Sung Sik Choi

Dominant Role of Peroxiredoxin/JNK Axis in Stemness Regulation during Neurogenesis from Embryonic Stem Cells

Redox balance has been suggested as an important determinant of “stemness” in embryonic stem cells (ESCs). In this study, we demonstrate that peroxiredoxin (Prx) plays a pivotal role in maintenance of ESC stemness during neurogenesis through suppression of reactive oxygen species (ROS)‐sensitive signaling. During neurogenesis, Prx I and Oct4 are expressed in a mutually dependent manner and their expression is abruptly downregulated by an excess of ROS. Thus, in Prx I−/− or Prx II−/− ESCs, rapid loss of stemness can occur due to spontaneous ROS overload, leading to their active commitment into neurons; however, stemness is restored by the addition of an antioxidant or an inhibitor of c‐Jun N‐terminal kinase (JNK). In addition, Prx I and Prx II appear to have a tight association with the mechanism underlying the protection of ESC stemness in developing teratomas. These results suggest that Prx functions as a protector of ESC stemness by opposing ROS/JNK cascades during neurogenesis. Therefore, our findings have important implications for understanding of maintenance of ESC stemness through involvement of antioxidant enzymes and may lead to development of an alternative stem cell‐based therapeutic strategy for production of high‐quality neurons in large quantity. Stem Cells 2014;32:998–1011

Efficacy of Pulsed Electromagnetic Therapy for Chronic Lower Back Pain: a Randomized, Double-blind, Placebo-controlled Study

This randomized, double-blind, placebo-controlled clinical trial studied the effectiveness of pulsed electromagnetic therapy (PEMT) in patients with chronic lower back pain. Active PEMT (n = 17) or placebo treatment (n = 19) was performed three times a week for 3 weeks. Patients were assessed using a numerical rating scale (NRS) and revised Oswestry disability scores for 4 weeks after therapy. PEMT produced significant pain reduction throughout the observation period compared with baseline values. The percentage change in the NRS score from baseline was significantly greater in the PEMT group than the placebo group at all three time-points measured. The mean revised Oswestry disability percentage after 4 weeks was significantly improved from the baseline value in the PEMT group, whereas there were no significant differences in the placebo group. In conclusion, PEMT reduced pain and disability and appears to be a potentially useful therapeutic tool for the conservative management of chronic lower back pain.

Measurement of Single-Cell Deformability Using Impedance Analysis on Microfluidic Chip

In this paper, we propose a microfluidic chip that measures the deformability of single cells by an impedance measurement method. The proposed chip is designed to differentiate the deformability of various cells by measuring the length of their stretched membrane indirectly according to the variation of the impedance after applying aspiration pressure to the cell membrane. The length of the stretched cell membrane is proportional to the applied pressure. Lengths of 18 and 21 µm were observed at the same suction pressure for human breast normal cells (MCF-10A) and caner cells (MCF-7), respectively. Electrical measurement was performed using an impedance analyzer at various frequencies. Results revealed that the impedance measurement method can be used to analyze the biomechanical characteristics of single cells, which indicates the state of malignancy of cells.

Association of Matrix metalloproteinase-3 with cardiogenic activity during Noggin-induced differentiation of mouse embryonic stem cells

BACKGROUND Despite the pluripotency of embryonic stem (ES) cells, their clinical applications have been hindered due to the lack of reliable differentiation methods. Recently, it was shown that Noggin could effectively induce cardiomyocyte differentiation by transient treatment of ES cells. METHODS To determine how Noggin may induce cardiac differentiation, we compared differentially expressed genes during Noggin-induced differentiation of ES cells using microarray analysis. We found Matrix metalloproteinase-3 (Mmp-3) expression was highly up-regulated by Noggin treatment. To understand the role of Mmp-3 in the cardiac differentiation of ES cells, we inhibited Mmp-3 activity by treating with a specific Mmp-3 inhibitor during Noggin-induced cardiac differentiation of ES cells. We also analyzed the expression levels of cardiac markers and the ratio of spontaneously beating embryoid bodies (EBs) in the presence of the Mmp-3 inhibitor. RESULTS We analyzed EB samples from zero, two, and four days with or without Noggin treatment, and found that the expression levels of 2 (0 day), 56 (2 days), and 805 (4 days) genes were altered with Noggin treatment. Up-regulation of Mmp-3 was closely associated with relative increases of cardiogenic, vasculogenic, and hematopoietic genes in EB treated with Noggin. By inhibiting Mmp-3 activity, we verified that Mmp-3 activation is partly responsible for both the expression of cardiac markers and the elevated ratio of spontaneously beating to non-beating EBs. CONCLUSIONS The concurrent expression of Mmp-3 with many cardiogenic genes and the specific inhibition of Mmp-3 revealed a critical role for Mmp-3 in Noggin-induced cardiac differentiation of ES cells.

