While most of us recover from influenza after a week, it can be a very severe disease, and even fatal in rare cases, with no reason for physicians to have expected such an outcome. By analysing the genome of a little girl who contracted a severe form of influenza at the age of two and a half years, researchers at the Laboratory of Human Genetics of Infectious Diseases (a joint French-American international laboratory), which brings together researchers from Inserm, Paris Descartes University, and physicians from the Paris public hospitals (AP-HP; Necker Hospital for Sick Children), working at the Imagine Institute, and from The Rockefeller University in New York, have discovered that she has a genetic mutation, unknown until now, that causes a subtle dysfunction in her immune system. More generally, these results show that genetic mutations could be the root cause of some severe forms of influenza in children, and indicate in any event that immune mechanisms missing in this little girl are needed for protection against this virus in humans. These results are published in the journal Science Express.

©fotolia

Seasonal influenza is an acute viral infection caused by the influenza virus. It is characterised by high fever, headaches, sore muscles, etc. Apart from vaccination, there is no treatment for it other than symptomatic (pain) treatment. In most cases, patients recover after a week, but in more vulnerable people influenza can cause acute respiratory distress, which is potentially fatal.

The main known risk factors for severe forms of influenza are some acquired comorbidities, such as chronic lung disease. However, the cause of most fatal cases remains unexplained, especially in children.

The absence of cases of severe influenza in patients with known acquired immunodeficiencies, which usually increase susceptibility to infections, is also surprising.

Given these different observations, the researchers at Jean-Laurent Cassanova’s and Laurent Abel’s laboratory, in Paris and New York, therefore formulated a hypothesis whereby severe influenza in healthy children might be the result of genetic errors.

To test this hypothesis, they sequenced the entire genome of a 7-year-old child who had contracted a severe form of influenza (influenza A virus strain H1N1), requiring her admission to a paediatric intensive care unit in January 2011, at the age of two and a half years. At the time, she showed no other known pathology that might have suggested greater vulnerability to the virus than that of other children.

This analysis, combined with analysis of her parents’ genomes, made it possible to show that the little girl had inherited a mutated allele of the gene encoding interferon regulatory factor (IRF7) from both of her parents. The latter is a transcription factor known to amplify the production of interferons in response to viral infection in mice and humans.

In contrast to her parents, in whom the mutation of a single allele of the gene is of no consequence, in the little girl, mutation of both alleles of the gene encoding IRF7 has led to its inactivation. The result: failure to produce interferons, disrupting her system of defence against influenza virus infection in a cascading manner.

By carrying out a comprehensive series of experiments on blood cells, particularly dendritic cells, and by generating lung cells from stem cells taken from the young girl, the researchers provided proof that the mutations observed in this little girl explain the development of severe influenza. Furthermore, this discovery demonstrates that interferon amplification dependent on IRF7 expression is needed for protection against influenza virus in humans. They now need to search for mutations in this or other genes in other children recruited following an episode of unexplained severe influenza.

Based on these initial observations, the researchers at Inserm believe that therapeutic strategies based on recombinant interferons, available in the pharmacopoeia, could help to combat severe forms of influenza in children.

This multidisciplinary study required multiple collaborations in Europe and the United States.

Basic research on blood pressure has led researchers from Inserm (Inserm Unit 1138, “Cordeliers Research Centre”) to obtain unexpected results: drugs used to treat hypertension (high blood pressure) reduce side effects from corticosteroid-based creams used to treat certain skin diseases.

This work is published in the Journal of Investigative Dermatology.

To switch the subtitles of the movie on or off during playback check/uncheck the Subtitles enabled option.

Corticosteroid-based dermatological creams are indicated for the symptomatic treatment of inflammatory skin conditions, such as atopic dermatitis and psoriasis, for example. However, they have frequent side effects, such as a slight burning sensation, and very often end by inducing skin atrophy (thinning of the skin, which becomes fragile), which is inconvenient for the patient, and for which there is presently no treatment.

