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Fluorescence microscopy performed on living cells is a valuable technique for elucidating patterns of cell growth in vitro over artificial biomaterials such as vascular grafts, and for in vivo studies such as identification and treatment of atherosclerotic plaques. Two fluorescent dyes of particular value for vital fluorescence studies are Rhodamine-123 and 3,3'-dioctadecylindocarbocyanine-labeled low density lipoprotein (DiI-LDL). We examined the toxicity of these two dyes and of dimethylsulfoxide (DMSO), a solvent used in Rhodamine-123 studies, on the growth of MRC5 human fetal fibroblasts in monolayer culture. Two parameters of cell growth were quantitated: Cell number (a measure of proliferation), and cell area (a measure of cell spreading), based on microscopic images obtained at the start and end of a 48-h growth period after brief exposure (0.5 h) to test solutions. We found that the recommended solvent for solubilization of Rhodamine-123, DMSO, caused cessation of cell proliferation and actual reduction in the area covered by adherent fibroblasts at concentrations of as low as 0.1% (vol:vol). Rhodamine-123 made up from an aqueous stock solution modestly retarded proliferation and spreading, and there was no significant effect of DiI-LDL on these parameters over prolonged periods of exposure (up to 24 h) in culture. These results demonstrate that the most toxic substance for growing fibroblasts was the solvent DMSO. We conclude that both the solvent vehicle and fluorescent dye should be carefully examined for potential toxicity before such dyes are used for vital fluorescence studies of living cells.

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Vital fluorescent staining of human endothelial cells, fibroblasts, and monocytes seeded on a variety of surfaces has been carried out to develop a method for studying cell growth and interactions on viable cells. Using special fluorochrome markers and monoclonal antibodies, it was possible to differentiate different cell types as they grew on polymer surfaces similar to ones that are presently used on artificial heart valves and vascular grafts.

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In 1959, a prosthetic mitral valve of flexible polyurethane with Teflon chordae tendineae was designed and fabricated. After a series of experiments in dogs carried out at the Clinic of Surgery at the National Heart Institute, on March 11, 1960, this valve was used as a total replacement of the mitral valve of a 44-year-old woman with mitral regurgitation. After an uneventful postoperative course, she was discharged from the hospital and did well thereafter, but died suddenly, presumably of an arrhythmia, 4 months after operation.

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Infraclavicular subclavian puncture may be performed with fluoroscopic observation of the needle trajectory. In 92 patients so implanted between July 1985 and May 1987 uneventful venous access was achieved in 90, one was unsuccessful and one patient had subcutaneous emphysema, a complication rate of 2.2%.

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Intraoperative closure of the median sternotomy after cardiac operations in patients with complications, including severe postoperative bleeding, impaired cardiac function caused by myocardial edema, and cardiac dilatation, may lead to a critical and possibly fatal deterioration of hemodynamic function. In an effort to prevent this complication, we delayed mediastinal closure in 15 patients, covering the wound temporarily with a sheet of rubber latex (Esmarch bandage). An oval patch of this material was sized and sutured to the skin edges with a continuous suture. This technique provided easy and fast access to the mediastinal structures in four of the 15 patients who required multiple surgical interventions in the early postoperative period. Delayed closure was indicated for severe bleeding in 10 patients, heart compression in four patients, and severe postbypass arrhythmias in one patient. Definitive closure of the chest was delayed until satisfactory hemostasis was achieved or the heart size returned to normal. Thirteen of the 15 patients were long-term survivors, none of them had wound infections, and their wounds healed well. Delayed closure of the median sternotomy was an effective and safe approach in these groups of critically ill patients.

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Both electrocautery and internal defibrillation are routinely used in cardiac operations. The cases of 5 patients are presented in whom backup-mode reversion or injury of permanently implanted pulse generators occurred during a cardiac procedure. The theoretical explanations for backup-mode reversion and generator or tissue injury are discussed, and recommendations are made for the management of patients with a pacemaker who are to undergo a cardiac operation.

