Mining data on usage of electronic nicotine delivery systems (ENDS) from YouTube videos.

OBJECTIVE: The objective was to analyse and compare puff and exhalation duration for individuals using electronic nicotine delivery systems (ENDS) and conventional cigarettes in YouTube videos. METHODS: Video data from YouTube videos were analysed to quantify puff duration and exhalation duration during use of conventional tobacco-containing cigarettes and ENDS. For ENDS, comparisons were also made between 'advertisers' and 'non-advertisers', genders, brands of ENDS, and models of ENDS within one brand. RESULTS: Puff duration (mean =2.4 s) for conventional smokers in YouTube videos (N=9) agreed well with prior publications. Puff duration was significantly longer for ENDS users (mean =4.3 s) (N = 64) than for conventional cigarette users, and puff duration varied significantly among ENDS brands. For ENDS users, puff duration and exhalation duration were not significantly affected by 'advertiser' status, gender or variation in models within a brand. Men outnumbered women by about 5:1, and most users were between 19 and 35 years of age. CONCLUSIONS: YouTube videos provide a valuable resource for studying ENDS usage. Longer puff duration may help ENDS users compensate for the apparently poor delivery of nicotine from ENDS. As with conventional cigarette smoking, ENDS users showed a large variation in puff duration (range =1.9-8.3 s). ENDS puff duration should be considered when designing laboratory and clinical trials and in developing a standard protocol for evaluating ENDS performance.

Effects of bone morphogenetic protein 2 on Id expression and neuroblastoma cell differentiation.

Bone morphogenetic proteins (BMPs) are secretory signal molecules that have a variety of regulatory functions during embryonic morphogenesis. BMP2 has been shown to induce differentiation in many cell types, mediated through the activation of its target genes: the inhibitors of differentiation (Id1-3) and key transcription factors. In this study, we investigated the effects of BMP2 on mouse neuroblastoma (Neuro2a) cell differentiation and regulation of the expression of Id1-3 and neural-specific transcription factors. Our results showed that BMP2 stimulation upregulated Id1-3 expression at the early stage of application by involvement of the Smad signaling pathway. BMP2 caused phosphorylation of Smad1/5/8 followed by upregulation of Id1-3. Co-incubation with Noggin, a BMP antagonist, or Smad1 siRNA transfection significantly inhibited phosphorylation of Smad1/5/8 and upregulation of Id protein. Furthermore, our results showed that BMP2-induced differentiation of Neuro2a cells into neurons by downregulating the expression of Id1-3 proteins and upregulating the expression of neural-specific transcriptional factors Dlx2, Brn3a, and NeuroD6. The results suggested that the transient upregulation of Id1-3 expression during the early phase of BMP stimulation may play a role in lineage specification and promote differentiation of neuroblastoma cells towards a neuronal phenotype. Subsequently, a coordinated increase in expression of proneural transcription factors and a decrease in Id1-3 expression may culminate in the transition from proliferation to neurogenesis and the terminal neuronal differentiation of neuroblastoma cells.

Reverse-phase HPLC purification for an extremely unstable glucuronide metabolite.

A pure ß-​D-​Glucopyranosiduronic acid metabolite (≥98.0 % purity and a single impurity ≤0.50 %) was requested for biological studies. Due to its unusual instability, the purification of the glucuronide metabolite was extremely challenging. Initially, the crude sample (89 % HPLC area purity) was purified on a Waters SunFire C8 OBD column with 40 mM ammonium acetate buffer and acetonitrile as the mobile phase under a gradient program. The purified glucuronide metabolite solid was obtained by evaporation and lyophilization. However, this procedure yielded the target compound with 97.6 % HPLC area purity and did not meet the requirements. Through the investigation, lyophilization was identified as the key step for the purity of the metabolite, and further lyophilization resulted in an increased amount of the degraded impurities. To better understand the compound, stability studies of the purified metabolite were conducted under sample media, organic solvent, acid, base, and light exposure. The compound was observed to be extremely unstable in water, acid, base and methanol, and sensitive to light, but relatively stable in ammonium acetate buffer (pH 5.0). Taking into account compound stability and the initial purification method, the improved purification procedure was successfully developed and the purified glucuronide metabolite was obtained with 99.2 % HPLC area purity and 0.39 % of the largest single impurity.

