Biomedical subjects

R M Nerem

Publications and source records attributed to R M Nerem.

At least 37 records · Page 2Linked to original sources

PubMed1996

Shear stress modulates expression of Cu/Zn superoxide dismutase in human aortic endothelial cells.

A major determinant of the level of cellular superoxide anion (O2-.) is the dismutation of O2-. to hydrogen peroxide by the enzyme superoxide dismutase (SOD). Three forms of SOD exist, but in endothelial cells, the major form outside of the mitochondria is the cytosolic copper/zinc-containing superoxide dismutase (Cu/Zn SOD). Since fluid shear stress is an important determinant of the function and structure of endothelial cells in vivo, we examined the effect of laminar shear stress on the expression of Cu/Zn SOD in cultured human aortic endothelial cells. Laminar shear stress of 0.6 to 15 dyne/cm2 increased Cu/Zn SOD mRNA in a time- and dose-dependent manner in human aortic endothelial cells. Shear stress also increased both Cu/Zn SOD protein content and the enzyme activity. Nuclear runon assays showed that nuclei from human aortic endothelial cells exposed to laminar shear stress had a 1.6-fold greater transcriptional activity of the Cu/Zn SOD gene compared with cells not exposed to shear, indicating that an increase in Cu/Zn SOD mRNA induced by laminar shear stress is at least in part mediated by increased transcription. In contrast, shear stress had no effect on Cu/Zn SOD mRNA levels in human aortic smooth muscle cells. These findings show that physiological levels of shear stress increase expression of Cu/Zn SOD in the endothelium. This adaptation to shear stress might augment the effect of locally produced NO. and thereby promote the antiatherogenic and anti-inflammatory properties of the endothelial cell.

Aorta↗

PubMed1996

Phosphorylation of endothelial nitric oxide synthase in response to fluid shear stress.

Endothelial cells release nitric oxide (NO) more potently in response to increased shear stress than to agonists which elevate intracellular free calcium concentration ([Ca2+]i). To determine mechanistic differences in the regulation of endothelial constitutive NO synthase (ecNOS), we measured NO production by bovine aortic endothelial cells exposed to shear stress in a laminar flow chamber or treated with Ca2+ ionophores in static culture. The kinetics of cumulative NO production varied strikingly: shear stress (25 dyne/cm2) stimulated a biphasic increase over control that was 13-fold at 60 minutes, whereas raising [Ca2+]i caused a monophasic 6-fold increase. We hypothesized that activation of a protein kinase cascade mediates the early phase of flow-dependent NO production. Immunoprecipitation of ecNOS showed a 210% increase in phosphorylation 1 minute after flow initiation, whereas there was no significant increase after Ca2+ ionophore treatment. Although ecNOS was not tyrosine-phosphorylated, the early phase of flow-dependent NO production was blocked by genistein, an inhibitor of tyrosine kinases. To determine the Ca2+ requirement for flow-dependent NO production, we measured [Ca2+]i with a novel flow-step protocol. [Ca2+]i increased with the onset of shear stress, but not after a step increase. However, the step increase in shear stress was associated with a potent biphasic increase in NO production rate and ecNOS phosphorylation. These studies demonstrate that shear stress can increase NO production in the absence of increased [Ca2+]i, and they suggest that phosphorylation of ecNOS may importantly modulate its activity during the imposition of increased shear stress.

Animals↗

PubMed1995

Shear stress modulates endothelial cell morphology and F-actin organization through the regulation of focal adhesion-associated proteins.

