Boosting CNS axon regeneration by harnessing antagonistic effects of GSK3 activity.

Implications of GSK3 activity for axon regeneration are often inconsistent, if not controversial. Sustained GSK3 activity in GSK3 knock-in mice reportedly accelerates peripheral nerve regeneration via increased MAP1B phosphorylation and concomitantly reduces microtubule detyrosination. In contrast, the current study shows that lens injury-stimulated optic nerve regeneration was significantly compromised in these knock-in mice.

Microglia Are Irrelevant for Neuronal Degeneration and Axon Regeneration after Acute Injury.

The role of microglia in degenerative and regenerative processes after damage of the nervous system remains ambiguous, partially due to the paucity of appropriate investigative methods. Here, we show that treatment with the pharmacological colony stimulating factor 1 receptor inhibitor PLX5622 specifically eliminated microglia in murine retinae and optic nerves with high efficiency. Interestingly, time course and extent of retinal ganglion cell (RGC) degeneration after optic nerve crush remained unaffected upon microglia depletion, although remnants of prelabeled apoptotic RGCs were not cleared from the retina in these animals.

Highly efficient transduction of primary adult CNS and PNS neurons.

Delivery and expression of recombinant genes, a key methodology for many applications in biological research, remains a challenge especially for mature neurons. Here, we report easy, highly efficient and well tolerated transduction of adult peripheral and central neuronal populations of diverse species in culture using VSV-G pseudo-typed, recombinant baculovirus (BacMam). Transduction rates of up to 80% were reliably achieved at high multiplicity of infection without apparent neuro-cytopathic effects.

Boosting Central Nervous System Axon Regeneration by Circumventing Limitations of Natural Cytokine Signaling.

Retinal ganglion cells (RGCs) do not normally regenerate injured axons, but die upon axotomy. Although IL-6-like cytokines are reportedly neuroprotective and promote optic nerve regeneration, their overall regenerative effects remain rather moderate. Here, we hypothesized that direct activation of the gp130 receptor by the designer cytokine hyper-IL-6 (hIL-6) might induce stronger RGC regeneration than natural cytokines.

Promotion of Functional Nerve Regeneration by Inhibition of Microtubule Detyrosination.

Unlabelled: Functional recovery of injured peripheral neurons often remains incomplete, but the clinical outcome can be improved by increasing the axonal growth rate. Adult transgenic GSK3α(S/A)/β(S/A) knock-in mice with sustained GSK3 activity show markedly accelerated sciatic nerve regeneration. Here, we unraveled the molecular mechanism underlying this phenomenon, which led to a novel pharmacological approach for the promotion of functional recovery after nerve injury.

Topical treatment with coenzyme Q10-containing formulas improves skin's Q10 level and provides antioxidative effects.

Ubiquinone (coenzyme Q10, Q10) represents an endogenously synthesized lipid-soluble antioxidant which is crucial for cellular energy production but is diminished with age and under the influence of external stress factors in human skin. Here, it is shown that topical Q10 treatment is beneficial with regard to effective Q10 replenishment, augmentation of cellular energy metabolism, and antioxidant effects. Application of Q10-containing formulas significantly increased the levels of this quinone on the skin surface.

Active mechanistic target of rapamycin plays an ancillary rather than essential role in zebrafish CNS axon regeneration.

The developmental decrease of the intrinsic regenerative ability of the mammalian central nervous system (CNS) is associated with reduced activity of mechanistic target of rapamycin (mTOR) in mature neurons such as retinal ganglion cells (RGCs). While mTOR activity is further decreased upon axonal injury, maintenance of its pre-injury level, for instance by genetic deletion of the phosphatase and tensin homolog (PTEN), markedly promotes axon regeneration in mammals. The current study now addressed the question whether active mTOR might generally play a central role in axon regeneration by analyzing its requirement in regeneration-competent zebrafish.

Characterization of optic nerve regeneration using transgenic zebrafish.

