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Inhibitors of angiotensin-converting enzyme prevent myointimal proliferation after vascular injury
Volume
12
Number 2 August 1990
Abstmzs
out loss of efficacy. The results demonstrate that heparin and Fragmin have similar antithrombotic efficacy for similar anti-Xa levels measured ex vivo. The observed risk reduction for bleeding with Fragmin therapy, although not statistically significant, might still offer a clinically important benefit that may be proved in larger studies. Joseph R. Durham, MD The Ohio State Univenity
College
ofMedicine
Recombinant gene expression in vivo within endotheliaI cells of the arterial wall Nabel EG, Plautz G, Boyce FM, Stanley JC, Nabel GJ. Science 1989;244: 1342-4. It has been proposed that genetically altered endothelial cells seeded in vivo could be used to transmit recombinant deo xyribonucleic acid products. This approach could provide anticoagulant, vasodilating or growth factor influence at local sites, or systemic intravascular secretion of gene products. In this article the authors present their initial experience with in vivo seeding of retrovirus-altered endothelial cells into the iliac artery of inbred Yucatan minipigs by use of a catheter delivery system. After establishing a primary endothelial cell line (jugular vein), subsets of cells were infected with a murine B-galactosidasetransducing retrovirus, which is replication defective. These cells could then be differentiated from noninfected cells by histochemical staining based on B-galactosidase expression. Syngeneic cells were introduced into the iliac arteries of nine minipigs after balloon catheter and protease denudation of the native endothelial cells. Seeded cells were incubated for 30 minutes in a protected space in the artery created by the inflation of proximal and distal balloons on a special catheter that contained an injection port between the balloons for cell installation. Local thrombosis was prevented by aspirin pretreatment and heparin anticoagulation at the time of inoculation. Nuclide labeling indicated that 2% to 11% of 2 x lo5 endothelial cells successfully attached to the arterial wall. After 2 to 4 weeks B-galactosidase staining was present on the intimal surface of all the retroviral infected, experimentally seeded vessels. This innovative study demonstrates that localized endothelial seeding on normal iliac arteries is possible after balloon catheter denudation of the native intima, and that molecular engineering can be used to modify the gene expression of these seeded cells. Since atherosclerosis can be readily established in the minipig, this model will allow future study of abnormal arterial surfaces. Seeded cells continued to express B-galactosidase up to 4 weeks after implantation, although it has not yet been established how long such cells will remain active before being replaced by native endothelial cells. Replication-defective retrovintses were successfully used in this study to eliminate the potential concern of replicating competent retroviruses after transplantation in vivo. This article provides a glimpse into the promising application of molecular medicine to pa-
225
tients with vascular disease, and it is reassuring to note the involvement of vascular surgeons in this effort. Jack L. Cronenwett, Dartnzouth-Hitchcock
A4D Medical
Center
Inhibitors of angiotensin-converting enzyme prevent myointimal proliferation after vascular injury Powell JS, Clozel J-P, Muller RKM, et al. Science 1989:245:186-S. This work represents another addition to the long series of carefully executed projects that we have come to expect from Powell et al. at Hoffmann-La Roche laboratories. The primary observation in this study was an impressive reduction in myointimal proliferation of deendothelialized rat carotid arteries when the angiotensinconverting enzyme inhibimr cilazapril was given. Cilazapril was administered 6 days before balloon injury of common carotid arteries, and neointimal development was measured by use of morphometric analysis with computerized digitalization 14 days after injury. Neointimal area in rats treated with cilazapril was less than one fifth that of control animals. The reduction in neointimal growth was most significant when cilazapril was begun before the production of endothelial damage and continued throughout the 14 day period after injury. Whereas most investigators would stop with this empiric data, this group continued investigation into the mechanism of myointimal suppression. The possibility of a direct inhibition of neointimal formation by cilazapril itself was eliminated through observation of a lack of cultured smooth muscle cell proliferation in media containing the drug or its active metabolites. The possibility that the observations were a result of the hypotensive actions of the medication was discounted by experiments with verapamil, wherein similar reductions in arterial pressure were achieved without corresponding effects on myointimal proliferation. Despite the elegance of this study, the mechanism of reduction in myointimal proliferation has not been fully elucidated. Although the authors state that their data support the hypothesis of a local angiotensin system modulating the myointimal proliferative response of the vascular wall, one may hypothesize other equally sound mechanisms for these observations. For instance, is it possible that the reduction in angiotensin to levels by cilazapril resulted in decreased spasm in the small rat carotid arteries, with corresponding reduction in shear rates and decreased platelet deposition on the deendothelialized surface? This reduction in platelet deposition might explain decreased myointimal proliferation on the basis of decreased plateletderived growth factor release at the luminal surface. In summary, although the exact mechanism of the authors’ observations remain obscure, the constraining effect of angiotensin-converting enzyme inhibition on myointima1 proliferation is impressive. Further studies are war-
ranted so that the effect of these agents may eventually evaluated in a controlled clinical trial. Kenneth Ouriel, University
be
MD
of Rochester
Medical
Center
Carotid arteriosclerosis in identica.I for cigarette smoking Haapanen A, Koskenvuo M, Kaprio Heikkila K. Circulation 1989;8: 10-6.