The researchers from Inserm formulated a hypothesis whereby this harmful effect might be related to the inappropriate activation by these creams of mineralocorticoid receptors located in the epidermis. These receptors, which are present in the kidney, heart, eye, and certain neurons in particular, reacted with aldosterone, a hormone that regulates the blood pressure. Moreover, previous studies also showed them to be highly sensitive to corticosteroids.

Application of corticosteroids to cultured skin causes it to become thinner: in 6 days, the thickness of the epidermis was reduced by one-third. The researchers then induced a pharmacological blockade of the receptors by adding specific antagonists to the corticosteroid treatment. The inability of the corticosteroid to bind to the mineralocorticoid receptors restores proliferation of the epidermal cells, and partially corrects epidermal atrophy.

From the clinical point of view, it turns out that spironolactone, a drug used for a very long time as an antihypertensive drug (and which has marketing authorisation), is an antagonist of the mineralocorticoid receptor. The researchers therefore tested a treatment based on spironolactone for 28 days in 23 healthy volunteers. Creams of different composition were applied to 4 areas of their arms:

– a cream containing a corticosteroid used in dermatology

– a cream containing spironolactone

– a combination of both drugs

– a placebo

The results obtained show that adding spironolactone to the corticosteroid significantly limits skin atrophy.

For Nicolette Farman, “This is a highly original piece of work, at the crossroads between endocrinology and dermatology, and brings together researchers in basic science and clinicians. Now it remains to reformulate this old drug for a new application, and test this product in patients with various skin diseases in order to confirm the reduction in side effects from corticosteroids without loss of efficacy.”

© fotolia

Scientists from the CNRS, INSERM and Université de Limoges, working in the Laboratoire Contrôle de la Réponse Immune B et Lymphoproliférations (CNRS/Université de Limoges)[1] have demonstrated that the production of type E immunoglobulins (IgE)[2]by B lymphocytes induces a loss in their mobility and the initiation of cell death mechanisms. These antibodies, present in small quantities, are the most powerful “weapons” in the immune system and can trigger extremely violent immune reactions or immediate allergies (asthma, urticaria, allergic shock) as soon as their levels rise, even slightly. These findings, published online in Cell reports on 12 February 2015, thus elucidate how our bodies restrict the production of IgE in order to prevent an allergic reaction.

B lymphocytes under a confocal microscope (x1000). The cytoskeleton (actin molecules is labeled with a green fluorescent probe (phalloidin FITC). B lymphocytes with an IgM on their surface display protuberances (pseudopods) which testify to their mobility, while IgE+ B lymphocytes lose theses structures and become immobile.
© CNRS Laboratoire Contrôle de la réponse immune B et lymphoproliférations 

Immunity is based on cells, B lymphocytes, which carry or secrete antibacterial or antiviral “weapons”, the immunoglobulins (IgG, IgM, IgA, IgE) or antibodies. Although these weapons of immunity offer protection, they can also sometimes turn on us. This is the case for the most effective of antibodies, IgE, where even infinitesimal traces (these IgE are 100,000 times less abundant than other antibodies) can trigger extremely violent allergic reactions.

The lymphocytes that produce IgM, IgG or IgA are numerous, easily identifiable and persistent (as “memory cells”). For hitherto unexplained reasons, the cells that produce IgE are rare and have thus been the subject of very little study. In order to understand the mechanisms controlling IgE, the scientists first of all used genetic engineering to force cells to produce these antibodies in large numbers. They then succeeded in demonstrating two principal control mechanisms. They showed that as soon as a B lymphocyte carries an IgE on its membrane, it “freezes”, swells, loses its pseudopods[3] and becomes incapable of moving, although lymphocytes are generally highly mobile. The scientists also revealed that the lymphocyte activates several mechanisms leading to apoptosis, or programed cell death. This self-destruction causes the rapid elimination of lymphocytes carrying IgE, while other cells in the immune system are able to survive for up to several years.