Sub/supercritical Fluid Chromatography Purification and Desalting of a Cyclic Dinucleotide STING Agonist.

An efficient and "endotoxin-free" purification of a cyclic dinucleotide (CDN) STING agonist was achieved to produce multigram quantities of pure BMT-390025, an active pharmaceutical ingredient (API), for toxicological studies. A two-step sub/supercritical fluid chromatography (SFC) procedure was developed for the achiral purification and desalting of the polar ionic CDN. A robust SFC process employing methanol-acetonitrile-water with ammonium acetate as co-solvent in CO2 on BEH 2-ethylpyridine was established and scaled up as the first step to achieve a successful purification. The desalting/salt-switching (i.e. removing acetate and acetamide) was conducted using methanol-water with ammonium hydroxide as co-solvent on the same column in the second step to convert the final API to the ammonium salt. Water with additive was essential to eliminating salt precipitation and improving the peak shape and resolution. Due to the extreme hydrophilicity of BMT-390025, 65% of co-solvent was needed to adequately elute the target in both steps. More than 40 g of crude API was purified and desalted producing >20 g of pure BMT-390025 as the ammonium salt which was obtained with a chemical purity of >98.5% and met the endotoxin requirement of <0.1 EU/mg. In addition, >80 g of its penultimate prior to the deprotection of the silyl group was purified at a high throughput of 6.3 g/h (0.42 g/day/g SP).

Large-scale supercritical fluid chromatography purification of unstable STING agonist intermediates.

A regioisomeric mixture of the nucleoside derivative, Intermediate 1, required resolution by preparative supercritical fluid chromatography (SFC) in order to obtain the desired regioisomer as a key intermediate in a STING agonist program. Various chiral columns and solvents including methanol, acetonitrile, isopropanol, and the mixture of acetonitrile and isopropanol as organic modifiers in carbon dioxide at different temperatures were screened to obtain the best regioisomeric resolution. A key issue associated with interconversion between the regioisomers via silyl migration during purification was investigated in methanol, acetonitrile, and the mixture of acetonitrile and isopropanol, and the optimal organic modifier in CO2 was established to mitigate the interconversion to an acceptable level (<5%). Taking into account peak resolution, throughput, interconversion and operation robustness, an efficient SFC method for large-scale purification was successfully developed and scaled up onto a 5 cm I. D. Chiralcel OJ-H column using 25% acetonitrile: isopropanol [1:1 (v/v)] with 0.1% ammonium hydroxide as the modifier in CO2 at a total flow rate of 270 mL/min and a temperature of 30°C. In addition, continual evaporation (i.e. every hour) of the desired isomer fraction stream post-separation ensured minimal further interconversion. A total of 258 grams were separated at a high throughput of 8.6 g/h. Regioisomeric purity of the desired isomer of Intermediate 1 was ≥98.2% and the recovery was ≥90.2%. A similar purification strategy was applied to the regioisomeric resolution of Intermediate 2, an analog of Intermediate 1. In total, 1028 grams of Intermediate 2 were processed at a high throughput of 12.5 g/h on a Viridis BEH 2-EP column. The regioisomeric purity of the desired isomer was ≥96.8% and the recovery was ≥90.7%.

The effect of water on the large-scale supercritical fluid chromatography purification of two factor XIa active pharmaceutical ingredients.

