IL-18, produced as a biologically inactive precursor, is processed by caspase-1 in LPS-activated macrophages. Here, we investigated caspase-1-independent processing of IL-18 in Fas ligand (FasL)-stimulated macrophages and its involvement in liver injury. Administration of Propionibacterium acnes (P. acnes) upregulated functional Fas expression on macrophages in an IFNgamma-dependent manner, and these macrophages became competent to secrete mature IL-18 upon stimulation with FasL. This was also the case for caspase-1-deficient mice. Administration of recombinant soluble FasL (rFasL) after P. acnes priming induced comparable elevation of serum IL-18 in parallel with elevated serum liver enzyme levels. However, liver injury was not induced in IL-18-deficient mice after rFasL administration. These results indicate a caspase-1-independent pathway of IL-18 secretion from FasL-stimulated macrophages and its critical involvement in FasL-induced liver injury.

Preimplantation development is marked by four major events: the transition of maternal transcripts to zygotic transcripts, compaction, the first lineage differentiation into inner cell mass and trophectoderm, and implantation. The scarcity of the materials of preimplantation embryos, both in size (diameter < 100 microm) and in quantity (only a few to tens of oocytes from each ovulation), has hampered molecular analysis of preimplantation embryos. Recent progress in RNA amplification methods and microarray platforms, including genes unique to preimplantation embryos, allow us to apply global gene expression profiling to the study of preimplantation embryos. Our gene expression profiling during preimplantation development revealed the distinctive patterns of maternal RNA degradation and embryonic gene activation, including two major transient waves of de novo transcription. The first wave corresponds to zygotic genome activation (ZGA). The second wave, mid-preimplantation gene activation (MGA), contributes dramatic morphological changes during late preimplantation development. Further expression profiling of embryos treated with inhibitors of transcription or translation revealed that the translation of maternal RNA is required for the initiation of ZGA, suggesting a cascade of gene activation from maternal RNA/protein sets to ZGA gene sets and thence to MGA gene sets. To date, several reports of microarray experiments using mouse and human preimplantation embryos have been published. The identification of a large number of genes and multiple signaling pathways involved at each developmental stage by such global gene expression profiling accelerates understanding of molecular mechanisms underlining totipotency/pluripotency and programs of early mammalian development.

Hepatoma-derived growth factor (HDGF) is highly expressed in tumor cells, and stimulates their proliferation. In the present study, we investigated the role of HDGF in tumorigenesis and elucidated the mechanism of action. Stable transfectants of NIH3T3 cells overexpressing HDGF did not show significant anchorage-independent growth in soft agar assay. However, these stable transfectants overexpressing HDGF generated sarcomatous tumors in nude mice. These tumors were red-colored macroscopically, and histologically showed a rich vascularity. Immunohistochemical analysis using CD31 antibody showed new vessel formation. Recombinant HDGF stimulated proliferation of human umbilical vein endothelial cells in a dose-dependent manner, and stimulated tubule formation. ncogenesis is induced by genetic alterations of various proteins involved in the cell cycle and cellular proliferation, especially by up-regulation of oncogenic genes and/or down-regulation of tumor suppressor genes. Overexpression of growth factors, receptors for growth factors, proteins related to signal transduction from the respective receptors and positive transcriptional regulatory factors, and suppressed expression of growth-inhibitory genes, including negative signal transducers and transcriptional regulatory factors, have all been associated with tumor formation in nude mice. [1][2][3][4][5][6][7][8][9] Hepatoma-derived growth factor (HDGF) is a heparin-binding protein purified from the conditioned media of Huh-7 hepatoma cells, which proliferate autonomously in a serum-free chemically defined medium.10, 11) HDGF contains a well-conserved N-terminal amino acid sequence, which is called the hath (homologous to amino terminus of HDGF) region. 11,12) HDGF is translocated to the nucleus via nuclear localization signals and its nuclear translocation is essential for induction of cell growth activity. 13,14) It is highly expressed in fetal tissues and may be involved in development of organs, including the cardiovascular system, kidney and liver. [15][16][17][18] HDGF has mitogenic activity for some hepatocellular carcinoma (HCC) cells, in addition to fibroblasts, endothelial cells, vascular smooth muscle cells and fetal hepatocytes. 10,11,13,15,16,18,19) It is abundantly expressed in tumor cell lines, and HDGF antisense oligonucleotides suppress the proliferation of hepatoma cells expressing HDGF endogenously. 19) In human and murine models of HCC, HDGF expression increases at an early stage, even before tumor formation. Furthermore, HDGF is more abundantly expressed in HCC than in adjacent non-tumor liver tissues.20) HDGF is a unique nuclear/growth factor, possibly playing an important role in the development and progression of cancer cells.Increasing evidence suggests that HDGF might be implicated in tumorigenesis. However, whether HDGF really induces transformation of cells and tumor formation in vivo is not known. Therefore, we established stable cell lines overexpressing HDGF from NIH3T3 fibroblasts by transfection, and investigated their oncogenic p...

