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The tumor microenvironment (TME) is formed by several immune cells. Notably, tumorassociated macrophages (TAMs) are existed in the TME that induce angiogenesis, metastasis, and proliferation of cancer cells. Recently, a point-mutated variant of IL-32θ was discovered in breast cancer tissues, which suppressed migration and proliferation through intracellular pathways. Although the relationship between cancer and IL-32 has been previously studied, the effects of IL-32θ on TAMs remain elusive. Recombinant human IL-32θ (rhIL-32θ) was generated using an Escherichia coli expression system. To induce M0 macrophage polarization, THP-1 cells were stimulated with PMA. After PMA treatment, the cells were cultured with IL-4 and IL-13, or rhIL-32θ. The mRNA level of M1 macrophage markers (IL-1β, TNFα, inducible nitric oxide synthase) were increased by rhIL-32θ in M0 macrophages. On the other hand, the M2 macrophage markers (CCL17, CCL22, TGFβ, CD206) were decreased by rhIL-32θ in M2 macrophages. rhIL-32θ induced nuclear translocation of the NF-κB via regulation of the MAPK (p38) pathway. In conclusion, point-mutated rhIL-32θ induced the polarization to M1-like macrophages through the MAPK (p38) and NF-κB (p65/p50) pathways.

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No abstract available.

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BACKGROUND: Systemic lupus erythematosus (SLE) is an autoimmune disease that is a significant source of morbidity and mortality when it manifests in the central nervous system. The early detection and treatment of neuropsychiatric SLE (NPSLE) is very important, but a confirmative diagnostic tool has yet to be developed. CASE REPORT: We report here a case of neuropsychiatric manifestations in a patient that were associated with SLE, and evidence of reversal of bilateral amygdala and parahippocampal lesions in the brain revealed by 18fluorodeoxy glucose-positron emission tomography. CONCLUSIONS: We are suggestive of 18fluorodeoxy glucose-positron emission tomography appear to be more sensitive in detecting subtle brain changes in NPSLE.

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Redox adaptation is an important concept that explains the mechanisms by which cancer cells survive under persistent endogenous oxidative stress and become resistant to certain anticancer agents. To investigate this concept, we determined the expression levels of peroxiredoxins (Prxs), antioxidant enzymes in drug-resistant non-small cell lung carcinoma cells. Prx II was remarkably increased only in A549/GR (gefitinib-resistant) cells compared with A549 cells, consistent with methylation/demethylation. Prx II was highly methylated in the A549 cells but was demethylated in the A549/GR cells. The elevated expression of Prx II resulted in the downregulation of reactive oxygen species (ROS) and cell death and upregulation of cell cycle progression in the A549/GR cells. When Prx II mRNA in the A549/GR cells was knocked down, the levels of ROS and apoptosis were significantly recovered to the levels of the controls. In addition, signaling molecules involved in apoptosis were increased in the A549/GR-shPrx II cells. There was no difference in the expression of MAPK/ERK between the A549/GR cells and A549/GR-shPrx II cells, but the phosphorylation of JNK was increased in the A549/GR cells and was markedly decreased in the A549/GR-shPrx II cells. Colony number and tumor growth were significantly decreased in the A549/GR-shPrx II cells compared with the A549/GR cells. Our findings suggest that Prx II has an important role in cancer cell survival via the modulation of signaling molecules involved in apoptosis and the phosphorylation of JNK by the downregulation of ROS levels in A549/GR cells.

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Redox adaptation is an important concept that explains the mechanisms by which cancer cells survive under persistent endogenous oxidative stress and become resistant to certain anticancer agents. To investigate this concept, we determined the expression levels of peroxiredoxins (Prxs), antioxidant enzymes in drug-resistant non-small cell lung carcinoma cells. Prx II was remarkably increased only in A549/GR (gefitinib-resistant) cells compared with A549 cells, consistent with methylation/demethylation. Prx II was highly methylated in the A549 cells but was demethylated in the A549/GR cells. The elevated expression of Prx II resulted in the downregulation of reactive oxygen species (ROS) and cell death and upregulation of cell cycle progression in the A549/GR cells. When Prx II mRNA in the A549/GR cells was knocked down, the levels of ROS and apoptosis were significantly recovered to the levels of the controls. In addition, signaling molecules involved in apoptosis were increased in the A549/GR-shPrx II cells. There was no difference in the expression of MAPK/ERK between the A549/GR cells and A549/GR-shPrx II cells, but the phosphorylation of JNK was increased in the A549/GR cells and was markedly decreased in the A549/GR-shPrx II cells. Colony number and tumor growth were significantly decreased in the A549/GR-shPrx II cells compared with the A549/GR cells. Our findings suggest that Prx II has an important role in cancer cell survival via the modulation of signaling molecules involved in apoptosis and the phosphorylation of JNK by the downregulation of ROS levels in A549/GR cells.