BMS-962212, a parenteral Factor XIa inhibitor, was scaled-up for toxicity studies. Two steps of supercritical fluid chromatography (SFC) were developed for the chiral resolution of the penultimate and achiral purification of final active pharmaceutical ingredient (API), BMS-962212. A robust SFC process using Chiralcel OD-H with methanol-acetonitrile as modifier in CO2 was established to achieve a stable and uninterrupted operation with reduced mobile phase viscosity and system pressure drop. More than 230 g of the racemic penultimate was chirally resolved to reach >99% chiral purity, ready for final tert-butyl ester deprotection to provide the API. There were a significant number of impurities in BMS-962212 generated from the final step that needed to be removed. In contrast to conventional SFC conditions, an SFC method exploiting water and ammonia as additives in both the mobile phase and sample solution was developed to accomplish purification and desalting (i.e. removing TFA) of the zwitterionic API in one step. Water as an additive eliminated salt precipitation and improved the resolution while ammonia contributed to the desalting, details of which will be discussed in this article. A throughput of 2 g/h was achieved, and >80 g of the crude API was purified. The same strategy was applied to another Factor XIa API (compound A) and its penultimate.

Discovery and Preclinical Pharmacology of an Oral Bromodomain and Extra-Terminal (BET) Inhibitor Using Scaffold-Hopping and Structure-Guided Drug Design.

Inhibition of the bromodomain and extra-terminal (BET) family of adaptor proteins is an attractive strategy for targeting transcriptional regulation of key oncogenes, such as c-MYC. Starting with the screening hit 1, a combination of structure-activity relationship and protein structure-guided drug design led to the discovery of a differently oriented carbazole 9 with favorable binding to the tryptophan, proline, and phenylalanine (WPF) shelf conserved in the BET family. Identification of an additional lipophilic pocket and functional group optimization to optimize pharmacokinetic (PK) properties culminated in the discovery of 18 (BMS-986158) with excellent potency in binding and functional assays. On the basis of its favorable PK profile and robust in vivo activity in a panel of hematologic and solid tumor models, BMS-986158 was selected as a candidate for clinical evaluation.

Humanized mice for immune checkpoint blockade in human solid tumors.

Immunotherapy, specifically research involving immune checkpoint blockers (ICBs), has become a popular trend in anticancer research over the last three years. Due to the difficulties and often poor translation of results from in-vitro models, in-vivo models have become more relevant than ever. With the discovery of NOD, Prkdcscid , and Il2rγ-/- mutations, patient-derived xenograft (PDX) mouse models were developed, providing an ideal environment for ICBs testing. By implanting a PDX with either CD34+ or peripheral blood mononuclear cells, we can create a human immune system capable of mounting a response against tumor burden. These animal models are currently being used to study molecular mechanisms, test drug efficacy, and trial drug combinations. Others have found use for these humanized mouse models as surrogates to represent otherwise uncommon diseases. Limitations remain with regards to what the models are capable of, but in the short amount of time between the development of these models and heightened interest in ICBs, these mice have already shown utility for future developments in the field of immunotherapy.

Molecular cloning and characterization of the zebrafish (Danio rerio) telomerase catalytic subunit (telomerase reverse transcriptase, TERT).

Telomerase is an enzyme composed of a catalytic subunit (TERT) and RNA template (TR), which specifically elongates telomeres and prevents cellular senescence. Although telomerase cannot be detected in most human somatic tissues, including the nervous system, it can be detected in teleost tissues. To facilitate the investigation of telomerase function in the teleost visual system, the coding sequence of zebrafish TERT is revealed and cloned. Immunoblot, immunohistochemistry, reverse transcription polymerase chain reaction (RT-PCR), and telomeric repeats amplification protocol (TRAP) assay are used to assess the expression of telomerase at mRNA, protein, and functional levels in zebrafish retina. Based on the amino acid sequence of mouse TERT, a full-length telomerase reverse transcriptase cDNA of zebrafish has been isolated and cloned. The deduced protein sequence contains 1,091 amino acid residues and a predicted molecular mass of 126 kDa. Multiple alignment shows that the protein sequence contains the conserved motifs and residues found in TERT of other species. RT-PCR and TRAP assay has detected TERT mRNA expression and telomerase activity, respectively, in all tissues examined, including the retina and the brain. The presence of telomerase activity indicates that a fully functional form of telomerase can be found in the retina. Immunohistochemistry reveals that most neurons in zebrafish retina express TERT in the cell nucleus. The presence of telomerase in different tissues may be associated with the indeterminate growth of teleost. However, teleost retinal neurons are post-mitotic and do not further divide under normal situation. The expression of telomerase activity and TERT in retina implies that telomerase has functions other than the elongation of telomere. These findings could provide new insights on telomerase function in the nervous system.

