Seon Jeong Kim

Suppression of transient receptor potential melastatin 7 channel induces cell death in gastric cancer

Ca2+ and Mg2+ have a fundamental role in many cellular processes and ion channels are involved in normal physiologic processes and in the pathology of various diseases. The aim here was to show that the presence and potential role of transient receptor potential melastatin 7 (TRPM7) channels in the growth and survival of AGS cells, the most common human gastric adenocarcinoma cell line. The patch‐clamp technique for whole‐cell recording was used in AGS cells. TRPM7‐specific small interfering RNAs were used for specific inhibition of TRPM7. Whole‐cell voltage‐clamp recordings revealed the TRPM7‐like currents that activated spontaneously following loss of intracellular Mg2+. The current had a non‐linear current–voltage relationship with the characteristic steep outward rectification associated with TRPM7 channels. Reverse transcription–polymerase chain reaction, western blotting, and immunoreactivity all showed abundant expression of TRPM7 messenger RNA and protein in AGS cells. Transfection of AGS cells with TRPM7 siRNA significantly reduced the expression of TRPM7 mRNA and protein as well as the amplitude of the TRPM7‐like currents. Furthermore, we found that Mg2+ is critical for the growth and survival in AGS cells. Blockade of TRPM7 channels by La3+ and 2‐APB or suppression of TRPM7 expression by siRNA inhibited the growth and survival of these cells. Human gastric adenocarcinoma cells express TRPM7 channel whose presence is essential for cell survival. The protein is a likely potential target for the pharmacological treatment of gastric cancer. (Cancer Sci 2008; 99: 2502–2509)

pH-Dependent Structures of an i-Motif DNA in Solution

We have investigated for the first time the structure of i-motif DNA in solution at various pH conditions by using synchrotron small-angle X-ray scattering technique. To facilitate direct structural comparison between solution structures of i-motif DNA at various pH values, we created atomic coordinates of i-motif DNA from a fully folded to unfolded atomic model. Under mild acidic conditions, the conformations for i-motif DNA appeared to be similar to that of the partially unfolded i-motif atomic model in overall shape, rather than the fully folded i-motif atomic model. Collectively, our observations indicate that i-motif DNA molecule is structurally dynamic over a wide pH range, adopting multiple conformations ranging from the folded i-motif structure to a random coil conformation. As the i-motif structure has been used as an important component in nanomachines, we can therefore believe that the structural evidence presented herein will promote the development of future DNA-based molecular-actuator devices.

Hydrogel-Assisted Polyaniline Microfiber as Controllable Electrochemical Actuatable Supercapacitor

Flexible, controllable, and stable electrochemical supercapacitors serving as actuators at low operating voltage combining the advantages of the high power of the dielectric capacitors and the high specific energy of rechargeable batteries are important in artificial muscle technology, hybrid electric vehicles, and in short-term power sources for mobile electronic devices [Baughman, Science, 300, 268 (2003); Winter and Brodd, Chem. Rev. (Washington, D.C.), 104, 4245 (2004); Ebron, et al., Science, 311, 1580 (2006)]. High capacitance, a surprising 99% inner charge contribution and actuation in the hydrogel-assisted actuatable electrochemical supercapacitor (HAES) microfiber fabricated through wet spinning of a chitosan solution, followed by the in situ chemical polymerization of aniline was made possible through the perfect utilization of the large surface area provided by the nanostructured polyaniline grown inside as well as on the surface of the fiber. The HAES electrodes with an actuation strain of 0.33% showed 703 F/g specific capacitance in 1 M methane sulfonic acid, and more than 3000 cycles durability. The change in impedance as well as capacitance was achieved by the controlled strain as a function of applied stress, which can establish a direct relationship between the actuation strain and specific capacitance of electrochemical supercapacitors.

Controlled magnetic nanofiber hydrogels by clustering ferritin.