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Acute hemorrhagic leukoencephalopathy is a rare condition associated with the influenza virus, but the role of influenza in this condition has not been clarified. We experienced a patient with influenza A and B virus co-infection who initially presented mild gastrointestinal symptoms and rapidly progressed to coma. Magnetic resonance imaging showed severe brain edema and multiple intracranial hemorrhagic lesions. The patient was treated with oseltamivir and corticosteroid, and her clinical course improved without neurological sequelae.

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Acute hemorrhagic leukoencephalopathy is a rare condition associated with the influenza virus, but the role of influenza in this condition has not been clarified. We experienced a patient with influenza A and B virus co-infection who initially presented mild gastrointestinal symptoms and rapidly progressed to coma. Magnetic resonance imaging showed severe brain edema and multiple intracranial hemorrhagic lesions. The patient was treated with oseltamivir and corticosteroid, and her clinical course improved without neurological sequelae.

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Acetyl-L-carnitine (ALC), an acetylated form of L-carnitine, is able to influence the activity of cholinergic neurons, cell membrane stabilization and enhancing mitochondrial function. A 52-year-old woman was referred to neurology clinic for memory impairment within 1 year. She was administered ALC as dose of 1,500 mg per day for improving memory decline. After 14 days from administrating ALC, she complained vivid dreams at every night. Vivid dream was disappeared after ceasing ALC. Another patient, a 72-year-old man, visited neurology clinic for cognitive decline for 2 years. After 20 days from administering ALC with dose of 1,500 mg per day, he also suffered from vivid dreams at every night. His previous stable sleep was also restored after ceasing ALC. ALC supplementation may present vivid dreams as a side effect. Possibility of vivid dream as a side effect should be considered during the management with oral ALC.

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Cerebral achromatopsia, which refers to a loss of color vision, is a rare complication of posterior circulation stroke. We report two patients who presented with achromatopsia and dyschromatopsia (incomplete form of achromatopsia) respectively after acute posterior cerebral artery infarction. Lingual and fusiform gyri within the occipito-temporal area are known to be responsible for color perception.

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BACKGROUND: Human telomerase reverse transcriptase (hTERT) is a catalytic enzyme that is required for telomerase activity (TA) and cancer progression. Telomerase inhibition or inactivation increases cellular sensitivity to UV irradiation, DNA-damaging agents, the tyrosine kinase inhibitor, imatinib, and pharmacological inhibitors, such as BIBR1532. hTERT is associated with apoptosis. Some patients show drug-resistance during anti-cancer drug treatment and the cancer cell acquire anti-apoptotic mechanism. Therefore, we attempted to study correlation between hTERT and drug-resistance. METHODS: To study the correlation between protein level and activity of hTERT and drug-resistance, Western blotting and telomerase repeat amplification protocol (TRAP) assays were performed. To investigate whether hTERT contributes to drug resistance in tumor cells, we transiently decreased hTERT levels using small interfering RNA (siRNA) in T24/R2 cells. RESULTS: hTERT knockdown increased Bax translocation into the mitochondria and cytochrome C release into the cytosol. Caspase inhibitors, especially Z-VAD-FMK, rescued this phenomenon, suggesting that the stability or expression of hTERT might be regulated by caspase activity. CONCLUSIONS: These data suggest that hTERT might be a target molecule for drug-resistant tumor therapy.

