In silico energetic and molecular dynamic simulations studies demonstrate potential effect of the point mutations with implications for protein engineering in BDNF.

Protein engineering by directed evolution is time-consuming. Hence, in silico techniques like FoldX-Yasara for ∆∆G calculation, and SNPeffect for predicting propensity for aggregation, amyloid formation, and chaperone binding are employed to design proteins. Here, we used in silico techniques to engineer BDNF-NTF3 interaction and validated it using mutations with known functional implications for NGF dimer. The structures of three mutants representing a positive, negative, or neutral ∆∆G involving two interface residues in BDNF and two mutations representing a neutral and positive ∆∆G in NGF, which is aligned with BDNF, were selected for molecular dynamics (MD) simulation. Our MD results conclude that the secondary structure of individual protomers of the positive and negative mutants displayed a similar or different conformation from the NTF3 monomer, respectively. The positive mutants showed fewer hydrophobic interactions and higher hydrogen bonds compared to the wild-type, negative, and neutral mutants with similar SASA, suggesting solvent-mediated disruption of hydrogen-bonded interactions. Similar results were obtained for mutations with known functional implications for NGF and BDNF. The results suggest that mutations with known effects in homologous proteins could help in validation, and in silico directed evolution experiments could be a viable alternative to the experimental technique used for protein engineering.

TrkB transmembrane domain: bridging structural understanding with therapeutic strategy.

TrkB (neuronal receptor tyrosine kinase-2, NTRK2) is the receptor for brain-derived neurotrophic factor (BDNF) and is a critical regulator of activity-dependent neuronal plasticity. The past few years have witnessed an increasing understanding of the structure and function of TrkB, including its transmembrane domain (TMD). TrkB interacts with membrane cholesterol, which bidirectionally regulates TrkB signaling. Additionally, TrkB has recently been recognized as a binding target of antidepressant drugs. A variety of different antidepressants, including typical and rapid-acting antidepressants, as well as psychedelic compounds, act as allosteric potentiators of BDNF signaling through TrkB. This suggests that TrkB is the common target of different antidepressant compounds. Although more research is needed, current knowledge suggests that TrkB is a promising target for further drug development.

Plasma Brain-Derived Neurotrophic Factor Levels in First-Episode and Recurrent Major Depression and before and after Bright Light Therapy in Treatment-Resistant Depression.

Brain-derived neurotrophic factor (BDNF) is implicated in the etiology and treatment response in major depressive disorder (MDD). However, peripheral BDNF concentrations have not been compared across different MDD stages. Bright light therapy (BLT) offers some potential in treatment-resistant depression (TRD), but its effects on BDNF levels are unknown. This study included a cross-sectional analysis of plasma BDNF concentration in females with TRD, unmedicated MDD patients, and healthy controls (HC), and measurements of longitudinal BLT effects on plasma BDNF levels in TRD patients. The present study included 55 drug-naïve, first-episode patients, 25 drug-free recurrent-episode MDD patients, 71 HC participants, and 54 TRD patients. Patients were rated by Hamilton Depression Rating Scale (HAMD)-17 and the Montgomery-Åsberg Depression Rating Scale (MADRS). Patients with TRD received BLT during 4 weeks. The total HAMD-17 and MADRS scores decreased following BLT. All patient groups had lower plasma BDNF than HC, but BDNF levels did not differ between first- and recurrent-episode BDNF patients and TRD patients before or after BLT. However, responders and remitters to BLT had higher post-treatment plasma BDNF concentrations than patients who did not achieve response or remission. The changes in plasma BDNF levels may be candidates for biomarkers of treatment response to BLT in TRD patients.

Effects of moderate-intensity aerobic exercise on serum BDNF and motor learning in the upper-limb in patients after chronic-stroke: A randomized, controlled feasibility study with embedded health economic evaluation.

