Minimized perfusion circuits: an alternative in the surgical treatment of Jehovah's Witnesses.

OBJECTIVES: Jehovah's Witnesses present a challenge to cardiac surgeons, as quality of care is not only defined by mortality and morbidity, but also by the avoidance of blood transfusions. Over the last years, minimized perfusion circuits (MPC) have contributed substantially to the achievement of this goal in our clinic. Presented is a retrospective analysis of our experience. METHODS: Twenty-nine Jehovah's Witnesses, aged 69 ± 10 years, have undergone cardiac surgery with a MPC in our institution since 2005. The ROCsafe (reservoir optional circuit) MPC was used in most of these patients (n=27) as it offers the unique possibility of a speedy integration of a reservoir in the event of a major air leak, thereby, negligating any safety concerns. RESULTS: There was no in-hospital or 30-day postoperative mortality. Mean ICU stay was 1.6 ± 2 days with a mean intubation time of 11.3 ± 9.1 hrs. Postoperative complications included one myocardial infarction with accompanying low cardiac output, one stroke, one transient delirium, one idiopathic thrombocytopenia and three re-operations (one sternal infection, one postoperative bleeding and one delayed tamponade). The mean postoperative hospital stay was 9.9 ± 2.3 days. Mean decrease in hemoglobin was 2.1 ± 1.3 g/dl during cardiopulmonary bypass and 3.4 ±1.4 g/dl at discharge. The lowest postoperative hemoglobin level was 9.3 ±1.8 (Range 6-12.9). CONCLUSIONS: These encouraging results emphasize the role MPCs can play in optimizing the quality of patient care. We hope that this report can serve as a stimulus for similar experiences.

Preclinical development of tissue-engineered vein valves and venous substitutes using re-endothelialised human vein matrix.

OBJECTIVE: The aim of the study was to evaluate the use of a decellularised scaffold and its re-endothelialisation in vitro in order to create human vascular substitutes containing venous valves. This research is clinically relevant particularly with regard to the development of venous (valve containing) transplants to replace a diseased femoral vein valve and/or obstructed veins. This technique may enable causal treatment of venous reflux and obstructions. MATERIALS AND METHODS: Valve-bearing segments of human allogeneic great saphenous veins (GSVs) were decellularised using sodium deoxycholic acid (SD) and treated with DNase I. Human venous endothelial cells (ECs) were enzymatically harvested from the GSV, expanded up to the 3rd passage using FCS (n=20) or human AB serum (hABS; n=8) supplemented media before used for re-seeding. In special bioreactors, 3D re-seeding of 28 decellularised GSV was performed with constant perfusion (A; n=8), bidirectional perfusion (B; n=8), bidirectional perfusion/reduced flow (C; n=2), static conditions (D; n=2), and bidirectional perfusion/reduced flow using hABS (E; n=8) instead of FCS. Decellularised GSV, scaled-up EC and 3D-seeded tissue-engineered valve containing neo-veins underwent immunohistochemical and PCR characterisation. RESULTS: Intact collagen and elastin networks as well as complete acellularity were shown after GSV decellularisation. In EC culture, supplementation with hABS led to a significantly higher expression of vWF compared to FCS (p=0.025). Additional EC markers such as CD 31, FLK-1 and VE-Cadherin were not altered. EC re-seeding using hABS supplemented medium (E) led to a confluent monolayer of cells that were immunohistochemically positive for FLK-1, CD 31, vWF and VE-Cadherin and by means of PCR after RNA preparation in 7 of 8 cases but was unsuccessful if FCS was used (A-D). In A-D cells presented as conglomerates positive for CD 31 and VE-Cadherin, suggesting sufficient intercellular contact but not cell-matrix contact. CONCLUSIONS: Treatment with SD and DNase enables complete decellularisation of human valve containing veins whereas 3D matrix components such as collagen and elastin remain preserved. The lumen of the scaffold including the valves can be successfully re-seeded with a human EC monolayer in a 3D bioreactor. There is substantial evidence that hABS and not FCS is essential for the completion of cell-matrix contacts in human veins.

Physiological effects of seven different blocks in glycolysis in Saccharomyces cerevisiae.

Saccharomyces cerevisiae mutants unable to grow and ferment glucose have been isolated. Of 45 clones isolated, 25 had single enzyme defects of one of the following activities: phosphoglucose isomerase (pgi), phosphofructokinase (pfk), triosephosphate isomerase (tpi), phosphoglycerate kinase (pgk), phosphoglyceromutase (pgm), and pyruvate kinase (pyk). Phosphofructokinase activities in crude extracts of the pfk mutant were only 2% of the wild-type level. However, normal growth on glucose medium and normal fermentation of glucose suggested either that the mutant enzyme was considerably more active in vivo or, alternatively, that 2% residual activity was sufficient for normal glycolysis. All other mutants were moderately to strongly inhibited by glucose. Unusually high concentrations of glycolytic metabolites were observed before the reaction catalyzed by the enzyme which was absent in a given mutant strain when incubated on glucose. This confirmed at the cellular level the location of the defect as determined by enzyme assays. With adh (lacks all three alcohol dehydrogenase isozymes) and pgk mutants, accumulation of the typical levels of hexosephosphates was prevented when respiration was blocked with antimycin A. A typical feature of all glycolytic mutants described here was the rapid depletion of the intracellular adenosine 5'-triphosphate pool after transfer to glucose medium. No correlation of low or high levels of fructose-1,6-bisphosphate with the degree of catabolite repression and inactivation could be found. This observation does not support the concept that hexose metabolites are directly involved in these regulatory mechanisms in yeast.