Yang-Kyu Choi

The antipsychotic agent chlorpromazine induces autophagic cell death by inhibiting the Akt/mTOR pathway in human U-87MG glioma cells.

2-Chloro-10-[3(-dimethylamino)propyl]phenothiazine mono hydrochloride (chlorpromazine; CPZ) is an antipsychotic agent that was originally developed to control psychotic disorders. The cytotoxic properties of the CPZ are well known, but its mechanism of action is poorly understood. In this study, we investigated the role of apoptosis and autophagy in CPZ-induced cytotoxicity in U-87MG glioma cells. CPZ treatment inhibited cell proliferation and long-term clonogenic survival. Additionally, CPZ triggered autophagy, as indicated by electron microscopy and accumulation of the membrane form of microtubule-associated protein 1 light chain 3 (LC3-II); however, CPZ did not induce apoptosis. Inhibition of autophagy by expression of Beclin 1 small interfering RNA (siRNA) in U-87MG cells attenuated CPZ-induced LC3-II formation. Furthermore, U-87MG cells expressing Beclin 1 siRNA attenuated CPZ-induced cell death. CPZ inhibited phosphatidylinositol 3-kinase (PI3K)/AKT/ mTOR pathway in U-87MG cells. Treatment with LY294002, a PI3K inhibitor, alone increased the accumulation of LC3-II and potentiated the effect of CPZ. In contrast, exogenous expression of AKT partially inhibited CPZ-induced LC3-II formation. When U-87MG cells were implanted into the brain of athymic nude mouse, CPZ triggered autophagy and inhibited xenograft tumor growth. These results provided the first evidence that CPZ-induced cytotoxicity is mediated through autophagic cell death in PTEN (phosphatase and tensin homolog deleted on chromosome 10)-null U-87MG glioma cells by inhibiting PI3K/AKT/mTOR pathway.

ANXA8 Down-regulation by EGF-FOXO4 Signaling Is Involved in Cell Scattering and Tumor Metastasis of Cholangiocarcinoma

BACKGROUND & AIMSnThe sarcomatoid change in cholangiocarcinoma (CC) contributes to more aggressive intrahepatic spread and widespread metastasis. Therefore, the aim of this study was to identify the molecular mechanisms of CC metastasis during tumor progression and sarcomatoid change.nnnMETHODSnUsing the subtraction suppression hybridization (SSH) method, we identified altered expression of the candidate gene ANXA8 and epidermal growth factor receptor (EGFR) in sarcomatoid CC cells. We assessed ANXA8 expression during the progression of CC in cells and tissues and examined its functional significance by performing in vitro cell experiments and using in vivo animal models.nnnRESULTSnANXA8 is highly expressed in human and hamster CCs but is down-regulated with tumor dedifferentiation. ANXA8 is transcriptionally down-regulated by epidermal growth factor (EGF), which is correlated with the morphologic changes of the epithelial-to-mesenchymal transition (EMT) in the CC cells. Furthermore, ectopic ANXA8 reverses the morphology of cells, and this is associated with focal adhesion kinase expression and altered F-actin dynamics. EGFR and its downstream targets, phosphatidylinositol-3-kinase and Akt, are linked to the phosphorylation of FOXO4, which leads to the inhibition of ANXA8 transcription. In addition, an in vitro cell invasion assay and in vivo spontaneous metastasis assay reveal that ANXA8 inhibits the cell migratory and metastatic characteristics of CC cells.nnnCONCLUSIONSnThese findings suggest that FOXO4 and ANXA8 play key roles in growth factor-mediated tumor progression and metastasis during the EMT change in CC.

Retrospective study of canine cutaneous tumors in Korea.

Over the 42 month period from January 2003 to June 2006, a total of 2,952 canine biopsy specimens were received from the Veterinary Medical Teaching Hospital of Seoul National University and from veterinary practitioners across the nation. Out of these, 748 (25.34%) cases were diagnosed as canine cutaneous tumors in the Department of Veterinary Pathology, College of Veterinary Medicine, Seoul National University, Korea. Thirty-eight different types of cutaneous tumors were identified and categorized into epithelial and melanocytic tumors (56.95%), mesenchymal tumors (38.90%), and hematopoietic tumors (4.14%) located in the skin. Among these, 69.25% were benign and 30.74% were malignant. The top ten most frequently diagnosed cutaneous tumors were epidermal and follicular cysts (12.70%), lipoma (11.36%), mast cell tumors (8.82%), cutaneous histiocytoma (7.49%), basal cell tumors (6.82%), sebaceous gland adenoma (6.68%), sebaceous gland hyperplasia (5.08%), hepatoid gland adenoma (3.61%), apocrine adenocarcinoma (3.07%), and fibroma (2.81%), in order of prevalence. They comprised 68.45% of all cutaneous tumors. These top ten cutaneous tumors were distributed on the trunk (30.08%), head and neck (20.9%), extremities (19.14%), anal and perianal area (8.59%), and tail (3.91%). The age of the dogs with the ten most frequent tumors had a mean age of 8.3 years, with a range of 2 months to 19 years. When all types of tumors were considered together in the entire population, there was no difference in incidence according to sex.

