Forsøksdyr: Alginate-cellulose and other alginate based microspheres; Novel microspheres as a mean to reduce the fibrosis currently limiting the use of alginate capsules in cell therapy

Godkjenningsdato 16.06.2020

Godkjenningsperiode 16.06.2020-15.06.2024

Alginates are biocompatible and non-toxic polysaccharides typically obtained from brown seaweeds. Furthermore, they form ionically crosslinked gels under physiological conditions allowing for applications within cell encapsulation and tissue engineering. In cell therapy, alginate gels restrict access to immune cells and has been proposed as a functional cure for Type 1 diabetes. Encapsulating insulin producing cells in alginate will mitigate the need for immunosuppressant therapy (adverse health effects). A current limitation of the use of alginates in cell therapy is fibrotic overgrowth, which causes graft failure following transplantation of pancreatic tissue.

This study aims to investigate the fibrotic host-response to microspheres of chemically modified alginates and to composite microspheres of alginate tunicate cellulose. Cellulose is a polysaccharide that is frequently used in conjunction with alginate in the production of implantable 3D-bioprinted constructs. Recently, the production of cellulose from tunicates has commenced in Norway. These celluloses show early promise, owing to an absence of impurities such as lignin and hemicellulose found in plant based celluloses and less endotoxins than those of bacterial origin.

Previously, we have investigated chemically modified alginates for the encapsulation of cells in cell therapy. Some of these candidate materials greatly reduced fibrosis. Herein we wish to further explore these materials in order to further mitigate fibrotic responses and to improve the recovery rate of alginate microspheres. The mouse strain C57BL/6J is known to provoke immune responses towards microcapsules and is relevant for fibrosis in monkeys and humans. Our hypothesis is that the new capsule designs will reduce the fibrotic reactions towards the biomaterial.

The animal study will consist of a pilot experiment using 30 mice to qualitatively assess the host-graft responses, and a subsequent main study of selected alginate capsules showing minimal/no fibrosis using up to 70 mice (8-12 animals per capsule design in total), giving a total of 100 mice. Empty capsules will be implanted into the peritoneal cavity of mice and explanted after two weeks - 4 months. The mild implantation procedure has few adverse effects. The procedure involves a small incision (maximum 1 cm) through the abdominal wall and peritoneum, where the capsules will be injected into the intraperitoneal cavity. The incision site will be closed with 1-3 stitches. Isofluran (2-3%) will be used as anaesthesia (reconvalence time is 5 minutes), and marcaine before implantation of capsules (0.05 ml subcutaneously at operation site) to hinder discomfort after the surgical treatment. The mice will be put in individual cages to mitigate stress responses. If the animals display abnormal behavior or signs of distress/pain (mouse grimace scale), inadequate wound healing at the site of injection before the end of experiment, the animal will be euthanized. The animals will be euthanized at the end of the experiment, before opening the animal and evaluation of capsules. The method of euthanization will be cervical dislocation with anaesthesia.

The current animal study will be correlated to investigations on the complement and cytokine responses in human whole blood, to study the transferability of the in vivo results to the in vitro findings.