Serum response factor is required for cell contact maintenance but dispensable for proliferation in visceral yolk sac endothelium

BACKGROUND: Endothelial-specific knockout of the transcription factor serum response factor (SRF) results in embryonic lethality by mid-gestation. The associated phenotype exhibits vascular failure in embryos as well as visceral yolk sac (VYS) tissues. Previous data suggest that this vascular failure is caused by alterations in cell-cell and cell-matrix contacts. In the current study, we sought to more carefully address the role of SRF in endothelial function and cell contact interactions in VYS tissues. RESULTS: Tie2-Cre recombinase-mediated knockout of SRF expression resulted in loss of detectable SRF from VYS mesoderm by E12.5. This loss was accompanied by decreased expression of smooth muscle alpha-actin as well as vascular endothelial cadherin and claudin 5, endothelial-specific components of adherens and tight junctions, respectively. Focal adhesion (FA) integrins alpha5 and beta1 were largely unchanged in contrast to loss of the FA-associated molecule vinculin. The integrin binding partner fibronectin-1 was also profoundly decreased in the extracellular matrix, indicating another aspect of impaired adhesive function and integrin signaling. Additionally, cells in SRF-null VYS mesoderm failed to reduce proliferation, suggesting not only that integrin-mediated contact inhibition is impaired but also that SRF protein is not required for proliferation in these cells. CONCLUSIONS: Our data support a model in which SRF is critical in maintaining functional cell-cell and cell-matrix adhesion in endothelial cells. Furthermore, we provide evidence that supports a model in which loss of SRF protein results in a sustained proliferation defect due in part to failed integrin signaling

Analysis of the Ex Vivo and In Vivo Antiretroviral Activity of Gemcitabine

Replication of retroviral and host genomes requires ribonucleotide reductase to convert rNTPs to dNTPs, which are then used as substrates for DNA synthesis. Inhibition of ribonucleotide reductase by hydroxyurea (HU) has been previously used to treat cancers as well as HIV. However, the use of HU as an antiretroviral is limited by its associated toxicities such as myelosuppression and hepatotoxicity. In this study, we examined the ribonucleotide reductase inhibitor, gemcitabine, both in cell culture and in C57Bl/6 mice infected with LP-BM5 murine leukemia virus (LP-BM5 MuLV, a murine AIDS model). Gemcitabine decreased infectivity of MuLV in cell culture with an EC50 in the low nanomolar range with no detectable cytotoxicity. Similarly, gemcitabine significantly decreased disease progression in mice infected with LP-BM5. Specifically, gemcitabine treatment decreased spleen size, plasma IgM, and provirus levels compared to LP-BM5 MuLV infected, untreated mice. Gemcitabine efficacy was observed at doses as low as 1 mg/kg/day in the absence of toxicity. Higher doses of gemcitabine (3 mg/kg/day and higher) were associated with toxicity as determined by a loss in body mass. In summary, our findings demonstrate that gemcitabine has antiretroviral activity ex vivo and in vivo in the LP-BM5 MuLV model. These observations together with a recent ex vivo study with HIV-1[1], suggest that gemcitabine has broad antiretroviral activity and could be particularly useful in vivo when used in combination drug therapy

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Endothelial-specific ablation of Serum Response Factor causes hemorrhaging, yolk sac vascular failure, and embryonic lethality-4

Sac tissues from mutant embryos showed disorganized vascular networks at E12.5 (B) that become more pronounced at E13.5 (D, F arrowheads). Blood pools (B, asterisks) were also observed in mutant yolk sac tissues.<p><b>Copyright information:</b></p><p>Taken from "Endothelial-specific ablation of Serum Response Factor causes hemorrhaging, yolk sac vascular failure, and embryonic lethality"</p><p>http://www.biomedcentral.com/1471-213X/8/65</p><p>BMC Developmental Biology 2008;8():65-65.</p><p>Published online 20 Jun 2008</p><p>PMCID:PMC2442838.</p><p></p

Endothelial-specific ablation of Serum Response Factor causes hemorrhaging, yolk sac vascular failure, and embryonic lethality-11

Os (B, D, F, H) exhibited internal hemorrhaging compared to wild-type littermates (A, C, E, G). This effect progressed with increasing gestational age. E14.5 mutant embryos were grossly necrotic and unsuitable for analysis. Labelled boxes in Fig. 1 correspond to images in Fig. 2.; Lv = liver. Arrowhead marks minimal hemorrhaging observed in E11.5 embryos.<p><b>Copyright information:</b></p><p>Taken from "Endothelial-specific ablation of Serum Response Factor causes hemorrhaging, yolk sac vascular failure, and embryonic lethality"</p><p>http://www.biomedcentral.com/1471-213X/8/65</p><p>BMC Developmental Biology 2008;8():65-65.</p><p>Published online 20 Jun 2008</p><p>PMCID:PMC2442838.</p><p></p