Gianni D. Angelini

Effects of external stenting on wall thickening in arteriovenous bypass grafts

Late occlusion of the saphenous vein graft appears to result in part from wall thickening as an adaptation to increased mean wall stress. Using an established pig model of arteriovenous bypass grafting, the effect of reducing wall stress with an external porous polytetrafluoroethylene stent was investigated. Segments of autologous saphenous vein were implanted by end-to-end anastomoses into both carotid arteries, with one graft supported by a stent 4 mm in diameter. Increases in graft wall dimensions were quantified 4 weeks later by computer-aided planimetry of transverse histological sections. The contribution of hyperplasia (i.e., cell proliferation) to the changes observed was further clarified by measurements of DNA concentration. All grafts showed an increase in external size, but this was restricted by stenting. All grafts also showed an increase in cross-sectional area of the tunica media and tunica intima that was only partly accounted for by an increase in DNA concentration, which indicated that both hyperplasia and hypertrophy had occurred. Stented grafts showed less enlargement of the media but greater enlargement of the intima. Overall wall size was therefore similar in stented and unstented grafts. Stented grafts showed less increase in DNA concentration than unstented grafts. In stented grafts, the residual luminal cross-sectional area was significantly less than in unstented grafts. The data show that external stenting reduces medial enlargement and hyperplasia but increases encroachment of the intima into the lumen. Because final luminal size is thought to be of paramount importance in maintaining long-term patency, external stenting is unlikely to be of benefit.

Comparison of response to injury in organ culture of human saphenous vein and internal mammary artery

Autologous saphenous vein grafts, unlike internal mammary artery grafts, suffer many late occlusions as a result of excessive proliferation of vascular smooth muscle cells and the superimposition of atheroma on the resulting thickened intima. We investigated the possible basis of this difference using organ cultures. Internal mammary artery segments and freshly isolated and surgically prepared saphenous vein segments were obtained from patients undergoing coronary artery bypass grafting. Internal mammary artery and freshly isolated vein segments showed a high degree of endothelial coverage and medial cell viability that were maintained during culture. Surgically prepared veins showed partial endothelial denudation and medial cell injury, both of which tended to be reversed during culture. Neointimal thickening was greater in surgically prepared vein (72 +/- 13 microns; n = 11) than in freshly isolated vein (44 +/- 8 microns; n = 10) or internal mammary artery (34 +/- 4 microns; n = 13) segments. The occurrence of proliferating cells in the medial layer was also significantly greater in surgically prepared vein (2.8 +/- 1.0/mm; n = 11) than in freshly isolated vein (0.8 +/- 0.3/mm; n = 9) or internal mammary artery (0.6 +/- 0.3/mm; n = 10) segments. The data show that although the smooth muscle proliferation was similar in undamaged saphenous vein and internal mammary artery, it was significantly greater in damaged vein. This implies that the greater intimal proliferation seen in saphenous vein grafts may arise not from intrinsic differences in arterial and venous smooth muscle cells but from a greater susceptibility to injury.

Should Thyroid Function Be Assessed Before Cardiopulmonary Bypass Operations

Thyroid function is depressed during and after cardiopulmonary bypass surgical procedures, and this may adversely affect myocardial performance. There is known to be a high prevalence of thyroid abnormalities in the elderly population, and many patients undergoing cardiac operations fall into this category. We have assessed thyroid function in 116 patients admitted for elective open heart procedures to determine the value of preoperative testing. Abnormalities in thyroid function were present in 13 (11.2%) of the patients studied, 3 of whom were receiving thyroxine therapy. One patient who had overt hypothyroidism died postoperatively of a large cerebral infarct; 11 had elevated thyrotropin levels with normal serum thyroxine levels; and 1 who had overtreated hypothyroidism suffered fast atrial fibrillation postoperatively. No other complications were observed. These findings indicate that thyroid function should be assessed preoperatively in patients already on thyroxine therapy. Whether thyroid function should be evaluated routinely in all patients before operations involving cardiopulmonary bypass is not clear. Although there is a high incidence of abnormal laboratory results, there were no apparent adverse effects on the surgical outcome.