Flow-related shear stress has been shown to modulate endothelial cell structure and function including F-actin microfilament organization. Focal adhesion-associated proteins such as vinculin, talin, and specific integrins may play a role in the modulation of these cytoskeletal and morphological changes. Double-label immunofluorescence studies indicated that, in static culture, alpha 5 beta 1 fibronectin receptors (alpha 5 beta 1 FNRs) and alpha v beta 3 vitronectin receptors (alpha v beta 3 VNRs) were found predominantly in the peripheral regions of bovine aortic endothelial cells (BAECs) corresponding to the localization of vinculin, talin, and actin microfilament terminations. In response to shear stress, concomitant with cell elongation and the appearance of stress fibers aligned with the direction of flow, there was a prominent localization of vinculin and alpha v beta 3 VNRs as the "upstream" end of the cells. Stress fiber terminations were clearly evident at these concentrations of focal adhesion-associated proteins. These data suggest that the upstream concentration of these proteins may direct shear stress-induced stress fiber formation and may function in the alignment of the fibers in the direction of flow. Levels of surface alpha v beta 3 VNRs were found to decrease in response to flow, possibly reflecting the decrease in numbers of "downstream" receptors. Unlike the arrangement of vinculin and alpha v beta 3 VNRs observed following exposure to flow, talin and alpha 5 beta 1 FNRs, in addition to being localized at the upstream end of the cell, were also evenly distributed throughout the rest of the cell. Surface levels of alpha 5 beta 1 FNRs increased in response to shear stress, perhaps providing an increased adherence of BAECs to the extracellular matrix through these receptors. These data suggest that focal adhesion-associated proteins play specific roles in the response of BAECs to shear stress.

Actins↗

PubMed1995

Elongation of confluent endothelial cells in culture: the importance of fields of force in the associated alterations of their cytoskeletal structure.

Studies using either animal models or in vitro flow systems have shown that the shape of large-vessel endothelial cells (ECs) was sensitive to the amplitude of the flow imposed on them. In order to better understand the morphological changes experienced by ECs when exposed to physical forces such as shear stress, the mechanical integrity of confluent bovine aortic ECs (BAECs) was anisotropically perturbed using the five following types of experiments: (i) slicing and partial scraping of BAEC monolayers; (ii) culture of BAECs on narrow strips of adhesive plastic; (iii) incubation of confluent BAECs with media containing low Ca2+ concentrations; (iv) culture of ECs on top of rectangular collagen gels; and (v) exposure of BAECs to laminar steady shear stress. In all five experimental systems, BAECs exhibited an elongated morphology and aligned their major axes in specific directions. In addition, a preferential alignment of actin microfilaments, vimentin intermediate filaments, and streaks of vinculin with the major axes of the cells often occurred concomitantly with BAEC elongation. In all five systems, the elongation of ECs was analyzed in terms of a mechanical deformation borne by the cytoskeleton, and possibly caused by anisotropic distribution of the forces experienced by the cell structure. In addition, the strain-stress and stiffness-stress relationships characterizing the elongation of BAECs exposed to steady flow were qualitatively similar to those computed for the uniaxial deformation of a spherical geodesic. Our findings suggest that the cytoskeleton of ECs plays an important role in the transduction of those forces which cause an elongation of ECs.

Animals↗

PubMed1995

Oscillatory shear stress and hydrostatic pressure modulate cell-matrix attachment proteins in cultured endothelial cells.

Endothelial cells (ECs) may behave as hemodynamic sensors, translating mechanical information from the blood flow into biochemical signals, which may then be transmitted to underlying smooth muscle cells. The extracellular matrix (ECM), which provides adherence and integrity for the endothelium, may serve an important signaling function in vascular diseases such as atherogenesis, which has been shown to be promoted by low and oscillating shear stresses. In this study, confluent bovine aortic ECs (BAECs), were exposed to an oscillatory shear stress or to a hydrostatic pressure of 40 mmHg for time periods of 12 to 48 h. Parallel control cultures were maintained in static condition. Although ECs exposed to hydrostatic pressure or to oscillatory flow had a polygonal morphology similar to that of control cultures, these cells possessed more numerous central stress fibers and exhibited a partial loss of peripheral bands of actin, in comparison to static cells. In EC cultures exposed to oscillatory flow or hydrostatic pressure, extracellular fibronectin (Fn) fibrils were more numerous than in static cultures. Concomitantly, a dramatic clustering of alpha 5 beta 1 Fn receptors and of the focal contact-associated proteins vinculin and talin occurred. Laminin (Ln) and collagen type IV formed a network of thin fibrils in static cultures, which condensed into thicker fibers when BAECs were exposed to oscillatory shear stress or hydrostatic pressure. The ECM-associated levels of Fn and Ln were found to be from 1.5- to 5-fold greater in cultures exposed to oscillatory shear stress or pressure for 12 and 48 h, than in static cultures. The changes in the organization and composition of ECM and focal contacts reported here suggest that ECs exposed to oscillatory shear stress or hydrostatic pressure may have different functional characteristics from cells in static culture, even though ECs in either environment exhibit a similar morphology.

