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Biomedical subjects

R N Ré

Publications and source records attributed to R N Ré.

14 recordsLinked to original sources
PubMed1996

Suppression of cellular proliferation using p53 DNA recognition site-related oligonucleotides.

A number of oligonucleotides were designed to bind through Hoogsteen triple helix or Watson-Crick hydrogen bonds to the p53 consensus sequence homology localized within the human nontranscribed rRNA spacer region. The oligomers, which bind in vitro to the consensus sequence homology, function as p53 analogues in cells deficient in wild-type p53 protein. Oligomers suppress proliferation of human colon cancer cells by three to eightfold, but only suppress proliferation of normal human mesangial cells or lung fibroblasts by less than 50%. On the basis of these studies, p53 analogues may be used therapeutically to selectively modify proliferation of transformed cells.

Amino Acid Sequence↗
PubMed1995

Distance constraints and stereospecific alignment requirements characteristic of p53 DNA-binding consensus sequence homologies.

We present evidence in favor of the position that some mutant p53 proteins retain the ability to trans-activate downstream genes through p53 DNA-binding consensus sequence (CS) homologies. We tested one cell line possessing high levels of mutant p53 and found that this mutant p53 is highly active in trans-activating one CS homology, moderately active in trans-activating a second sequence and inactive in modulating a third sequence. We tested a second cell line, also possessing high levels of mutant p53 and found the same pattern of activation. In addition we find that inter-motif distance [represented by N in RRRCWWGYYY(N)RRRCWWGYYY] is very important in determining the relative binding affinity of a given CS homology for wild-type or mutant p53. Our studies suggest that stereospecific alignment of the DNA-binding motifs within the CS may favor binding of wild-type p53 while misalignment may favor binding of mutant p53. Furthermore, we find that the maximum distances at which p53 DNA-binding CS homologies are functionally active vary for different sequences. Introduction of as few as 200 bp between one CS homology and the downstream TATA box can eliminate a 45-fold p53-mediated transactivation. We present evidence that the composition of the DNA which flanks a p53 DNA-binding consensus sequence may also modulate trans-activation.

Amino Acid Sequence↗
PubMed1995

Retroviral suicide vector does not inhibit neointimal growth in a porcine coronary model of restenosis.

We attempted to attenuate neointimal formation following vascular injury using a retroviral suicide vector. Epicardial coronary arteries of adult miniature swine were injured by deployment of oversized tantalum stents. One week later, injured segments were exposed to packaged retroviral constructs with or without the herpes simplex virus thymidine kinase gene. Ganciclovir treatments were initiated 48 hours later in both control and experimental swine and continued for two weeks. Four weeks following vascular injury, both control and experimental arteries were harvested and histologically prepared for image analysis. Despite adequate marker gene expression, there was no significant difference in the neointimal area or neointimal/media ratio between control and experimental groups. While the HSVtk ganciclovir system attenuates cell proliferation in other systems, retroviral vector targeting of vascular smooth muscle cells for elimination may be too inefficient to prevent restenosis following angioplasty.

Animals↗
PubMed1995

Identification and antisense inhibition of a renin-angiotensin system in transgenic cardiomyocytes.

Cardiac myocytes (AT-1 cells) derived from heart tumors of mice transgenic for an atrial natriuretic factor promoter, SV40 large T-antigen DNA transgene, demonstrate properties consistent with normal cardiac myocytes but retain the capacity to proliferate in culture. We studied the renin-angiotensin system (RAS) and related growth regulation of these cells because AT-1 cells (or transgenically similar cells) may be useful to repair injured myocardium. This study reveals two separate and distinct findings: 1) AT-1 cells proliferate or hypertrophy in response to angiotensin II (ANG II), depending on their competence to proceed through the cell cycle; and 2) AT-1 cells possess components of a RAS, and angiotensinogen antisense experiments suggest that the RAS is functional in these cells. Specifically, AT-1 cells proliferate in response to ANG II in low-serum medium but hypertrophy in response to ANG II when first treated with mitomycin C (at a concentration that inhibits DNA replication but is not cytotoxic). The ANG II-mediated proliferative and hypertrophic responses are inhibited by DuP 753. In addition, there is a significant increase in the protein-to-DNA ratio of cells, which are proliferation-inhibited in the absence of ANG II treatment (20%, P < 0.05). DuP 753 also inhibits this hypertrophy, suggesting that these cells possess a functional RAS. AT-1 cells contain mRNAs for angiotensin-converting enzyme, renin, angiotensinogen, and the AT1 receptor as determined by sequence analysis of polymerase chain reaction amplification products. Antisense oligonucleotides complementary to the angiotensinogen mRNA specifically inhibit angiotensinogen mRNA accumulation and proliferation of AT-1 cells. In summary, these cells contain a growth-regulating RAS, suggesting that such a system may play a significant role in left ventricular hypertrophy.

