Publications by authors named "Hyeon-Ki Jang"

20 Publications

  • Page 1 of 1

Quantitative assessment of engineered Cas9 variants for target specificity enhancement by single-molecule reaction pathway analysis.

Nucleic Acids Res 2021 Sep 23. Epub 2021 Sep 23.

Department of Chemistry, Seoul National University, Seoul 08826, Republic of Korea.

There have been many engineered Cas9 variants that were developed to minimize unintended cleavage of off-target DNAs, but detailed mechanism for the way they regulate the target specificity through DNA:RNA heteroduplexation remains poorly understood. We used single-molecule FRET assay to follow the dynamics of DNA:RNA heteroduplexation for various engineered Cas9 variants with respect to on-target and off-target DNAs. Just like wild-type Cas9, these engineered Cas9 variants exhibit a strong correlation between their conformational structure and nuclease activity. Compared with wild-type Cas9, the fraction of the cleavage-competent state dropped more rapidly with increasing base-pair mismatch, which gives rise to their enhanced target specificity. We proposed a reaction model to quantitatively analyze the degree of off-target discrimination during the successive process of R-loop expansion. We found that the critical specificity enhancement step is activated during DNA:RNA heteroduplexation for evoCas9 and HypaCas9, while it occurs in the post-heteroduplexation stage for Cas9-HF1, eCas9, and Sniper-Cas9. This study sheds new light on the conformational dynamics behind the target specificity of Cas9, which will help strengthen its rational designing principles in the future.
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http://dx.doi.org/10.1093/nar/gkab858DOI Listing
September 2021

High-purity production and precise editing of DNA base editing ribonucleoproteins.

Sci Adv 2021 Aug 27;7(35). Epub 2021 Aug 27.

Department of Chemistry and Research Institute for Convergence of Basic Sciences, Hanyang University, Seoul 04763, South Korea.

Ribonucleoprotein (RNP) complex-mediated base editing is expected to be greatly beneficial because of its reduced off-target effects compared to plasmid- or viral vector-mediated gene editing, especially in therapeutic applications. However, production of recombinant cytosine base editors (CBEs) or adenine base editors (ABEs) with ample yield and high purity in bacterial systems is challenging. Here, we obtained highly purified CBE/ABE proteins from a human cell expression system and showed that CBE/ABE RNPs exhibited different editing patterns (i.e., less conversion ratio of multiple bases to single base) compared to plasmid-encoded CBE/ABE, mainly because of the limited life span of RNPs in cells. Furthermore, we found that off-target effects in both DNA and RNA were greatly reduced for ABE RNPs compared to plasmid-encoded ABE. We ultimately applied NG PAM-targetable ABE RNPs to in vivo gene correction in retinal degeneration 12 () model mice.
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http://dx.doi.org/10.1126/sciadv.abg2661DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8397273PMC
August 2021

Safe scarless cassette-free selection of genome-edited human pluripotent stem cells using temporary drug resistance.

Biomaterials 2020 12 1;262:120295. Epub 2020 Sep 1.

College of Pharmacy, Seoul National University, Seoul, South Korea. Electronic address:

An efficient gene-editing technique for use in human pluripotent stem cells (hPSCs) has great potential value in regenerative medicine, as well as in drug discovery based on isogenic human disease models. However, the extremely low efficiency of gene editing in hPSCs remains as a major technical hurdle. Previously, we demonstrated that YM155, a survivin inhibitor developed as an anti-cancer drug, induces highly selective cell death in undifferentiated hPSCs. In this study, we demonstrated that the high cytotoxicity of YM155 in hPSCs, which is mediated by selective cellular uptake of the drug, is due to the high expression of SLC35F2 in these cells. Knockout of SLC35F2 with CRISPR-Cas9, or depletion with siRNAs, made the hPSCs highly resistant to YM155. Simultaneous editing of a gene of interest and transient knockdown of SLC35F2 following YM155 treatment enabled the survival of genome-edited hPSCs as a result of temporary YM155 resistance, thereby achieving an enriched selection of clonal populations with gene knockout or knock-in. This precise and efficient genome editing approach took as little as 3 weeks and required no cell sorting or the introduction of additional genes, to be a more feasible approach for gene editing in hPSCs due to its simplicity.
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http://dx.doi.org/10.1016/j.biomaterials.2020.120295DOI Listing
December 2020

Current trends in gene recovery mediated by the CRISPR-Cas system.

