Do-Sim Park

Vital roles of mTOR complex 2 in Notch-driven thymocyte differentiation and leukemia

Rictor is essential in Notch-driven T-ALL pathogenesis.

An efficient method for the rapid establishment of Epstein‐Barr virus immortalization of human B lymphocytes

Abstract. Several methods have been developed for the immortalization of B lymphocytes by Epstein‐Barr virus (EBV). We developed an efficient method which reduces the time from culture initiation to immortalization and cryopreservation. Two infections of EBV to lymphocytes, and the use of phorbol ester‐induced EBV stock significantly improved immortalization efficiency and reduced the time between initiation and immortalization and cryopreservation. The resulting cell bank was used to produce DNA for genetic studies focusing on the genes involved in immune and autistic disorders.

Distribution of hepatitis B virus genotypes in Korea

BACKGROUND/AIMS Considering the incidence of prevailing hepatitis B virus (HBV) genotypes in neighboring nations, the predominance of genotype C in Korea is exceptional and needs to be confirmed by nationwide investigation. METHODS A total of 510 HBsAg (+) or HBeAg (+) serum samples was collected from subjects in several cities and harbors throughout the Korean peninsula for genotype (A-G)-specific multiplex PCR analysis. Another 40 serum samples from chronic HBV carriers from Iksan city were selected for sequencing of the entire HBV genome. Phylogenetic analysis was performed with 22 whole genomic sequences of Korean HBV strains enrolled in GenBank. RESULTS An amplicon was found in 377 specimens and genotype C occupied 98.1% (370 cases); none of the other genotypes were found. A mixed pattern of genotypes B and C was seen in seven specimens (1.9%), of which five were tested using PCR targeting the X fragment; no genotype B bands were found. With the exception of 1 case, which was subgenotype A2, whole sequences of Korean HBV strains (n=62) belonged to subgenotype C2. CONCLUSIONS The prevailing HBV genotype in Korea is C2; the other genotypes occur only rarely. Future studies should include confirmation of the detection of genotypes other than C.

Catalposide, a compound isolated from Catalpa Ovata, attenuates induction of intestinal epithelial proinflammatory gene expression and reduces the severity of trinitrobenzene sulfonic acid-induced colitis in mice

Certain irinoid-producing plants have been used as herbal anti-inflammatory remedies. Here we evaluated whether catalposide (CATP), a single compound isolated from irinoid-producing plant Catalpa ovata, has a potential for preventing or ameliorating diseases characterized by mucosal inflammation. Preliminary microarray-based gene expression test revealed that CATP, which alone did not significantly affect expression of any of the >8,000 genes analyzed, attenuated the expression of tumor necrosis factor-alpha (TNF-alpha)-induced proinflammatory genes including interleukin-8 (IL-8) in human intestinal epithelial HT-29 cells. Down-regulation of IL-8 mRNA accumulation was also reflected by the decreased IL-8 secretion in CATP-treated HT-29 cells. The signal transduction study revealed that CATP significantly attenuates TNF-alpha-mediated p38 and extracellular signal-regulated kinase (ERK) phosphorylation. Further, CATP reduced NF-kappaB-mediated transcriptional activation as well as Ikappa-Balpha degradation. To establish the in vivo relevance of these findings, we examined whether CATP could affect intestinal inflammation in vivo using the mouse model of trinitrobenzene sulfonic acid (TNBS)-induced inflammatory colitis. Intrarectal administration of CATP dramatically reduced the weight loss, colonic damage, and mucosal ulceration that characterize TNBS colitis. Moreover, CATP suppressed the expression of TNF-alpha, interleukin-1beta, and intercellular adhesion molecule-1 along with the inhibition of NF-kappa B p65 translocation into nucleus in TNBS colitis. Collectively, current results demonstrate that CATP may be an effective agent for the treatment of diseases characterized by mucosal inflammation.

Relationship between the Echocardiographic Epicardial Adipose Tissue Thickness and Serum Adiponectin in Patients with Angina.

