[Is the determination of the defibrillation threshold in patients with an implantable cardioverter-defibrillator still required?]. / Wird die Bestimmung der Defibrillationsschwelle bei Patienten mit implantierbarem Kardioverter/Defibrillator noch benötigt?

BACKGROUND: Intraoperative testing of implantable cardioverter-defibrillators (ICDs) is time consuming and associated with risks. In the present study, we elucidated whether the initial implantation of an ICD with high energy output makes intraoperative defibrillation threshold testing (DFTT) unnecessary even though antiarrhythmic (AA) therapy is needed in the future. METHODS: A total of 111 patients (94 men, 17 women) receiving an ICD with subsequent AA therapy (mexiletine, amiodarone, sotalol, flecainide) were analyzed retrospectively. DFT was performed during ICD implantation and after AA drug therapy. In a second step, DFT results from the study cohort were analyzed for implantation of virtual ICDs with either low (≤ 30 J, LOD), intermediate (34 J, IOD), or high energy output (36 J, HOD). RESULTS: In the study cohort, all patients reached the safety margin (SM) of 10 J between DFT and maximal shock energy of the ICD. After loading of AA agents, 6 patients (12%) with a LOD, 3 patients (11%) with an IOD, and 3 (13%) patients with a HOD failed the 10 J SM. Using virtual ICDs, 6 (5.5%) patients with a LOD, 1 patient (1%) with an IOD, and no patients with a HOD would have failed the 10 J SM. After loading of AA agents, 18 patients (16%) with a virtual LOD, 12 patients (10.8%) with an IOD, and still 9 patients (8%) with a HOD would have failed the 10 J SM. CONCLUSION: Our results demonstrate that the 10 J SM would have been achieved intraoperatively in all patients with virtual HOD ICDs. Thus, determination of the DFT during implantation does not seem to be obligatory. However, in patients receiving AA agents, DFT testing is still required.

Indications for predismissal testing with arrhythmia-induction in patients receiving an implantable cardioverter defibrillator.

UNLABELLED: Arrhythmia induction during implantation of cardioverter defibrillators (ICD) is a standard procedure. However, controversy exists regarding the need for routine arrhythmia induction before discharge from hospital (pre-hospital discharge (PHD) test). In order to reduce the number of tests we identified risk factors that predict relevant ICD malfunction. METHODS AND RESULTS: 965 patients receiving a first device implantation (n=724) or device/system replacement (n=241) between 1998 and 2004 were analysed. During implantation 176 (18%) complications (intraoperative undersensing of induced arrhythmias, unsuccessful arrhythmia-therapy or low DFT safety margin) occurred. Frequent (>4 times) intraoperative lead repositioning due to low sensing values was present in 44 patients (5%). 9% of the patients with first ICD implantation, 21% with device replacement and 27% with system replacement developed complications during PHD testing with arrhythmia induction. Intraoperative complications, although corrected during implantation, were independent risk factors for malfunction during PHD testing (p<0.05). Additional predictors for malfunction were intraoperative lead repositioning (>4 times) and a history of both VF and VT (p<0.05). Patients without intraoperative complications rarely developed malfunction during PHD testing (3.7% first device, 6.25% system replacement). Only in patients undergoing device replacement was a higher risk for failure (13%) evident. No risk factors could be identified for these subgroups. CONCLUSION: Routine arrhythmia induction during PHD is recommended in ICD patients with intraoperative complications, although corrected during implantation, as well as frequent intraoperatives lead repositioning. Patients undergoing device/system replacement uncomplicated implantation are not generally at low risk for device failure.

Effect of testosterone loading on the kinetic of faecal testosterone excretion in mallards.

Intestinal passage time of coloured fodder and testosterone turnover were examined by faecal steroid analysis in mallards in the reproductive and postrefractory period. In the latter, the discharge of coloured fodder began 36 minutes after ingestion in males, and 56 minutes in females. During reproduction the discharge began 93 minutes and 112 minutes after ingestion in males and females, respectively. Total passage time was similar in the reproductive and postrefractory period in both sexes. After intraperitoneal testosterone injection, faecal samples were collected for 8 hours and testosterone levels were measured using RIA. In the postrefractory period, 1-2 hours after testosterone loading a strong increase of faecal testosterone content developed in males, meanwhile a slighter testosterone peak appeared in females. During reproduction testosterone excretion began 1.5-2 hours after injection in both sexes but in females its increase was smaller. The duration of response to testosterone loading was 5 hours in both periods and both sexes. Intensive excretion after T loading appeared earlier in males than in females, but total passage time finished at the same time: 5 hours after loading. The character of testosterone excretion was corresponding to the passage of fodder-chimus-faeces in the reproductive and postrefractory period in both sexes.