Exp Mol Med 2020 07 10;52(7):1016-1027. Epub 2020 Jul 10.

Department of Chemistry and Research Institute for Convergence of Basic Sciences, Hanyang University, Seoul, 04763, South Korea.

The CRISPR-Cas system has undoubtedly revolutionized the genome editing field, enabling targeted gene disruption, regulation, and recovery in a guide RNA-specific manner. In this review, we focus on currently available gene recovery strategies that use CRISPR nucleases, particularly for the treatment of genetic disorders. Through the action of DNA repair mechanisms, CRISPR-mediated DNA cleavage at a genomic target can shift the reading frame to correct abnormal frameshifts, whereas DNA cleavage at two sites, which can induce large deletions or inversions, can correct structural abnormalities in DNA. Homology-mediated or homology-independent gene recovery strategies that require donor DNAs have been developed and widely applied to precisely correct mutated sequences in genes of interest. In contrast to the DNA cleavage-mediated gene correction methods listed above, base-editing tools enable base conversion in the absence of donor DNAs. In addition, CRISPR-associated transposases have been harnessed to generate a targeted knockin, and prime editors have been developed to edit tens of nucleotides in cells. Here, we introduce currently developed gene recovery strategies and discuss the pros and cons of each.
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http://dx.doi.org/10.1038/s12276-020-0466-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8080666PMC
July 2020

Anti-Atherogenic Effect of Stem Cell Nanovesicles Targeting Disturbed Flow Sites.

Small 2020 04 2;16(16):e2000012. Epub 2020 Apr 2.

Department of Medical Engineering, Yonsei University College of Medicine, Seoul, 03722, Republic of Korea.

Atherosclerosis development leads to irreversible cascades, highlighting the unmet need for improved methods of early diagnosis and prevention. Disturbed flow formation is one of the earliest atherogenic events, resulting in increased endothelial permeability and subsequent monocyte recruitment. Here, a mesenchymal stem cell (MSC)-derived nanovesicle (NV) that can target disturbed flow sites with the peptide GSPREYTSYMPH (PREY) (PMSC-NVs) is presented which is selected through phage display screening of a hundred million peptides. The PMSC-NVs are effectively produced from human MSCs (hMSCs) using plasmid DNA designed to functionalize the cell membrane with PREY. The potent anti-inflammatory and pro-endothelial recovery effects are confirmed, similar to those of hMSCs, employing mouse and porcine partial carotid artery ligation models as well as a microfluidic disturbed flow model with human carotid artery-derived endothelial cells. This nanoscale platform is expected to contribute to the development of new theragnostic strategies for preventing the progression of atherosclerosis.
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http://dx.doi.org/10.1002/smll.202000012DOI Listing
April 2020

i-Silence, Please! An Alternative for Gene Disruption via Adenine Base Editors.

Mol Ther 2020 02 17;28(2):348-349. Epub 2020 Jan 17.

Department of Chemistry, Hanyang University and Research Institute for Convergence of Basic Sciences, Hanyang University, Seoul 04763, South Korea. Electronic address:

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http://dx.doi.org/10.1016/j.ymthe.2020.01.009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7001083PMC
February 2020

A Disposable Photovoltaic Patch Controlling Cellular Microenvironment for Wound Healing.

Int J Mol Sci 2018 Oct 4;19(10). Epub 2018 Oct 4.

Department of BioNano Technology, Gachon University, Seongnam 13120, Korea.

Electrical stimulation (ES) is known to affect the wound healing process by modulating skin cell behaviors. However, the conventional clinical devices that can generate ES for promoting wound healing require patient hospitalization due to large-scale of the extracorporeal devices. Herein, we introduce a disposable photovoltaic patch that can be applied to skin wound sites to control cellular microenvironment for promoting wound healing by generating ES. In vitro experiment results show that exogenous ES could enhance cell migration, proliferation, expression of extracellular matrix proteins, and myoblast differentiation of fibroblasts which are critical for wound healing. Our disposable photovoltaic patches were attached to the back of skin wound induced mice. Our patch successfully provided ES, generated by photovoltaic energy harvested from the organic solar cell under visible light illumination. In vivo experiment results show that the patch promoted cutaneous wound healing via enhanced host-inductive cell proliferation, cytokine secretion, and protein synthesis which is critical for wound healing process. Unlike the current treatments for wound healing that engage passive healing processes and often are unsuccessful, our wearable photovoltaic patch can stimulate regenerative activities of endogenous cells and actively contribute to the wound healing processes.
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http://dx.doi.org/10.3390/ijms19103025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6213857PMC
October 2018

Therapeutic Angiogenesis via Solar Cell-Facilitated Electrical Stimulation.

