A SAGE-based screen for genes expressed in sub-populations of neurons in the mouse dorsal root ganglion

<p>Abstract</p> <p>Background</p> <p>The different sensory modalities temperature, pain, touch and muscle proprioception are carried by somatosensory neurons of the dorsal root ganglia. Study of this system is hampered by the lack of molecular markers for many of these neuronal sub-types. In order to detect genes expressed in sub-populations of somatosensory neurons, gene profiling was carried out on wild-type and TrkA mutant neonatal dorsal root ganglia (DRG) using SAGE (serial analysis of gene expression) methodology. Thermo-nociceptors constitute up to 80 % of the neurons in the DRG. In TrkA mutant DRGs, the nociceptor sub-class of sensory neurons is lost due to absence of nerve growth factor survival signaling through its receptor TrkA. Thus, comparison of wild-type and TrkA mutants allows the identification of transcripts preferentially expressed in the nociceptor or mechano-proprioceptor subclasses, respectively.</p> <p>Results</p> <p>Our comparison revealed 240 genes differentially expressed between the two tissues (P < 0.01). Some of these genes, CGRP, Scn10a are known markers of sensory neuron sub-types. Several potential markers of sub-populations, Dok4, Crip2 and Grik1/GluR5 were further analyzed by quantitative RT-PCR and double labeling with TrkA,-B,-C, c-ret, parvalbumin and isolectin B4, known markers of DRG neuron sub-types. Expression of Grik1/GluR5 was restricted to the isolectin B4+ nociceptive population, while Dok4 and Crip2 had broader expression profiles. Crip2 expression was however excluded from the proprioceptor sub-population.</p> <p>Conclusion</p> <p>We have identified and characterized the detailed expression patterns of three genes in the developing DRG, placing them in the context of the known major neuronal sub-types defined by molecular markers. Further analysis of differentially expressed genes in this tissue promises to extend our knowledge of the molecular diversity of different cell types and forms the basis for understanding their particular functional specificities.</p

A transcription factor code defines nine sensory interneuron subtypes in the mechanosensory area of the spinal cord

Interneurons in the dorsal spinal cord process and relay innocuous and nociceptive somatosensory information from cutaneous receptors that sense touch, temperature and pain. These neurons display a well-defined organization with respect to their afferent innervation. Nociceptive afferents innervate lamina I and II, while cutaneous mechanosensory afferents primarily innervate sensory interneurons that are located in lamina III-IV. In this study, we outline a combinatorial transcription factor code that defines nine different inhibitory and excitatory interneuron populations in laminae III-IV of the postnatal cord. This transcription factor code reveals a high degree of molecular diversity in the neurons that make up laminae III-IV, and it lays the foundation for systematically analyzing and manipulating these different neuronal populations to assess their function. In addition, we find that many of the transcription factors that are expressed in the dorsal spinal cord at early postnatal times continue to be expressed in the adult, raising questions about their function in mature neurons and opening the door to their genetic manipulation in adult animals

Crude oil to chemicals: light olefins from crude oil

[EN] The possibility to fulfill the increasing market demand and producers' needs in processing crude oil, a cheap and universally available feedstock, to produce petrochemicals appears to be a very attractive strategy. Indeed, many petrochemicals are produced as side streams during crude oil refining, which primary goal remains transportation fuel production. Availability of some critical feedstocks may then depend on local refining policy. In order to improve flexibility, it has been proposed to directly crack crude oil to produce petrochemicals, in particular light olefins (ethylene, propylene, butenes), using technologies derived from fluid catalytic cracking. This paper attempts to review the main research works done on the topic in the literature in the last five decades, focussing on process as well as catalyst technology, with a special interest for fluid catalytic cracking (FCC) based technology that can be used towards maximizing chemicals from crude oil. Factors investigated include use of severe cracking conditions, on-purpose additives (from ZSM5 to more exotic, metal doped additives), recycle streams and multiple riser systems.The authors thank Saudi Aramco for material and financial support. Financial support by the Spanish Government-MINECO through program "Severo Ochoa" (SEV 2012-0267), Consolider Ingenio (2010-Multicat, CSD-2009-0050), MAT2012-31657, CTQ2015-70126-R (MINECO/FEDER), by the European Union through ERC-AdG-2014-671093-SynCatMatch and by the Generalitat Valenciana through the Prometeo program (PROMETEOII/2013/011) is also acknowledged.Corma Canós, A.; Corresa Mateu, E.; Mathieu ., Y.; Sauvanaud ., LL.; Al-Bogami, S.; Al-Ghrami, M.; Bourane, A. (2017). Crude oil to chemicals: light olefins from crude oil. Catalysis Science & Technology. 7(1):12-46. https://doi.org/10.1039/c6cy01886fS12467

