Original Article
Thermo-responsive polymeric nanoparticles for enhancing neuronal differentiation of human induced pluripotent stem cells

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Abstract

We report thermo-responsive retinoic acid (RA)-loaded poly(N-isopropylacrylamide)-co-acrylamide (PNIPAM-co-Am) nanoparticles for directing human induced pluripotent stem cell (hiPSC) fate. Fourier transform infrared spectroscopy and 1H nuclear magnetic resonance analysis confirmed that RA was efficiently incorporated into PNIAPM-co-Am nanoparticles (PCANs). The size of PCANs dropped with increasing temperatures (300-400 nm at room temperature, 80-90 nm at 37 °C) due to its phase transition from hydrophilic to hydrophobic. Due to particle shrinkage caused by this thermo-responsive property of PCANs, RA could be released from nanoparticles in the cells upon cellular uptake. Immunocytochemistry and quantitative real-time polymerase chain reaction analysis demonstrated that neuronal differentiation of hiPSC-derived neuronal precursors was enhanced after treatment with 1-2 μg/ml RA-loaded PCANs. Therefore, we propose that this PCAN could be a potentially powerful carrier for effective RA delivery to direct hiPSC fate to neuronal lineage.

From the Clinical Editor

The use of induced pluripotent stem cells (iPSCs) has been at the forefront of research in the field of regenerative medicine, as these cells have the potential to differentiate into various terminal cell types. In this article, the authors utilized a thermo-responsive polymer, Poly(N-isopropylacrylamide) (PNIPAM), as a delivery platform for retinoic acid. It was shown that neuronal differentiation could be enhanced in hiPSC-derived neuronal precursor cells. This method may pave a way for future treatment of neuronal diseases.

Graphical Abstract

Thermo-responsive polymeric nanoparticle-mediated neuronal differentiation from the human induced pluripotent stem cells.

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Section snippets

Synthesis of PCANs

To synthesize PCANs, 500 mg NIPAM (Sigma-Aldrich, St. Louis, MO, USA), 13.5 mg N,N′-Methylenebisacrylamide (MBA, Sigma-Aldrich), 100 mg sodium dodecyl sulfate (SDS, Sigma-Aldrich), and 12.5 mg Am (Sigma-Aldrich) were added to 50 ml distilled water. The solution was mixed with nitrogen gas at room temperature for 20 minutes. Potassium persulfate (37.5 mg, KPS, Dae Jung, Siheung, Gyonggi, Korea) was added to initiate polymerization. The solution was then allowed to react with nitrogen gas at 70 °C for 5 

Synthesis and analysis of RA-loaded PCANs

PCANs were synthesized through a radical polymerization reaction (Figure 1, A). TEM images revealed the spherical morphology of PCANs (Figure 1, B). DLS analysis showed that the diameter of PCANs was approximately 300-400 nm at room temperature. When the temperature was raised above the LCST (35 °C), the diameter of the nanoparticles shrank to 80-90 nm (Figure 1, C). The size distribution of PCANs determined by DLS analysis indicated that D10, D50, and D90 were 43.49 ± 0.41 nm, 65.09 ± 1.27 nm, and 98.38

Discussion

The thermo-responsive property is one of the major features of RA-loaded PCANs. At temperatures above the LCST, the hydrogen bonds between the amide groups of PCANs and water molecules were broken, and the coil-to-globule transition began. This caused the phase transition of the PCANs from hydrophilic to hydrophobic. This phase transition induced a deswelling of PCANs (Figure 1, D), leading to RA release from PCANs in a temperature-dependent manner (Figure 1, E). The surface charge of

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    This research was supported by a grant from the Korea Health Technology R&D Project through the Korea Health Industry Development Institute, funded by the Ministry of Health & Welfare, Republic of Korea (Grant number HI14C3347); by the Bio & Medical Technology Development Program of the National Research Foundation (NRF) funded by the Ministry of Science, ICT & Future Planning (2015M3A9D7030461); by a grant (NRF-2010-0020409) from the National Research Foundation of Korea (NRF), funded by the MSIP, Republic of Korea; and in part by a grant (HI14C1588) from the Korea Health Technology R&D Project, funded by the Ministry of Health and Welfare, Republic of Korea.

    1

    These authors contributed equally to this work.

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