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Intramyocardial pH and temperature data recorded in 100 patients undergoing cardiac operations were analyzed to elucidate the effects of ventricular fibrillation and reflow. All patients underwent a single period of aortic clamping. Systemic hypothermia (25 degrees C) and intermittent cold crystalloid K+ cardioplegia were employed for myocardial protection. Baseline myocardial pH was 6.88 +/- 0.03 at a temperature of 36.5 degrees +/- 0.2 degree C. During the period of hypothermic ventricular fibrillation prior to aortic clamping, ventricular fibrillation did not affect myocardial pH in 45 patients (Group 1). In 21 patients (Group 2), it caused a significant drop in intramyocardial pH despite cooling. Group 2 patients had a higher incidence of valvular heart disease and left ventricular hypertrophy. They also exhibited low intramyocardial pH values during the subsequent periods of aortic clamping and reflow, indicating inadequate myocardial protection. During the period of reflow, reperfusion acidosis (pH less than 6.8 at 32 degrees C) was encountered in 39 patients (Group B) as opposed to 37 patients (Group A) whose pH remained well above 6.8 during that period. Group B patients had a higher incidence of valvular heart disease and left ventricular hypertrophy, tended to have more ischemic anterior walls prior to cardiopulmonary bypass, sustained longer periods of aortic clamping, had intramyocardial pH evidence of suboptimal protection during aortic clamping, were affected more adversely by ventricular fibrillation during reflow, and tended to have a higher operative mortality. Thus: Depending on the underlying myocardial disease, the adequacy of protection during aortic clamping, and the conditions of reflow, intramyocardial pH in man can fall significantly during ventricular fibrillation and reflow. The metabolic correlate of injury with reflow is a reperfusion acidosis that can reach as low as pH 5.98. When encountered, reperfusion acidosis can be minimized by prompt defibrillation.

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Interrelationships among silicone poppet wear, cloth wear, and tissue ingrowth were investigated in 14 retrieved Braunwald-Cutter heart valve prostheses following implantation of 37 to 118 (mean 83) months. Six aortic valves (mean 81 months) had severe cloth and poppet wear. In three the poppet had escaped. The lesser wear of the strut covering on the eight mitral valves (mean 84, range 37 to 108 months) was generally functionally insignificant. Mean decrease in mitral poppet diameter was 0.4% (range 0% to 1.5%), in contrast to a mean of 5.8% for aortic poppets. Histologic examination of the cloth/tissue complex demonstrated well-collagenized tissue ingrowth in areas of intact fabric with focal endothelial lining. Functionally trivial calcific deposits were often noted deep in the tissue coating, adjacent to cloth fibers or the strut metal. These results suggest that the mitral Braunwald-Cutter prosthesis need not be electively replaced without specific indication. A model is presented which explains the favorable clinical course demonstrated for mitral recipients and provides a rationale for the disparate clinicopathological behavior of mitral and aortic Braunwald-Cutter prostheses. Although inconsequential in this setting, the focal microcalcification noted in all mitral prostheses implanted for more than 72 months may have implications for the development of clinical cardiac assist devices for long-term application.

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Intramyocardial pH and temperature were continuously measured in the anteroseptal region in 40 patients undergoing aortic cross-clamping during cardiac operations. Myocardial protection was achieved with systemic cooling (25 degrees C) and multidose potassium cardioplegia (4 degrees C). A clinical myocardial preservation score was devised based on intraoperative and postoperative need for inotropic support, postoperative creatine kinase isoenzyme (CK-MB) and electrocardiographic changes, and radionuclide ventriculography. The patients were divided into three groups according to their preservation scores. Group I (n = 17) with good preservation (scores 0 to 2), Group II (n = 15) with fair preservation (scores 3 to 8), and Group III (n = 8) with poor preservation (scores 9 to 15). Baseline intramyocardial pH was similar in all groups (mean +/- SEM = 6.77 +/- 0.03). With the administration of cold potassium cardioplegia, intramyocardial pH rose above baseline in all three groups. The magnitude of this rise related directly to the adequacy of preservation and to the duration of the cross-clamp period. Patients with lowest preservation scores and shortest cross-clamp periods had the highest intramyocardial pH. In contrast, there was no relationship between myocardial temperature during cross-clamp and either intramyocardial pH or the preservation score. The integrated mean intramyocardial pH during cross-clamp was found to be the parameter that correlated most with the adequacy of preservation. The correlation between intramyocardial pH and myocardial temperature during the period of cross-clamping related to the length of this period; it was good (r = 0.76, p less than 0.01) in periods of 40 minutes or less and very poor in periods exceeding 60 minutes (r = 0.27, p greater than 0.10). It is concluded that (1) the magnitude of rise in intramyocardial pH during the period of aortic cross-clamping is a good indicator of the adequacy of myocardial preservation; (2) during periods of aortic cross-clamping exceeding 40 minutes, myocardial temperature is a poor indicator of adequacy of preservation, since progressive tissue acidosis may occur despite low myocardial temperatures; and (3) techniques and solutions that can effectively reduce the progression of tissue acidosis will, in most likelihood, enhance our ability to protect the ischemic myocardium during cardioplegic arrest.

