Search PubMed⌕ Search

Biomedical subjects

R M Nerem

Publications and source records attributed to R M Nerem.

At least 19 recordsLinked to original sources
PubMed1992

Peptide amidating enzymes are present in cultured endothelial cells.

Carboxy-terminal amidation is a prevalent post-translational modification necessary for the bioactivity of many peptides. We now report that the two enzymes essential for amidation, peptidylglycine alpha-monooxygenase (PAM) and peptidylamidoglycolate lyase (PGL), are present in both the cytosol and membrane fractions of cultured bovine aortic endothelial cells. Endothelial PAM exhibits ascorbate-dependent turnover and is inactivated by the mechanism-based inactivator, 4-phenyl-3-butenoic acid (PBA), whereas PGL activity is independent of ascorbate and is not affected by PBA. These enzymological characteristics correspond to those of amidating enzymes from other tissues. These results suggest a heretofore unrecognized role for alpha-amidated peptides in cardiovascular function.

Amidine-Lyases↗
PubMed1992

Tissue engineering in the USA.

Tissue engineering is the application of the principles and methods of engineering and the life sciences towards the development of biological substitutes to restore, maintain or improve functions. It is an area which is emerging in importance worldwide. In the USA it has been actively fostered by the National Science Foundation, both through research grants and the sponsorship of a series of workshops starting in 1988. This brief review of activities in the USA focuses on cell culture technology as a foundation for tissue engineering and then discusses examples of applications. These include artificial skin and the use of encapsulated cells in the development of bioartificial organs. Also discussed is the reconstitution of a blood vessel in culture, both for use in basic research and for implantation as an artificial blood vessel in bypass surgery. In conclusion, other potential applications are mentioned as well as generic areas of technology for future development.

Artificial Organs↗
PubMed1992

Vascular fluid mechanics, the arterial wall, and atherosclerosis.

Atherosclerosis, a disease of large- and medium-size arteries, is the chief cause of death in the United States and in most of the western world. Severe atherosclerosis interferes with blood flow; however, even in the early stages of the disease, i.e. during atherogenesis, there is believed to be an important relationship between the disease processes and the characteristics of the blood flow in the arteries. Atherogenesis involves complex cascades of interactions among many factors. Included in this are fluid mechanical factors which are believed to be a cause of the highly focal nature of the disease. From in vivo studies, there is evidence of hemodynamic influences on the endothelium, on intimal thickening, and on monocyte recruitment. In addition, cell culture studies have demonstrated the important effect of a cell's mechanical environment on structure and function. Most of this evidence is for the endothelial cell, which is believed to be a key mediator of any hemodynamic effect, and it is now well documented that cultured endothelial monolayers, in response to a fluid flow-imposed laminar shear stress, undergo a variety of changes in structure and function. In spite of the progress in recent years, there are many areas in which further work will provide important new information. One of these is in the engineering of the cell culture environment so as to make it more physiologic. Animal studies also are essential in our efforts to understand atherogenesis, and it is clear that we need better information on the pattern of the disease and its temporal development in humans and animal models, as well as the specific underlying biologic events.(ABSTRACT TRUNCATED AT 250 WORDS)

Arteriosclerosis↗
PubMed1992

Flow-induced calcium transients in single endothelial cells: spatial and temporal analysis.

Endothelial cells (EC) are uniquely situated to respond to hemodynamic forces. Because flow-mediated release of endothelial-derived relaxing factors is associated with increased EC intracellular calcium ([Ca2+]i), we sought to determine the effects of fluid shear stress on EC [Ca2+]i. Cells were subjected to flow in parallel-plate flow chambers and glass capillary tubes, and single cell [Ca2+]i was measured using fura-2. Upon initiation of flow (shear stress of 30 dyn/cm2), [Ca2+]i increased within 30 s to a peak value (approximately 4 times basal) and then decreased slowly to a plateau (approximately 2 times basal) that persisted for greater than 5 min. A striking finding was that the increases in [Ca2+]i were nonhomogeneous; the nuclear region and a periplasma membrane region were higher than the cytosol. After flow cessation, the increase in [Ca2+]i could be elicited repeatedly by resumption of flow. Removing extracellular Ca2+ did not eliminate the response. In contrast to EC, rat aortic smooth muscle cells showed no flow-mediated increase in [Ca2+]i. The complexity of EC [Ca2+]i response to flow suggests regulation of [Ca2+]i by several mechanisms that may serve a role in both short- and long-term EC responses to flow.

Animals↗
PubMed1992

Molecular cloning and characterization of the constitutive bovine aortic endothelial cell nitric oxide synthase.

