Drosophila wing development in the absence of dorsal identity.

The developing wing disc of Drosophila is divided into distinct lineage-restricted compartments along both the anterior/posterior (A/P) and dorsal/ventral (D/V) axes. At compartment boundaries, morphogenic signals pattern the disc epithelium and direct appropriate outgrowth and differentiation of adult wing structures. The mechanisms by which affinity boundaries are established and maintained, however, are not completely understood. Compartment-specific adhesive differences and inter-compartment signaling have both been implicated in this process. The selector gene apterous (ap) is expressed in dorsal cells of the wing disc and is essential for D/V compartmentalization, wing margin formation, wing outgrowth and dorsal-specific wing structures. To better understand the mechanisms of Ap function and compartment formation, we have rescued aspects of the ap mutant phenotype with genes known to be downstream of Ap. We show that Fringe (Fng), a secreted protein involved in modulation of Notch signaling, is sufficient to rescue D/V compartmentalization, margin formation and wing outgrowth when appropriately expressed in an ap mutant background. When Fng and alphaPS1, a dorsally expressed integrin subunit, are co-expressed, a nearly normal-looking wing is generated. However, these wings are entirely of ventral identity. Our results demonstrate that a number of wing development features, including D/V compartmentalization and wing vein formation, can occur independently of dorsal identity and that inter-compartmental signaling, refined by Fng, plays the crucial role in maintaining the D/V affinity boundary. In addition, it is clear that key functions of the ap selector gene are mediated by only a small number of downstream effectors.

Chip is an essential cofactor for apterous in the regulation of axon guidance in Drosophila.

LIM-homeodomain transcription factors are expressed in subsets of neurons and are required for correct axon guidance and neurotransmitter identity. The LIM-homeodomain family member Apterous requires the LIM-binding protein Chip to execute patterned outgrowth of the Drosophila wing. To determine whether Chip is a general cofactor for diverse LIM-homeodomain functions in vivo, we studied its role in the embryonic nervous system. Loss-of-function Chip mutations cause defects in neurotransmitter production that mimic apterous and islet mutants. Chip is also required cell-autonomously by Apterous-expressing neurons for proper axon guidance, and requires both a homodimerization domain and a LIM interaction domain to function appropriately. Using a Chip/Apterous chimeric molecule lacking domains normally required for their interaction, we reconstituted the complex and rescued the axon guidance defects of apterous mutants, of Chip mutants and of embryos doubly mutant for both apterous and Chip. Our results indicate that Chip participates in a range of developmental programs controlled by LIM-homeodomain proteins and that a tetrameric complex comprising two Apterous molecules bridged by a Chip homodimer is the functional unit through which Apterous acts during neuronal differentiation.

Axon routing across the midline controlled by the Drosophila Derailed receptor.

In nervous systems with symmetry about the midline, many neurons project axons from one side to the other. Although several of the components controlling midline crossing have been identified, little is known about how axons choose the appropriate pathway when crossing. For example, in the Drosophila embryo axons cross the midline in one of two distinct tracts, the anterior or posterior commissure (AC or PC, respec tively). Here we show that the Derailed (Drl) receptor tyrosine kinase is expressed by neurons that project in the AC, and that in the absence of Drl such neurons often project abnormally into the PC. Conversely, misexpression of Drl in PC neurons forces them to cross in the AC. The behaviour of Drl-misexpressing neurons and the in vivo binding pattern of a soluble Drl receptor probe indicate that Drl acts as a guidance receptor for a repellent ligand present in the PC. Our results show that Drl is a novel component in the control of midline crossing.

The LIM homeodomain protein dLim1 defines a subclass of neurons within the embryonic ventral nerve cord of Drosophila.