In contrast to the adult mammalian central nervous system (CNS), fish are able to functionally regenerate severed axons upon injury. Although the zebrafish is a well-established model vertebrate for genetic and developmental studies, its use for anatomical studies of axon regeneration has been hampered by the paucity of appropriate tools to visualize re-growing axons in the adult CNS. On this account, we used transgenic zebrafish that express enhanced green fluorescent protein (GFP) under the control of a GAP-43 promoter.

Quantitative X-ray Elemental Imaging in Plant Materials at the Subcellular Level with a Transmission Electron Microscope: Applications and Limitations.

Energy-dispersive X-ray microanalysis (EDX) is a technique for determining the distribution of elements in various materials. Here, we report a protocol for high-spatial-resolution X-ray elemental imaging and quantification in plant tissues at subcellular levels with a scanning transmission electron microscope (STEM). Calibration standards were established by producing agar blocks loaded with increasing KCl or NaCl concentrations.

The nano-scale mechanical properties of the extracellular matrix regulate dermal fibroblast function.

Changes in the mechanical properties of dermis occur during skin aging or tissue remodeling and affect the activity of resident fibroblasts. With the aim to establish elastic culture substrates that reproduce the variable softness of dermis, we determined Young's elastic modulus E of human dermis at the cell perception level using atomic force microscopy. The E of dermis ranged from 0.

Functional characterisation of the maturation of the blood-brain barrier in larval zebrafish.

Zebrafish are becoming increasingly popular as an organism in which to model human disease and to study the effects of small molecules on complex physiological and pathological processes. Since larvae are no more than a few millimetres in length, and can live in volumes as small as 100 microliters, they are particularly amenable to high-throughput and high content compound screening in 96 well plate format. There is a growing literature providing evidence that many compounds show similar pharmacological effects in zebrafish as they do in mammals, and in particular humans.

Populus euphratica XTH overexpression enhances salinity tolerance by the development of leaf succulence in transgenic tobacco plants.

Populus euphratica is a salt-tolerant tree species that develops leaf succulence after a prolonged period of salinity stress. In the present study, a putative xyloglucan endotransglucosylase/hydrolase gene (PeXTH) from P. euphratica was isolated and transferred to tobacco plants.

Temperature-induced lipocalin (TIL) is translocated under salt stress and protects chloroplasts from ion toxicity.

Temperature-induced lipocalins (TIL) have been invoked in the defense from heat, cold and oxidative stress. Here we document a function of TIL for basal protection from salinity stress. Heterologous expression of TIL from the salt resistant poplar Populus euphratica did not rescue growth but prevented chlorophyll b destruction in salt-exposed Arabidopsis thaliana.

Do growth-stimulated retinal ganglion cell axons find their central targets after optic nerve injury? New insights by three-dimensional imaging of the visual pathway.

Retinal ganglion cells (RGCs) do not normally regenerate injured axons. However, several strategies to transform RGCs into a potent regenerative state have been developed in recent years. Intravitreal CNTF application combined with conditional PTEN and SOCS3 deletion or zymosan-induced inflammatory stimulation together with cAMP analogue injection and PTEN-deletion in RGCs induce long-distance regeneration into the optic nerve of adult mice.

CXCL12/SDF-1 facilitates optic nerve regeneration.

Mature retinal ganglion cells (RGCs) do not normally regenerate injured axons, but undergo apoptosis soon after axotomy. Besides the insufficient intrinsic capability of mature neurons to regrow axons inhibitory molecules located in myelin of the central nervous system as well as the glial scar forming at the site of injury strongly limit axon regeneration. Nevertheless, RGCs can be transformed into a regenerative state upon inflammatory stimulation (IS), enabling these neurons to grow axons into the injured optic nerve.

Glaucoma and optic nerve repair.

Glaucoma is a leading cause of irreversible blindness worldwide and causes progressive visual impairment attributable to the dysfunction and death of retinal ganglion cells (RGCs). Progression of visual field damage is slow and typically painless. Thus, glaucoma is often diagnosed after a substantial percentage of RGCs has been damaged.