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BACKGROUND: Mac-2 binding protein (Mac-2BP) is a secreted glycoprotein from the culture fluid of several human cancer cells, especially breast, lung, and gastric cells. Mac-2BP plays a role in immune response and cell adhesion activity in patients with various cancer and infectious diseases. In this study, we attempted to identify the regulators of Mac-2BP expression at the transcriptional level. METHODS: To determine the effect of epidermal growth factor (EGF) to Mac-2BP expression in gastric cancers, we constructed the different lengths of Mac-2BP promoter plasmids and measured the promoter activity and Mac-2BP expression. In addition to investigating the role of signal transducer and activator of transcription3 (STAT3) or human telomerase reverse transcriptase (hTERT) as a regulator of Mac-2BP, we transfected the small interfering RNA (siRNA) specific for STAT3 or hTERT, and Mac-2BP level was observed by a quantitative ELISA. RESULTS: EGF treatment could suppress the Mac-2BP transcription in HEK293 or gastric cancer cell lines (SNU-638 or AGS). In 5'-deleted promoter experiment, pGL3-Mac Pro-2377 transfected cells showed a decreased luciferase activity compared to pGL3-Mac Pro-2277. We also identified that (-2,366/-2,356) on Mac-2BP promoter is a putative STAT3 binding site and suppression of STAT3 with STAT3 specific siRNA increased the Mac-2BP level, suggesting the role of STAT3 as a negative regulator, in contrast to hTERT, which is known as a positive regulator. CONCLUSIONS: EGF signal is critical for the Mac-2BP expression, and more importantly, STAT3 could work as a negative regulator, while hTERT as a positive regulator in Mac-2BP transcription.

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BACKGROUND: The aim of our study was to optimize and establish erythropoietin (EPO) enzyme linked immunosorbent assay (ELISA) system. METHODS: We prepared several monoclonal and polyclonal antibodies specific to human-EPO. The best combinations of antibodies for coating and detecting antibodies were selected for the establishment of ELISA. We tested several methods such as a competitive EIA and a sandwich ELISA. RESULTS: The best sandwich ELISA was optimized compared to competitive EIA when purified polyclonal antibody (PoAb) was used as a coating antibody and biotinylated PoAb as a detecting antibody. This sandwich ELISA easily detected EPO when PoAb pairs were used compared to the ELISA using monoclonal antibody and PoAb. There were no significant differences between the effects of various blocking solutions on the performance of sandwich ELISA using biotinylated antibody. The ELISA system using PBST containing 3% BSA as a blocking solution can sensitively detect EPO (10 mU/mL) in a broad range of EPO concentrations (10-2,000 mU/mL) and there were cross-reactions with other cytokines). CONCLUSIONS: EPO can be easily determined by using biotinylated PoAb as a detecting antibody and another PoAb as a coating antibody.

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Xenotransplantation, the transplantation of cells, tissues or organs between individuals of different species, would resolve the current shortage of organs, but rejection remains the major hurdle to successful xenotransplantation. In the present study, we analyzed mixed lymphocyte reactions (MLRs) and used 51Cr release assays in order to identify the proliferation and expansion of mouse CD8+ cytotoxic T lymphocyte cells against PK15, PK15/pIL-18 or PK15/mIL-18 cells. In addition, we identified T cell populations in mouse splenocytes and lymph node cells using two-color flow cytometry. It was found that the CD8+T cells of xenograft recipients proliferated extensively and that the survival rates of populations of PK15/mIL-18 or PK15/pIL-18 cells were higher than untransfected controls. Moreover, CD3+T cells were increased in mice injected with PK15 cells or PK15/pIL-18 cells but PK15/pIL-18 cell numbers were lower in lymph nodes than untransfected controls. CD8+T cells numbers were reduced in the lymph nodes of PK15/pIL-18 injected mice. These results suggest that porcine IL-18 regulates anti-pig cellular rejection in C57BL/6 mice.

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OBJECTIVE: Attention deficit hyperactivity disorder is known to be a disorder with high genetic trait. Recently the relationship between alleles frequency distribution and ADHD, has been actively researched. In Korea, the relationship between the genetic type and alleles for COMT (Catechol-O-methyltransferase) gene, has been studied in ADHD patients. METHODS: Thirty three patients diagnosed with ADHD according to the DSM-IV diagnostic criteria were selected for the study. The diagnosis and clinical features were confirmed by Korean version Child behavior check list, Korean version Conner's parent rating scale, Attention deficit Diagnostic System, Korean version Spielberger state-trait anxiety scale etc. For the control group, the parents of patients were chosen. Blood samples were taken from the 99 subjects. DNA was extracted from blood lymphocytes, PCR was performed for COMT NlaIII VNTR Polymorphism. Allele and genotype frequencies were compared using the Chi-square test. For the family-based analyses, we used the TDT and HHRR method. RESULTS AND CONCLUSION: In comparing the ADHD transmitted group and the not transmitted group, No significant difference was seen between the COMT genetic type and alleles distribution. As a result, it is viewed that there is not relationship between ADHD and the dopamine transporter gene, but final decision is indefinite. Follow up studies with larger patient or pure subgroups are expected.