Members of the LIM homeodomain family of transcription factors have been implicated in specifying cell identity in a range of species. In Drosophila three LIM homeobox genes, apterous, lim3 and isl, have been shown to control axon pathfinding of subsets of neurons within the embryo. Here we describe the isolation and characterization of another LIM homeobox gene in Drosophila termed dlim1, a homolog of the vertebrate Lim-1 gene. The sequence and expression of dLim1 is highly related to its vertebrate homologs. Within the Drosophila embryo, dLim1 is expressed in the head primordia, the brain lobes, and in distinct sets of motorneurons and interneurons within the ventral nerve cord. Comparatively in vertebrates, Lim-1 (Lhx1) along with Lim-3 (Lhx3), Gsh-4 (Lhx4), Isl-1 and Isl-2 are expressed in developing motorneurons along the spinal column, where their overlapping expression suggests a role for these genes in the establishment of specific motorneuron subtypes. dLim1 is absent from all cells expressing Isl, Lim3, and Apterous, indicating that these proteins function independently within the Drosophila embryo. To investigate the function of dlim1, we generated loss-of-function mutations within the locus. Our findings show that dlim1 is an essential gene that when mutated results in lethality near the larval-pupal boundary. In contrast to vertebrate Lim-1, dlim1 has no apparent role in anterior patterning of the Drosophila embryo. Our analysis shows that dlim1 has been evolutionarily conserved, however the Drosophila lim1 gene exhibits unique properties that distinguishes it from its vertebrate homologs.

Chip and apterous physically interact to form a functional complex during Drosophila development.

LIM homeodomain (LIM-HD) proteins play key roles in a variety of developmental processes throughout the animal kingdom. Here we show that the LIM-binding protein Chip acts as a cofactor for the Drosophila LIM-HD family member Apterous (Ap) in wing development. We define the domains of Chip required for LIM-HD binding and for homodimerization and show that mutant proteins deleted for these domains act in a dominant-negative fashion to disrupt Ap function. Our results support a model for multimeric complexes containing Chip and Ap in transcriptional regulation. This model is confirmed by the activity of a chimeric fusion between Chip and Ap that reconstitutes the complex and rescues the ap mutant phenotype.

Function and specificity of LIM domains in Drosophila nervous system and wing development.

LIM domains are found in a variety of proteins, including cytoplasmic and nuclear LIM-only proteins, LIM-homeodomain (LIM-HD) transcription factors and LIM-kinases. Although the ability of LIM domains to interact with other proteins has been clearly established in vitro and in cultured cells, their in vivo function is unknown. Here we use Drosophila to test the roles of the LIM domains of the LIM-HD family member Apterous (Ap) in wing and nervous system development. Using a rescuing assay of the ap mutant phenotype, we have found that the LIM domains are essential for Ap function. Furthermore, expression of LIM domains alone can act in a dominant-negative fashion to disrupt Ap function. The Ap LIM domains can be replaced by those of another family member to generate normal wing structure, but LIM domains are not interchangeable during axon pathfinding of the Ap neurons. This suggests that the Ap LIM domains mediate different protein interactions in different developmental processes, and that LIM domains can participate in conferring specificity of target gene selection.

Benefits of glucose and oxygen in multidose cold cardioplegia.

We tested the effects of glucose and oxygen in cardioplegic solutions on myocardial protection in the isolated perfused working rat heart. Recovery from 2 hours' hypothermic (8 degrees C) cardioplegic arrest was examined in 93 hearts. Cardioplegic solution, which was delivered every 15 minutes, was supplemented with glucose 28 mmol/L as a substrate or sucrose 28 mmol/L as a nonmetabolizable osmotic control; it was equilibrated with either 98% oxygen or 98% nitrogen, both with 2% carbon dioxide. Four combinations of hyperkalemic cardioplegic solution were studied: nitrogen-sucrose, nitrogen-glucose, oxygen-sucrose, and oxygen-glucose. During hypothermic arrest, oxygenation of cardioplegic solution greatly reduced myocardial lactate production and prevented ischemic contracture as indicated by coronary vascular resistance. Glucose increased lactate production modestly but significantly only when the cardioplegic solution was nitrogenated. Although end-arrest myocardial adenosine triphosphate and creatine phosphate were greatly increased by oxygenation of cardioplegic solution (p less than 0.005), we could not detect improved preservation of these high-energy phosphates by glucose. Averaged over reperfusion, percent recovery of cardiac output for the nitrogen-sucrose, nitrogen-glucose, oxygen-sucrose, and oxygen-glucose solutions was 32.3% +/- 6.1%, 45.9% +/- 4.6%, 44.5% +/- 4.6%, and 62.2% +/- 4.5%, respectively. Oxygenation of the glucose solution or addition of glucose to the oxygenated solution significantly improved recovery of cardiac output. The benefits of glucose and oxygen were additive, so that the oxygen-glucose cardioplegic solution provided the best functional recovery. We conclude that the addition of glucose to the fully oxygenated multidose cold cardioplegic solution improves functional recovery without increasing lactate production during arrest.

