scholarly journals Immunolocalization of a neuronal growth-dependent membrane glycoprotein.

1985 ◽  
Vol 101 (5) ◽  
pp. 1990-1998 ◽  
Author(s):  
I Wallis ◽  
L Ellis ◽  
K Suh ◽  
K H Pfenninger
Keyword(s):  
Spinal Cord ◽  
Nervous System ◽  
Growth Cone ◽  
Cortical Neurons ◽  
Axonal Growth ◽  
Staining Intensity ◽  
Olfactory Nerve ◽  
Membrane Glycoprotein ◽  
Neural Cells ◽  
Developing Spinal Cord

Monoclonal antibody (mAb) 5B4 recognizes in the rat a large, developmentally regulated membrane glycoprotein. The larger form of this antigen (185-255 kD) occurs in the developing nervous system and is present in membranes of nerve growth cones, as determined by analysis of a growth cone particle fraction. An immunochemical characterization of this antigen and of a smaller form (140 kD), sparsely present in the mature nervous system, has been described (Ellis, L., I. Wallis, E. Abreu, and K. H. Pfenninger, 1985, J. Cell. Biol., 101:1977-1989). The present paper reports on the localization by immunofluorescence of 5B4 antigen in cultured cortical neurons, developing spinal cord, and the mature olfactory system. In culture, mAb 5B4 stains only neurons; it is sparsely present in neurons at the onset of sprouting while, during sprouting, it appears to be concentrated at the growth cone and in regions of the perikaryon. In the developing spinal cord, 5B4 labeling is faintly detectable on embryonic day 11 but is intense on fetal day 13. At this stage, the fluorescence is observed in regions of the cord where axonal growth is occurring, while areas composed of dividing or migrating neural cells are nonfluorescent. With maturation of the spinal cord, this basic pattern of fluorescence persists initially, but the staining intensity decreases dramatically. In the adult, faint fluorescence is detectable only in gray matter, presumably indicating the presence of the 140 kD rather than the fetal antigen. The only known structure of the adult mammalian nervous system where axonal growth normally occurs is the olfactory nerve. mAb 5B4 intensely stains a variable proportion of olfactory axons in the mucosa as well as in the olfactory bulb. Based on both immunochemical and immunofluorescence data, the 5B4 antigen of 185-255 kD is associated specifically with growing neurons, i.e., neurons that are generating neurites.

scholarly journals Exosome-Mediated siRNA pool Inhibits Axonal Growth Inhibitor in Cortical Neurons of Spinal Cord Injury in Rats

2020 ◽  
Author(s):  
Qi Liao ◽  
Jiang-Hua Ming ◽  
Ge-Liang Hu
Keyword(s):  
Spinal Cord ◽  
Spinal Cord Injury ◽  
Cell Proliferation ◽  
Cortical Neurons ◽  
Axon Growth ◽  
Axonal Growth ◽  
Growth Inhibitor ◽  
Protective Role ◽  
Primary Cortical Neurons ◽  
Cord Injury

Abstract Background: As exosomes have been confirmed as a reservoir of siRNAs involved in certain diseases, the current study aims to investigate whether exosomal-siRNA could exert a protective role in spinal cord injury (SCI). Methods and Results: Exosomes in our experiment were isolated from lysosomal membrane-associated protein 2b (Lamp2b) overexpression HEK 293T cells, and purity of exosomes was characterized by the expression of CD9, CD47, and CD63 via western blot. Furthermore, the siRNA pool contains four siRNAs including siRNA-NgR, siRNA-LINGO-1, siRNA-Troy, and siRNA-PTEN was loaded to the exosomes, which indicated a significant role for the siRNA pool in reducing the expression of axon growth inhibitory factors. Upon the completion of loading into exosomes (exo-siRNA pool), the exo-siRNA pool was injected into primary cortical neurons of the SCI model in rats before cell proliferation and Rho expression were determined With the results revealed that purified addition could be applied to future experiments. The exo-siRNA pooled transfection caused downregulation of axon growth suppressors in primary cortical neurons including Nogo receptors (NgR), leucine-rich repeats and immunoglobulin domain-containing protein 1 (LINGO-1), Troy, and phosphatase and tenson homolog (PTEN). Cell proliferation and Rho expression of primary cortical neurons inhibited the expression of axonal growth inhibitors in rats with SCI by transfecting exogenous Sirna. Conclusion: This study confirmed that exosomes derived from Lamp2b overexpression HEK 293T cells facilitated both the recovery of functions and the survival of neurons when being loaded with the siRNA pool.

