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Abstract Study of germination using fresh seeds and seeds from an aerial seed bank of a young Verbascum phlomoides population revealed the existence of primary dormancy that blocks immediate germination and of substantial variations in the subsequent germination capability. The heterogeneity of desynchronized individual responses in a species with a high germination capacity would make it possible for the descendants of one or more individuals to occupy a disturbed site. Exposure to cold followed by a return to the light accelerating germinative response would appear to enhance late spring germination. [ABSTRACT FROM AUTHOR] more...

Background: The use of morphometric implants in total knee arthroplasty (TKA) has shown better early clinical outcomes compared to conventional implants. The primary objective of this study is to evaluate the functional outcome and the implant survivorship of a morphometric TKA at a minimum of 5 years of follow-up., Methods: From May 2012 to June 2015, all patients undergoing primary TKA with a single design of morphometric posterior-stabilized prosthesis (Persona; Zimmer) in a prospective observational single-center study were evaluated. The Knee Society Scoring System (KSS) and the Knee Injury and Osteoarthritis Outcome Score (KOOS) were completed preoperatively, 1, 3, and 5 years postoperatively. Kaplan-Meier was used to calculate survivorship of the implants. The average follow-up was 75 months., Results: In total, 237 TKAs were performed in 235 patients with a mean age of 73 years (49-90). The KSS Knee Score increased from 44.7 (13-64) preoperatively to 93.6 (71-100), the KSS Function Score from 45.8 (17-69) to 92.2 (51-98), and the KSS Satisfaction Score from 26.6 (16-51) to 41 (35-55) at 5 years of follow-up. Similarly, for the KOOS score, a significant improvement of all the subscales was observed at 5 years of follow-up. Implant survival without reoperation at 5 years of follow up was 98.72% (95% confidence interval 0.95-1.00)., Conclusion: This is the first study demonstrating that significant improvements of the functional scores with good survivorship can be achieved at a minimum of 5 years of follow-up with TKA using morphometric implants., (Copyright © 2021 Elsevier Inc. All rights reserved.) more...

Neurons innervating peripheral tissues display complex responses to peripheral nerve injury. These include the activation and suppression of a variety of signalling pathways that together influence regenerative growth and result in more or less successful functional recovery. However, these responses can be offset by pathological consequences including neuropathic pain. Calcium signalling plays a major role in the different steps occurring after nerve damage. As part of our studies to unravel the roles of injury-induced molecular changes in dorsal root ganglia (DRG) neurons during their regeneration, we show that the calcium calmodulin kinase CaMK1a is markedly induced in mouse DRG neurons in several models of mechanical peripheral nerve injury, but not by inflammation. Intrathecal injection of NRTN or GDNF significantly prevents the post-traumatic induction of CaMK1a suggesting that interruption of target derived factors might be a starter signal in this de novo induction. Inhibition of CaMK signalling in injured DRG neurons by pharmacological means or treatment with CaMK1a siRNA resulted in decreased velocity of neurite growth in vitro. Altogether, the results suggest that CaMK1a induction is part of the intrinsic regenerative response of DRG neurons to peripheral nerve injury, and is thus a potential target for therapeutic intervention to improve peripheral nerve regeneration. more...

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. more...

Growing evidence indicates that collagens perform crucial functions during the development and organization of the nervous system. Collagen XXVIII is a recently discovered collagen almost exclusively expressed in the peripheral nervous system (PNS). In this study, we show that this collagen is associated with nonmyelinated regions of the PNS. With the notable exception of type II terminal Schwann cell in the hairy skin, collagen XXVIII surrounds all nonmyelinating glial cells studied. This includes satellite glial cells of the dorsal root ganglia, terminal Schwann cells type I around mechanoceptors in the skin, terminal Schwann cells around proprioceptors in the muscle spindle or at the neuromuscular junction and olfactory ensheathing cells. Collagen XXVIII is also detected at nodes of Ranvier where the myelin sheath of myelinated fibers is interrupted and is thus a distinctive component of the PNS nodal gap. The correlation between the absence of myelin and the presence of collagen XXVIII is confirmed in a mouse model of Charcot-Marie-Tooth characterized by dysmyelinated nerve fibers, in which enhancement of collagen XXVIII labeling is observed. more...