The constitutive endothelial cell nitric oxide synthase (NOS) importantly regulates vascular homeostasis. To gain understanding of this enzyme, a pEF BOS cDNA library of 5 x 10(5) clones was prepared from bovine aortic endothelial cells (BAEC) and screened with a 2.8-kb cDNA BamHI fragment of rat brain NOS. Clone pBOS13 was found to express NO synthase activity when transfected into COS-7 cells. Sequence analysis revealed sequences compatible with binding domains for calcium/calmodulin, flavin mononucleotide, flavin adenine nucleotide and NADPH. The deduced amino acid sequence revealed a protein with a relative mol mass of 133,286, which is 58% homologous to the rat cerebellar NOS and 51% homologous to the mouse macrophage NOS. The amino-terminal portion of the protein exhibits several characteristics peculiar to the endothelial cell NOS. These include a proline-rich region and several potential sites for proline-directed phosphorylation as well as a potential substrate site for acyl transferase. Northern hybridization to mRNA from cultured BAEC revealed an abundant 4.8-kb message, which was not increased by coincubation with tumor necrosis factor alpha, but was markedly increased by exposure to shear stress for 24 h. The unique features of the endothelial cell NO synthase, particularly in the amino terminal portion of the molecule, may provide for novel regulatory influences of enzyme activity and localization.

Amino Acid Oxidoreductases↗
PubMed1991

The pathogenesis of atherosclerosis: an overview.

In this unifying hypothesis directed to the etiology and pathogenesis of atherosclerosis, the importance of focal arterial lesion-prone sites has been emphasized. Key initial participants in these sites include the focal intimal influx and accumulation of low-density lipoprotein (LDL) and a preferential recruitment of blood monocytes. Both are further enhanced in the presence of hyperlipidemia, when the quantity of intimal LDL and the oxidative potential of the intima exceed the capacity of macrophages to remove, via the non-down-regulating scavenger receptor, cytotoxic anionic (Ox-LDL) macromolecules. Foam cells, pathognomonic of the fatty streak, form during the receptor-mediated uptake of Ox-LDL by the macrophages. Interstitial free radicals and the excess of Ox-LDL particles injure and kill cells, including the foam cells, with the formation of the necrotic extracellular lipid core, a key transitional step in lesion progression. Monocyte-macrophage recruitment to the intima is likely to be regulated not only by a multiplicity of endothelial adhesive cytokines, integrins, and selectins, but also by the monocyte-specific chemoattractant, MCP-1, constitutively synthesized and secreted by intimal smooth muscle and endothelial cells. Its synthesis and secretion is augmented by mildly oxidized LDL. Free radicals, pivotal in the oxidation of LDL, and derived from activated macrophages, and also endothelial and smooth muscle cells. Smooth muscle cells migrate from the media through the intimal endothelial layer (IEL) and proliferate under the regulation of a number of mitogens, including platelet-derived growth factor (PDGF). Collagen synthesis by smooth muscle cells is substantial. Lymphocytes, as a source of interferons, invade the plaque and are present in the adventitia in substantial numbers, likely representing an autoimmune response in the later stages of plaque development. Platelets and mural thrombosis directly contribute to subsequent plaque growth, particularly after plaque rupture or fissure and disruption of the thromboresistant endothelial cells (EC). Plaque regression in all likelihood involves the conversion of the inert pool of extracellular lipid to a metabolically active intracellular pool and subsequent clearance by the high-density lipoprotein mediated reverse cholesterol transport system. The atherogenic cascades so described conceptually represent arterial inflammatory and healing processes occurring in a hyperlipidemic environment. Many components of pathogenesis are the targets for modulation by genetic, hemodynamic and selected risk factors. The prevention and treatment of the disease should logically target reduction in plasma LDL levels, the inhibition of the oxidative modification of lipoproteins, including LDL, by free radical scavengers, and augmentation of the reverse cholesterol transport system.

Animals↗
PubMed1991

Cellular engineering.

Cellular engineering applies the principles and methods of engineering to the problems of cell and molecular biology of both a basic and applied nature. As biomedical engineering has shifted from the organ and tissue level to the cellular and sub-cellular level, cellular engineering has emerged as a new area. A cornerstone of much of this activity is cell culture technology, i.e., the ability to grow living cells in the artificial environment of a laboratory. Cellular engineering includes the role of engineering in both basic cell biology research and in the making of products which use living cells, e.g., tissue engineering and bioprocess engineering. The former involves the use of living cells in the development of biological substitutes for the restoration or replacement of function, and the latter the use of living cells to manufacture a biochemical product, e.g., through the use of recombinant DNA technology. In fact, as biomedical engineering has expanded to include the cellular level, and bioprocess engineering has shifted in interest from microbial organisms to include mammalian cells, there are intellectual issues in which an interest is shared by these two formerly separate areas of engineering activity. Cellular engineering thus transcends the field of biomedical engineering.

Biomedical Engineering↗
PubMed1991

A numerical calculation of flow in a curved tube model of the left main coronary artery.