A Systematic Approach to Assess the Activity and Classification of PCSK9 Variants

Background: Gain of function (GOF) mutations of PCSK9 cause autosomal dominant familial hypercholesterolemia as they reduce the abundance of LDL receptor (LDLR) more efficiently than wild-type PCSK9. In contrast, PCSK9 loss of function (LOF) variants are associated with a hypocholesterolemic phenotype. Dozens of PCSK9 variants have been reported, but most remain of unknown significance since their characterization has not been conducted. Objective: Our aim was to make the most comprehensive assessment of PCSK9 variants and to determine the simplest approach for the classification of these variants. Methods: The expression, maturation, secretion, and activity of nine well-established PCSK9 variants were assessed in transiently transfected HEK293 cells by Western blot and flow cytometry. Their extracellular activities were determined in HepG2 cells incubated with the purified recombinant PCSK9 variants. Their binding affinities toward the LDLR were determined by solid-phase immunoassay. Results: LDLR expression increased when cells were transfected with LOF variants and reduced when cells were transfected with GOF variants compared with wild-type PCSK9. Extracellular activities measurements yielded exactly similar results. GOF and LOF variants had increased, respectively reduced, affinities for the LDLR compared with wild-type PCSK9 with the exception of one GOF variant (R218S) that showed complete resistance to inactivation by furin. All variants were expressed at similar levels and underwent normal maturation and secretion patterns except for two LOF and two GOF mutants. Conclusions: We propose that transient transfections of HEK293 cells with a plasmid encoding a PCSK9 variant followed by LDLR expression assessment by flow cytometry is sufficient to reliably determine its GOF or LOF status. More refined experiments should only be used to determine the underlying mechanism(s) at hand.This work was supported by the Basque Government (Grupos Consolidados IT-1264-19). GL is supported by the Agence Nationale de la Recherche (Paris, France) Program Grant CHOPIN (CHolesterol Personalized Innovation) ANR-16-RHUS-0007 and Project Grant KRINGLE2 ANR-20-CE14-0009 as well as by La Fondation De France (FDF-00096274). U.G-G. was supported by Fundación Biofísica Bizkaia. A.B.-V. was supported by Programa de especialización de Personal Investigador Doctor en la UPV/EHU (2019) 2019-2020. A.L.-S. was supported by a grant PIF (2019–2020), Gobierno Vasco, and partially supported by Fundación Biofísica Bizkaia. KC and AKJ received a scholarship from the European Union (European Regional Development Fund INTERREG V) and the Région Réunion (Saint-Denis, Réunion, France)

Regulation of the Na,K-ATPase Gamma-Subunit FXYD2 by Runx1 and Ret Signaling in Normal and Injured Non-Peptidergic Nociceptive Sensory Neurons

Dorsal root ganglia (DRGs) contain the cell bodies of sensory neurons which relay nociceptive, thermoceptive, mechanoceptive and proprioceptive information from peripheral tissues toward the central nervous system. These neurons establish constant communication with their targets which insures correct maturation and functioning of the somato-sensory nervous system. Interfering with this two-way communication leads to cellular, electrophysiological and molecular modifications that can eventually cause neuropathic conditions. In this study we reveal that FXYD2, which encodes the gamma-subunit of the Na,K-ATPase reported so far to be mainly expressed in the kidney, is induced in the mouse DRGs at postnatal stages where it is restricted specifically to the TrkB-expressing mechanoceptive and Ret-positive/IB4-binding non-peptidergic nociceptive neurons. In non-peptidergic nociceptors, we show that the transcription factor Runx1 controls FXYD2 expression during the maturation of the somato-sensory system, partly through regulation of the tyrosine kinase receptor Ret. Moreover, Ret signaling maintains FXYD2 expression in adults as demonstrated by the axotomy-induced down-regulation of the gene that can be reverted by in vivo delivery of GDNF family ligands. Altogether, these results establish FXYD2 as a specific marker of defined sensory neuron subtypes and a new target of the Ret signaling pathway during normal maturation of the non-peptidergic nociceptive neurons and after sciatic nerve injury