A neuroprotective herbal mixture inhibits caspase-3-independent apoptosis in retinal ganglion cells.

It was previously demonstrated that Menta-FX, a mixture of Panax quinquefolius L. (PQE), Ginkgo biloba (GBE), and Hypericum perforatum extracts (HPE), enhances retinal ganglion cell survival after axotomy. However, the mechanisms of neuroprotection remain unknown. The aim of this study is to elucidate the neuroprotective mechanisms of Menta-FX. Since PQE, GBE and HPE have all been observed to display anti-oxidative property, the involvement of anti-oxidation in Menta-FX's neuroprotective effect was investigated. Menta-FX lowered nitric oxide (NO) content in axotomized retinas without affecting nitric oxide synthase activity, suggesting that Menta-FX possibly exhibited a NO scavenging property. In addition, the effect of Menta-FX on the frequency of axotomy-induced nuclear fragmentation and caspase-3 activation was investigated. Menta-FX treatment significantly reduced nuclear fragmentation in axotomized retinas. Surprisingly, Menta-FX had no effect on caspase-3 activation, but selectively lowered caspase-3-independent nuclear fragmentation in axotomized retinal ganglion cells. In addition, inhibition of PI3K activity by intravitreal injection of wortmannin, a phosphoinositide-3 kinase (PI3K) inhibitor, completely abolished the neuroprotective effect of Menta-FX, indicating that Menta-FX's neuroprotective effect was PI3K-dependent. Data here suggest that Menta-FX displayed a PI3K-dependent, selective inhibition on a caspase-3-independent apoptotic pathway in axotomized RGCs, thus, highlighting the potential use of herbal remedies as neuroprotective agents for other neurodegenerative diseases.

Downregulation of Aquaporin 9 Exacerbates Beta-amyloid-induced Neurotoxicity in Alzheimer's Disease Models In vitro and In vivo.

Alzheimer's disease (AD) is the most common cause of dementia in the elderly, characterized by progressive cognitive dysfunction. Aquaporin 9 (AQP9) is an aquaglyceroporin membrane channel shown biophysically to conduct water, glycerol, and other small solutes. In our study, we reported for the first time an age-associated decrease in AQP9 mRNA and protein expressions in both hippocampus and cerebral cortex of APPswe/PS1dE9 (Tg) AD mice at 3, 6 and 10 months of age. Consistently, we observed a dose-dependent downregulation of AQP9 expression in PC12 cells after treatment with amyloid-beta protein 1-40 (Aß1-40). Pre-treatment with AQP9 small interfering RNA led to a more severe neurotoxicity in PC12 cells in response to Aß1-40. Furthermore, we corroborated that the active participation of AQP9 in AD progression is associated with Aß-induced apoptosis both in vitro and in vivo. Taken together, our results reveal an important role of AQP9 in Aß-induced pathogenesis of AD which deserves further investigation.

Expression of telomerase reverse transcriptase in adult goldfish retina.

Telomerase, a specialized reverse transcriptase that maintains telomere during cell division, is commonly associated with cell proliferation. Increasing evidence suggests that telomerase may bear functions other than telomere elongation. We investigated whether telomerase is expressed in the continuously growing goldfish retina. Telomeric repeat amplification protocol (TRAP) assay reveals telomerase activity in goldfish retina. Reverse transcription-polymerase chain reaction (RT-PCR) and western blot show that telomerase catalytic subunit (TERT) is expressed at both mRNA and protein levels. Localization of TERT by immunohistochemistry indicates prominent expression of TERT in the outer nuclear layer, the inner nuclear layer, and, in a small population of cells, in the ganglion cell layer. Coexpression of TERT with proliferative cell nuclear antigen (PCNA) immunoreactivity is found in rod progenitor cells. These results suggest the role of telomerase in vertebrate central nervous system (CNS) other than telomere maintenance, such as regulation of cell cycle progression and maintenance of retinal cell phenotypes.