91 Expression Pattern of Neuron-Specific Red Fluorescence Protein in Nervous System of Transgenic Dog with Human Synapsin I Promoter

2018 ◽  
Vol 30 (1) ◽  
pp. 185
Author(s):  
H. J. Oh ◽  
M. J. Kim ◽  
G. A. Kim ◽  
E. M. N. Setyawan ◽  
S. H. Lee ◽  
...  
Keyword(s):  
Spinal Cord ◽  
Nervous System ◽  
Neurodegenerative Disorders ◽  
Peripheral Nerves ◽  
Synapsin I ◽  
Neural Cells ◽  
Tissue Samples ◽  
Red Fluorescence ◽  
Fluorescence Protein ◽  
Red Fluorescence Protein

Neurodegenerative disorders, such as Alzheimer’s disease, affect neurons in large areas within the central nervous system. The selective expression of disease-causing and therapeutic genes in susceptible regions and cell types is critical to the generation of animal models. In our previous studies, we reported production of a transgenic dog by nuclear transfer using donor cells that had been stably transfected by vector containing the red fluorescence protein (RFP) and human synapsin I (SynI) promoter, a neuron-specific promoter. Here, we evaluated whether RFP expression in transgenic dog with SynI (SynI dog) had high neuronal specificity and strong transcriptional activity. For confirmation of neuron-specific RFP expression, tissue samples were obtained from a killed 4-year-old SynI dog. Quantification of RFP in heart, kidney, liver, lung, spleen, cerebrum, cerebrum, cerebellum, midbrain, hippocampus, peripheral nerves, skin, and spinal cord of SynI dog was analysed using ELISA (Cell Biolabs, San Diego, CA, USA). Moreover, the distribution of RFP activity in neural system of the SynI dog was determined by immunohistochemistry. The first antibody, rabbit polyclonal RFP antibody (1:200, ab62341, Abcam, Cambridge, MA, USA) was applied, and antibody labelling was visualised by incubation with avidin-biotinylated horseradish peroxidase complex (1:1,000; ABC Elite, Vector Laboratories, Burlingame, CA, USA). In the 4-year-old SynI dog, RFP was observed only in neuronal tissues including brain, spinal cord, and peripheral nerves, but was not detected in non-neuronal tissue such as heart, kidney, and skin. In addition, robust RFP expression was observed in the neurons of the peripheral nerve, spinal cord, and prefrontal cortex. In the hippocampus and cerebellum, the RFP-expressing cells appeared heterogeneous in hippocampus neurons and purkinje cells. In conclusion, we report that human SynI promoter is functional in neural cells of dogs. A neural specific-transgene expressed dog could be applied as a research tool in the study of neurodegenerative disorders. This research was supported by RDA (#PJ010928032017), Korea IPET (#316002-05-2-SB010), NRF (#2016R1D1A1B03932198), and Research Institute for Veterinary Science, the BK21 plus program.

Analysis of subcellular structural tension in axonal growth of neurons

2018 ◽  
Vol 29 (2) ◽  
pp. 125-137 ◽  
Author(s):  
Yi Chen Guo ◽  
Yu Xuan Wang ◽  
Yan Ping Ge ◽  
Lu Jia Yu ◽  
Jun Guo
Keyword(s):  
Nervous System ◽  
Functional Recovery ◽  
Growth Cone ◽  
System Development ◽  
Axonal Growth ◽  
Nervous System Development ◽  
Axonal Elongation ◽  
The Core ◽  
Pathological Conditions ◽  
New Material

AbstractThe growth and regeneration of axons are the core processes of nervous system development and functional recovery. They are also related to certain physiological and pathological conditions. For decades, it has been the consensus that a new axon is formed by adding new material at the growth cone. However, using the existing technology, we have studied the structural tension of the nerve cell, which led us to hypothesize that some subcellular structural tensions contribute synergistically to axonal growth and regeneration. In this review, we classified the subcellular structural tension, osmotic pressure, microfilament and microtubule-dependent tension involved controllably in promoting axonal growth. A squeezing model was built to analyze the mechanical mechanism underlying axonal elongation, which may provide a new view of axonal growth and inspire further research.

scholarly journals Leptin Increases Axonal Growth Cone Size in Developing Mouse Cortical Neurons by Convergent Signals Inactivating Glycogen Synthase Kinase-3β

2006 ◽  
Vol 281 (18) ◽  
pp. 12950-12958 ◽  
Author(s):  
Alessandra Valerio ◽  
Valentina Ghisi ◽  
Marta Dossena ◽  
Cristina Tonello ◽  
Antonio Giordano ◽  
...  
Keyword(s):  
Growth Cone ◽  
Cortical Neurons ◽  
Glycogen Synthase ◽  
Axonal Growth ◽  
Glycogen Synthase Kinase ◽  
Glycogen Synthase Kinase 3Β ◽  
Axonal Growth Cone

scholarly journals Programmed cell senescence in the mouse developing spinal cord and notochord