Direct activation of TGF‐β1 transcription by androgen and androgen receptor complex in Huh7 human hepatoma cells and its tumor in nude mice

Importance of androgen for promotion of hepatocelullar carcinoma (HCC) has long been supported by clinical and experimental evidences. However, mechanisms involved in the carcinogenesis have not yet been fully elucidated. Moreover, unbalanced expression of TGF‐β1 during tumor progression results in prooncogenic rather than growth inhibition. To investigate the effect of androgen on transcriptional regulation of TGF‐β1, we isolated rat TGF‐β1 promoter, based on our previous report (GenBank AF249327), and examined regulation of its promoter activity by dihydrotestosterone in Huh7, LNCaP, and PC3 cells. Several putative transcription factor‐binding sites were found, but no TATA box. When the full‐length (−4784 to +68) and variously deleted promoter DNAs were evaluated, the promoter region spanning from −2732 to −1203 showed the highest activity towards dihydrotestosterone in a dose‐dependent manner in both Huh7 and PC3 cells with androgen receptor (AR) expression. Putative androgen response sequence half site (5′‐TGTCCT‐3′) was identified to be located within −1932 to −1927, proved by mutant (5′‐AGACCT‐3′) analysis and chromatin immunoprecipitation (ChIP) assay. AR mediated upregulation of TGF‐β1 expression was confirmed by HCC developed in nude mice with AR‐overexpressed Huh7‐cells. This work presents in vivo and in vitro evidences of activation of TGF‐β1 expression by androgen and AR, and implicates the modulation of hepatocarcinogenesis by AR through the regulation of TGF‐β1 expression. J. Cell. Biochem.

Sexual activity counteracts the suppressive effects of chronic stress on adult hippocampal neurogenesis and recognition memory

Adult neurogenesis can be influenced by a variety of factors. Stress is one of the most potent inhibitors of hippocampal neurogenesis. Stress effects on adult hippocampal neurogenesis are affected differently by environmental factors, including social interaction. Sexual behavior between males and females in a social context has been suggested to influence neurogenesis and enhance hippocampal cell proliferation. However, the mechanisms of action of sexual interaction, the possible changes relative to stress state, and its effects on learning and memory remain uncertain. The current study examined the influence of sexual interaction on neurological responses in adult male mice and the function of sexual interaction relative to recognition memory in stress states. Changes in the expression of neurotrophic and transcription factors were assessed in reference to stress and/or sexual behaviors. The survival of newly generated cells and their rate of differentiation into neurons were determined in the hippocampus of chronically stressed and/or sexually experienced mice. Finally, to evaluate whether sexual experience alters adult hippocampal function, we tested learning and memory in a recognition memory task. The results demonstrated that sexual activity increased the expression of brain-derived neurotrophic factor, tyrosine kinase B, and cAMP response element-binding factor. Furthermore, the results supported the view that sexual interaction could be helpful for buffering adult hippocampal neurogenesis and recognition memory function against the suppressive actions of chronic stress.

IL-10 suppresses bactericidal response of macrophages against Salmonella Typhimurium.

We report, herein, an attempt to determine whether an IL-10-induced immunological state affects the response of macrophages against Salmonella Typhimurium (ST). Pretreatment with mrIL-10 induced the intracellular invasion of ST into macrophages in a dose-dependent manner. It also activated AKT phosphorylation, cyclin D1, Bcl-XL, and COX-2 upon ST infection, which may correlate with Salmonella’s survival within the macrophages. However, I-κB phosphorylation was shown to be inhibited, along with the expression of TNF-α and MIP-2α mRNA. Therefore, IL-10 not only suppresses the bactericidal response of macrophages against ST, but also ultimately causes infected macrophages to function as hosts for ST replication.