BACKGROUND: Brain-derived neurotrophic factor (BDNF) promotes activity-dependent neuroplasticity and is released following aerobic-exercise. OBJECTIVE: Feasibility and efficacy of 1.Moderate-Intensity Cycle-Ergometer-Training (MI-ET) and 2.Low-Intensity Circuit-Training (LI-CT) on BDNF-serum-concentration in chronic-stroke and consequently efficacy of motor-learning in varying BDNF-concentrations (neuroplasticity being the substrate for motor-learning) via upper-limb robotic-training (RT) in both groups. METHODS: Randomised-control feasibility-study. 12-week, 3x/week intervention, 17 chronic-stroke-survivors randomized into: (1) MI-ET&RT or (2) LI-CT&RT. Both groups completed 40 mins MI-ET or LI-CT followed by 40 mins RT. Feasibility outcomes: (1) screening and enrollment-rates, (2) retention-rates, (3) adherence: (i) attendance-rates, (ii) training-duration, (4) adverse events. Primary clinical outcomes: 1. serum-BDNF changes pre-post training (immediate) and pre-training basal-levels over 12-weeks (long-term). 2.upper-limb performance with Action-Research-Arm-Test (ARAT). Additionally, feasibility of an embedded health economic evaluation (HEE) to evaluate health-costs and cost-effectiveness. OUTCOMES: cost-questionnaire return-rates, cost-of-illness (COI) and Health-Utitility-Index (HUI). RESULTS: 21.5% of eligible and contactable enrolled. 10 randomized to MI-ET and 7 to LI-CT. 85% of training-sessions were completed in MI-ET (306/360) and 76.3% in LI-CT-group (165/216). 12-weeks: Drop-outs MI-ET-10%, LI-CT-43%. CLINICAL OUTCOMES: No significant changes in immediate or long-term serum-BDNF in either group. Moderate-intensity aerobic-training did not increase serum-BDNF post-stroke. Individual but no group clinically-relevant changes in ARAT-scores. HEE outcomes at 12-weeks: 100% cost-questionnaires returned. Group-costs baseline and after treatment, consistently favouring MI-ET group. COI: (1-year-time-frame): MI-ET 67382 SD (43107) Swiss-Francs and LI-CT 95701(29473) Swiss-Francs. CONCLUSION: The study is feasible with modifications. Future studies should compare high-intensity versus moderate-intensity aerobic-exercise combined with higher dosage arm-training.

Longitudinal Changes in BDNF and MMP-9 Protein Plasma Levels in Children after Cochlear Implantation.

Congenitally deaf children who undergo cochlear implantation before 1 year of age develop their auditory skills faster than children who are implanted later. In this longitudinal study, a cohort of 59 implanted children were divided into two subgroups according to their ages at implantation-below or above 1 year old-and the plasma levels of matrix metalloproteinase-9 (MMP-9), brain-derived neurotrophic factor (BDNF), and pro-BDNF were measured at 0, 8, and 18 months after cochlear implant activation, while auditory development was simultaneously evaluated using the LittlEARs Questionnaire (LEAQ). A control group consisted of 49 age-matched healthy children. We identified statistically higher BDNF levels at 0 months and at the 18-month follow-ups in the younger subgroup compared to the older one and lower LEAQ scores at 0 months in the younger subgroup. Between the subgroups, there were significant differences in the changes in BDNF levels from 0 to 8 months and in LEAQ scores from 0 to 18 months. The MMP-9 levels significantly decreased from 0 to 18 months and from 0 to 8 months in both subgroups and from 8 to 18 months only in the older one. For all measured protein concentrations, significant differences were identified between the older study subgroup and the age-matched control group.

LC-MS/MS determination of GTS-201, a dipeptide mimetic of the brain-derived neurotrophic factor, and neurotransmitter metabolites with application to a pharmacokinetic study in rats.

