Zinc metalloprotease ProA of Legionella pneumophila increases alveolar septal thickness in human lung tissue explants by collagen IV degradation.

ProA is a secreted zinc metalloprotease of Legionella pneumophila causing lung damage in animal models of Legionnaires' disease. Here we demonstrate that ProA promotes infection of human lung tissue explants (HLTEs) and dissect the contribution to cell type specific replication and extracellular virulence mechanisms. For the first time, we reveal that co-incubation of HLTEs with purified ProA causes a significant increase of the alveolar septal thickness. This destruction of connective tissue fibres was further substantiated by collagen IV degradation assays. The moderate attenuation of a proA-negative mutant in A549 epithelial cells and THP-1 macrophages suggests that effects of ProA in tissue mainly result from extracellular activity. Correspondingly, ProA contributes to dissemination and serum resistance of the pathogen, which further expands the versatile substrate spectrum of this thermolysin-like protease. The crystal structure of ProA at 1.48 Å resolution showed high congruence to pseudolysin of Pseudomonas aeruginosa, but revealed deviations in flexible loops, the substrate binding pocket S1 ' and the repertoire of cofactors, by which ProA can be distinguished from respective homologues. In sum, this work specified virulence features of ProA at different organisational levels by zooming in from histopathological effects in human lung tissue to atomic details of the protease substrate determination.

Effect of minimized perfusion circuit on brain injury markers carnosinase and brain-type fatty binding protein in coronary artery bypass grafting patients.

A minimized perfusion circuit (MPC) has proven to be superior to the conventional circulatory perfusion bypass (CCPB) as it reduces the blood-material interaction and hemodilution. Until now not much is known about impact these different perfusion systems have on the brain. The objective of this study is to determine carnosinase and brain-type fatty binding protein (BFABP) activity as novel specific biomarkers for ischemic brain tissue damage and how their activity differs during and after MPC and CCPB as well as to compare the inflammatory response of both perfusion systems. In a prospective pilot study, 28 patients undergoing coronary artery bypass grafting were randomly divided into an MPC group (n = 14) and a CCPB group (n = 14). Blood samples were taken before, during, and after operation until the fifth postoperative day. The brain biomarker carnosinase was determined by measuring the rate of histidine production from the substrate homocarnosine, whereas BFABP and interleukin-6 were determined by enzyme-linked immunosorbent assay (ELISA). C-reactive protein (CRP) and endothelin-1 were determined by enzyme immunoassay. The mean serum carnosinase activity was significantly higher in MPC (0.57 ± 0.34 nM histidine/mL/min) as compared with the CCPB group (0.36 ± 0.13 nM histidine/mL/min) at the end of operation (P = 0.02). The BFABP did not show any difference between the two groups in the immediate postoperative period until the second postoperative day. From that time point onward, it showed a steep increase in the CCPB group (581.3 ± 157.11 pg/mL) as compared with the concentrations in the MPC group (384.6 ± 39 pg/mL) (P = 0.04). The inflammation markers interleukin-6 and CRP showed a similar pattern in both groups without significant difference. In contrast, the leukocyte count on operation day and endothelin-1 on the first postoperative day were significantly higher in the CCPB group (P = 0.01, P = 0.03, respectively). MPC showed a significant higher and stable serum carnosinase activity during extracorporeal circulation as compared with the CCPB due to less hemodilution and a better preserved oxygen capacity. As a consequence, the antioxidant stress during MPC is limited as compared with CCPB, which means less brain tissue damage reflected by a lower BFABP release. Except endothelin-1 and leukocyte count, the inflammatory response of the MPC and CCPB was equal.

Autotransfusion management during and after cardiopulmonary bypass alters fibrin degradation and transfusion requirements.

