Henry J. Mann

Extracorporeal application of a gel-entrapment, bioartificial liver: Demonstration of drug metabolism and other biochemical functions

Metabolic activity of a gel-entrapment, hollow fiber, bioartificial liver was evaluated in vitro and during extracorporeal hemoperfusion in an anhepatic rabbit model. The bioartificial liver contained either 100 million rat hepatocytes (n = 12), fibroblasts (n = 3), or no cells (n = 7) during hemoperfusion of anhepatic rabbits. Eight other anhepatic rabbits were studied without hemoperfusion as anhepatic controls, and three sham rabbits served as normal controls. Albumin production rates (mean ± SEM) were similar during in vitro (17.0 ± 2.8 μg/h) and extracorporeal (18.0 ± 4.0 μg/h) application of the hepatocyte bioartificial liver. Exogenous glucose requirements were reduced (p < 0.01) and euglycemia was prolonged (p < 0.001) in anhepatic rabbits treated with the hepatocyte bioartificial liver. The maximum rate of glucose production by the hepatocyte bioartificial liver ranged from 50-80 μg/h. Plasma concentrations of aromatic amino acids, proline, alanine, and ammonia were normalized in anhepatic rabbits during hepatocyte hemoperfusion. Gel-entrapped hepatocytes in the bioartificial liver performed sulfation and glucuronidation of 4-methylumbelliferone. P450 activity was demonstrated during both in vitro and extracorporeal application of the BAL device by the formation of 3-hydroxy-lidocaine, the major metabolite of lidocaine biotransformation by gel-entrapped rat hepatocytes. In summary, a gel-entrapment, bioartificial liver performed multiple hepatocyte-specific functions without adverse side effects during extracorporeal application in an anhepatic, small animal model. With its potential for short term support of acute liver failure, scale-up of the current bioartificial liver device is indicated for further investigations in large animal, preclinical trials.

Extended liver-specific functions of porcine hepatocyte spheroids entrapped in collagen gel.

The potential use of porcine hepatocytes in a bioartificial liver device requires large quantities of viable and highly active cells. To facilitate the scaling up of the system, liver specific activities of hepatocytes should be maximized. One way of enhancing the specific activities is to cultivate hepatocytes as multicellular spheroids. Freshly isolated porcine hepatocytes form spheroids when cultivated in suspended cultures. These spheroids exhibit higher activities for a number of liver specific functions compared to hepatocytes cultivated as monolayers. However, these activities decreased in a few days in culture. Entrappment of spheroids in collagen gel sustained their metabolic activities at a stable level over 21 days. Production of albumin and urea by spheroid hepatocytes entrapped in collagen gels were 2 to 3 times higher than those by freshly isolated single cells. P-450 activity was demonstrated by metabolism of lidocaine to its main metabolite, monoethylglycinexylidide. Phase II drug metabolism was demonstrated by glucuronidation of 4-methylumbelliferone. This work shows that porcine hepatocyte spheroids entrapped in collagen maintain differentiated functions for an extended time period. Such hepatocyte spheroid entrappment system may facilitate the development of a bioartificial liver support device.

The science of implementation: changing the practice of critical care.

Purpose of reviewFew would disagree that evidence from clinical research should be brought to the bedside in an efficient and equitable manner. Unfortunately, this common agreement does not result in practice change at the bedside where delayed and variable implementation is common. Recognition of this gap has resulted in a new discipline called implementation science that seeks to understand the reasons for slow adoption of clinical therapeutics and to discover effective strategies that accelerate practice change. This article reviews implementation theory and strategies and their effectiveness and relevance to critical care. Recent findingsThe absence of a proven effective framework for implementing clinical practice change has resulted in a patchwork of interventions in ambulatory and acute care medicine. There is an increasing appreciation that interventions should be undertaken only after careful, theory-based examination of the source and strength of the evidence, the organizational and professional context in which the change will be made, and the availability of facilitating methods. Barriers to implementing sepsis management programs have been identified and, in some cases, overcome. SummaryChanging clinical practice is sometimes as difficult as the basic science and clinical trials work that led to the discovery of beneficial therapies. Investigators are now beginning to develop and test more theory-based implementation models that are relevant to the clinical environment. A proportion of the resources used in developing an ICU guideline or protocol must be dedicated to the implementation strategy for successful adoption. ICUs are ideal organizations to test new approaches in implementation science. Intensive care professionals should insist that their practice environment have both a culture that is supportive of adopting new practices and adequate resources to implement them into patient care.

A technique for porcine hepatocyte harvest and description of differentiated metabolic functions in static culture.

