Belgio, Beatrice (Politecnico di Milano)


"Introduction: Age-related macular degeneration (AMD) is a major cause of blindness worldwide, affecting 170 million people1. AMD consists in a degeneration of the central retina due to age-related changes in Bruch’s membrane (BM) and in the retinal pigment epithelium (RPE). Currently, periodic intravitreal injections of anti-vascular endothelial growth factor drugs are the gold standard therapy in the management of AMD. However, these drugs are unable to restore tissue functionality. Retinal tissue engineering may be helpful in providing better solutions. This work aims at designing a functional BM.

Methodology: We fabricated electrospun nanofibrous membranes composed of Bombyx mori silk fibroin (BMSF) and polycaprolactone (PCL). We investigated both aligned and randomly oriented nanofibrous mats to study the influence of fibre orientation on tissue regeneration. We evaluated the following scaffold properties: fibre morphology, thickness, permeability, mechanical properties, in vitro cytocompatibility, and the formation of mature epithelial monolayers. We employed ARPE-19 cells and zonula occludens (ZO)-1 staining for in vitro studies.

Results: BMSF/PCL membranes with aligned and random fibres were successfully produced by electrospinning. The thickness of aligned and randomly oriented fibrous mats was 58.6 µm and 40 µm, respectively. The randomly oriented fibrous membrane displayed an average fibre diameter of 938±223 nm, whereas the aligned fibres showed a slightly higher average diameter. Mechanical properties were investigated as they are crucial for BM structural role and surgical applications. The elastic modulus of the aligned and randomly oriented fibrous mats was 43.4±9.1 and 13±3.6 MPa, respectively. Transport is another important function of native BM; therefore, membrane hydraulic conductivity (Lp) was assessed. We found a Lp of 2.35×10-8 and 20x10-10 m/(Pa*s) for the aligned and randomly oriented fibrous membranes, respectively. Moreover, we proved the biocompatibility of the mats and the formation of a functional epithelial monolayer. The expression of ZO-1 on the aligned fibrous membrane presented a higher staining intensity than on the randomly oriented fibrous scaffolds suggesting that the fibre orientation promotes a proper cell organization.

Conclusion: An ideal substrate for constructing a prosthetic BM with attached RPE cells has yet to be found. Both the obtained mats showed similar properties to human BM, which has a fibrillar network, a elastic modulus ranging from 6 to 14 MPa, and a hydraulic conductivity ranging from 20 to 100 x10-10 m/(Pa*s)2. Currently, we are carrying out additional studies to confirm that oriented fibres enhance biological tissue regeneration. The successful outcome of this study will inform the treatment of an optimal substrate for RPE transplantation.

Acknowledgments: We thank Leonardino SRL for scaffold technical support.

1. Wong W. L., et al. Lancet Glob. Health 2(2), 106-116 (2014).
2. Chan, W.H., et al. IOVS, 48, 2187 (2007)."


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