[SAMHD1 acts at stalled replication forks to prevent interferon induction]. / SAMHD1 agit sur les fourches de réplication bloquées pour empêcher l'induction d'interféron.

DNA replication is an extremely complex process, involving thousands of replication forks progressing along chromosomes. These forks are frequently slowed down or stopped by various obstacles, such as secondary DNA structures, chromatin-acting proteins or a lack of nucleotides. This slowing down, known as replicative stress, plays a central role in tumour development. Complex processes, which are not yet fully understood, are set up to respond to this stress. Certain nucleases, such as MRE11 and DNA2, degrade the neo-replicated DNA at the level of blocked forks, allowing the replication to restart. The interferon pathway is a defense mechanism against pathogens that detects the presence of foreign nucleic acids in the cytoplasm and activates the innate immune response. DNA fragments resulting from genomic DNA metabolism (repair, retrotransposition) can diffuse into the cytoplasm and activate this pathway. A pathological manifestation of this process is the Aicardi-Goutières syndrome, a rare disease characterized by chronic inflammation leading to neurodegenerative and developmental problems. In this encephalopathy, it has been suggested that DNA replication may generate cytosolic DNA fragments, but the mechanisms involved have not been characterized. SAMHD1 is frequently mutated in the Aicardi-Goutières syndrome as well as in some cancers, but its role in the etiology of these diseases was largely unknown. We show that cytosolic DNA accumulates in SAMHD1-deficient cells, particularly in the presence of replicative stress, activating the interferon response. SAMHD1 is important for DNA replication under normal conditions and for the processing of stopped forks, independent of its dNTPase activity. In addition, SAMHD1 stimulates the exonuclease activity of MRE11 in vitro. When SAMHD1 is absent, degradation of neosynthesized DNA is inhibited, which prevents activation of the replication checkpoint and leads to failure to restart the replication forks. Resection of the replication forks is performed by an alternative mechanism which releases DNA fragments into the cytosol, activating the interferon response. The results obtained show, for the first time, a direct link between the response to replication stress and the production of interferons. These results have important implications for our understanding of the Aicardi-Goutières syndrome and cancers related to SAMHD1. For example, we have shown that MRE11 and RECQ1 are responsible for the production of DNA fragments that trigger the inflammatory response in cells deficient for SAMHD1. We can therefore imagine that blocking the activity of these enzymes could decrease the production of DNA fragments and, ultimately, the activation of innate immunity in these cells. In addition, the interferon pathway plays an essential role in the therapeutic efficacy of irradiation and certain chemotherapeutic agents such as oxaliplatin. Modulating this response could therefore be of much wider interest in anti-tumour therapy.

La réplication de l'ADN est un processus extrêmement complexe, impliquant des milliers de fourches de réplication progressant le long des chromosomes. Ces fourches sont fréquemment ralenties ou arrêtées par différents obstacles, tels que des structures secondaires de l'ADN, des protéines agissant sur la chromatine ou encore un manque de nucléotides. Ce ralentissement, qualifié de stress réplicatif, joue un rôle central dans le développement tumoral. Des processus complexes, qui ne sont pas encore totalement connus, sont mis en place pour répondre à ce stress. Certaines nucléases, comme MRE11 et DNA2, dégradent l'ADN néorépliqué au niveau des fourches bloquées, ce qui permet le redémarrage des réplisomes. La voie interféron est un mécanisme de défense contre les agents pathogènes qui détecte la présence d'acides nucléiques étrangers dans le cytoplasme et active la réponse immunitaire innée. Des fragments d'ADN issus du métabolisme de l'ADN génomique (réparation, rétrotransposition) peuvent diffuser dans le cytoplasme et activer cette voie. Une manifestation pathologique de ce processus est le syndrome d'Aicardi-Goutières, une maladie rare caractérisée par une inflammation chronique générant des problèmes neurodégénératifs et développementaux. Dans le cadre de cette encéphalopathie, il a été suggéré que la réplication de l'ADN pouvait générer des fragments d'ADN cytosoliques, mais les mécanismes impliqués n'avaient pas été caractérisés. SAMHD1 est fréquemment muté dans le syndrome d'Aicardi-Goutières ainsi que dans certains cancers, mais son rôle dans l'étiologie de ces maladies était jusqu'à présent largement inconnu. Nous montrons que de l'ADN cytosolique s'accumule dans les cellules déficientes pour SAMHD1, particulièrement en présence de stress réplicatif, activant la réponse interféron. Par ailleurs, SAMHD1 est important pour la réplication de l'ADN en conditions normales et pour le processing des fourches arrêtées, indépendamment de son activité dNTPase. De plus, SAMHD1 stimule l'activité exonucléase de MRE11 in vitro. Lorsque SAMHD1 est absent, la dégradation de l'ADN néosynthétisé est inhibée, ce qui empêche l'activation du checkpoint de réplication et entraine un défaut de redémarrage des fourches de réplication. De plus, la résection des fourches de réplication est réalisée par un mécanisme alternatif qui libère des fragments d'ADN dans le cytosol, activant la réponse interféron. Les résultats obtenus montrent, pour la première fois, un lien direct entre la réponse au stress réplicatif et la production d'interférons. Ces résultats ont des conséquences importantes dans notre compréhension du syndrome d'Aicardi Goutières et des cancers liés à SAMHD1. Par exemple, nous avons démontré que MRE11 et RECQ1 sont responsables de la production des fragments d'ADN qui déclenchent la réponse inflammatoire dans les cellules déficientes pour SAMHD1. Nous pouvons donc imaginer que bloquer l'activité de ces enzymes pourrait diminuer la production des fragments d'ADN et, in fine, l'activation de l'immunité innée dans ces cellules. Par ailleurs, la voie interférons joue un rôle essentiel dans l'efficacité thérapeutique de l'irradiation et de certains agents chimiothérapiques comme l'oxaliplatine. Moduler cette réponse pourrait donc avoir un intérêt beaucoup plus large en thérapie anti-tumorale.