Current bioartificial liver devices are based on the use of a large mass of hepatocytes exhibiting differentiated metabolic function. The pig has become a source of interest for the acquisition of such cells-however, harvesting a large mass of highly viable cells has met with difficulty. This study describes a technique for harvesting large quantities of hepatocytes at viabilities greater than 90% and also describes several features documenting differentiated function. Pigs, 6 to 10 kg body weight, underwent in situ two-step whole liver perfusion (ethylene glycol tetraacetic acid and collagenase) and ex vivo cell harvest. Harvests yielded an average of 19.5 billion cells with an average viability of 94.6%. Hepatocytes were then entrapped in type I collagen (3 x 10(5) cells/well) and cultured in serum-free media for 5 days. Pig hepatocytes produced stable amounts of albumin and maintained cytochrome P-450 and glucuronidation activity over 5 days, as shown by the metabolism of lidocaine and 4-methylumbelliferone. These data indicate that pig hepatocytes can be harvested with high yields and can retain viability and differentiated function over at least 5 days of culture, and therefore should prove to be an excellent source of hepatocytes for bioartificial liver devices.

Update on the management of cardiogenic shock

Purpose of reviewCardiogenic shock is a life-threatening emergency that occurs frequently with acute coronary syndromes. If rapid myocardial reperfusion following acute myocardial infarction is not obtained, either with thrombolytics or by revascularization, cardiogenic shock frequently develops and the mortality rate is high. This review summarizes recent advances in the pathophysiology, incidence and treatment of cardiogenic shock. Particular attention is given to pharmacologic advances. Recent findingsCardiogenic shock continues to occur in 5–10% of patients who suffer a myocardial infarction and the mortality remains over 50% in most studies. Treatment preference is referral to a cardiac center capable of reperfusion using multiple therapies. While no delay in reperfusion is acceptable, emphasis on implementing supportive treatment such as vasopressors, inotropes, and fluids remains critical. There is a wide variance in treatment standards despite established guidelines. Overall mortality from cardiogenic shock has decreased but the incidence remains unchanged. SummaryEmerging pharmacological interventions designed to counteract the underlying proinflammatory pathophysiologic mechanisms may, in combination with early revascularization, result in improved patient outcomes, but there is no magic bullet on the horizon. Attention to the timeliness of transport and treatment of patients with a focus on revascularization is required for cardiogenic shock patients.

Pharmacokinetic analysis verifies P450 function during in vitro and in vivo application of a bioartificial liver

Lidocaine is a sensitive substrate for evaluating liver P450 function. In this study, metabolism of lidocaine by xenogeneic hepatocytes in a hollow fiber, bioartificial liver was measured under in vitro conditions (n = 6) and in an anhepatic rabbit model. Animals in the treatment group (n = 6) received hemoperfusion by a bioartificial liver that contained 100 million rat hepatocytes. Other anhepatic rabbits received no hemoperfusion (n = 3) or a bioartificial liver with no cells (n = 3). Lidocaine clearance was 7.0 +/- 0.6 ml/min, and the half-life of lidocaine was 5.6 +/- 0.8 hr under in vitro conditions. Conversion of lidocaine to 3-hydroxy-lidocaine was confirmed in vitro and accounted for 46% of lidocaine elimination in the hepatocyte bioartificial liver. During in vivo application of the bioartificial liver, pharmacokinetic parameters of lidocaine metabolism, including drug half-life and metabolite formation, were significantly improved in anhepatic rabbits. 3-Hydroxy-lidocaine profiles verified the activity of a P450 isozyme expressed preferentially by rat hepatocytes in the bioartificial liver. We conclude that hepatic P450 activity was provided by xenogeneic hepatocytes during in vitro and in vivo applications of a bioartificial liver.

Cefamandole distribution in serum, adipose tissue, and wound drainage in morbidly obese patients

Distribution and elimination of cefamandole 2 g iv were studied in 11 morbidly obese patients during a gastric bypass operation and again on the first postoperative day. Serum, subcutaneous adipose tissue, wound drainage, and urine were analyzed by high performance liquid chromatography for cefamandole and pharmacokinetic parameters from the intraoperative period were compared to those obtained postoperatively. Total body clearance was significantly greater (p < 0.001) postoperatively (297 ml/min) than intraoperatively (254 ml/min). Volume changes were unpredictable but the elimination rate constant tended to increase postoperatively. Renal clearance and percentage of urinary recovery were significantly increased (p < 0.01) postoperatively. The patients had a mean (± SD) volume of the central compartment of 10.3 (± 2.3) L, volume at steady state of 18.3 (± 3.9) L, and elimination rate constant of 1.67 (± 0.63) h−1. Tissue concentrations of cefamandole were highest during the first hour after drug administration and were < 1 μg/g after 3.5 hours. Mean wound drainage concentrations ranged between 10 and 12 μg/ml during a dosing interval and dropped to 7 μg/ml 12 hours after the last dose. Intraoperative dosing of cefamandole is required to maintain subcutaneous adipose tissue concentrations > 1 μg/g during procedures longer than three hours in morbidly obese patients. A postoperative dose of cefamandole 2 g iv q6h will provide sustained and therapeutic concentrations in the wound drainage of morbidly obese patients.

