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Pathophysiology of osteoarthritis (OA) is characterized by progressive loss of articular cartilage in the knee-joints. To impart regenerative ability in lowly metabolizing chondrocytes, the bone marrow stem cells (BMSCs) has recently been recognized as a superior alternative treatment for OA. However, study of primary BMSCs-mediated chondrogenesis is difficult due to progressive cellular aging and replicative senescence. To obtain a therapeutic cell population for OA, BMSCs were immortalized by human papilloma virus (HPV)-16 E6/E7 along with mCherry luciferase (mCL), a gene marker for non-invasive imaging, and designated as iBMSCs-mCL. Next, their cell morphology, population doubling time (PDT) and colony forming ability (CFU) were evaluated. Furthermore, pluripotency and immunophenotypic markers were investigated. To deduce therapeutic ability, iBMSCs-mCL were intra-articularly injected into right knee of anterior cruciate ligament transaction (ACLT)-OA mice model and tracked through non-invasive bioluminescence imaging. Cell morphology of iBMSCs-mCL was similar to parental BMSCs. PDT and CFU ability of iBMSCs-mCLs were significantly increased. Pluripotency and immunophenotypic markers were highly expressed in iBMSC-mCL. Long-term survival and tri-lineage differentiation particularly chondrogenic potential of iBMSCs-mCL were also demonstrated in vitro and then in vivo which was monitored through non-invasive imaging. Intensive bioluminescent signals in iBMSCs-mCL administered knee-joint indicated a marked in vivo survival and proliferation of iBMSCs-mCL. Immunohistochemical staining for type II collagen (IHC of Col II) and alcian blue &amp; safranin o staining of proteoglycans also corroborated cartilage regeneration by iBMSCs-mCL. Conclusively, iBMSCs-mCL maintains stemness and in vivo cartilage regeneration potential suggesting a promising avenue for development of OA therapeutics.

Non-invasive in vivo molecular imaging of intra-articularly transplanted immortalized bone marrow stem cells for osteoarthritis treatment