Ho Joong Sung

Suppressing effect of resveratrol on the migration and invasion of human metastatic lung and cervical cancer cells

The antioxidant 3,4′,5 tri-hydroxystilbene (resveratrol), a phytoalexin found in grapes, shows cancer preventive activities, including inhibition of migration and invasion of metastatic tumors. However, the molecular mechanism underlying the effect of resveratrol on tumor metastasis, especially in human metastatic lung and cervical cancers is not clear. A non-cytotoxic dosage of resveratrol causes a reduction in the generation of reactive oxygen species, and suppresses phorbol 12-myristate 13-acetate (PMA)-induced invasion and migration in both A549 and HeLa cells. Resveratrol also decreases both the expression and the enzymatic activity of matrix metalloproteinase-9 (MMP-9), and the promoter activity of PMA-stimulated MMP-9 is also inhibited. However, resveratrol does not affect either the expression or the proteolytic activity of MMP-2. Our results also show that resveratrol suppresses the transcription of MMP-9 by the inhibition of both NF-κB and AP-1 transactivation. These results indicate that resveratrol inhibits both NF-κB and AP-1 mediated MMP-9 expression, leading to suppression of migration and invasion of human metastatic lung and cervical cancer cells. Resveratrol has potential for clinical use in preventing invasion by human metastatic lung and cervical cancers.

Regulation of Human LZIP Expression by NF-κB and Its Involvement in Monocyte Cell Migration Induced by Lkn-1

Human LZIP is a transcription factor that is involved in leukocyte cell mobility. Expression of LZIP is known to differentially regulate monocyte cell migration induced by CCR1-dependent chemokines. However, its transcriptional regulation has not been characterized. Our results indicate that Lkn-1 induces LZIP expression in a time- and dose-dependent manner, and the induction of LZIP shows an immediate early response to Lkn-1. We identified and cloned ∼1.4 kb of the LZIP promoter from a human genomic DNA. To identify regulatory elements controlling restricted expression of LZIP, deletion mutants were constructed from the 1469-bp LZIP promoter region (–1219/+251) linked to the luciferase reporter gene. Maximal promoter activity was contained within 613 bp from the tentative transcription initiation site and was sharply reduced in a truncated construct (–338/+251). This promoter sequence contained consensus NF-κB- and Sp-1-binding sites. Results from an inhibitor assay showed that NF-κB is involved in Lkn-1-induced LZIP expression, but Sp-1 is not. We also demonstrated that NF-κB binds to the LZIP promoter and that the binding is specific, as revealed by an electrophoretic mobility shift assay and a mutation analysis. Chemotaxis analysis showed that LZIP expression because of the NF-κB subfamily is specifically involved in Lkn-1-induced chemotaxis. Our findings suggest that transcription factor NF-κB plays an important role in regulation of LZIP expression, and LZIP expression regulates the monocyte cell migration induced by Lkn-1.

Inhibitory effect of Trolox on the migration and invasion of human lung and cervical cancer cells.

The antioxidant 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) is implicated in migration and invasion of metastatic tumors. However, the molecular mechanism underlying the effect of Trolox on metastatic cancer cells is not known. We found that a non-cytotoxic dose of Trolox decreased phorbol 12-myristate 13-acetate (PMA)-induced invasion and migration of both A549 and HeLa cancer cells. We also found that Trolox suppressed both the expression and the proteolytic activity of matrix metalloproteinase-9 (MMP-9), and that the promoter activity of PMA-induced MMP-9 was inhibited by Trolox. Our results show that Trolox inhibits the transcriptional activity of MMP-9 by suppression of NF-κB transactivation. These results indicate that Trolox inhibits NF-κB-mediated MMP-9 expression, leading to the suppression of migration and invasion in lung and cervical cancer cells. Trolox is a potential agent for clinical use in preventing the invasion and metastasis of human malignant lung and cervical cancers.

N-acetyl cysteine suppresses the foam cell formation that is induced by oxidized low density lipoprotein via regulation of gene expression

Foam cells derived from macrophages have been implicated as markers of early stage atherosclerosis development. In this study, we found that N-acetyl cysteine (NAC), a well-known inhibitor of reactive oxygen species (ROS), decreased the generation of ROS and suppressed foam cell formation in the presence of oxidized low density lipoprotein through down-regulation of cluster of differentiation 36 expression. We investigated gene expression profiles in order to determine the effects of NAC on foam cell formation using a microarray analysis. The level of apolipoprotein E, which is involved in lipid efflux, was increased and the levels of the antioxidant genes glutathione peroxidase 1 and 3 were also increased. The expression levels of the oxidative stress response and the DNA repair genes were decreased. These results were confirmed using quantitative real-time PCR. Our results indicate that oxidative stress plays an important role in foam cell formation, and that regulation of oxidation using antioxidants is a potential therapeutic method for blocking atherosclerosis development.

