New BDNF and NT-3 Cyclic Mimetics Concur with Copper to Activate Trophic Signaling Pathways as Potential Molecular Entities to Protect Old Brains from Neurodegeneration.

A low level of Neurotrophins (NTs), their Tyrosine Kinase Receptors (Trks), Vascular Endothelial Growth Factors (VEGFs) and their receptors, mainly VEGFR1 and VEGFR2, characterizes AD brains. The use of NTs and VEGFs as drugs presents different issues due to their low permeability of the blood-brain barrier, the poor pharmacokinetic profile, and the relevant side effects. To overcome these issues, different functional and structural NT mimics have been employed. Being aware that the N-terminus domain as the key domain of NTs for the binding selectivity and activation of Trks and the need to avoid or delay proteolysis, we herein report on the mimicking ability of two cyclic peptide encompassing the N-terminus of Brain Derived Growth Factor (BDNF), (c-[HSDPARRGELSV-]), cBDNF(1-12) and of Neurotrophin3 (NT3), (c-[YAEHKSHRGEYSV-]), cNT3(1-13). The two cyclic peptide features were characterized by a combined thermodynamic and spectroscopic approach (potentiometry, NMR, UV-vis and CD) that was extended to their copper(II) ion complexes. SH-SY5Y cell assays show that the Cu2+ present at the sub-micromolar level in the complete culture media affects the treatments with the two peptides. cBDNF(1-12) and cNT3(1-13) act as ionophores, induce neuronal differentiation and promote Trks and CREB phosphorylation in a copper dependent manner. Consistently, both peptide and Cu2+ stimulate BDNF and VEGF expression as well as VEGF release; cBDNF(1-12) and cNT3(1-13) induce the expression of Trks and VEGFRs.

The Role of the Second Extracellular Loop of Norepinephrine Transporter, Neurotrophin-3 and Tropomyosin Receptor Kinase C in T Cells: A Peripheral Biomarker in the Etiology of Schizophrenia.

The neurobiology of schizophrenia is multifactorial, comprising the dysregulation of several biochemical pathways and molecules. This research proposes a peripheral biomarker for schizophrenia that involves the second extracellular loop of norepinephrine transporter (NEText), the tropomyosin receptor kinase C (TrkC), and the neurotrophin-3 (NT-3) in T cells. The study of NEText, NT-3, and TrkC was performed in T cells and plasma extracted from peripheral blood of 54 patients with schizophrenia and 54 healthy controls. Levels of NT-3, TrkC, and NET were significantly lower in plasma and T cells of patients compared to healthy controls. Co-immunoprecipitation (co-IPs) showed protein interactions with Co-IP NEText-NT-3 and Co-IP NEText-TrkC. Computational modelling of protein-peptide docking by CABS-dock provided a medium-high accuracy model for NT-3-NEText (4.6935 Å) and TrkC-NEText (2.1365 Å). In summary, immunocomplexes reached statistical relevance in the T cells of the control group contrary to the results obtained with schizophrenia. The reduced expression of NT-3, TrkC, and NET, and the lack of molecular complexes in T cells of patients with schizophrenia may lead to a peripheral dysregulation of intracellular signaling pathways and an abnormal reuptake of norepinephrine (NE) by NET. This peripheral molecular biomarker underlying schizophrenia reinforces the role of neurotrophins, and noradrenergic and immune systems in the pathophysiology of schizophrenia.

Surface modification of neurotrophin-3 loaded PCL/chitosan nanofiber/net by alginate hydrogel microlayer for enhanced biocompatibility in neural tissue engineering.

This study prepared a novel three-dimensional nanocomposite scaffold by the surface modification of PCL/chitosan nanofiber/net with alginate hydrogel microlayer, hoping to have the privilege of both nanofibers and hydrogels simultaneously. Bead free randomly oriented nanofiber/net (NFN) structure composed of chitosan and polycaprolactone (PCL) was fabricated by electrospinning method. The low surface roughness, good hydrophilicity, and high porosity were obtained from the NFN structure. Then, the PCL/chitosan nanofiber/net was coated with a microlayer of alginate containing neurotrophin-3 (NT-3) and conjunctiva mesenchymal stem cells (CJMSCs) as a new stem cell source. According to the cross-sectional FESEM, the scaffold shows a two-layer structure with interconnected pores in the range of 20 µm diameter. The finding revealed that the surface modification of nanofiber/net by alginate hydrogel microlayer caused lower inflammatory response and higher proliferation of CJMSCs than the unmodified scaffold. The initial burst release of NT-3 was 69% in 3 days which followed by a sustained release up to 21 days. The RT-PCR analysis showed that the expression of Nestin, MAP-2, and ß-tubulin III genes were increased 6, 5.4, and 8.8-fold, respectively. The results revealed that the surface-modified biomimetic scaffold possesses enhanced biocompatibility and could successfully differentiate CJMSCs to the neuron-like cells.

