Rénald Delanoue, Inserm Researcher, and his colleagues at the Institute of Biology Valrose in Nice (Inserm-CNRS-Université Côte d’Azur) have identified the missing links in the process that regulates the size of an organism based on the richness of its diet. Their research was conducted on Drosophila, an insect that seems very distant from humans, but the study of which has nonetheless enabled many advances in biomedical research. This work is published in the 30 September 2016 issue of the journal Science.

The size of an organism depends on its nutrient intake during development. In the event of a nutrient deficiency during this period, animals modify their growth and become adults of small size, while retaining the correct proportions. This coupling between nutrition and growth involves hormones from the insulin family and IGFs (insulin growth factors); however, the molecular mechanisms that govern this regulation are still not well understood.

Work done by Inserm researchers at the Institute of Biology Valrose (Inserm-CNRS-UCA) has enabled the identification of the substances on which this coupling is based at molecular level in the fruit fly (Drosophila melanogaster). Despite 700 million years of evolutionary divergence, this insect is a relevant model for biomedical research, because it possesses the same physiological processes as mammals.

For example, it is interesting to know that the fat body in Drosophila fulfils the same roles as the liver and adipose tissues in humans. The neurosecretory insulin producing cells (IPCs) are located in the larval brain of the insect, and correspond functionally to the pancreatic β-cells in humans.

Using the Drosophila model, these researchers have already shown that the coupling of nutrition and growth requires communication between these two organs (ndlr fat body and IPCs). Depending on the quantity of amino acids available in the diet, the fat body sends different signals to the brain. The IPCs are able to interpret these, and to secrete the appropriate amount of insulin. A low level of amino acids induces a reduction in insulin secretion and a slowing in growth, and vice versa.

It remained necessary to identify the nature of these remotely acting signals, and the IPC molecule capable of interpreting them to determine the quantity of insulin to be secreted. To do this, the researchers inhibited, one by one, the known receptors on IPCs, and identified the Methuselah receptor, inhibition of which blocks insulin secretion.

The Stunted protein, which binds to this receptor, was known, but had not been linked to the regulation of insulin secretion. And for a reason! This protein is usually found mainly inside the cells, and plays a role in ATP synthesis. It was therefore surprising to find it freely circulating in the haemolymph of the insect (the equivalent of the bloodstream). The researchers showed that the level of circulating Stunted varies with the quantity of amino acids present in the diet. Its suppression disrupts insulin secretion, and gives rise to adults of small size. Finally, they also demonstrated that this “signalling” function of Stunted in communication between two organs is a novel one, and independent of its function in ATP synthesis.

These results, although very fundamental, could nonetheless guide the study of the molecular circuits of certain diseases such as diabetes, obesity or some forms of cancer that depend on hormones and receptors from the same family as those described in Drosophila. Stunted proteins, which have been found on the surface of many cell types, could also play a signalling role in humans.

The fluorescent protein GFP (green) is expressed in insulin producing neurons, IPCs (dotted white lines). Secretion of Drosophila insulins (Dilp2, in red) is regulated in these specialised neurons by the presence of the Methuselah receptor. The absence of this membrane protein in Mth- IPCs leads to blocking of insulin (Dilp2) secretion and its accumulation in IPCs (bottom row), compared with the control (top row).

(c) Rénald Delanoue/Inserm

The first embryonic division, which follows gamete fusion (oocyte and spermatozoon), starts the development of a new individual, the genesis of a functional adult body. This division is symmetric in the one-cell embryo stage (also known as the zygote); it leads to the formation of two daughter cells of identical size. Conversely, it is asymmetric in the oocyte, which has the same size and shape as the zygote. Why? What directs the zygote to divide symmetrically, while the oocyte divides asymmetrically during meiosis? Such are the questions pondered by Marie-Emilie Terret, a researcher at Inserm, and Marie-Hélène Verlhac, a researcher at CNRS and director of the Asymmetric Divisions in Oocytes team at the Center for Interdisciplinary Research in Biology (CIRB; Inserm/CNRS/Collège de France)[1]. By combining biology, physics and mathematics, the researchers succeeded in demonstrating, in mice, the regulatory mechanics that determine, in a very short time, the geometry and hence the fate (symmetric or asymmetric division) of the cell. In future, the elements arising from this work may contribute to improving the efficacy of in vitro fertilisation.

Details of these results are published today in the journal Nature communications.

