Bumrae Cho

Generation and Characterization of Human Heme Oxygenase-1 Transgenic Pigs

Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1), an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs). Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

A Dynamic Model of a PEM Fuel Cell System

The steady states and dynamics of a fuel cell system are influenced by both operating conditions and the frequency characteristics of electrochemical reactions in a cell. A dynamic model of a practical fuel cell system, based on an analytical theory and empirical data, is proposed. The model consists of three parts, such as an open circuit voltage model including fuel flow and reaction in channels, an equivalent circuit model caused by voltage losses of electrochemical reactions in a cell, and an air compressor model (a fuel supply model among auxiliary systems in a fuel cell system). It will be useful to design the power system and control strategy and to analyze the system dynamics in a fuel cell power system. The model is verified by experiments of 1kW PEM FC system. The dynamics of its OCV(open circuit voltage) by fuel flow effect, and the frequency characteristics of reaction dynamics in a cell are verified by experiments.

Immunosuppressive mechanisms of embryonic stem cells and mesenchymal stem cells in alloimmune response.

Although both embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs) are known to have immunosuppressive effects, the mechanisms of immunosuppression are still controversial. Both types of stem cells suppressed not only the proliferation but also survival of CD4(+) T cells in vitro. They suppressed secretion of various cytokines (IL-2, IL-12, IFN-γ, TNF-α, IL-4, IL-5, IL-1β, and IL-10), whereas there was no change in the levels of TGF-β or IDO. Classic and modified transwell experiments demonstrated that immunosuppressive activities were mainly mediated by cell-to-cell contact. Granzyme B in the ESCs played a significant role in their immunosuppression, whereas PDL-1, Fas ligand, CD30 or perforin was not involved in the contact-dependent immunosuppression. However, none of the above molecules played a significant role in the immunosuppression by the MSCs. Interestingly, both stem cells increased the proportion of Foxp3(+) regulatory T cells. Our results showed that both ESCs and MSCs suppressed the survival as well as the proliferation of T cells by mainly contact-dependent mechanisms and increased the proportion of regulatory T cells. Granzyme B was involved in immunosuppression by the ESCs in a perforin-independent manner.

Modeling and control of the single-phase photovoltaic grid-connected cascaded H-bridge multilevel inverter

This paper presents the modeling and control of the single-phase photovoltaic grid-connected five-level cascaded H-bridge multilevel inverter. For the unity power factor, the proportional and integral current controller with the duty ratio feed-forward compensation method is used. And in order to track the maximum power point and to reduce the partial shading due to stacked photovoltaic modules, each DC voltage is stably controlled to their maximum power points by dedicated voltage controllers of each H-bridge module. The modeling approach and the control loop design method are provided in this paper. The proposed control schemes are validated from experimental results of the 2-kW prototype hardware.

Generation of soluble human tumor necrosis factor-α receptor 1-Fc transgenic pig.

Background. Acute humoral xenograft rejection (AHXR) is an important barrier to xenograft survival. Human tumor necrosis factor-&agr; (hTNF-&agr;) is one of the essential mediators of AHXR and induces activation of porcine endothelial cells (PECs), resulting in upregulation of major histocompatibility complex molecules, adhesion molecules, and proinflammatory chemokines. We investigated whether introduction of a soluble human tumor necrosis factor receptor I-Fc (shTNFRI-Fc) fusion gene can suppress activation of PECs and, more importantly, produced shTNFRI-Fc transgenic pigs. Methods. The shTNFRI-Fc gene expression vector was constructed and inserted into PECs. The inhibitory effects of shTNFRI-Fc were tested by luciferase assay, reverse-transcriptase polymerase chain reaction, and flow cytometry. A shTNFRI-Fc transgenic pig was generated by somatic cell nuclear transfer. The expression of shTNFRI-Fc in the transgenic pig was evaluated by PCR, western blot, enzyme-linked immunosorbent assay, and immunohistochemistry. The inhibitory effects of shTNFRI-Fc in the serum obtained from the transgenic pig were also tested. Results. In comparison with control green fluorescent protein, shTNFRI-Fc protein showed much stronger inhibitory effects on NF-&kgr;B activation in the HEK293-NF-&kgr;B-luciferase reporting cell line, expression of chemokines and adhesion molecules in PECs, and TNF-&agr;-mediated cytotoxicity. We successfully generated shTNFRI-Fc transgenic pig. Sera obtained from the transgenic pig inhibited induction of chemokines, and E-selectin in PECs stimulated with Human TNF-&agr;. Conclusions. We have generated transgenic pigs producing shTNFRI-Fc protein that can inhibit TNF-&agr;-mediated activation of PECs. Because TNF-&agr; is an important mediator of xenograft rejection, the use of xenografts that can produce shTNFRI-Fc proteins de novo could be an effective approach in overcoming a considerable component of the xenograft rejection process, especially AHXR.

