Jeong Ah Kim

Enhancement of neurite outgrowth in PC12 cells by iron oxide nanoparticles

Despite the many potential therapeutic applications of iron oxide nanoparticle such as its use as an imaging and targeting tool, its biological effects have not yet been extensively characterized. Herein, we report that iron oxide nanoparticles taken up by PC12 cells can enhance neurite outgrowth. PC12 cells exposed to both iron oxide nanoparticles and nerve growth factor (NGF) synergistically increased the efficiency of neurite outgrowth in a dose-dependent manner. This may have resulted from the activation of cell adhesion molecules that are associated with cell-matrix interactions through iron. Immunoblotting assays also revealed that both neural specific marker protein and cell adhesion protein expression were upregulated by iron oxide nanoparticles compared with non-treated cells via activation of the mitogen-activated protein kinase (MAPK) signaling pathway. Our findings point to the possibility that iron oxide nanoparticles can affect cell-substrate interactions and regulate cell behaviors, which provides clinical insights into potential neurologic and therapeutic applications of iron oxide nanoparticles.

Droplet-based microfluidic system to form and separate multicellular spheroids using magnetic nanoparticles

The importance of creating a three-dimensional (3-D) multicellular spheroid has recently been gaining attention due to the limitations of monolayer cell culture to precisely mimic in vivo structure and cellular interactions. Due to this emerging interest, researchers have utilized new tools, such as microfluidic devices, that allow high-throughput and precise size control to produce multicellular spheroids. We have developed a droplet-based microfluidic system that can encapsulate both cells and magnetic nanoparticles within alginate beads to mimic the function of a multicellular tumor spheroid. Cells were entrapped within the alginate beads along with magnetic nanoparticles, and the beads of a relatively uniform size (diameters of 85% of the beads were 170-190 μm) were formed in the oil phase. These beads were passed through parallel streamlines of oil and culture medium, where the beads were magnetically transferred into the medium phase from the oil phase using an external magnetic force. This microfluidic chip eliminates additional steps for collecting the spheroids from the oil phase and transferring them to culture medium. Ultimately, the overall spheroid formation process can be achieved on a single microchip.

Regulation of morphogenesis and neural differentiation of human mesenchymal stem cells using carbon nanotube sheets

In order to successfully utilize stem cells for therapeutic applications in regenerative medicine, efficient differentiation into a specific cell lineage and guidance of axons in a desired direction is crucial. Here, we used aligned multi-walled carbon nanotube (MWCNT) sheets to differentiate human mesenchymal stem cells (hMSCs) into neural cells. Human MSCs present a preferential adhesion to aligned CNT sheets with longitudinal stretch parallel to the CNT orientation direction. Cell elongation was 2-fold higher than the control and most of the cells were aligned on CNT sheets within 5° from the CNT orientation direction. Furthermore, a significant, synergistic enhancement of neural differentiation was observed in hMSCs cultured on the CNT sheets. Axon outgrowth was also controlled using nanoscale patterning of CNTs. This CNT sheet provides a new cellular scaffold platform that can regulate morphogenesis and differentiation of stem cells, which could open up a new approach for tissue and stem cell regeneration.

Simultaneous and multiplexed detection of exosome microRNAs using molecular beacons

Simultaneous and multiplexed detection of microRNAs (miRNAs) in a whole exosome is developed, which can be utilized as a PCR-free efficient diagnosis method for various diseases. Exosomes are small extracellular vesicles that contain biomarker miRNAs from parental cells. Because they circulate throughout bodily fluids, exosomal biomarkers offer great advantages for diagnosis in many aspects. In general, PCR-based methods can be used for exosomal miRNA detection but they are laborious, expensive, and time-consuming, which make them unsuitable for high-throughput diagnosis of diseases. Previously, we reported that single miRNA in the exosomes can be detected specifically using an oligonucleotide probe or molecular beacon. Herein, we demonstrate for the first time that multiple miRNAs can be detected simultaneously in exosomes using miRNA-targeting molecular beacons. Exosomes from a breast cancer cell line, MCF-7, were used for the production of exosomes because MCF-7 has a high level of miR-21, miR-375, and miR-27a as target miRNAs. Molecular beacons successfully hybridized with multiple miRNAs in the cancer cell-derived exosomes even in the presence of high human serum concentration. In addition, it is noteworthy that the choice of fluorophores for multiplexing biomarkers in an exosome is crucial because of its small size. The proposed method described in this article is beneficial to high-throughput analysis for disease diagnosis, prognosis, and response to treatment because it is a time-, labor-, and cost-saving technique.

Detection of exosome miRNAs using molecular beacons for diagnosing prostate cancer

Abstract Prostate cancer is the fifth leading cause of cancer-related deaths among males worldwide. However, the biomarker for diagnosing prostate cancer that is used currently has limitations that must be overcome. Recently, several studies have demonstrated that the cancer liquid biopsy can be implemented by using exosome miRNAs. However, the current methods for the detection of exosome miRNAs are time-consuming, expensive, and laborious. Thus, we investigated a novel method for diagnosing prostate cancer that involves the use of molecular beacons for the in situ detection of miRNAs in exosomes from prostate cancer cells. We chose miRNA-375 and miRNA-574-3p as the target miRNAs for prostate cancer, and these markers in exosomes produced by prostate cancer cells including DU145 and PC-3 were successfully detected using molecular beacons. High fluorescent signals were obtained from MB and miRNA hybridization in exosomes in a concentration-dependent manner. In addition, exosome miRNAs can be detected even in the presence of human urine, so this method can be applied directly using human urine to perform liquid biopsies for prostate cancer. Overall, the in situ detection of exosome miRNAs using molecular beacons can be developed as a simple, cost effective, and non-invasive liquid biopsy for diagnosing prostate cancer.