Tandem quadruplication of HMA4 in the zinc (Zn) and cadmium (Cd) hyperaccumulator noccaea caerulescens

Zinc (Zn) and cadmium (Cd) hyperaccumulation may have evolved twice in the Brassicaceae, in Arabidopsis halleri and in the Noccaea genus. Tandem gene duplication and deregulated expression of the Zn transporter, HMA4, has previously been linked to Zn/Cd hyperaccumulation in A. halleri. Here, we tested the hypothesis that tandem duplication and deregulation of HMA4 expression also occurs in Noccaea. A Noccaea caerulescens genomic library was generated, containing 36,864 fosmid pCC1FOSTM clones with insert sizes ~20–40 kbp, and screened with a PCR-generated HMA4 genomic probe. Gene copy number within the genome was estimated through DNA fingerprinting and pooled fosmid pyrosequencing. Gene copy numbers within individual clones was determined by PCR analyses with novel locus specific primers. Entire fosmids were then sequenced individually and reads equivalent to 20-fold coverage were assembled to generate complete whole contigs. Four tandem HMA4 repeats were identified in a contiguous sequence of 101,480 bp based on sequence overlap identities. These were flanked by regions syntenous with up and downstream regions of AtHMA4 in Arabidopsis thaliana. Promoter-reporter b-glucuronidase (GUS) fusion analysis of a NcHMA4 in A. thaliana revealed deregulated expression in roots and shoots, analogous to AhHMA4 promoters, but distinct from AtHMA4 expression which localised to the root vascular tissue. This remarkable consistency in tandem duplication and deregulated expression of metal transport genes between N. caerulescens and A. halleri, which last shared a common ancestor >40 mya, provides intriguing evidence that parallel evolutionary pathways may underlie Zn/Cd hyperaccumulation in Brassicaceae

Cannabinoid CB2 receptor (CB2R) stimulation delays rubrospinal mitochondrial-dependent degeneration and improves functional recovery after spinal cord hemisection by ERK1/2 inactivation.

Spinal cord injury (SCI) is a devastating condition of CNS that often results in severe functional impairments for which there are no restorative therapies. As in other CNS injuries, in addition to the effects that are related to the primary site of damage, these impairments are caused by degeneration of distal regions that are connected functionally to the primary lesion site. Modulation of the endocannabinoid system (ECS) counteracts this neurodegeneration, and pharmacological modulation of type-2 cannabinoid receptor (CB2R) is a promising therapeutic target for several CNS pathologies, including SCI. This study examined the effects of CB2R modulation on the fate of axotomized rubrospinal neurons (RSNs) and functional recovery in a model of spinal cord dorsal hemisection (SCH) at the cervical level in rats. SCH induced CB2R expression, severe atrophy, and cell death in contralateral RSNs. Furthermore, SCH affected molecular changes in the apoptotic cascade in RSNs - increased cytochrome c release, apoptosome formation, and caspase-3 activity. CB2R stimulation by its selective agonist JWH-015 significantly increased the bcl-2/bax ratio, reduced cytochrome c release, delayed atrophy and degeneration, and improved spontaneous functional recovery through ERK1/2 inactivation. These findings implicate the ECS, particularly CB2R, as part of the endogenous neuroprotective response that is triggered after SCI. Thus, CB2R modulation might represent a promising therapeutic target that lacks psychotropic effects and can be used to exploit ECS-based approaches to counteract neuronal degeneration

The Legal Landscape for Advanced Therapies: Material and Institutional Implementation of European Union Rules in France and the United Kingdom

In 2007, the European Union adopted a lex specialis, Regulation (EC) No. 1394/2007 on advanced therapy medicinal products (ATMPs), a new legal category of medical product in regenerative medicine. The regulation applies to ATMPs prepared industrially or manufactured by a method involving an industrial process. It also provides a hospital exemption, which means that medicinal products not regulated by EU law do not benefit from a harmonized regime across the European Union but have to respect national laws. This article describes the recent EU laws, and contrasts two national regimes, asking how France and the United Kingdom regulate ATMPs which do and do not fall under the scope of Regulation (EC) No. 1394/2007. What are the different legal categories and their enforceable regimes, and how does the evolution of these highly complex regimes interact with the material world of regenerative medicine and the regulatory bodies and socioeconomic actors participating in it