twins
discordant
J, Kesaniemi
YA,
The authors report an in vitro experimental study in which a 20 MHz pulsed Doppler was used to detect flow disturbances and define velocity profiles in a prosthetic conduit under conditions of a steady flow with varying velocities. Both Fast-Fourier transform spectrum analysis and frequency tracking with a demodulator were evaluated for their ability to identify and quantitate flow disturbances and turbulence. To identify the onset of flow disturbances, ensemble averaging was used with spectrum analysis, whereas root mean square fluctuations were used with the frequency tracker. Velocity profiles were calculated from spectrum analysis by use of the Doppler equation. The authors report that the frequency tracking method was preferred for detection and quantitation of flow disturbances because of its ease of use and greater versatility. This technique is intended to be applied to the study of flow disturbances and turbulence in small diameter arterial prostheses, and the authors postulate that these factors are important in the pathogenesis of graft thrombosis. Although the paper is replete with technical detail, the relationship of these flow disturbances to graft thrombosis remains speculative, particularly in this steady flow system. In a pulsatile in vivo flow system with varying outflow resistance and systemic blood pressure, it is likely that the events surrounding graft thrombosis are much more complex and may bear little relation to the presence of minor degrees of flow disturbance in the conduit itself. In addition, the authors have not clearly demonstrated that this research instrument provides any advantage over more widely available commercial Doppler devices, which can be used in vivo. Nevertheless, the article should be of interest to those involved in research to flow mechanics and prosthetic grafts and the ultrasonic detection and quantitation of flow disturbances.
This interesting article has selected a clever method to examine the influence of life-long smoking on the development of carotid arteriosclerosis. The study originated in Finland where the Finnish Twin Cohort compromises all Finnish sex-matched twin pairs born before 1958 and alive in 1967. From 78 identical pairs (61 male and 17 female) with the highest discordance in the life-long dose of cigarette smoking, the authors were able to recruit 49 pairs with a mean age of 52 years. Smoking history was obtained in 1975, 1981, and 1986. The mean life-long smoking dose of the smoking co-twins was 20 pack-years. The smoking and nonsmoking co-twins had similar blood pressures, total plasma cholesterol, body mass index, and psychosocial factors. A significant difference was found in the use of alcohol, which was greater among smoking individuals. Duplex sonography of the carotid arteries was then performed with a 7.5 MHz real-time transducer and a pulsed Doppler. Carotid artery stenosis (narrowing of area of the lumen from 15% to 60%) was found in significantly more smoking twins than nonsmoking co-twins (18% vs 4%). Not only was the total area of carotid plaque larger in smoking co-twins, but the thickness of the inner layer of the carotid arteries was more marked in smoking co-twins. The size of piaques and the degree of inner layer thickening correlated with the dose of smoking but was not statistically significant. The association of smoking with carotid arteriosclerosis was highly significant even after the adjustment for age, total plasma cholesterol level, diastolic blood pressure, and body mass index in multiple logistic regression analyses. The authors emphasize that rhe life-long dose of smoking in the current sample was clearly lower than that of the age- and sex-matched Finnish population. Consequently, they feel that their results probably underestimate the real effect of smoking on carotid arteriosclerosis. The importance of this study lies in its novel method of studying the association of cigarette smoking and carotid arteriosclerosis and in the results that indicate that smoking is a strong factor in the development of carotid atheroma independent of genetic factors.
William M. Blackhar, Jr., MD Univemity of South Florida ColLe8e of Medicine
John W. Hallett, Mayo Clinic
Pulsed Doppler ultrasound system for the measurement of velocity distributions and flow disturbances in arterial prostheses Black RA, How TV. J Biomed Eng 1989;11:35-42.
