Panax ginseng roots have long been used as a medicinal herb in oriental countries. We have investigated anti-proliferative effects of lipid soluble Panax ginseng components on human renal cancer cell lines. Petroleum ether extract of Panax ginseng roots (GX-PE) or its partially purified preparation (7:3 GX) was added to cultures of three human renal cell carcinoma (RCC) cell lines, A498, Caki-1, and CURC II. Proliferation of RCC cells was estimated by a [3H]thymidine incorporation assay and cell cycle distribution was analyzed by flow cytometry. GX-PE, 7:3 GX, panaxydol and panaxynol inhibited proliferation of all three RCC cell lines in a dose dependent manner in vitro with an order of potency, 7:3 GX > panaxydol > panaxynol = GX-PE. Additive effect of interleukin 4 was also demonstrated, most prominently in Caki-1 which responded poorly to GX-PE alone. Analysis of cell cycle in CURC II and Caki-1 treated with GX-PE demonstrated increase in G1 phase population and corresponding decrease in S phase population. The present study demonstrated that proliferation of human RCC cell lines were inhibited by lipid soluble components of Panax ginseng roots by blocking cell cycle progression at G1 to S phase transition.
Alkanes ; Alkynes/therapeutic use ; Antineoplastic Agents/therapeutic use ; Antineoplastic Agents, Phytogenic/therapeutic use* ; Carcinoma, Renal Cell/drug therapy* ; Cell Cycle/drug effects ; Fatty Alcohols/therapeutic use ; Ginseng/therapeutic use* ; Ginseng/chemistry ; Human ; Interleukin-4/therapeutic use ; Kidney Neoplasms/drug therapy* ; Plant Extracts/therapeutic use ; Plant Roots/therapeutic use ; Plant Roots/chemistry

Interleukin (IL)-4 inhibits proliferation of several human cancer cell lines in vitro. Although IL-4 is known to regulate proliferation of lymphocytes by modulating p27KIP1 expression, the mechanism involved in the IL-4-induced growth inhibition of nonhematopoietic cancer cells has not been fully elucidated. Previously, we reported that IL-4 suppressed proliferation of human renal cell carcinoma (RCC) cell lines in vitro. Here, we show that IL-4 inhibits cell cycle progression at the G1 phase in Caki-1 cells by increasing the expression of p21WAF1 and interferon regulatory factor (IRF)-1, and decreasing the cyclin dependent kinase (CDK) 2 activity. Up-regulation of p21WAF1 and IRF-1 expression is transcriptional, but independent of p53. The levels of p21WAF1 and IRF-1 proteins were enhanced as early as 1 h after IL-4 treatment. CDK2 activity started to decline at 4 h after IL-4 treatment, and by 24 h, was ~50% of the control. Neither the protein expressions of p27KIP1 and p16INK4a, nor the phosphorylation level of pRb was changed. The importance of p21WAF1 and IRF-1 in the growth inhibition induced by IL-4 was confirmed by antisense oligonucleotide transfection. Both of p21WAF1 and IRF-1 antisense oligonucleotides prevented IL-4-mediated growth inhibition by ~30% compared to the respective sense oligonucleotides. In summary, our study indicated that p21WAF1 and IRF-1 mediate the growth inhibitory effect of IL-4 in human RCC cells.
CDC2-CDC28 Kinases/metabolism ; Carcinoma, Renal Cell/genetics/*metabolism ; Cell Cycle/drug effects ; Cell Cycle Proteins/genetics/*metabolism ; Cell Line, Tumor ; Cell Proliferation/drug effects ; DNA-Binding Proteins/genetics/*metabolism ; Gene Expression/drug effects ; Humans ; Interleukin-4/*pharmacology ; Kidney Neoplasms/genetics/*metabolism ; Oligonucleotides, Antisense/genetics ; Phosphoproteins/genetics/*metabolism ; Research Support, Non-U.S. Gov't

