Xeno-free and minimally invasive approach for ocular therapies based on mesenchymal stem cells

Abstract

Purpose : The therapeutic potential of mesenchymal stem cells (MSCs) has been heralded by their multipotentiality and immunomodulatory capacity. The potential use of MSCs at the ocular surface is based on two translational objectives: limiting the progression of corneal inflammatory disease and promoting corneal tissue regeneration. However, current MSCs treatments include arduous cell isolation protocols that include xenogeneic components or allogeneic scaffolds for MSC delivery, constraining its use due to challenges in the manufacture of clinical grade products. In this study, we introduce a xeno-free and minimally invasive technique for MSC isolation, expansion and delivery using Plasma Rich in Growth Factors technology (PRGF-Endoret®)

Methods : PRGF-Endoret® eye drops were obtained using the Endoret® kit in Ophthalmology. Whole blood was collected by venipuncture from healthy donors and PRGF was used as culture media supplement and as a PRGF-based scaffold for MSCs delivery after plasma activation with CaCl₂.
Human adipose tissue samples were obtained from peripatellar fat pads of multiorganic donors after informed written consent. Adipose fragments of 1-2 mm² were cultured, without previous enzymatic digestion, on a 12-well plate using DMEM supplemented with 10% PRGF and 1% antibiotics. MSCs were characterized by flow cytometry against surface protein markers and were subjected to in vitro differentiation to osteoblast, adipocytes, and chondrocytes.
MSCs were seeded on PRGF scaffolds and cultured for 48 hours. Cultured membranes were grafted in the subcutaneous space of nude mice; and after 21 and 35 days, animals were euthanized and tissue samples were subjected to immunohistochemical analysis

Results : MSCs expressed normal phenotype as assessed by flow cytometry and retained plasticity to differentiate in to a variety of mesenchymal lineages when cultured using xeno-free culture media. After in vivo implantation, MSCs were located in the subdermal space suggesting that MSCs may still retain immunoregulatory properties up to 35 days upon implantation

Conclusions : The present results could accelerate the translation of MSCs from the bench to the treatment of ocular surface disease due to its ease to be upgraded to clinical grade manufacture, setting the future direction of standardization of MSC-based therapies using a xeno-free and minimally invasive approach for MSCs isolation, expansion and delivery

This is a 2020 ARVO Annual Meeting abstract.