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We purified phytoestrogens from Pueraria root (Pueraria mirifica from Thailand and Pueraria lobata from Korea), which is used as a rejuvenating folk medicine in Thailand and China. Dried, powdered plant material was extracted with 100% ethanol and further separated by concentration, filtration, and thin layer silica gel chromatography. Using the fractions obtained during separation, we first investigated their cytotoxicity in several cancer cell lines from various tissues. The ethanol-extracted components (PE1, PE4) had significant antiproliferative effects on breast cancer cell lines, including MCF-7, ZR-75-1, MDA-MB-231, SK-BR-3, and Hs578T. Second, we compared these results with the cytotoxic effects of known flavonoids, sterols, and coumarins from Pueraria root. The known compounds were not as effective, and occurred in a different polarity region on HPLC. Third, further separation resulted in the isolation of eight different components (Sub PE-A to -H). One of these, PE-D, affected the growth of some breast cancer cell lines (MCF-7, MDA-MB-231) in a dose- and time-dependent manner, as well as the growth of ovarian (2774) and cervical cancer cells (HeLa). Finally, a transfection assay showed that this component had an estrogenic effect similar to 17beta-estradiol, which activates both estrogen receptor a (ER alpha) and ER beta. The NMR analysis determined that spinasterol (stigmasta-7, 22-dien-3beta-ol) is an active cytotoxic component of Pueraria root.

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BACKGROUND: Bcl-2 family proteins play a central role in regulating apoptosis. In human, over 20 members of this family have been identified to date. Bfl-1, a member of the Bcl-2 family, has been known to retard apoptosis in various cell lines. However, the function of Bfl-1 remains unclear. METHODS: In order to investigate the Bfl-1 function, we employed yeast two-hybrid system to identify the proteins which are capable of interacting with Bfl-1. The interaction of inhibitor kappaB kinase-beta (IKK-beta) and Bfl-1 was confirmed using glutathione S-transferase pull down assays. To determine which regions of IKK-beta were required for interaction with Bfl-1, we constructed 12 deletion mutants of IKK-beta and 5 deletion mutants of Bfl-1. RESULTS: Bfl-1 interacted with the C-terminal region of IKK-beta which is a subunit of IKK complex, and IKK-beta activity is very important in the NF-kappaB related pathway. In addition, the amino acids 673-745 of IKK-beta were important for Bfl-1 interactions, and amino acids 1-484 of Bfl-1, including Bcl-2 homology domains (BH1, BH2, BH3, BH4), were crucial for IKK-beta interactions. CONCLUSIONS: IKK beta C-terminus contains many serine residues as binding partner of Bfl-1. Our results suggested that Bfl-1 is involved in the NF-kappaB activation through interaction of IKK-beta and Bfl-1. Further studies need to be performed to understand functions of the IKK-beta and Bfl-1 associated with the regulation of the NF-kappaB activation pathway.

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BACKGROUND: The aim of our study was to investigate the diagnostic value of interleukin-18 (IL-18) and other cytokines in chronic liver disease patients with hepatitis B virus (HBV), including chronic hepatitis B infection (CHB), liver cirhhosis (LC) with HBV, and hepatocellular carcinoma (HCC) with HBV. METHODS: Serum levels of IL-18, interferon-gamma (INF-gamma), IL-4 and tumor necrosis factoralpha (TNF-alpha) were evaluated using ELISA in 87 patients with liver diseases with HBV (asymptomatic carriers, 9; CHB, 19; LC, 40; and HCC, 19), 13 patients with non-hepatic diseases (NHD), and 9 normal controls. RESULTS: Serum IL-18 levels of CHB, LC with HBV and HCC with HBV patients were higher than those of healthy controls and asymptomatic carriers (P<0.05). Serum IFN-gamma and IL-4 levels were not significantly increased in all groups, but TNF-alpha levels were increased in LC and HCC patients (P<0.05). CONCLUSIONS: Determination of IL-18 may be useful in assessing the severity and predicting the prognosis of liver dieases with HBV.