Actins↗

PubMed1995

Effects of shear on endothelial cell calcium in the presence and absence of ATP.

The purpose of this study was to characterize the effect of various shear conditions on endothelial cell intracellular calcium ([Ca2+]i). Bovine aortic endothelial cells (BAEC) were loaded with Fluo-3 and exposed to flow in a parallel plate flow chamber designed for confocal microscopy. The flow medium was medium 199 (M-199), which was prepared with and without adenosine triphosphate (ATP). In the presence of ATP, initiation of flow at a shear stress of 2.5 dyn/cm2 evoked a strong, sustained elevation of [Ca2+]i that gradually returned to baseline levels over 10 to 15 min. By contrast, in the absence of ATP, initiation of flow at 2.5 dyn/cm2 produced only transient increases in [Ca2+]i in a small proportion of the cells. As shear rate was increased from 2.5 to 15 dyn/cm2 in this medium, both the relative fluorescence of the monolayer and the proportion of cells across the monolayer that displayed calcium transients increased in a dose-dependent fashion. In conclusion, the response of an endothelial cell monolayer to increasing levels of shear is not only to increase [Ca2+]i within individual cells, but to increase the duration of response and the number of cells responding at the onset of shear. This recruitment of larger numbers of cells at higher levels of shear may represent a novel signaling mechanism within the endothelium.

Adenosine Triphosphate↗

PubMed1995

Calcium responses of endothelial cell monolayers subjected to pulsatile and steady laminar flow differ.

The vascular endothelium is the primary transducer of hemodynamically imposed mechanochemical events. In this study, we measured the intracellular free calcium concentration ([Ca2+]i) using the fluorescent probe fura 2 and ratiometric digital imaging in cultured bovine aortic endothelial cells (BAEC) subjected to various laminar flow patterns. These were steady shear stress (0.2-70 dyn/cm2) and three types of sinusoidal pulsatile shear stress (nonreversing: 40 +/- 20 dyn/cm2; reversing: 20 +/- 40 dyn/cm2; and purely oscillatory: 0 +/- 20 dyn/cm2; flow frequencies: 0.4, 1.0, and 2.0 Hz) in a serum-containing medium. The most dramatic finding was failure of a purely oscillatory flow to increase [Ca2+]i in BAEC monolayers. In contrast, steady flow, as well as nonreversing and reversing pulsatile flows, increased [Ca2+]i. The dynamics of the response were dependent on the flow pattern. Both internal Ca2+ release and extracellular Ca2+ entry were involved in these [Ca2+]i increases. Also, switching from either a steady nonreversing pulsatile or reversing pulsatile flow back to a static condition resulted in a [Ca2+]i increase. However, switching from an oscillatory flow to a static condition did not induce any changes in average [Ca2+]i. This study shows that endothelial cells are able to sense different flow environments in terms of [Ca2+]i signaling and is relevant to further studies of the influence of hemodynamic forces on vascular pathophysiology.

Animals↗

PubMed1995

Regulation of endothelial cell nitric oxide synthase mRNA expression by shear stress.

Shear stress enhances expression of Ca(2+)-calmodulin-sensitive endothelial cell nitric oxide synthase (ecNOS) mRNA and protein in bovine aortic endothelial cells (BAEC). The present studies were performed to investigate mechanisms responsible for regulation of ecNOS mRNA expression by shear stress and to determine if this induction of ecNOS mRNA is accompanied by an enhanced nitric oxide (NO) production. Shear stresses of 15 dyn/cm2 for 3-24 h resulted in a two- to threefold increase of ecNOS mRNA content quantified by Northern analysis in BAEC. Shear stresses (1.2-15 dyn/cm2) for 3 h resulted in an induction of ecNOS mRNA in a dose-dependent manner. In human aortic endothelial cells, shear stresses of 15 dyn/cm2 for 3 h also resulted in ecNOS mRNA induction. In BAEC, this induction in ecNOS mRNA was prevented by coincubation with actinomycin D (10 micrograms/ml). The K+ channel antagonist tetraethylammonium chloride (3 mM) prevented increase in ecNOS mRNA in response to shear stress. The ecNOS promotor contains putative binding domains for AP-1 complexes, potentially responsive to activation of protein kinase C (PKC). However, selective PKC inhibitor calphostin C (100 nM) did not inhibit ecNOS induction by shear stress. Finally, production of nitrogen oxides under both basal conditions and in response to the calcium ionophore A-23187 (1 microM) by BAEC exposed to shear stress was increased approximately twofold compared with cells not exposed to shear stress. These data suggest that ecNOS mRNA expression is regulated by K+ channel opening, but not by activation of PKC, and that shear not only enhances ecNOS mRNA expression but increases capacity of endothelial cells to release NO.

