Molecular and cellular neurosciences
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Mol. Cell. Neurosci. · Jun 2012
Disruption of E3 ligase NEDD4 in peripheral neurons interrupts axon outgrowth: Linkage to PTEN.
Exploiting molecules and pathways important in developmental axon behaviour may offer new insights into regenerative behaviour of adult peripheral neurons after injury. In previous work, we have provided evidence that inhibition or knockdown of PTEN (phosphatase and tensin homolog deleted on chromosome ten) dramatically increases adult peripheral axon outgrowth, especially in preconditioned neurons (Christie et al., 2010). PTEN appears to operate as an endogenous brake to regeneration. ⋯ A significant role for this unique co-expression was observed with fluorescently tagged siRNA inhibition of Nedd4, which decreased neurite outgrowth, an impact associated with greater expression of PTEN and that was completely reversed with application of a PTEN inhibitor. Overall, our results suggest an important role for Nedd4 regulation of PTEN in the response of peripheral neurons to injury. By degrading PTEN among other potential actions, Nedd4 supports axonal outgrowth whereas its inhibition facilitates PTEN inhibition of regeneration.
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Mol. Cell. Neurosci. · Apr 2012
AAV8(gfp) preferentially targets large diameter dorsal root ganglion neurones after both intra-dorsal root ganglion and intrathecal injection.
Adeno-associated viral vectors (AAV) are increasingly used to deliver therapeutic genes to the central nervous system (CNS) where they promote transgene expression in post mitotic neurones for long periods with little or no toxicity. In adult rat dorsal root ganglia (DRG), we investigated the cellular tropism of AAV8 containing the green fluorescent protein gene (gfp) after either intra-lumbar DRG or intrathecal injection and showed that transduced DRG neurones (DRGN) expressed GFP irrespective of the delivery route, while non-neuronal cells were GFP(-). After intra-DRG delivery of AAV8(gfp), the mean DRGN transduction rate was 11%, while intrathecal delivery transduced a mean of 1.5% DRGN. ⋯ Microglia and astrocytes were highly ramified with increased GFAP(+) immunoreactivity (i.e. activated) in the neuropil around GFP(+) DRG axon projections within the cord after intra-DRG injection. This study showed that after both intra-DRG and intrathecal delivery, strong preferential AAV8 tropism exists for large DRGN unassociated with cell death, but GFP(+) axons projecting in the spinal cord induced local glial activation. These results open up opportunities for targeted delivery of therapeutics such as neurotrophic factors to the injured spinal cord.
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Mol. Cell. Neurosci. · Apr 2012
Homocysteine has anti-inflammatory properties in a hypercholesterolemic rat model in vivo.
Inflammation is a hallmark in many neurodegenerative diseases like Alzheimer's disease or vascular dementia. Cholesterol and homocysteine are both vascular risk factors which have been associated with dementia, inflammation and blood-brain barrier dysfunction. In previous studies we found that hypercholesterolemia but not hyperhomocysteinemia induced inflammation in rats in vivo. ⋯ To study the potential protective effect of homocysteine, inflammation was induced in organotypic rat brain cortex slices and primary microglial cells by treatment with different inflammatory stimuli (e.g. lipopolysaccharide or tissue plasminogen activator). Tissue plasminogen activator-induced inflammation was counteracted by homocysteine. In conclusion, our data demonstrate that homocysteine significantly ameliorates cholesterol-induced inflammation and blood-brain barrier disruption but not the memory impairment, possibly involving a tissue plasminogen activator-related mechanism.
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Mol. Cell. Neurosci. · Mar 2012
Leak K⁺ channel mRNAs in dorsal root ganglia: relation to inflammation and spontaneous pain behaviour.
Two pore domain potassium (K2P) channels (KCNKx.x) cause K⁺ leak currents and are major contributors to resting membrane potential. Their roles in dorsal root ganglion (DRG) neurons normally, and in pathological pain models, are poorly understood. Therefore, we examined mRNA levels for 10 K2P channels in L4 and L5 rat DRGs normally, and 1 day and 4 days after unilateral cutaneous inflammation, induced by intradermal complete Freund's adjuvant (CFA) injections. ⋯ Ipsilateral SFL duration during inflammation was positively correlated with ipsilateral TASK1 and TASK3 mRNAs, and contralateral TASK1, TRESK and TASK2 mRNAs. Thus changes in K2P mRNA levels occurred during inflammation and for 4 K2P channels were associated with spontaneous pain behaviour (SFL). K2P channels and their altered expression are therefore associated with inflammation-induced pain.
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Mol. Cell. Neurosci. · Mar 2012
Examination of mesenchymal stem cell-mediated RNAi transfer to Huntington's disease affected neuronal cells for reduction of huntingtin.
Huntington's disease (HD) is a fatal, autosomal dominant neurodegenerative disorder caused by an expanded trinucleotide (CAG) repeat in exon 1 of the huntingtin gene (Htt). This expansion creates a toxic polyglutamine tract in the huntingtin protein (HTT). Currently, there is no treatment for either the progression or prevention of the disease. ⋯ MSC expressing shRNA antisense to GFP were found to decrease expression of GFP in SH-SY5Y cells after co-culture when assayed by flow cytometry. Additionally MSC expressing shRNA antisense to HTT were able to decrease levels of mutant HTT expressed in both U87 and SH-SY5Y target cells when assayed by Western blot and densitometry. These results are encouraging for expanding the therapeutic abilities of both RNAi and MSC for future treatments of Huntington's disease.