-stranded RNA that results from its replication inside cells. When human immune system proteins detect this virus-specific RNA, they sound an alarm to signal the rest of the immune system to respond. The new research describes how the VP35 protein of the Marburg virus binds to the viral double-stranded RNA and hides it to prevent the alarm from being sounded.
The new research also revealed a surprise. Images from the Marburg virus reveal the VP35 protein spirals around the double-stranded RNA, enveloping it completely. This is in contrast to previous images of the similar VP35 protein from Ebola virus that showed it only capping the ends of the RNA, leaving the center of the RNA helix exposed for possible recognition.
In addition to Ollmann Saphire and Bale, the article, “Marburg virus VP35 can both fully coat the backbone and cap the ends of dsRNA for interferon antagonism,” was authored by Jean-Philippe Julien, Zachary A. Bornholdt, Michelle A. Zandonatti, Gerard J.A. Kroon, Christopher R. Kimberlin, Ian J. MacRae, and Ian A. Wilson of The Scripps Research Institute, and Peter Halfmann, John Kunert, and Yoshihiro Kawaoka of the University of Wisconsin.
Support for the research was provided by grants from the Burroughs Wellcome Fund and The Skaggs Institute for Chemical Biology at Scripps Research.
Note: Materials may be edited for content and length. For further information, please contact the source cited above.
Bale S, Julien J-P, Bornholdt ZA, Kimberlin CR, Halfmann P, et al. Marburg Virus VP35 Can Both Fully Coat the Backbone and Cap the Ends of dsRNA for Interferon Antagonism. PLoS Pathog. PLoS Pathogens, 2012; 8(9): e1002916 DOI: 10.1371/journal.ppat.1002916
As far as bacteria are concerned, other living creatures are just another niche to exploit, which means that pretty much every animal and plant has a set of bacterial pathogens that come along with it. These bacteria have made the animal in question their speciality, and are highly adapted to live inside their hosts. While these bacteria often make the host ill, or less fit, or sometimes dead, the longer they live with their host, overall, the less they damage it. After all, it’s no help to the bacteria if their home drops down dead right after they’ve moved in.
A great example of this appeared in PLoS Pathogens this month (reference 1), concerning the bacteria Wolbachia. These bacteria infect insects and other arthropods and are much beloved of journalists (well, compared to other insect bacteria at least) because one of their effects is to stop insects producing male offspring (so only female survive to pass on the bacterial genome), which gives journalists the opportunity to write silly headlines.
An electron micrograph of an insect cell, with three Wolbachia bacteria inside (the large circular blobs with white lines surrounding them). Image from reference 2. (up)
As well as passing from females onto their offspring, Wolbachia can also be transmitted horizontally, that is between insects in the same generation. In its normal host the Wolbachia is not hugely damaging (apart from removing all males from the population) but when transmitted to a new species it causes various unpleasant nervous system complications, often leading to death. Clearly, the bacteria are more virulent when they encounter a new species. However when the bacterial infection was closely examined, it was found that infected individuals of both species contained the same number of bacteria. It wasn’t just that the new species couldn’t respond to the infection, it was in fact the way they responded that was doing the damage.
As it turns out, the reason Wolbachia are more dangerous in new species isn’t because the bacteria go wild in the unexplored territory, rather it’s because the new host doesn’t know how to deal with them. The insects that are used to dealing with the presence of the bacteria have developed ways to contain the infection, or just tolerate it. New species however, tend to panic, particularly as the bacteria tend to congregate in the gonads (sex organs) and the central nervous system, which even insects understand are bad places to have bacteria.
As the bacteria are found inside cells, the best way for an insect immune system to get rid of them, is by destroying the cells that house the bacteria. Which, as previously mentioned, are mainly the gonads and the central nervous system. When the Wolbachia get into a new species, the first response of the insect is to quickly and efficiently destroy any cells which have bacteria inside them. As a consequence the unfortunate insect basically destroys its own brain, leading to various unpleasant symptoms and death.
The carpenter ant, Camponotus pennsylvanicus. Many species of Camponotus are infected with Wolbachia. Image from reference 3. (up)
Even in insects, the immune system is vital to defend animals from bacterial, fungal, and viral attacks. However it’s fascinating to see the cases where the immune system (even ‘primitave’ immune systems that consist of nothing more than infected cells quickly being removed) can lead to issues by over-reacting to a threat. The best response to the Wolbachia is for the insects to learn to deal with it, rather than to attempt to launch counter-attacks which can be damaging for the animal as a whole.
Adopted from: Rat Lab blog
2: (2004) Genome Sequence of the Intracellular Bacterium Wolbachia. PLoS Biol 2(3): e76. doi:10.1371/journal.pbio.0020076
3: Wernegreen JJ (2004) Endosymbiosis: Lessons in Conflict Resolution. PLoS Biol 2(3): e68. doi:10.1371/journal.pbio.0020068