u-0126 and Huntington-Disease

Huntington's disease (HD) is a neurodegenerative disease that causes deficits in neurite outgrowth, which suggests that enhancement of neurite outgrowth is a potential direction by which to improve HD. Our previous publications showed that fibroblast growth factor 9 (FGF9) provides anti-apoptosis and anti-oxidative functions in striatal cell models of HD through the extracellular signal-regulated kinases (ERK) pathway, and FGF9 also stimulates cytoskeletons to enhance neurite outgrowth via nuclear factor kappa B (NF-kB) signaling. In this study, we further demonstrate the importance of the ERK pathway for the neurite outgrowth induced by FGF9 in HD striatal models.. FGF9 was treated with ERK (U0126) or NF-kB (BAY11-7082) inhibitors in STHdh. Here, we show that suppression of ERK signaling significantly inhibits FGF9-induced neurite outgrowth, cytoskeletal markers, and synaptic proteins in HD striatal cells. In addition, we also show suppression of ERK signaling significantly decreases FGF9-induced NF-kB activation, whereas suppression of NF-kB does not decrease FGF9-induced ERK signaling. These results suggest that FGF9 activates ERK signaling first, stimulates NF-kB upregulation, and then enhances neurite outgrowth in HD striatal cells.. We elucidate the more detailed mechanisms of neurite outgrowth enhanced by FGF9 in these HD striatal cells. This study may provide insights into targeting neurite outgrowth for HD therapy.

Topics: Animals; Butadienes; Cell Line; Cells, Cultured; Corpus Striatum; Enzyme Inhibitors; Fibroblast Growth Factor 9; Humans; Huntingtin Protein; Huntington Disease; MAP Kinase Signaling System; Mice; Mice, Transgenic; Neurites; Neuronal Outgrowth; NF-kappa B; Nitriles; Nuclear Proteins; Oxidative Stress; Recombinant Proteins; Signal Transduction; Sulfones

Mitochondrial dysfunction is a major cytopathology in Huntington's disease (HD), a fatal and inherited neurodegenerative disease. However, the molecular mechanisms by which the disease-causing gene, mutant Huntingtin (mtHtt), affects mitochondrial function remains elusive. This study aims to determine the role that Mitogen-activated protein kinase 1 (MAPK1) plays in the over-activation of Dynamin-related protein 1 (Drp1), the mitochondrial fission protein, which leads to mitochondrial dysfunction and neurodegeneration seen in HD. We show that MAPK1 binds to and phosphorylates Drp1 in vitro. Drp1 phosphorylation at serine 616 is increased in HD knock-in mouse derived striatal cells, which is abolished by treatment with U0126, a potent inhibitor of MEK1/2. A phosphorylation-deficient mutant of Drp1, Drp1S616A, corrects mitochondrial fragmentation associated with HD. Treatment with U0126 also reduces mitochondrial fragmentation, but has no additional effect in correcting aberrant mitochondrial morphology in cells expressing Drp1S616A. Finally, treatment with U0126 reduces mitochondrial depolarization and mitochondrial superoxide production in HD mutant striatal cells when compared to wildtype cells. This study suggests that in HD, MAPK1 activation leads to the aberrant mitochondrial fission and mitochondrial function by phosphorylating Drp1. Therefore, inhibition of Drp1-mediated excessive mitochondrial fission might be a strategy for development of therapy for treating HD.

Topics: Animals; Butadienes; Corpus Striatum; Dynamins; GTP Phosphohydrolases; Huntington Disease; Membrane Potential, Mitochondrial; Mice; Microtubule-Associated Proteins; Mitochondria; Mitochondrial Proteins; Mitogen-Activated Protein Kinase 1; Models, Biological; Mutant Proteins; Nitriles; Phosphorylation; Phosphoserine; Protein Binding; Reactive Oxygen Species

