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Biomedical subjects

Ki-Young Lee

Publications and source records attributed to Ki-Young Lee.

At least 19 recordsLinked to original sources
PubMed2005

PDK1 nucleates T cell receptor-induced signaling complex for NF-kappaB activation.

Activation of the transcription factor NF-kappaB after engagement of the T cell receptor (TCR) is important for T cell proliferation and activation during the adaptive immune response. Recent reports have elucidated a signaling pathway that involves the protein kinase C (PKC), the scaffold protein CARD11 (also called CARMA-1), the caspase recruitment domain (CARD)-containing protein Bcl10, and the paracaspase (protease related to caspases) MALT1 as critical intermediates linking the TCR to the IkappaB kinase (IKK) complex. However, the events proximal to the TCR that initiate the activation of this signaling pathway remain poorly defined. We demonstrate that 3-phosphoinositide-dependent kinase 1 (PDK1) has an essential role in this pathway by regulating the activation of PKC and through signal-dependent recruiting of both PKC and CARD11 to lipid rafts. PDK1-associated PKC recruits the IKK complex, whereas PDK1-associated CARD11 recruits the Bcl10-MALT1 complex, thereby allowing activation of the IKK complex through Bcl10-MALT1-dependent ubiquitination of the IKK complex subunit known as NEMO (NF-kappaB essential modifier). Hence, PDK1 plays a critical role by nucleating the TCR-induced NF-kappaB activation pathway in T cells.

3-Phosphoinositide-Dependent Protein Kinases↗
PubMed2005

Novel inhibitors targeted to methionine aminopeptidase 2 (MetAP2) strongly inhibit the growth of cancers in xenografted nude model.

Inhibition of angiogenesis is emerging as a promising strategy for the treatment of cancer. In our study reported here, the effects of 4 highly potent methionine aminopeptidase 2 (MetAP2) inhibitors, IDR-803, IDR-804, IDR-805 and CKD-732 (designed by structure-based molecular modeling), on angiogenesis and tumor growth were assessed. Concentrations of these inhibitors as low as 2.5 nM were able to inhibit the growth of human umbilical vein endothelial cells (HUVEC) by as much as 50%, arresting growth in the G1 stage of mitosis. An intracellular accumulation of p21(WAF1/Cip1) protein was also observed. Furthermore, at higher concentrations (25 nM) of these 4 MetAP2 inhibitors, a significant induction of apoptosis was apparent in the same HUVEC cultures. As a result of these findings, the possible anticancer effects of these inhibitors were examined, utilizing the SNU-398 hepatoma cell line. Interestingly, pretreatment with these inhibitors led to an increased number of apoptotic cells of up to 60% or more, compared to untreated controls. Moreover, utilizing an in vivo xenografted murine model, these inhibitors suppressed the growth of engrafted tumor. In conclusion, these 4 inhibitory compounds potently exert an antiangiogenic effect to inhibit the growth of cancers in vivo and could potentially be useful for the treatment of a variety of cancers.

Aminopeptidases↗
PubMed2004

GTP-dependent secretion from neutrophils is regulated by Cdk5.

We have previously shown evidence for the existence of a calcium-independent, GTP-regulated mechanism of secretion from neutrophils, but this secretory mechanism remains to be fully elucidated. Cyclin-dependent kinase 5 (Cdk5), the various substrates of which include Munc18 and synapsin 1, has been implicated in neuronal secretion. Although the Cdk5 activator, p35, and Cdk5-p35 activity are primarily associated with neurons, we report here that p35 also exists in neutrophils and that an active Cdk5-p35 complex is present in these cells. Cdk5-p35 activity in human neutrophils is mostly localized in secretory granules, which show an increase in Cdk5-p35 level and activity upon GTP stimulation. The potent Cdk5 inhibitor, roscovitine, completely blocks GTP-stimulated granule Cdk5 activity, which accompanies lactoferrin secretion from neutrophil-specific granules. Roscovitine also inhibits GTP-induced lactoferrin secretion and surface localization of the secretion markers, CD63 and CD66b, to a certain extent. Furthermore, neutrophils from wild-type mice treated with roscovitine and neutrophils from p35(-/-) mice exhibit comparable surface expression levels of both CD63 and CD66b upon GTP stimulation. Although our data suggest that other molecules control GTP-induced secretion from neutrophils, it is clear that Cdk5-p35 is required to elicit the maximum GTP-induced secretory response. Our observation that multiple proteins in neutrophil granules serve as specific substrates of Cdk5 further supports the premise that the kinase is a key component of the GTP-regulated secretory apparatus in neutrophils.