Mechanically strengthened hybrid peptide-polyester hydrogel and potential applications in spinal cord injury repair.

ADA16 peptide hydrogels have been broadly used in tissue engineering due to their good biocompatibility and nanofibrous structure mimicking the native extracellular matrix (ECM). However, the low mechanical strength often fails them as implantable scaffolds. To improve the mechanical stability of the RADA16 peptide hydrogel, a photocrosslinkable diacrylated poly(ϵ-caprolactone)-b-poly(ethylene glycol)-b-poly(ϵ-caprolactone) triblock copolymer (PCECDA) was physically combined with RADA16 peptide pre-modified with cell adhesive Arg-Gly-Asp sequence (RADA16-RGD). Consequently, an interpenetrating network, RADA16-RGD/PCECDA, was formed with highly enhanced mechanical property. The storage modulus (G') of RADA16-RGD/PCECDA (6% w/v, mass ratio mRADA16-RGD/mPCECDA = 1:5) hybrid hydrogel was elevated to ∼2000 Pa, compared to the RADA16-RGD (1% w/v) hydrogel alone (∼700 Pa). Furthermore, this hybrid hydrogel retained the nanofibrous structure from RADA16-RGD peptide, but underwent much slower degradation than RADA16-RGD alone. In vitro, the hybrid hydrogel exhibited excellent cytocompatibility and promoted the differentiation of the seeded neural stem cells. Finally, the RADA16-RGD/PCECDA hydrogel demonstrated capability in reducing cavitation, glial scar formation and inflammation at the lesion sites of hemi-sectioned spinal cord injury model in rats, which holds great potential for application in neural tissue engineering and regenerative medicine.

A combinatorial approach for spinal cord injury repair using multifunctional collagen-based matrices: development, characterization and impact on cell adhesion and axonal growth.

Spinal cord injury is a devastating condition of the central nervous system, in which traditional treatments are largely ineffective due to the complex nature of the injured tissue. Therefore, biomaterial-based systems have been developed as possible alternative strategies to repair the damaged tissue. In the present study, we aimed to design bioactive agent loaded scaffolds composed of two layers with distinct physical properties to improve tissue regeneration. An electrospun layer with aligned nanofibers was made of collagen (Col) Type-I, poly(lactide-co-glycolide) (PLGA) and laminin to promote cell attachment of mesenchymal-like stem cells towards the direction of fibers, while a Col-based second layer was fabricated by plastic compression to act as a releasing system for NT-3 and chondroitinase ABC, so that axonal growth could be stimulated. Results showed that a source of mesenchymal stem cell (MSC)-like cells, adipose tissue-derived stem cells cultured on the fibrous layer of the matrices were able to adhere and proliferate, where the aligned fibers promoted the cell growth in an organized way. Furthermore, the bilayered matrices also promoted dorsal root ganglion neurite outgrowth. The bilayered matrice with Col/PLGA + laminin top layer appears to promote higher neurite growth. Collectively, the designed constructs show promising structural properties and biological performance for being employed as a scaffold for engineering the spinal cord tissue.

Copper complexes of synthetic peptides mimicking neurotrophin-3 enhance neurite outgrowth and CREB phosphorylation.

In this work we report on the synthesis and physiochemical/biological characterization of a peptide encompassing the first thirteen residues of neurotrophin-3 (NT-3). The protein capability to promote neurite outgrowth and axonal branching by a downstream mechanism that involves the increase of the cAMP response element-binding level (CREB) was found for the NT3(1-13) peptide, thus validating its protein mimetic behaviour. Since copper ions are also involved in neurotransmission and their internalization may be an essential step in neuron differentiation and CREB phosphorylation, the peptide and its copper complexes were characterized by potentiometric and spectroscopic techniques, including UV-visible, CD and EPR. To have a detailed picture of the coordination features of the copper complexes with NT3(1-13), we also scrutinized the two peptide fragments encompassing the shorter sequences 1-5 and 5-13, respectively, showing that the amino group is the main anchoring site for Cu(ii) at physiological pH. The peptide activity increased in the presence of copper ions. The effect of copper(ii) addition is more marked for NT3(1-13) than the other two peptide fragments, in agreement with its higher affinity for metal ions. Confocal microscopy measurements carried out on fluorescently labelled NT3(1-13) indicated that copper ions increase peptide internalization.