Lung cancer is one of the deadliest and commonly diagnosed neoplasms. Early diagnosis of this disease is critical for improving clinical outcome and prognosis. Because the early stages of lung cancer often produce no symptoms, it is necessary to identify biomarkers for early detection, prognostic evaluation, and recurrence monitoring of the cancer. To identify potential lung cancer biomarkers, we analyzed the differential protein secretion from transformed bronchial epithelial cells (1198 and 1170-I) as compared to immortalized normal bronchial epithelial cells (BEAS-2B) and non-transformed cells (1799) all of which are derived from BEAS-2B and represent multistage bronchial epithelial carcinogenesis. The proteins recovered from the conditioned media of the cells were separated on two-dimensional gels. There was little difference between the secretome of the BEAS-2B and 1799 cells, whereas the patterns between the transformed 1198 and 1170-I cells and non-transformed 1799 cells were significantly different. Using mass spectrometry and database search, we identified 20 proteins including protein gene product 9.5 (PGP9.5), translationally controlled tumor protein (TCTP), tissue inhibitors of metalloproteinases-2 (TIMP-2), and triosephosphate isomerase (TPI), that were either increased or decreased simultaneously in conditioned media of both 1198 and 1170-I cells. Furthermore, levels of PGP9.5, TCTP, TIMP-2, and TPI were significantly increased not only in the conditioned media of both transformed cell lines when compared to those of BEAS-2B and 1799 cells, but also in plasmas and tissues from lung cancer patients when compared to those in normal controls. We suggest the PGP9.5, TCTP, TIMP-2, and TPI as promising candidates for lung cancer serum biomarkers.
Aged ; Aged, 80 and over ; Bronchi/cytology ; Cell Line, Transformed ; Culture Media, Conditioned ; Epithelial Cells/*metabolism ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Lung Neoplasms/*diagnosis/metabolism ; Male ; Middle Aged ; Proteomics ; Respiratory Mucosa/cytology ; Tumor Markers, Biological/*metabolism

Amelogenesis imperfecta (AI) occurs either in isolation or in association with other dental abnormalities and systemic disorder. A rare syndrome associating AI with nephrocalcinosis was named as Enamel Renal Syndrome (ERS; OMIM #204690). This syndrome is characterized by severe enamel hypoplasia, failed tooth eruption, intra pulpal calcifications, enlarged gingiva, and nephrocalcinosis. Nephrocalcinosis is a condition where calcium salts are deposited in renal tissue, and this may lead to critical kidney complications. This rare syndrome shows pathognomonic oral characteristics that are easily detectable at an early age, which proceeds the onset of renal involvement. Pediatric dentists are the first oral health practitioners whom ERS patients will meet at early age. The role of pediatric dentists is critically important for early diagnosis and referral of patients to both nephrologists for renal assessment and geneticists for identification of causative mutation and diagnosis. Early detection of renal involvement may provide chances to prevent further undesired renal complications.

PURPOSE: Reactive oxygen species have been recognized as a common signaling mediator in diverse stimuli-induced apoptosis and hydrogen peroxide is a natural one of those reactive oxygen species. This study was performed to investigate the role of caspases in hydrogen peroxide-induced apoptosis of HL 60 cells. METHODS: Apoptosis was induced in HL 60 cells by treating 50microM hydrogen peroxide for 2, 4, and 6 hrs and induction of apoptosis was confirmed by flow cytometry and DNA fragmentation analysis. Caspase substrate assay was used to show the activity of caspases and then protein levels of caspase and its substrate were analyzed using immunoblotting. RESULTS: During the apoptosis, caspase substrates assay showed the increased activity of caspase-3, -7, -10, but not that of caspase-8 nor caspase-9, and immunoblotting analysis showed decreasing procaspase-3 protein with the progression of apoptosis. Furthermore, with progression of apoptosis, analysis of caspase substrates showed retinoblastoma protein decreased while cleaved 89kD fragment of poly (ADP-ribosyl) polymerase protein increased. CONCLUSION: These results suggest that hydrogen peroxide-induced apoptosis in HL 60 cells is not associated with the activation of caspase-8 nor caspase-9. Rather, caspase-3 is directly activated and responsible for hydrogen peroxide-induced apoptosis of HL 60 cells.
Apoptosis* ; Caspase 3* ; Caspase 8 ; Caspase 9 ; Caspases ; DNA Fragmentation ; Flow Cytometry ; HL-60 Cells* ; Humans* ; Hydrogen Peroxide ; Hydrogen* ; Immunoblotting ; Leukemia* ; Reactive Oxygen Species ; Retinoblastoma Protein