Endothelial nitric oxide synthase is expressed in amacrine cells of developing human retinas.

PURPOSE: To examine the expression and cellular distribution pattern of endothelial nitric oxide synthase (eNOS) in the developing human retina and to compare its expression with that in rats. METHODS: Expression of eNOS was examined by immunohistochemistry in retinas of humans ranging from 8.5 to 28 weeks of gestation (WG) and of rats. RESULTS: In the developing human retina, eNOS expression was first detected in the proximal margin of the neuroblastic layer in the incipient fovea-surrounding area at 12 WG. At 17 to 28 WG, eNOS-immunoreactive cells were located in the innermost part of the inner nuclear layer and in the ganglion cell layer, expanding to both temporal and nasal retinas and the processes projecting into the inner plexiform layer. These eNOS-positive cells coexpressed syntaxin and glutamate decarboxylase, and are probably GABAergic amacrine cells. The onset of eNOS expression in developing amacrine cells, however, preceded the invasion of retinal vasculature, long before vascular function involving these cells can be expected, suggesting that eNOS has a role not only in vasoregulation but also in retinal development. From 20 WG on, eNOS was also detected in the photoreceptors adjacent to the fovea. eNOS expression in amacrine cells and photoreceptors was observed in the central-to-peripheral and temporal-to-nasal gradients. However, in the developing rat retina, eNOS was expressed exclusively in the vascular endothelial cells. CONCLUSIONS: The results support that eNOS plays a role, not only in the regulation of vascular function but also in the process of retinal development in humans.

CNTF and BDNF have similar effects on retinal ganglion cell survival but differential effects on nitric oxide synthase expression soon after optic nerve injury.

PURPOSE: To investigate the effect of ciliary neurotrophic factor (CNTF) and brain-derived neurotrophic factor (BDNF) on retinal ganglion cell (RGC) survival and nitric oxide synthase (NOS) expression in the retina during the early phase of optic nerve (ON) injury, and to examine whether intraperitoneal application of the NOS scavenger nitro-l-arginine (l-NA) could protect the injured RGCs. METHODS: RGCs were retrogradely labeled with granular blue 3 days before the ON was intraorbitally transected. RGC survival was examined 1 week after ON transection and intraocular injection of CNTF and/or BDNF, or 1 to 2 weeks after daily intraperitoneal injection of the NOS inhibitor l-NA. NOS expression was examined by NADPH-diaphorase histochemistry and neuronal NOS (nNOS) immunohistochemistry, and nNOS-positive cells were identified by various staining approaches. RESULTS: Both CNTF and BDNF significantly increased RGC survival 1 week after ON injury. In the ganglion cell layer (GCL), CNTF did not increase the number of NADPH-diaphorase positive ((+)) cells but appeared to reduce the intensity of NADPH-diaphorase staining, whereas BDNF increased the number of NADPH-diaphorase(+) cells and also appeared to enhance the intensity of NADPH-diaphorase staining. In the GCL, amacrine cells but not RGCs were nNOS(+). Some macrophages were also nNOS(+). In contrast, no amacrine cells were nNOS(+) in the inner nuclear layer. Daily intraperitoneal injection of l-NA at appropriate concentrations promoted RGC survival for 1 or 2 weeks after ON injury. CONCLUSIONS: Both CNTF and BDNF protected RGCs after ON injury. CNTF and BDNF acted differently on NOS expression in the GCL. Intraperitoneal injections of l-NA at appropriate dosages enhance RGC survival.

Different optic nerve injury sites result in different responses of retinal ganglion cells to brain-derived neurotrophic factor but not neurotrophin-4/5.