Protection of the hypertrophied myocardium by crystalloid cardioplegia.

Patients with left ventricular hypertrophy (LVH) have a worse outcome after cardiac surgery than those without hypertrophy. We studied protection of hearts with LVH in an isolated rat heart model using multidose, cold, oxygenated cardioplegia. LVH was produced by banding the abdominal aorta in young rats. Six weeks after banding, this produced a 31% increase in the left ventricular dry weight/body weight ratio compared to two age-matched control groups comprising sham-operated and nonoperated animals. The recovery of cardiac output after arrest was higher in LVH (82 +/- 4% of prearrest) than in sham-operated (69 +/- 4%) or nonoperated (66 +/- 3%) control groups. The improved functional recovery in LVH occurred although there were no differences among the groups in myocardial adenosine triphosphate (ATP) and phosphocreatine (PCr) prior to arrest, at the end of arrest, or after reperfusion. Glycogen levels were also similar among the three groups prior to arrest and after reperfusion but were highest in LVH after arrest. Myocardial oxygen consumption (MVO2) and efficiency, expressed as cardiac output/MVO2, were similar among the groups prior to arrest. Myocardial efficiency after reperfusion declined in all groups but was best preserved in LVH. We also compared the sensitivity of hypertrophied and control hearts to the deleterious effects of calcium in cardioplegia. Calcium in the cardioplegia increased myocardial lactate production during arrest in a dose-related fashion and depressed myocardial levels of ATP, PCr, and glycogen at end arrest in all groups. Cardiac output recovery was also depressed by calcium but was still best in LVH. We conclude that the hypertrophied myocardium is well protected by standard cardioplegia and that calcium in cardioplegia does not preferentially depress recovery in LVH.

Relation of myocardial protection to cardioplegic solution pH: modulation by calcium and magnesium.

The relationship between myocardial preservation and cardioplegic solution pH was assessed in isolated, perfused rat hearts. A base solution without calcium or magnesium and the same solution containing 0.2 mmol/L ionized calcium or 16 mmol/L magnesium or both ions were studied at several values of pH between 6.8 and 8.7. Hearts were arrested at 8 degrees C by multidose infusions of these bicarbonate-buffered solutions bubbled with oxygen and a varying percentage of carbon dioxide to control pH. Diastolic tone (left ventricular balloon) and adenosine triphosphate (ATP) depletion during arrest both increased as the cardioplegic solution became more alkaline. Calcium increased these effects of pH. Magnesium weakened the effect of pH on diastolic tone, maintained ATP at all pH levels, and inhibited the effects of calcium on the relationships of pH to diastolic tone and ATP. When data from all solutions were considered together, ATP depletion was shown to be linearly related to diastolic tone. Calcium depressed functional recovery (left ventricular developed pressure during reperfusion expressed as a percentage of its prearrest value) at all pH levels. With the other solutions, recovery was similar and best within a broad and relatively alkaline pH range. With the solution containing calcium and magnesium, at pH levels of 8.28 +/- 0.02, 7.87 +/- 0.03, 7.58 +/- 0.02, 7.41 +/- 0.01, 7.06 +/- 0.02, and 6.80 +/- 0.01, recovery at 5 minutes of reperfusion was 101.4% +/- 3.7%, 102.9% +/- 2.8%, 107.3% +/- 3.7%, 102.8% +/- 2.9%, 91.8% +/- 3.6%, and 94.3% +/- 3.5%, respectively. This effect of alkalinity was short-lived. Extreme alkalinity of the base, acalcemic solution produced the calcium paradox, as reported previously. Good preservation of ATP by the most acid solutions did not predict good functional recovery. Magnesium increased the persistence of frequent extrasystoles during early reperfusion, but the effect was attenuated by calcium. The data support the inclusion of magnesium in cardioplegic solutions, particularly when they contain calcium, show that cardioplegic solution pH can have major effects on the arrested heart, and suggest that a relatively alkaline pH may modestly benefit functional recovery.

