Friedrich Wilhelm Hehrlein

Gene Polymorphism but not Catalytic Activity of Angiotensin I–Converting Enzyme Is Associated With Coronary Artery Disease and Myocardial Infarction in Low-Risk Patients

BACKGROUND An insertion/deletion (I/D) polymorphism of the angiotensin I-converting enzyme (ACE) gene has been postulated to be associated with an increased risk of coronary artery disease (CAD) and myocardial infarction (MI). METHODS AND RESULTS In the present study, the effects of I/D gene polymorphism and of ACE activity on CAD and MI were investigated in 920 individuals who underwent coronary angiography for diagnostic purposes. In the total population and in all CAD and MI groups, a strong association was observed between the gene polymorphism and ACE activities; DD genotypes had approximately twofold higher ACE activities than II genotypes. Although classic risk and protective factors of CAD and MI were identified, associations of ACE genotype and of ACE activity to CAD and MI were not detected in the total population. Among subjects defined to be at lower risk of MI by low body mass index and low cigarette consumption, however, an association of the DD genotype with MI was found. Exclusion of individuals with triglyceride levels > 140 mg/dL and cholesterol levels > 180 mg/dL revealed an association of the DD genotype with CAD. An association of the ACE activity with CAD or MI could not be demonstrated in any of the low-risk populations. CONCLUSIONS Increased ACE activity obviously is not a risk factor of CAD or MI. The importance of the deletion polymorphism for the development of CAD and MI may be restricted to individuals without classic risk factors.

ACE I:D gene polymorphism: presence of the ACE D allele increases the risk of coronary artery disease in younger individuals

BACKGROUND Presence of the D allele or homozygosity for the deletion (D) allele of the ACE insertion/deletion (I/D) polymorphism has been discussed as potent risk factor for coronary artery disease (CAD) and myocardial infarction (MI). METHODS AND RESULTS In 2267 male Caucasians the relation of the ACE I/D gene polymorphism to CAD and MI were investigated. An association of the D allele to CAD was detected in younger subjects (e.g. < 61.7 years, mean value), but not in older patients (e.g. > or = 61.7 years). Additional exclusion of individuals with other cardiovascular risk factors (e.g. high BMI) produced an even stronger association of the D allele to CAD. In contrast, a relation of this polymorphism to non-fatal MI was only observed in older subjects; additional limitation to individuals without cardiovascular risk factors (e.g. BMI and/or diabetes) yielded a further enhancement of this association to MI. In younger subjects (e.g. < 61.7 years) the gene polymorphism was not related to non-fatal MI even after exclusion of additional risk factors. CONCLUSIONS The present large case-control study strengthens the assumption of an association of the ACE D allele with the risk of ischemic heart disease.

Angiotensinogen T174M and M235T gene polymorphisms are associated with the extent of coronary atherosclerosis

BACKGROUND The relations of the angiotensinogen (AGT) T174M and M235T gene polymorphisms to the risk of coronary heart disease (CHD) have been investigated in only a few studies with conflicting results. RESULTS Therefore, we analysed the relationship of the AGT gene polymorphisms to the presence and extent of CHD in 2250 male Caucasians whose coronary anatomy was defined by means of coronary angiography. The relative frequencies of the T and M alleles of the T174M and of the M235T gene variation did not significantly differ between patients without or with single-, double- or triple-vessel disease and between subjects without or with myocardial infarction (MI). In contrast the mean CHD score--defined by Gensini--was higher within MM homozygotes of the T174M gene variation than within TT genotypes; TM subjects had intermediate values. In M235T genotypes, mean CHD scores were similar in the total sample and in older individuals (> or = 62 years), whereas in younger individuals (< 62 years) a higher CHD score was found within AGT 235 T allele carriers than within MM homozygotes. In younger individuals with high apoAI plasma levels, the mean CHD score was clearly higher within TT homozygotes of the M235T gene variation than within MM genotypes; MT subjects had intermediate values. An interaction between both angiotensinogen gene polymorphisms on the extent of CHD or on the risk of non-fatal MI were not observed when the M allele of AGT T174M was combined either with the T allele or the TT genotype of M235T. CONCLUSIONS The present study strengthens the hypothesis of an association of both angiotensinogen gene polymorphisms with the extent of coronary heart disease.