Brain-derived neurotrophic factor (BDNF) is a member of the neurotrophin family with diverse psychopharmacological effects including antidepressant and anxiolytic actions. However, the clinical use of BDNF is limited due to its poor pharmacokinetic properties. The development of low-molecular-weight BDNF mimetics passing through the blood-brain barrier is an emerging strategy for improved managing psychiatric diseases. The present study characterizes a novel dipeptide mimetic of the 2nd BDNF loop named GTS-201, which exhibits psychotropic properties in experimental animal models of anxiety and alcohol dependence. The aim of this work was to study the pharmacokinetics of GTS-201 in rats at a saturating dosage of 5 mg/kg applied by the intraperitoneal route and to characterize the effects on neurotransmitter levels in the blood and brain. The maximum concentration (Cmax) of GTS-201 in the plasma (867 ± 69 ng/ml) was recorded at 35 ± 7.7 min after administration (Tmax) with a half-elimination period (T1/2) of 19.5 ± 1.8 min, while in the brain tissue Cmax was 14.92 ± 3.11 ng/ml, Tmax was 40.0 ± 7.7 min and T1/2 were 87.5 ± 12.7 min. The relative tissue availability of the GTS-201 for the brain reached 2.9%. At the dose applied, GTS-201 induced a significant increase of serotonin (5-fold) and dopamine levels in the brain tissue (8-fold) along with a decrease in cortisol content in blood plasma 45 min after acute administration. In summary, GTS-201 crosses the blood-brain barrier after acute administration and affects the activity of serotonergic and dopaminergic systems, which may underlie its neuropsychotropic effects described previously.

Acute Normobaric Hypoxia Lowers Executive Functions among Young Men despite Increase of BDNF Concentration.

BACKGROUND: Decreased SpO2 during hypoxia can cause cognitive function impairment, and the effects of acute hypoxia on high-order brain functions such as executive processing remain unclear. This study's goal was to examine the impact of an acute normobaric hypoxia breathing session on executive function and biological markers. METHODS: Thirty-two healthy subjects participated in a blind study performing two sessions of single 30 min breathing bouts under two conditions (normoxia (NOR) and normobaric hypoxia (NH), FIO2 = 0.135). The Stroop test was applied to assess cognitive function. RESULTS: No significant difference was observed in the Stroop interference in the "reading" part of the test in either condition; however, there was a significant increase in the "naming" part under NH conditions (p = 0.003), which corresponded to a significant decrease in SpO2 (p < 0.001). There was a significant increase (p < 0.013) in the brain-derived neurotrophic factor (BDNF) level after NH conditions compared to the baseline, which was not seen in NOR. In addition, a significant drop (p < 0.001) in cortisol levels in the NOR group and a slight elevation in the NH group was noticed. CONCLUSIONS: According to these findings, acute hypoxia delayed cognitive processing for motor execution and reduced the neural activity in motor executive and inhibitory processing. We also noted that this negative effect was associated with decreased SpO2 irrespective of a rise in BDNF.

The role of BDNF exon I region methylation in the treatment of depression with sertraline and its clinical diagnostic value.

BACKGROUND: Brain-derived neurotrophic factor (BDNF) is considered to be one of the best candidate genes for depression. However, whether sertraline treatment affects the methylation level of this gene remains unknown. METHODS: Fifty-three patients with depression and 51 healthy controls were included in the study. The methylation level of BDNF exon I was determined in blood samples from these subjects. The Hamilton Depression Scale was used to evaluate the depression status of patients. Single nucleotide polymorphism detection was used for genotyping, and a receiver operating characteristic (ROC) curve was used to evaluate the predictive value of the methylation level of this locus in patients with depression. RESULTS: There was a significant difference in the methylation level of BDNF exon I between the control and depression groups. No effect of sertraline monotherapy on BDNF methylation was found in subjects with depression. Moreover, no interaction was found between BDNF genotype and the per cent methylation of BDNF exon I. However, methylation at this site was positively correlated with diurnal variation and retardation scores. Blood homocysteine concentrations were significantly reduced by sertraline treatment. No influence of genotype on serum BDNF concentration was found in subjects with depression. The ROC curve showed that methylation of BDNF exon I may be used to distinguish patients from healthy people, to a certain extent. CONCLUSION: Methylation of BDNF exon I may be used as a biomarker of depression and may be a therapeutic target for previously untreated depression.

