Biomedical subjects

D S Ettenson

Publications and source records attributed to D S Ettenson.

8 recordsLinked to original sources

PubMed2000

Endothelial heparan sulfate is necessary but not sufficient for control of vascular smooth muscle cell growth.

The state of the endothelial cell (EC) determines the nature of its control of vascular smooth muscle cell (vSMC) biology. Conditioned medium from postconfluent ECs inhibits vSMC proliferation, whereas subconfluent conditioned medium from the same ECs has a stimulatory effect. We and others have identified confluent endothelial cells' production of heparan sulfate proteoglycans (HSPG) as critical to vSMC growth control. The question that arises is whether the stimulation that is observed with subconfluent cells is from (1) aberrant HSPG production, (2) elaboration of noninhibitory species of HSPG, or (3) production of other factors, such as mitogens, which counteract the inhibitory HSPG to stimulate vSMCs. We studied the relative effects of conditioned medium produced by both subconfluent and postconfluent EC cultures on vSMC growth. Conditioned medium was fractionated into nonproteoglycan (non-PG) and proteoglycan (PG) components by anion-exchange chromatography. The PG fractionation profile and the antiproliferative activity of the HSPGs isolated from both subconfluent and postconfluent EC-conditioned media were similar. However, the HSPG fraction alone could not approach the inhibitory potential of unfractionated conditioned medium from postconfluent EC cultures. Non-PG proteins produced by the endothelial cultures had no effect on vSMC growth on their own. Yet, when they were mixed together with HSPG fractions, from either subconfluent or postconfluent EC cultures, the full growth effects were returned. Non-PG protein fractions from postconfluent cultures with HSPG fractions gave maximal inhibition of vSMC growth, whereas non-PG protein fractions from subconfluent EC cultures with HSPG fractions produced the maximal stimulation. Thus, whereas the net stimulatory or inhibitory effect on vSMC growth of EC-conditioned medium is density dependent, this effect does not result from a difference in the antiproliferative heparan sulfate component but rather from non-PG proteins that interact with the heparan sulfates.

Animals↗

PubMed2000

Local drug delivery: an emerging approach in the treatment of restenosis.

Very limited success has been demonstrated with systemic pharmacological treatment to reduce the incidence of restenosis following angioplasty in patients. The lack of success of many of the pharmacotherapeutic agents in reducing the restenosis rates post-angioplasty and following stent implementation is believed to arise from inadequate concentrations of the agents at the lesion site. This has led to the development of various local delivery devices that would ideally deliver and retain adequate amounts of drug to the vessel wall for sufficient periods of time to ensure a therapeutic effect without inducing further injury or compromising blood flow. Local dosing would avoid systemic toxicity, and the use of modified balloon catheters or coated stents might enable percutaneous approaches.

Angioplasty, Balloon, Coronary↗

PubMed1997

Heparin/heparan sulfate chelation inhibits control of vascular repair by tissue-engineered endothelial cells.

The relative importance of heparin-like compounds in mediating vascular repair is unclear. We investigated how protamine, a chelator of heparin, affected endothelial cell inhibition of vascular smooth muscle cell growth and intimal hyperplasia. The 52% (P < 0.001) reduction in smooth muscle cell proliferation produced by postconfluent endothelial cell-conditioned medium was entirely reversed by pretreatment of medium with heparinase and heparitinase and was inhibited in a dose-dependent fashion by the coadministration of protamine. Pretreatment of conditioned medium with heparinase and heparitinase largely prevented protamine's mitogenic activity, suggesting that protamine affects growth by interacting with heparin-like compounds. Perivascular implantation of polymerengrafted endothelial cells reduced neointima formation in denuded rat carotid arteries by 92% (P < 0.001) and cell proliferation by 81% (P < 0.001). Coadministration of protamine abolished the inhibitory potential of the cell implants, resulting in a nearly twofold exacerbation of intimal hyperplasia compared with controls (P < 0.001). Thus heparin-like molecules are essential to the biochemical regulation of vascular repair provided by endothelial cells, and the continued routine clinical use of heparin chelators, like protamine, may be questionable.

Animals↗

PubMed1995

Basic fibroblast growth factor is a signal for the initiation of centrosome redistribution to the front of migrating endothelial cells at the edge of an in vitro wound.

Rapid, efficient repair of the endothelium following focal endothelial wounding and denudation is regulated by a complex series of cellular processes. Directed cell migration, an early essential event in repair, is thought to be initiated by centrosome redistribution toward the front of the cell prior to the onset of migration. As such, centrosomal polarity may be an important regulatory event in directed endothelial cell migration. Little is known about the regulation of this process. To study this further, in vitro wounds were created down the middle of confluent porcine aortic endothelial monolayers by mechanical denudation. Conditioned media collected 1 hour after wounding contained basic fibroblast growth factor (bFGF). Antibodies directed against bFGF added to the cultures at the time of wounding significantly inhibited cell migration and transiently inhibited centrosome redistribution. When transcription was transiently inhibited with actinomycin D, present at 1 hour before and for 1 hour after wounding, the cells moved more slowly (5.2 +/- 2.8 versus 22.7 +/- 5.7 microns/h for control), taking five times longer for the wound to close. Throughout this period, centrosomes did not reorient to the front of the cells. When either recombinant bFGF or conditioned medium collected from control cultures at 1 hour after wounding was added 23 hours after actinomycin D was washed out (at which time RNA synthesis returned to control levels), the centrosomes redistributed to the front of the cells, and cells migrated at a rapid rate (17.2 +/- 4.2 microns/h), similar to control. However, the recombinant bFGF or conditioned media had no effect when added immediately after actinomycin D was removed, ie, when RNA synthesis was still inhibited.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Endothelial wounds with disruption in cell migration repair primarily by cell proliferation.