Human LZIP induces monocyte CC chemokine receptor 2 expression leading to enhancement of monocyte chemoattractant protein 1/CCL2-induced cell migration

Chemokines and chemokine receptors play a role in migration of circulating leukocytes to the region of inflammation. Human LZIP is an uncharacterized transcription factor and is known to participate in leukotactin (Lkn)-1/CCL15-induced cell migration. We investigated the role of human LZIP in expression of CC chemokine receptors (CCRs) and its involvement in monocyte migration. RNase protection analysis showed that LZIP increased mRNA expression of CCR2 and CCR1 in THP-1 cells. Surface expressions of both CCR2 and CCR1 were also increased by LZIP. Results from an electrophoretic mobility shift assay showed that LZIP binds to the C/EBP element in the CCR2 promoter. LZIP also enhanced the chemotactic activities of monocyte chemoattractant protein-1/CCL2 and Lkn-1. These results suggest that LZIP regulates expression of chemokine receptors that are involved in monocyte migration.

Triglyceride (TG) down-regulates expression of MCP-1 and CCR2 in PMA-derived THP-1 macrophages

Triglycerides (TGs) are implicated in the development of atherosclerosis. A key contributing factor for atherosclerosis is the migration of macrophages to atherosclerotic lesions. MCP-1 is a major chemoattractant for macrophages to atherosclerotic lesions. We examined the expression profile of MCP-1 and CCR2 in THP-1 macrophages in response to TG treatment by RT-PCR analysis. Chemical inhibitors were used to identify cell signaling pathway(s) involved in regulation of MCP-1 and CCR2 expression. We found that treatment of THP-1 macrophages with TG down-regulated MCP-1 expression in a time and dose-dependent manner. PMA treatment alone did not affect MCP-1 expression. Using chemical inhibitors of cell signaling pathways, we found that the NF-κB inhibitor inhibited TG-induced down-regulation of MCP-1. CCR2 expression decreased after TG treatment in THP-1 macrophages and the PKC inhibitor alleviated TG-induced down-regulation of CCR2. Our results provide further insights into the role of TG on macrophages during atherosclerosis.

Effects of MTNR1B variants on fasting glucose levels in a Korean population

Fasting glucose level is the most basic and widely used indicator of diabetes. Several genome wide association studies have reported that the gene encoding melatonin receptor 1B (MTNR1B) exerts a major effect on serum fasting glucose levels. We tested for the association between single nucleotide polymorphisms (SNPs) in the MTNR1B gene and fasting glucose levels in a Korean population consisting of 8,229 subjects taken from two community-based cohorts. The mean age of the subjects in the study population was 51.9 years. For this study, we selected 363 SNPs located in the MTNR1B gene, which is located on chromosome 11. Multivariate linear regression models were used to test for genotypic effects on fasting glucose levels while adjusting for age and sex under an additive model. The MTNR1B SNP most highly associated with fasting glucose levels was rs10830962 (p=1.95×10−5), followed by rs3847554 (p=3.16×10−4). Replication of these initial findings is important to better understand the correlation between MTNR1B variations and their effects, especially in Asian populations.

Gene and protein expression profiles in a mouse model of collagen-induced arthritis

The risk of rheumatoid arthritis (RA), an autoimmune disease, in the elderly population increases along with that of atherosclerosis, cardiovascular disease, type 2 diabetes, and Alzheimers disease. Identifying specific biomarkers for RA can clarify the underlying molecular mechanisms and can aid diagnosis and patient care. To this end, the present study investigated the genes and proteins that are differentially expressed in RA using a mouse collagen-induced arthritis (CIA) model. We performed gene microarray and proteome array analyses using blood samples from the mice and found that 50 genes and 24 proteins were upregulated and 48 genes were downregulated by more than 2-fold in the CIA model relative to the control. The gene microarray and proteome array results were validated by evaluating the expression levels of select genes and proteins by real-time PCR and western blotting, respectively. We found that the level of integrin α2, which has not been previously reported as a biomarker of RA, was significantly increased in CIA mice as compared to controls. These findings provide a set of novel biomarkers that can be useful for diagnosing and evaluating the progression of RA.

Melatonin modulates adiponectin expression on murine colitis with sleep deprivation

AIM To determine adiponectin expression in colonic tissue of murine colitis and systemic cytokine expression after melatonin treatments and sleep deprivation. METHODS The following five groups of C57BL/6 mice were used in this study: (1) group I, control; (2) group II, 2% DSS induced colitis for 7 d; (3) group III, 2% DSS induced colitis and melatonin treatment; (4) group IV, 2% DSS induced colitis with sleep deprivation (SD) using specially designed and modified multiple platform water baths; and (5) group V, 2% DSS induced colitis with SD and melatonin treatment. Melatonin (10 mg/kg) or saline was intraperitoneally injected daily to mice for 4 d. The body weight was monitored daily. The degree of colitis was evaluated histologically after sacrificing the mice. Immunohistochemical staining and Western blot analysis was performed using anti-adiponectin antibody. After sampling by intracardiac punctures, levels of serum cytokines were measured by ELISA. RESULTS Sleep deprivation in water bath exacerbated DSS induced colitis and worsened weight loss. Melatonin injection not only alleviated the severity of mucosal injury, but also helped survival during stressful condition. The expression level of adiponectin in mucosa was decreased in colitis, with the lowest level observed in colitis combined with sleep deprivation. Melatonin injection significantly (P < 0.05) recovered the expression of adiponectin. The expression levels of IL-6 and IL-17 were increased in the serum of mice with DSS colitis but decreased after melatonin injection. CONCLUSION This study suggested that melatonin modulated adiponectin expression in colonic tissue and melatonin and adiponectin synergistically potentiated anti-inflammatory effects on colitis with sleep deprivation.