The human cholinesterase (ChE) gene from a patient with acholinesterasemia was cloned and analyzed. By using ChE cDNA as a probe, four independent clones were isolated from a genomic library constructed from the patient's DNA. Sequencing analysis ofall of the four clones revealed that exon 2 of the ChE gene was disrupted by a 342-base-pair (bp) insertion of Alu element, including a poly(A) tract of 38 bp, which showed 93% sequence homology with a current type of human Alu consensus sequence. Southern blot analysis showed that the Alu insertion occurred in both alleles of the patient and was inherited in the patient's family. This Alu insertion was flanked by 15-bp of target site duplication in exon 2 corresponding to positions 1062-1076 of ChE cDNA, indicating that an Alu element could have been integrated by retrotransposition. Thus, this case provides an important clue to the mechanism of-inactivation of a gene by integration of a retrotransposon.

Cyclooxygenase 2 (COX-2) has been suggested to be associated with liver carcinogenesis. Several reports have shown that NSAIDs inhibit the growth of hepatocellular carcinoma cell lines. There is little evidence of how COX-2 inhibitors regulate the proliferation of hepatocellular carcinoma cells or the mechanism involved. In our study, we investigated the growth-inhibitory mechanism of a selective COX-2 inhibitor, NS-398, in 4 hepatocellular carcinoma cell lines by studying cell growth, COX-2 and proliferating cell nuclear antigen (PCNA) expression, cell cycle distribution and the evidence of apoptosis. NS-398 inhibited the growth of all 4 cell lines in a time-and dose-dependent manner and the inhibitory effects were independent of the level of COX-2 protein expression. PCNA expression was downregulated by NS-398 in a dose-independent manner. NS-398 caused cell cycle arrest in the S phase with a reduction in cell numbers and cell accumulation in the G0/G1 phase, for all 4 cell lines. No evidence of apoptosis was observed in our present study. Key words: selective COX-2 inhibitor; cell growth; cell cycle; hepatocellular carcinoma cellsCyclooxygenases (COX) are key rate-limiting enzymes involved in the conversion of arachidonic acid to prostaglandin H2, the precursor of various compounds including prostaglandins (PGs), prostacyclin and thromboxanes. There are at least 2 isoforms of COX: COX-1 and COX-2. COX-1 is expressed constitutively in a wide variety of tissues whereas COX-2 is highly inducible and is expressed in response to a variety of proinflammatory agents and cytokines. 1,2 Overexpression of COX-2 has been demonstrated in various tumor tissues such as colon cancer, pancreatic cancer and hepatocellular carcinoma. [3][4][5][6][7] Although several studies have shown the upregulation of COX-2 in cirrhotic tissues adjacent to hepatocellular carcinoma (HCC) and welldifferentiated HCC, the precise role of COX-2 in carcinogenesis remains unclear. 7,8 A few reports have shown that NSAIDs inhibited the growth of various cancer cell lines including those derived from hepatocellular carcinoma. 9 -11 The question arises whether COX-2 also is a target for the prevention or treatment of hepatocellular carcinoma, as it is for colon cancer. 12 There is little evidence, however, of how COX-2 inhibitors regulate the proliferation of hepatocellular carcinoma cells or the mechanism involved. In our study, we investigated the growth-inhibitory mechanism of a selective COX-2 inhibitor, NS-398, in hepatocellular carcinoma cell lines. MATERIAL AND METHODS Cell lines and cell cultureFour human hepatocellular carcinoma cell lines, PLC/PRF5, HepG2, Mahlavu and HuH-7, were obtained from the American Type Culture Collection. Cells (2 ϫ 10 5 cells) were grown for 24 hr in DMEM (PLC/PRF5 and HepG2) (ICN; Biomedicals Inc., Aurora, MI), minimum essential Eagle's medium (Mahlavu) (ICN) and RPMI 1640 (HuH-7) (ICN) supplemented with 10% FBS at 37°C under 5% CO 2 /95% air in 100 mm 2 cell culture dishes. Then, NS-398 (Cayman Chemical, Ann Ar...