Rapid analysis of proteomic biomarkers expressed in human endometrial stromal cells during decidualization.

Decidualization of human endometrial stromal (ES) cells plays a critical role in successful uterine implantation. Therefore, monitoring of the behavior of human ES cells may provide the clue for early detection of a uterine abnormality such as sterility and abortion. Monitoring of decidualization in vitro cell culture system fundamentally depends on expression of the definite biomarkers. In this study, we tried to uncover novel marker proteins of 8-bromo-cyclic adenosine monophosphate (8-Br-cAMP)-induced decidualization in human ES cells using the surface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS). Samples were divided into three groups; control human ES cells (n=7), ES cells treated with 8-Br-cAMP (n=7 per each treatment, treated for 3, 6, 9, or 12 days), and cells from which 8-Br-cAMP was withdrawn for 3 days (n=7) or 6 days (n=7) after 8-Br-cAMP treatment for 6 days. Differential expressions between non-decidual control cells and 8-Br-cAMP-induced decidual cells were observed in the peaks of 9787.058 Da, 10115.45 Da, and 24031.25 Da, detected by H4 ProteinChip, and in the peaks of 10833.08 Da, 22440.88 Da, and 32777.38 Da, detected by CM10 ProteinChip. The expression patterns of these decidual markers are expected to provide invaluable information in monitoring cellular development, and further identification of these proteins may hopefully offer precious means for clinical research and therapeutic purposes.

Cloning and expression of alpha class glutathione S-transferase gene from the small hermaphroditic fish Rivulus marmoratus (Cyprinodontiformes, Rivulidae).

In order to assess its potential as a biomarker of aquatic pollution, an alpha class glutathione S-transferase gene (GSTalpha gene) was cloned from the small hermaphroditic fish Rivulus marmoratus. The R. marmoratus GSTalpha gene spanned 1.3 kb, consisting of 6 exons encoding 221 amino acid residues. It showed high similarity to zebrafish GST. We named this R. marmoratus GSTalpha gene as rm-GSTalpha. The cDNA of the rm-GSTalpha gene was also investigated for its phylogeny, tissue-specific and chemical-induced expression. Rm-GSTalpha was subcloned into a 6 x His-tagged pCRT7 TOPO TA expression vector to produce the recombinant 6 x His-tagged rm-GST protein. This will be used in future to raise an rm-GSTalpha antibody for use in the study of phase II metabolism involved in detoxification. We also exposed R. marmoratus to 300 microg/l of 4-nonylphenol in water, and found approximately 4-fold induction of R. marmoratus GSTalpha mRNA in the treated animals. In this paper, we discuss the characteristics of the R. marmoratus GSTalpha gene as well as its potential use in relation to environmental pollution.

Genomic cloning and expression of vitellogenin gene from the self-fertilizing fish Rivulus marmoratus (Cyprinodontiformes, Rivulidae).

We cloned the vitellogenin gene from the self-fertilizing fish Rivulus marmoratus, and sequenced 12,326 bp. The number of exons of R. marmoratus and rainbow trout vitellogenin genes were different, and also the splicing junctions are different throughout most of the exons and introns but the amino acid similarity of R. marmoratus vitellogenin gene to other species was rather high. In promoter region of R. marmoratus vitellogenin gene, there were several E2 binding sites and the estrogen response element (ERE). We discuss here the gene structure and expression of R. marmoratus vitellogenin gene.

Comprehensive transcriptome analysis of differentiation of embryonic stem cells into midbrain and hindbrain neurons.

Neurogenesis is one of the most complex events in embryonic development. However, little information is available regarding the molecular events that occur during neurogenesis. To identify regulatory genes and underlying mechanisms involved in the differentiation of embryonic stem (ES) cells to neurons, gene expression profiling was performed using cDNA microarrays. In mouse ES cells, we compared the gene expression of each differentiated cell stage using a five-stage lineage selection method. Of 10,368 genes, 1633 (16%) known regulatory genes were differentially expressed at least 2-fold or greater at one or more stages. At stage 3, during which ES cells differentiate into neural stem cells, modulation of nearly 1000 genes was observed. Most of transcription factors (Otx2, Ebf-3, Ptx3, Sox4, 13, 18, engrailed, Irx2, Pax8, and Lim3), signaling molecules (Wnt, TGF, and Shh family members), and extracellular matrix/adhesion molecules (collagens, MAPs, and NCAM) were up-regulated. However, some genes which may play important roles in maintaining the pluripotency of ES cells (Kruppel-like factor 2, 4, 5, 9, myeloblast oncogene like2, ZFP 57, and Esg-1) were down-regulated. The many genes identified with this approach that are modulated during neurogenesis will facilitate studies of the mechanisms underlying ES cell differentiation, neural induction, and neurogenesis.

Teratogenic effects of N-ethyl-N-nitrosourea (ENU) on larvae of the self-fertilizing fish Rivulus marmoratus (Cyprinodontiformes, Rivulidae).

We showed that N-Ethyl-N-Nitrosourea (ENU) induces teratogenesis in larvae of the self-fertilizing fish Rivulus marmoratus. We discuss this and the issue of carcinogenesis caused by ENU.