Silver nanoparticles conjugated with Neurotrophin 3 upregulate myelin gene transcription pathway.

Mathematical modeling is the art of converting problems from the biological area into handy mathematical formulations whose theoretical and numerical analysis provides understandings about the directions and solutions to the particular problem. Recently, the combination therapy treatments have been revealed exceptionally fruitful by using mathematical modeling technique. The human nervous system is composed of axons, covered by the myelin sheath. Axons carry signals and promote myelin development. The abnormalities in myelination formation due to mutations in myelin gene result in memory disorders and impaired cognitive activities. The ERBb gene family is responsible for causing abnormalities in myelin gene. Using this knowledge, the pathway of mutated myelin gene was retrieved and its model was developed. Modeling and simulation analysis was performed to determine the level of expression of several genes. The Neurotrophin 3 ligand-coated with silver nanoparticle was induced in the model to normalize the transcription of myelin gene. It was observed that the myelin gene expression level increases from 0 after two days of NT3 induction and reaches to the maximum level on the 10th day of drug induction along with an increase in ERBb expression. This research work can be used in the future as a part of drug discovery and formulation.

The Promotion of Neural Regeneration in A Rat Facial Nerve Crush Injury Model Using Collagen-Binding NT-3.

Traumatic facial nerve injury, an important cause of facial paralysis, has a number of adverse effects, including facial muscle dysfunction and facial asymmetry. It has been demonstrated in our previous work that native human NT-3 fused with a collagen-binding domain (CBD-NT-3) could bind to collagen, specifically to exert neurotrophic effects, promoting axonal regeneration. To evaluate the effect of CBD-NT-3 in inducing facial nerve regeneration and functional recovery, the differing effects of CBD-NT-3 and native neurotrophin-3 (NAT-NT-3) were observed using the results of facial nerve functional recovery, electrophysiological testing, and axonal and myelin changes in a rat model of facial nerve crush injury. The rats were injected in the epineurium in crushed fibers of the facial nerve with CBD-NT-3, NAT-NT-3, and PBS respectively. After 4 weeks, the CBD-NT-3 group demonstrated significantly more ordered growth of axons and nerve functional recovery than the NAT-NT-3 group. The results suggest that CBD-NT-3 considerably enhances facial nerve regeneration and functional recovery.

Sustained delivery of bioactive neurotrophin-3 to the injured spinal cord.

Spinal cord injury is a debilitating condition that currently lacks effective clinical treatment. Neurotrophin-3 (NT-3) has been demonstrated in experimental animal models to induce axonal regeneration and functional improvements, yet its local delivery remains challenging. For ultimate clinical translation, a drug delivery system is required for localized, sustained, and minimally invasive release. Here, an injectable composite drug delivery system (DDS) composed of biodegradable polymeric nanoparticles dispersed in a hyaluronan/methyl cellulose hydrogel was injected into the intrathecal space to achieve acute local delivery to the spinal cord after a thoracic clip compression injury. NT-3 was encapsulated in the DDS and released in vitro for up to 50 d. With a single injection of the DDS into the intrathecal space of the injured spinal cord, NT-3 diffused ventrally through the cord and was detectable in the spinal cord for at least 28 d therein. Delivery of NT-3 resulted in significant axon growth with no effect on the astroglial response to injury in comparison with vehicle and injury controls. NT-3 treatment promoted functional improvements at 21 d according to the Basso Beattie Bresnahan locomotor scale in comparison with the DDS alone. The sustained delivery of bioactive NT-3 to the injured spinal cord achieved in this study demonstrates the promise of this DDS for central nervous system repair.

Electroacupuncture Promotes the Differentiation of Transplanted Bone Marrow Mesenchymal Stem Cells Preinduced With Neurotrophin-3 and Retinoic Acid Into Oligodendrocyte-Like Cells in Demyelinated Spinal Cord of Rats.