We have fabricated biocompatible nanofiber hydrogels with diverse sizes of ferritin clusters according to the mixing temperature of solutions employing electrospinning. Poly(vinyl alcohol) (PVA) was used as a polymeric matrix for fabricating nanocomposites. By thermal means we controlled the interaction between the host PVA hydrogel and the protein shell on ferritin bionanoparticles to vary the size and concentration of ferritin clusters. The clustering of ferritin was based on the partial unfolding of a protein shell of ferritin. By studying the magnetic properties of the PVA/ferritin nanofibers according to the mixing temperature of the PVA/ferritin solutions, we confirmed that the clustering process of the ferritin was related to changes in the superparamagnetic properties and magnetic resonance imaging (MRI) contrast of the PVA/ferritin nanofibers. PVA/ferritin nanofiber hydrogels with diverse spatial distributions of ferritin nanoparticles are applicable as MRI-based noninvasive detectable cell culture scaffolds and as artificial muscles because of their improved superparamagnetic properties.

DNA Hydrogel Fiber with Self‐Entanglement Prepared by Using an Ionic Liquid

DNA hydrogels have a wide range of biomedical applications in tissue engineering and drug-delivery systems. There are two ways to create hydrogel structures: one is enzymecatalyzed assembly of synthetic DNA and the other is by crosslinking natural DNA chemically. For natural DNA, formaldehyde and metal compounds such as arsenic, chromate, and nickel are widely used as crosslinkers. However, these modified DNA hydrogels are unsafe to apply in biological systems because the crosslinkers have potentially adverse side effects, with some being carcinogens. Besides this, these DNA hydrogels are difficult to form into hydrogel fibers by using conventional spinning methods in the absence of chemical crosslinking. In solution, DNA resembles modular proteins such as titin, silk, and polysaccharides. The very flexible linear DNA strands and their noncovalent assemblies can form compacted interwound supercoils in bulk aqueous solution with cationic salts. Alternatively, they can roll into soluble clusters of toroids. In concentrated DNA solutions in poor solvents, rodlike multiple-chain bundling occurs and, simultaneously, single or multiple loops form knots with themselves or with adjacent loops through a nucleation-growth pathway. Thus, these condensates are seen primarily as intertwined aggregates of toroids. We were inspired by the spinning processes used by insects (for example, silkworms and spiders) to develop spinning conditions to create the desired DNA hydrogel fibers. It is known that the last process to occur in insect spinning is the formation of a dragline in air. It can be considered that the air effectively removes water through evaporation to produce dense, dried fibers. To replicate this process in wet spinning, we need to ensure that the coagulation solvent does not fill the space created in the spinning droplet by the exiting water. In addition, the diffusion rates of the coagulation solvent and water must be controlled to prevent the formation of a dense skin on the fiber, which could trap water and create a porous structure. If the coagulation solvent also contains crosslinking cations, then the concentrated DNA solution can form hydrogel fibers with intertwined toroidal entanglements. We have found that room-temperature hydrophilic ionic liquids (RTILs) can produce suitable conditions. The feasibility of using RTILs was suggested by our previous work, which showed that 100% of RTILs will absorb water even when bound to a polymer network. Moreover, some RTILs can create low-pH conditions when in contact with water, and such acidic conditions have been used to promote coagulation in the wet spinning of DNA fibers. The cations present in RTILs condense the DNA as a matter of course. In this work, we prepared a DNA hydrogel fiber in a single step by injecting aqueous DNA solution into a coagulation bath of an RTIL.

Tough Supersoft Sponge Fibers with Tunable Stiffness from a DNA Self-Assembly Technique

Tough and soft: Highly porous, spongelike materials self-assemble by calcium ion condensation of DNA-wrapped carbon nanotubes (SWNTs-DNA; see picture, IL = ionic liquid). The toughness, modulus, and swellability of the electrically conductive sponges can be tuned by controlling the density and strength of interfiber junctions. The sponges have compliances similar to the softest natural tissue, while robust interfiber junctions give high toughness.