Increased expression of disintegrin-metalloproteinases ADAM-15 and ADAM-9 following upregulation of integrins α5β1 and αvβ3 in atherosclerosis

Regulation of αvβ3 and α5β1 integrin function plays a crucial role in atherosclerosis. Possible regulators of integrin–matrix interactions are integrin‐binding ADAMs (proteins with a disintegrin‐ and metalloproteinase‐domain), like ADAM‐15 and ADAM‐9. Molecular interactions between ADAM‐15, α5β1, and αvβ3 have been demonstrated. ADAM‐9 and ADAM‐15 were found to be interdependently regulated. This study, therefore, investigated whether the upregulation of integrins α5β1 and αvβ3 was correlated with the expression of integrin‐binding ADAMs in atherosclerotic processes. Human arterial and venous vascular smooth muscle cells (VSMCs) were incubated with PDGF over different time intervals up to a 3‐day culture period. mRNA concentrations, quantified by real‐time RT‐PCR and normalized to PBGD, of integrins αvβ3 and α5β1 were strongly increased after a 12‐h PDGF‐incubation in arterial and venous VSMC. ADAM‐15 and ADAM‐9 mRNA production showed a corresponding increase following integrin upregulation after a 24‐h incubation period. Western blot anaylsis revealed an increased protein expression of integrins and ADAMs in PDGF‐stimulated VSMC. Additionally, mRNA concentrations of atherosclerotic and normal human specimens were quantified by real‐time RT‐PCR. mRNA of ADAMs and integrins was significantly increased in atherosclerotic arteries compared to normal arteries. Immunohistochemistry of these specimens showed an increased expression and codistribution of both ADAMs and integrins in atherosclerosis. In conclusion, upregulation of ADAM‐15 and ADAM‐9 in atherosclerosis appears to follow an increase in α5β1 and αvβ3 integrins. Since α5β1 and αvβ3 are known to promote smooth muscle cell migration and proliferation, upregulation of ADAM‐15 and ADAM‐9 could balance integrin–matrix interactions and cell migration, thus modulating neointima progression. J. Cell. Biochem. 89: 808–823, 2003.

Insulin and local growth factor PDGF induce intimal hyperplasia in bypass graft culture models of saphenous vein and internal mammary artery.

OBJECTIVE In arteriosclerosis and bypass graft stenosis, intimal proliferation is controlled by local and systemic growth factors, such as platelet derived growth factor (PDGF) or insulin. Intimal hyperplasia can be produced in organ culture models. Our aim was to compare neointima formation in two organ culture models of internal mammary artery (IMA) and saphenous vein (SV), with special reference to the influence of systemic and local growth stimuli. METHODS Rings of freshly isolated human SV and IMA were cultured over a 3-, 6- or 8-day period. They were distributed into five groups of incubation protocols: incubation with 10% serum; insulin 50 ng/ml and 100 ng/ml; PDGF-BB 5 ng/ml and 10 ng/ml. Frozen sections of cultured rings and pre-culture segments were subjected to elastic stain and immunohistochemistry. Antibodies directed against beta-actin and smooth muscle alpha-actin were used to characterize smooth muscle cell phenotype and against proliferating cell nuclear antigen (PCNA) to demonstrate proliferating cells. RESULTS Growth factor incubation caused massive intimal hyperplasia with increased elastic fibers in SV and intimal smooth muscle cell as well as matrix accumulation in IMA. Intimal thickening, PCNA and beta-actin expression reached their maximum on day 6 of culture. In both culture models, serum, insulin and PDGF caused increasing intimal thickening, with more pronounced effects in SV. CONCLUSIONS These organ culture models demonstrate the effects of insulin and PDGF on intimal hyperplasia in IMA and SV representing models for arteriosclerosis and bypass graft stenosis and stressing the role of insulin and growth factors for neointima development.

Association of the insertion/deletion gene polymorphism of the apolipoprotein B signal peptide with myocardial infarction.