Transplantation of bone marrow mesenchymal stem cells (MSCs) promotes functional recovery in multiple sclerosis (MS) patients and in a murine model of MS. However, there is only a modicum of information on differentiation of grafted MSCs into oligodendrocyte-like cells in MS. The purpose of this study was to transplant neurotrophin-3 (NT-3) and retinoic acid (RA) preinduced MSCs (NR-MSCs) into a demyelinated spinal cord induced by ethidium bromide and to investigate whether EA treatment could promote NT-3 secretion in the demyelinated spinal cord. We also sought to determine whether increased NT-3 could further enhance NR-MSCs overexpressing the tyrosine receptor kinase C (TrkC) to differentiate into more oligodendrocyte-like cells, resulting in increased remyelination and nerve conduction in the spinal cord. Our results showed that NT-3 and RA increased transcription of TrkC mRNA in cultured MSCs. EA increased NT-3 levels and promoted differentiation of oligodendrocyte-like cells from grafted NR-MSCs in the demyelinated spinal cord. There was evidence of myelin formation by grafted NR-MSCs. In addition, NR-MSC transplantation combined with EA treatment (the NR-MSCs + EA group) reduced demyelination and promoted remyelination. Furthermore, the conduction of cortical motor-evoked potentials has improved compared to controls. Together, our data suggest that preinduced MSC transplantation combined with EA treatment not only increased MSC differentiation into oligodendrocyte-like cells forming myelin sheaths, but also promoted remyelination and functional improvement of nerve conduction in the demyelinated spinal cord.

The effects of controlled release of neurotrophin-3 from PCLA scaffolds on the survival and neuronal differentiation of transplanted neural stem cells in a rat spinal cord injury model.

Neural stem cells (NSCs) have emerged as a potential source for cell replacement therapy following spinal cord injury (SCI). However, poor survival and low neuronal differentiation remain major obstacles to the use of NSCs. Biomaterials with neurotrophic factors are promising strategies for promoting the proliferation and differentiation of NSCs. Silk fibroin (SF) matrices were demonstrated to successfully deliver growth factors and preserve their potency. In this study, by incorporating NT-3 into a SF coating, we successfully developed NT-3-immobilized scaffolds (membranes and conduits). Sustained release of bioactive NT-3 from the conduits for up to 8 weeks was achieved. Cell viability was confirmed using live/dead staining after 14 days in culture. The efficacy of the immobilized NT-3 was confirmed by assessing NSC neuronal differentiation in vitro. NSC neuronal differentiation was 55.2 ± 4.1% on the NT-3-immobilized membranes, which was significantly higher than that on the NT-3 free membrane. Furthermore, 8 weeks after the NSCs were seeded into conduits and implanted in rats with a transected SCI, the conduit+NT-3+NSCs group achieved higher NSC survival (75.8 ± 15.1%) and neuronal differentiation (21.5 ± 5.2%) compared with the conduit+NSCs group. The animals that received the conduit+NT-3+NSCs treatment also showed improved functional outcomes, as well as increased axonal regeneration. These results indicate the feasibility of fabricating NT-3-immobilized scaffolds using the adsorption of NT-3/SF coating method, as well as the potential of these scaffolds to induce SCI repair by promoting survival and neuronal differentiation of transplanted NSCs.

Combinatorial assembly of small molecules into bivalent antagonists of TrkC or TrkA receptors.

A library of peptidomimetics was assembled combinatorially into dimers on a triazine-based core. The pharmacophore corresponds to ß-turns of the neurotrophin polypeptides neurotrophin-3 (NT-3), nerve growth factor (NGF), or brain-derived neurotrophic factor (BDNF). These are the natural ligands for TrkC, TrkA, and TrkB receptors, respectively. The linker length and the side-chain orientation of each monomer within the bivalent mimics were systematically altered, and the impact of these changes on the function of each ligand was evaluated. While the monovalent peptidomimetics had no detectable binding or bioactivity, four bivalent peptidomimetics (2c, 2d, 2e, 3f) are selective TrkC ligands with antagonistic activity, and two bivalent peptidomimetics (1a, 1b) are TrkC and TrkA ligands with antagonistic activity. All these bivalent compounds block ligand-dependent receptor activation and cell survival, without affecting neuritogenic differentiation. This work adds to our understanding of how the neurotrophins function through Trk receptors, and demonstrates that peptidomimetics can be designed to selectively disturb specific biological signals, and may be used as pharmacological probes or as therapeutic leads. The concept of altering side-chain, linker length, and sequence orientation of a subunit within a pharmacophore provides an easy modular approach to generate larger libraries with diversified bioactivity.

Combined use of spinal cord-mimicking partition type scaffold architecture and neurotrophin-3 for surgical repair of completely transected spinal cord in rats.

