OC-001 THE ISOLATION, CULTURE AND CHARACTERISATION OF HUMAN RECTUS SHEATH FIBROBLASTS: THE BIRTH OF TISSUE ENGINEERING THE ABDOMINAL WALL AND A LIVING MESH?

Abstract

Abstract Aim To isolate human rectus sheath fibroblasts (hRSFs) from tissue samples, and characterise those cells - establishing their capacity to contract collagen matrix. Methods Samples of posterior rectus sheath were taken from 3 patients undergoing stoma formation. Samples underwent collagenase digestion, and isolated cells grown in culture media. Cultures were grown to confluence at 2nd passage, then cryopreserved. Cells were successfully thawed and re-animated at 3rd passage. 3 Cell lines of hRSFs and one of human dermal fibroblasts (HDFs) were all studied. Cell lines were assessed for their morphology and underwent immunofluorescent staining for Vimentin and α-smooth muscle actin. Cell lines were also seeded into collagen matrices to assess stromal contraction. Results Our protocol produced confluent clusters of fibroblasts within 2 weeks, and healthy cultures of cells into their 3rd passage within 6 weeks. Cells were successfully cryopreserved in liquid nitrogen after 3rd passage and were thawed and reanimated thereafter. Notable differences in morphology were observed between different cell lines – all of which stained positively for vimentin and α-smooth muscle actin. All cell lines induced a different speed of contraction within a collagen matrix from the HDFs achieving 82% contraction after 5 days, to 54% in the slowest RSF group. Conclusion Our work is the first to describe the isolation and characterisation of human rectus sheath fibroblasts and prove the concept of their introduction into 3D tissue models. This work opens exciting future prospects of both forming 3D models of rectus sheath tissue and in vitro mesh testing.