The Del allele of the apolipoprotein B (apoB) signal peptide (SP) insertion/deletion (Ins/Del) polymorphism has been shown to be associated with elevated plasma levels of apoB, cholesterol and low density lipoprotein. It was the aim of the present study to analyse the relation of this gene variation to the risk of coronary artery disease (CAD) and of myocardial infarction (MI) in a population of 2259 male Caucasians, whose coronary anatomy was defined by means of coronary angiography. ApoB SP DelDel genotypes had significantly higher apoB plasma concentrations than InsIns homozygotes (P = 0.0001) and InsDel heterozygotes (P = 0.002); however, the apoB plasma levels of InsIns and InsDel genotypes were essentially the same (P = 0.54). Similar observations were made with respect to ApoB SP genotype-dependent cholesterol plasma concentrations. Since the apoB plasma level was not only associated with the apoB SP Ins/Del gene variation but also to the extent of coronary artery disease (P <0.0001), individuals with an InsIns genotype and without CAD had the lowest and subjects with a DelDel genotype and triple vessel disease the highest apoB plasma levels (P <0.0001). An association of the apoB SP Ins/Del gene variation with CAD was not detected, neither in the total population nor in low risk groups. In contrast, the gene variation was associated with MI (P <0.05). An Odds ratio of 1.18 (95% CI, 1.01-1.39) associated with the Del allele was detected in the total sample (P <0.02). In a subpopulation of individuals with low plasma triglyceride levels ( <154 mg/dl; mean value) and an DD genotype of the angiotensin I-converting enzyme insertion/deletion gene polymorphism an Odds ratio of 2.01 (1.42-3.05) was calculated (P <0.001). The present study presents evidence for a statistically significant difference in the development of MI between genotype classes of the apoB SP Ins/Del gene polymorphism.

Selective cerebral perfusion via innominate artery in aortic arch replacement without deep hypothermic circulatory arrest

To attain satisfactory results in aortic arch surgery a reliable method of cerebral protection, avoidance of emboli, and control of hemorrhage is mandatory. Deep hypothermic circulatory arrest is the most common technique at present but gives only a limited period of protection, whereas a complicated aortic arch operation may require more time than anticipated. Therefore the selective cannulation and perfusion of the innominate artery has not been widely used until now because it is uncertain whether the left hemisphere of the brain is adequately perfused. Between 1990 and 1995, 21 of 69 patients within the last 36 months, consisting of 15 men and 6 women averaging 45±13.4 years, underwent operative treatment for aneurysm (n=9) or type A dissection (n=12) involving the aortic valve and aortic arch; selective innominate perfusion (SCP [i]) in moderate hypothermia (28°C) for brain protection was used.Extended perioperative monitoring included bilateral somatosensory-evoked potentials (SEP), transcranial Doppler sonography (TCD), a computer-aided topographical electroencephalometry (CATEEM), and analysis of the arterial and venous oxygen saturation and desaturation. Mean time periods were 229.7±56.5 minutes for extracorporeal circulation, 151.7±34.1 minutes for aortic cross-clamping, and 67.05±34.03 for selective cerebral perfusion via the innominate artery. Not once did the intraoperative monitoring reveal hints of cerebral damage due to inadequate perfusion. All patients survived surgery but two could not be weaned from the respirator; one died 2 days and the other 6 days after the operation due to multiple organ failure (MOF). Another two patients died after 13 days due to untreatable septic syndrome with pulmonary insufficiency. All four patients died within 30 days, during which time they had aortic dissection involving the complete aortic arch and severe aortic valvular incompetence (grade IV). There was no late death and follow-up time of 19.76±8.04 months revealed an overall mortality rate of 19%. Only temporary neurological affections (left-sided hemiparesis) were found in two patients (9.5%). Additionally, we observed neuropsychological disturbances in one of these.Our first experience with selective cerebral perfusion via innominate artery and the attendant CATEEM monitoring for assessment of adequate bilateral cerebral perfusion suggests that this method is a useful addition to the armamentarium in complicated aortic arch surgery.

Expression of sodium pump isoforms and other sodium or calcium ion transporters in the heart of hypertensive patients.

