Dramatic neurological and biological effects by botulinum neurotoxin type A on SH-SY5Y neuroblastoma cells, beyond the blockade of neurotransmitter release

ABSTRACT Clostridial neurotoxins potently and specifically inhibit neurotransmitter release in defined cell types by a mechanism that involves cleavage of specific components of the vesicle docking/fusion complex, the SNARE complex. A derivative of the type A neurotoxin fromClostridium botulinum (termed LHN/A) that retains catalytic activity can be prepared by proteolysis. The LHN/A, however, lacks the putative native binding domain (HC) of the neurotoxin and is thus unable to bind to neurons and effect inhibition of neurotransmitter release. Here we report the chemical conjugation of LHN/A to an alternative cell-binding ligand, wheat germ agglutinin (WGA). When applied to a variety of cell lines, including those that are ordinarily resistant to the effects of neurotoxin, WGA-LHN/A conjugate potently inhibits secretory responses in those cells. Inhibition of release is demonstrated to be ligand mediated and dose dependent and to occur via a mechanism involving endopeptidase-dependent cleavage of the natural botulinum neurotoxin type A substrate. These data confirm that the function of the HC domain of C. botulinumneurotoxin type A is limited to binding to cell surface moieties. The data also demonstrate that the endopeptidase and translocation functions of the neurotoxin are effective in a range of cell types, including those of nonneuronal origin. These observations lead to the conclusion that a clostridial endopeptidase conjugate that can be used to investigate SNARE-mediated processes in a variety of cells has been successfully generated.

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Dramatic neurological and biological effects by Botulinum Neurotoxin Type A on SH-SY5Y neuroblastoma cells, beyond the blockade of neurotransmitter release

10.21203/rs.2.22629/v2 ◽

2020 ◽

Author(s):

Lei Wang ◽

Carol S. Ringelberg ◽

Bal R. Singh

Keyword(s):

Nervous System ◽

Signaling Pathways ◽

Neurotransmitter Release ◽

Site Effects ◽

Botulinum Neurotoxin ◽

System Development ◽

Neuroblastoma Cells ◽

Type A ◽

Nervous System Development ◽

Time Points

Abstract Background Gene expression profile analysis on mammalian cell lines and animal models after exposure to botulinum neurotoxin (BoNT) have been investigated in several studies in recent years. This provides a powerful tool to identify critical signaling pathways involved in the biological and inflammatory responses to BoNT and helps to determine the mechanism of the function of botulinum toxins. One of the key clinical characteristics of BoNT is its prolonged on-site effects. The role of BoNT on the blockage of neurotransmitter acetylcholine release in the neuromuscular junction has been well established. However, the effects of the treatment time of BoNT on the human cellular model and its potential mechanism remain to be defined. This study is aimed to use the gene microarray technology to compare the two physiological critical time points of BoNT type A (BoNT/A) treatment of human neuroblastoma cells and to advance our understanding of the profound biological influences that toxin molecules play in the neuronal cellular system. Results SH-SY5Y cells were treated with BoNT/A for 4 and 48 hours which represent the time needed for the entrance of toxin into the cells and the time necessary for the initial appearance of the on-site effects after BoNT application. A comparison of the two time points identified 122 functional groups that are significantly changed. The top five groups are the alternative splicing, phosphoprotein, nucleus, cytoplasm, and acetylation. Furthermore, after 48 hours there were 744 genes significantly up-regulated and 624 genes significantly down-regulated (p‹ 0.01). These genes fell into the following neurological and biological annotation groups: Nervous system development, proteinaceous extracellular matrix, signaling pathways regulating pluripotency of stem cells, cellular function and signal transduction, and apoptosis. We have also noticed that the up-regulated groups contained neuronal cell development, nervous system development, and metabolic processes while the down-regulated groups contained many chromosomes and cell cycle categories. Conclusions The effects of BoNT/A on neuronal cells extend beyond blocking the neurotransmitter release, and that BoNT/A is a multifunctional molecule that can evoke profound cellular responses which warrant a more in-depth understanding of the mechanism of the toxin’s effects after administration.

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Effects of peripheral botulinum neurotoxin type A injection on behavioral responses and neurotransmitter release of hypernociceptive rats

Toxicon ◽

10.1016/j.toxicon.2018.11.166 ◽

2018 ◽

Vol 156 ◽

pp. S69

Author(s):

Victor R. Muñoz Lora ◽

Giancarlo de la Torre Canales ◽

Juliana Clemente Napimoga ◽

Henrique Balassini Abdalla ◽

Celia Rizzatti ◽

...