Decreased BDNF levels are a major contributor to the embryonic phenotype of huntingtin knockdown zebrafish.

Huntington's disease (HD) is an autosomal dominant, neurodegenerative condition caused by a CAG trinucleotide repeat expansion that is translated into an abnormally long polyglutamine tract in the protein huntingtin. Genetic and transgenic studies suggest that the mutation causes disease predominantly via gain-of-function mechanisms. However, loss of normal huntingtin function resulting from the polyglutamine expansion might also contribute to the pathogenesis of HD.

Zebrafish neurolin-a and -b, orthologs of ALCAM, are involved in retinal ganglion cell differentiation and retinal axon pathfinding.

Neurolin-a and Neurolin-b (also called alcam and nlcam, respectively) are zebrafish orthologs of human ALCAM, an adhesion protein of the immunoglobulin superfamily with functions in axon growth and guidance. Within the developing zebrafish retina, onset and progression of Neurolin-a expression parallels the pattern of retinal ganglion cell (RGC) differentiation. By using a morpholino-based knockdown approach, we show that Neurolin-a (but not Neurolin-b) is necessary for a crucial step in RGC differentiation.

Loss of PINK1 function affects development and results in neurodegeneration in zebrafish.

Parkinson's disease (PD) is the second most prevalent neurodegenerative disorder in the Western world. PTEN (phosphatase/tensin homolog on chromosome 10)-induced putative kinase 1 (PINK1), a putative kinase that is mutated in autosomal recessive forms of PD, is also implicated in sporadic cases of the disease. Although the mutations appear to result in a loss of function, the roles of this protein and the pathways involved in PINK1 PD are poorly understood.

Analysis of the reticulon gene family demonstrates the absence of the neurite growth inhibitor Nogo-A in fish.

Reticulons (RTNs) are a family of evolutionary conserved proteins with four RTN paralogs (RTN1, RTN2, RTN3, and RTN4) present in land vertebrates. While the exact functions of RTN1 to RTN3 are unknown, mammalian RTN4-A/Nogo-A was shown to inhibit the regeneration of severed axons in the mammalian central nervous system (CNS). This inhibitory function is exerted via two distinct regions, one within the Nogo-A-specific N-terminus and the other in the conserved reticulon homology domain (RHD).

The neuronal growth and regeneration associated Cntn1 (F3/F11/Contactin) gene is duplicated in fish: expression during development and retinal axon regeneration.

The Cntn1 (Contactin/F3/F11) cell adhesion molecule is involved in axon growth and guidance, fasciculation, synapse formation, and myelination in birds and mammals. We identified Cntn1 genes in goldfish, zebrafish, and fugu, and provide evidence for a fish-specific duplication leading to Cntn1a and Cntn1b. Our analyses suggest a subfunctionalization for the Cntn1 paralogs in zebrafish compared to other vertebrates which have a single Cntn1 gene.

Identification of two NOGO/RTN4 genes and analysis of Nogo-A expression in Xenopus laevis.

Myelin-associated axon growth inhibitors such as Nogo-A/RTN4-A impair axon regeneration in the adult mammalian central nervous system (CNS). Here, we describe the cloning and expression of two independent Xenopus laevis rtn4 orthologs. As in mammals, alternative transcripts are generated both through differential splicing and promoter usage, giving rise to Xenopus nogo-A, -B, -C and to a new isoform, nogo-N/rtn4-N.

Identification of Nogo-66 receptor (NgR) and homologous genes in fish.

The Nogo-66 receptor NgR has been implicated in the mediation of inhibitory effects of central nervous system (CNS) myelin on axon growth in the adult mammalian CNS. NgR binds to several myelin-associated ligands (Nogo-66, myelin associated glycoprotein, and oligodendrocyte-myelin glycoprotein), which, among other inhibitory proteins, impair axonal regeneration in the CNS of adult mammals. In contrast to mammals, severed axons readily regenerate in the fish CNS.