ACS Appl Mater Interfaces 2017 Nov 27;9(44):38344-38355. Epub 2017 Oct 27.

Department of BioNano Technology, Gachon University , Seongnam 13120, Republic of Korea.

Cell therapy has been suggested as a treatment modality for ischemic diseases, but the poor survival and engraftment of implanted cells limit its therapeutic efficacy. To overcome such limitation, we used electrical stimulation (ES) derived from a wearable solar cell for inducing angiogenesis in ischemic tissue. ES enhanced the secretion of angiogenic growth factors and the migration of mesenchymal stem cells (MSCs), myoblasts, endothelial progenitor cells, and endothelial cells in vitro. In a mouse ischemic hindlimb model, ES generated by a solar cell and applied to the ischemic region promoted migration of MSCs toward the ischemic site and upregulated expression of angiogenic paracrine factors (vascular endothelial, basic fibroblast, and hepatocyte growth factors; and stromal cell-derived factor-1α). Importantly, solar cell-generated ES promoted the formation of capillaries and arterioles at the ischemic region, attenuated muscle necrosis and fibrosis, and eventually prevented loss of the ischemic limb. Solar cell ES therapy showed higher angiogenic efficacy than conventional MSC therapy. This study shows the feasibility of using solar cell ES as a novel treatment for therapeutic angiogenesis.
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http://dx.doi.org/10.1021/acsami.7b13322DOI Listing
November 2017

In vivo monitoring of angiogenesis in a mouse hindlimb ischemia model using fluorescent peptide-based probes.

Amino Acids 2016 07 20;48(7):1641-54. Epub 2016 Apr 20.

Center for Theragnosis, Biomedical Research Institute, Korea Institute of Science and Technology, 39-1 Hawolgok-dong, Seongbuk-gu, Seoul, 136-791, Republic of Korea.

Vascular endothelial growth factor receptor (VEGFR) and matrix metalloproteinase (MMP) are up-regulated in ischemic tissue and play pivotal roles in promoting angiogenesis. The purpose of the present study was to evaluate two fluorophore-conjugated peptide probes specific to VEGFR and MMP for dual-targeted in vivo monitoring of angiogenesis in a murine model of hindlimb ischemia. To this end, VEGFR-Probe and MMP-Probe were developed by conjugating distinct near-infrared fluorophores to VEGFR-binding and MMP substrate peptides, respectively. VEGFR-Probe exhibited specific binding to VEGFR on HUVECs, and self-quenched MMP-Probe produced strong fluorescence intensity in the presence of MMPs in vitro. Subsequently, VEGFR-Probe and MMP-Probe were successfully utilized for time course in vivo visualization of VEGFR or MMP, respectively. Simultaneous visualization provided information regarding the spatial distribution of these proteins, including areas of co-localization. This dual-targeted in vivo imaging approach will be useful for understanding the detailed mechanism of angiogenesis and for evaluating therapeutic angiogenesis.
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http://dx.doi.org/10.1007/s00726-016-2225-0DOI Listing
July 2016

Therapeutic angiogenesis using tumor cell-conditioned medium.

Biotechnol Prog 2016 03 23;32(2):456-64. Epub 2016 Jan 23.

School of Chemical and Biological Engineering, Seoul National University, Seoul, 151-744, Republic of Korea.