During evolution, our bodies have thus developed several self-restriction mechanisms around one of their most powerful immune “weapons”, IgE. Because a cell carrying IgE can no longer move, it can only survive for a brief period — just long enough to play a short-lived protective role against parasites, toxins and poisons. It then self-destructs by committing a sort of “hara-kiri” which strongly reduces IgE production and hence the triggering of allergies. The scientists now wish to explore in more detail the different molecular pathways governing this self-restriction. Indeed, these may constitute numerous new therapeutic targets whose pharmacological activation could block allergies, or even permit the reduction of other pathological B lymphocytes, such as those involved in lymphomas.

Internalization of membrane IgE (spontaneous endocytosis) contributing to weak IgE expression and the death of these cells. Visualization under a confocal microscope (x1000) of IgE+ B lymphocytes at 37 °C with anti-IgE antibodies. This labeling is able to demonstrate the internalization of membrane IgE indicated in blue (right). 
© CNRS Contrôle de la réponse immune B et lymphoproliférations

A study has focused on the evolutionary history of the mycobacterium that causes tuberculosis, and more specifically on the Beijing lineage associated with the spread of multidrug resistant forms of the disease in Eurasia. While confirming the East-Asian origin of this lineage, the results also indicate that this bacterial population has experienced notable variations coinciding with key events in human  history. They also demonstrate that two multidrug resistant (MDR) clones of this lineage started to spread concomitantly with the collapse of the public health system in the former Soviet Union, thus highlighting the need to sustain efforts to control tuberculosis. Finally, this work has made it possible to identify new potential targets for the treatment and diagnosis of this disease. This study was carried out by scientists at the Centre d’Infection et d’Immunité de Lille (CNRS/Institut Pasteur de Lille/Inserm/Université de Lille) and the Institut de Systématique, Evolution, Biodiversité (CNRS/Muséum national d’Histoire naturelle/UPMC/EPHE), working in collaboration with a large international consortium[1]. Its findings were published on 19 January in Nature Genetics.

Tuberculosis bacilli visualized through scanning electron microscopy © Jean-Pierre Tissier (INRA, Villeneuve d’Ascq) et Franco Menozzi (Institut Pasteur de Lille)

The results of these genetic analyses indicate that the Beijing lineage emerged nearly 7,000 years ago in a region lying between northeastern China, Korea and Japan, whence it radiated throughout the world in successive waves associated with historical movements of human populations from east to west. In the modern era, the bacterial population first saw its size increase during the industrial revolution and the First World War. These phases of growth were probably linked to increases in human population density and deprivation respectively associated with these events. The only phase of  decline observed thereafter was concomitant with the widespread use of antibiotics during the 1960s. This decline ended in the late 1980s, and was  related to the HIV/AIDS epidemic and the onset of multidrug resistance.

This study has also shown that the more recent epidemic spread of the two  strains most closely associated with MDR in Central Asia and Eastern Europe coincided with the collapse of the public health system in the former Soviet Union. These findings underline the importance of maintaining highly  efficient disease control systems and developing new and more effective methods for  diagnosis and treatment.

In this context, the scientists identified a series of mutations and genes that might be connected with epidemic episodes and multidrug resistance. These genes constitute potential targets for treatment and for the development of new and more rapid diagnostic methods for multidrug resistance, based on  genomic sequencing.

A clinical trial project, coordinated by Inserm, involving the testing of a preventive vaccine against Ebola has been selected by the European Commission. The protocol plans to include participants throughout Europe and Africa to evaluate immune response and tolerance to a vaccine strategy named “prime boost”, based on the use of two candidate vaccines developed by Janssen, a pharmaceutical company of Johnson & Johnson.

© Fotolia

The development and rapid access to treatment and candidate vaccines are among the WHO’s recommendations to stop the transmission of the Ebola virus and prevent the international spread of infection. Since the beginning of the epidemic, the French and international scientific community has been actively working towards these objectives.

