Therapeutic Effects of rAAV-Mediated Concomittant Gene Transfer and Overexpression of TGF-β and IGF-I on the Chondrogenesis of Human Bone-Marrow-Derived Mesenchymal Stem Cells.

Stephanie Morscheid,Jagadeesh Kumar Venkatesan,Ana Rey-Rico,Magali Cucchiarini,Gertrud Schmitt,Henning Madry

doi:10.3390/ijms20102591

Stephanie Morscheid, Jagadeesh Kumar Venkatesan + Show 4 more

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https://doi.org/10.3390/ijms20102591

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Abstract

Application of chondroreparative gene vectors in cartilage defects is a powerful approach to directly stimulate the regenerative activities of bone-marrow-derived mesenchymal stem cells (MSCs) that repopulate such lesions. Here, we investigated the ability of combined recombinant adeno-associated virus (rAAV) vector-mediated delivery of the potent transforming growth factor beta (TGF-β) and insulin-like growth factor I (IGF-I) to enhance the processes of chondrogenic differentiation in human MSCs (hMSCs) relative to individual candidate treatments and to reporter (lacZ) gene condition. The rAAV-hTGF-β and rAAV-hIGF-I vectors were simultaneously provided to hMSC aggregate cultures (TGF-β/IGF-I condition) in chondrogenic medium over time (21 days) versus TGF-β/lacZ, IGF-I/lacZ, and lacZ treatments at equivalent vector doses. The cultures were then processed to monitor transgene (co)-overexpression, the levels of biological activities in the cells (cell proliferation, matrix synthesis), and the development of a chondrogenic versus osteogenic/hypertrophic phenotype. Effective, durable co-overexpression of TGF-β with IGF-I via rAAV enhanced the proliferative, anabolic, and chondrogenic activities in hMSCs versus lacZ treatment and reached levels that were higher than those achieved upon single candidate gene transfer, while osteogenic/hypertrophic differentiation was delayed over the period of time evaluated. These findings demonstrate the potential of manipulating multiple therapeutic rAAV vectors as a tool to directly target bone-marrow-derived MSCs in sites of focal cartilage defects and to locally enhance the endogenous processes of cartilage repair.

Highlights

Articular cartilage lesions are prevalent clinical issues that impede the whole musculoskeletal system and for which there is no definitive cure to date [1,2]
Successful recombinant adeno-associated virus (rAAV)-Mediated TGF-β and insulin-like growth factor I (IGF-I) Co-Overexpression in human MSCs (hMSCs) Aggregates hMSC aggregates were first transduced over time with rAAV according to the study design (Figure 1) in order to evaluate the ability of the vectors to promote the co-overexpression of the candidate TGF-β and IGF-I genes (TGF-β/IGF-I) relative to independent gene application (TGF-β/lacZ or IGF-I/lacZ) and control treatment
Based on the perception that combined treatments may be more beneficial than single approaches [12,21], we tested the hypothesis that concomitant administration of two highly chondrogenic genes (TGF-β and IGF-I) stimulates the pro-chondrogenic activities of human mesenchymal stem cells (MSCs) relative to single and control gene conditions

Summary

Introduction

Articular cartilage lesions are prevalent clinical issues that impede the whole musculoskeletal system and for which there is no definitive cure to date [1,2]. Cartilage gene therapy that aims at activating the intrinsic chondroreparative activities of bone-marrow-derived mesenchymal stem cells (MSCs) [8,9,10] may be envisaged by directly providing therapeutic gene vehicles in sites of cartilage injury [11,12] especially using clinically relevant recombinant adeno-associated virus (rAAV) vectors [13] that support the safe, effective, and durable transduction of these cells (up to 100% efficiencies for at least 3 weeks) without alteration of their chondrogenic potential [14,15,16,17,18,19,20] Such a strategy has been reported using a number of therapeutic (chondrogenic) candidates such as the transforming growth factor beta (TGF-β) [14,18], basic fibroblast growth factor (FGF-2) [16], insulin-like growth factor I (IGF-I) [19], and the sex-determining region Y-type high-mobility group box 9 transcription factor (SOX9) alone [17] or combined with TGF-β [20], promoting the chondrogenic differentiation of the cells only to a certain extent and showing the need to explore new setups and combinations that might be more valuable than single treatments [12,21]

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