Stem cell-conditioned medium (CM), which contains angiogenic factors that are secreted by stem cells, represents a potential therapy for ischemic diseases. Along with stem cells, tumor cells also secrete various angiogenic factors. Here, tumor cells as a cell source of CM for therapeutic angiogenesis was evaluated and the therapeutic efficacy of tumor cell CM in mouse hindlimb ischemia models was demonstrated. CM obtained from a human fibrosarcoma HT1080 cell line culture was compared with CM obtained from a human bone marrow-derived mesenchymal stem cell (MSC) culture. HT1080 CM contained higher concentrations of angiogenic factors compared with MSC CM, which was attributable to the higher cell density that resulted from a much faster growth rate of HT1080 cells compared with MSCs. For use in in vitro and in vivo angiogenesis studies, HT1080 CM was diluted such that HT1080 CM and MSC CM would have the same cell number basis. The two types of CMs induced the same extent of human umbilical vein endothelial cell (HUVEC) proliferation in vitro. The injection of HT1080 CM into mouse ischemic limbs significantly improved capillary density and blood perfusion compared with the injection of fresh medium. Although the therapeutic outcome of HT1080 CM was similar to that of MSC CM, the preparation of CM by tumor cell line culture would be much more efficient due to the faster growth and unlimited life-time of the tumor cell line. These data suggest the potential application of tumor cell CM as a therapeutic modality for angiogenesis and ischemic diseases. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:456-464, 2016.
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http://dx.doi.org/10.1002/btpr.2226DOI Listing
March 2016

Mesenchymal Stem Cells Aggregate and Deliver Gold Nanoparticles to Tumors for Photothermal Therapy.

ACS Nano 2015 Oct 11;9(10):9678-90. Epub 2015 Sep 11.

School of Chemical Engineering, Sungkyunkwan University , Suwon 440-746, Republic of Korea.

Gold nanoparticles (AuNPs) have been extensively studied for photothermal cancer therapy because AuNPs can generate heat upon near-infrared irradiation. However, improving their tumor-targeting efficiency and optimizing the nanoparticle size for maximizing the photothermal effect remain challenging. We demonstrate that mesenchymal stem cells (MSCs) can aggregate pH-sensitive gold nanoparticles (PSAuNPs) in mildly acidic endosomes, target tumors, and be used for photothermal therapy. These aggregated structures had a higher cellular retention in comparison to pH-insensitive, control AuNPs (cAuNPs), which is important for the cell-based delivery process. PSAuNP-laden MSCs (MSC-PSAuNPs) injected intravenously to tumor-bearing mice show a 37-fold higher tumor-targeting efficiency (5.6% of the injected dose) and 8.3 °C higher heat generation compared to injections of cAuNPs after irradiation, which results in a significantly enhanced anticancer effect.
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http://dx.doi.org/10.1021/acsnano.5b02207DOI Listing
October 2015

pH-triggered release of manganese from MnAu nanoparticles that enables cellular neuronal differentiation without cellular toxicity.

Biomaterials 2015 Jul 9;55:33-43. Epub 2015 Apr 9.

School of Chemical and Biological Engineering, Seoul National University, Seoul 151-744, Republic of Korea; Interdisciplinary Program for Bioengineering, Seoul National University, Seoul 151-744, Republic of Korea; Bio-MAX Institute, Institute of Chemical Processes, Engineering Research Institute, Seoul National University, Seoul 151-744, Republic of Korea. Electronic address:

At high concentrations, manganese (Mn) promotes cellular neurodevelopment but causes toxicity. Here, we report that Mn ion at high concentrations can be delivered to pheochromocytoma 12 (PC12) cells using gold nanoparticles (AuNPs) to enhance cellular neurodevelopment without toxicity. Mn(2+) release from AuNPs was designed to be pH-responsive so that low pH condition of the cell endosomes can trigger in situ release of Mn(2+) from AuNPs after cellular uptake of Mn-incorporated AuNPs (MnAuNPs). Due to the differences in reduction potentials of Mn and Au, only Mn ionized and released while Au remained intact when MnAuNPs were uptaken by cells. Compared to PC12 cells treated with a high concentration of free Mn(2+), PC12 cells treated with an equal concentration of MnAuNPs resulted in significantly enhanced cellular neurodevelopment with decreased apoptosis and necrosis. Treatment with a high concentration of free Mn(2+) led to an abrupt consumption of a large amount of ATP for the intracellular transport of Mn(2+) through the ion channel of the cell membrane and to mitochondrial damage caused by the high intracellular concentration of Mn(2+), both of which resulted in cell necrosis and apoptosis. In contrast, MnAuNP-treated cells consumed much smaller amount of ATP for the intracellular transport of MnAuNPs by endocytosis and showed pH-triggered in situ release of Mn(2+) from the MnAuNPs in the endosomes of the cells, both of which prevented the cell death caused by ATP depletion and mitochondrial damage. To our knowledge, this is the first report on the use of AuNPs as a vehicle for pH-responsive, intracellular delivery of metal ion, which may open a new window for drug delivery and clinical therapy.
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http://dx.doi.org/10.1016/j.biomaterials.2015.03.025DOI Listing
July 2015

A dual delivery of substance P and bone morphogenetic protein-2 for mesenchymal stem cell recruitment and bone regeneration.