Jr., MD
12
Number 2 August 1990
Abstmzs
out loss of efficacy. The results demonstrate that heparin and Fragmin have similar antithrombotic efficacy for similar anti-Xa levels measured ex vivo. The observed risk reduction for bleeding with Fragmin therapy, although not statistically significant, might still offer a clinically important benefit that may be proved in larger studies. Joseph R. Durham, MD The Ohio State Univenity
College
ofMedicine
Recombinant gene expression in vivo within endotheliaI cells of the arterial wall Nabel EG, Plautz G, Boyce FM, Stanley JC, Nabel GJ. Science 1989;244: 1342-4. It has been proposed that genetically altered endothelial cells seeded in vivo could be used to transmit recombinant deo xyribonucleic acid products. This approach could provide anticoagulant, vasodilating or growth factor influence at local sites, or systemic intravascular secretion of gene products. In this article the authors present their initial experience with in vivo seeding of retrovirus-altered endothelial cells into the iliac artery of inbred Yucatan minipigs by use of a catheter delivery system. After establishing a primary endothelial cell line (jugular vein), subsets of cells were infected with a murine B-galactosidasetransducing retrovirus, which is replication defective. These cells could then be differentiated from noninfected cells by histochemical staining based on B-galactosidase expression. Syngeneic cells were introduced into the iliac arteries of nine minipigs after balloon catheter and protease denudation of the native endothelial cells. Seeded cells were incubated for 30 minutes in a protected space in the artery created by the inflation of proximal and distal balloons on a special catheter that contained an injection port between the balloons for cell installation. Local thrombosis was prevented by aspirin pretreatment and heparin anticoagulation at the time of inoculation. Nuclide labeling indicated that 2% to 11% of 2 x lo5 endothelial cells successfully attached to the arterial wall. After 2 to 4 weeks B-galactosidase staining was present on the intimal surface of all the retroviral infected, experimentally seeded vessels. This innovative study demonstrates that localized endothelial seeding on normal iliac arteries is possible after balloon catheter denudation of the native intima, and that molecular engineering can be used to modify the gene expression of these seeded cells. Since atherosclerosis can be readily established in the minipig, this model will allow future study of abnormal arterial surfaces. Seeded cells continued to express B-galactosidase up to 4 weeks after implantation, although it has not yet been established how long such cells will remain active before being replaced by native endothelial cells. Replication-defective retrovintses were successfully used in this study to eliminate the potential concern of replicating competent retroviruses after transplantation in vivo. This article provides a glimpse into the promising application of molecular medicine to pa-
225
tients with vascular disease, and it is reassuring to note the involvement of vascular surgeons in this effort. Jack L. Cronenwett, Dartnzouth-Hitchcock
A4D Medical
Center
Inhibitors of angiotensin-converting enzyme prevent myointimal proliferation after vascular injury Powell JS, Clozel J-P, Muller RKM, et al. Science 1989:245:186-S. This work represents another addition to the long series of carefully executed projects that we have come to expect from Powell et al. at Hoffmann-La Roche laboratories. The primary observation in this study was an impressive reduction in myointimal proliferation of deendothelialized rat carotid arteries when the angiotensinconverting enzyme inhibimr cilazapril was given. Cilazapril was administered 6 days before balloon injury of common carotid arteries, and neointimal development was measured by use of morphometric analysis with computerized digitalization 14 days after injury. Neointimal area in rats treated with cilazapril was less than one fifth that of control animals. The reduction in neointimal growth was most significant when cilazapril was begun before the production of endothelial damage and continued throughout the 14 day period after injury. Whereas most investigators would stop with this empiric data, this group continued investigation into the mechanism of myointimal suppression. The possibility of a direct inhibition of neointimal formation by cilazapril itself was eliminated through observation of a lack of cultured smooth muscle cell proliferation in media containing the drug or its active metabolites. The possibility that the observations were a result of the hypotensive actions of the medication was discounted by experiments with verapamil, wherein similar reductions in arterial pressure were achieved without corresponding effects on myointimal proliferation. Despite the elegance of this study, the mechanism of reduction in myointimal proliferation has not been fully elucidated. Although the authors state that their data support the hypothesis of a local angiotensin system modulating the myointimal proliferative response of the vascular wall, one may hypothesize other equally sound mechanisms for these observations. For instance, is it possible that the reduction in angiotensin to levels by cilazapril resulted in decreased spasm in the small rat carotid arteries, with corresponding reduction in shear rates and decreased platelet deposition on the deendothelialized surface? This reduction in platelet deposition might explain decreased myointimal proliferation on the basis of decreased plateletderived growth factor release at the luminal surface. In summary, although the exact mechanism of the authors’ observations remain obscure, the constraining effect of angiotensin-converting enzyme inhibition on myointima1 proliferation is impressive. Further studies are war-
ranted so that the effect of these agents may eventually evaluated in a controlled clinical trial. Kenneth Ouriel, University
be
MD
of Rochester
Medical
Center
Carotid arteriosclerosis in identica.I for cigarette smoking Haapanen A, Koskenvuo M, Kaprio Heikkila K. Circulation 1989;8: 10-6.