Inserm has established an academic partnership with the London School of Hygiene and Tropical Medicine (LSHTM), as well as an industrial partnership (Crucell Holland BV; one of Janssen’s companies), The University of Oxford and the Centre Muraz to develop a vaccination from Phase 1 to Phase 3 that combines two vaccines derived from viral vectors (Ad26.ZEBOV et MVA-BN-Filo[1]). This project has been selected for funding by the European Commission under the second call for IMI2 (Innovative Medicines Initiative) projects. As part of this programme, Inserm will conduct Phase 2 trials in Europe and Africa, coordinated by Prof. Rodolphe Thiébaut.

These Phase 2 trials will evaluate the tolerance and quality of the immune response to the vaccine strategy. These tests will supplement data from Phase 1 trials that are currently in progress in the United States, England and are soon to be conducted in Africa. The data will be available in March 2015 and will be critical to evaluating effectiveness in areas at risk.

“This strategy, unlike conventional immunization protocols that involve one or more administrations of the same vaccine, is based on the concept of a vaccination involving several steps with two different vectors that will expose the organism to the same antigens in several ways. This is a new approach in the development of a vaccine against Ebola” stated Prof. Yves Levy, Inserm CEO.

The Grant Agreement is currently being finalised. Definitive information on the project, including the budget, will be published once the agreement has been signed.

[1] Both viral vectors used are conventional vectors that are already widely used in humans in several vaccines for other infectious diseases.

The complete genomes of 16 anopheline mosquito species from the five continents have just been sequenced. Ten years of research have enabled an international consortium, coordinated by the University of Michigan and University of Notre Dame (United States) and including researchers from the French Institute for Development Research (IRD) and Inserm, to publish in the 27 November 2014 issue of the journal Science the DNA sequence for these mosquitoes, which are vectors of malaria. These results lead the way to the development of comparative genomic studies, and will make it possible to improve strategies for vector control, which are essential to controlling the disease in the absence of a vaccine.

© IRD/ M. Dukhan : female mosquito : Anopheles sundaicus

Malaria causes over 600,000 deaths a year, mainly in Sub-Saharan Africa. The parasites, from the genus Plasmodium, are transmitted by mosquitoes of the genus Anopheles. Of the 450 Anopheles species on the planet, only a dozen are responsible for most of the transmission to humans.

In order to improve vector control strategies, researchers have devoted many years to decoding the genome of Anopheles species. That of the African mosquito, Anopheles gambiae, a major vector of malaria, has been available since 2002. Those of its South American (Anopheles darlingi) and Indian (Anopheles stephensi) homologues have been the subject of recent publications, in 2013 and 2014 respectively. Until now, the lack of knowledge on the genetic resources of other Anopheles species has restricted comparisons that would enable the identification the key features determining the ability of some mosquitoes to transmit the parasites.

10 years of North/South partnership

Led by the University of Michigan’s Broad Institute and the University of Notre Dame’s Eck Institute, over a period of 10 years the international consortium enlisted over a hundred researchers from 50 research institutes from the northern (United States, Europe) and southern (Africa, Asia, South America, Oceania, Australia) hemispheres.

The researchers studied Anopheles specimens from many regions in the world: Africa, Asia, Asia Minor, Central America and Oceania. They combined the most recent techniques for sequencing, assembly of genomes and of expressed genes, and chromosome mapping, using innovative methodologies for sequence analysis and genome comparison. Using this multidisciplinary approach, the researchers succeeded in sequencing and annotating the complete genomes of 16 new Anopheles species.

Strong genetic evolution in Anopheles

The sequenced genomes turned out to be very different, both in their composition and their general organisation. Thus their size varies from 135 to 275 million base pairs. Between 10,000 and 16,000 genes were identified per species. The first comparative analyses showed strong genetic peculiarities, including a high rate of molecular evolution compared with other insects (particularly Drosophila), and substantial genomic plasticity, with much gain and loss of genes (or entire groups of genes) in the course of evolution.

