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Recombinant α2(IV)NC1 domain of type IV collagen is an effective regulator of retinal capillary endothelial cell proliferation and inhibits pre-retinal neovascularisation

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Abstract

Background

A recombinant form of the α2(IV)NC1 domain of type IV collagen has been shown to have potent anti-angiogenic activity although this peptide has not been studied in the context of proliferative retinopathies. In the current investigation we examined the potential for α2(IV)NC1 to regulate retinal microvascular endothelial cell function using a range of in vitro and in vivo assay systems.

Materials and methods

α2(IV)NC1 at concentrations between 0.1 and 1 μg/ml was added to retinal microvascular endothelial cells (RMECs) followed by assessment of cell attachment, proliferation and survival. This agent was also tested within a novel in vitro three-dimensional retinal angiogenesis assay and the number of angiogenic sprouts quantified. α2(IV)NC1 was also delivered intra-vitreally to mice with oxygen-induced proliferative retinopathy (OIR) and neovascularisation evaluated in comparison with vehicle-treated controls.

Results

RMECs treated with α2(IV)NC1 (0.1, 0.5 and 1 μg/ml) showed delayed attachment at 3 h post-seeding, although this deficit had been restored at the 6-h time point. BrdU assay of DNA replication revealed that confluent RMECs treated with α2(IV)NC1 showed no measurable response in comparison with vehicle-treated controls. By contrast, proliferation of sub-confluent RMECs was significantly reduced by α2(IV)NC1 at 0.5 μg/ml (P<0.01). α2(IV)NC1 also induced apoptosis in RMECs and inhibited angiogenesis of pre-existing retinal vascular networks in vitro (P<0.001). Intra-vitreal injection of α2(IV)NC1 in the OIR model significantly inhibited pre-retinal neovascularisation compared with vehicle-treated controls (P<0.001).

Conclusion

α2(IV)NC1 inhibits angiogenesis in the retinal microvasculature. This recombinant protein has potential for the treatment of neovascularisation in proliferative retinopathies.

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Authors and Affiliations

  1. Centre for Vision Science, School of Biomedical Science, Queen’s University Belfast, Royal Victoria Hospital, Grosvenor Road, Belfast, BT12 6BA, Northern Ireland, UK

    Gary Coleman, Tom A. Gardiner & Alan W. Stitt

  2. BioStratum Incorporated, Durham, NC, USA

    Ariel Boutaud

Authors
  1. Gary Coleman
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  2. Tom A. Gardiner
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  3. Ariel Boutaud
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  4. Alan W. Stitt
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Corresponding author

Correspondence to Alan W. Stitt.

Additional information

BioStratum Inc. did not sponsor this research in any way. None of the authors are paid consultants with this company.

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Coleman, G., Gardiner, T.A., Boutaud, A. et al. Recombinant α2(IV)NC1 domain of type IV collagen is an effective regulator of retinal capillary endothelial cell proliferation and inhibits pre-retinal neovascularisation. Graefe's Arch Clin Exp Ophthalmol 245, 581–587 (2007). https://doi.org/10.1007/s00417-006-0396-1

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  • DOI: https://doi.org/10.1007/s00417-006-0396-1

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