Jungwook Kim

Entrapped doxorubicin nanoparticles for the treatment of metastatic anoikis-resistant cancer cells

Metastasized and chemoresistant secondary breast cancer treatment commonly shows very low efficacy. A new efficient treatment method is required to overcome the limitation against the secondary breast cancer. In this study, anoikis-resistant breast cancer cells, MDA-MB-231 and MCF-7 were developed as models of chemoresistant and metastatic breast cancer. Doxorubicin encapsulating human serum albumin nanoparticles (HSA+DOX NPs) were fabricated to confirm the benefits of nanoparticles at the treatment of anoikis-resistant breast cancer cells. The side population (SP) fraction in the anoikis-resistant cancer cells was higher than the parental cells. HSA+DOX NPs were more cytotoxic to anoikis-resistant cancer cells than free doxorubicin. The confocal microscope images demonstrated HSA+DOX NPs to deliver more doxorubicin into cells compared to the free doxorubicin by bypassing the drug efflux pump systems of anoikis-resistant cancer cells. In this study, a nanomedicine-based drug delivery carrier shows a potential in treating a metastasized and chemoresistant breast cancer.

The regulation of 2,3-butanediol synthesis in Klebsiella pneumoniae as revealed by gene over-expressions and metabolic flux analysis

A variety of microorganism species are able naturally to produce 2,3-butanediol (2,3-BDO), although only a few of them are suitable for consideration as having potential for mass production purposes. Klebsiella pneumoniae (K. pneumoniae) is one such strain which has been widely studied and used industrially to produce 2,3-BDO. In the central carbon metabolism of K. pneumoniae, the 2,3-BDO synthesis pathway is dominated by three essential enzymes, namely acetolactate decarboxylase, acetolactate synthase, and butanediol dehydrogenase, which are encoded by the budA, budB, and budC genes, respectively. The mechanisms of the three enzymes have been characterized with regard to their function and roles in 2,3-BDO synthesis and cell growth (Blomqvist et al. in J Bacteriol 175(5):1392–1404, 1993), while a few studies have focused on the cooperative mechanisms of the three enzymes and their mutual interactions. Therefore, the K. pneumoniae KCTC2242::ΔwabG wild-type strain was utilized to reconstruct seven new mutants by single, double, and triple overexpression of the three enzymes key to this study. Subsequently, continuous cultures were performed to obtain steady-state metabolism in the organisms and experimental data were analyzed by metabolic flux analysis (MFA) to determine the regulation mechanisms. The MFA results showed that the seven overexpressed mutants all exhibited enhanced 2,3-BDO production, and the strain overexpressing the budBA gene produced the highest yield. While the enzyme encoded by the budA gene produced branched-chain amino acids which were favorable for cell growth, the budB gene enzyme rapidly enhanced the conversion of acetolactate to acetoin in an oxygen-dependent manner, and the budC gene enzyme catalyzed the reversible conversion of acetoin to 2,3-BDO and regulated the intracellular NAD+/NADH balance.

Observation of 2,3-butanediol biosynthesis in Lys regulator mutated Klebsiella pneumoniae at gene transcription level

Microorganisms that produce 2,3-butanediol (2,3-BDO) are mostly mixed acid fermentation microorganisms, and they synthesize 2,3-BDO in order to suppress medium acidification. The 2,3-BDO operon (budBAC) is activated by the LysR regulator protein derived from the budR. In this study, the effect of the budR on 2,3-BDO-biosynthesis was observed at gene transcription level. The Klebsiella pneumoniae strains (wabG-deleted strain (SGSB100), budR over-expressed strain (SGSB101), and the budR-deleted strain (SGSB102)) were constructed. The resulting strains were cultivated in unified conditions. Samples were obtained at the lag-, log-, and stationary-phase of cell growth, and gene transcription levels of the budR, 2,3-BDO-biosynthesis-related (budB, budA, and budC), and acid-biosynthesis-related (ldhA and ack) genes were observed. During the lag-phase of cell growth in SGSB101, the budR transcription level increased approximately 8-fold, and 2,3-BDO production increased approximately 2-fold, when compared to SGSB100. Also in SGSB101 the transcription level of the acid-biosynthesis-related genes (ldhA and ack) increased approximately up to 11-fold during the lag-phase of cell growth compared to SGSB100. On contrast, in SGSB102 budR transcription was not detected, and the transcription level of the acid-biosynthesis-related genes (ldhA and ack) decreased approximately 70-fold during the lag-phase of cell growth compared to SGSB100. This is by far the first observation of 2,3-BDO regulation mechanism at gene transcription level in K. pneumoniae, and therefore may be useful for understanding and improving 2,3-BDO biosynthesis metabolic network.

Synthetic Hydrogels with Stiffness Gradients for Durotaxis Study and Tissue Engineering Scaffolds

Migration of cells along the right direction is of paramount importance in a number of in vivo circumstances such as immune response, embryonic developments, morphogenesis, and healing of wounds and scars. While it has been known for a while that spatial gradients in chemical cues guide the direction of cell migration, the significance of the gradient in mechanical cues, such as stiffness of extracellular matrices (ECMs), in directed migration of cells has only recently emerged. With advances in synthetic chemistry, micro-fabrication techniques, and methods to characterize mechanical properties at a length scale even smaller than a single cell, synthetic ECMs with spatially controlled stiffness have been created with variations in design parameters. Since then, the synthetic ECMs have served as platforms to study the migratory behaviors of cells in the presence of the stiffness gradient of ECM and also as scaffolds for the regeneration of tissues. In this review, we highlight recent studies in cell migration directed by the stiffness gradient, called durotaxis, and discuss the mechanisms of durotaxis. We also summarize general methods and design principles to create synthetic ECMs with the stiffness gradients and, finally, conclude by discussing current limitations and future directions of synthetic ECMs for the study of durotaxis and the scaffold for tissue engineering.

The Mechanical Aspects of Formation and Application of PDMS Bilayers Rolled into a Cylindrical Structure

A polydimethylsiloxane (PDMS) film with its surface being oxidized by a plasma treatment or a UV-ozone (UVO) treatment, that is, a bilayer made of PDMS and its oxidized surface layer, is known to roll into a cylindrical structure upon exposure to the chloroform vapor due to the mismatch in the swelling ratio between PDMS and the oxidized layer by the chloroform vapor. Here we analyzed the formation of the rolled bilayer with the mechanical aspects: how the mismatch in the swelling ratio of the bilayer induces rolling of the bilayer, why any form of trigger that breaks the symmetry in the in-plane stress level is needed to roll the bilayer uniaxially, why the rolled bilayer does not unroll in the dry state when there is no more mismatch in the swelling ratio, and how the measured curvature of rolled bilayer matches well with the prediction by the theory. Moreover, for the use of the rolled bilayer as the channel of the microfluidic device, we examined whether the rolled bilayer deforms or unrolls by the flow of the aqueous solution that exerts the circumferential stress on the rolled bilayer.