Separation of Human Breast Cancer and Epithelial Cells by Adhesion Difference in a Microfluidic Channel

A simple, label-free microfluidic cell purification method is presented for separation of cancer cells by exploiting difference in cell adhesion. To maximize the adhesion difference, three types of polymeric nanostructures (50nm pillars, 50nm perpendicular and 50nm parallel lines with respect to the direction of flow) were fabricated using UVassisted capillary moulding and included inside a polydimethylsiloxane (PDMS) microfluidic channel bonded onto glass substrate. The adhesion force of human breast epithelial cells (MCF10A) and human breast carcinoma (MCF7) was measured independently by injecting each cell line into the microfluidic device followed by culture for a period of time (e.g., one, two, and three hours). Then, the cells bound to the floor of a microfluidic channel were detached by increasing the flow rate of medium in a stepwise fashion. It was found that the adhesion force of MCF10A was always higher than that of MCF cells regardless of culture time and surface nanotopography at all flow rates, resulting in a label-free detection and separation of cancer cells. For the cell types used in our study, the optimum separation was found for 2 hours culture on 50nm parallel line pattern followed by flow-induced detachment at a flow rate of 300 ㎕/min.

Blueberry protects LPS-stimulated BV-2 microglia through inhibiting activities of p38 MAPK and ERK1/2

The effects of blueberry extract (BE) were investigated on activities of p38 mitogen-activated protein kinase (MAPK) and extracellular extracellular signal regulated kinase 1/2 (ERK1/2) in mouse BV-2 microglia stimulated lipopolysaccharide (LPS). BV-2 microglia were cultured for 24 h in the presence of 1 μg/mL LPS either with or without BE preincubation for 0, 1, and 12 h. BE relieved repression of cell proliferation, and reduced cell death. These alterations caused by BE addition accompanied reduction of radical oxygen species (ROS). There were also significant decreases in the levels of both transcripts, and proteins of inducible nitric oxide synthase (iNOS) and peroxiredoxin 1 (Prx1) genes in the BE-treated cells, suggesting that BE inhibit the ROS-induced gene expression. The antioxidant effects of BE appeared to involve p38 MAPK and ERK1/2 signaling pathways since BE reduced both kinase activities by inhibiting phosphorylation. Taken these results together, BE protects LPS-stimulated BV-2 microglia by reducing cell death, ROS accumulation, and activities of p38 MAPK and ERK1/2.

Dual Reporter Genes Enabling Cell Tracing with Viable and Reliable Selection of Various Cell Types

Dual reporter genes driven by either a ubiquitous cytomegalovirus (CMV) or a neuro-specific tubulin α1 promoter (Tα1) were constructed. The new genes, CMV (pCMV-GL) or Tα1 promoter-driven GFP–LacZ (pTα1-GL), robustly expressed the fused GFP–LacZ protein reporting constitutive expressions in various cell types including CHO cells, loach and chicken embryos, and neuro-specific expression in differentiating mouse embryonic stem cells, respectively. The dual reporter genes thus provide a versatile tool for the studies of gene expression, cell lineage within the embryo and possibly the fate of stem cells in transplantation experiment, thus facilitating different analyses depending on the experimental purposes.

A microfluidic flow sensor for measuring cell adhesion

We present a simple, biomarker-free microfluidic device for separating cancer cells from a mixed solution of normal and cancer cells by difference in adhesion force. A polydimethylsiloxane (PDMS) microfluidic chip was fabricated onto glass substrate using standard soft lithography. Three types of polyurethane acrylate (PUA) nanostructure (50 nm pillar, 50 nm perpendicular groove, 50 nm horizontal groove with respect to the direction of flow) were included inside the microfluidic channel by UV-assisted capillary molding. For cell types, MCF7 (breast cancer cell line) and MCF10A (breast normal cell line) were used. To find the optimum condition for separation, each cell line was injected into the microfluidic device and cultured for 1 h, 2 h, and 3 h, respectively, followed by cell detachment by flow of medium solution with increasing flow rate. The adhesion force of MCF10A was stronger than that of MCF7. MCF10A cells cultured onto the nanopatterned surface were more spread than those cultured onto the glass surface. Furthermore, the presence of nanopatterns increased the ratio of adhesion force of normal and cancer cells and thus and the separation efficiency. The optimum culture condition was 2 h onto the nanopattern and flow rate was ~ 300 mul/min.

Erratum: Dual reporter genes enabling cell tracing with viable and reliable selection of various cell types (Biotechnology Letters (2006) 28, (287-293) DOI: 10.1007/s10529-005-5715-9)

Dual reporter genes driven by either a ubiquitous cytomegalovirus (CMV) or a neuro-specific tubulin alpha1 promoter (Talpha1) were constructed. The new genes, CMV (pCMV-GL) or Talpha1 promoter-driven GFP-LacZ (pTalpha1-GL), robustly expressed the fused GFP-LacZ protein reporting constitutive expressions in various cell types including CHO cells, loach and chicken embryos, and neuro-specific expression in differentiating mouse embryonic stem cells, respectively. The dual reporter genes thus provide a versatile tool for the studies of gene expression, cell lineage within the embryo and possibly the fate of stem cells in transplantation experiment, thus facilitating different analyses depending on the experimental purposes.