Neuroprotective effects of NDEELNK from sea cucumber ovum against scopolamine-induced PC12 cell damage through enhancing energy metabolism and upregulation of the PKA/BDNF/NGF signaling pathway.

The aim of the study was to evaluate the neuroprotective function of sea cucumber ovum peptide-derived NDEELNK and explore the underlying molecular mechanisms. NDEELNK exerted the neuroprotective effect by improving the acetylcholine (ACh) level and reducing the acetylcholinesterase (AChE) activity in PC12 cells. By molecular docking, we confirmed that the NDEELNK backbone and AChE interacted through hydrophobic and hydrogen bonds in contact with the amino acid residues of the cavity wall. NDEELNK increased superoxide dismutase (SOD) activity and decreased reactive oxygen species (ROS) production, thereby reducing mitochondrial dysfunction and enhancing energy metabolism. Our results demonstrated that NDEELNK supplementation alleviated scopolamine-induced PC12 cell damage by improving the cholinergic system, increasing energy metabolism and upregulating the expression of phosphorylated protein kinase A (p-PKA), brain-derived neurotrophic factor (BNDF) and nerve growth factor (NGF) signaling proteins in in vitro experiments. These results demonstrated that the sea cucumber ovum peptide-derived NDEELNK might play a protective role in PC12 cells.

Low-Molecular Weight BDNF Mimetic, Dimeric Dipeptide GSB-106, Reverses Depressive Symptoms in Mouse Chronic Social Defeat Stress.

A mimetic of the BDNF loop 4, bis (N-monosuccinyl-L-seryl-L-lysine) hexamethylenediamide, named GSB-106, was designed and synthesized in our scientific group. The compound activated TrkB, MAPK/ERK, PI3K/AKT, and PLCγ in in vitro experiments. In vivo experiments with rodents revealed its antidepressant-like activity in the forced swim and the tail suspension tests at the dose range of 0.1-5.0 mg/kg (i.p., p.o.). However, GSB-106 was not studied in depression models modulating major depression in humans. In the present study, the GSB-106 antidepressant-like activity was revealed in mice at the depression model induced by 28-day social defeat stress with 21-days oral administration (0.1 mg/kg) after stress. At the same time, GSB-106 restored reduced locomotor activity and completely eliminated the anhedonia manifestations. The compound also restored reduced levels of synaptophysin and CREB in the hippocampus. In addition, the Trk receptor antagonist K252A, and the PLC inhibitor U73122, were found to completely block the antidepressant-like activity of GSB-106 in the forced swimming test in mice. Thus, the present results demonstrate the dipeptide BDNF mimetic GSB-106 reversed depressive-like behavior and restored hippocampal neuroplasticity in a rodent depression model. These effects of GSB-106 are probably regulated by TrkB signaling.

ERRγ ligand HPB2 upregulates BDNF-TrkB and enhances dopaminergic neuronal phenotype.

