Extensive genetic diversity and substructuring among zebrafish strains revealed through copy number variant analysis.

Copy number variants (CNVs) represent a substantial source of genomic variation in vertebrates and have been associated with numerous human diseases. Despite this, the extent of CNVs in the zebrafish, an important model for human disease, remains unknown. Using 80 zebrafish genomes, representing three commonly used laboratory strains and one native population, we constructed a genome-wide, high-resolution CNV map for the zebrafish comprising 6,080 CNV elements and encompassing 14.6% of the zebrafish reference genome. This amount of copy number variation is four times that previously observed in other vertebrates, including humans. Moreover, 69% of the CNV elements exhibited strain specificity, with the highest number observed for Tubingen. This variation likely arose, in part, from Tubingen's large founding size and composite population origin. Additional population genetic studies also provided important insight into the origins and substructure of these commonly used laboratory strains. This extensive variation among and within zebrafish strains may have functional effects that impact phenotype and, if not properly addressed, such extensive levels of germ-line variation and population substructure in this commonly used model organism can potentially confound studies intended for translation to human diseases.

Characterization of the gene structure, functional significance, and clinical application of RNF180, a novel gene in gastric cancer.

BACKGROUND: By using genome-wide methylation screening, the authors identified ring finger protein 180 (RNF180) as preferentially methylated in cancer. This study was undertaken to clarify its structure and functional role in gastric cancer. METHODS: The transcription start site and core functional promoter region of RNF180 were revealed by 5' rapid amplification of cDNA ends and luciferase activity assays. Promoter methylation was detected by combined bisulfite restriction analysis and bisulfite genomic sequencing. Cell growth was detected by colony formation assay, apoptosis by annexin V assay, and RNF180 target genes by cDNA microarray. RESULTS: The authors revealed the transcription start site of RNF180 gene and identified the functional core promoter region (-202/+372) in the CpG island, which could be silenced by in vitro methylation assay. RNF180 was silenced in 6 of 7 gastric cancer cell lines and significantly down-regulated in primary gastric cancers compared with adjacent normal tissues (P = .001). Loss of gene expression was associated with promoter methylation. Re-expression of RNF180 suppressed cell growth (P < .001) and induced apoptosis (P < .05), which were mediated by up-regulating the antiproliferation regulators MTSS1 and CDKN2A and the proapoptotic mediator TIMP3. Promoter methylation of RNF180 was detected in 76% (150 of 198) of primary gastric cancers and 55% (11 of 20) of intestinal metaplasia, but in none of 23 normal gastric tissues. Methylated RNF180 DNA was detected in the plasma of 56% of gastric cancer patients, but not in healthy controls (P = .003). Patients with low or loss of RNF180 expression had significantly poorer overall survival. CONCLUSIONS: RNF180 is a novel potential tumor suppressor in gastric carcinogenesis and has potential clinical utility as a biomarker for gastric cancer patients.

NF-κB targets miR-16 and miR-21 in gastric cancer: involvement of prostaglandin E receptors.

Cigarette smoke is one of the risk factors for gastric cancer and nicotine has been reported to promote tumor growth. Deregulation of microRNA (miRNA) and cyclooxygenase-2 (COX-2) expressions are hallmarks of many cancers including gastric cancer. Here, we used an miRNA array platform covering a panel of 95 human miRNAs to examine the expression profile in nicotine-treated gastric cancer cells. We found that miR-16 and miR-21 were upregulated upon nicotine stimulation, transfection with anti-miR-16 or anti-miR-21 significantly abrogated cell proliferation. In contrast, ectopic miR-16 or miR-21 expression exhibited a similar stimulatory effect on cell proliferation as nicotine. Nicotine-mediated IkappaBα degradation and nuclear factor-kappa B (NF-κB) translocation dose-dependently. Knockdown of NF-κB by short interfering RNA (siRNA) or specific inhibitor (Bay-11-7085) markedly suppressed nicotine-induced cell proliferation and upregulation of miR-16 and miR-21. Interestingly, NF-κB-binding sites were located in both miR-16 and miR-21 gene transcriptional elements and we showed that nicotine enhanced the binding of NF-κB to the promoters of miR-16 and miR-21. Furthermore, activation of COX-2/prostaglandin E2 (PGE2) signaling in response to nicotine was mediated by the action of prostaglandin E receptors (EP2 and EP4). EP2 or EP4 siRNA or antagonists impaired the nicotine-mediated NF-κB activity, upregulation of miR-16 and miR-21 and cell proliferation. Taken together, these results suggest that miR-16 and miR-21 are directly regulated by the transcription factor NF-κB and yet nicotine-promoted cell proliferation is mediated via EP2/4 receptors. Perhaps this study may shed light on the development of anticancer drugs to improve the chemosensitivity in smokers.