Animals↗
PubMed2004

OGT functions as a catalytic chaperone under heat stress response: a unique defense role of OGT in hyperthermia.

Protein O-GlcNAcylation is proceeded by O-linked GlcNAc transferase (OGT) in nucleocytoplasm and is involved in many biological processes although its physiological role is not clearly defined. To identify the functional significance of O-GlcNAcylation, we investigated heat stress effects on protein O-GlcNAcylation. Here, we found that protein O-GlcNAcylation was significantly increased in vivo during acute heat stress in mammalian cells and simultaneously, the enhanced protein O-GlcNAcylation was closely associated with cell survival in hyperthermia. Our results demonstrate that hyperthermal cytotoxicity may considerably be facilitated under the condition of insufficient level of protein O-GlcNAcylation inside cells. Furthermore, OGT reaction might be crucial for triggering thermotolerance to recover hyperthermal sensitivity without particular induction of heat shock proteins (hsps). Thus, we propose that OGT can respond rapidly to heat stress through the enhancement of nucleocytoplasmic protein O-GlcNAcylation for a rescue from the early phase of hyperthermal cytotoxicity.

Acylation↗
PubMed2004

L6 myoblast differentiation is modulated by Cdk5 via the PI3K-AKT-p70S6K signaling pathway.

Cdk5 regulates myogenesis but the signaling cascade through which Cdk5 modulates this process remains to be characterized. Here, we investigated whether PI3K, Akt, p70S6K, p38 MAPK, p44/42 MAPK, and Egr-1 serve as upstream regulators of Cdk5 during L6 myoblast differentiation. Upon serum reduction, we found that besides elevated expression of Cdk5 and its activator, p35, and increased Cdk5/p35 activity, Egr-1, Akt, p70S6K, and p38 MAPK activity were upregulated in differentiating L6 cells. However, p44/42 MAPK was downregulated and SAPK/JNK was unaffected. LY294002, a PI3K inhibitor, blocked the activation of Akt and p70S6K, indicating that Akt and p70S6K activation is linked to PI3K activation. The lack of LY294002 effect on p38 MAPK suggests that p38 MAPK activation is not associated with PI3K activation. Rapamycin, a specific inhibitor of FRAP/mTOR (the upstream kinase of p70S6K), also blocked p70S6K activation, indicating the involvement of FRAP/mTOR activation. LY294002 and rapamycin also blocked the enhancement of Egr-1 level, Cdk5 activity, and myogenin expression, suggesting that upregulation of these factors is coupled to PI3K-p70S6K activation. Overexpression of dominant-negative-Akt also reduced Cdk5/p35 activity and myogenin expression, indicating that the PI3K-p70S6K-Egr-1-Cdk5 signaling cascade is linked to Akt activation. SB2023580, a p38 MAPK inhibitor, had no effect on p70S6K, Egr-1, or Cdk5 activity, suggesting that p38 MAPK activation lies in a pathway distinct from the PI3K-Akt-p70S6K-Egr-1 pathway that we identify as the upstream modulator of Cdk5 activity during L6 myoblast differentiation.

Animals↗
PubMed2004

Effect of enflurane on the baroreflex control of heart rate in decerebrate rats.

Volatile anesthetics alter the arterial baroreflex (BRX) but its mechanisms are poorly understood. This study was designed to determine the effect of 1 and 2 minimal alveolar concentrations (MAC) of enflurane on the BRX parameters in unanesthetized brain stem-intact and decerebrate rats. Under enflurane anesthesia, the femoral artery and both femoral vein were catheterized for pressor (phenylephrine) and depressor (nitroprusside) drug delivery and continuous blood pressure measurements. Decerebration was performed at midcollicular level. BRX tests were performed in 3 time periods; before enflurane (conscious brain-intact), during 1 or 2 MAC enflurane exposure 1 hour after a sham operation or a decerebration operation, and 2 hours after the termination of enflurane (zero enflurane). Mean arterial pressure (MAP) and heart rate (HR) were fitted to a sigmoid logistic equation, the Boltzman equation. The curve of best fit was obtained with a computer program. 1 MAC and 2 MAC of enflurane shifted MAP-HR baroreflex curves to the left in the all groups and significantly attenuated the baroreflex range. The slope of conscious intact period and zero enflurane period of each group did not change significantly, but during the enflurane period the slope was significantly lowered. Enflurane depressed the baroreflex sensitivity (slope) and the HR range in a similar dose-dependent manner in both brain stem-intact and decerebrate rats. Such results draw into question whether the suprapontine sites contribute to enflurane's actions on cardiovascular autonomic regulation.