Brain derived neurotrophic factor (BDNF) promotes maturation of dopaminergic (DAergic) neurons in the midbrain and positively regulates their maintenance and outgrowth. Therefore, understanding the mechanisms regulating the BDNF signaling pathway in DAergic neurons may help discover potential therapeutic strategies for neuropsychological disorders associated with dysregulation of DAergic neurotransmission. Because estrogen-related receptor gamma (ERRγ) is highly expressed in both the fetal nervous system and adult brains during DAergic neuronal differentiation, and it is involved in regulating the DAergic neuronal phenotype, we asked in this study whether ERRγ ligand regulates BDNF signaling and subsequent DAergic neuronal phenotype. Based on the X-ray crystal structures of the ligand binding domain of ERRγ, we designed and synthesized the ERRγ agonist, (E)-4-hydroxy-N'-(4-(phenylethynyl)benzylidene)benzohydrazide (HPB2) (Kd value, 8.35 µmol/L). HPB2 increased BDNF mRNA and protein levels, and enhanced the expression of the BDNF receptor tropomyosin receptor kinase B (TrkB) in human neuroblastoma SH-SY5Y, differentiated Lund human mesencephalic (LUHMES) cells, and primary ventral mesencephalic (VM) neurons. HPB2-induced upregulation of BDNF was attenuated by GSK5182, an antagonist of ERRγ, and siRNA-mediated ERRγ silencing. HPB2-induced activation of extracellular-signal-regulated kinase (ERK) and phosphorylation of cAMP-response element binding protein (CREB) was responsible for BDNF upregulation in SH-SY5Y cells. HPB2 enhanced the DAergic neuronal phenotype, namely upregulation of tyrosine hydroxylase (TH) and DA transporter (DAT) with neurite outgrowth, both in SH-SY5Y and primary VM neurons, which was interfered by the inhibition of BDNF-TrkB signaling, ERRγ knockdown, or blockade of ERK activation. HPB2 also upregulated BDNF and TH in the striatum and induced neurite elongation in the substantia nigra of mice brain. In conclusion, ERRγ activation regulated BDNF expression and the subsequent DAergic neuronal phenotype in neuronal cells. Our results might provide new insights into the mechanism underlying the regulation of BDNF expression, leading to novel therapeutic strategies for neuropsychological disorders associated with DAergic dysregulation.

Comparative Genomics of the BDNF Gene, Non-Canonical Modes of Transcriptional Regulation, and Neurological Disease.

Alternative splicing of genes in the central nervous system is ubiquitous and utilizes many different mechanisms. Splicing generates unique transcript or protein isoforms of the primary gene that result in shortened, lengthened, or reorganized products that may have distinct functions from the parent gene. Learning and memory genes respond selectively to a variety of environmental stimuli and have evolved a number of complex mechanisms for transcriptional regulation to act rapidly and flexibly to environmental demands. Their patterns of expression, however, are incompletely understood. Many activity-inducible genes generate transcripts by alternative splicing that have an unknown physiological or behavioral function. One such gene codes for the protein brain-derived neurotrophic factor (BDNF). BDNF is a neurotrophin whose expression is essential for cellular growth, synaptogenesis, and synaptic plasticity. It is an important model gene because of its complex structure and the variety of transcriptional mechanisms it displays for expression in response to external stimuli. Some of these are unexpected, or non-canonical, transcriptional control mechanisms that require further exploration in an activity-dependent context. In this review, a comparative genomics approach is taken to highlight the different forms of BDNF gene transcription including potential autoregulatory mechanisms. Modes of BDNF control have general implications for understanding the origins of several neurological disorders that are associated with reduced BDNF function.

Eucommia Leaf Extract Induces BDNF Production in Rat Hypothalamus and Enhances Lipid Metabolism and Aerobic Glycolysis in Rat Liver.

BACKGROUND: Mutations in the brain-derived neurotrophic factor (BDNF) gene and its receptor, tyrosine receptor kinase B (TrkB), have been reported to cause severe obesity in rodents. Our previous study demonstrated that the oral administration of 5% Eucommia leaf extract (ELE) or ELE aroma treatment (ELE aroma) produced anti-obesity effects. OBJECTIVE: In this study, we investigated the effects of ELE on glycolysis and lipid metabolism in male Sprague-Dawley rats, as well as the effects of ELE on BDNF in rat hypothalamus. METHODS AND RESULTS: A significant reduction and a reduction tendency in the respiratory quotient were observed in association with 5% ELE and ELE aroma treatment, respectively. Furthermore, RT-qPCR results showed significant increases in Cpt2, Acad, Complex II, and Complex V mRNA levels in the liver with both treatments. In addition, in rat hypothalamus, significant elevations in BDNF, Akt, PLCγ proteins and CREB phosphorylation were observed in the 5% ELE group and the ELE aroma group. Furthermore, the Ras protein was significantly increased in the ELE aroma group. On the other hand, significant dephosphorylation of ERK1/2 was observed by the western blotting in the 5% ELE group and the ELE aroma group. CONCLUSION: These findings suggest that the ELE treatment enhances the lipid metabolism and increases the aerobic glycolytic pathway, while ELE-induced BDNF may affect such energy regulation. Therefore, ELE has the possibility to control metabolic syndrome.