In this study, we investigated whether brain-derived neurotrophic factor (BDNF) and neurotrophin-4/5 (NT-4/5) can achieve prolonged protection on retinal ganglion cells (RGCs) and whether site of axon injury modulates RGC response to neurotrophins. Two optic nerve (ON) injury paradigms, proximal and distal transections, were used. Autologous sciatic nerves were grafted onto ON stump in some animals to provide a suitable environment for axons to regrow. Multiple intravitreal injections of saline, BDNF, or NT-4/5 were performed. Immunohistochemistry was used to determine the proportion of RGCs that were expressing trkB. Twenty days after proximal injury, both BDNF and NT-4/5 promoted RGC survival; this protection diminished 30 days after injury. One month after distal injury, BDNF, but not NT-4/5, promoted RGC survival (by 2-fold). No difference in the proportion of trkB expressing RGCs among the viable ones was seen between the two injury models or after BDNF treatment. Interestingly, the mean size of RGC somata was larger after proximal injury than distal injury. This study demonstrates that (1) RGCs respond differently to neurotrophins under different injury conditions, (2) BDNF but not NT-4/5 significantly enhances survival of distally but not proximally injured RGCs over a prolonged period.

c-Jun expression in surviving and regenerating retinal ganglion cells: effects of intravitreal neurotrophic supply.

PURPOSE: To investigate c-jun expression in surviving and axon-regenerating retinal ganglion cells (RGCs) and the effect of intravitreal neurotrophic supply on c-jun expression. METHODS: All animals underwent optic nerve transection (ONT) 0.5 mm behind the eyeball. Some animals underwent a replacement of the optic nerve with an autologous sciatic nerve graft (SNG) to allow axonal regrowth. To provide a neurotrophic supply, a peripheral nerve (PN) segment or brain-derived neurotrophic factor (BDNF)/ciliary neurotrophic factor (CNTF) was applied intravitreally. The time course of c-jun expression was first examined in both surviving and regenerating RGCs. Then, c-jun expression was examined in surviving and regenerating RGCs 3 weeks after intravitreal BDNF/CNTF treatment. Animals with vehicle eye injection were used as the control. Fluorescent dye was used for retrograde labeling of surviving (applied behind the eyeball) and regenerating (applied at the distal end of the SNG) RGCs. All retinas were immunohistochemically stained for c-jun. RESULTS: c-Jun was not detected in normal RGCs, but weak expression was seen in surviving RGCs after ON injury. The proportion of c-jun-positive (+) RGCs among surviving cell population was 52.6% to 86.5% 2 to 6 weeks after ONT. Among regenerating RGCs, more than 80% expressed c-jun in all treatment groups, a proportion that was significantly higher after CNTF treatment (90.7%). In addition, c-jun expression was much stronger in intensity and the c-jun(+) nuclei were much larger in regenerating than in surviving RGCs. CONCLUSIONS: c-Jun expression in RGCs was upregulated after injury. Most regenerating RGCs were c-jun(+), and the intensity of c-jun expression was higher in regenerating than in surviving RGCs. CNTF also upregulated c-jun expression in RGCs.

Expression of trkA, trkB, and trkC in injured and regenerating retinal ganglion cells of adult rats.

PURPOSE: To investigate changes in percentage of tyrosine kinase (trk)A-, trkB-, and trkC-immunopositive ((+)) retinal ganglion cells (RGCs) at various times after optic nerve (ON) axotomy; the proportion of RGCs regenerating axons into peripheral nerve (PN) grafts that are trkA(+), trkB(+), and trkC(+); whether intravitreal PN-ON implants affect trk immunoreactivity; and the levels of trk mRNAs in ON-injured retinas. METHODS: The ON was transected intraorbitally. Proportions of trkA(+), trkB(+), and trkC(+) RGCs and levels of trk mRNAs were studied by using immunocytochemistry and Northern blot methods, respectively, in injured and RGC-regenerating retinas. RESULTS: In normal retinas, only small numbers of trkB(+) and trkC(+), but not trkA(+), RGCs were seen. The optic fiber layer was intensively immunolabeled with trkB. After ON injury, the proportions of trkA(+), trkB(+), and trkC(+) RGCs rapidly increased and reached their peaks by 3 to 5 days. During the next 3 weeks, the proportion of trkA(+) or trkB(+) RGCs gradually decreased, but the proportion of trkC(+) RGCs remained high. Intravitreal implants of PN+ON segments transiently but significantly suppressed injury-induced increases in all these trk(+) RGC proportions for approximately 5 days. In contrast, 3 days after ON injury, quantitative retinal expression of trkA mRNA, and to a lesser extent trkC mRNA, was downregulated, whereas trkB mRNA expression remained unaffected. Higher proportions of trkA(+) and trkB(+) RGCs and higher levels of all trk mRNAs were seen in regenerating RGCs and retinas, respectively. CONCLUSIONS: This study provides a kinetic analysis of expression of trk in RGCs and retinas after ON injury and during regeneration.