The effects of calcium and magnesium in hyperkalemic cardioplegic solutions on myocardial preservation.

Sustained left ventricular pressure development during each infusion of a cold calcium-containing hyperkalemic cardioplegic solution has been observed in rat hearts. The present study was undertaken to relate such contraction (i.e., increase in resting pressure) to myocardial preservation and to the calcium and magnesium contents of a crystalloid hyperkalemic cardioplegic solution. Isolated perfused rat hearts with a left ventricular isovolumic balloon were arrested at 8 degrees C by the fully oxygenated cardioplegic solution infused every 15 minutes for 2 hours. Cardioplegic solutions containing ionized calcium in concentrations of 0, 0.1, or 1.2 mmol/L were each studied with (groups 2, 4, and 6) and without (groups 1, 3, and 5) the addition of magnesium (16 mmol/L). Hearts arrested by the cardioplegic solution with no calcium or magnesium (group 1) developed a pressure (averaged over the second to eighth infusion and expressed as percent prearrest left ventricular pressure) of 6.0% +/- 0.4% during cardioplegic infusions. This solution maintained end-arrest myocardial adenosine triphosphate (13.1 +/- 1.0 nmol/mg dry weight) and phosphocreatine (21.7 +/- 2.8 nmol/mg dry weight) contents near the prearrest contents and preserved left ventricular function at 95% +/- 3% of prearrest developed left ventricular pressure at 15 minutes of reperfusion at 37 degrees C. Calcium (groups 3 and 5) increased pressure development during cardioplegic infusions (10.4% +/- 0.5% and 15.1% +/- 0.9%), depleted adenosine triphosphate (7.2 +/- 1.0 and 7.4 +/- 0.9) and phosphocreatine (13.3 +/- 1.8 and 10.7 +/- 1.5), and depressed left ventricular functional recovery (71% +/- 1% and 73% +/- 3%). Magnesium alone (group 2) decreased pressure development during cardioplegic infusions (3.0% +/- 0.3%), maintained adenosine triphosphate (15.6 +/- 0.9), augmented phosphocreatine (38.3 +/- 1.2), and preserved left ventricular function (99% +/- 4%). Magnesium added to calcium (groups 4 and 6) prevented the calcium-induced increased pressure development during cardioplegic infusions (4.0% +/- 0.5% and 6.7% +/- 0.6%), maintained adenosine triphosphate (13.6 +/- 1.4 and 14.9 +/- 0.7), augmented phosphocreatine (31.3 +/- 1.6 and 32.2 +/- 2.4), and ameliorated the depression of functional recovery (82% +/- 2% and 86% +/- 2%). These data suggest that left ventricular pressure development during arrest contributed to calcium-induced energy depletion and impairment of functional recovery and that these deleterious effects were inhibited by magnesium. The inhibitory effects of magnesium on left ventricular pressure development were rapidly reversed on reperfusion. The data support the addition

Myocardial preservation related to magnesium content of hyperkalemic cardioplegic solutions at 8 degrees C.