Keyword(s):

Neurotransmitter Release ◽

Botulinum Neurotoxin ◽

Behavioral Responses ◽

Type A ◽

Botulinum Neurotoxin Type A

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Dramatic neurological and biological effects by Botulinum Neurotoxin Type A on SH-SY5Y neuroblastoma cells, beyond the blockade of neurotransmitter release

10.21203/rs.2.22629/v1 ◽

2020 ◽

Author(s):

Lei Wang ◽

Carol S. Ringelberg ◽

Bal R. Singh

Keyword(s):

Nervous System ◽

Signaling Pathways ◽

Neurotransmitter Release ◽

Site Effects ◽

Botulinum Neurotoxin ◽

System Development ◽

Neuroblastoma Cells ◽

Type A ◽

Nervous System Development ◽

Time Points

Abstract Background Gene expression profile analysis on mammalian cell lines and animal models after exposure to botulinum neurotoxin (BoNT) have been investigated in several studies in recent years. This provides a powerful tool to identify critical signaling pathways involved in the biological and inflammatory responses to BoNT and helps to determine the mechanism of the function of botulinum toxins. One of the key clinical characteristics of BoNT is its prolonged on-site effects. The role of BoNT on the blockage of neurotransmitter acetylcholine release in the neuromuscular junction has been well established. However, the effects of the treatment time of BoNT on the human cellular model and its potential mechanism remain to be defined. This study is aimed to use the gene microarray technology to compare the two physiological critical time points of BoNT type A (BoNT/A) treatment of human neuroblastoma cells and to advance our understanding of the profound biological influences that toxin molecules play in the neuronal cellular system. Results SH-SY5Y cells were treated with BoNT/A for 4 and 48 hours which represent the time needed for the entrance of toxin into the cells and the time necessary for the initial appearance of the on-site effects after BoNT application. A comparison of the two time points identified 122 functional groups that are significantly changed. The top five groups are the alternative splicing, phosphoprotein, nucleus, cytoplasm, and acetylation. Furthermore, after 48 hours there were 744 genes significantly up-regulated and 624 genes significantly down-regulated (p‹ 0.01). These genes fell into the following neurological and biological annotation groups: Nervous system development, proteinaceous extracellular matrix, signaling pathways regulating pluripotency of stem cells, cellular function and signal transduction, and apoptosis. We have also noticed that the up-regulated groups contained neuronal cell development, nervous system development, and metabolic processes while the down-regulated groups contained many chromosomes and cell cycle categories. Conclusions The effects of BoNT/A on neuronal cells extend beyond blocking the neurotransmitter release, and that BoNT/A is a multifunctional molecule that can evoke profound cellular responses which warrant a more in-depth understanding of the mechanism of the toxin’s effects after administration.

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BOTULINUM NEUROTOXIN TYPE A AS A PROBE FOR STUDYING NEUROTRANSMITTER RELEASE

Biomedical Aspects of Botulism ◽

10.1016/b978-0-12-447180-1.50009-2 ◽

1981 ◽

pp. 47-64 ◽

Cited By ~ 6

Author(s):

J.O. Dolly ◽

C.K. Tse ◽

J.D. Black ◽

R.S. Williams ◽

D. Wray ◽

...

Keyword(s):

Neurotransmitter Release ◽

Botulinum Neurotoxin ◽

Type A ◽

Botulinum Neurotoxin Type A

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Treatment of Immature Scars with Botulinum Toxin

Textbook on Scar Management ◽

10.1007/978-3-030-44766-3_26 ◽

2020 ◽

pp. 219-226

Author(s):

Alexandra Chambers

Keyword(s):

Clinical Practice ◽

Botulinum Toxin ◽

Botulinum Neurotoxin ◽

Biological Effects ◽

Muscle Tension ◽

Type A ◽

Inhibitory Effects ◽

Botulinum Neurotoxin Type A ◽

Tissue Healing ◽

Therapeutic Choice

AbstractUsing Botulinum neurotoxin type A (BoNTA) has yielded promising results in the treatment of immature scars. The biological effects of the toxin on tissue healing appear to be complex and multidimensional and still require additional research. Nevertheless, it is clear that not only does BoNTA reduce muscle tension at the edges of wounds, but it also provides anti-inflammatory effects, promotes angiogenesis and healing, and exerts mediatory or inhibitory effects on a variety of cells. In clinical practice, this pluripotency of BoNTA has been recognized as a therapeutic choice for both prophylaxis and treatment of excessive scarring.

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