Adverse hemodynamic effects of pericardial closure soon after open heart operation

The short-term hemodynamic effects of pericardial closure on cardiac function were studied during steady-state anesthesia and ventilation in 10 patients (6 men) (mean age, 59 +/- 9 years) who underwent an open-heart valve operation. Observations were made after the heart was decannulated, both while the pericardium was open and after it had been closed, and then after closure of the chest after the pericardium had been reopened by removing the pericardial suture through the chest wall. The effect of closing the pericardium before closing the chest was an immediate reduction in cardiac output (thermodilution) of 1.39 +/- 0.24 L/min from 5.09 +/- 0.40 L/min (p less than 0.001). The heart rate remained stable, but there was a decrease in stroke volume of 29% and an increase in systemic vascular resistance of 34% (both, p less than 0.01). The mean arterial pressure increased slightly by 2% (not significant). Opening the pericardium (1.5 to 2 hours after the end of the operation) while the chest remained closed was followed by an increase in cardiac output of 1.33 +/- 0.15 L/min from 4.12 +/- 0.62 L/min (p less than 0.001). As the heart rate and the mean blood pressure changed insignificantly, there was an increase in stroke volume of 15 +/- 3 mL from 53 +/- 5 mL and a reduction in systemic vascular resistance of 473 +/- 83 dyne . s . cm-5 from 1,721 +/- 181 dyne.s.cm-5 (both, p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

A surgical technique that preserves human saphenous vein functional integrity

We investigated the effect of a novel surgical preparative technique for human saphenous vein by using the concentration of adenosine triphosphate and the adenosine triphosphate/diphosphate ratio to quantify medial integrity and by using stimulated rates of prostacyclin production to quantify endothelial function. Freshly isolated vein had an adenosine triphosphate concentration of 358 +/- 54 nmol.g-1 wet weight and an adenosine triphosphate/diphosphate ratio of 2.89 +/- 0.13 (n = 12); it produced prostacyclin in response to fluid shear at a rate of 14.3 +/- 2.0 pg.min-1.mg-1 wet weight (n = 12). Surgically prepared vein obtained on completion of the last proximal anastomosis had been distended with the patients own arterial pressure using a side-arm connected to the aortic cannula. This vein had an adenosine triphosphate concentration of 413 +/- 70 nmol.g-1 wet weight and an adenosine triphosphate/diphosphate ratio of 2.74 +/- 0.44 (n = 11), and it produced prostacyclin at a rate of 13.1 +/- 0.2 pg.min-1.mg-1 wet weight (n = 12). All values were indistinguishable from those in freshly isolated vein. The results demonstrate that this simple technique for distention at arterial pressure preserved medial and endothelial function.