Role of calmodulin and myosin light chain kinase in the activation of carbachol-activated cationic current in murine ileal myocytes.

We investigated the effect of calmodulin (CaM) and myosin light chain kinase (MLCK) on murine ileal myocytes using the whole-cell patch-clamp technique. Under the voltage clamp, at the holding potential of -60 mV, 50 micromol/L carbachol (CCh) induced inward currents (I CCh), and spontaneous decay of I CCh occurred. The peak inward currents induced by the repetitive application of CCh (50 micromol/L) tended to decrease in amplitude. Intracellular application of 0.2 mmol/L guanosine 5-O-(gamma-thio)triphosphate (GTP gammaS) from the patch electrode induced an inward current at a holding potential of -60m V, and the peak inward currents induced by the repetitive application of Cs tended to decrease slightly in amplitude. The amplitude of I CCh was reduced by pretreatment either with W-7, trifluoroperazine, W-5, and melittin (CaM inhibitors) or with ML-7 and ML-9 (selective MLCK inhibitors), and the inhibitory effects were reversible. However, when we pretreated with 50 micromol/L W-7 or 5 micromol/L ML-7 on GTP gammaS-induced inward currents, almost no inhibition was observed in the inward currents. Application of both Rho kinase inhibitor and MLCK inhibitor inhibited GTP gammaS-induced currents. We conclude that CaM and MLCK modulate the activation process of I CCh in murine ileal myocytes and suggest that the classical type transient receptor potential (TRPC) channel 5 might be a candidate for nonselective cationic currents (NSCC) activated by muscarinic stimulation in gastrointestinal smooth muscle cells.

Switchable redox activity by proton fuelled DNA nano-machines

The switching electrochemical property of an SWNT/DNA hybrid can be produced through reversible conformational changes between the closed and open state originating from the pH-responding i-motif DNA which significantly improves its molecular switching and stability by hydrophobic interactions with SWNTs.

Functional characteristics of TRPC4 channels expressed in HEK 293 cells

The classical type of transient receptor potential (TRPC) channel is a molecular candidate for Ca2+-permeable cation channels in mammalian cells. Because TRPC4 and TRPC5 belong to the same subfamily of TRPC, they have been assumed to have the same physiological properties. However, we found that TRPC4 had its own functional characteristics different from those of TRPC5. TRPC4 channels had no constitutive activity and were activated by muscarinic stimulation only when a muscarinic receptor was co-expressed with TRPC4 in human embryonic kidney (HEK) cells. Endogenous muscarinic receptor appeared not to interact with TRPC4. TPRC4 activation by GTPγS was not desensitized. TPRC4 activation by GTPγS was not inhibited by either Rho kinase inhibitor or MLCK inhibitor. TRPC4 was sensitive to external pH with pKa of 7.3. Finally, TPRC4 activation by GTPγS was inhibited by the calmodulin inhibitor W-7. We conclude that TRPC4 and TRPC5 have different properties and their own physiological roles.

Electrical properties of polyaniline and multi-walled carbon nanotube hybrid fibers.

We have fabricated for the first time one-dimensional multiwalled carbon nanotube (MWNT) nanocomposite fibers with improved electrical properties using electrospinning. Polyaniline (PANi) and poly(ethylene oxide) (PEO) were used as a conducting and a nonconducting matrix, respectively, for hybrid nanofibers including MWNTs. The hybrid nanofibers fabricated by electrospinning had a length of several centimeters and a diameter ranging from approximately 100 nm to approximately 1 microm. Transmission electron microscopic analysis confirmed that the MWNTs were successfully oriented along the fiber axis without any severe aggregation during electrospinning. The hybrid nanofibers showed an enhanced electrical conductance with increasing MWNT content up to 0.5 wt%, and compared to PANi/PEO fibers, they also showed a stable linear ohmic behavior. These hybrid conducting nanofibers can be applied to chemical and biosensors that require a high sensitivity.