Analyses carried out on certain groups of genes involved in key elements of anopheline biology—such as reproduction, immune system, insecticide resistance, composition of the cuticle or saliva, odour perception or hormonal communication—enabled the researchers to identify a certain number of specific genes and traits acquired during evolution, which underpin the emergence of anthropophily[1] and parasite transmission in anophelines.

The knowledge of this genetic material improves the understanding of the mechanisms by which the vectors adapt to humans and their environment. The researchers now have new avenues of research for making their vector control strategies more effective, and controlling the transmission of malaria.

[1] Describes organisms (plant or animal) that live in contact with humans or in places they frequent.

Erosive oral lichen planus (OLP) is an auto-immune disease affecting skin and mucous membranes which results in an abnormal immune response against mucocutaneous cells. Today, scientists at the Institut Pasteur, Inserm, Paul Sabatier University (Toulouse) and the CNRS have proven that the immune cells involved in OLP are the same as those activated during an immune response to human papillomavirus (type HPV-16). This suggests a link between OLP and HPV. The results of this study were published in the Journal of Investigative Dermatology.

Erosive oral lichen planus (OLP) is an inflammatory disease which is considered auto-immune because of the abnormal immune response it triggers against other cells in the body. The condition affects mucous membranes around the mouth and genitals causing lesions and the destruction of skin cells called keratinocytes. Because current treatments are only partially effective, the condition is chronic and, although rare (between 0.1 and 4% of the overall population is affected), OLP can have serious side effects such as pain, difficulty eating, and cancer.

Papillomavirus© Institut Pasteur

Little was known about the underlying biological mechanisms of OLP until teams led by Marie-Lise Gougeon (Institut Pasteur), Nicolas Fazilleau (Inserm, Paul Sabatier University, CNRS) and Hervé Bachelez (Sorbonne Paris Cité, Paris Diderot University) demonstrated that the immune response which leads to the destruction of mucosal cells involves the same lymphocytes responsible for the immune response to human papillomavirus (HPV). This would suggest a link between OLP and infection by the HPV-16 strain, a virus known to be responsible for genital warts and cervical cancer.

Initial analyses of lesional tissue and blood samples from OLP patients revealed the presence of cytotoxic lymphocytes around the destroyed cells. This prompted scientists to try to characterize the specific role and origin of these lymphocytes.

For the ten patients enrolled in a study by Manuelle Viguier (Institut Pasteur, Sorbonne Paris Cité, Paris Diderot University), analyses showed an abnormally high population of a particular type of lymphocyte specific for HPV-16: T CD8 (type Vβ3).

The proportion of this type of lymphocyte was then measured during the different phases of OLP (outbreaks or remission stages). This allowed the scientists to show that the number of these cells decreased during periods of clinical remission and multiplied during outbreaks.

One of the theories put forward by the scientists was that keratinocytes in OLP patients might express an autoantigen (an endogenous normal tissue constituent) very similar to the HPV-16 antigen. This could be a source of confusion for T lymphocytes which, having already been exposed to HPV, might mistake the surface antigen on keratinocytes of OLP patients for the HPV antigen and trigger a cytotoxic immune response against keratinocytes.

This research indicates that the auto-immune disease OLP could involve T CD8 lymphocytes specific to HPV-16. This is the first time a link has been established between infection by HPV-16 and an auto-immune disease.

These results open up new therapeutic possibilities for treating severe forms of OLP. The Institut Pasteur has filed to patent this research in hopes that use of the HPV vaccine will be expanded to include OLP.

This study was financed by the French Dermatological Society, the Fondation ARC (Cancer Research Foundation), the French Cancer League, the French National Cancer Institute, the Midi-Pyrénées Regional Council and the Marie Curie International Re-integration Grant.