This study investigates whether the addition of magnesium to a hyperkalemic cardioplegic solution containing 0.1 mM ionized calcium improves myocardial preservation, and whether there is an optimal magnesium concentration in this solution. Isolated perfused rat hearts were arrested for two hours by this cardioplegic solution, which was fully oxygenated and infused at 8 degrees C every 15 minutes to simulate clinical conditions. The cardioplegic solution contained either 0, 2, 4, 8, 16, or 32 mM magnesium. At end-arrest, the myocardial creatine phosphate concentration (nanomoles per milligram of dry weight) was 20.7 +/- 2.1, 22.9 +/- 1.7, 24.8 +/- 2.0, 31.3 +/- 1.4, 33.1 +/- 1.8, and 31.6 +/- 0.8, respectively, in hearts given cardioplegic solution containing these magnesium concentrations. Thus, the concentration of creatine phosphate was significantly higher at end-arrest when the cardioplegic solution contained 8, 16, or 32 mM than 0 or 2 mM magnesium (p less than 0.002) or 4 mM magnesium (p less than 0.02), and highest with 16 mM magnesium. Also, creatine phosphate was more sensitive to the magnesium concentration of the cardioplegic solution than was end-arrest adenosine triphosphate levels, which did not differ among the experimental groups. Aortic flow, expressed as a percentage of prearrest aortic flow, was 60.3 +/- 5.0, 70.2 +/- 5.5, 71.6 +/- 4.4, 71.8 +/- 4.8, 81.0 +/- 5.0, and 71.8 +/- 5.3, respectively. The addition of magnesium to the cardioplegic solution improved recovery of aortic flow (p less than 0.05, 16 mM versus 0 mM magnesium). We conclude from these data that with deep myocardial hypothermia and at an ionized calcium concentration of 0.1 mM, the addition of magnesium, over a broad concentration range, improved preservation of myocardial creatine phosphate and, at a concentration of 16 mM, improved aortic flow. The optimal magnesium concentration in the cardioplegic solution was 16 mM.

Calcium-induced ventricular contraction during cardioplegic arrest.

Cardiac arrest induced by hyperkalemic perfusion is generally considered to represent a state of complete electromechanical arrest. However, high-energy phosphate concentrations and ventricular function decrease with increasing cardioplegic calcium concentrations, possibly because of elevated resting muscle tone produced by calcium influx. We examined isolated rat hearts containing an isovolumic intraventricular balloon for the presence of contractile activity during the administration at 10 degrees C of a cardioplegic solution containing potassium, 20 mEq/L. Significant left ventricular pressure was developed (35.6% +/- 4.3% of prearrest systolic pressure) during administration of a solution containing a calcium concentration of 1.0 mmol/L and far less (9.7% +/- 1.6% of prearrest systolic pressure) with a calcium-free cardioplegic solution. The muscle contraction diminished with repeated doses, was increased by increasing cardioplegic calcium content, and was inhibited by magnesium. Adenosine triphosphate and creatine phosphate concentrations were 9.0 +/- 1.4 and 7.0 +/- 0.9 nmol/mg dry weight immediately after infusion of 15 ml of a hypoxic cardioplegic solution containing calcium, versus 13.3 +/- 1.3 (p less than 0.02) and 31.9 +/- 3.5 nmol/mg dry weight (p less than 0.0001) after a hypoxic acalcemic solution was given. When repeated doses of a hypoxic cardioplegic solution containing calcium in a concentration of 1.0 mmol/L were given at 15 minute intervals at 10 degrees C, ischemic contracture (a sustained development of ventricular pressure, mean 51% +/- 4% of prearrest systolic pressure) resulted within 1 hour. Coronary vascular resistance was increased during the muscle contractions induced by calcium-containing solutions, markedly so during contracture. Calcium-related mechanical activity was also observed during hypothermic cardioplegic arrest in five of six isolated isovolumic canine hearts. We conclude that hearts remain potentially active mechanically during cold hyperkalemic arrest and undergo energetically wasteful contraction when stimulated with calcium-containing hyperkalemic cardioplegic solutions.

