Etude de l'interaction entre ADN et apatite analogue au minéral osseux et dentaire : implications pour la préservation de l'ADN ancien, son extraction, son analyse

Ce mémoire contribue à l'exploration des mécanismes physico-chimiques qui sous-tendent la préservation post mortem de l'ADN ancien au sein de restes squelettiques dans un contexte anthropologique ou médico-légal. Nous avons étudié le rôle potentiellement joué par la matrice minérale, composée d'apatite phosphocalcique, au sein d'un système complexe. Cette complexité provient tant des propriétés intrinsèques du tissu que de l'influence de paramètres dits " environnementaux " (température, pH, force ionique, activités enzymatiques) dans la préservation de l'ADN. Trois volets d'exploration ont été retenus. Une étude in situ a été réalisée via la caractérisation physico-chimique de spécimens osseux et dentaires anciens utilisés comme substrats d'ADN. Nos résultats contribuent à l'identification d'indicateurs physico-chimiques de la diagénèse, tels que la quantité de matière organique résiduelle, l'état de maturation de l'apatite, ou encore la taille de cristallites. L'exploration in vitro de processus d'adsorption/désorption d'ADN sur une apatite biomimétique de synthèse préalablement caractérisée a été initiée, permettant des essais de modélisation des interactions physico-chimiques observées. Nos données corroborent l'hypothèse d'une conservation exceptionnelle de l'ADN dans le temps par le biais d'une adsorption sur l'apatite, avancée empiriquement par de nombreux paléogénéticiens. Sous un angle plus appliqué, un kit de purification d'ADN, utilisé pour l'ADN ancien, a été caractérisé afin de mieux comprendre l'affinité de l'ADN pour des surfaces inorganiques et identifier les éventuels paramètres influençant le rendement et la qualité des extraits d'ADN ancien.This thesis contributes to explore the physico-chemical mechanisms underlying the preservation of ancient DNA within skeletal remains, arising from anthropological or forensic contexts. The complexity of such systems is not only related to the intrinsic features of hard tissues, but also to the effect of environmental parameters (temperature, pH, ionic strength, microbiological activity). In particular, we focused on the potential key role played by the mineral matrix, composed of calcium phosphate apatite. We addressed this topic from three viewpoints. First, an in situ study was achieved by means of physico-chemical characterization of bone and tooth remains, previously used as DNA substrates. Our results contribute to identify diagenetic indicators, such as the amount of residual organic matter, the maturation state of the apatite phase, or else mean crystallite dimensions. Next, we initiated a study of DNA adsorption/desorption processes on synthetic biomimetic apatite, pointing to the existence of strong physico-chemical interactions, that we have attempted to model. Our data substantiate the empirical hypothesis often alleged by paleogenetics researchers - but not experimentally demonstrated - explaining the exceptional preservation of DNA over time thanks to an adsorption phenomenon onto apatite within hard tissues. From a more applied perspective, the components of a DNA purification kit, used for ancient DNA, have been examined from a physico-chemical point of view with the goal to better understand DNA affinity toward inorganic surfaces and identify key parameters which may allow improving the yield and the quality of ancient DNA extracts

Heme Drives Susceptibility of Glomerular Endothelium to Complement Overactivation Due to Inefficient Upregulation of Heme Oxygenase-1

Atypical hemolytic uremic syndrome (aHUS) is a severe disease characterized by microvascular endothelial cell (EC) lesions leading to thrombi formation, mechanical hemolysis and organ failure, predominantly renal. Complement system overactivation is a hallmark of aHUS. To investigate this selective susceptibility of the microvascular renal endothelium to complement attack and thrombotic microangiopathic lesions, we compared complement and cyto-protection markers on EC, from different vascular beds, in in vitro and in vivo models as well as in patients. No difference was observed for complement deposits or expression of complement and coagulation regulators between macrovascular and microvascular EC, either at resting state or after inflammatory challenge. After prolonged exposure to hemolysis-derived heme, higher C3 deposits were found on glomerular EC, in vitro and in vivo, compared with other EC in culture and in mice organs (liver, skin, brain, lungs and heart). This could be explained by a reduced complement regulation capacity due to weaker binding of Factor H and inefficient upregulation of thrombomodulin (TM). Microvascular EC also failed to upregulate the cytoprotective heme-degrading enzyme heme-oxygenase 1 (HO-1), normally induced by hemolysis products. Only HUVEC (Human Umbilical Vein EC) developed adaptation to heme, which was lost after inhibition of HO-1 activity. Interestingly, the expression of KLF2 and KLF4—known transcription factors of TM, also described as possible transcription modulators of HO-1- was weaker in micro than macrovascular EC under hemolytic conditions. Our results show that the microvascular EC, and especially glomerular EC, fail to adapt to the stress imposed by hemolysis and acquire a pro-coagulant and complement-activating phenotype. Together, these findings indicate that the vulnerability of glomerular EC to hemolysis is a key factor in aHUS, amplifying complement overactivation and thrombotic microangiopathic lesions