Animals↗

PubMed1995

Changes in organization and composition of the extracellular matrix underlying cultured endothelial cells exposed to laminar steady shear stress.

BACKGROUND: In blood vessels, the extracellular matrix (ECM) underlying the endothelium supports endothelial cell (EC) attachment, spreading, migration, and proliferation. The structure and composition of the ECM may be modulated by hemodynamic shear stress, which may play a role in the pathogenesis of vascular diseases such as atherosclerosis. EXPERIMENTAL DESIGN: In this study, in vitro effects of fluid shear stress on the ECM of EC were investigated. Cultured bovine aortic EC (BAEC) were exposed to a steady laminar shear stress of 30 dyn/cm2 from 3 to 48 hours, using a parallel-plate flow chamber. Parallel control cultures were maintained under static conditions. The organization of fibronectin (Fn), laminin (Ln), collagen type IV (Col IV), and vitronectin (Vn) was analyzed by immunofluorescence microscopy. Changes in the profile of proteins present in the deoxycholate-insoluble ECM fraction of EC were determined using two-dimensional gel electrophoresis, and the levels of Fn, Ln, and Vn were determined by Western blotting. RESULTS: Fn, Ln, and Col IV exhibited both a granular pattern in cell perinuclear areas and a fibrillar pattern localized underneath EC. On exposure of bovine aortic EC to shear stress, Fn fibrils grouped into thicker tracts of fibrils, and there was a tendency for some of these tracks of fibrils to align with the direction of flow. Ln and Col IV also grouped into thicker fibers, which, in contrast to Fn, were randomly oriented. Vn exhibited a diffuse granular pattern, which did not change in response to shear stress. Consistent increases in the levels of four unidentified acidic proteins (mol wt/pI = 52/4.9, 70/4.7, 70/5.5, and 110/4.4) were observed after 3 to 6 hours of exposure to flow. The level of Fn present in the ECM was decreased twofold 12 hours after exposure of the cell monolayer to flow, and then increased after 24 and 48 hours. The level of Ln showed a twofold increase after 24 and 48 hours of flow, whereas the level of Vn was not altered by shear stress. CONCLUSIONS: These changes in organization and composition observed in the ECM of cultured EC may play a significant role in shear stress-induced morphologic alterations in EC and may represent relevant events in the initiation of atherosclerotic lesions by influencing both EC and smooth muscle cell function.

Amino Acid Sequence↗

PubMed1994

Tissue engineering a blood vessel: regulation of vascular biology by mechanical stresses.

Important to the tissue engineering of a substitute blood vessel is an understanding of those factors which regulate vascular biology. A major factor in this regulation is the mechanical environment imposed by the hemodynamics of the vascular system. In this the vascular endothelium plays a critical role, and over the past two decades much has been learned about the influence of hemodynamics on vascular endothelial biology, to a large degree using cell culture to study the effects of flow and cyclic stretch. In our laboratory, such studies are now being extended through the development of a model of the arterial wall involving the co-culture of endothelial cells and smooth muscle cells. The development of such a model and its use in the study of hemodynamic effects represents necessary steps in the evolution of approaches to tissue engineering a blood vessel.

Animals↗

PubMed1994

Effect of flow on the process of endothelial cell division.

