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In the context of stroke-induced brain damage, the molecular and biochemical mechanisms involving retraction and collapse of the axonal network remain unclear. One of the early morphological changes accompanying excitotoxicity-induced neuronal death in cultured neurons is the retraction/collapse of the neurite network, which indicates that axonal damage occurs before the emergence of typical morphological hallmarks of neuronal death (Deckwerth and Johnson 1994;Raff et al. 2002). Typically, axonal degeneration is manifested by irregular blebbing of axons with thinning and fragmentation, followed by retraction and collapse of the axonal network. While axonal damage was regarded as an outcome of the death process occurring within the cell body, more importantly, it Address correspondence and reprint requests to Sheng T. Hou, Institute for Biological Sciences, National Research Council Canada, 1200 Montreal Road, Bldg M54, Ottawa, ON, Canada, K1A 0R6. E-mail: sheng.hou@nrc-cnrc.gc.caAbbreviations used: AIPII, autocamtide-2-related inhibitory peptide; AKT, protein kinase B; CaMKII, Ca 2+ /calmodulin-dependent protein kinase II; CRMP, collapsin response mediator protein; DAPI, 4¢,6-diamidino-2-phenylindole; EGFP, enhanced green fluorescent protein; GSK, glycogen synthase kinase; MAP2, microtubule-associated protein 2; MAPK, mitogen-activated protein kinase; MCAO, middle cerebral artery occlusion; MK801, dizocilpine; PI3K, phosphoinositide 3-kinase; PSD, post-synaptic density; TUNEL, terminal deoxynucleotidyl transferase dUTP nick end labeling. AbstractIntracellular calcium influx through NMDA receptors triggers a cascade of deleterious signaling events which lead to neuronal death in neurological conditions such as stroke. However, it is not clear as to the molecular mechanism underlying early damage response from axons and dendrites which are important in maintaining a network essential for the survival of neurons. Here, we examined changes of axons treated with glutamate and showed the appearance of bIII-tubulin positive varicosities on axons before the appearance of neuronal death. Dizocilpine blocked the occurrence of varicosities on axons suggesting that these microstructures were mediated by NMDA receptor activities. Despite early increased expression of pCaMKII and pMAPK after just 10 min of glutamate treatment, only inhibitors to Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) and calpain prevented the occurrence of axonal varicosities. In contrast, inhibitors to Rho kinase, mitogen-activated protein kinase and phosphoinositide 3-kinase were not effective, nor were they able to rescue neurons from death, suggesting CaMKII and calpain are important in axon survival. Activated CaMKII directly phosphorylates collapsin response mediator protein (CRMP) 2 which is independent of calpain-mediated cleavage of CRMP2. Over-expression of CRMP2, but not the phosphorylation-resistant mutant CRMP2-T555A, increased axonal resistance to glutamate toxicity with reduced numbers of varicosities. The levels of both pCRMP2 ...

As a well-known neurotrophic factor, nerve growth factor (NGF) has also been extensively recognized for its acceleration of healing in cutaneous wounds in both animal models and randomized clinical trials. However, the underlying mechanisms accounting for the therapeutic effect of NGF on skin wounds are not fully understood. NGF treatment significantly accelerated the rate of wound healing by promoting wound reepithelialization, the formation of granulation tissue, and collagen production. To explore the possible mechanisms of this process, the expression levels of CD68, VEGF, PCNA, and TGF-β1 in wounds were detected by immunohistochemical staining. The levels of these proteins were all significantly raised in NGF-treated wounds compared to untreated controls. NGF also significantly promoted the migration, but not the proliferation, of dermal fibroblasts. NGF induced a remarkable increase in the activity of PI3K/Akt, JNK, ERK, and Rac1, and blockade with their specific inhibitors significantly impaired the NGF-induced migration. In conclusion, NGF significantly accelerated the healing of skin excisional wounds in rats and the fibroblast migration induced by NGF may contribute to this healing process. The activation of PI3K/Akt, Rac1, JNK, and ERK were all involved in the regulation of NGF-induced fibroblast migration.

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