Growth Factor-free Chondrogenic Differentiation from Induced Pluripotent Stem Cells using Minicircle Vectors

The human degenerative cartilage has low regenerative potential. Chondrocyte transplantation offers a promising strategy for cartilage treatment and regeneration. Currently chondrogenesis using human pluripotent stem cells are accomplished using human recombinant growth factors. Here, we differentiated human induced pluripotent stem cells (hiPSCs) into chondrocytes and cartilage pellet using minicircle vectors. Minicircles are used as a non-viral gene delivery system for gene therapy in various diseases. Non-viral gene delivery can produce growth factors without integrating into the host genome. Minicircle vectors containing bone morphogenetic protein 2 (BMP2) and transforming growth factor, beta 3 (TGFβ3) were successfully generated and delivered to hiPSC-derived outgrowth (OG) cells. Cell pellets generated using minicircle-transfected OG cells successfully differentiated into chondrogenic lineage. Chondrogenic pellets transfected with growth factor-encoding minicircles effectively recovered osteochondral defect in rat models. Taken together, this work shows the potential application of minicircles in cartilage regeneration using hiPSCs.

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Generation of induced pluripotent stem cells using elastin like polypeptides as a non-viral gene delivery system

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease ◽

10.1016/j.bbadis.2019.01.031 ◽

2020 ◽

Vol 1866 (4) ◽

pp. 165405 ◽

Cited By ~ 4

Author(s):

Chang Hyun Lee ◽

Rohan S.J. Ingrole ◽

Harvinder Singh Gill

Keyword(s):

Stem Cells ◽

Gene Delivery ◽

Induced Pluripotent Stem Cells ◽

Delivery System ◽

Pluripotent Stem Cells ◽

Viral Gene ◽

Gene Delivery System ◽

Induced Pluripotent ◽

Viral Gene Delivery

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Simple Generation of Human Induced Pluripotent Stem Cells Using Poly-β-amino Esters As the Non-viral Gene Delivery System

Journal of Biological Chemistry ◽

10.1074/jbc.m110.168013 ◽

2011 ◽

Vol 286 (14) ◽

pp. 12417-12428 ◽

Cited By ~ 50

Author(s):

Núria Montserrat ◽

Elena Garreta ◽

Federico González ◽

Jordán Gutiérrez ◽

Cristina Eguizábal ◽

...

Keyword(s):

Stem Cells ◽

Gene Delivery ◽

Induced Pluripotent Stem Cells ◽

Delivery System ◽

Pluripotent Stem Cells ◽

Viral Gene ◽

Gene Delivery System ◽

Amino Esters ◽

Induced Pluripotent ◽

Viral Gene Delivery

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Chondrogenic Differentiation from Induced Pluripotent Stem Cells Using Non-Viral Minicircle Vectors

Cells ◽

10.3390/cells9030582 ◽

2020 ◽

Vol 9 (3) ◽

pp. 582 ◽

Cited By ~ 5

Author(s):

Yeri Alice Rim ◽

Yoojun Nam ◽

Narae Park ◽

Hyerin Jung ◽

Kijun Lee ◽

...

Keyword(s):

Stem Cells ◽

Growth Factors ◽

Transforming Growth Factor Beta ◽

Pluripotent Stem Cells ◽

Transforming Growth Factor ◽

Cartilage Regeneration ◽

Bone Morphogenetic Protein 2 ◽

Viral Gene ◽

Morphogenetic Protein ◽

Induced Pluripotent

Human degenerative cartilage has low regenerative potential. Chondrocyte transplantation offers a promising strategy for cartilage treatment and regeneration. Currently, chondrogenesis using human pluripotent stem cells (hiPSCs) is accomplished using human recombinant growth factors. Here, we differentiate hiPSCs into chondrogenic pellets using minicircle vectors. Minicircles are a non-viral gene delivery system that can produce growth factors without integration into the host genome. We generated minicircle vectors containing bone morphogenetic protein 2 (BMP2) and transforming growth factor beta 3 (TGFβ3) and delivered them to mesenchymal stem cell-like, hiPSC-derived outgrowth (OG) cells. Cell pellets generated using minicircle-transfected OG cells successfully differentiated into the chondrogenic lineage. The implanted minicircle-based chondrogenic pellets recovered the osteochondral defects in rat models. This work is a proof-of-concept study that describes the potential application of minicircle vectors in cartilage regeneration using hiPSCs.

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A miniature dialysis-culture device allows high-density human-induced pluripotent stem cells expansion from growth factor accumulation

Communications Biology ◽

10.1038/s42003-021-02848-x ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Fuad Gandhi Torizal ◽

Qiao You Lau ◽

Masato Ibuki ◽

Yoshikazu Kawai ◽

Masato Horikawa ◽

...

Keyword(s):

Stem Cells ◽

Shear Stress ◽

Growth Factors ◽

Growth Factor ◽

Induced Pluripotent Stem Cells ◽

Pluripotent Stem Cells ◽

High Density ◽

Induced Pluripotent ◽

Dialysis Culture ◽

Factor Accumulation

AbstractThree-dimensional aggregate-suspension culture is a potential biomanufacturing method to produce a large number of human induced pluripotent stem cells (hiPSCs); however, the use of expensive growth factors and method-induced mechanical stress potentially result in inefficient production costs and difficulties in preserving pluripotency, respectively. Here, we developed a simple, miniaturized, dual-compartment dialysis-culture device based on a conventional membrane-culture insert with deep well plates. The device improved cell expansion up to approximately ~3.2 to 4×107 cells/mL. The high-density expansion was supported by reduction of excessive shear stress and agglomeration mediated by the addition of the functional polymer FP003. The results revealed accumulation of several growth factors, including fibroblast growth factor 2 and insulin, along with endogenous Nodal, which acts as a substitute for depleted transforming growth factor-β1 in maintaining pluripotency. Because we used the same growth-factor formulation per volume in the upper culture compartment, the cost reduced in inverse proportional manner with the cell density. We showed that growth-factor-accumulation dynamics in a low-shear-stress environment successfully improved hiPSC proliferation, pluripotency, and differentiation potential. This miniaturised dialysis-culture system demonstrated the feasibility of cost-effective mass production of hiPSCs in high-density culture.

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