Tissue Eng Part A 2015 Apr 23;21(7-8):1275-87. Epub 2015 Jan 23.

1 School of Chemical and Biological Engineering, Seoul National University , Seoul, Republic of Korea.

Implantation of ex vivo expanded and osteogenically differentiated mesenchymal stem cells (MSCs) for bone regeneration has drawbacks for clinical applications, such as poor survival of implanted cells and increased treatment expenses. As a new approach for bone regeneration that can circumvent these limitations, we propose dual delivery of substance P (SP) and bone morphogenetic protein-2 (BMP-2) to facilitate endogenous stem cell recruitment to bone defects by SP and subsequent in situ osteogenic differentiation of those cells by BMP-2. A heparin-conjugated fibrin (HCF) gel enabled dual delivery with fast release of SP and slow release of BMP-2, which would be ideal for prompt recruitment of endogenous stem cells in the first stage and time-consuming osteogenic differentiation of the recruited stem cells in the second stage. The HCF gels with SP and/or BMP-2 were implanted into mouse calvarial defects for 8 weeks. Local delivery of SP to the calvarial defects using HCF gel was more effective in recruiting MSCs to the calvarial defects than intraperitoneal or intravenous administration of SP. Many of the cells recruited by SP underwent osteogenic differentiation through local delivery of BMP-2. The efficacy of in vivo bone regeneration was significantly higher in the SP/BMP-2 dual delivery group. The dual delivery of SP and BMP-2 using the HCF gel therefore has potential as an effective bone regeneration strategy.
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http://dx.doi.org/10.1089/ten.TEA.2014.0182DOI Listing
April 2015

Conditioned medium of adipose-derived stromal cell culture in three-dimensional bioreactors for enhanced wound healing.

J Surg Res 2015 Mar 6;194(1):8-17. Epub 2014 Nov 6.

School of Chemical and Biological Engineering, Seoul National University, Seoul, Republic of Korea; Institute of Bioengineering, Institute of Chemical Processess, Seoul National University, Seoul, Republic of Korea. Electronic address:

Background: It was previously shown that human adipose-derived stromal cell (hADSC)-conditioned medium (CM) promotes wound healing. An essential part of the wound healing process is neovascularization in the wound bed.

Materials And Methods: We hypothesized that CM prepared from hADSCs cultured as spheroids in three-dimensional suspension bioreactors (spheroid CM) would contain much higher concentrations of angiogenic growth factors secreted by hADSCs, induce a higher extent of neovascularization in the wound bed, and improve wound healing as compared with CM prepared by conventional monolayer culture (monolayer CM).

Results: The concentrations of angiogenic growth factors (i.e., vascular endothelial growth factor, basic fibroblast growth factor, and hepatocyte growth factor) in spheroid CM were 20- to 145-fold higher than those in monolayer CM. Either fresh medium, monolayer CM, or spheroid CM was administered to full-thickness wounds created on the dorsal aspects of athymic mice. The monolayer CM promoted wound healing as compared with fresh medium or no treatment. Importantly, wound closure was faster, and dermal and epidermal regeneration was improved in the spheroid CM-treated mice compared with that in the monolayer CM-treated mice.

Conclusions: The improved wound healing by spheroid CM may be attributed, at least in part, to enhanced neovascularization in the wound beds. The spheroid-based CM approach showed potential as a therapy for skin wound repair.
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http://dx.doi.org/10.1016/j.jss.2014.10.053DOI Listing
March 2015

Efficacious and clinically relevant conditioned medium of human adipose-derived stem cells for therapeutic angiogenesis.

Mol Ther 2014 Apr 13;22(4):862-72. Epub 2013 Jan 13.

1] School of Chemical and Biological Engineering, Seoul National University, Seoul, Korea [2] Interdisciplinary Program for Bioengineering, Seoul National University, Seoul, Korea [3] Institute of Chemical Processes, Seoul National University, Seoul, Korea.