At the one-cell stage, the embryo very closely resembles an oocyte: a round, isolated cell, similar in size to an oocyte. The geometry of a cell’s division is determined by the position of the microtubule spindle, the machinery that carries and separates the chromosomes. In most animal cells, the centrosomes organise the microtubule network, which is essential to the formation and positioning of the division spindle. But oocytes and zygotes lack centrosomes. A major difference between these two types of cells, however, lies in the geometry of their divisions. Indeed, during meiosis oocytes divide in an extremely asymmetric manner in terms of size, allowing the formation of a single enormous oocyte and the expulsion of “polar bodies” containing the excess genetic material. Conversely, the zygote divides in a perfectly symmetric manner, leading to the formation of two daughter cells of identical size.

Mouse oocyte and embryo at the one-cell stage. The top images show the actin networks; the bottom images show the microtubule spindles with aligned chromosomes.

(c) Marie-Emilie Terret

The geometry of division is determined by the position of the spindle: eccentric in oocytes, centred in zygotes. The Asymmetric Divisions in Oocytes team had previously shown that the eccentric position of the division spindle in the oocyte depends on the mechanics of the actin networks. In the work published today, the team of researchers shows that the centred position of the division spindle in the zygote is also due to the mechanics of the actin networks, but controlled differently.

Three steps are required for this symmetric division:

1. The coarse centring of the male and female pronuclei, requiring a network of actin and myosin-Vb.
2. The fine centring of the division spindle, requiring strong rigidity in the oocyte.
3. The passive maintenance of the spindle in the centre of the cell.

The mechanics of actin/myosin networks therefore make it possible to change from an asymmetric division to a symmetric division, a change in geometry needed for the transition from oocyte to embryo.

The research team is already formulating hypotheses regarding the mode of action of actin, which plays a role in the physical characteristics of the cell membrane (rigid or soft), which in turn influence the geometry of division. “Our next work will involve the more detailed study of interactions between actin and microtubules in an attempt to understand their respective roles in the architecture of the cell at the moment of division, and the potential involvement of other intermediary proteins,” explains Marie-Emilie Terret.

A better understanding of the physical characteristics and behaviour of the oocyte during division, whether fertilised or not, has the potential to contribute useful new elements to medically assisted conception. During in vitro fertilisation (IVF), for instance, the storage temperature of the oocytes can have an impact on the quality of the actin networks, and consequently affect division, and hence the formation of a zygote.

Increasing the efficacy of IVF might therefore represent a long-term objective for this research team, one of the few in France working on this theme.

[1] In collaboration with researchers from the Laboratory for Analysis and Modelling for Biology and Environment (LAMBE; CNRS/CEA/University of Évry Val d’Essonne/Cergy Pontoise University), the Theoretical Physics of Condensed Matter Laboratory (LPTMC; CNRS/UPMC) and the Curie Physical Chemistry Laboratory (CNRS/Institut Curie/UPMC).

Researchers at Inserm and Paris Descartes University have just taken an important step in research on stem cells and dental repair. They have managed to isolate dental stem cell lines and to describe the natural mechanism by which they repair lesions in the teeth. This fundamental discovery will make it possible to initiate unprecedented therapeutic strategies to mobilise the resident dental stem cells and magnify their natural capacity for repair.

These results are published in the journal Stem Cells.

The tooth is a mineralised organ, implanted in the mouth by a root. The “living” part of the tooth or dental cavity is the dental pulp (in yellow in the photograph shown opposite) composed of vessels and nerves. Around it is a hard substance, the dentine or ivory, which is in turn covered by an even harder tissue, the enamel. When a dental lesion appears, the dormant stem cells in the pulp awaken and try to repair the tooth by an unknown process.

3D modelling of a teeth. The dental pulp is in yellow. ©Inserm/ Chappard, Daniel

In this study, the researchers from Inserm and Paris Descartes University at Unit 1124, “Toxicology, Pharmacology and Cellular Signaling,” have succeeded in extracting and isolating tooth stem cells by working on the pulp from the mouse molar.

The researchers were thus able to analyse the cells in detail, and identify 5 specific receptors for dopamine and serotonin on their surface, two neurotransmitters that are essential to the body (see schema on page 2).