twins
discordant
J, Kesaniemi
YA,
The authors report an in vitro experimental study in which a 20 MHz pulsed Doppler was used to detect flow disturbances and define velocity profiles in a prosthetic conduit under conditions of a steady flow with varying velocities. Both Fast-Fourier transform spectrum analysis and frequency tracking with a demodulator were evaluated for their ability to identify and quantitate flow disturbances and turbulence. To identify the onset of flow disturbances, ensemble averaging was used with spectrum analysis, whereas root mean square fluctuations were used with the frequency tracker. Velocity profiles were calculated from spectrum analysis by use of the Doppler equation. The authors report that the frequency tracking method was preferred for detection and quantitation of flow disturbances because of its ease of use and greater versatility. This technique is intended to be applied to the study of flow disturbances and turbulence in small diameter arterial prostheses, and the authors postulate that these factors are important in the pathogenesis of graft thrombosis. Although the paper is replete with technical detail, the relationship of these flow disturbances to graft thrombosis remains speculative, particularly in this steady flow system. In a pulsatile in vivo flow system with varying outflow resistance and systemic blood pressure, it is likely that the events surrounding graft thrombosis are much more complex and may bear little relation to the presence of minor degrees of flow disturbance in the conduit itself. In addition, the authors have not clearly demonstrated that this research instrument provides any advantage over more widely available commercial Doppler devices, which can be used in vivo. Nevertheless, the article should be of interest to those involved in research to flow mechanics and prosthetic grafts and the ultrasonic detection and quantitation of flow disturbances.
This interesting article has selected a clever method to examine the influence of life-long smoking on the development of carotid arteriosclerosis. The study originated in Finland where the Finnish Twin Cohort compromises all Finnish sex-matched twin pairs born before 1958 and alive in 1967. From 78 identical pairs (61 male and 17 female) with the highest discordance in the life-long dose of cigarette smoking, the authors were able to recruit 49 pairs with a mean age of 52 years. Smoking history was obtained in 1975, 1981, and 1986. The mean life-long smoking dose of the smoking co-twins was 20 pack-years. The smoking and nonsmoking co-twins had similar blood pressures, total plasma cholesterol, body mass index, and psychosocial factors. A significant difference was found in the use of alcohol, which was greater among smoking individuals. Duplex sonography of the carotid arteries was then performed with a 7.5 MHz real-time transducer and a pulsed Doppler. Carotid artery stenosis (narrowing of area of the lumen from 15% to 60%) was found in significantly more smoking twins than nonsmoking co-twins (18% vs 4%). Not only was the total area of carotid plaque larger in smoking co-twins, but the thickness of the inner layer of the carotid arteries was more marked in smoking co-twins. The size of piaques and the degree of inner layer thickening correlated with the dose of smoking but was not statistically significant. The association of smoking with carotid arteriosclerosis was highly significant even after the adjustment for age, total plasma cholesterol level, diastolic blood pressure, and body mass index in multiple logistic regression analyses. The authors emphasize that rhe life-long dose of smoking in the current sample was clearly lower than that of the age- and sex-matched Finnish population. Consequently, they feel that their results probably underestimate the real effect of smoking on carotid arteriosclerosis. The importance of this study lies in its novel method of studying the association of cigarette smoking and carotid arteriosclerosis and in the results that indicate that smoking is a strong factor in the development of carotid atheroma independent of genetic factors.
William M. Blackhar, Jr., MD Univemity of South Florida ColLe8e of Medicine
John W. Hallett, Mayo Clinic
Pulsed Doppler ultrasound system for the measurement of velocity distributions and flow disturbances in arterial prostheses Black RA, How TV. J Biomed Eng 1989;11:35-42.
Jr., MD