Multicenter implementation of a consensus-developed, evidence-based, spontaneous breathing trial protocol.

Objective:Evidence-based practice recommendations abound, but implementation is often unstructured and poorly audited. We assessed the ability of a peer network to implement an evidence-based best practice protocol and to measure patient outcomes. Design:Consensus definition of spontaneous breathing trial followed by implementation in eight academic medical centers. Setting:Six medical, two surgical, and two combined medical/surgical adult intensive care units among eight academic medical centers. Study Population:Patients initiating mechanical ventilation through an endotracheal tube during a 12-wk interval formed the study population. Interventions:Adoption and implementation of a common spontaneous breathing trial protocol across multiple intensive care units. Measurements and Main Results:Seven hundred five patients had 3,486 safety screens for conducting a spontaneous breathing trial; 2072 (59%) patients failed the safety screen. Another 379 (11%) patients failed a 2-min tolerance screen and 1,122 (34%) patients had a full 30–120 min spontaneous breathing trial performed. Seventy percent of eligible patients were enrolled. Only 55% of passing spontaneous breathing trials resulted in liberation from mechanical ventilatory support before another spontaneous breathing trial was performed. Conclusions:Peer networks can be effective in promoting and implementing evidence-based best practices. Implementation of a best practice (spontaneous breathing trial) may be necessary for, but by itself insufficient to achieve, consistent and timely liberation from ventilator support.

Labetalol for the control of elevated blood pressure following coronary artery bypass grafting

In a multicenter study, the efficacy and safety of intravenous (IV) labetalol for the control of elevated blood pressure were studied in the intensive care unit (ICU) in 65 patients within 4 hours following coronary artery bypass grafting (CABG). Patients with pre-existing ventricular dysfunction, bradycardia, bronchospastic disease, or postoperative complications were excluded. All patients were monitored with a thermodilution pulmonary artery catheter. Entry criteria were a systolic blood pressure (SBP) greater than 140 mm Hg or diastolic blood pressure (DBP) greater than 90 mm Hg for at least five minutes. Intravenous labetalol was loaded incrementally (5, 10, 20, and 40 mg at 10-minute intervals) to a maximum cumulative dose of 75 mg, until either SBP decreased 10% or DBP decreased 10% and was less than 90 mm Hg. Responders were entered into a 6-hour maintenance period, and received 5 to 40 mg of IV labetalol every 10 minutes as needed for blood pressure control. Hemodynamic data and temperature were recorded at baseline, just before each dose of labetalol during the loading period, and at the end of the maintenance period. Alternative therapy was given in the case of nonresponse or adverse events. Intravenous labetalol successfully controlled post-CABG hypertension in 55 of 65 patients (85%); of these, 46 responded to 35 mg or less. Although 28 patients required no further labetalol in the maintenance period, in the others dosage varied from 5 to 400 mg. Reductions in SBP and DBP were associated with moderate reductions in pulse pressure (SBP-DBP) and heart rate (HR). Cardiac index decreased by 18.5%, with a 12.5% decrease in stroke index and 8.1% decrease in HR. Systemic vascular resistance did not increase significantly. Four patients (6%) developed hypotension related to IV labetalol. There was one death due to perioperative myocardial infarction, which was unrelated to labetalol use. The mechanism of action of IV labetalol in controlling hypertension after CABG surgery seems to be moderate negative inotropy and chronotropy. Its alpha-blocking effects seem to be important in preventing reflex vasoconstriction. This is directly opposite to the primary vasodilator effect found when IV labetalol is used to control nonsurgical hypertension. Because of these actions, labetalol should be avoided or used with caution in patients with preoperative and postoperative cardiac dysfunction. In patients with normal left ventricular function, IV labetalol appears to be a safe, effective agent in controlling post-CABG hypertension, with the added potential benefit of enhanced myocardial oxygen balance.

Cimetidine-Associated Thrombocytopenia:

A case of thrombocytopenia associated with cimetidine administration in a 65-year-old white female with recurrent adenocarcinoma is reported. Her platelet count fell from 273 000/mm3 to 35 000/mm3 after cimetidine therapy was started. Upon discontinuation of cimetidine, the platelet count increased to 164000/mm3. Later in her hospital course, the patient again received cimetidine and her platelet count fell to 37 000/mm3. The patient was never leukopenic during her hospitalization. This is the only reported case of thrombocytopenia in the absence of leukopenia that is documented by a rechallenge with cimetidine.