Semi-intact ex vivo approach to investigate spinal somatosensory circuits

The somatosensory input that gives rise to the perceptions of pain, itch, cold and heat are initially integrated in the superficial dorsal horn of the spinal cord. Here, we describe a new approach to investigate these neural circuits in mouse. This semi-intact somatosensory preparation enables recording from spinal output neurons, while precisely controlling somatosensory input, and simultaneously manipulating specific populations of spinal interneurons. Our findings suggest that spinal interneurons show distinct temporal and spatial tuning properties. We also show that modality selectivity — mechanical, heat and cold — can be assessed in both retrogradely labeled spinoparabrachial projection neurons and genetically labeled spinal interneurons. Finally, we demonstrate that interneuron connectivity can be determined via optogenetic activation of specific interneuron subtypes. This new approach may facilitate key conceptual advances in our understanding of the spinal somatosensory circuits in health and disease

The cellular and synaptic architecture of the mechanosensory dorsal horn

The deep dorsal horn is a poorly characterized spinal cord region implicated in processing low-threshold mechanoreceptor (LTMR) information. We report an array of mouse genetic tools for defining neuronal components and functions of the dorsal horn LTMR-recipient zone (LTMR-RZ), a role for LTMR-RZ processing in tactile perception, and the basic logic of LTMR-RZ organization. We found an unexpectedly high degree of neuronal diversity in the LTMR-RZ: seven excitatory and four inhibitory subtypes of interneurons exhibiting unique morphological, physiological, and synaptic properties. Remarkably, LTMRs form synapses on between four and 11 LTMR-RZ interneuron subtypes, while each LTMR-RZ interneuron subtype samples inputs from at least one to three LTMR classes, as well as spinal cord interneurons and corticospinal neurons. Thus, the LTMR-RZ is a somatosensory processing region endowed with a neuronal complexity that rivals the retina and functions to pattern the activity of ascending touch pathways that underlie tactile perception

Direct crude oil cracking for producing chemicals: Thermal cracking modeling

[EN] The direct cracking of crude oil is an interesting option for producing cheaply large amounts of petrochemicals. This may be carried out with catalyst and equipment similar to that of catalytic cracking, but at a temperature range between that of standard catalytic cracking and steam cracking. Thermal cracking will play a role in the conversion, but is rarely disclosed in experimental or modeling work. Thus, a crude oil and its fractions were thermally cracked and the products yields were modeled using a 9 lumps cracking scheme. It was found that heavy fraction cracks twice as fast as diesel fraction and ten times faster than gasoline fraction, with activation energies in the 140-200 kJ/mol range. Selectivity to ethylene, propylene and butenes were found similar in the operating range explored.The authors thank Saudi Aramco for its material and financial support. Financial support by the Spanish Government-MINECO through programs "Severo Ochoa" (SEV 2012-0267) and CTQ2015-70126-R and by the Generalitat Valenciana through the Prometeo program (PROMETEOII/2013/011) is also acknowledged.Corma Canós, A.; Sauvanaud, LL.; Mathieu, Y.; Al-Bogami, S.; Bourane, A.; Al-Ghrami, M. (2018). Direct crude oil cracking for producing chemicals: Thermal cracking modeling. Fuel. 211:726-736. https://doi.org/10.1016/j.fuel.2017.09.099S72673621

New Gas-Phase Catalytic Oxidative Processes for Desulfurization of Diesel Fuel

An effective gas-phase oxidative desulfurization (ODS) process was proposed. The process was studied in a laboratory reactor with a proprietary catalyst at 300-400 ºС and ambient pressure with model fuels represented by thiophene, dibenzothiophene(DBT) and 4,6-dimethyldibenzothiophene (DMDBT) dissolved in octane, isooctane or toluene. The reactivity of different sulfur containing molecules in ODS was shown to increase in the sequence: thiophene<DBT<DMDBT. The main sulfur containing product of oxidation of these compounds was SO2. During the gas-phase ODS both processes of sulfur species oxidation and processes of their adsorption were observed and studied. Based on the conducted studies, different ODS process designs comprising its integration with adsorption and regeneration processes and with conventional hydrodesulfurization (HDS) process were proposed. One scheme is based on alternating regimes of ODS and catalyst regeneration in two reactors: sulfur is removed from organic feedstock by oxidation and adsorption in one reactor while simultaneous regeneration of the catalyst that has accumulated sulfur compounds takes place in another reactor. Two other schemes are based on joint use of ODS and HDS. The conventional HDS process is most effective for removal of low-boiling sulfur containing compounds reactive with respect to hydrogen, while removal of refractory sulfur compounds, such as DMDBT is more easily achieved by gas phase ODS. Thus the combination of these processes is expected to be most efficient for deep desulfurization of diesel fuel