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Femorofemoral arterial crossover grafts were used in three patients to avoid or correct limb ischemia during intra-aortic balloon counterpulsation. In retrospec, these grafts should have been used in two earlier patients in whom the balloon was removed prematurely. They are applied to the femoral artery just distal to the site of insertion of the balloon or the the balloon access graft. Removal of the graft at the time of the balloon removal is optional.

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Experimental studies carried out in our laboratory suggest that it is possible to develop a family of stent-supported tissue valve substitutes suitable for use in tissue and annuli of the hearts of small children in tissue annulus sizes ranging from 12 to 22 mm. Either glutaraldehyde-preserved, stent-supported primate tissue aortic valves or tissue leaflet valves constructed from dura mater preserved in 98 per cent glycerine can be used. In both instances, hemodynamics assessment of the valve substitutes in a mock circulation indicated that function was acceptable at the cardiac outputs normal for infants and children during the first few years of life. Stent-supported dura mater valves 16 nm. in diameter have been used to replace the mitral valve in 2 infants 7 and 8 months of age with complete atrioventricular canal defects and mitral regurgitation, one of whom survives with demonstratable satisfactory hemodynamic function of the valve substitute.

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It is now possible to construct vascular grafts of expanded polytetrafluoroethylene having complex configurations. In this series of experiments a double tapered graft was used to create a bypass shunt between the aorta and the pulmonary artery in rhesus monkeys and between the aorta and the iliac artery in dogs. When the synthetic material was placed under a slight degree of tension, a high degree of patency was achieved in animals killed three to six months following implantation. The use of this approach in neonates with cyanotic congenital heart disease necessitating the creation of systemic-pulmonary artery shunts is suggested as a new method to achieve a controlled lumen size and an increased patency rate, while simultaneously decreasing the incidence of congestive heart failure or of late stricture of the pulmonary artery.

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A technique for accelerating the development of tissue layers on fabrics implanted in the bloodstream has been explored in dogs and monkeys. Endothelial cell seeding of the fabrics with cells derived from the endocardial surface of the atrium together with an autogenous plasma coating was shown to accelerate organization of fabric-covered devices implanted in the bloodstream of adult primates. At the same time, this technique appears simple and rapid enough to be practical in a clinical setting. The results suggest that this approach might be useful in accelerating the development of tissue layers on prosthetic valves implanted in the heart.

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Eighty-eight operations for correction of intracardiac congenital heart defects were performed using local cardiac hypothermia for protection of the ischemic myocardium. Twenty-six patients underwent repair of tetralogy of Fallot, 23 had patch closure of ventricular septal defect, 24 had correction of various types of congenital aortic stenosis, and 15 were operated upon for other complex lesions. The overall operative mortality was 5.6%. Ischemia times ranged from 9 to 119 minutes (mean, 48 minutes). Ischemic arrest protected by local cardiac hypothermia provides an optimal operative field, permitting repair of uncomplicated intracardiac defects in a precise, unhurried manner. No hemodynamic abnormalities attributable to the technique were encountered.

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