A conductive dual-network hydrogel composed of oxidized dextran and hyaluronic-hydrazide as BDNF delivery systems for potential spinal cord injury repair.

Spinal cord injury (SCI) often causes neuronal death and axonal degeneration. In this study, we report a new strategy for preparing injectable and conductive polysaccharides-based hydrogels that could sustainably deliver brain-derived neurotrophic factor (BDNF) for SCI repair. We used poly(lactic-co-glycolic acid) (PLGA) as a carrier to encapsulate BDNF. The resulting microspheres were then modified with tannic acid (TA). The polysaccharides-based hydrogel composed of oxidized dextran (Dex) and hyaluronic acid-hydrazide (HA) was mixed with TA-modified microspheres to form the ultimate BDNF@TA-PLGA/Dex-HA hydrogel. Our results showed that the hydrogel had properties similar to natural spinal cords. Specifically, the hydrogel had soft mechanical properties and high electrical conductivity. The cross-sectional morphology of the hydrogel exhibited a continuous and porous structure. The swelling and degradation behaviors of the Dex-HA hydrogel in vitro indicated the incorporation of TA into hydrogel matrix could improve the stability of the hydrogel matrix as well as extend the release time of BDNF from the matrix. Furthermore, results from immunostaining and real-time PCR demonstrated that BDNF@TA-PLGA/Dex-HA hydrogel could promote the differentiation of neural stem cells (NSCs) into neurons and inhibit astrocyte differentiation in vitro. These results show the great potential of this hydrogel as a biomimetic material in SCI regeneration.

3D printable Sodium alginate-Matrigel (SA-MA) hydrogel facilitated ectomesenchymal stem cells (EMSCs) neuron differentiation.

Ectomesenchymal stem cells (EMSCs) are typical adult stem cells obtained from the cranial neural crest. They have the potential to differentiate into various cell types, such as osseous cells, neurons and glial cells. Three-dimensional (3 D) printing is a novel method to construct biological structures by rapid prototyping. Previously, our group reported on the stemness and multi-lineage differentiation potential of EMSCs on gels. However, the exploration of EMSCs in 3 D printing and then evaluation of the growth and neuronal differentiation of EMSCs on extruded 3 D printable hybrid hydrogels has not been reported. Therefore, the current study explored the novel hybrid Sodium alginate-Matrigel (SA-MA) hydrogel extruded 3 D printing to design an in vitro scaffold to promote the differentiation and growth of EMSCs. In addition, the physical properties of the hydrogel were characterized and its drug-releasing property determined. Notably, the results showed that the construct exhibited a sustain-released effect of growth factor BDNF in accordance with the Higuchi equation. Moreover, the cell survival rate on the 3 D printed scaffold was 88.22 ± 1.13% with higher neuronal differentiation efficiency compared with 2 D culture. Thus, SA-MA's ability to enhanced EMSCs neuronal differentiation offers a new biomaterial for neurons regeneration in the treatment of spinal cord injury.

Novel design of (PEG-ylated)PAMAM-based nanoparticles for sustained delivery of BDNF to neurotoxin-injured differentiated neuroblastoma cells.