UNLABELLED: The coagulation-fibrinolytic profile during cardiopulmonary bypass (CPB) has been widely documented. However, less information is available on the possible persistence of these alterations when autotransfusion is used in management of perioperative blood loss. This study was designed to explore the influence of autotransfusion management on intravascular fibrin degradation and postoperative transfusions. Thirty patients, undergoing elective primary isolated coronary bypass grafting, were randomly allocated either to a control group (group A; n=15) or an intervention group (group B; n=15) in which mediastinal and residual CPB blood was collected and processed by a continuous autotransfusion system before re-infusion. Intravascular fibrin degradation as indicated by D-dimer generation was measured at five specific intervals and corrected for hemodilution. In addition, chest tube drainage and need for homologous blood were monitored. D-dimer generation increased significantly during CPB in group A, from 312 to 633 vs. 291 to 356 ng/mL in group B (p = .001). The unprocessed residual blood (group A) revealed an unequivocal D-dimer elevation, 4131 +/- 1063 vs. 279 +/- 103 ng/mL for the processed residual in group B (p < .001). Consequently, in the first post-CPB period, the intravascular fibrin degradation was significantly elevated in group A compared with group B (p = .001). Twenty hours postoperatively, no significant difference in D-dimer levels was detected between both groups. However, a significant intra-group D-dimer elevation pre- vs. postoperative was noticed from 312 to 828 ng/mL in group A and from 291 to 588 ng/mL in group B (p < .01 for both). Postoperative chest tube drainage was higher in the patients from group A, which also had the highest postoperative D-dimer levels. Patients in group A perceived a higher need for transfusions of red cells suspensions postoperatively. These data clearly indicate that autotransfusion management during and after CPB suppresses early postoperative fibrin degradation. KEYWORDS: cardiopulmonary bypass, cardiotomy suction, coronary surgery, autotransfusion, fibrin degradation.

Analysis of mortality within the first six months after coronary reoperation.

BACKGROUND: Identify risk factors associated with mortality following repeat coronary revascularization (redoCABG) within the first 6 months following surgery. METHODS: Five hundred forty-one redoCABG patients (1987 to 1998) were studied by univariant and multivariant analysis. Mortality was assessed at three different points: hospital mortality (A) (36/541, 6.7%); mortality at 6 months (C) (75/541, 13.9%); and outpatient perioperative mortality, which is a death occurring from the time of hospital discharge to 6 months postoperatively (B) (39/541, 7.2%). RESULTS: Diabetes, hypertension, peripheral vascular disease, renal insufficiency, lung disease, myocardial infarction (MI) before the first operation, MI between the first and redoCABG, lack of sinus rhythm, no IMA graft, acute/emergency operation, perfusion time, and perioperative MI were all identified as risk factors related to early mortality. MI before the first operation, antegrade cardioplegia, and the time period 1987 to 1992 all influenced hospital mortality (A). Diabetes, hypertension, renal insufficiency, lung disease, and valvular heart disease all influenced the outpatient mortality up to 6 months. Independent predictive factors for early mortality were: age more than 69 years; diabetes; vascular insufficiency; chronic lung disease; MI between first and redoCABG; no IMA-graft; acute preoperative MI; emergency operation; perfusion time; perioperative MI; and the time period 1987 to 1992. Risk factors for in-hospital death included MI between the first and redoCABG, cardiopulmonary bypass time, and the time period 1987 to 1992. Diabetes is an important risk factor during the outpatient perioperative phase. Emergency surgery and perioperative MI predict mortality regardless of the time period (A, B, or C). CONCLUSIONS: Early mortality after redoCABG is influenced by many variables during the first 6 months following surgery. Understanding these factors and their time course may better help to assess the true risk associated with reoperation for recurrent coronary insufficiency.

Preoperative prediction of early mortality in redocoronary artery surgery.

OBJECTIVES: Construct a predictive model for early mortality in coronary reoperations (RECABG). METHODS: Five hundred and forty one RECABG (1987-1998) were studied by univariate and multivariate analysis. Stepwise selective procedure (p<0.05) was used to identify a subset of variables with prognostic value for early mortality. This subset was used to calculate a prognostic score 'S' and a predicted probability 'P' for early mortality, P=1/1+e(-S). Sensitivity analysis was used for evaluation. RESULTS: The best predictive variables for early mortality were diabetes, vascular-, lung-disease, a myocardial infarction between the primary and the RECABG, acute- and emergency operation and the operative period. The prognostic accuracy (receiver operating characteristics curve (ROC) area) was 80%. Observed probabilities compare well with the predicted probabilities, and patients were classified in low risk (5%), intermediate risk (15%), high risk (30%) and very high risk (40%). A predicted probability of > or =0.40 was used as cut-off point for the prognostic test. The specificity of this test was 97%, sensitivity 33%, predictive value of a positive test 63% and 90% for a negative test. CONCLUSIONS: The results show that individual patients presented for RECABG, can be stratified according to their early mortality risk. This information can be used to inform the patient, and also to discus the opportunity of the RECABG.

Preoperative prediction of prolonged stay in the intensive care unit for coronary bypass surgery.