Analyse du principe d'Équivalence en Substance en tant que concept directeur pour l'évaluation de l'innocuité des végétaux génétiquement modifiés

Le principe d'Équivalence en Substance (É.S.) est développé en 1993 par l'OCDE et implémenté en 1996 par la FAO/OMS, pour remédier au manque de régulation au niveau de la gestion des risques concernant les végétaux à caractère nouveau (VCN). Ce concept est un outil de comparaison des composantes biochimiques permettant de faire le parallèle entre un produit transgénique et un homologue issu de l'agriculture traditionnelle, dont l'innocuité est prouvée; une É.S. permet alors de conclure à l'innocuité du VCN. Cependant, il semble difficile de parvenir à établir une équivalence alors qu'aucun critère n'est défini. La substance qui doit être équivalente n'est pas non plus déterminée et peut étymologiquement porter à confusion. Comment comprendre ce concept devant le paradoxe qu'est la vente d'un produit"similaire, mais différent"? Ce principe est très limité. Il a été démontré que seule, la composition biochimique ne peut définir la toxicité d'un aliment, il faut alors chercher ailleurs que là vers où nous guide l'É.S. L'homme entretient aussi une dynamique sociale particulière avec la nourriture. C'est pourquoi d'autres aspects tels que l'équivalence éthique et l'équivalence quantitative doivent être pris en compte pour évaluer un VCN. Comment alors doit-on comprendre ce principe? Ne l'avons-nous tout simplement pas mal interprété et utilisé de manière erronée en tant que processus plutôt qu'en tant que simple outil? Certes, mais il est aussi insuffisant et demande à être échangé au profit d'une méthode plus"pratique"

Distinct regulation of nNOS and iNOS by CB2 receptor in remote delayed neurodegeneration

Hemicerebellectomy results in remote delayed degeneration of precerebellar neurons. We have reported that such a lesion induces type 2 cannabinoid receptor (CB(2)) expression in precerebellar neurons and that stimulation of CB(2), but not CB(1), has neuroprotective effects. In this study, we found that in the same model, the CB(2) agonist JWH-015 enhances neuronal nitric oxide synthase (nNOS) expression in axotomized neurons and that CB(2)-mediated neuroprotection is abrogated by pharmacological inhibition of nNOS. JWH-015 prevented the axotomy-induced upregulation of inducible NOS (iNOS) in astrocytes but had no effect on endothelial NOS (eNOS). In addition, we observed that JWH-015 significantly reduces hemicerebellectomy-induced neuroinflammatory responses and oxidative/nitrative stress. With regard to the signaling pathways of CB(2)/nNOS-mediated neuroprotection, we noted nNOS-dependent modulation of the expression of anti-oxidative (Hsp70) and anti-apoptotic (Bcl-2) proteins. These findings shed light on the interactions between the endocannabinoid and nitrergic systems after focal brain injury, implicating distinct functions of nNOS activation and iNOS inhibition in CB(2) signaling, which protect neurons from axotomy-induced cell death