Brain-derived neurotrophic factor (BDNF) is essential for the development and function of human neurons, therefore it is a promising target for neurodegenerative disorders treatment. Here, we studied BDNF-based electrostatic complex with dendrimer nanoparticles encapsulated in polyethylene glycol (PEG) in neurotoxin-treated, differentiated neuroblastoma SH-SY5Y cells, a model of neurodegenerative mechanisms. PEG layer was adsorbed at dendrimer-protein core nanoparticles to decrease their cellular uptake and to reduce BDNF-other proteins interactions for a prolonged time. Cytotoxicity and confocal microscopy analysis revealed PEG-ylated BDNF-dendrimer nanoparticles can be used for continuous neurotrophic factor delivery to the neurotoxin-treated cells over 24 h without toxic effect. We offer a reliable electrostatic route for efficient encapsulation and controlled transport of fragile therapeutic proteins without any covalent cross-linker; this could be considered as a safe drug delivery system. Understanding the polyvalent BDNF interactions with dendrimer core nanoparticles offers new possibilities for design of well-ordered protein drug delivery systems.

Zinc induced structural changes in the intrinsically disordered BDNF Met prodomain confer synaptic elimination.

Human brain derived neurotrophic factor (BDNF) encodes a protein product consisting of a C-terminal mature domain (mature BDNF) and an N-terminal prodomain, which is an intrinsically disordered protein. A common single nucleotide polymorphism in humans results in a methionine substitution for valine at position 66 of the prodomain, and is associated with memory deficits, depression and anxiety disorders. The BDNF Met66 prodomain, but not the Val66 prodomain, promotes rapid structural remodeling of hippocampal neurons' growth cones and dendritic spines by interacting directly with the SorCS2 receptor. While it has been reported that the Met66 and Val66 prodomains exhibit only modest differences in structural propensities in the apo state, here we show that Val66 and Met66 prodomains differentially bind zinc (Zn). Zn2+ binds with higher affinity and more broadly impacts residues on the Met66 prodomain compared to the Val66 prodomain as shown by NMR and ITC. Zn2+ binding to the Met66 and Val66 prodomains results in distinct conformational and macroscopic differences observed by NMR, light scattering and cryoEM. To determine if Zn2+ mediated conformational change in the Met66 prodomain is required for biological effect, we mutated His40, a Zn2+ binding site, and observed a loss of Met66 prodomain bioactivity. As the His40 site is distant from the known region of the prodomain involved in receptor binding, we suggest that Met66 prodomain bioactivity involves His40 mediated stabilization of the multimeric structure. Our results point to the necessity of a Zn2+-mediated higher order molecular assembly of the Met66 prodomain to mediate neuronal remodeling.

Self-assembling peptide hydrogels functionalized with LN- and BDNF- mimicking epitopes synergistically enhance peripheral nerve regeneration.