BACKGROUND It is still unknown whether increased cardiac adiposity is related to the risk factors of coronary artery disease (CAD). We measured epicaridal adopose tissue (EAT) and mediastinal adipose tissue (MAT) using echocardiography and examined their correlations with CAD and serum adiponectin. METHODS One hundred fifty three patients who underwent elective coronary angiography for chest pain were measured cardiac adiposity by transthoracic echocardiography. The correlations of cardiac adipose tissue with the presence and severity of CAD and the serum adiponectin level were examined. RESULTS EAT was thicker in patients with CAD (1.8+/-1.4 vs. 3.8+/-1.9 mm, p<0.001), but MAT was not different according to the presence of CAD (2.9+/-2.8 vs. 3.5+/-2.5 mm, p=0.121). EAT showed a significant positive correlation with age (r=0.225, p=0.005), homocystein (r=0.289, p=0.001), fasting glucose (r=0.167, p=0.042), and fibrinogen (r=0.218, p=0.009), and a significant negative correlation with serum adiponectin (r=-0.194, p=0.016). EAT thickness (OR 11.53, 95% CI; 3.61-36.84, p<0.001) and low serum adiponectin (OR 2.88, 95% CI; 1.02-8.15, p=0.046) were independent predictors of obstructive CAD. However, MAT thickness was not associated with CAD. CONCLUSION EAT was associated with the severity and risk factors of CAD and correlated with serum adiponectin level. In contrast with EAT, MAT was not associated with CAD and adiponectin.

Sequence Motifs in IL-4Rα Mediating Cell-Cycle Progression of Primary Lymphocytes

IL-4 signaling through the IL-4Rα chain regulates the development and proliferation of the Th2 lineage of effector CD4+ T cells. Analyses of the IL-4R in factor-dependent cell lines led to the development of two apparently conflicting models of the primary structural determinants of IL-4R-mediated proliferative signaling. In one model, proliferation was dependent on the first conserved tyrosine in the cytoplasmic tail (Y1), while in the second, proliferation was independent of cytoplasmic tyrosines. We found that in activated primary T cells, mutation of only the Y1 residue resulted in a modest decrease in IL-4-induced S phase entry, a further decrease in cell-cycle completion, and a complete failure of IL-4 to induce p70S6 kinase phosphorylation. Consistent with a role for the PI3K/mammalian target of rapamycin pathway in mediating cytokine acceleration of G2/M transit, pretreatment of activated T cells with rapamycin resulted in only a modest decrease in IL-4-induced S phase entry, but a total block of cell-cycle completion. Strikingly, IL-4Rα chains that lacked all cytoplasmic tyrosines were competent to signal for STAT5 phosphorylation, mediated efficient S phase entry, and promoted cell-cycle progression. The ability of tyrosine-deficient IL-4Rs to mediate proliferative signaling and STAT phosphorylation was absolutely dependent on the presence of an intact ID-1 region. These findings show that IL-4Rα lacking cytoplasmic tyrosine residues is competent to induce ID-1-dependent proliferation, and indicate that IL-4 can promote G2/M progression via activation of the mammalian target of rapamycin pathway initiated at the Y1 residue.

TAGLN2 regulates T cell activation by stabilizing the actin cytoskeleton at the immunological synapse.

TAGLN2 stabilizes cortical F-actin and thereby maintains F-actin contents at the immunological synapse, which allows T cell activation following T cell receptor stimulation.

Hepatitis B Virus X Protein Regulates Hepatic Glucose Homeostasis via Activation of Inducible Nitric Oxide Synthase

Dysregulation of liver functions leads to insulin resistance causing type 2 diabetes mellitus and is often found in chronic liver diseases. However, the mechanisms of hepatic dysfunction leading to hepatic metabolic disorder are still poorly understood in chronic liver diseases. The current work investigated the role of hepatitis B virus X protein (HBx) in regulating glucose metabolism. We studied HBx-overexpressing (HBxTg) mice and HBxTg mice lacking inducible nitric oxide synthase (iNOS). Here we show that gene expressions of the key gluconeogenic enzymes were significantly increased in HepG2 cells expressing HBx (HepG2-HBx) and in non-tumor liver tissues of hepatitis B virus patients with high levels of HBx expression. In the liver of HBxTg mice, the expressions of gluconeogenic genes were also elevated, leading to hyperglycemia by increasing hepatic glucose production. However, this effect was insufficient to cause systemic insulin resistance. Importantly, the actions of HBx on hepatic glucose metabolism are thought to be mediated via iNOS signaling, as evidenced by the fact that deficiency of iNOS restored HBx-induced hyperglycemia by suppressing the gene expression of gluconeogenic enzymes. Treatment of HepG2-HBx cells with nitric oxide (NO) caused a significant increase in the expression of gluconeogenic genes, but JNK1 inhibition was completely normalized. Furthermore, hyperactivation of JNK1 in the liver of HBxTg mice was also suppressed in the absence of iNOS, indicating the critical role for JNK in the mutual regulation of HBx- and iNOS-mediated glucose metabolism. These findings establish a novel mechanism of HBx-driven hepatic metabolic disorder that is modulated by iNOS-mediated activation of JNK.