Using stem cell-conditioned medium (CM) might be a viable alternative to stem cell transplantation, which is often hampered by low grafting efficiency and potential tumorigenesis, but the concentrations of angiogenic growth factors in CM are too low for therapeutic use and some components of the medium are not for human use. We used three-dimensional (3D) spheroid culture of human adipose-derived stem cells (ADSCs) with clinically relevant medium composed of amino acids, vitamins, glucose, and human serum to produce clinically relevant CM containing angiogenic and/or antiapoptotic factors such as vascular endothelial cell growth factor, fibroblast growth factor 2, hepatocyte growth factor, and chemokine (C-X-C motif) ligand 12. The concentrations of these factors were 23- to 27-fold higher than that in CM produced by conventional monolayer culture. Compared with injection of either monolayer culture CM or human ADSC, injection of spheroid culture CM to an ischemic region in mice significantly enhanced endothelial cell growth, CD34(+)/PTPRC(-) (endothelial progenitor) cell mobilization from bone marrow, and bone marrow cell homing to the ischemic region, resulting in improved blood vessel density, limb salvage, and blood perfusion in a mouse hindlimb ischemia model. The stem cell CM developed in this study will likely be an effective alternative to conventional stem cell transplantation therapy.
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http://dx.doi.org/10.1038/mt.2013.301DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3982496PMC
April 2014

Therapeutic angiogenesis by a myoblast layer harvested by tissue transfer printing from cell-adhesive, thermosensitive hydrogels.

Biomaterials 2013 Nov 6;34(33):8258-68. Epub 2013 Aug 6.

Department of Bioengineering, College of Engineering, Hanyang University, 17 Haengdang-dong, Seongdong-gu, Seoul 133-791, Republic of Korea.

Peripheral arterial disease (PAD) is characterized by the altered structure and function of arteries caused by accumulated plaque. There have been many studies on treating this disease by the direct injection of various types of therapeutic cells, however, the low cell engraftment efficiency and diffusion of the transplanted cells have been major problems. In this study, we developed an approach (transfer printing) to deliver monolayer of cells to the hindlimb ischemic tissue using thermosensitive hydrogels, and investigated its efficacy in long term retention upon transplantation and therapeutic angiogenesis. We first investigated the in vitro maintenance of robust cell-cell contacts and stable expression of the ECM proteins in myoblast layer following transfer printing process. In order to confirm the therapeutic effect of the myoblasts in vivo, we cultured a monolayer of C2C12 myoblasts on thermosensitive hydrogels, which was then transferred to the hindlimb ischemia tissue of athymic mice directly from the hydrogel by conformal contact. The transferred myoblast layer was retained for a longer period of time than an intramuscularly injected cell suspension. In addition, the morphology of the mice and laser Doppler perfusion (28 days after treatment) supported that the myoblast layer enhanced the therapeutic effects on the ischemic tissue. In summary, the transplantation of the C2C12 myoblast layer using a tissue transfer printing method could represent a new approach for the treatment of PAD by therapeutic angiogenesis.
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http://dx.doi.org/10.1016/j.biomaterials.2013.07.071DOI Listing
November 2013

Modulation of BMP-2-induced chondrogenic versus osteogenic differentiation of human mesenchymal stem cells by cell-specific extracellular matrices.

Tissue Eng Part A 2013 Jan 16;19(1-2):49-58. Epub 2012 Nov 16.

School of Chemical and Biological Engineering, Seoul National University, Seoul, Republic of Korea.

Bone morphogenetic protein-2 (BMP-2) is known to induce both osteogenic and chondrogenic commitment of human mesenchymal stem cells (hMSCs). However, factors influencing BMP-2-dependent chondrogenic and osteogenic differentiation have not been investigated. In this study, we demonstrated that extracellular microenvironments, in the form of cell-derived matrices, play important roles in determining the specific lineage commitment of hMSCs in the presence of BMP-2. Extracellular matrices (ECMs) derived from osteoblasts and chondrocytes were utilized to regulate cell differentiation. Osteogenic and chondrogenic differentiation of hMSCs cultured on the two different cell-derived ECMs were assessed by quantitative real-time-polymerase chain reaction, immunocytochemistry, and western blot analysis. To minimize the effects of the cell-adhesion proteins contained in serum on the ECMs, hMSCs were cultured in serum-free osteogenic or chondrogenic differentiation medium. Fibronectin-, collagen type I-, or collagen type II-coated substrates were utilized as ECM controls. The ECM specific to each cell type promoted lineage-specific commitment of hMSCs in the presence of BMP-2, that is, osteoblast- and chondrocyte-derived ECM promoted osteogenic and chondrogenic commitment, respectively. Therefore, cell-specific ECMs are capable of modulating the BMP-2-induced osteogenic and chondrogenic differentiation of hMSCs.
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http://dx.doi.org/10.1089/ten.TEA.2012.0245DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3530938PMC
January 2013

Efficient formation of cell spheroids using polymer nanofibers.