OBJECTIVES: To construct a predictive model for a prolonged stay in the intensive care unit (ICU) for coronary artery bypass graft surgery (CABG). METHODS: Eight hundred and eighty-eight patients undergoing CABG were studied by univariate and multivariate analysis. Prolonged stay in the ICU was defined as >/=3 days stay. Stepwise selective procedure (P/=0.40 was used as cut-off point for the prognostic test. The specificity of this test for prolonged stay in the ICU was 99%; sensitivity 9%; positive predictive value 60%; and negative predictive value 89%. CONCLUSIONS: The results show that individual patients presented for CABG, can be stratified according to their risk for prolonged stay >/=3 days in the ICU.

Changing profile of patients undergoing redo-coronary artery surgery.

OBJECTIVE: To outline the changes of the patient population undergoing redo-coronary artery bypass surgery (RECABG). METHODS: Preoperative data of 582 first RECABGs, January 1987-June 2000 were analyzed. Group A: 1987-1991 (n=193); group B: 1992-1995 (n=201); and group C: 1996-June 2000 (n=188). RESULTS: These 582 RECABGs represent about 8.5% of the performed bypass surgeries during this period. Mean age (P=<0.001), percentage of patients with kidney disease (P=0.001), a preoperative PTCA (P<0.001) increased. Significant more elective operations (P<0.001) and lesser urgent operations (P=0.001) were performed in group C. There is a significant different distribution of vessel disease over the three periods (P=0.04). Significant more patients had a patent IMA graft (P<0.001). The angina-free period after CABG (P<0.001), the time period between both operations (P<0.001) and the period between the new onset of angina and the RECABG (P=0.012) increased significantly. Despite the importance of progression of atherosclerosis of any sort decrease significantly (P<0.001) over the whole study, there is a significant increase of patients reoperated for isolated progression of the disease in the native coronary arteries (P<0.001) in group C. The impact of late GF was not different for the total group (P=0.82), however, the percentage of RECABGs for isolated late GF increased significantly between group A and B (P<0.001) but there was a decrease from B to C, however, not significant (P=0.067). CONCLUSION: During the past few years there is a trend in RECABG of older patients, with more coexisting diseases. There are an increasing number of patients with patent arterial grafts, an increase of the event-free period after the CABG, the period between the CABG and RECABG and of the time period between the onset of new angina and the RECABG. Although the impact of atherosclerosis, as angiographic indication, decreased over the total group, there is the significant increase of the percentage of patients reoperated because of isolated progression of atherosclerosis in the native coronary arteries.

Conductivity-based hematocrit measurement during cardiopulmonary bypass.

OBJECTIVE: In a recent clinical study on the reliability of a point-of-care (POC) analyzer, we described a downward bias in hematocrit measurement during cardiopulmonary bypass leading potentially to overtreatment with packed red cells. We hypothesized that the detected deviation is caused by variations in electrolyte concentration rather than by colloids used. METHODS: Blood was sampled from patients before cardiac surgery to obtain undiluted anticoagulated whole blood samples (n = 53). From each sample, four dilution series covering a hematocrit range of 15-30% were made using NaCl (0.9%), modified gelatine (4%), hydroxyethylstarch (6%), or a potassium-based (16 mEq/l) solution, respectively. In each dilution series, hematocrit was measured by POC and via the "golden standard" microcentrifugal method to determine whether the deviation of the POC-analyzer to the microcentrifuge was dependent on the type and dilution level of the solution used. RESULTS: In contrast to the colloid-based dilution series, the crystalloids revealed a significant downward bias of the POC-analyzer with respect to the microcentrifuge (p < 0.05). Due to the correction algorithm for sodium in the POC-analyzer, this deviation was nearly constant for NaCl (mean of difference: -1.8 +/- 0.1%), but increased significantly in case of the potassium-based solution (up to -8.2 +/- 0.4% after 1.5-times dilution). The starch- and gelatine-based solutions led to a significant upward bias (p < 0.05) that increased with progressing dilution (up to 1.2 +/- 0.1% for hydroxyethylstarch and up to 1.3 +/- 0.1% for modified gelatine after 1.5-times dilution). CONCLUSIONS: Conductivity-based POC hematocrit measurement suffers from biases due to changes of the plasma constituents. The downward bias in hematocrit as often seen during cardiopulmonary bypass is driven by changes of different electrolyte concentration rather than by colloids used per se.

Diaphragm single-fiber weakness and loss of myosin in congestive heart failure rats.