Low-threshold mechanoreceptor neurons (LTMs) of the dorsal root ganglia (DRG) are essential for touch sensation. They form highly specialized terminations in the skin and display stereotyped projections in the spinal cord. Functionally defined LTMs depend on neurotrophin signaling for their postnatal survival and functioning, but how these neurons arise during development is unknown. Here, we show that specific types of LTMs can be identified shortly after DRG genesis by unique expression of the MafA transcription factor, the Ret receptor and coreceptor GFRalpha2, and find that their specification is Ngn2 dependent. In mice lacking Ret, these LTMs display early differentiation defects, as revealed by reduced MafA expression, and at later stages their central and peripheral projections are compromised. Moreover, in MafA mutants, a discrete subset of LTMs display altered expression of neurotrophic factor receptors. Our results provide evidence that genetic interactions involving Ret and MafA progressively promote the differentiation and diversification of LTMs., (2009 Elsevier Inc. All rights reserved.) more...

Dorsal root ganglia (DRG) neurons exhibit a wide molecular heterogeneity in relation to the various sensory modalities (mechanoception, thermoception, nociception) that they subserve. Finding markers of subpopulations is an important step in understanding how these neurons convey specific information. We identified fibroblast growth factor homologous factor 1 (FHF1) in a search for markers of subpopulations of DRG neurons. FHFs constitute a family of four factors that share some structural properties with fibroblast growth factors (FGFs) but are functionally distinct. They are expressed in specific subsets of neurons and are involved in the modulation of sodium channel activity. The pattern of expression of FHF1 in the DRG was determined during development, in the adult and after axotomy. We show that in the adult, FHF1 is expressed in two populations, one composed of nociceptors and another in which no neurotrophic factor receptors were detected (panTrk-/c-Ret-). Interestingly, in the nociceptors, FHF1 expression was restricted to a subset of TrkA+/calcitonin gene-related peptide (CGRP)-positive neurons. Neurofilament 200 (NF-200) and peripherin labeling indicates that 70% of the FHF1-expressing neurons contribute to A-fibers and 30% to C-fibers. FHF1 interacts with the Na(v)1.9 sodium channel isoform, which is strongly expressed in cRet+/isolectin-B4 binding neurons, but we show that FHF1 is not expressed in the cRet+/IB4+ subclass and that it does not colocalize with Na(v)1.9. Our results argue strongly against the possibility that FHF1 has a modulatory effect on this channel in cRet+/IB4+ neurons, but FHF1 could play a role in a distinct subset of TrkA+/CGRP+ nociceptors., (Copyright 2008 Wiley-Liss, Inc.) more...

Neural stem cells cultured with fibroblast growth factor 2 (FGF2)/epidermal growth factor (EGF) generate clonal expansions called neurospheres (NS), which are widely used for therapy in animal models. However, their cellular composition is still poorly defined. Here, we report that NS derived from several embryonic and adult central nervous system (CNS) regions are composed mainly of remarkable cells coexpressing radial glia markers (BLBP, RC2, GLAST), oligodendrogenic/neurogenic factors (Mash1, Olig2, Nkx2.2), and markers that in vivo are typical of the oligodendrocyte lineage (NG2, A2B5, PDGFR-alpha). On NS differentiation, the latter remain mostly expressed in neurons, together with Olig2 and Mash1. Using cytometry, we show that in growing NS the small population of multipotential self-renewing NS-forming cells are A2B5(+) and NG2(+). Additionally, we demonstrate that these NS-forming cells in the embryonic spinal cord were initially NG2(-) and rapidly acquired NG2 in vitro. NG2 and Olig2 were found to be rapidly induced by cell culture conditions in spinal cord neural precursor cells. Olig2 expression was also induced in astrocytes and embryonic peripheral nervous system (PNS) cells in culture after EGF/FGF treatment. These data provide new evidence for profound phenotypic modifications in CNS and PNS neural precursor cells induced by culture conditions. more...

The non-genomic role of neuroactive steroids on [Ca2+]i transients induced by GABA receptor activation was investigated in cultured dorsal root ganglia (DRG) neurones at embryonic stage E13. [Ca2+]i measurements were performed with Fura-2 fast fluorescence microfluorimetry. Application of the GABAA receptor agonist muscimol (Musci) evoked an increase in [Ca2+]i, confirming the excitatory effect of GABA at this embryonic stage. The muscimol-induced [Ca2+]i response was inhibited by progesterone (Proges) and its primary metabolite allopregnanolone (Allo) in a rapid, reversible and dose-dependent manner. These calcium transients were suppressed in the absence of external Ca2+ or in the presence of Ni2+ + Cd2+ suggesting an involvement of voltage-activated Ca2+ channels. In contrast, none of these steroids affected the resting [Ca2+]i nor exhibited any inhibitory effect on 50 mM KCl-induced [Ca2+]i increases. In view of the well-established potentiation of GABAA receptor by direct binding of neurosteroids, the inhibitory effects described in this study seem to involve distinct mechanisms. This new inhibitory effect of progesterone is observed at low and physiological concentrations, is rapid and independent of RU38486, an antagonist of the classic progesterone receptor, probably involving a membrane receptor. Using RT-PCR, we demonstrated the expression of progesterone receptor membrane component 1 (Pgrmc1), encoding 25-Dx, a membrane-associated progesterone binding protein in DRG neurones at different stages of development. In conclusion, we describe for the first time a rapid effect of progestins on embryonic DRG neurones involving an antagonistic effect of progesterone and allopregnanolone on GABAA receptors. more...