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a prototypic halogenated aromatic hydrocarbon (HAH), is known as one of the most potent toxicants. At least a part of its toxic effects appears to be derived from its ability to induce TNF-alpha production. However, the signaling pathway of TCDD that leads to TNF-alpha expression has not been elucidated. In this study, we investigated the signaling mechanism of TCDD-induced TNF-alpha expression in PMA-differentiated THP-1 macrophages. TCDD induced both mRNA and protein expression of TNF-alpha in a dose- and time-dependent manner. Alpha-Naphthoflavone (NF), an aryl hydrocarbon receptor (AhR) inhibitor, prevented the TCDD-induced expression of TNF-alpha at both mRNA and protein levels. Genistein, a protein tyrosine kinase (PTK) inhibitor, and PD153035, an EGFR inhibitor, also blocked the increase of TNF-alpha expression by TCDD, indicating the role of EGFR in TCDD-induced TNF-alpha expression. On the other hand, PP2, a c-Src specific inhibitor, did not affect TCDD-induced TNF-alpha expression. EGFR phosphorylation was detected as early as 5 min after TCDD treatment. TCDD-induced EGFR activation was AhR-dependent since co-treatment with alpha-NF prevented it. ERK was found to be a downstream effector of EGFR activation in the signaling pathway leading to TNF-alpha production after TCDD stimulation. Activation of ERK was observed from 30 min after TCDD treatment. PD98059, an inhibitor of the MEK-ERK pathway, completely prevented the TNF-alpha mRNA and protein expression induced by TCDD, whereas inhibitors of JNK and p38 MAPK had no effect. PD153035, an EGFR inhibitor, as well as alpha-NF significantly reduced ERK phosphorylation, suggesting that ERK activation by TCDD was mediated by both EGFR and AhR. These results indicate that TNF-alpha production by TCDD in differentiated THP-1 macrophages is AhR-dependent and involves activation of EGFR and ERK, but not c-Src, JNK, nor p38 MAPK. A signaling pathway is proposed where TCDD induces sequential activation of AhR, EGFR and ERK, leading to the increased expression of TNF-alpha.
Animals ; Benzoflavones/pharmacology ; Cell Differentiation ; Cell Line, Tumor ; Enzyme Activation ; Genistein/pharmacology ; Hazardous Substances/*toxicity ; Humans ; MAP Kinase Signaling System/drug effects/physiology ; Macrophages/*metabolism ; Mice ; Phosphorylation ; Pyrimidines/pharmacology ; Quinazolines/pharmacology ; RNA, Messenger/metabolism ; Receptor, Epidermal Growth Factor/antagonists & inhibitors/metabolism ; Receptors, Aryl Hydrocarbon/antagonists & inhibitors ; Signal Transduction ; Tetrachlorodibenzodioxin/*toxicity ; Tumor Necrosis Factor-alpha/*biosynthesis ; src-Family Kinases/antagonists & inhibitors/metabolism

