Neural Regenerative Strategies Incorporating Biomolecular Axon Guidance Signals

McCormick, Aleesha M.; Leipzig, Nic D.

doi:10.1007/s10439-011-0505-0

Cited by 26 publications

(18 citation statements)

References 184 publications

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“…Antagonizing semaphorins might be beneficial; for example, SEMA3A&F are involved in inhibiting axonal regeneration in the injured adult central nervous system [27,28,29,30]. In addition, SEMA3A&F that are secreted from neurons in the avascular hypoxic retina enhance aberrant neovascularization and impair neuroretinal function [14,15].…”

Section: Resultsmentioning

confidence: 99%

Gleevec/imatinib, an ABL2 kinase inhibitor, protects tumor and endothelial cells from semaphorin-induced cytoskeleton collapse and loss of cell motility

Procaccia

Nakayama

Shimizu

et al. 2014

Biochemical and Biophysical Research Communications

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Class 3 semaphorins are axonal guidance mediators and regulators of angiogenesis and tumor progression Semaphorin 3A and 3F (SEMA3A&F) act by depolymerizing F-actin, resulting in cytoskeleton collapse A key signaling step is that SEMA3A&F activates ABL2 tyrosine kinase, which activates p190RhoGAP, which in turn inactivates RhoA, thereby diminishing stress fiber formation and ensuing cell migration We now demonstrate that Gleevec (imatinib, STI571), an ABL2 tyrosine kinase inhibitor, abrogates SEMA3A&F-induced stress fiber loss in glioblastoma cells and endothelial cells and diminishes their ability to inhibit migration. On the other hand, Sutent (sunitinib), a receptor tyrosine kinase inhibitor, did not rescue SEMA3A&F-induced collapsing activity. These results describe a novel property of Gleevec, its ability to antagonize semaphorins.

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Section: Resultsmentioning

confidence: 99%

Gleevec/imatinib, an ABL2 kinase inhibitor, protects tumor and endothelial cells from semaphorin-induced cytoskeleton collapse and loss of cell motility

Procaccia

Nakayama

Shimizu

et al. 2014

Biochemical and Biophysical Research Communications

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“…Figure 1 depicts the main procedures involved in engineering recombinant proteins. With this expression system we have created many biotinylatable proteins, such as interferon-gamma, platelet-derived growth factor, and bone-morphogenetic protein [21][22][23] , but we will focus on two proteins that we designed for axon guidance, NGF (29 kDa) and Sema3A (91 kDa) 10 (for review see reference 24 ). Biotinylation is a common technique for identification, immobilization and isolation of labeled proteins utilizing the well-known biotin-streptavidin interaction [25][26][27] .…”

Section: Introductionmentioning

confidence: 99%

Expression, Isolation, and Purification of Soluble and Insoluble Biotinylated Proteins for Nerve Tissue Regeneration

McCormick

Jarmusik

Endrizzi

et al. 2014

JoVE

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Recombinant protein engineering has utilized Escherichia coli (E. coli) expression systems for nearly 4 decades, and today E. coli is still the most widely used host organism. The flexibility of the system allows for the addition of moieties such as a biotin tag (for streptavidin interactions) and larger functional proteins like green fluorescent protein or cherry red protein. Also, the integration of unnatural amino acids like metal ion chelators, uniquely reactive functional groups, spectroscopic probes, and molecules imparting post-translational modifications has enabled better manipulation of protein properties and functionalities. As a result this technique creates customizable fusion proteins that offer significant utility for various fields of research. More specifically, the biotinylatable protein sequence has been incorporated into many target proteins because of the high affinity interaction between biotin with avidin and streptavidin. This addition has aided in enhancing detection and purification of tagged proteins as well as opening the way for secondary applications such as cell sorting. Thus, biotin-labeled molecules show an increasing and widespread influence in bioindustrial and biomedical fields. For the purpose of our research we have engineered recombinant biotinylated fusion proteins containing nerve growth factor (NGF) and semaphorin3A (Sema3A) functional regions. We have reported previously how these biotinylated fusion proteins, along with other active protein sequences, can be tethered to biomaterials for tissue engineering and regenerative purposes. This protocol outlines the basics of engineering biotinylatable proteins at the milligram scale, utilizing a T7 lac inducible vector and E. coli expression hosts, starting from transformation to scale-up and purification.

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“…Often along their paths in vivo , the axonal growth cones encounter choice points regulated by the synchronized effect of numerous receptor-ligand interactions. The molecular guideposts governing these choice points are multifaceted, including bound cell adhesion molecules and soluble and immobilized chemoattractive and chemorepulsive signals (McCormick and Leipzig, 2012). The same cues mediating neuronal axon development are thought to be important for functional recovery following traumatic injury.…”

Section: Introductionmentioning

confidence: 99%

Sensory axon guidance with semaphorin 6A and nerve growth factor in a biomimetic choice point model

et al. 2014

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The direct effect of guidance cues on developing and regenerating axons in vivo is not fully understood, as the process involves a multiplicity of attractive and repulsive signals, presented both as soluble and membrane-bound ligands. A better understanding of axon guidance is critical to functional recovery following injury to the nervous system through improved outgrowth and mapping of damaged nerves. Due to their implications as inhibitors to CNS regeneration, we investigated the repulsive properties of Semaphorin 6A and Ephrin-B3 on E15 rat dorsal root ganglion explants, as well as possible interactions with soluble gradients of chemoattractive nerve growth factor. We employed a 3D biomimetic in vitro choice point model, which enabled the simple and rapid preparation of patterned gel growth matrices with quantifiable presentation of guidance cues in a specifiable manner that resembles the in vivo presentation of soluble and/or immobilized ligands. Neurites demonstrated an inhibitory response to immobilized Sema6A by lumbosacral DRG explants, while no such repulsion was observed for immobilized Ephrin-B3 by explants at any spinal level. Interestingly, Sema6A inhibition could be partially attenuated in a concentration-dependent manner through the simultaneous presentation of soluble NGF gradients. The in vitro model described herein represents a versatile and valuable investigative tool in the quest for understanding developmental processes and improving regeneration following nervous system injury.

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Neural Regenerative Strategies Incorporating Biomolecular Axon Guidance Signals

Cited by 26 publications

References 184 publications

Gleevec/imatinib, an ABL2 kinase inhibitor, protects tumor and endothelial cells from semaphorin-induced cytoskeleton collapse and loss of cell motility

Gleevec/imatinib, an ABL2 kinase inhibitor, protects tumor and endothelial cells from semaphorin-induced cytoskeleton collapse and loss of cell motility

Expression, Isolation, and Purification of Soluble and Insoluble Biotinylated Proteins for Nerve Tissue Regeneration

Sensory axon guidance with semaphorin 6A and nerve growth factor in a biomimetic choice point model

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