Effect of simvastatin on the resistance to EGFR tyrosine kinase inhibitors in a non-small cell lung cancer with the T790M mutation of EGFR.

Although non-small cell lung cancer (NSCLC) tumors with activating mutations in the epidermal growth factor receptor (EGFR) are highly responsive to EGFR tyrosine kinase inhibitors (TKIs) including gefitinib and erlotinib, development of acquired resistance is almost inevitable. Statins show antitumor activity, but it is unknown whether they can reverse EGFR-TKIs resistance in NSCLC with the T790M mutation of EGFR. This study investigated overcoming resistance to EGFR-TKI using simvastatin. We demonstrated that addition of simvastatin to gefitinib enhanced caspase-dependent apoptosis in T790M mutant NSCLC cells. Simvastatin also strongly inhibited AKT activation, leading to suppression of β-catenin activity and the expression of its targets, survivin and cyclin D1. Both insulin treatment and AKT overexpression markedly increased p-β-catenin and survivin levels, even in the presence of gefitinib and simvastatin. However, inhibition of AKT by siRNA or LY294002 treatment decreased p-β-catenin and survivin levels. To determine the role of survivin in simvastatin-induced apoptosis of gefitinib-resistant NSCLC, we showed that the proportion of apoptotic cells following treatment with survivin siRNA and the gefitinib-simvastatin combination was greater than the theoretical additive effects, whereas survivin up-regulation could confer protection against gefitinib and simvastatin-induced apoptosis. Similar results were obtained in erlotinib and simvastatin-treated HCC827/ER cells. These findings suggest that survivin is a key molecule that renders T790M mutant NSCLC cells resistant to apoptosis induced by EGFR-TKIs and simvastatin. Overall, these data indicate that simvastatin may overcome EGFR-TKI resistance in T790M mutant NSCLCs via an AKT/β-catenin signaling-dependent down-regulation of survivin and apoptosis induction.

Levels of Soluble Receptor for Advanced Glycation End Products in Acute Ischemic Stroke without a Source of Cardioembolism

Background and Purpose Low levels of soluble receptor for advanced glycation end products (sRAGE) are associated with three conventional vascular risk factors (3Fs: diabetes, hypertension, and hypercholesterolemia), nondiabetic coronary artery disease, and Alzheimers disease. However, the association between sRAGE and acute ischemic stroke (AS), especially AS without a source of cardioembolism, has not yet been established. Methods Patients with AS without a source of cardioembolism (n=259) and age-matched controls (n=300) were grouped according to the presence of 3Fs: AS patients with and without 3Fs (3Fs+ AS and 3Fs- AS, respectively) and controls with and without 3Fs (3Fs+ control and 3Fs- control, respectively). Levels of sRAGE were analyzed among the four groups. Results sRAGE was significantly higher in the controls than in the AS patients (855 pg/mL vs. 690 pg/mL, p<0.01). sRAGE was significantly higher in 3Fs- controls (996 pg/mL, p<0.05) than in 3Fs+ controls (721 pg/mL), and in AS group regardless of the 3Fs (629 pg/mL in 3Fs- and 705 pg/mL in 3Fs+). The lowest tertile of sRAGE was associated with an increased risk of AS in the 3Fs- group [adjusted odds ratio (OR) 4.0, 95% confidence interval (CI) 1.6-10.3, p<0.01] but not in the 3Fs+ group. The level of sRAGE was also correlated with neurological severity in the 3Fs- AS group (r=-0.32, p<0.05) but not in the 3Fs+ AS group. Conclusions Low plasma levels of sRAGE is a potential biomarker for the risk of AS and may reflect the neurological severity of the condition, especially in subjects without identifiable conventional risk factors.