Generation of Human Induced Pluripotent Stem Cells from Anterior Cruciate Ligament

Human induced pluripotent stem cells (hiPSCs) are reprogrammed somatic cells and are an excellent cell source for tissue engineering applications, disease modelling and for understanding human development. HiPSC lines have now been generated from a diverse range of somatic cell types and have been reported to retain an epigenetic memory of their somatic origin. To date the reprogramming of a true ligament has not been reported. The aim of this study is to generate iPSCs from human anterior cruciate ligament cells. Anterior cruciate ligament cells from 3 above‐knee amputation donors, with donor matched dermal fibroblasts were tested for reprogramming using an existing dermal fibroblast reprogramming protocol. Anterior cruciate ligament cells were however more sensitive than donor matched dermal fibroblast to TGFβ; displaying marked contraction, increased proliferation and increased TNC and COMP expression in vitro, which hindered reprogramming to iPSCs. Modification of the protocol by scoring the cell monolayer or by removal of TGFβ during anterior cruciate ligament reprogramming resulted in emerging colonies being easier to identify and extract, increasing reprogramming efficiency. Following 30 passages in culture, the generated anterior cruciate ligament derived iPSCs displayed pluripotency markers, normal karyotype and can successfully differentiate to cells of the 3 embryonic germ layers. This study illustrates it is possible to generate hiPSCs from ligament, and identifies optimised ligament reprogramming conditions. Anterior cruciate ligament derived iPSCs may provide a promising cell source for ligament and related tissue engineering applications.