Angiotensin II↗
PubMed1994

Sea change.

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Animals↗
PubMed1994

Hemodynamics in transgenic mice with overexpression of atrial natriuretic factor.

The circulatory effects associated with lifelong plasma atrial natriuretic factor (ANF) elevation were examined by generating transgenic mice, which constitutively express a fusion gene consisting of the transthyretin promoter and the ANF structural gene. These mice have chronically elevated ANF levels as compared with their nontransgenic siblings. Transgenic animals exhibited immunoreactive ANF levels that were nearly fivefold higher than those measured in nontransgenic littermates. Systemic and regional hemodynamics and blood volumes were explored by using modifications of the reference microsphere and dilution techniques. Mean arterial pressure was reduced by 24 mm Hg, associated with a 27% reduction in total heart weight. This chronic reduction in blood pressure was due to a 21% reduction in total peripheral resistance, whereas cardiac output, stroke volume, and heart rate were not significantly altered, despite a 15% elevation in plasma volume. Transgenic mice displayed reductions of 35%, 33%, 32%, and 19% in muscle, skin, brain, and renal vascular resistance, respectively, whereas coronary and splanchnic resistances were not significantly altered. The findings complement earlier data from chronically infused normotensive mammals and suggest that these mice are an excellent model for investigating the effects of lifelong ANF elevation.

Animals↗
PubMed1993

Prior arterial injury enhances luciferase expression following in vivo gene transfer.

We determined the time course of gene expression following DNA/Lipofectin transfection of normal or previously injured arterial segments using direct intraluminal infusion following surgical exposure. Constructs possessing the firefly luciferase cDNA regulated by Simian virus 40, Rous sarcoma virus, or alpha-actin promoter were incubated together with Lipofectin for 30 minutes. Arterial segments were assayed for luciferase activity following harvest at 2-21 days. Without prior injury, luciferase activity was only 2.5-fold greater than background two days following gene transfer. Arterial injury three days before gene transfer resulted in luciferase activity 12.5-fold over background levels. This observation has clinical implications with regard to gene therapy following angioplasty, a procedure that is associated with endothelial cell denudation and smooth muscle cell proliferation. Maintenance of gene expression for several days could ameliorate the early smooth muscle migration and proliferation following arterial injury.

Actins↗
PubMed1992

Microsphere and dilution techniques for the determination of blood flows and volumes in conscious mice.

Although the mouse is the most commonly used transgenic species, little is known regarding cardiovascular and fluid homeostasis in this animal. Therefore, the reference microsphere and dilution techniques were adapted for the measurement of cardiac output (CO), regional blood flows, and intravascular fluid volumes in the conscious mouse. Previously acclimatized C3H mice were studied 4-5 h after surgery and recovery from anesthesia. Approximately 40,000 85Sr-labeled microspheres were injected into the left ventricle while a reference sample was withdrawn at one of two rates from the femoral artery. 51Cr and 125I were used for the determination of blood volume (BV), plasma volume (PV), and Fcells ratio (whole body hematocrit/large vessel hematocrit). CO and BV in the conscious mouse were 16 +/- 1.4 ml/min and 2.3 +/- 0.1 ml, respectively. Anesthesia lowered heart rate, blood pressure, PV, and altered the distribution of CO. Two successive injections of 15,000-20,000 microspheres were tolerated in the mouse without an increase in total peripheral resistance. The results indicate that the microsphere and indicator dilution techniques can be applied to study cardiovascular and fluid homeostasis in the mouse.

Animals↗
PubMedDate not supplied

The interaction of insulin and angiotensin II on the regulation of human neuroblastoma cell growth.

This laboratory has previously reported that angiotensin II is a growth factor for human SH-SY5Y neuroblastoma cells, and that a variety of converting enzyme inhibitors and angiotensin II antagonists reduce thymidine incorporation into the DNA of these cells. In the present study, insulin, at 5 micrograms/mL, was found to stimulate thymidine incorporation in SH-SY5Y cells. The insulin effect was only partially inhibited by the converting enzyme inhibitors enalapril, quinapril, and quinaprilat, whereas it was markedly or totally blunted by the angiotensin II antagonists DuP753 and PD123177. In additional studies, IGF-1 (50 ng/mL) significantly stimulated thymidine incorporation into these cells in a fashion indistinguishable from that of insulin. Taken together, these studies are consistent with the suggestion that insulin at high concentrations and IGF at low concentrations enhance the proliferative response of these cells to angiotensin II. The differential effects of converting enzyme inhibition and angiotensin II antagonism on cell proliferation could be explained if converting enzyme inhibition results in low, but effective, levels of angiotensin II in the culture medium, whereas the angiotensin II antagonists effectively block angiotensin II at its receptor. Finally, in this system, both the AT1 receptor blocking agent DuP 753 and the AT2 receptor blocking agent PD123177 appear to be effective.

Angiotensin II↗