Anesthetics, Inhalation↗
PubMed2004

Bcl-2 and Bcl-xL are important for the induction of paclitaxel resistance in human hepatocellular carcinoma cells.

In this study we have investigated the mechanism underlying resistance to the chemotherapeutic drug paclitaxel in tumors of hepatocellular carcinoma (HCC) patients. Treatment with paclitaxel led to potent inhibition of growth of Hep3B hepatoma cells, but did not affect the growth properties of SNU-368 and SNU-398 cell lines that were established from primary HCC tumors. The growth inhibitory effect induced by paclitaxel correlated with levels of intracellular p21 and resulted in cell cycle arrest at the G2/M phase. However, paclitaxel treatment did not alter intracellular p53 levels. Instead, SNU-398 cells express high levels of the anti-apoptotic Bcl-2 and Bcl-x(L) proteins and the level of Bcl-x(L) could be further induced upon paclitaxel treatment. In contrast, Hep3B cells express pro-apoptotic members of the Bcl family and fail to induce Bcl-x(L) upon paclitaxel treatment. Therefore, these results strongly suggest that Bcl-2 and Bcl-x(L) play an important role in mediating resistance to paclitaxel.

Animals↗
PubMed2004

Cdk5/p35 expression in the mouse ovary.

Cyclin-dependent kinase 5 (Cdk5) is primarily associated with brain development but it is also implicated in lens and muscle differentiation. We found that Cdk5 is also expressed in mouse ovary, and explored the possibility that it plays a role in that tissue. We show by Western blotting and immunohistochemistry that the known Cdk5 activator, p35, is also present in the mouse ovary. Cdk5 and p35 were detected in oocytes at all stages of the follicle. While Cdk5 was present in the cytoplasm and nucleus of the oocyte, p35 was observed only in the cytoplasm. Both proteins were detected in the cytoplasm of luteinized cells in the corpus luteum. Immunoprecipitation and histone H1 kinase assays revealed that they form an ovarian complex with considerable kinase activity. Phosphorylation assays showed that several ovarian proteins are substrates for Cdk5/p35 in vitro. Together our findings suggest that p35-associated Cdk5 activity plays an important role in the ovary, where it may regulate cell differentiation and apoptosis as it does in the brain.

Animals↗
PubMed2003

Outer dense fibers serve as a functional target for Cdk5.p35 in the developing sperm tail.

Cdk5 is ubiquitously expressed in all tissues, but its activators, p35 and p39, are principally found in brain, and Cdk5 activity has mostly been associated with brain development, particularly neuronal differentiation and migration. Here we show that the p35 transcript and protein are also present in the testis, and an active Cdk5.p35 complex exists in this tissue as well. Cdk5 and p35 are prominently observed in elongating spermatid tails, particularly over the tail outer dense fibers (ODF). The appearance of Cdk5.p35 proceeds from the proximal to the distal end of elongating spermatids, coinciding with the proximal to distal assembly of ODF along the length of the tail axoneme. Incidentally, increased Cdk5.p35 activity is observed in isolated elongating spermatids and at a time when elongating spermatids appear in the developing testis, suggesting a role for Cdk5.p35 in spermiogenesis. The presence of Cdk5 and p35 in ODF isolated from rat sperm tails implies a strong association among these proteins. In vitro ODF phosphorylation by Cdk5.p35 and decreased in vivo sperm tail ODF phosphorylation in p35-deficient mice indicate that Cdk5.p35 is an integral component of ODF and that ODF is a functional Cdk5.p35 target in the testis. Our results demonstrate for the first time that Cdk5.p35 may participate in the regulation of sperm tail development via a mechanism involving ODF phosphorylation. Apparently, as in brain development, Cdk5.p35 plays a part in testis development.

Animals↗
PubMed2003

Cyclin E in breast tumors is cleaved into its low molecular weight forms by calpain.

