Proceedings of The Physiological Society

University of Oxford (2011) Proc Physiol Soc 23, C15

Oral Communications

VEGF-A165b is neuroprotective in in vitro models of diabetic and chemotherapy-induced sensory neuropathy and prevents ipsilateral DRG ATF-3 upregulation after nerve injury in vivo

N. Beazley-Long1, R. P. Hulse2, S. J. Harper1, D. O. Bates1, L. F. Donaldson2

1. Microvasular Research Laboratories, Physiology and Pharmacology, The University of Bristol, Bristol, United Kingdom. 2. School of Physiology and Pharmacology, The University of Bristol, Bristol, United Kingdom.


Alternative splicing of the Vascular Endothelial Growth Factor-A (VEGF-A) C-terminus results in the production of either pro-angiogenic (VEGF-Axxx) or anti-angiogenic (VEGF-Axxxb) isoforms (Bates et al., 2002). Over the last decade, the neuronal effects of VEGF-A have been recognised. VEGF-A165 promotes neurogenesis, axon extension and branching, and is neuroprotective in both the central and peripheral nervous systems. We previously identified a novel neuroprotective role for VEGF-A165b, the predominant VEGF-Axxxb isoform, for hippocampal neurones against L-glutamate-induced excitotoxicity, in vitro. Sensory neuronal damage and loss in diabetes and on chemotherapy lead to chronic pain and parastheias, severely affecting quality of life. We aimed to identify if VEGF-A165b is neuroprotective for peripheral sensory neurones in models of diabetic and chemotherapy-induced neuropathy, and in a traumatic nerve injury model. Adult rat (male Wistar, ~250g) dorsal root ganglion neurones were removed, dissociated mechanically and with collagenase, and plated onto coverslips. 30μg/mL 5-fluoro-2’-deoxyuridine was added to inhibit glial proliferation. After overnight pre-incubation with ±2.5nM VEGF-A165b cells were treated with 0, 5, 10, 20 μg/mL oxaliplatin (24h), or 0, 20, 40mM additional D-glucose (6h). The percentage of neurones positive for activated caspase-3 was determined by immunofluorescent staining. Treatment with 40mM glucose, or 20μg/mL oxaliplatin significantly increased the percentage of activated caspase-3-positive neurones compared to control (1-way ANOVA; 40mM glucose: 52.9±6.2% vs. 26.5±3.6%, p<0.05, n=3; 20μg/mL oxaliplatin 68.7±3.4% vs. 48.9±1.5%, p<0.01, n=3). Treatment with 2.5nM VEGF-A165b prevented this increase in the percentage of activated caspase-3-positive neurones induced by 40 mM hyperglycaemia (VEGF-A165b 34.8%±2.4, vehicle 52.9%±6.2, p<0.05, n=3) or 20μg/mL oxaliplatin (VEGF-A165b; 51.7±0.6%, vehicle 68.7±3.4%, p<0.01, n=3). To determine whether VEGF-A was neuroprotective in an in vivo nerve injury model, male C57Bl6 mice (25-30g) underwent partial saphenous nerve injury (Hulse et al., 2008) under brief isofluorane anaesthesia (2-3% isoflurane in oxygen). Nerve injury significantly increased immunofluorescence for the neuronal damage marker ATF-3 in ipsilateral L4 DRG (Kruskal-Wallis + Dunn’s, p<0.05, n=3) compared to sham, 12 days post surgery. Biweekly i.p. injection of 8ng/g VEGF-A165 or 20ng/g VEGF-A165b prevented the increase in ATF-3 immunofluorescence in ipsilateral L4 DRG. VEGF-A165b is neuroprotective for peripheral sensory neurones in vitro and in vivo. VEGF-A165b therapy may have potential as a preventative intervention or treatment for peripheral sensory neuropathies.

Where applicable, experiments conform with Society ethical requirements