Diagenesis of archaeological bone and tooth

An understanding of the structural complexity of mineralised tissues is fundamental for exploration into the field of diagenesis. Here we review aspects of current and past research on bone and tooth diagenesis using the most comprehensive collection of literature on diagenesis to date. Environmental factors such as soil pH, soil hydrology and ambient temperature, which influence the preservation of skeletal tissues are assessed, while the different diagenetic pathways such as microbial degradation, loss of organics, mineral changes, and DNA degradation are surveyed. Fluctuating water levels in and around the bone is the most harmful for preservation and lead to rapid skeletal destruction. Diagenetic mechanisms are found to work in conjunction with each other, altering the biogenic composition of skeletal material. This illustrates that researchers must examine multiple diagenetic pathways to fully understand the post-mortem interactions of archaeological skeletal material and the burial environment

Heme Oxygenase 1: A Defensive Mediator in Kidney Diseases

The incidence of kidney disease is rising, constituting a significant burden on the healthcare system and making identification of new therapeutic targets increasingly urgent. The heme oxygenase (HO) system performs an important function in the regulation of oxidative stress and inflammation and, via these mechanisms, is thought to play a role in the prevention of non-specific injuries following acute renal failure or resulting from chronic kidney disease. The expression of HO-1 is strongly inducible by a wide range of stimuli in the kidney, consequent to the kidney’s filtration role which means HO-1 is exposed to a wide range of endogenous and exogenous molecules, and it has been shown to be protective in a variety of nephropathological animal models. Interestingly, the positive effect of HO-1 occurs in both hemolysis- and rhabdomyolysis-dominated diseases, where the kidney is extensively exposed to heme (a major HO-1 inducer), as well as in non-heme-dependent diseases such as hypertension, diabetic nephropathy or progression to end-stage renal disease. This highlights the complexity of HO-1’s functions, which is also illustrated by the fact that, despite the abundance of preclinical data, no drug targeting HO-1 has so far been translated into clinical use. The objective of this review is to assess current knowledge relating HO-1’s role in the kidney and its potential interest as a nephroprotection agent. The potential therapeutic openings will be presented, in particular through the identification of clinical trials targeting this enzyme or its products

Préservation de l'ADN ancien dans les vestiges osseux et dentaires (rôle et interaction de l'hydroxyapatite)

STRASBOURG-Medecine (674822101) / SudocSudocFranceF

Heme Oxygenase 1: A Defensive Mediator in Kidney Diseases

International audienceThe incidence of kidney disease is rising, constituting a significant burden on the healthcare system and making identification of new therapeutic targets increasingly urgent. The heme oxygenase (HO) system performs an important function in the regulation of oxidative stress and inflammation and, via these mechanisms, is thought to play a role in the prevention of non-specific injuries following acute renal failure or resulting from chronic kidney disease. The expression of HO-1 is strongly inducible by a wide range of stimuli in the kidney, consequent to the kidney's filtration role which means HO-1 is exposed to a wide range of endogenous and exogenous molecules, and it has been shown to be protective in a variety of nephropathological animal models. Interestingly, the positive effect of HO-1 occurs in both hemolysis- and rhabdomyolysis-dominated diseases, where the kidney is extensively exposed to heme (a major HO-1 inducer), as well as in non-heme-dependent diseases such as hypertension, diabetic nephropathy or progression to end-stage renal disease. This highlights the complexity of HO-1's functions, which is also illustrated by the fact that, despite the abundance of preclinical data, no drug targeting HO-1 has so far been translated into clinical use. The objective of this review is to assess current knowledge relating HO-1's role in the kidney and its potential interest as a nephroprotection agent. The potential therapeutic openings will be presented, in particular through the identification of clinical trials targeting this enzyme or its products

MECANISMES DE RECONNAISSANCE ET DE TRANSPORT DE CATIONS METALLIQUES PAR DES LIGANDS D'INTERET BIOLOGIQUE

STRASBOURG-Sc. et Techniques (674822102) / SudocSudocFranceF

C3-dependent effector functions of complement

International audienceC3 is the central effector molecule of the complement system, mediating its multiple functions through different binding sites and their corresponding receptors. We will introduce the C3 forms (native C3, C3 [H2 O], and intracellular C3), the C3 fragments C3a, C3b, iC3b, and C3dg/C3d, and the C3 expression sites. To highlight the important role that C3 plays in human biological processes, we will give an overview of the diseases linked to C3 deficiency and to uncontrolled C3 activation. Next, we will present a structural description of C3 activation and of the C3 fragments generated by complement regulation. We will proceed by describing the C3a interaction with the anaphylatoxin receptor, followed by the interactions of opsonins (C3b, iC3b, and C3dg/C3d) with complement receptors, divided into two groups: receptors bearing complement regulatory functions and the effector receptors without complement regulatory activity. We outline the molecular architecture of the receptors, their binding sites on the C3 activation fragments, the cells expressing them, the diversity of their functions, and recent advances. With this review, we aim to give an up-to-date analysis of the processes triggered by C3 activation fragments on different cell types in health and disease contexts