Disseminated peritoneal leiomyomatosis after laparoscopic supracervical hysterectomy with characteristic molecular cytogenetic findings of uterine leiomyoma.

Disseminated peritoneal leiomyomatosis (DPL) is a rare condition characterized by scattered smooth muscle nodules over the peritoneal surfaces. The pathogenesis of DPL remains unclear. Herein, we report a case of DPL occurring 7 years after laparoscopic supracervical hysterectomy with morcellation for uterine leiomyomata (UL). We analyzed both the original UL and the subsequent DPL by molecular cytogenetics to assess the role of chromosomal abnormalities in DPL pathobiology. Interestingly, all of the chromosomal aberrations detected in this case of DPL, including r(1)(p34.3q41), del(3)(q23q26.33), and t(12;14)(q14.3;q24.1), are characteristic chromosomal abnormalities detected in UL. Fluorescence in situ hybridization analysis of the initial UL confirmed an interstitial deletion spanning at least 3q24 and 3q25.1, suggesting that functional alteration of a potential gene in this chromosomal region may play a role in DPL development from UL. With the increasing rate of hysterectomy through laparoscopic approach to UL, the unique complications of laparoscopy with morcellation, especially seeding and proliferation of tumor cells over abdominal organs and peritoneum, are becoming more significant and may necessitate review of current surgical protocols to prevent future seeding of the pelvic region with tumor particles.

Transgenic cyclooxygenase-2 expression and high salt enhanced susceptibility to chemical-induced gastric cancer development in mice.

Cyclooxoygenase (COX)-2 overexpression is involved in gastric carcinogenesis. While high-salt intake is a known risk factor for gastric cancer development, we determined the effects of high salt on gastric chemical carcinogenesis in COX-2 transgenic (TG) mice. COX-2 TG mice were developed in C57/BL6 strain using the full-length human cox-2 complementary DNA construct. Six-week-old COX-2 TG and wild-type (WT) littermates were randomly allocated to receive alternate week of N-methyl-N-nitrosourea (MNU, 240 p.p.m.) in drinking water or control for 10 weeks. Two groups of mice were further treated with 10% NaCl during the initial 10 weeks. All mice were killed at the end of week 50. Both forced COX-2 overexpression and high-salt intake significantly increased the frequency of gastric cancer development in mice as compared with WT littermates treated with MNU alone. However, no additive effect was observed on the combination of high salt and COX-2 expression. We further showed that MNU and high-salt treatment increased chronic inflammatory infiltrates and induced prostaglandin E(2) (PGE(2)) production in the non-cancerous stomach. Whereas high-salt treatment markedly increased the expression of inflammatory cytokines (tumor necrosis factor-alpha, interferon-gamma, interleukin (IL)-1 beta and IL-6) in the gastric mucosa, COX-2 overexpression significantly altered the cell kinetics in the MNU-induced gastric cancer model. In conclusion, both high salt and COX-2 overexpression promote chemical-induced gastric carcinogenesis, possibly related to chronic inflammation, induction of PGE(2), disruption of cell kinetics and induction of inflammatory cytokines.