Disruption of vascular integrity results in activation of endothelial cells to initiate repair. Cytoskeletal reorganization, cell spreading, migration, and proliferation are important processes which regulate rapid and efficient repair. Cell proliferation has been thought to be a secondary event and dependent to a large extent on prior cell migration. We used a model of dysfunctional repair to test the hypothesis that a reduction in migration will result in a reduction in cell proliferation. The extent of cell proliferation was studied in an in vitro repair model, in which endothelial function is studied during the closure of a 1500-microns mechanically induced wound in a confluent monolayer. Dysfunctional repair is induced in this model by transiently inhibiting endothelial cell transcription. Wounds are incubated with actinomycin D at the time of wounding for 2 hr, which results in wound closure taking about 4.5 times longer than normal. Cell proliferation is measured by 5-bromo-2'-deoxyuridine incorporation. In actinomycin D-treated wounds, the rate of cell migration is reduced, and for the first 120 hr very little cell proliferation occurs and it involves the first 4 rows of cells adjacent to the wound edge. This is in comparison to normal wounds in which proliferation is prominent by 24 hr after wounding, also involving the first 4 rows of cells and by 48 hr, proliferation extends to involve the first 10 rows of cells. In dysfunctional wounds, however, it takes 144 hr after wounding before the first 10 rows of cells begin to actively proliferate. In addition, proliferation occurs deeper in the monolayer and this level is maintained over the next 144 hr, when the wound closes. Thus, when migration is reduced, proliferation becomes prominent and persists to complete repair. However, the overall rate of repair is reduced.

Animals↗

PubMed1993

In vitro large-wound re-endothelialization. Inhibition of centrosome redistribution by transient inhibition of transcription after wounding prevents rapid repair.

Rapid, efficient re-endothelialization of large wounds is characterized by a specific sequence of cytoskeletal events that occur after wounding. Wounds 1.5 mm wide were created down the middle of confluent porcine aortic endothelial monolayers to study regulation of repair. The wounded cultures were incubated for short periods with cycloheximide or actinomycin D to test the hypothesis that transient inhibition of translation and transcription at the time of wounding disrupts rapid repair by interfering with centrosome redistribution to the front of the cell, an early event associated with cell migration. Although centrosome reorientation did not occur when protein synthesis was inhibited with 20 micrograms/mL cycloheximide for 1 hour before and for up to 4 hours after wounding, reorientation did occur by 2 hours after cycloheximide was washed out. The times taken for the wound to close for cycloheximide-treated and control cells did not differ (60 +/- 1.1 vs 60 +/- 0.8 hours). When transcription was inhibited with 0.25 micrograms/mL actinomycin D for 1 hour before and for 1 hour after wounding, re-endothelialization was dramatically reduced. The time taken for the wound to close was almost five times longer (288 +/- 5.3 hours) than for control cells. The cells moved very slowly, maintaining a flattened, spread-out shape, as opposed to being elongated. The centrosomes did not reorient to the front of the cell throughout the entire period. However, addition of actinomycin D for 2 hours when centrosomes had already moved to the front of the cells (4 hours after wounding) did not reduce subsequent wound repair (60 +/- 1.3 hours). This study supports our hypothesis that centrosome redistribution is essential for efficient wound repair and suggests that redistribution is regulated by transcription of essential gene(s) that is induced immediately after wounding by an unknown short-lived signal. Two possible signals are the loss of cell contact and/or a soluble substance released from the cells at the time of wounding. When the signal is unable to induce transcription, dysfunctional repair occurs by a very slow centrosome-independent process.

Animals↗

PubMed1992

Centrosomes, microtubules, and microfilaments in the reendothelialization and remodeling of double-sided in vitro wounds.

The maintenance of endothelial monolayer integrity is an important function of aortic endothelial cells. Our study was designed to test two hypotheses related to the repair of the wounded endothelial monolayer. First, that the reappearance of the highly ordered cobblestone monolayer after wound closure is associated with specific sequential changes in the cytoskeletal system. Second, that there are different patterns of reendothelialization depending on whether microfilaments or microtubules are disrupted. One and a half millimeter wide wounds were created down the middle of confluent endothelial monolayers so that there were two wound edges facing each other. During the initiation of repair, the centrosomes of the cells on both sides of the wound reorientated to the front of the cell. The dense peripheral band of actin microfilaments disappeared, the cells elongated and migrated as a uniform sheet with wound closure occurring within 60 hours. The rate of closure remained constant until the migrating fronts met. The cytoskeletal changes observed and the rate of closure were similar to those we reported in a single edge wound. At closure, however, there was a transient piling up of cells which disappeared after 24 to 36 hours. Within 36 to 48 hours after closure, the centrosomes became randomly distributed around the nucleus. By 40 to 48 hours, the dense peripheral band started to reappear and the cells returned to a cobblestone appearance 72 to 96 hours after closure. Thus, the remodeling of the confluent monolayer after wound closure occurs in association with a specific series of cytoskeletal changes. When the microfilaments were disrupted with cytochalasin B, cell migration still occurred but it took up to four times longer for closure. These cells were initially flatter than normal and contained only a few microfilament fibers; however, the microtubule system was intact and centrosome reorientation occurred, but at a slower rate. However, when the microtubules were disrupted with colchicine, either at the onset of wounding or during repair, neither centrosome reorientation nor cell migration occurred. Thus wound closure of small-sized wounds require the presence of intact microtubules, whereas the additional presence of microfilaments results in a more rapid and efficient system of reendothelialzation.

Actin Cytoskeleton↗

PubMed1990

Endothelial repair.

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Actin Cytoskeleton↗