2020 ◽  
Author(s):  
Jorge Antolio Domínguez-Bautista ◽  
Pilar Sarah Acevo-Rodríguez ◽  
Susana Castro-Obregón
Keyword(s):  
Spinal Cord ◽  
Nervous System ◽  
Cellular Senescence ◽  
Cell Senescence ◽  
Surrounding Tissue ◽  
Galactosidase Activity ◽  
Mouse Development ◽  
Different Types ◽  
Beta Galactosidase ◽  
Developing Spinal Cord

AbstractProgrammed cell senescence is a cellular process that seems to contribute to morphogenesis during embryo development, in addition to cell proliferation, migration, differentiation and programmed cell death, and has been observed in evolutionary distant organisms like mammals, amphibians and fish. Programmed cell senescence is a phenotype similar to stress-induced cellular senescence, characterized by the expression of cell cycle inhibitors such as CDKN1A/p21, increased activity of a lysosomal enzyme with beta-galactosidase activity (coined senescence-associated beta-galactosidase) and, most importantly, secretion of growth factors, interleukins, chemokines, metalloproteases, etc., collectively known as a senescent-associated secretory phenotype that instructs surrounding tissue. How wide is the distribution of programmed cell senescence during mouse development and its specific mechanisms to shape the embryo are still poorly understood. Here, we investigated whether markers of programmed cell senescence are found in the developing mouse spinal cord and notochord. We found discrete areas and developmental windows with high senescence-associated beta galactosidase in both spinal cord and notochord; expression of CDKN1A/p21 was documented in epithelial cells of the spinal cord and the notochord. Treatment of mice embryos developed ex-utero in the presence of the senolytic ABT-263 resulted in decrease senescence-associated beta-galactosidase activity and number of motoneurons. Our data suggest that several cell types undergo programmed cell senescence in developing spinal cord and notochord contributing to morphogenesis.Contribution to the Field StatementCellular senescence is a state in which cells no longer divide but have the remarkable ability to secrete signaling molecules that alter the tissue where they reside. In adults, this state is typically induced by stress situations that cause DNA damage so cells with altered genome do not multiply. Senescent cells also form when a tissue is injured; they help to regenerate damaged tissue and contribute to wound healing. Phagocytic cells eliminate them when their function is done, having a transient existence. During vertebrate development some cells acquire a very similar phenotype, coined programmed cell senescence, and interestingly they have been found in regions that organize the pattern of development of some organs. How wide is the distribution of programmed cell senescence during development and how they help to shape the embryo are still poorly understood. We discovered in mice embryos different types of cells with senescent features located in particular regions of the developing nervous system: where motoneurons form and in a region that secrete molecules that instruct the embryo where different types of neurons will be created. We propose that programed cell senescence contributes to the morphogenesis of the nervous system.

scholarly journals Development of lipidoid nanoparticles for siRNA delivery to neural cells

2021 ◽  
Author(s):  
Purva Khare ◽  
Kandarp M. Dave ◽  
Yashika S Kamte ◽  
Muthiah A. Manoharan ◽  
Lauren A. O'Donnell ◽  
...  
Keyword(s):  
Nervous System ◽  
Cortical Neurons ◽  
Sirna Delivery ◽  
Dorsal Root Ganglion Neurons ◽  
Neural Cells ◽  
Significant Toxicity ◽  
Delivery Platform ◽  
The Central Nervous System ◽  
Ganglion Neurons ◽  
Transfection Agent

Lipidoid nanoparticles (LNPs) are the delivery platform in Onpattro, the first FDA-approved siRNA drug. LNPs are also the carriers in the Pfizer-BioNTech and Moderna COVID-19 mRNA vaccines. While these applications have demonstrated that LNPs effectively deliver nucleic acids to hepatic and muscle cells, it is unclear if LNPs could be used for delivery of siRNA to neural cells, which are notoriously challenging delivery targets. Therefore, the purpose of this study was to determine if LNPs could efficiently deliver siRNA to neurons. Because of their potential utility in either applications in the central nervous system and the peripheral nervous system, we used both cortical neurons and sensory neurons. We prepared siRNA-LNPs using C12-200, a benchmark ionizable cationic lipidoid along with helper lipids. We demonstrated using dynamic light scattering that the inclusion of both siRNA and PEG-lipid provided a stabilizing effect to the LNP particle diameters and polydispersity indices by minimizing aggregation. We found that siRNA-LNPs were safely tolerated by primary dorsal root ganglion neurons. Flow cytometry analysis revealed that Cy5 siRNA delivered via LNPs into rat primary cortical neurons showed uptake levels similar to Lipofectamine RNAiMAX, the gold standard commercial transfection agent. However, LNPs demonstrated a superior safety profile whereas the Lipofectamine-mediated uptake was concomitant with significant toxicity. Fluorescence microscopy demonstrated a time-dependent increase in the uptake of LNP-delivered Cy5 siRNA in a human cortical neuron cell line. Overall, our results suggest that LNPs are a viable platform that can be optimized for delivery of therapeutic siRNAs to neural cells.