Huntington's disease (HD) is a heritable neurodegenerative disorder, and there is no cure for HD to date. A type of fibroblast growth factor (FGF), FGF9, has been reported to play prosurvival roles in other neurodegenerative diseases, such as Parkinson's disease and Alzheimer's disease. However, the effects of FGF9 on HD is still unknown. With many similarities in the cellular and pathological mechanisms that eventually cause cell death in neurodegenerative diseases, we hypothesize that FGF9 might provide neuroprotective functions in HD.. In this study, STHdhQ7/Q7 (WT) and STHdhQ111/Q111 (HD) striatal knock-in cell lines were used to evaluate the neuroprotective effects of FGF9. Cell proliferation, cell death and neuroprotective markers were determined via the MTT assay, propidium iodide staining and Western blotting, respectively. The signaling pathways regulated by FGF9 were demonstrated using Western blotting. Additionally, HD transgenic mouse models were used to further confirm the neuroprotective effects of FGF9 via ELISA, Western blotting and immunostaining.. Results show that FGF9 not only enhances cell proliferation, but also alleviates cell death as cells under starvation stress. In addition, FGF9 significantly upregulates glial cell line-derived neurotrophic factor (GDNF) and an anti-apoptotic marker, Bcl-xL, and decreases the expression level of an apoptotic marker, cleaved caspase 3. Furthermore, FGF9 functions through ERK, AKT and JNK pathways. Especially, ERK pathway plays a critical role to influence the effects of FGF9 toward cell survival and GDNF production.. These results not only show the neuroprotective effects of FGF9, but also clarify the critical mechanisms in HD cells, further providing an insight for the therapeutic potential of FGF9 in HD.

Topics: Animals; Apoptosis; bcl-X Protein; Butadienes; Caspase 3; Cell Line; Extracellular Signal-Regulated MAP Kinases; Fibroblast Growth Factor 9; Glial Cell Line-Derived Neurotrophic Factor; Huntington Disease; JNK Mitogen-Activated Protein Kinases; MAP Kinase Signaling System; Mice; Mice, Transgenic; Nitriles; Proto-Oncogene Proteins c-akt; Visual Cortex

The therapeutic potential of cannabinoids in Huntington's disease (HD) has been investigated by several groups with complex and sometimes contrasting results. We sought to examine key points of intersection between cannabinoid receptor 1 (CB(1)) signalling, survival and the formation of mutant huntingtin aggregates in HD.. Using a simplified pheochromocytoma (PC12) cell model of HD expressing exon 1 of wild-type or mutant huntingtin, we assayed cell death and aggregate formation using high-throughput cytotoxicity and image-based assays respectively.. CB(1) activation by HU210 conferred a small but significant level of protection against mutant huntingtin-induced cell death. Pertussis toxin uncoupled HU210 from the inhibition of cAMP, preventing rescue of cell death. Phosphorylation of extracellular signal-regulated kinase (ERK) was also critical to CB(1)-mediated rescue. Conversely, treatments that elevated cAMP exacerbated mutant huntingtin-induced cell death. Despite opposing effects on HD cell survival, both HU210 and compounds that elevated cAMP increased the formation of mutant huntingtin aggregates. The increase in aggregation by HU210 was insensitive to Pertussis toxin and UO126, suggesting a G-protein alpha subtype s (G(s))-linked mechanism.. We suggest that the CB(1) receptor, through G-protein alpha subtype i/o (G(i/o))-linked, ERK-dependent signal transduction, is a therapeutic target in HD. However the protective potential of CB(1) may be limited by promiscuous coupling to G(s), the stimulation of cAMP formation and increased aggregate formation. This may underpin the poor therapeutic efficacy of cannabinoids in more complex model systems and suggest that therapies that are selective for the G(i/o), ERK pathway may be of most benefit in HD.

Topics: Animals; Butadienes; Cell Culture Techniques; Cell Death; Cell Line, Transformed; Cerebellum; Cyclic AMP; Dronabinol; GTP-Binding Protein alpha Subunits; Humans; Huntingtin Protein; Huntington Disease; Mice; Mitogen-Activated Protein Kinase 3; Nerve Tissue Proteins; Neuroprotective Agents; Nitriles; Nuclear Proteins; PC12 Cells; Pertussis Toxin; Piperidines; Pyrazoles; Rats; Receptor, Cannabinoid, CB1; Receptors, Dopamine; Rimonabant