The regenerative capacity of the peripheral nervous system is closely related to the role that Schwann cells (SCs) play in construction of the basement membrane containing multiple extracellular matrix proteins and secretion of neurotrophic factors, including laminin (LN) and brain-derived neurotrophic factor (BDNF). Here, we developed a self-assembling peptide (SAP) nanofiber hydrogel based on self-assembling backbone Ac-(RADA)4-NH2 (RAD) dual-functionalized with laminin-derived motif IKVAV (IKV) and a BDNF-mimetic peptide epitope RGIDKRHWNSQ (RGI) for peripheral nerve regeneration, with the hydrogel providing a three-dimensional (3D) microenvironment for SCs and neurites. Methods: Circular dichroism (CD), atomic force microscopy (AFM), and scanning electron microscopy (SEM) were used to characterize the secondary structures, microscopic structures, and morphologies of self-assembling nanofiber hydrogels. Then the SC adhesion, myelination and neurotrophin secretion were evaluated on the hydrogels. Finally, the SAP hydrogels were injected into hollow chitosan tubes to bridge a 10-mm-long sciatic nerve defect in rats, and in vivo gene expression at 1 week, axonal regeneration, target muscular re-innervation, and functional recovery at 12 weeks were assessed. Results: The bioactive peptide motifs were covalently linked to the C-terminal of the self-assembling peptide and the functionalized peptides could form well-defined nanofibrous hydrogels capable of providing a 3D microenvironment similar to native extracellular matrix. SCs displayed improved cell adhesion on hydrogels with both IKV and RGI, accompanied by increased cell spreading and elongation relative to other groups. RSCs cultured on hydrogels with IKV and RGI showed enhanced gene expression of NGF, BDNF, CNTF, PMP22 and NRP2, and decreased gene expression of NCAM compared with those cultured on other three groups after a 7-day incubation. Additionally, the secretion of NGF, BDNF, and CNTF of RSCs was significantly improved on dual-functionalized peptide hydrogels after 3 days. At 1 week after implantation, the expressions of neurotrophin and myelin-related genes in the nerve grafts in SAP and Autograft groups were higher than that in Hollow group, and the expression of S100 in groups containing both IKV and RGI was significantly higher than that in groups containing either IKV or RGI hydrogels, suggesting enhanced SC proliferation. The morphometric parameters of the regenerated nerves, their electrophysiological performance, the innervated muscle weight and remodeling of muscle fibers, and motor function showed that RAD/IKV/RGI and RAD/IKV-GG-RGI hydrogels could markedly improve axonal regeneration with enhanced re-myelination and motor functional recovery through the synergetic effect of IKV and RGI functional motifs. Conclusions: We found that the dual-functionalized SAP hydrogels promoted RSC adhesion, myelination, and neurotrophin secretion in vitro and successfully bridged a 10-mm gap representing a sciatic nerve defect in rats in vivo. The results demonstrated the synergistic effect of IKVAV and RGI on axonal regrowth and function recovery after peripheral nerve injury.

Behavioral Effects of Dimeric Dipeptide BDNF Mimetic GSB-106 in a Rat Model of Depressive-Like State.

The effects of GSB-106, a low-molecular mimetic of BDNF loop 4, that represents a substituted dimeric dipeptide bis (N-monosuccinyl-L-seryl-L-lysine) hexamethylenediamide, on cognitive and motor impairments in a model of a depressive-like state in rats caused by unavoidable electric foot-shock were studied using active avoidance and open-field tests. GSB-106 (0.5 mg/kg, per os, 10 days) completely restored the number of avoidance reactions that was reduced in rats exposed to foot-shock and the percentage of trained rats in active avoidance training. In the open-field test, the peptide restored reduced horizontal activity and the number of explored holes. Thus, GSB-106 corrected impaired learning and memory, as well as locomotor activity and exploratory behavior in a model of depression in rats.

Neurogenic Niche Conversion Strategy Induces Migration and Functional Neuronal Differentiation of Neural Precursor Cells Following Brain Injury.

Glial scars formed after brain injuries provide permissive cues for endogenous neural precursor/stem cells (eNP/SCs) to undergo astrogenesis rather than neurogenesis. Following brain injury, eNP/SCs from the subventricular zone leave their niche, migrate to the injured cortex, and differentiate into reactive astrocytes that contribute to glial scar formation. In vivo neuronal reprogramming, directly converting non-neuronal cells such as reactive astrocytes or NG2 glia into neurons, has greatly improved brain injury repair strategies. However, reprogramming carries a high risk of future clinical applications such as tumorigenicity, involving virus. In this study, we constructed a neural matrix to alter the adverse niche at the injured cortex, enabling eNP/SCs to differentiate into functional neurons. We found that the neural matrix functioned as a "glial trap" that largely concentrated and limited reactive astrocytes to the core of the lesion area, thus altering the adverse niche. The eNP/SCs migrated toward the injured cortex and differentiated into functional neurons. In addition, regenerated neurites extended across the boundary of the injured cortex. Mice treated with the neural matrix demonstrated significant behavioral recovery. For the first time, we induced eNP/SC-derived functional neurons in the cortex after brain injury without the use of viruses, microRNAs, or small molecules. Our novel strategy of applying this "glial trap" to obtain functional neurons in the injured cortex may provide a safer and more natural therapeutic alternative to reprogramming in future clinical applications.