The sodium pump (Na(+),K(+)-ATPase; EC 3.6.1.37) of animal cell membranes is the enzyme responsible for the maintenance of membrane potential, for the function of secondary active transporters, and for osmoregulation of the cell. Since inhibition of the enzyme by cardiac glycosides results in increased contractility of the heart muscle and increased blood pressure, we were interested in whether there is a correlation between hypertension and expression of the various isoforms of the sodium pump. In addition, we also examined the expression of the isoforms of the sarcoplasmic and plasma membrane Ca(2+)-ATPase, the Na(+)/Ca(2+)- and Na(+)/H(+)-exchangers, and Na(+) channel and Ca(2+) channel isoforms. Total mRNA was isolated from 50 mg tissue from the right atrium of hypertensive and normotensive patients who were undergoing cardiac surgery. After reverse transcription and subsequent amplification of ion transporter-specific cDNA fragments by polymerase chain reaction (PCR) in the presence of [alpha-(32)P]dCTP, quantification of the amplified fragments was carried out by the Phosphorimager technique. The data obtained show that the alphal subunit mRNA is expressed similarly in normotensive and hypertensive patients. The amount of alpha2 subunit mRNA, however, is increased 5-fold in hypertensive patients. In the same group, the amount of alpha3 isoform is also significantly increased, although not as dramatically as the alpha2 isoform. Besides the Na(+),K(+)-ATPase isoforms, a significant increase in the expression of mRNA for the Na(+)/Ca(2+)-exchanger and the plasma membrane Ca(2+)-ATPase isoforms was detected. It is possible that the observed changes in mRNA expression for these ion transporters reflect compensatory mechanisms to overcome a defective Na(+) and Ca(2+) metabolism in the tissues of hypertensive patients or reflect defects directly involved in the cause of hypertension. The expression of mRNA for all other transporters investigated was unaltered.

The fibrin-stabilizing factor as a topical means for leg ulcer healing: Biochemical and experimental properties and clinical results

The topical application of factor XIII (Fibrogammin HS) is a new concept in the treatment of ulcerative leg disease. In addition to the mode of application, the utilization of a fibrin-stabilizing effect in ulcer treatment is a completely different strategy, since many other medications used for local wound treatment contain a fibrinolytic component. Of the 29 inpatients treated topically during the last 3 years, 17, 12 female and 5 male averaging 60.8±13.6 years, suffered ulcerative leg disease due to a postthrombotic syndrome. The average period since first clinical manifestation of venous ulcer was 3.5±2.1 years. Thirteen patients (76.4%) with chronic ulceration showed such a distinct improvement of topical site after an average period of 3.02±1.05 weeks with topically applied factor XIII (Fibrogammin HS) combined with compression bandaging that they were discharged for further ambulant treatment. Apart from a wound surface reduction and a clinical improvement of granulation, we observed a marked reduction of bleeding and secretion tendency within the ulcus area, especially in patients with severe venous insufficiency. Our hypothesis was that high doses of topically applied factor XIII could improve barrier function of endothelial cells in the wound area. For experimental investigation, porcine aortic endothelial cells were cultured on filter membranes to confluent monolayers. To monitor the barrier function, permeability of the monolayer was continuously measured as flux of trypan blue-labeled albumin. Confluent endothelial monolayers exposed to factor XIII (Fibrogammin HS) showed a significant (n=8,p<0.05) reduction of albumin flux compared with controls. After induction of hyperpermeability in endothelial monolayers by incubation with potassium cyanide (KCN) and 2-deoxyglucose (2-DG), we found a significant increase (up to 250%) in albumin flux compared with controls (n=8,p<0.05). This hyperpermeability was significantly reduced to 50% (n=8,p<0.05) in monolayers that had been preincubated with Fibrogammin HS. We conclude that F XIII reduces permeability of endothelial monolayers. This effect is due to the stabilization of the endothelial barrier function and likely caused by cross-linking of suitable substrates from endothelial cell membrane or extracellular matrix. Because of our good clinical results in improvement of granulation and reduction of secretion during topical treatment of venous ulcer and the demonstrable effect on endothelial permeability, F XIII may play an important role in wound healing of chronic ulcerative leg disease.

Coronary Endarterectomy in Patients with Diffuse Coronary Disease

Diffuse multivessel disease can limit a complete revascularization in patients undergoing coronary bypass surgery. It is considered that coro nary endarterectomy (CEA) offers a reasonable chance for this special group of patients. Among 2415 patients undergoing coronary bypass surgery, a CEA of the right and/or left coronary system had to be performed in 397 patients with diffuse multivessel disease. Completeness of the procedure was controlled intraoperatively by dye perfusion and angioscopy. One hun dred one patients gave informed con sent to be reinvestigated clinically and by angiography at an average of forty-three months after surgery. The thirty days mortality after additional CEA was slightly in creased at 3.8%. Also the periopera tive infarction rate increased at 9.3 % . The long-term analysis re vealed a late mortality of 5.5%. Cor onary angiography showed a patency rate of the endarterectomized vessels of 81.2 %; 91% of the patients being reinvestigated showed an improved or asymptomatic clinical status. Despite a slightly increased risk, CEA proved to be a feasible therapy in this strictly limited group of pa tients, in whom conventional bypass surgery offers poor chances and who otherwise would even be candidates for transplantation.