scholarly journals Fetal Spinal Cord Tissue in Mini-Guidance Channels Promotes Longitudinal Axonal Growth after Grafting into Hemisected Adult Rat Spinal Cords

1999 ◽  
Vol 6 (4) ◽  
pp. 103-121 ◽  
Author(s):  
Norman I. Bamber ◽  
Huaying Li ◽  
Patrick Aebischer ◽  
Xiao Ming Xu
Keyword(s):  
Spinal Cord ◽  
Nervous System ◽  
Axonal Growth ◽  
Electron Microscopic Level ◽  
Myelinated Axons ◽  
Double Labeling ◽  
Freeze Thaw ◽  
Adult Rat ◽  
Graft Interface ◽  
Guidance Channels

Solid fetal spinal cord (FSC) tissue, seeded into semipermeable mini-guidance channels, was tested for the ability to promote axonal growth across the gap created by a midthoracic (T8) hemisection in adult rats. Fetal thoracic spinal cords, at embryonic days 13 to 15, were harvested and gently aspirated into mini-guidance channels (1.25 mm in diameter and 3.0 mm in length). Care was taken to maintain the rostro-caudal orientation of the FSC. In control rats, the FSC-channel congraft struct was exposed to 5 freeze/thaw cycles to produce non-viable grafts before implantation into the hemisected cord. All cases revealed intact tissue cables of various diameters spanning the rostro-caudal extent of the lesion cavity, with integration of host-graft tissues at both interfaces. Immunofluorescence results indicated that numerous neurofilament-positive axons were present within the FSC tissue cable. Double-labeling of a subpopulation of these axons with calcitonin generelated peptide indicated their peripheral nervous system (PNS) origin. Descending serotonergic and noradrenergic axons were found in the proximity of the rostral host-graft interface, but were not observed to grow into the FSC-graft. Anterograde tracing of propriospinal axons with Phaseolus vulgaris-leucoagglutinin demonstrated that axons had regenerated into the FSC-graft and had traveled longitudinally to the distal end of the channel. Few axons were observed to cross the distal host-graft interface to enter the host spinal cord. Cross-sectional analysis at the midpoint of the tissue cable stained with toluidine blue demonstrated a significant increase (P<0.01) in myelinated axons in viable FSC grafts (1455±663, mean±S.E.M.; n=6) versus freeze-thaw control grafts (155±50; n=5). In addition to the myelinated axons, many unmyelinated axons were observed in the tissue cable at the electron microscopic level. Areas resembling the PNS with typical Schwann cells, as well as those resembling the central nervous system with neurons and central neuropil, were also seen. In freeze-thaw control grafts, neither viable neurons nor central neuropil were observed. Retrograde tracing with Fast Blue and Diamidino Yellow demonstrated that neurons within the FSC graft extended axons into the host spinal cord at least for 2 mm from both the rostral and caudal host-graft interfaces. We conclude that viable FSC grafts within semipermeable guidance channels may serve both as a permissive bridge for longitudinally directed axonal growth and a potential relay for conveying information across a lesion site in the adult rat spinal cord.

β-Mannosidosis: Myelin and oligodendrocyte lesions in brain and spinal cord

1984 ◽  
Vol 42 ◽  
pp. 694-695
Author(s):  
Kathryn L. Lovell ◽  
Margaret Z. Jones
Keyword(s):  
Spinal Cord ◽  
Nervous System ◽  
Genetic Disorder ◽  
Neurological Deficits ◽  
Axonal Spheroids ◽  
South Wales ◽  
Pendular Nystagmus ◽  
Myelin Deficits ◽  
Recessive Defect ◽  
Brain And Spinal Cord

Caprine β-mannosidosis, an autosomal recessive defect of glycoprotein catabolism, is associated with a deficiency of tissue and plasma -mannosidase and with tissue accumulation and urinary excretion of oligosaccharides, including the trisaccharide Man(β1-4)GlcNAc(βl-4)GlcNAc and the disaccharide Man(β1-4)GlcNAc. This genetic disorder is evident at birth, with severe neurological deficits including a marked intention tremor, pendular nystagmus, ataxia and inability to stand. Major pathological characteristics described in Nubian goats in Michigan and in Anglo-Nubian goats in New South Wales include widespread cytoplasmic vacuolation in the nervous system and viscera, axonal spheroids, and severe myelin paucity in the brain but not spinal cord or peripheral nerves. Light microscopic examination revealed marked regional variation in the severity of central nervous system myelin deficits, with some brain areas showing nearly complete absence of myelin and other regions characterized by the presence of 25-50% of the control number of myelin sheaths.

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