Abundant levels of the hyperactive low molecular weight (LMW) forms of cyclin E contribute to deregulation of Cdk2 in breast tumors, but the mechanism through which they arise is not fully understood. Here, we explored the hypothesis that post-translational processing by a protease generates the LMW forms of cyclin E in breast tumors. In ZR75 tumor cell lysates, calcium-induced cyclin E truncation into peptides corresponding in size with LMW forms of cyclin E in tumor tissues. Calpeptin inhibited calcium-stimulated cyclin E truncation, indicating that cleavage resulted from activity of the calcium-dependent protease, calpain. Consistently, calcium+calpain caused truncation of cyclin E immunoprecipitated from tumor cells and tissues. Calcium also caused truncation of the calpain regulatory subunit in tumor cell lysates, indicating that elevated calpain activity accompanies cyclin E truncation. Increased levels of the calpain small subunit were also observed in breast tumors, and significant amounts of its proteolyzed forms indicated increased calpain activity. While elastase also caused cyclin E truncation, the cleavage pattern was distinct from that generated by calpain, suggesting discrete mechanisms in regulating the formation of LMW cyclin E in breast tumors. Treatment of ZR75 cultures with calcium+A23187 recapitulated the formation of the calcium/calpain-induced LMW forms of cyclin E. Altered calcium homeostasis and/or inability of the endogenous calpain inhibitor to control the activity of high levels of the calpain small subunit may contribute to increased calpain activity in breast tumors, causing abundant levels of LMW cyclin E.

Breast Neoplasms↗
PubMed2003

Tumor eradication by hepatitis B virus X antigen-specific CD8+ T cells in xenografted nude mice.

We have previously reported several CTL epitopes derived from the hepatitis B viral X Ag (HBx). In this study, we evaluated whether HBx-specific CTLs can be effectively used in adoptive cancer immunotherapy. To validate the possibility, four peptides containing a HLA-A2.1-restricted binding consensus motif were identified from the HBx protein and tested for their ability to activate CTL from PBMCs isolated from chronic carriers of HBV (n = 12). We selected two highly potent epitopes, HBx 52-60 (HLSLRGLFV) and HBx 115-123 (CLFKDWEEL), that are capable of inducing Ag-specific cytotoxic T cells in patient PBMCs. For adoptive immunotherapy using HBx-specific CTLs, we generated CTL clones restricted to the HBx 52-60 or HBx 115-123 peptide using a limiting dilution technique. LC-46, an HBx 52-60-specific clone, is CD62L(-)CD69(+)CD45RO(+)CD45RA(-)CD25(dim) and is stained by IFN-gamma (approximately 92%), IL-2 (30%), and TNF-alpha (56%), but not by IL-5, IL-10, IL-12, or TNF-beta, indicating that the cells are fully activated T cytotoxic 1-type cells. When LC-46 cells were adoptively transferred into xenografted nude mice bearing human hepatomas expressing HLA-A2.1 molecules and intracellular HBx proteins, the tumors were eradicated. Taken together, our data provide solid evidence for the feasibility of adoptive immunotherapy with HBx-sensitized CTLs in hepatitis disease, including hepatocellular carcinoma (HCC).

Animals↗
PubMed2003

Postchallenge hyperglycemia but not hyperinsulinemia is associated with angiographically documented coronary atherosclerosis in Korean subjects.

Although hyperinsulinemia has attracted considerable attention as a possible risk factor for coronary artery disease (CAD), previous studies have not shown consistent results. Hyperglycemia could be an alternative explanation for the association between type 2 diabetes and atherosclerosis. Since previous studies have been mostly lacking coronary angiographic data, we analyzed the relationship between the presence and severity of coronary atherosclerosis based on angiography and hyperinsulinemia or hyperglycemia. Two hundred and thirty subjects underwent coronary angiography and a 75-g oral glucose tolerance test. Age, sex, waist-to-hip ratio, postchallenge 1-h and 2-h glucose levels, plasma triglyceride and HDL-cholesterol levels were different between those with or without CAD. However, there was no significant difference in the plasma insulin levels, area of insulin under the curve, and the ratio of the insulin- and glucose areas between the groups with and without CAD. Multiple logistic regression analysis including fasting-, 1-h, and 2-h glucose values and a variety of atherosclerosis risk factors showed that age, sex and postchallenge 2-h glucose levels were independent determinants of the presence of CAD. These results suggest that coronary atherosclerosis might be associated with postchallenge hyperglycemia, but not with hyperinsulinemia in Korean subjects.

Adult↗
PubMed2003

ASK1-p38 MAPK/JNK signaling cascade mediates anandamide-induced PC12 cell death.

