The generation and functional characterization of induced pluripotent stem cells from human intervertebral disc nucleus pulposus cells

Abstract

Disc degenerative disease (DDD) is believed to originate in the nucleus pulposus (NP) region therefore, it is important to obtain a greater number of active NP cells for the study and therapy of DDD. Human induced pluripotent stem cells (iPSCs) are a powerful tool for modeling the development of DDD in humans, and have the potential to be applied in regenerative medicine. NP cells were isolated from DDD patients following our improved method, and then the primary NP cells were reprogramed into iPSCs with Sendai virus vectors encoding 4 factors. Successful reprogramming of iPSCs was verified by the expression of surface markers and presence of teratoma. Differentiation of iPSCs into NP-like cells was performed in a culture plate or in hydrogel, whereby skin fibroblast derived-iPSCs were used as a control. Results demonstrated that iPSCs derived from NP cells displayed a normal karyotype, expressed pluripotency markers, and formed teratoma in nude mice. NP induction of iPSCs resulted in the expression of NP cell specific matrix proteins and related genes. Non-induced NP derived-iPSCs also showed some NP-like phenotype. Furthermore, NP-derived iPSCs differentiate much better in hydrogel than that in a culture plate. This is a novel method for the generation of iPSCs from NP cells of DDD patients, and we have successfully differentiated these iPSCs into NP-like cells in hydrogel. This method provides a novel treatment of DDD by using patient-specific NP cells in a relatively simple and straightforward manner.

Are specific gene expressions of extracellular matrix and nucleus pulposus affected by primer cell cultures prepared from intact or degenerative intervertebral disc tissues?

“…The authors' literature review suggests that there are no studies indicating a potential treatment modality for IVD (3,9,23,34).…”

“…They observed that IPSCSs, which originate from NP cells, induced the synthesizing of pluripotent markers in normal karyotype, NPcell specific matrix protein and related genes. They maintained that IPSCSs, which were produced from NPC cells that are peculiar to IVD cases, could be treated by a carrier scaffold (such as hydrogel) in IVD cases (34).…”

iASPP overexpression is associated with clinical outcome in spinal chordoma and influences cellular proliferation, invasion, and sensitivity to cisplatinin vitro

“…The primary cell culture method of NP was similar to others [ 55 ]. Briefly, human intervertebral disc NP tissues derived from the patients with LDH or LSS were rinsed and minced into small fragments of about 1 mm 3 .…”

Versatility of Induced Pluripotent Stem Cells (iPSCs) for Improving the Knowledge on Musculoskeletal Diseases

“…Since then, a wide variety of studies have reported the derivation of specific cell types from iPSCs, generating robust and reproducible phenotypes in vitro that reflect diseases intrinsic to the cellular level [ 35 ]. Specifically, evidence that reprogrammed cells can be differentiated into osteoblasts [ 54 , 55 , 56 ], chondrocytes [ 57 , 58 , 59 , 60 ], myoblasts [ 61 ], nucleus pulposus cells [ 47 , 48 , 49 ] and tenocytes [ 62 ] has recently impacted musculoskeletal research and changed orthopedic medicine. Nevertheless, the use of iPSCs for studying the wide variety of musculoskeletal conditions has been explored to a limited extent ( Table 1 ) in comparison with conditions affecting nervous or cardiac pathologies, even when investigating genetic disorders where symptoms affect several organs or systems [ 63 , 64 , 65 , 66 ].…”

“…Efficient and robust differentiation of iPSCs into tissue-specific cell types is crucial both for disease modeling and for regenerative medicine. Should the reader seek a more detailed explanation about iPSC differentiation protocols, please see the previously published studies on osteogenic [ 3 , 30 , 43 , 44 , 45 ], chondrogenic [ 3 , 27 , 30 , 46 ], nucleus pulposus [ 47 , 48 , 49 ] and/or myogenic [ 3 , 30 , 50 ] differentiation of iPSCs.…”