A body of evidence has suggested that tissue-engineered nerve grafts hold promise for the surgical repair of spinal cord injuries. In this study, a novel nerve graft was prepared to be implantated into a 5 mm gap which was caused by a complete transection of the rat spinal cord. The graft was featured by incorporation of neurotrophin-3 into a chitosan-based tube scaffold with a spinal cord-mimicking, partition-type architecture, which was prepared based on the morphometric insights of normal spinal cord anatomy. A set of behavioral, functional, and histological examinations were carried out to evaluate the repair. Results from Basso, Beattie, and Bresnahan tests, motor evoked potential measurements, anterograde tracing, and histological analyses suggested that the combined application of chitosan as the scaffold biomaterial, a spinal cord-mimicking partition-type as the scaffold architecture, and neurotrophin-3 (NT-3) as the bioactive component might probably create synergetic promotion on spinal cord regeneration. This composite nerve graft yielded significantly better results in axonal regeneration and function restoration as compared to its scaffold alone or other types of hollow tube scaffold alone.

Enhanced neurotrophin-3 bioactivity and release from a nanoparticle-loaded composite hydrogel.

Neurotrophin-3 (NT-3) has shown promise in regenerative strategies after spinal cord injury; however, sustained local delivery is difficult to achieve by conventional methods. Controlled release from poly(lactic-co-glycolic acid) (PLGA) nanoparticles has been studied for numerous proteins, yet achieving sustained release of bioactive proteins remains a challenge. To address these issues, we designed a composite drug delivery system comprised of NT-3 encapsulated in PLGA nanoparticles dispersed in an injectable hydrogel of hyaluronan and methyl cellulose (HAMC). A continuum model was used to fit the in vitro release kinetics of an NT-3 analog from a nanoparticle formulation. Interestingly, the model suggested that the linear drug release observed from composite HAMC was due to a diffusion-limiting layer of methyl cellulose on the particle surface. We then studied the effects of processing parameters and excipient selection on NT-3 release, stability, and bioactivity. Trehalose was shown to be the most effective additive for stabilizing NT-3 during sonication and lyophilization and PLGA itself was shown to stabilize NT-3 during these processes. Of four excipients tested, 400g/mol poly(ethylene glycol) was the most effective during nanoparticle fabrication, with 74% of NT-3 detected by ELISA. Conversely, co-encapsulation of magnesium carbonate with NT-3 was the most effective in maintaining NT-3 bioactivity over 28 days according to a cell-based axonal outgrowth assay. Together, the modeling and optimized processing parameters provide insight critical to designing a controlled bioactive release formulation for ultimate testing in vivo.

The inorganic perspectives of neurotrophins and Alzheimer's disease.

The recent metal hypothesis represents an attempt of a new interpretation key of Alzheimer's disease (AD) to overcome the limits of amyloid cascade. Neurons need to maintain metal ions within a narrow range of concentrations to avoid a detrimental alteration of their homeostasis, guaranteed by a network of specific metal ion transporters and chaperones. Indeed, it is well known that transition metal ions take part in neuromodulation/neurotrasmission. In addition, they are prominent factors in the development and exacerbation of neurodegeneration. Neurotrophins are proteins involved in development, maintenance, survival and synaptic plasticity of central and peripheral nervous systems. A neurotrophin hypothesis of AD has been proposed, whereas the link between neurotrophic factor, the amyloid cascade and biometals has not been taken into account. As a matter of fact, there is a significant overlap between brain areas featured by metal ion dys-homeostasis, and those where the neurotrophins exert their biological activity. Metal ions can directly modulate their activities, through conformational changes, and/or indirectly by activating their downstream signaling in a neurotrophin-independent mode. The focus of this review is on the molecular aspects of Zn(2+) and Cu(2+) interactions with neurotrophins, with the aim to shed light on the intricate mechanisms involving metallostasis and proteostasis in AD.

The therapeutic potential of human multipotent mesenchymal stromal cells combined with pharmacologically active microcarriers transplanted in hemi-parkinsonian rats.