In this study we investigated the effect of flow and the associated shear stress on the process of endothelial cell division. A shear stress of 7 N/m2 (70 dyne/cm2) was applied to a monolayer of bovine aortic endothelial cells (BAECs) by using a parallel-plate flow-chamber system. Dividing cells under flow conditions were qualitatively compared with those under static-culture conditions. In addition, the duration of some phases of the cell cycle (ie, mitosis and cytokinesis) was measured for both static and flow conditions. Dividing BAECs in static cultures bowed upward; however, when cells were preconditioned by at least 6 hours of exposure to flow, they stayed relatively attached to the substrate during the entire process of cell division. Cell area significantly decreased during division for both static-culture and flow conditions. Finally, although significant changes in the duration of phases were observed in single experiments during which cells were exposed to shear stress, any differences disappeared when the data for a large number of cells were combined. Hence, on average, flow and its associated shear stress did not affect the duration of the different phases of endothelial cell division.

Animals↗

PubMed1993

Polarized secretion of IGF-I and IGF-I binding protein activity by cultured aortic endothelial cells.

Insulin-like growth factor-I (IGF-I) secretion by the vascular endothelium has been proposed to play a role in the regulation of vascular smooth muscle cell proliferation. Because vascular smooth muscle cells are adjacent to the abluminal surface of the endothelium, we tested the hypothesis that secretion of IGF-I by endothelial cells is polarized. Porcine aortic endothelial cells were cultured on permeable membranes and IGF-I measured by radioimmunoassay. Basal secretion exceeded apical secretion by a ratio of 2.3 +/- 0.2:1.0 (P < 0.05). We also identified 35 kDa IGF-I binding protein activity that is preferentially secreted on the basal surface of endothelial cells. We conclude that both IGF-I and IGF-I binding protein activity secretion by endothelial cells is polarized towards the basal surface of the endothelium. A polarized secretion mechanism for IGF-I may be of importance in the normal growth and differentiation of the vasculature as well as in the development of vascular pathology.

Animals↗

PubMed1993

Hemodynamics and vascular endothelial biology.

The vascular endothelium, as the interface between flowing blood and the underlying vessel wall, not only resides in a hemodynamic environment, but also is a mediator of flow-related influences. In recent years, particularly with the advent of cell culture studies, much has been learned about the effects of this mechanical environment. This brief review focuses on these more recent cell culture studies, emphasizing the effects of flow and the associated shear stress on vascular endothelial biology. Included are endothelial cell shape and orientation, cytoskeletal components, endocytosis, proliferation, and signal recognition/transduction mechanisms. Of particular interest are mechanical effects on the release of vasoactive substances, and recent studies of prostacyclin, endothelin, and nitric oxide are discussed. Through this review it will be demonstrated that flow has a major influence on the synthesis and release of such substances.

Animals↗

PubMed1993

Hemodynamics and the vascular endothelium.

The endothelium, once thought to be a passive, non-thrombogenic barrier, is now recognized as being a dynamic participant in vascular biology and pathobiology. Part of its dynamic nature is due to the influence of the mechanical environment imposed by the hemodynamics of the vascular system. Over the past two decades much has been learned about the influence of hemodynamics on the vascular endothelium. This has been in part through in vivo experiments; however, in the past 15 years a number of laboratories have turned to the application of in vitro cell culture systems to investigate the influence of flow and cyclic stretch on the biology of vascular endothelium. Taken together these studies demonstrate that flow and the associated shear stress modulate both endothelial cell structure and function. Cell culture studies employing cyclic stretch provide similar evidence. Furthermore, these effects of mechanical environment extend to the gene expression level, with there being a differential regulation of mRNA. A critical question is how does an endothelial cell recognize the mechanical environment in which it resides and, having done so, how is this transduced into the changes in structure and function observed? Although our knowledge of the recognition events remains limited, studies on signal transduction in response to a mechanical stimulus indicate that many of the second messengers known to be triggered by chemical agonists also are involved in transducing a mechanical signal.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Fluid shear stress stimulates platelet-derived growth factor expression in endothelial cells.