In order to shed light on transcriptional networks involved in adult peripheral nerve repair program, we propose for the first time an organization of the transcriptional dynamics of the mouse dorsal root ganglia (DRG) following a sciatic nerve lesion. This was done by a non-hierarchical bioinformatical clustering of four Serial Analysis of Gene Expression libraries performed on DRG at embryonic day E13, neonatal day P0, adult and adult 3 days post-sciatic nerve section. Grouping genes according to their expression profiles shows that a combination of down-regulation of genes expressed at the adult stages, re-expression of embryonic genes and induction of a set of de novo genes takes place in injured neurons. Focusing on this latter event highlights Ddit3, Timm8b and Oazin as potential new injury-induced molecular actors involved in a stress response pathway. Their association with the traumatic state was confirmed by real-time PCR and in situ hybridization investigations. Clustering analysis allows us to distinguish developmental re-programming events from nerve-injury-induced processes and thus provides a basis for molecular understanding of transcriptional alterations taking place in the DRG after a sciatic nerve lesion. more...

Flavors are commonly used in pharmaceutical oral solutions and oral suspensions to mask drug bitterness and to make the formulation more palatable. Flavor analysis during product development is typically performed by human organoleptic analysis, which is often expensive and less objective. A novel approach using a metal oxide sensor-based instrument (electronic-nose) for headspace analysis was explored to replace human sensory perception for consistent qualitative and quantitative analysis of flavors in a pharmaceutical formulation. The use of the electronic-nose technique to qualitatively distinguish among six common flavoring agents (raspberry, red berry, strawberry, pineapple, orange, and cherry) in placebo formulations was demonstrated. The instrument was also employed to identify unknown flavors in drug formulation placebos. Raspberry flavor samples from different lots made by the same manufacturer, as well as freshly prepared and aged samples, were also distinguished by electronic-nose. Therefore, the instrument can potentially be used for identity testing of different flavor raw materials and the flavored solution formulations. The electronic-nose was also employed successfully for quantitative analysis of flavors in an oral solution formulation. The quantitative method might be used to assay the flavor concentration during release testing of the oral solution formulation or to monitor flavor shelf-life in the marketed container. It can also be implemented for packaging selection for the formulation in order to ensure the flavor shelf-life. Chemometric methodologies including principal component analysis (PCA), discriminant factorial analysis (DFA), and partial least squares (PLS), were used for data processing and identification. more...

By regulating voltage-dependent Ca2+ influx and intracellular Ca2+ homeostasis, electrical activity plays a central role in motoneuron development. Dissociated cultures of purified embryonic rat motoneurons were used to explore the molecular mechanisms by which Ca2+ influx control [Ca2+]i transients in these neurons. Thapsigargin (250 nm) and cyclopiazonic acid (10 micro m), which deplete Ca2+ stores in the endoplasmic reticulum, decrease by 30% the depolarization-induced [Ca2+]i transients in motoneurons without affecting voltage-activated calcium currents. This thapsigargin-sensitive intracellular Ca2+ pool differs from other previous described Ca2+ stores that are sensitive to ryanodine or caffeine, inositol triphosphate, insulin and from mitochondrial Ca2+ pools. Thapsigargin affected the Cav2.1 P-type Ca2+ channel component of the depolarization-induced [Ca2+]i transient in motoneurons but spared [Ca2+]i transient induced by Cav1 L-type and Cav2.2 N-type Ca2+ channel components, suggesting a close functional relationship between Cav2.1 subunit and this unique thapsigargin-sensitive Ca2+ store. Altogether the present results demonstrate a new pathway, used by embryonic motoneurons, to regulate Ca2+ signalling through voltage-activated (Cav2.1) Ca2+ channels. more...