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BACKGROUND: Biomedical products such as viral vaccines can be contaminated with hazardous viruses during manufacturing processes and storage, thus causing harmful side effects. To assure the safety of biomedical products, highly effective and sensitive methods should be available to detect contaminating viruses. In this study, we performed recovery tests to determine the limit of detection of HIV-1. METHODS: An HIV-1 plasmid preparation was serially diluted and spiked into various culture media (DMEM, RPMI-1640, IMDM, GICM, and SDM) containing 10% fetal bovine serum (FBS). The HIV-1 plasmid was detected by PCR alone or a combination of PCR and ELISA (PCR/ELISA). RESULTS: When spiked into DMEM, RPMI, and IMDM, less than 4x10(-2) ng of HIV-1 plasmid was not detectable as HIV-1 PCR products in agarose gel. Intra- and inter-assays (n=6) showed that the PCR-ELISA system could detect PCR products diluted as much as 1, 875 times from HIV-1 plasmid serially spiked in various media. CONCLUSIONS: The PCR/ELISA system can be useful for the detection of trace amounts of hazardous viruses which may be present as contaminants in biological products.

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PURPOSE: In this study, a possible suppressive effect of a flavon extracted from Artemisia absinthium on a mouse collagen-induced arthritis (CIA) model was investigated. METHODS: DBA/1 mice were injected intradermally with emulsified chicken type II collagen. Three weeks after immunization, a flavon was introduced p.o. everyday. Clinical incidences of arthritis and arthritis index were measured. Measurement of anti-collagen antibodies and a stimulation index of the splenocytes of the mice were measured. IL-10 and TNF-alpha in the supernatants of the mice sera were measured by ELISA. mRNA expression for IL-10 and TNF-alpha in the splenocytes were tested. RESULTS: Flavon extracted from Artemisia absinthium appears to be an effective suppressor of CIA in mice. The serum anti-collagen antibody level and stimulation index of the cultured splenocytes showed no significant differences among the three experimental groups. Also serum IL-10 and TNF-alpha levels did not show any significant differences among the three experimental groups. An increased expression of mRNA for IL-10 was observed in the splenocytes treated with flavon. CONCLUSION: With these results, flavon extracted from Artemisia absinthium appears to have a suppressive effect of CIA. The mechanism of the suppressive effect of flavon extracted from Artemisia absinthium may be from a stimulation of IL-10 production.

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BACKGROUND: The fas (CD95/Apo-1)/Fas ligand (FasL) system is reported to be involved in the suppression and stimulation of immune responses, and the reactive oxygen species (ROS) play a key role in the mechanism for resisting Fas-induced apoptosis of tumor cells. In this work, we investigated the effect of endogenous interleukin (IL)-18 on the regulation of immune related factors such as Fas/Fas ligand and intercellular adhesion molecules (ICAM), and of the ROS level in IL-18 receptor (IL-18R) transfected C-33A cells. METHODS: The cervical cancer cell line C-33A was transfected with IL-18R (C-33A/IL-18R). For the detection of pro-inflammatory cytokines in C-33A/IL-18R, reverse-transcriptase (RT) polymerase chain reaction (PCR), in situ enzyme-linked immunosorbent assay (ELISA), Western blot, and Northern blot analyses were performed. The level of p53 was determined by Western blot. Intracellular ROS, ICAM-1, FasL, and apoptosis in C-33A/IL-18R were measured by flow cytometry. RESULTS: In situ ELISA and RT-PCR showed that, among pro-inflammatory cytokines, IL-18 was induced in C-33A/IL-18R whereas there appeared no induction of the IL-1alpha, IL-1beta, tumor necrosis factor (TNF)-alpha, and IL-6. IL-18R transfection showed a slight enhancement of the Fas via upregulation of intracellular ROS and IL-18 in C-33A cells whereas there was no effect on the expression of p53, ICAM-1 and FasL. However, treatment with the agonistic anti-Fas antibody showed that the enhanced surface Fas was not functional or was not enough to induce apoptosis and the C-33A/IL-18R cells escaped still resistant to Fas-mediated apoptosis. CONCLUSIONS: IL-18R transfection induced IL-18 expression and enhanced ROS and Fas expression in C-33A cells. These results show that C-33A/IL-18R cells escaped from immunuosurveillance by failure to express ICAM-1 adhesion molecules and Fas ligand, and are resistant to Fas-mediated apoptosis.

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