Health inequalities in old age: the relative contribution of material, behavioral and psychosocial factors in a German sample.

Background: Whereas the association between education and health in later life is well described, investigations about the underlying mechanisms of these health inequalities are scarce. This study examines the relative contribution of material, behavioral and psychosocial factors to health inequalities in older Germans. Methods: Data were drawn from the fifth wave of the Survey of Health, Ageing and Retirement in Europe (SHARE). The analytic sample included 3246 participants aged 60-85 years. We examined the independent and indirect contribution of material, behavioral and psychosocial factors to the association between education and self-rated health based on logistic regression models. Results: Material factors were most important as they were additionally working through behavioral and psychosocial factors whereas the independent contribution of behavioral and psychosocial factors was much lower than suggested in the separate analyses of the three explanatory pathways. Conclusions: Policy interventions that focus on the improvement of material living conditions might reduce health inequalities in old age. In studies on the underlying mechanisms of health inequalities, material, behavioral and psychosocial factors should be modeled as inter-related predictors as the separate analysis does not reveal their actual contribution so that the relevance of single explanatory pathways might be overestimated.

Analgesic doses of intrathecal but not intravenous clonidine increase acetylcholine in cerebrospinal fluid in humans.

Epidural clonidine increases acetylcholine (ACh) concentrations in cerebrospinal fluid (CSF) in humans, and experiments in animals support a cholinergic link in spinal alpha2-adrenoceptor-mediated antinociception. The purpose of the present study was to evaluate whether intravenous (I.V.) clonidine is also able to increase CSF ACh in humans. Accordingly, we studied 20 patients scheduled for resection of an acoustic neuroma under general anesthesia. Anesthesia was induced with propofol and maintained with propofol and N2O. After induction, an intrathecal catheter was inserted at the L3-4 interspace. Patients were then assigned, in a random, blind manner to receive either a bolus of 1 microg/kg intrathecal (I.T.) clonidine and an I.V. infusion of saline (n = 10) or an I.V. infusion of 4 microg/kg clonidine given in 20 min and an I.T. injection of saline (n = 10). CSF samples for ACh and clonidine concentration determination were drawn immediately before I.T. injection (time -20), at the end of the I.V. injection (time 0), then every 10 min thereafter. CSF ACh concentrations were determined by high-pressure liquid chromatography and CSF clonidine by radioimmunoassay. There was no significant difference between the groups with respect to age, gender, weight, and ASA physical status. I.T. but not I.V. administration of clonidine increased the CSF ACh concentration. We conclude that I.V. administration of four times the dose of clonidine delivered spinally failed to induce a significant increase of ACh in the CSF. These observations indicate that the analgesic effects observed after I.V. clonidine administration are not mediated by a cholinergic mechanism at the spinal level.