Digital load current feed-forward control method for a dc-dc converter

This paper describes a new digital feed-forward control method to enhance the dynamic performance of a dc-dc converter. The feed-forward duty ratio is calculated based on the charge balance of the output capacitor using the load current, and it is combined with the output of the digital PED feedback compensator. The feed-forward duty ratio propagates to the consecutive switching cycles during the load transient interval to immediately supply the required energy. The propagation length is decided by comparing the feed-forward duty ratio with the switching period. The performance improvement of the proposed digital control method is experimentally verified on a 200 W buck dc-dc converter.

Constant resistance control of solar array regulator using average current mode control

In this paper, a single current control loop to perform the MPPT and battery charging control as well as the current sharing control for the parallel connected SAR is presented. It transforms the effective load characteristic of the SAR seen by the SA from the constant power load into the constant resistance load. Thus, the system becomes large-signally stable for the entire operation region of the SA. The proposed method is simple and flexible since it is not affected by the characteristic of the SAR. Although it uses the nonlinear control to transform the effective load characteristic of SAR, it can be easily implemented using digital controller, which may be already available for the MPPT controller without additional circuits. For the design and the stability analysis of the proposed method, the small signal analysis is performed and the Z.O.H effect is analyzed. The experimental results from a 180W prototype hardware show that the proposed control scheme has a simple implementation structure and can stabilize the system in the entire region of SA.

Expression Analysis of Combinatorial Genes Using a Bi-Cistronic T2A Expression System in Porcine Fibroblasts

In pig-to-primate xenotransplantation, multiple transgenic pigs are required to overcome a series of transplant rejections. The generation of multiple transgenic pigs either by breeding or the introduction of several mono-cistronic vectors has been hampered by the differential expression patterns of the target genes. To achieve simultaneous expression of multiple genes, a poly-cistronic expression system using the 2A peptide derived from the Thosea asigna virus (T2A) can be considered an alternative choice. Before applying T2A expression system to pig generation, the expression patterns of multiple genes in this system should be precisely evaluated. In this study, we constructed several bi-cistronic T2A expression vectors, which combine target genes that are frequently used in the xenotransplantation field, and introduced them into porcine fibroblasts. The proteins targeted to the same or different subcellular regions were efficiently expressed without affecting the localization or expression levels of the other protein. However, when a gene with low expression efficiency was inserted into the upstream region of the T2A sequences, the expression level of the downstream gene was significantly decreased compared with the expression efficiency without the insertion. A small interfering RNA targeting one gene in this system resulted in the significant downregulation of both the target gene and the other gene, indicating that multiple genes combined into a T2A expression vector can be considered as a single gene in terms of transcription and translation. In summary, the efficient expression of a downstream gene can be achieved if the expression of the upstream gene is efficient.

A Cost Effective PDP Sustainer using Two-Winding Transformer with Hybrid Operation

In this paper, a new PDP sustainer using transformer energy recovery networks is proposed. The two-winding transformer gives the proposed topology several merits, such as a lower number of devices, simple gate driver, ZVS-on of the main switches, ZCS-off of the resonant switches and diodes and high energy recovery efficiency. The turns ratio realizes the voltage-applied overdriving technique as well as the current-injected overdriving technique. The hybrid driving technique using both methods is researched, which is useful for a faster transition time, hence providing higher contrast ratio and luminous efficiency. The proposed circuit is verified with simulation and experimental results of a 42-inch PDP.

Human thrombomodulin regulates complement activation as well as the coagulation cascade in xeno-immune response.

With the introduction of the α1, 3‐galactosyltransferase gene‐knockout (GT‐KO) pig and its pivotal role in preventing hyperacute rejection (HAR), coagulation remains a considerable obstacle yet to be overcome in order to provide long‐term xenograft survival. Thrombomodulin (TBM) plays a critical anticoagulant and anti‐inflammatory role in its part of the protein C pathway. Many studies have demonstrated the strong anticoagulant effects of TBM in xenotransplantation, but its complement regulatory effects have not been appropriately examined. Here, we investigate whether TBM can regulate complement activation as well as coagulation in response to xenogeneic stimuli.