Oxygenation of cardioplegic solutions. Potential for the calcium paradox.

Oxygenation of crystalloid cardioplegic solutions is beneficial, yet bicarbonate-containing solutions equilibrated with 100% oxygen become highly alkaline as carbon dioxide is released. In the isolated perfused rat heart fitted with an intraventricular balloon, we recently observed a sustained contraction related to infusion of cardioplegic solution. In the same model, to record these contractions, we studied myocardial preservation by multidose bicarbonate-containing cardioplegic solutions in which first the calcium content and then the pH was varied. An acalcemic cardioplegic solution (Group 1) and the same solution with calcium provided by adding calcium chloride (Group 2) or blood (Group 3) were equilibrated with 100% oxygen. Ionized calcium concentrations were 0, 0.10 +/- 0.06, and 0.11 +/- 0.07 mmol/L and pH values were 8.74 +/- 0.07, 8.54 +/- 0.08, and 8.40 +/- 0.07, all highly alkaline. Hearts were arrested for 2 hours at 8 degrees +/- 2.5 degrees C and reperfused for 1 hour at 37 degrees C. At end-arrest, myocardial adenosine triphosphate was depleted in all three groups, significantly in Groups 2 and 3. In Group 1 the calcium paradox developed upon reperfusion, with contracture (left ventricular end-diastolic pressure = 60 +/- 7 mm Hg), creatine kinase release up to 620 +/- 134 U/L, a profound further decrease in adenosine triphosphate to 1.9 +/- 1.7 nmol/mg dry weight, and either greatly impaired or no functional recovery (17% +/- 10% of prearrest developed pressure). Three hearts in this group released creatine kinase during arrest and did not resume beating during reperfusion. In Groups 2 and 3, the calcium paradox did not occur; functional recovery was 61% +/- 4% and 71% +/- 9% at 5 minutes of reperfusion. In two additional groups (4 and 5), the pH of the acalcemic cardioplegic solution was decreased by equilibration with 2% and 5% carbon dioxide in oxygen to 7.53 +/- 0.03 and 7.11 +/- 0.02. Contractions during arrest were smaller than in Groups 1, 2, and 3; adenosine triphosphate was maintained during arrest; functional recovery was 101% +/- 3% and 96% +/- 4% at 5 minutes of reperfusion. We conclude that acalcemic solutions with carbon dioxide are superior to highly alkaline calcium-containing solutions. If oxygenation of cardioplegic solutions, of proved value, causes severe alkalinity, then calcium paradox may result even with hypothermia. This hazard is prevented by adding calcium or blood to the solution or carbon dioxide to the oxygen used for equilibration.

Maximal oxygenation of dilute blood cardioplegic solution.

The content of dissolved O2 (the major source of O2 for the myocardium) of dilute blood cardioplegic solution (dBCS) varied widely when oxygenated at 4 degrees C by surface flow of O2 in a Bentley BCR-3500 cardiotomy reservoir. We have modified the system to consistently deliver maximally oxygenated dBCS to the heart. Laboratory studies indicated that bubbling O2 through a 16-gauge intravenous catheter in a central Luer-Lok port of the cardiotomy reservoir provided contents of dissolved O2 that were consistently near maximal. We then studied 17 patients in the operating room. The first 6 patients received dBCS oxygenated with 100% O2 with a high dissolved O2 content of 3.2 +/- 0.2 ml/dl. However, the pH of the dBCS became highly alkaline (7.83 +/- 0.11 at 37 degrees C). Therefore, in the remaining 11 patients, 2% CO2 was added to the O2. The dissolved O2 content remained high (3.3 +/- 0.1 ml/dl), and the pH was in a more physiological range (7.35 +/- 0.09 at 37 degrees C). We conclude that consistently maximal oxygenation of a dBCS at a more physiological pH can be achieved by this method.