Synergistic effect of Nogo-neutralizing antibody IN-1 and ciliary neurotrophic factor on axonal regeneration in adult rodent visual systems.

The presence of Nogo axon regeneration inhibitory molecules in the central nervous system (CNS) and the counteracting effect of IN-1 antibodies have been widely reported. In this study, we examined the effect of IN-1-producing hybridoma cells on axon regeneration in adult rodent retinal ganglion cells (RGCs) after various types of optic nerve (ON) injury, evaluating therein whether ciliary neurotrophic factor (CNTF) potentiated the effect of IN-1. We found that application of IN-1 alone failed to enhance regeneration of intracranially or intraorbitally transected RGC axons in a peripheral nerve (PN) graft. IN-1 hybridoma cells also failed to significantly promote intraorbitally crushed ON axons to reenter the distal part of the ON. However, a combined application of IN-1 and CNTF had a synergistic effect in both intracranial PN and intraorbital ON crush paradigms. This study suggests that the action of IN-1 antibodies in promoting axon regeneration in the CNS could be more effective when coupled with other appropriate factors.

Enhanced survival and regeneration of axotomized retinal ganglion cells by a mixture of herbal extracts.

The aim of this study is to investigate the effects of Panax quinquefolius L. extract (PQE), Ginkgo biloba extract (GBE), and Hypericum perforatum extract (HPE), in combination or alone, on the survival and regeneration of axotomized retinal ganglion cells (RGCs) in an optic nerve transection model in adult hamsters. Unilateral transection of the optic nerve was performed to evaluate the effects of herbal extracts on the survival of axotomized RGCs. Effects of the herbal extracts on axonal regeneration of axotomized RGCs, on the other hand, were studied by attaching a peripheral nerve graft onto the transected ocular stump to induce regeneration. Operated animals received daily oral administration of vehicle or herbal extracts (PQE, GBE, and HPE), alone or in combination, for 7 and 21 days, respectively, in the survival and regeneration experiments. Surviving and regenerating RGCs were retrogradely labeled with Fluoro-Gold. The eyes were then enucleated and the retinas were flat-mounted for the counting of the labeled RGCs. Treatment with PQE, GBE and HPE alone failed to offer neuroprotection to injured RGCs. However, treatment with Menta-FX, a mixture of PQE, GBE, and HPE, significantly augmented RGC survival 7 days postaxotomy. Treatment with Menta-FX also induced a significant (87%) increase in the number of regenerating RGCs 21 days after optic nerve transection. This study demonstrates that herbs can act as a potential neuroprotective agent for damaged RGCs. It also suggests that the therapeutic value of herbal remedies can be maximized by the use of mixtures of appropriate herbs.

Development of the regenerative capacity of postnatal axotomized rat spinal motoneurons.

The present study examined whether a peripheral nerve (PN) graft can rescue developing motoneurons from degeneration and determined when immature motoneurons begin to express a regenerative capacity. Transplantation of a PN graft was unable to rescue motoneurons from degeneration if spinal root avulsion was performed in animals younger than P14. However, this procedure did enhance motoneuron survival when root avulsion was performed at P14 or later. Immature (P1 or P7) motoneurons were unable to regenerate their axons into the transplanted PN graft following root avulsion, whereas in older animals (P14-P28) motoneurons were able to regenerate axons into the PN graft. The percentage of regenerated motoneurons increased from P21 to P28 and was similar to that of adult animals. Therefore, the regenerative capacity of rat spinal motoneurons first begins at about P14, which seems to be critical.