The flow pattern in the left main coronary artery has been calculated using an idealized geometry and by numerically solving the full Navier-Stokes equations for a Newtonian fluid. Two different forms for the entrance velocity profile were used, one a time-varying, flat profile and the other a time-varying, less flat velocity profile. The results obtained demonstrate the presence of secondary motions for conditions simulating flow in the left main coronary artery, with maximum secondary flow velocities being on the order of three to four percent of the maximum axial velocity. This secondary flow phenomenon has an important influence on the wall shear stress distribution, in spite of the fact that there is virtually no alteration in the axial velocity profile. The maximum ratio of the outer wall shear stress to that on the inner wall is 1.4 at a Reynolds number of Re = 270, and it increases with increasing Reynolds number, reaching a value of 1.7 at Re = 810. Although there are significant differences in the results in the immediate vicinity of the inlet for the two different forms of the entrance velocity profile used, this difference does not persist far into the tube. Independent of the choice of the entrance velocity profile, it appears that there will be significant secondary flow effects on the wall shear stress.

Blood Flow Velocity↗
PubMed1991

Effects of pulsatile flow on cultured vascular endothelial cell morphology.

Endothelial cells (EC) appear to adapt their morphology and function to the in vivo hemodynamic environment in which they reside. In vitro experiments indicate that similar alterations occur for cultured EC exposed to a laminar steady-state flow-induced shear stress. However, in vivo EC are exposed to a pulsatile flow environment; thus, in this investigation, the influence of pulsatile flow on cell shape and orientation and on actin microfilament localization in confluent bovine aortic endothelial cell (BAEC) monolayers was studied using a 1-Hz nonreversing sinusoidal shear stress of 40 +/- 20 dynes/cm2 (type I), 1-Hz reversing sinusoidal shear stresses of 20 +/- 40 and 10 +/- 15 dynes/cm2 (type II), and 1-Hz oscillatory shear stresses of 0 +/- 20 and 0 +/- 40 dynes/cm2 (type III). The results show that in a type I nonreversing flow, cell shape changed less rapidly, but cells took on a more elongated shape than their steady flow controls long-term. For low-amplitude type II reversing flow, BAECs changed less rapidly in shape and were always less elongated than their steady controls; however, for high amplitude reversal, BAECs did not stay attached for more than 24 hours. For type III oscillatory flows, BAEC cell shape remained polygonal as in static culture and did not exhibit actin stress fibers, such as occurred in all other flows. These results demonstrate that EC can discriminate between different types of pulsatile flow environments.(ABSTRACT TRUNCATED AT 250 WORDS)

Adaptation, Physiological↗
PubMed1990

Parametric analysis of flow in the intramyocardial circulation.

A simple mathematical model of the intramyocardial circulation has been utilized to provide a better understanding of coronary blood flow. The model includes three myocardial layers, each characterized by a three-parameter windkessel with one capacitance and two resistances. The effects of the beating heart are taken into account by means of an intramyocardial pump and the possible collapse of the vessels by an elevated backpressure. The three basic parameters that govern the flow are a normalized time constant, tau, the total resistance, Rt, and a parameter, alpha, which specifies the resistance distribution in the intramyocardial circulation. Both the normal beating heart and prolonged diastole have been investigated analytically as well as numerically. It is shown that each of these parameters has its own special significance. Calculated pressure-flow relationships and zero-flow pressures for the case of prolonged diastole show a high sensitivity to tau and alpha.

Animals↗
PubMed1990

Vascular endothelial cell proliferation in culture and the influence of flow.

The influence of laminar shear stress on cell proliferation was investigated for subconfluent bovine aortic endothelial cell monolayers seeded on either glass or Thermanox. The effect of both steady and pulsatile shear stress was studied. For bovine aortic endothelial cells on Thermanox exposed to steady flow, shear stress levels greater than 15 dyne/cm2 resulted in a dose-related reduction in the rate of cell proliferation. At 90 dyne/cm2, the rate of proliferation was virtually totally arrested for 48 h, but then resumed. Pulsatile shear stress produced an exaggeration of the effect observed in response to steady shear stress. Bovine aortic endothelial cells seeded on glass, exhibited a similar but more sensitive response, with a significant reduction in growth rate observed after 24 h at shear stress levels greater than 5 dyne/cm2 and a near cessation of proliferation at 13 dyne/cm2.

Animals↗
PubMed1990

Application of the micropipette technique to the measurement of cultured porcine aortic endothelial cell viscoelastic properties.

The viscoelastic deformation of porcine aortic endothelial cells grown under static culture conditions was measured using the micropipette technique. Experiments were conducted both for control cells (mechanically or trypsin detached from the substrate) and for cells in which cytoskeletal elements were disrupted by cytochalasin B or colchicine. The time course of the aspirated length into the pipette was measured after applying a stepwise increase in aspiration pressure. To analyze the data, a standard linear viscoelastic half-space model of the endothelial cell was used. The aspirated length was expressed as an exponential function of time. The actin microfilaments were found to be the major cytoskeletal component determining the viscoelastic response of endothelial cells grown in static culture.