Biodegradation of polycyclic aromatic hydrocarbons in the surface ocean

Tesi en modalitat de compendi de publicacionsApèndix "Women in Science: Scientific contributions of female scientists relevant to this thesis" p. 320-321Increasing quantities of organic pollutants (OP) are being released to the environment, posing a threat to Earth’s life system. In the marine environment, OP pollution caused by oil spill accidents receives a lot of academic and societal attention. However, the magnitude of semi-volatiles OP introduced by atmospheric deposition and by maritime currents is orders of magnitude larger. Little is known, however, about the cycling of these background OP in the oceans and their effects to marine ecosystems. It is believed that an important sink of OP in the marine environment must be microbial biodegradation, since it is widely recognized their capacity to consume many OPs. However, neither the magnitude of biodegradation, the identity of the main degraders nor OP effects to microorganisms, is known. Polycyclic aromatic hydrocarbons (PAH) and other semi-volatile hydrocarbons are among the most abundant OP in the marine environment. Their wide spectrum of physicochemical properties make PAH family an ideal surrogate to study the biogeochemistry of OP in seawater. This thesis focuses on the biodegradation activities under background concentrations of PAH in the upper ocean, and the co-occurring microbial responses to this exposure. The goals of this work are: 1) to get insights into microbial PAH biodegradation under realistic conditions in different upper ocean environments by means of biogeochemical, molecular and genomic approaches, 2) identify the main players and describe the main metabolisms co-occurring along with biodegradation in the interaction between PAH and microorganisms by means of physiological measurements and metatranscriptomic approaches, and 3) localize marine hot spots of PAH biodegradation and describe some of the main physicochemical influencing factors of biodegradation. For these purposes we did short term incubations with background concentrations of PAH, mimicking those conditions found in the surface ocean in several oceanic provinces with contrasted physicochemical conditions. We used molecular biology and analytical chemistry techniques to calculate biodegradation taxes and evaluate the changes in the composition and genetic expression profiles of microbial communities exposed to PAH ambient levels. We found that PAH microbial metabolic capacity is a widespread trait in the global upper ocean although PAH biodegradation rates span in a wide range of values. These findings strongly suggest a fundamental role of microbial degradation as a relevant driver of PAH fate in the upper ocean. We observed that PAH at background concentrations after short term exposure (24-48 hrs) had an impact on microbial communities compositions and functionality. The main taxa responsible for PAH biodegradation at background concentrations varied depending on the site and the habitat, and included many different groups. The battery of genes expressed during PAH exposure included PAH degrading genes plus genes involved in stress response and detoxifying strategies among others. Comparison of results from the different sites identified new environmental drivers affecting the efficiency of PAH biodegradation in the upper ocean beyond those already described and those that are commonly overlooked: bacterial life-style (PA vs free-living), microbial community pre-exposure history to organic pollutants and the habitat (specially the SML). Overall, this thesis adds new information and field-based evidence regarding the microbial controls over the fate and transport of the background PAH concentrations in the surface ocean. In a context of increasing emissions and global change, it becomes crucial to be able to forecast the fate of OP once in the ocean. This requires understanding at a mechanistic level how do microbial metabolic processes and the pool of OP in the ocean intertwine, in order to elucidate a more updated, and always evolving, global carbon cycle.Quantitats creixents de contaminants orgànics (CO) s’alliberen al medi ambient, cosa que suposa una amenaça per al sistema de vida de la Terra. En el medi marí, la contaminació causada per vessaments de petroli accidentals rep molta atenció tant acadèmica com social. No obstant això, la magnitud de la contaminació per CO semi-volàtils que arriba als oceans de manera difusa és ordres de magnitud més gran en comparació als vessaments accidentals. Actualment hi ha poca informació sobre el cicle d'aquests CO a la columna d’aigua i sobre quins són els efectes que causen sobre els ecosistemes marins. Es creu que un destí important dels CO al medi marí són els processos de biodegradació per part dels microorganismes, ja que és àmpliament reconeguda la seva capacitat per consumir molts CO. Tanmateix, no es coneix ni la magnitud de la biodegradació ni la identitat dels principals degradadors, ni tampoc els efectes dels CO als microorganismes. Els hidrocarburs aromàtics policíclics (HAP) i altres hidrocarburs semi-volàtils es troben entre els CO més abundants en el medi marí. L’ampli espectre de propietats fisicoquímiques dels HAP els converteixen en un model ideal per estudiar la biogeoquímica dels CO a l'aigua de mar. Aquesta tesi es centra en les activitats de biodegradació sota concentracions ambientals realistes de HAP a les aigües marines superficials i les respostes microbianes concomitants a aquesta exposició. Els objectius d'aquest treball són: 1) explorar la biodegradació microbiana de HAP en condicions realistes en diferents entorns d’aigües superficials marines mitjançant tècniques biogeoquímiques, moleculars i genòmiques, 2) identificar els principals actors de la degradació i descriure els principals metabolismes que coexisteixen juntament amb la biodegradació mitjançant mesures fisiològiques i tècniques metatranscriptòmiques i 3) localitzar “punts calents” de biodegradació de HAP en hàbitats marins i descriure alguns dels principals factors fisicoquímics que influeixen en la biodegradació. Per tal d’assolir els objectius, hem realitzat incubacions de curta durada amb aigua superficial marina amb nivells de HAP ambientals, imitant les concentracions que es troben a l'oceà superficial en diverses províncies oceàniques i amb condicions fisicoquímiques contrastades. En particular, el treball de camp es va desenvolupar a l'Antàrtida Marítima, al NE del Pacífic subàrtic i al NO del Mediterrani. Es va utilitzar tècniques de química analítica i biologia molecular per obtenir les taxes de biodegradació i avaluar els canvis en la composició i els perfils d'expressió gènica de les comunitats microbianes exposades a nivells ambientals de HAP. Els resultats indiquen que la capacitat microbiana de biodegradació dels HAP és un tret estès a l'oceà superficial a escala global, tot i que les taxes de biodegradació dels HAP tenen una gran variabilitat. Aquestes troballes suggereixen el paper fonamental de la biodegradació microbiana com a un del principals processos que modulen el destí dels HAP a l'oceà superficial. En els experiments d’incubació vam observar que les exposicions a curt termini (24-48 hores) amb concentracions ambientals de HAP tenen un impacte en la composició i la funcionalitat de les comunitats microbianes. Els principals tàxons responsables de la biodegradació dels HAP a concentracions ambientals variaven segons el lloc i l'hàbitat, i inclouen molts grups diferents. Durant els experiments de curta exposició les poblacions de bacteris hidrocarbonoclàstics (BHC), és a dir, els bacteris especialitzats en l'ús d'hidrocarburs com a font de carboni i energia, van esdevenir actors clau en la biodegradació. La bateria de gens expressats durant l'exposició als HAP inclou gens de degradació de HAP, a banda d’altres gens implicats en donar resposta a l'estrès i altres estratègies de desintoxicació.Postprint (published version