Fluid flow and the associated shear stress play a critical role in vascular growth and remodeling. Recent data suggest that increased endothelial cell expression of platelet-derived growth factor (PDGF) A- and B-chain by flow may participate in these events. In the present study, we examined the mechanism for flow-induced PDGF expression, focusing on protein kinase C (PKC). Bovine aortic endothelial cells were exposed to flow (shear stress = 30 dyn/cm2) in a parallel-plate flow chamber. Increases in PDGF B-chain, but not PDGF A-chain, were observed within 3 h, maximal within 6 h (13-fold increase), and sustained for 24 h. PKC appeared to be involved because phorbol 12-myristate 13-acetate induced PDGF B-chain mRNA. Activation of PKC alone, however, was insufficient to induce PDGF mRNA because the selective PKC activator, 1-oleoyl-2-acetyl-sn-glycerol, did not induce PDGF expression. A PKC-independent pathway was suggested by the fact that inhibition of PKC (downregulation with phorbol 12,13-dibutyrate or exposure to staurosporine) failed to block PMA or flow-induced PDGF B-chain expression. These results demonstrate flow-induced PDGF B-chain expression in endothelial cells that appears to be mediated, in part, by a PKC-independent pathway.

Animals↗

PubMed1993

Flow-related responses of intracellular inositol phosphate levels in cultured aortic endothelial cells.

In vitro and in vivo evidence indicates that hemodynamic wall shear stress evokes a diversity of biological responses in vascular endothelial cells, ranging from cell shape changes to alterations in low density lipoprotein receptor expression. The signal transduction mechanisms by which the level of fluid mechanical shear stress is recognized by the endothelial cell and translated into these diverse biological responses remain to be elucidated. The present study focuses on the association between the onset of elevated shear stress and activation of the phosphoinositide signal transduction pathway, as measured by the intracellular release of inositol phosphates, in cultured bovine aortic endothelial cells (BAECs). BAECs were seeded, grown to confluence on large polyester sheets, and preincubated with 0.3 microCi/ml [3H]inositol for 24 hours before insertion in parallel-plate flow chambers for exposure to high shear stress (HS) at 30 dynes/cm2 or low shear stress (LS) at < 0.5 dyne/cm2 for periods ranging from 15 seconds to 24 hours. The induction of HS was associated with an early, transient but significant increase (142%, HS/LS x 100%) in inositol trisphosphate (IP3) measured at 15 seconds of shear stress exposure followed by a major peak in IP3 (189%) observed at 5 minutes after HS onset. After these initial increases, IP3 levels returned to near resting levels within 30 minutes of continued HS exposure and then continued to decline to significantly lower (75%) levels relative to LS-treated cells within 4 hours and remained lower throughout the remainder of the 24-hour HS exposure. LS-treated cells exhibited no significant changes in inositol phosphate levels throughout the 24-hour exposure periods. Exposure of BAECs to shear stress of 60 dynes/cm2 resulted in an approximately fourfold increase in IP3 levels (396%) measured at 5 minutes, almost double the levels measured in cells exposed to 30 dynes/cm2 for 5 minutes. Pretreatment of BAECs for 30 minutes with 5 mM neomycin, an inhibitor of phosphoinositide metabolism, before HS exposure inhibited both the early increases in inositol phosphates and subsequent cell elongation and alignment observed in untreated BAECs simultaneously exposed to HS without inhibiting protein synthesis. These results indicate that the exposure of cultured BAECs to elevated wall shear stress is associated with an early biphasic IP3 increase followed by a resetting of intracellular inositol phosphate concentrations to levels below that observed in static cultured BAECs. Furthermore, neomycin inhibition of this IP3 response to shear stress is associated with an inhibition of one of the major endothelial biological responses to shear stress, i.e., cell shape change and orientation.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1993

Endothelial cell signaling and cytoskeletal changes in response to shear stress.

Endothelial cells exposed to a fluid shear stress both in vivo and in vitro show an alignment with the direction of flow and an elongation of cells with a concomitant reorganization of their F-actin microfilament network. Specialized regions of the plasma membrane known as focal contacts are sites of transmembrane linkages between the actin microfilament bundles and the extracellular matrix (ECM) where cytoskeletal organization and, hence, cellular morphological changes may be modulated. Focal contact-associated proteins such as fibronectin receptors, vitronectin receptors and vinculin were shown in this study to play specific roles in the modulation of the cytoskeletal and cell shape changes. Data is also presented suggesting that protein kinase C is part of the cellular signaling system involved in shear stress-induced cytoskeletal reorganization.

Animals↗

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↗