Biotechnol Lett 2012 May 30;34(5):795-803. Epub 2011 Dec 30.

School of Chemical and Biological Engineering, Seoul National University, 56-1, Silim-dong, Gwanak-Gu, Seoul, 151-744, Republic of Korea.

Spheroid culture has been used for suspension cultures of anchorage-dependent cells. In this study, we developed a new method for the suspension cultures of anchorage-dependent animal cells using polymer nanofibers. Poly(lactic-co-glycolic acid) nanofibers (785 nm in average fiber-diameter, 88 μm in average fiber-length) fabricated by the electrospinning method were added to each suspension culture of human embryonic kidney 293 cells and human dermal fibroblasts. As compared to no addition of nanofibers to the suspension cultures, nanofibers enhanced cell spheroid formation, thereby reducing cell death resulting from a lack of cell adhesion. Efficient formation of spheroids in the presence of polymer nanofibers may be useful for the suspension cultures of anchorage-dependent cells.
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http://dx.doi.org/10.1007/s10529-011-0836-9DOI Listing
May 2012

3,4-dihydroxyphenylalanine-assisted hydroxyapatite nanoparticle coating on polymer scaffolds for efficient osteoconduction.

Tissue Eng Part C Methods 2012 Apr 13;18(4):245-51. Epub 2011 Dec 13.

School of Chemical and Biological Engineering, Seoul National University, Seoul, Republic of Korea.

For bone regeneration applications, scaffolds made from a composite of a biodegradable polymer and ceramic have advantages over scaffolds made from only one component (biodegradable polymer or ceramic alone). In this study, a simple and rapid method was developed to induce hydroxyapatite (HA) nanoparticle adsorption on polyglycolic acid (PGA) scaffold surfaces. PGA meshes were coated with HA nanoparticles by immersing the scaffolds in a buffer solution containing 3,4-dihydroxyphenylalanine (DOPA), a critical, functional element in mussel adhesive protein known to strongly bind to various materials. Substantial HA coating on PGA scaffolds was achieved within 24 hours of immersion, as determined according to selective staining of ceramic particles, scanning electron microscopy, X-ray photoelectron spectroscopy, and energy-dispersive spectroscopy. To evaluate the osteoconduction efficacy of the scaffolds in vivo, PGA scaffolds, DOPA-coated PGA scaffolds, PGA scaffolds immersed in HA solution, and HA- and DOPA-coated PGA (HA-DOPA-PGA) scaffolds were implanted in critical-sized defects in mouse skulls for 8 weeks. Micro-computed tomography and histological analyses showed that bone regeneration in vivo was far more extensive on HA-DOPA-PGA scaffolds than on the other scaffolds. DOPA offers an efficient and simple method of HA coating on polymer scaffolds. HA-polymer composite scaffolds fabricated using this method could be useful as bone graft.
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http://dx.doi.org/10.1089/ten.TEC.2011.0373DOI Listing
April 2012

Modulation of stem cell differentiation with biomaterials.

Int J Stem Cells 2010 May;3(2):80-4

Interdisciplinary Program of Bioengineering, Seoul National University, Seoul, Korea ; School of Chemical and Biological Engineering, Seoul National University, Seoul, Korea.

Differentiation of stem cells can be controlled with interactions with microenvironments of the stem cells. The interactions contain various signals including soluble growth factor signal, cell adhesion signal, and mechanical signal, which can modulate differentiation of stem cells. Biomaterials can provide these types of signals to induce desirable cellular differentiation. Biomaterials can deliver soluble growth factors locally to stem cells at a controlled rate for a long period. Stem cell adhesion to specific adhesion molecules presented by biomaterials can induce specific differentiation. Mechanical signals can be delivered to stem cells seeded onto biomaterial scaffolds. These approaches would be invaluable for direction of stem cell differentiation and in vivo tissue regeneration using stem cells.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4021801PMC
http://dx.doi.org/10.15283/ijsc.2010.3.2.80DOI Listing
May 2010
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