Diaphragm weakness commonly occurs in patients with congestive heart failure (CHF) and is an independent predictor of mortality. However, the pathophysiology of diaphragm weakness is poorly understood. We hypothesized that CHF induces diaphragm weakness at the single-fiber level by decreasing myosin content. In addition, we hypothesized that myofibrillar Ca(2+) sensitivity is decreased and cross-bridge kinetics are slower in CHF diaphragm fibers. Finally, we hypothesized that loss of myosin in CHF diaphragm weakness is associated with increased proteolytic activities of caspase-3 and the proteasome. In skinned diaphragm single fibers of rats with CHF, induced by left coronary artery ligation, maximum force generation was reduced by approximately 35% (P < 0.01) compared with sham-operated animals for slow, 2a, and 2x fibers. In these CHF diaphragm fibers, myosin heavy chain content per half-sarcomere was concomitantly decreased (P < 0.01). Ca(2+) sensitivity of force generation and the rate constant of tension redevelopment were significantly reduced in CHF diaphragm fibers compared with sham-operated animals for all fiber types. The cleavage activity of the proteolytic enzyme caspase-3 and the proteasome were approximately 30% (P < 0.05) and approximately 60% (P < 0.05) higher, respectively, in diaphragm homogenates from CHF rats than from sham-operated rats. The present study demonstrates diaphragm weakness at the single-fiber level in a myocardial infarct model of CHF. The reduced maximal force generation can be explained by a loss of myosin content in all fiber types and is associated with activation of caspase-3 and the proteasome. Furthermore, CHF decreases myofibrillar Ca(2+) sensitivity and slows cross-bridge cycling kinetics in diaphragm fibers.

Caffeine prevents protection in two human models of ischemic preconditioning.

OBJECTIVES: We studied whether caffeine impairs protection by ischemic preconditioning (IP) in humans. BACKGROUND: Ischemic preconditioning is critically dependent on adenosine receptor stimulation. We hypothesize that the adenosine receptor antagonist caffeine blocks the protective effect of IP. METHODS: In vivo ischemia-reperfusion injury was assessed in the thenar muscle by 99mTc-annexin A5 scintigraphy. Forty-two healthy volunteers performed forearm ischemic exercise. In 24 subjects, this was preceded by a stimulus for IP. In a randomized double-blinded design, the subjects received caffeine (4 mg/kg) or saline intravenously before the experiment. At reperfusion, 99mTc-annexin A5 was administered intravenously. Targeting of annexin was quantified by region-of-interest analysis, and expressed as percentage difference between experimental and contralateral hand. In vitro, we assessed recovery of contractile function of human atrial trabeculae, harvested during heart surgery, as functional end point of ischemia-reperfusion injury. Field-stimulated contraction was quantified at baseline and after simulated ischemia-reperfusion, in a paired approach with and without 5 min of IP, in the presence (n=13) or absence (n = 17) of caffeine (10 mg/l). RESULTS: Ischemic preconditioning reduced annexin targeting in the absence of caffeine (from 13 +/- 3% to 7 +/- 1% at 1 h, and from 19 +/- 2% to 9 +/- 3% at 4 h after reperfusion, p = 0.006), but not after caffeine administration (targeting 11 +/- 2% and 16 +/- 3% at 1 and 4 h). In vitro, IP improved post-ischemic functional recovery in the control group, but not in the caffeine group (8 +/- 3% vs. -8 +/- 5%, p=0.003). CONCLUSIONS: Caffeine abolishes IP in 2 human models at a dose equivalent to the drinking of 2 to 4 cups of coffee. (The Effect of Caffeine on Ischemic Preconditioning; http://clinicaltrials.gov/ct/show/NCT00184912?order=1; NCT00184912).

Delayed diagnosis of a post-traumatic ventricular aneurysm?

A 52-year-old man was developed pleural effusion and congestive heart failure after a routine orthopedic operation. A compression of atrium and right ventricle, by a calcified mass was discovered. The patient remembered having a blunt chest trauma 34 years before. We believe that the mass, an old hematoma, which was resected, was the result of on occult post-traumatic ventricular aneurysm.

Preoperative prediction of early mortality and morbidity in coronary bypass surgery.

OBJECTIVE: A scoring system to predict early mortality and morbidity in CABG, distinguishing low and high risk patients. METHODS: 563 patients (1998) served as development dataset, 969 patients as validation set. Univariate and logistic regression analysis was used to identify risk factors. RESULTS: Gender, hypertension, pulmonary disease, reoperation, age, operative status and left-ventricular function were predictive variables for early mortality. The area under the ROC curve was 0.81. We identified a low risk, mortality of 1.8% and a high-risk group, mortality of 13.4%. Diabetes, hypertension, kidney and lung disease, reoperation, operative status and left ventricular function were predictive variables for morbidity. The area under the ROC curve was 0.73. We identified a low risk, morbidity of 17%, and a high-risk group, morbidity of 41%. CONCLUSION: This scoring system is a simple system identifying a low and high-risk group for morbidity and early mortality.