Therapeutic effects of lentivirus-mediated shRNA targeting of cyclin D1 in human gastric cancer

BackgroundGastric cancer is the second most common cause of cancer-related death in males and the fourth in females. Traditional treatment has poor prognosis because of recurrence and systemic side effects. Therefore, the development of new therapeutic strategies is an important issue. Lentivirus-mediated shRNA stably inhibits target genes and can efficiently transduce most cells. Since overexpressed cyclin D1 is closely related to human gastric cancer progression, inhibition of cyclin D1 using specific targeting could be an effective treatment method of human gastric cancer.MethodsThe therapeutic effect of lentivirus-mediated shRNA targeting of cyclin D1 (ShCCND1) was analyzed both in vitro and in vivo experiments.ResultsIn vitro, NCI-N87 cells with downregulation of cyclin D1 by ShCCND1 showed significant inhibition of cell proliferation, cell motility, and clonogenicity. Downregulation of cyclin D1 in NCI-N87 cells also resulted in significantly increased G1 arrest and apoptosis. In vivo, stable NCI-N87 cells expressing ShCCND1 were engrafted into nude mice. Then, the cancer-growth inhibition effect of lentivirus was confirmed. To assess lentivirus including ShCCND1 as a therapeutic agent, intratumoral injection was conducted. Tumor growth of the lentivirus-treated group was significantly inhibited compared to growth of the control group. These results are in accordance with the in vitro data and lend support to the mitotic figure count and apoptosis analysis of the tumor mass.ConclusionThe lentivirus-mediated ShCCND1 was constructed, which effectively inhibited growth of NCI-N87-derived cancer both in vitro and in vivo. The efficiency of shRNA knockdown and variation in the degree of inhibition is mediated by different shRNA sequences and cancer cell lines. These experimental results suggest the possibility of developing new gastric cancer therapies using lentivirus-mediated shRNA.

Difference in Resistance to Streptococcus pneumoniae Infection in Mice

Streptococcus pneumoniae is a major pathogen that causes various diseases, including pneumonia and sepsis, as millions of people suffer from S. pneumoniae infection worldwide. To better understand the immune and inflammatory responses to S. pneumoniae, we produced murine models. To investigate the differences between intranasal and intratracheal infection, BALB/c mice were infected with S. pneumoniae D39 intranasally or intratracheally. Mice showed no significant differences in survival rates, body weight changes, and bacterial loads. To investigate resistance and susceptibility among mouse strains, BALB/c, C57BL/6J, tumor necrosis factor-α (TNF-α) knockout, and interleukin-10 (IL-10) knockout mice were infected with S. pneumoniae D39 via intranasal or intravenous routes. In this study, BALB/c and C57BL/6J mice were resistant, IL-10 knockout mice were intermediate, and TNF-α knokout mice were susceptible to S. pneumoniae infection. These data show that intranasal and intratracheal infection induced similar results after S. pneumoniae infection, and the genetic background of mice must be considered when studying S. pneumoniae infection in vivo.

Effects of Korean Red Ginseng extract on busulfan-induced dysfunction of the male reproductive system

Background Anticancer agents induce a variety of adverse effects when administered to cancer patients. Busulfan is a known antileukemia agent. When administered for treatment of leukemia in young patients, busulfan could cause damage to the male reproductive system as one of its adverse effects, resulting in sterility. Methods We investigated the effects of Korean Red Ginseng extract (KRGE) on busulfan-induced damage and/or dysfunction of the male reproductive system. Results We found that administration of busulfan to mice: decreased testis weight; caused testicular histological damage; reduced the total number of sperm, sperm motility, serum testosterone concentration; and eventually, litter size. Preadministration of KRGE partially attenuated various busulfan-induced damages to the male reproductive system. These results indicate that KRGE has a protective effect against busulfan-induced damage to the male reproduction system. Conclusion The present study shows a possibility that KRGE could be applied as a useful agent to prevent or protect the male reproductive system from the adverse side effects induced by administration of anticancer agents such as busulfan.

Expression of hepatitis B virus x protein in hepatocytes suppresses CD8 T cell activity.

Background CD8+ T cells contribute to the clearance of Hepatitis B virus (HBV) infection and an insufficient CD8+ T cell response may be one of the major factors leading to chronic HBV infection. Since the HBx antigen of HBV can up-regulate cellular expression of several immunomodulatory molecules, we hypothesized that HBx expression in hepatocytes might affect CD8+ T cell activity. Methods We analyzed the activation and apoptosis of CD8+ T cells co-cultured with primary hepatocytes rendered capable of expressing HBx by recombinant baculovirus infection. Results Expression of HBx in hepatocytes induced low production of interferon-γ and apoptosis of CD8+ T cells, with no effect on CD8 T cell proliferation. However, transcriptional levels of H-2K, ICAM-1 and PD-1 ligand did not correlate with HBx expression in hepatocytes. Conclusion Our results suggest that HBx may inhibit CD8+ T cell response by regulation of interferon-γ production and apoptosis.