Optimal myocardial preservation with an acalcemic crystalloid cardioplegic solution.

The effect of the calcium and oxygen contents of a hyperkalemic glucose-containing cardioplegic solution on myocardial preservation was examined in the isolated working rat heart. The cardioplegic solution was delivered at 4 degrees C every 15 minutes during 2 hours of arrest, maintaining a myocardial temperature of 8 degrees +/- 2 degrees C. Hearts were reperfused in the Langendorff mode for 15 minutes and then resumed the working mode for a further 30 minutes. Groups of hearts were given the oxygenated cardioplegic solution containing an ionized calcium concentration of 0, 0.25, 0.75, or 1.25 mmol/L or the same solution nitrogenated to reduce the oxygen content and containing 0 or 0.75 mmol ionized calcium per liter. The myocardial adenosine triphosphate concentrations at the end of arrest in these six groups of hearts were 15.6 +/- 1.2, 9.5 +/- 0.5, 8.2 +/- 1.1, 4.9 +/- 1.8, 10.1 +/- 2.0, and 1.6 +/- 0.4 nmol/mg dry weight, respectively. At 5 minutes of working reperfusion, the percentages of prearrest aortic flow were 80 +/- 2, 62 +/- 4, 33 +/- 6, 37 +/- 5, 48 +/- 7 and 46 +/- 8, respectively. The differences among the groups in adenosine triphosphate concentrations and in functional recovery diminished during reperfusion. In hearts given the hypoxic calcium-containing solution, there was a marked increase in coronary vascular resistance during the administration of successive doses of cardioplegic solution, which was rapidly reversible upon reperfusion. These data indicate that hearts given the acalcemic oxygenated solution had better adenosine triphosphate preservation during arrest and better functional recovery than hearts in any other group. Addition of calcium to the oxygenated cardioplegic solution decreased adenosine triphosphate preservation and functional recovery. Oxygenation of the acalcemic solution increased adenosine triphosphate preservation and functional recovery. The lowest adenosine triphosphate levels at end arrest were observed in hearts given the hypoxic calcium-containing solution. In the setting of hypothermia and multidose administration, the addition of calcium to a cardioplegic solution resulted in increased energy depletion during arrest and depressed recovery.

Transfusion of predonated autologous blood in elective cardiac surgery.

Despite blood conservation techniques, the average transfusion requirement in patients undergoing elective cardiac surgical procedures remains 1 to 3 units. We studied the efficacy of predonated autologous blood in decreasing homologous transfusion in two matched groups of 58 patients each. Group 1 received homologous blood perioperatively, and Group 2 was transfused with predonated autologous blood. An average of 1.97 units was predonated in Group 2 over 18 days. This resulted in a decline in whole blood hemoglobin concentration of 2.2 gm/dl. No complications resulted from phlebotomy in this ambulatory population consisting predominantly of patients with coronary artery disease. Transfusion of an average of 1.7 units of autologous blood in Group 2 reduced the volume of homologous transfusion by 46% compared with Group 1 (p less than .01). In Group 1, 38% of patients required no homologous transfusion compared with 64% in Group 2 (p less than .02). There were no complications related to autologous blood transfusion. Total transfusion requirement was related to the length of cardiopulmonary bypass. We conclude that autologous predonation is a simple, safe, and cost-effective method of reducing homologous transfusion and thereby decreasing the risk of transfusion-related reactions and infections.

Microsphere reference flow samples during systemic flow adjustment.