Anandamide is a neuroimmunoregulatory molecule that triggers apoptosis in a number of cell types including PC12 cells. Here, we investigated the molecular mechanisms underlying anandamide-induced cell death in PC12 cells. Anandamide treatment resulted in the activation of p38 mitogen-activated protein kinase (MAPK), c-Jun N-terminal kinase (JNK), and p44/42 MAPK in apoptosing cells. A selective p38 MAPK inhibitor, SB203580, or dn-JNK, JNK1(A-F) or SAPKbeta(K-R), blocked anandamide-induced cell death, whereas a specific inhibitor of MEK-1/2, U0126, had no effect, indicating that activation of p38 MAPK and JNK is critical in anandamide-induced cell death. An important role for apoptosis signal-regulating kinase 1 (ASK1) in this event was also demonstrated by the inhibition of p38 MAPK/JNK activation and death in cells overexpressing dn-ASK1, ASK1 (K709M). Conversely, the constitutively active ASK1, ASK1DeltaN, caused prolonged p38 MAPK/JNK activation and increased cell death. These indicate that ASK1 mediates anandamide-induced cell death via p38 MAPK and JNK activation. Here, we also found that activation of p38 MAPK/JNK is accompanied by cytochrome c release from the mitochondria and caspase activation (which can be inhibited by SB203580), suggesting that anandamide triggers a mitochondrial dependent apoptotic pathway. The caspase inhibitor, zVAD, and the mitochondrial pore opening inhibitor, cyclosporine A, blocked anandamide-induced cell death but not p38 MAPK/JNK activation, suggesting that activation of these kinases may occur upstream of mitochondrial associated events.

Animals↗
PubMed2003

Ebselen inhibits NO-induced apoptosis of differentiated PC12 cells via inhibition of ASK1-p38 MAPK-p53 and JNK signaling and activation of p44/42 MAPK and Bcl-2.

Ebselen, a selenium-containing heterocyclic compound, prevents ischemia-induced cell death. However, the molecular mechanism through which ebselen exerts its cytoprotective effect remains to be elucidated. Using sodium nitroprusside (SNP) as a nitric oxide (NO) donor, we show here that ebselen potently inhibits NO-induced apoptosis of differentiated PC12 cells. This was associated with inhibition of NO-induced phosphatidyl Serine exposure, cytochrome c release, and caspase-3 activation by ebselen. Analysis of key apoptotic regulators during NO-induced apoptosis of differentiated PC12 cells showed that ebselen blocks the activation of the apoptosis signaling-regulating kinase 1 (ASK1), and inhibits phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-jun N-terminal protein kinase (JNK). Moreover, ebselen inhibits NO-induced p53 phosphorylation at Ser15 and c-Jun phosphorylation at Ser63 and Ser73. It appears that inhibition of p38 MAPK and p53 phosphorylation by ebselen occurs via a thiol-redox-dependent mechanism. Interestingly, ebselen also activates p44/42 MAPK, and inhibits the downregulation of the antiapoptotic protein Bcl-2 in SNP-treated PC12 cells. Together, these findings suggest that ebselen protects neuronal cells from NO cytotoxicity by reciprocally regulating the apoptotic and antiapoptotic signaling cascades.

Animals↗
PubMed2003

Cdk7 functions as a cdk5 activating kinase in brain.

Activity of Cdk5, which is essential for CNS development, depends on its association with p35 (or truncated p25) or p39, but the regulatory mechanisms whereby Cdk5 is activated have not yet been fully elucidated. Studies on PC12 cells showed that Cdk5/p25 activity is modulated by an unidentified kinase that phosphorylates Cdk5 at Ser159 and enhances its kinase activity (Sharma et al., 1999 PNAS USA 96,11156). In this study, we have partially purified a bovine brain enzyme that can phosphorylate the Cdk5(149-163) peptide but not the Cdk5(149-163)A159 peptide. The enzyme, which contains Cdk7, cyclin H, and Mat1, specifically phosphorylates Ser159 of Cdk5(149-163) and enhances Cdk5/p25 activity. The active Cdk7-containing enzyme also phosphorylates and activates wt Cdk5 but not mutated Cdk5(Ser159Ala). Likewise, Cdk7 or cyclin H immunoprecipitate from mouse brain specifically phosphorylates wt Cdk5 at Ser159 and enhances Cdk5/p25 activity. Conversely, blocked Cdk7 immunoprecipitate does not phosphorylate nor activate Cdk5. In addition, the Cdk7 substrate, CTD of RNAPII, causes a dose-dependent decline in Cdk5 activation by Cdk7. These findings indicate that Cdk7 functions as a Cdk5 activating kinase in brain. Indeed, elevated Cdk5 activity in the developing E18 mouse brain coincides with increased Cdk7 kinase activity.

Amino Acid Sequence↗