Mechanism of red cell 2,3-diphosphoglycerate increase in neonatal lambs

The tenfold increase in red cell 2,3-diphosphoglycerate (DPG) concentration that occurs during the first 5 days of life in lambs is an important adaptation to extrauterine life. In lambs, DPG reduces hemoglobin oxygen affinity by the Bohr effect. Our data on 10 neonatal lambs suggest that the biochemical mechanism underlying this DPG increase involves the following: (1) a rise in plasma glucose from 40 to 100 mg/dl in the first 48 hr of life, which allows for increased glucose consumption in the highly glucose-permeable neonatal RBC; (2) a transitory rise in blood pH begins at birth, peaks at about 20 hr, and falls slightly; (3) the pH increase coincides with a threefold increase in RBC fructose-1,6-diphosphate (FDP) concentration due, we believe, to pH activation of phosphofructokinase; (4) glycolytic intermediates after the glyceraldehyde-3-phosphate dehydrogenase (GAPD) step do not rise in the first 24 hr of life, possibly due to insufficient inorganic phosphate (Pi), a substrate of GAPD; (5) plasma Pi increases from about 7 mg/dl at birth to 11 mg/dl at 72 hr, activates the GAPD, and FDP levels decline; and (6) the in vitro activity of the DPG synthetic enzyme, DPG mutase, is increased 12-fold in neonatal compared to adult RBC. We conclude that the postnatal rise in DPG is explained at least in part by the sequential effects of these metabolic changes.     

A Negative Direct Antiglobulin Test with Strong IgG Red Cell Autoantibodies Present in the Serum of a Patient with Autoimmune Haemolytic Anaemia

The case is presented of a boy with an autoimmune haemolytic anaemia of 10 years duration. He had a positive direct antiglobulin test with IgG and complement detected on the red cells and with IgG autoantibodies in the serum. During a recent episode of severe haemolysis, the Hb level fell to 3.8 g/dl and the direct antiglobulin test became negative although his autoantibodies still reacted with all the red cells in a panel. The serum reacted more strongly with C- and e-positive cells. The Rh phenotype of the patient was CcDee as it had always been. Possible explanations of the unexpected findings are discussed.     

Interaction of single-chain urokinase with its receptor induces the appearance and disappearance of binding epitopes within the resultant complex for other cell surface proteins

Binding of urokinase-type plasminogen activator (uPA) to its glycosylphosphatidylinositol-anchored receptor (uPAR) initiates signal transduction, adhesion, and migration in certain cell types. To determine whether some of these activities may be mediated by associations between the uPA/uPAR complex and other cell surface proteins, we studied the binding of complexes composed of recombinant, soluble uPA receptor (suPAR) and single chain uPA (scuPA) to a cell line (LM-TK- fibroblasts) that does not express glycosylphosphatidylinositol (GPI)-anchored proteins to eliminate potential competition by endogenous uPA receptors. scuPA induced the binding of suPAR to LM-TK- cells. Binding of labeled suPAR/scuPA was inhibited by unlabeled complex, but not by scuPA or suPAR added separately, indicating cellular binding sites had been formed that are not present in either component. Binding of the complex was inhibited by low molecular weight uPA (LMW-uPA) indicating exposure of an epitope found normally in the isolated B chain of two chain uPA (tcuPA), but hidden in soluble scuPA. Binding of LMW-uPA was independent of its catalytic site and was associated with retention of its enzymatic activity. Additional cell binding epitopes were generated within suPAR itself by the aminoterminal fragment of scuPA, which itself does not bind to LM-TK- cells. When scuPA bound to suPAR, a binding site for alpha 2-macroglobulin receptor/LDL receptor-related protein (alpha 2 MR/LRP) was lost, while binding sites for cell-associated vitronectin and thrombospondin were induced. In accord with this, the internalization and degradation of cell-associated tcuPA and tcuPA-PAI- 1 complexes proceeded less efficiently in the presence of suPAR. Further, little degradation of suPAR was detected, suggesting that cell- bound complex dissociated during the initial stages of endocytosis. Thus, the interaction of scuPA with its receptor causes multiple functional changes within the complex including the dis-appearance of an epitope in scuPA involved in its clearance from the cell surface and the generation of novel epitopes that promote its binding to proteins involved in cell adhesion and signal transduction.     

Catheter-ablation of ventricular tachycardia in patients with coronary artery disease: influence of the endocardial substrate size on clinical outcome

Ablation of symptomatic ventricular tachycardia (VT) in patients with coronary artery disease is frequently performed using the three dimensional mapping system CARTO. In the amplitude map, bipolar potentials of <1.5 mV are considered abnormal and represent damaged myocardium due to previous infarction. This pathological electrical area can be arrhythmogenic, serving as the substrate for reentrant VT. The purpose of this study was to correlate the size of the endocardial substrate with the success of VT catheter ablation. Included in this retrospective analysis were 69 consecutive patients with coronary artery disease who underwent ablation for symptomatic clinical VT using CARTO. The voltage maps were analyzed and the area with abnormal bipolar electrograms (<1.5 mV) was determined using geometric approximation models. The area of abnormal electrograms was divided into three sizes: small (≤15 cm²; 11 patients), medium (16–99 cm²; 50 patients), and large (≥100 cm²; 8 patients). Patient characteristics were not different between the three substrate groups in regard to age, tachycardia cycle length, or number of radiofrequency applications, however differed significantly between the small, medium and large group in regard to left ventricular ejection fraction (44 ± 12% vs. 32 ± 9% vs. 21 ± 7%, respectively; P = 0.001). Overall, there was a significant correlation between myocardial infarction locations and endocardial substrate sizes (P = 0.031), such that 73% of small substrates were found after inferior myocardial infarctions, and 100% of large substrates after anterior and multiple myocardial infarctions (P = 0.003). After ablation, inducibility of ventricular arrhythmias was more rare in patients with small substrates compared to patients with medium or large substrates (small substrates: 9%, medium and large substrates: 43%, P = 0.043). Although during follow-up of 25 ± 17 months (1 day to 72 months) there was no significant difference between endocardial substrate sizes in regard to recurrence rates (small: 27%, medium: 38%, large: 50%, P = 0.588), patients with a small substrate did not have fast VT or ventricular fibrillation (VF), in contrast to 30% and 38% of patients with medium and large substrates, respectively. We conclude that in patients with coronary artery disease a small area of low amplitude bipolar potentials (≤15 cm²) was seen more often after inferior myocardial infarction than after anterior and multiple infarctions. After ablation, patients with small substrates were rarely inducible and showed a more benign course during follow-up (trend towards fewer arrhythmia recurrences and no fast VT or VF). As a result smaller arrhythmogenic substrates appear to be better amenable to catheter ablation than larger substrates.