Multipotent mesenchymal stromal cells (MSCs) raise great interest for brain cell therapy due to their ease of isolation from bone marrow, their immunomodulatory and tissue repair capacities, their ability to differentiate into neuronal-like cells and to secrete a variety of growth factors and chemokines. In this study, we assessed the effects of a subpopulation of human MSCs, the marrow-isolated adult multilineage inducible (MIAMI) cells, combined with pharmacologically active microcarriers (PAMs) in a rat model of Parkinson's disease (PD). PAMs are biodegradable and non-cytotoxic poly(lactic-co-glycolic acid) microspheres, coated by a biomimetic surface and releasing a therapeutic protein, which acts on the cells conveyed on their surface and on their microenvironment. In this study, PAMs were coated with laminin and designed to release neurotrophin 3 (NT3), which stimulate the neuronal-like differentiation of MIAMI cells and promote neuronal survival. After adhesion of dopaminergic-induced (DI)-MIAMI cells to PAMs in vitro, the complexes were grafted in the partially dopaminergic-deafferented striatum of rats which led to a strong reduction of the amphetamine-induced rotational behavior together with the protection/repair of the nigrostriatal pathway. These effects were correlated with the increased survival of DI-MIAMI cells that secreted a wide range of growth factors and chemokines. Moreover, the observed increased expression of tyrosine hydroxylase by cells transplanted with PAMs may contribute to this functional recovery.

The repair of the injured adult rat hippocampus with NT-3-chitosan carriers.

The injury of the CA1 region of the adult rat hippocampus causes cognitive impairment. In this study, animal models were established by mechanically injuring the CA1 region of the adult rat hippocampus, and into the injured area were implanted chitosan carriers loaded either with or without NT-3. Immunohistochemical and nerve tracer methods were adopted to observe the role of the above-mentioned carriers in repairing the injured brain and to observe the scar formation after the injury, and Morris water maze (MWM) tests were performed to evaluate the recovery degree of the cognitive function. The results showed that NT-3-chitosan carriers stimulated regeneration of a large amount of NF-positive nerve fiber and neuron-like cells into the injured area. The newly regenerated NF-positive nerve fibers in the injured area rebuilt a neural circuit with the contralateral CA1 region via corpus callosum. Comparison of the lesion control rats and the treated rats indicates that the chitosan carriers loaded either with or without NT-3 may significantly improve the cognitive function after the hippocampus injury.

Conducting polymers, dual neurotrophins and pulsed electrical stimulation--dramatic effects on neurite outgrowth.

In this study the synergistic effect of delivering two neurotrophins simultaneously to encourage neuron survival and neurite elongation was explored. Neurotrophin-3 (NT-3) and brain-derived neurotrophic factor (BDNF) were incorporated into polypyrrole (PPy) during electrosynthesis and the amounts incorporated and released were determined using iodine-125 ((125)I) radio-labelled neurotrophins. Neurite outgrowth from cochlear neural explants grown on the conducting polymer was equivalent to that on tissue culture plastic but significantly improved with the incorporation of NT-3 and BDNF. Neurite outgrowth from explants grown on polymers containing both NT-3 and BDNF showed significant improvement over PPy doped only with NT-3, due to the synergistic effect of both neurotrophins. Neurite outgrowth was significantly improved when the polymer containing both neurotrophins was electrically stimulated. It is envisaged that when applied to the cochlear implant, these conducting and novel polymer films will provide a biocompatible substrate for storage and release of neurotrophins to help protect auditory neurons from degradation after sensorineural hearing loss and encourage neurite outgrowth towards the electrodes.

A peptidomimetic of NT-3 acts as a TrkC antagonist.

Neurotrophins are a family of growth factors that regulate the peripheral and central nervous system. We designed and tested a mini-library of small molecules peptidomimetics based on beta-turns of the neurotrophin growth factor polypeptides NT-3, which is the natural ligand for TrkC receptors. Biological studies identified a peptidomimetic 2Cl that exhibited selective antagonism of TrkC. 2Cl reduces TrkC activation and signaling promoted by NT-3, and selectively blocks ligand-dependent cell survival. 2Cl also blocks ligand-independent TrkC activation and signals that take place when the receptor is over-expressed. This work adds to our understanding of how the neurotrophins function through Trk receptors, and demonstrates that peptidomimetics can be designed to selectively disturb neurotrophin-receptor interactions, and receptor activation.

[Oligomeric organization of recombinant human neurotrophins expressed in Escherichia coli cells].

Genes of human neurotrophins NGF, BDNF, NT-3 were cloned, and the corresponding proteins and their fragments were expressed in Escherichia coli BL-21 (DE3lambda) cells. Their intracellular localization was determined. The conditions for isolation and purification of the target recombinant proteins and for folding of BDNF and NT-3 precursors were selected. The recombinant proprecursors of human neurotrophines have been shown to possess complex oligomeric structure.