The presence of these receptors on the surface of these stem cells indicated that they had the ability to respond to the presence of dopamine and serotonin in the event of a lesion. The researchers naturally wondered what cells might be the source of these neurotransmitters, a warning signal. It turns out that the blood platelets, activated by the dental lesion, are responsible for releasing a large quantity of serotonin and dopamine. Once released, these neurotransmitters then recruit the stem cells to repair the tooth by binding to their receptors (see schema on page 2). The research team was able to confirm this result by observing that dental repair was absent in rats with modified platelets that do not produce serotonin or dopamine, i.e. in the absence of the signal.

“In stem cell research, it is unusual to be simultaneously able to isolate cell lines, identify the markers that allow them to be recognised (here the 5 receptors), discover the signal that recruits them (serotonin and dopamine), and discover the source of that signal (blood platelets). In this work, we have been able, unexpectedly, to explore the entire mechanism,” explains Odile Kellermann, leader of the team from Inserm and Paris Descartes University, and the main author of this work.

To take things a stage further, the researchers tried to characterise the different receptors they found. One of the 5 receptors does not seem to affect the repair process. On the other hand, the other 4 turn out to be strongly involved in the repair process. In vivo blocking of just one of them is enough to prevent dental repair.

“Currently, dentists use pulp capping materials (calcium hydroxide) and tricalcium phosphate-based biomaterials to repair the tooth and fill lesions. Our results lead us to imagine unprecedented therapeutic strategies aimed at mobilising the resident pulpal stem cells in order to magnify the natural reparative capacity of teeth without use of replacement materials,” concludes Odile Kellermann.

The foundations have been laid for extending this research done in rodents to stem cells of the human tooth in order to initiate new strategies for repairing teeth.

© Inserm / Odile Kellermann, Anne Baudry

How can a specialized cell change its identity? A team from the Institut de Génétique et de Biologie Moléculaire et Cellulaire (CNRS/INSERM/Université de Strasbourg) investigated a 100% effective natural example of this phenomenon, which is called transdifferentiation. This process, by which some cells lose their characteristics and acquire a new identity, could be more generally involved in tissue or organ regeneration in vertebrates, and is a promising research avenue for regenerative medicine. This study identifies the role of epigenetic factors involved in this conversion, underlines the dynamic nature of the process, and shows the key mechanisms for effective transdifferentiation. This work, conducted in collaboration with the Institut Curie1, was published on August 15, 2014 in Science.

How can a specialized cell change its identity? A team from the Institut de Génétique et de Biologie Moléculaire et Cellulaire (CNRS/INSERM/Université de Strasbourg) investigated a 100% effective natural example of this phenomenon, which is called transdifferentiation. This process, by which some cells lose their characteristics and acquire a new identity, could be more generally involved in tissue or organ regeneration in vertebrates, and is a promising research avenue for regenerative medicine. This study identifies the role of epigenetic factors involved in this conversion, underlines the dynamic nature of the process, and shows the key mechanisms for effective transdifferentiation. This work, conducted in collaboration with the Institut Curie1, was published on August 15, 2014 in Science.

Our body is constituted of cells that acquired characteristics during development and that fulfill a precise function in each organ: we call these differentiated cells. Generally cells maintain their specificity until they die, but it has been proven that some cells can change state and acquire new functions. This is rare but is found in many species and is called “transdifferentiation”.

The team studied this process in C. elegans, a small transparent nematode, where a rectal cell transforms naturally into a motor neuron. This change from one cell type into another occurs without cell division, by a succession of well defined steps that always lead to the same result. The researchers investigated the factors that make the conversion process so stable.

The team had elucidated the role of several transcription factors2 in this transdifferentiation.

The respective role of genetic and epigenetic factors in biological processes is a hotly debated subject. This work shows how each of these factors acts in transdifferentiation: transcription factors handle initiation and progress whereas epigenetic factors guarantee the constant result. The study even goes further, showing that under “normal” conditions, the epigenetic factors are incidental (even when they are absent the conversion occurs relatively efficiently) but that they are indispensable when there are environmental stressors. So they have a crucial role in maximizing the mechanism’s efficacy and ensuring that it remains stable in the face of external variations.

Transdifferentiation is a phenomenon that is poorly understood. It may be involved in the organ regeneration that we observe in some organisms, for example newts, which can reconstruct their eye lens after injury. These results bring key new information to help us understand how to control this process and may open the path to promising therapies, in particular in the field of regenerative medicine.

© Elodie Legrand et Sophie Jarriault
We can compare this process to the layers of an onion. Transcription factors are at the heart of process efficiency, while epigenetic factors form the outer layers that protect the mechanism from attacks and environmental change.