PURPOSE: Growth inhibitory effects of lipid soluble components of the Korean red ginseng and the antineoplastic mechanism against human melanoma cell lines were investigated. To examine molecular mechanism of growth inhibitory effects of GX-PE, we analyzed the effect of GX-PE on cell cycle progression and expression of cell cycle regulatory factors such as retinoblastoma gene product (Rb), p27 (Kip1), p21 (WAF1), cdk2, cdk4 and cyclin D1 which are known to regulate cell cycle progression. MATERIALS AND METHODS: Petroleum ether extract of the Korean red ginseng (GX-PE) was added to cultures of three human melanoma cell lines, SK-MEL-1, SK-MEL-2, and SK-MEL-5. Proliferation was measured by 3H-thymidine incorporation assay. Cell cycle and expression of cell cycle regulatory factors were analyzed by flow cytometry and Western blotting, respectively. RESULTS: Growth of melanoma cells was inhibited by GX-PE in proportion to the concentration. GX-PE significantly inhibited cell cycle progression at G1 phase. GX-PE increased expression of negative cell cycle regulators, i.e., p27 (Kip1) in SK-MEL-2 and p21 (WAF1) and Rb in SK-MEL-1. CONCLUSION: These results suggest that GX-PE inhibits proliferation of melanoma cells at a G1-S transition point of the cell cycle. The effect of GX-PE is most likely due to induction of negative cell cycle regulatory factors.
Blotting, Western ; Cell Cycle* ; Cell Line* ; Cyclin D1 ; Ether ; Flow Cytometry ; G1 Phase ; Genes, Retinoblastoma ; Humans* ; Melanoma* ; Panax* ; Petroleum

15-deoxy-delta12,14-PGJ2(15d-PGJ2) is a natural ligand that activates the peroxisome proliferators-activated receptor (PPAR) gamma, a member of nuclear receptor family implicated in regulation of lipid metabolism and adipocyte differentiation. Recent studies have shown that 15d-PGJ2 is the potent anti-inflammatory agent functioning via PPARgamma-dependent and -independent mechanisms. Most postulated mechanisms for anti-inflammatory action of PPARgamma agonists are involved in inhibiting NF-kappaB signaling pathway. We examined the possibility that IL-6 signaling via the Jak-Stat pathway is modulated by 15d-PGJ2 in lymphocytes and also examined whether the inhibition of IL-6 signaling is dependent of PPARgamma. 15d-PGJ2 blocked IL-6 induced Stat1 and Stat3 activation in primary human lymphocytes, Jurkat cells and immortalized rheumatoid arthritis B cells. Inhibition of IL-6 signaling was induced rapidly within 15 min after treatment of 15d-PGJ2. Other PPARgamma-agonists, such as troglitazone and ciglitazone, did not inhibit IL-6 signaling, indicating that 15d-PGJ2 affect the IL-6-induced Jak-Stat signaling pathway via PPARgamma-independent mechanism. Although cycloheximide reversed 15d-PGJ2-mediated inhibition of Stat3 activation, actinomycin D had no effect on 15d-PGJ2-mediated inhibition of IL-6 signaling, indicating that inhibition of IL-6 signaling occur independent of de novo gene expression. These results show that 15d-PGJ2 specifically inhibit Jak-Stat signaling pathway in lymphocytes, and suggest that 15d-PGJ2 may regulate inflammatory reactions through the modulation of different signaling pathway other than NF-kappaB in lymphocytes.
Arthritis, Rheumatoid/metabolism/pathology ; Chromans/pharmacology ; Cycloheximide/pharmacology ; DNA-Binding Proteins/*metabolism ; Dactinomycin/pharmacology ; Gene Expression Regulation ; Humans ; Hypoglycemic Agents/pharmacology ; Interleukin-6/*pharmacology ; Jurkat Cells/metabolism/pathology ; Lymphocytes/cytology/*drug effects/*metabolism ; NF-kappa B/metabolism ; PPAR gamma/metabolism ; Phosphorylation ; Prostaglandin D2/*analogs & derivatives/pharmacology ; Protein Synthesis Inhibitors/pharmacology ; Research Support, Non-U.S. Gov't ; *Signal Transduction ; Thiazolidinediones/pharmacology ; Trans-Activators/*metabolism