Animals↗
PubMed1990

Hemodynamic influences on vascular endothelial biology.

The vascular endothelium resides in a unique biomechanical stress environment resulting from the hemodynamics of the system. In vivo studies indicate that there are regional differences in endothelial biology and that this may be due to the influence of the local hemodynamic environment. To investigate this further, cell culture studies have been conducted using well-defined mechanical stress environments. To study flow effects, we have employed a parallel plate chamber in which endothelial cell monolayers are exposed to laminar flow. In such experiments and concomitant with changes in morphology, there are a variety of other alterations in cell function, including a decrease in the rate of cell proliferation for subconfluent monolayers. Changes in cell behavior due to the direct effect of pressure and in cultured cells which are in a cyclical stress field also have been observed. In the recognition/transduction of such a mechanical signal, the pathway may possibly include a membrane event linked to the control of intracellular calcium. It may be that the same signaling mechanisms are involved both in cytoskeletal/shape changes and in the control of the cell's growth program and, in exercising such an influence, hemodynamics may have an important role in the response of the arterial wall to injury and the resulting repair and/or disease processes.

Animals↗
PubMed1989

Pathophysiology of the atherogenic process.

Atherosclerosis is conceptually defined as the result of a multiplicity of interactive cascades among injurious stimuli and the healing responses of the arterial wall, occurring concurrently within a hyperlipidemic environment. In this discussion, the inflammatory nature of the disease is emphasized. Four aspects of the pathophysiology of atherogenesis are addressed: (1) The role(s) of fluid mechanical or hemodynamic stresses in the focal initiation and/or augmentation of lesions is discussed in terms of the influence of shear stress on endothelial cellular geometry, compliance, membrane anisotropy (r), low-density lipoprotein (LDL)-receptor expression, intracellular potential and replication; (2) mechanisms of blood monocyte recruitment to the arterial intima, including the roles of chemoattractants such as smooth muscle cell-derived chemotactic factor and oxidized LDL; (3) the alternate or "scavenger" receptor pathway of the macrophage and its pivotal roles in foam cell formation and plaque pathogenesis; and (4) the emerging significance of various lipoprotein modifications, and in particular, the oxidative modification of LDL, which facilitates the uptake of the cytotoxic oxidized LDL via the scavenger receptor, thus providing a non-down-regulating mechanism for foam cell formation and plaque development. Evidence indicates that the antioxidant drug probucol prevents the oxidative modification of LDL, thereby retarding atherogenesis independently of cholesterol reduction.

Animals↗
PubMed1989

The study of rheological effects on vascular endothelial cells in culture.

A number of cell culture studies have been reported on the influence of shear stress on vascular endothelial cells. Although through such studies much has been learned about the effect of an endothelial cell's hydrodynamic environment on its structure and function, the reports indicate significant differences in methodology. Using cell shape as an indicator of differences that might result from differing methodologies, an investigation of the influence of selected variables has been carried out. The results presented indicate that not only are such variables as the level of shear stress and the duration of exposure important, but also substrate, media composition, characteristics of the cell itself, and the nature of the flow, e.g. whether it is steady state or pulsatile.

Animals↗
PubMed1988

Correlation of laser-Doppler-velocity measurements and endothelial cell shape in a stenosed dog aorta.

Laser-Doppler-velocity measurements were carried out in an elastic 1:1 true-to-scale silicone rubber model of a dog aorta with stenosis. The model was constructed from a cast of a severely stenosed dog aorta (71% of its area). The stenosis in the dog aorta was prepared by wrapping a cotton band around the aorta. This band was tightened until the presence of a thrill or a bruit was felt distal to the band. Twelve weeks later the animal was sacrificed and a cast was prepared from the aorta. From this vascular cast, the cross-sectional area was calculated. Endothelial cell geometry and orientation was studied using computerized analysis to determine the cell area and shape index. An elastic silicone rubber model was prepared from the cast to measure the velocity profiles and to estimate the local wall shear stress. Velocity measurements were done at steady and pulsatile flow using a Newtonian aqueous-glycerol solution and a non-Newtonian blood-like fluid. From those velocity measurements the velocity gradients near the wall were determined and the shear stress calculated. The flow distal to the stenosis separates from the wall at physiological conditions. The endothelial cells are smaller and more elongated in the throat; distal to the stenosis they are larger and rounder. The shape index distribution along the stenosed aorta is correlated with the level of wall shear stress. It is shown that even low changes in the wall shear stress have an influence on the orientation of the endothelial cells.

Animals↗