Serum-Free organ culture of vascular tissues

Dear Editor: The aim of organ culture is to retain the original structural relationship of different and similar cells and their interactive function in order to study the effect of exogenous stimuli on further development (4). However, most organ culture systems described in the literature have a high requirement for serum and other undefined biological fluids (6,7) which can create problems when assessing the role of growth factors in regulating tissue growth and differentiation. To overcome this we attempted to develop a serum-free organ culture of vascular tissue. This was then used to evaluate the role of growth factors and cytokines in pathological conditions such as coronary atherosclerosis, restonosis after angioplasty and coronary saphenous vein bypass graft failure. Growth factors released from cells intrinsic to the vessel wall (5) are believed to cause smooth muscle cells to proliferate and secrete extracellular matrix components which contribute to vessel wall thickening and luminal narrowing. Our hypothesis is that during short-term serum-free culture if vessel viability is maintained, soluble bioeffectors such as peptides may be released into the medium. Analysis of the conditioned media for mitogenic activity can then be performed. We report here our experience of short-term serum-free organ culture of human coronary arteries and porcine arteriovenous bypass grafts. To our knowledge, there is no previous evidence in the literature on organ culture of vascular tissue in a chemically defined media free from both serum and hormones. Human coronary arteries were obtained from the discarded hearts of transplant recipients (3) and saphenous vein grafts from a pig model of myointimal hyperplasia developed in our laboratory (1). Vessels were carefully dissected and immediately transported to the laboratory (23 ° C) in sterile RPMI 1640 culture media containing 20 mmol/liter HEPES buffer (Flow Labs, High Wycombe, UK) supplemented with penicillin (100/xg/ml), streptomycin (100 U/ml), gentamycin (2.5/,tg/ml), amphotericin (2.5 ttg/ml), glutamine (2 mM), all from Flow Labs, High Wycombe, UK and 4 U/ml whole heparin (CP Pharmaceuticals, Wrexham, UK). Vessels were then washed several times in RPMI 1640 medium, dissected free from excess fatty tissue and adventitia, cut open with scissors along their upper aspect and pinned open endothelial surface uppermost using minuten pins (Watkins and Doncaster, Kent) on top of a square of polyester gauze (2 cm X 1 cm) supported on sylgard resin in the base of a glass petri dish (3,7). Artery and graft segments were then cultured for 24 hours in 6 ml of RPMI 1640 medium containing 2 g/liter sodium bicarbonate (Flow Labs, High Wycombe, UK) supplemented with antibiotics as above. Cultures were maintained at 37 ° C in a humidified atmosphere of 5% v/v CO2 in air and labeled with [3H]thymidine [1 #Ci/ml (80 Ci/mmol), Amersham International, UK] at the start of the culture period in order to assess and locate cell proliferation. In a separate experiment arteries and grafts were cultured in medium containing 30% fetal bovine serum (J-Bio Chemicals, France). Segments of artery and graft were snap frozen in liquid nitrogen (following preparation for culture and also after culture) for measurement of adenosine nucleotide concentrations using high performance liquid chromatography (2). Tissue ATP concentrations and ATP/ADP ratios which have previously been used to quantify the viability of vascular tissue (2,3,7) were unchanged in arteries and grafts following culture in serumfree medium or in medium containing 30% fetal bovine serum (Table 1). Transmission electron microscopy of ultra-thin sections confirmed these findings. An internal elastic lamina separated a monolayer of endothelial cells from several layers of medial smooth muscle cells and extracellular matrix. No necrotic areas were seen. Cell proliferation (DPM/ttgDNA) occurred to a similar extent in vessels cultured in the presence and absence of serum (human coronary arteries; 596 + 80 (n = 14) and 450 + 50 (n = 9), pig grafts; 892.8 _+ 113 (n -17)and 948.7 + 339 (n = 6), both n.s. Autoradiography of cultured arteries and grafts showed dividing cells predominantly in the intima with only occasional dividing cells in the media. Conditioned media from the serum-free experiments was stored at 8 0 ° C and its mitogenic potential investigated using a Swiss 3T3 fibroblast bioassay by determining the incorporation of 1/.tCi/ ml [aH]tbymidine (80 Ci/mmol) into a trichloroacetic acid insoluble cell fraction. Briefly, cells (ATCC CCL 92, Flow Labs, High Wy-

Organ culture of human coronary artery following balloon angioplasty

Intimal smooth muscle cell proliferation is the primary cause of restenosis following balloon angioplasty. Its underlying basis and progression remain unclear. The authors developed an organ culture of human coronary artery subjected to balloon angioplasty in order to investigate the cellular and molecular basis of intimal proliferation in a preparation that maintained the anatomic relationships of the vessel wall.Artery segments obtained from the explanted hearts of transplant recipients were maintained at 37°C in culture medium containing 30% fetal bovine serum for fourteen days. Balloon angioplasty produced partial endothelial denudation and medial smooth muscle cell damage, both of which tended to be reversed after fourteen days in culture. Transverse histologic sections of cultured artery showed the development of a new intima containing smooth muscle cells identified by immunocytochemistry with anti-α-actin. Labeling of cultures with [3H] thymidine showed proliferating cells in the neointima.The data demonstrate that intimal proliferation occurs in organ culture of human coronary artery subjected to balloon angioplasty. They also suggest the possibility that the smooth muscle cells in the neointimal layer are the result of both migration and proliferation.

Congenital ventricular septal defect presenting as rupture of the ventricular septum subsequent to myocardial infarction

A 74-year-old female presented in cardiogenic shock four weeks following a severe episode of ischaemic chest pain. Physical examination was suggestive of a ventricular septal defect, and this was confirmed by cross-sectional echocardiography and right heart catheterisation. As the resting electrocardiogram was consistent with an extensive anterior myocardial infarct of indeterminate age, a diagnosis of ventricular septal defect subsequent to infarction was made and the patient taken to the operating theatre for urgent repair. At operation, however, the defect was found to be congenital in origin.