Regulation of genetically engineered (GE) mosquitoes as a public health tool: a public health ethics analysis

Background: In recent years, genetically engineered (GE) mosquitoes have been proposed as a public health measure against the high incidence of mosquito-borne diseases among the poor in regions of the global South. While uncertainties as well as risks for humans and ecosystems are entailed by the open-release of GE mosquitoes, a powerful global health governance non-state organization is funding the development of and advocating the use of those bio-technologies as public health tools. In August 2016, the US Food and Drug Agency (FDA) approved the uncaged field trial of a GE Aedes aegypti mosquito in Key Haven, Florida. The FDA’s decision was based on its assessment of the risks of the proposed experimental public health research project. The FDA is considered a global regulatory standard setter. So, its approval of the uncaged field trial could be used by proponents of GE mosquitoes to urge countries in the global South to permit the use of those bio-technologies. Method: From a public health ethics perspective, this paper evaluates the FDA’s 2016 risk assessment of the proposed uncaged field trial of the GE mosquito to determine whether it qualified as a realistic risk evaluation. Results: The FDA’s risk assessment of the proposed uncaged field trial did not proximate the conditions under which the GE mosquitoes would be used in regions of the global South where there is a high prevalence of mosquito-borne diseases. Conclusion: Given that health and disease have political-economic determinants, whether a risk assessment of a product is realistic or not particularly matters with respect to interventions meant for public health problems that disproportionately impact socio-economically marginalized populations. If ineffective public health interventions are adopted based on risk evaluations that do not closely mirror the conditions under which those products would actually be used, there could be public health and ethical costs for those populations