Gene transfer into vascular cells

AbstractThe goal of gene therapy is to introduce foreign deoxyribonucleic acid (DNA) into somatic cells to correct or prevent disorders caused by the malfunction of genes within a diseased individual. Overexpression of recombinant genes at specific sites within the vasculature can provide insights into vascular biology and potential treatments for various cardiovascular disorders such as restenosis. Methods for the introduction of foreign DNA into endothelial and vascular smooth muscle cells have been developed recently. These include the genetic modification of endothelium in vitro and implantation in vivo on arterial segments, direct infection of the arterial wall in vivo with a replication-defective retroviral vector expressing a recombinant gene and direct transfer of genes into vascular cells in vivo with use of liposomes. Although still in its formative stages, gene transfer into the vasculature holds promise as a potential treatment for vascular diseases, including atherosclerosis and restenosis. This approach may also provide insight into the role of specific gene products in the development of pathologic lesions

Adoptive immunotherapy of cancer with polyclonal, 10(8)-fold hyperexpanded, CD4(+ )and CD8(+ )T cells

T cell-mediated cancer immunotherapy is dose dependent and optimally requires participation of antigen-specific CD4(+ )and CD8(+ )T cells. Here, we isolated tumor-sensitized T cells and activated them in vitro using conditions that led to greater than 10(8)-fold numerical hyperexpansion of either the CD4(+ )or CD8(+ )subset while retaining their capacity for in vivo therapeutic efficacy. Murine tumor-draining lymph node (TDLN) cells were segregated to purify the CD62L(low )subset, or the CD4(+ )subset thereof. Cells were then propagated through multiple cycles of anti-CD3 activation with IL-2 + IL-7 for the CD8(+ )subset, or IL-7 + IL-23 for the CD4(+ )subset. A broad repertoire of TCR Vβ families was maintained throughout hyperexpansion, which was similar to the starting population. Adoptive transfer of hyper-expanded CD8(+ )T cells eliminated established pulmonary metastases, in an immunologically specific fashion without the requirement for adjunct IL-2. Hyper-expanded CD4(+ )T cells cured established tumors in intracranial or subcutaneous sites that were not susceptible to CD8(+ )T cells alone. Because accessibility and antigen presentation within metastases varies according to anatomic site, maintenance of a broad repertoire of both CD4(+ )and CD8(+ )T effector cells will augment the overall systemic efficacy of adoptive immunotherapy

Safety and Short-Term Toxicity of a Novel Cationic Lipid Formulation for Human Gene Therapy

Overview summary Although several viral vectors have been widely applied to the treatment of human disease, the development of nonviral vectors is still in their infancy. In this report, a novel cationic lipid, DMRIE/DOPE, has been incorporated into the DNA–liposome formulation that improves transfection efficiencies and allows up to 1,000-fold higher concentrations of DNA to be administered in vivo. In this paper, the safety and toxicity of this formulation is described in two species, mice and pigs, suggesting that it may prove useful for human gene therapy.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/63224/1/hum.1993.4.6-781.pd

REVEILLE8 and PSEUDO-REPONSE REGULATOR5 Form a Negative Feedback Loop within the Arabidopsis Circadian Clock

Circadian rhythms provide organisms with an adaptive advantage, allowing them to regulate physiological and developmental events so that they occur at the most appropriate time of day. In plants, as in other eukaryotes, multiple transcriptional feedback loops are central to clock function. In one such feedback loop, the Myb-like transcription factors CCA1 and LHY directly repress expression of the pseudoresponse regulator TOC1 by binding to an evening element (EE) in the TOC1 promoter. Another key regulatory circuit involves CCA1 and LHY and the TOC1 homologs PRR5, PRR7, and PRR9. Purification of EE–binding proteins from plant extracts followed by mass spectrometry led to the identification of RVE8, a homolog of CCA1 and LHY. Similar to these well-known clock genes, expression of RVE8 is circadian-regulated with a dawn phase of expression, and RVE8 binds specifically to the EE. However, whereas cca1 and lhy mutants have short period phenotypes and overexpression of either gene causes arrhythmia, rve8 mutants have long-period and RVE8-OX plants have short-period phenotypes. Light input to the clock is normal in rve8, but temperature compensation (a hallmark of circadian rhythms) is perturbed. RVE8 binds to the promoters of both TOC1 and PRR5 in the subjective afternoon, but surprisingly only PRR5 expression is perturbed by overexpression of RVE8. Together, our data indicate that RVE8 promotes expression of a subset of EE–containing clock genes towards the end of the subjective day and forms a negative feedback loop with PRR5. Thus RVE8 and its homologs CCA1 and LHY function close to the circadian oscillator but act via distinct molecular mechanisms

Validation of the Block Walk Method for Assessing Physical Activity occurring on Sidewalks/Streets

The block walk method (BWM) is one of the more common approaches for assessing physical activity (PA) performed on sidewalks/streets; however, it is non-technical, labor-intensive, and lacks validation. This study aimed to validate the BWM and examine the potential for using a wearable video device (WVD) to assess PA occurring on sidewalks/streets. Trained observers (one wearing and one not wearing the WVD) walked together and performed the BWM according to a previously developed protocol along routes in low, medium, and high walkable areas. Two experts then reviewed the videos. A total of 1150 (traditional) and 1087 (video review) individuals were observed during 900 min of observation. When larger numbers of individuals were observed, the traditional method overestimated the overall number of people as well as those walking and sitting/standing, while underestimating the number of runners. Valid estimates of PA occurring on sidewalks/streets can be obtained by the traditional BWM in low and medium walkability areas and/or with non-common activities (cycling); however, its validity is questionable when sidewalks/streets use volume is high. The use of WVDs in PA assessment has the potential to establish new levels of accuracy, reduce resource requirements, and open up the possibility for retrospective analysis

Recombinant Growth Factor Gene Expression in Vascular Cells in Vivo

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72999/1/j.1749-6632.1994.tb12050.x.pd