Regional myocardial blood flow measurements in the right heart bypass preparation can be particularly valuable, since this preparation provides control of the main hemodynamic determinants of coronary blood flow. We examined the validity of aortic reference flow samples in relation to coronary samples during continuous systemic flow adjustment for aortic pressure control in six dogs on right heart bypass, anesthetized with chloralose and urethan. Microsphere concentrations were compared in paired reference flow samples drawn from the aortic arch and from a coronary artery for 119 left atrial microsphere injections. During left subclavian artery infusion and during femoral artery infusion at rates above 2,000 ml/min, there were high percentage errors in microsphere concentration between paired samples, consistent with aortic sample dilution by systemically infused blood. In 52 injections during withdrawal or femoral infusion below 2,000 ml/min, at cardiac outputs of 390-4,800 ml/min, the percentage error was 0.001 +/- 1.18% (SE); the absolute value of this error was below 20% in 96%, and below 10% in 77% of these injections. Linear regression related these coronary to aortic microsphere concentrations by the equation Y = 1.005X - 1.64, r = 0.997, Sy.x = 13.2 (5.9%). (Sy.x represents the standard deviation from regression.) These data indicate that valid aortic reference flow samples can be obtained within specific hemodynamic conditions during systemic flow adjustment in the right heart bypass preparation.

The superiority of cold oxygenated dilute blood cardioplegia.

It has been clearly shown, both in a laboratory model and in humans, that oxygenation of crystalloid cardioplegic solutions markedly enhances myocardial preservation. The addition of a small volume of red cells to a crystalloid perfusate improves capillary perfusion. Based on these results, we have changed our cardioplegic solution from cold crystalloid to cold oxygenated dilute blood. In the present study we retrospectively evaluate the results of 400 operative procedures to determine whether the addition of oxygenation and a small volume of blood to the cardioplegic solution enhances myocardial protection in the clinical setting. Two hundred consecutive patients who underwent operation with cardioplegic arrest using a cold crystalloid cardioplegic solution (group 1) were compared with a subsequent 200 patients who underwent operation with cold oxygenated dilute blood cardioplegia (group 2). Patients in group 2, who received cold oxygenated dilute blood cardioplegia, had a significantly reduced need for postoperative intraaortic balloon pump counterpulsation and for atrioventricular pacing. Also, patients in group 2 had a lower incidence of perioperative myocardial infarction and had improved early outcome. None of the 200 patients in group 2 had electrocardiographic evidence of perioperative infarction. We conclude that cold oxygenated dilute blood cardioplegia provides better preservation than does a nonoxygenated crystalloid solution during elective ischemic arrest, because a cold crystalloid solution is able to deliver oxygen and the red cells are able to enhance capillary perfusion.

Effect of preload on ischaemic and non-ischaemic left ventricular regional function.

The response to preload of ischaemic and non-ischaemic regions of the left ventricle was studied in 14 dogs undergoing right heart bypass with mean aortic pressure and heart rate held constant. Regional function was measured by sonomicrometry before and after coronary artery occlusion. In the ischaemic region, as expected, there was paradoxical systolic lengthening (that is, systolic shortening was negative) but as stroke volume was progressively increased end diastolic length increased, whereas end systolic length changed little; thus systolic lengthening decreased (systolic shortening increased). Ischaemic regions that were dyskinetic at low stroke volumes were virtually akinetic at high stroke volumes. Additional studies showed that this response was not attributable to increased regional blood flow at high preloads and occurred over a wide range of heart rates and mean aortic pressures. Plots of systolic shortening against end diastolic length, expressing the regional Frank-Starling relation, were well described by linear regression in both ischaemic and non-ischaemic regions, although a few of these relations were better described by higher order polynomials. The slopes of these relations in the ischaemic region were 0.86(0.05) before and 0.83(0.06) after ligation, reflecting a small effect of preload on end systolic length. The data suggest that when contractility and afterload are constant preload determines the magnitude and in certain instances the sign of systolic shortening. In any ischaemic regions incapable of developing force the positive slope of the Frank-Starling relation is attributable to myocardial passive elastic properties. Paradoxical lengthening does not, however, necessarily indicate the absence of active force development; positive and negative values of systolic shortening describe a continuous spectrum of regional contractility. Thus the effects of preload and contractility on systolic shortening in ischaemic as well as non-ischaemic myocardium require differentiation.