[Efficiency potential in the pacemaker/implantable cardioverter defibrillator outpatient clinic]. / Effizienzpotential in der Nachsorge von Schrittmachern und implantierbaren Kardioverter Defibrillatoren.

The aim of the present study was to elucidate whether the duration of a technical follow-up (FU) of a pacemaker (PM)/implantable cardioverter defibrillator (ICD) has an impact on cost-effectiveness in the outpatient clinic. We determined the time required for a complete FU of devices from three different manufacturers. In 130 patients (70 VVI/DDD-PM, 60 VVI/DDD-ICD) with either a PM (Phylos, Chorum/Talent, Kappa, EnPulse) or an ICD (Belos, Alto or GEM) the time was recorded for a complete FU including determination of lead impedance, sensing and pacing threshold. The time for activation of individual menue buttons was excluded. On the basis of time required for FU, cost-units (CU) were calculated for 2000 FU/year and for a presumed device longevity (PM 7 years, ICD 5 years). For VVI-PM, the duration of FU was almost identical for devices from different manufacturers (105+/-11 s to 125+/-8 s; p=n.s.). However, analysis of DDD-PM revealed marked differences (140+/-25 s vs 282+/-23 s, p<0.05). Time for FU of ICDs varied between 108+/-5 s and 207+/-21 s (p<0.05) in VVI-ICDs and between 129+/-8 ms and 225+/-23 s (p<0.05) in DDD-ICDs. The total savings could be 55 000 CU in VVI- and 53 333 CU in DDD-ICDs. For full automatic DDD-pacemakers (EnPulse) time for FU could be reduced to 58+/-3 s (p<0.05). Differences in FU times were caused by problems with telemetry, delay during booting of the programmer, interrogation at the beginning and at the end of FU and for sensing tests. Improving not only programmers and devices but also test automaticity could significantly increase cost-efficiency in the outpatient clinic.

Differential expression and imprinting status of Ins1 and Ins2 genes in extraembryonic tissues of laboratory mice.

There are two functional insulin genes in the mouse genome. The Ins2 gene is imprinted and expressed monoallelically from the paternal allele in the yolk sac. In the present study we have re-examined the imprinting status of Ins1. We found that Ins1 is not expressed in the yolk sac of several laboratory mouse strains. The asynchrony of replication at the wild type locus was significantly lower than at imprinted loci and was more similar to non-imprinted loci. Finally, we have taken the advantage of the Ins1(neo) allele created by homologous recombination to examine the allelic usage at this locus. We observed that the neo gene inserted at the Ins1 locus was expressed from both the paternally and the maternally transmitted allele. Therefore, the Ins1 gene does not share any of the basic properties of imprinted genes. On the basis of these data, we concluded that Ins1 locus is unlikely to be imprinted in common laboratory mice.

A comparative methodical study of the faecal steroid analysis on birds: looking for a valid method of testosterone determination.

In a comparative study, a relatively simple and high sensitivity method was developed for analysis of testosterone-equivalent(s) in the faeces of different bird species. To determine the recovery of extractions and purifications, tritium-labelled testosterone was added to the wet samples. Then the samples were treated with sodium dodecil sulphate (SDS), an emulsificator to "open-up" the complex, lipid-coated particles of faecal samples. This emulsification resulted in the decrease of the quantity of interfering substances after diethyl-ether extraction and the linearity of the measured testosterone equivalents from aliquots in the range of 2 and 10 mg of faeces. In the RIA, we applied a group specific polyclonal testosterone antibody which cross-reacted with reduced metabolites and at a certain level with sulphate conjugates as well. The use of Helix enzymes did not modified significantly the results of the analysis relating to a low level of conjugated androgens in the faecal extracts. The biological validity of the method was tested on domestic cockerels, where between the plasma and faecal testosterone values a four hours phase shift was observed, with a correlation of 0.6355. This method is suitable for "non invasive", behavioural-ethological studies.

T-cell epitope analysis on the autoantigen phogrin (IA-2beta) in the nonobese diabetic mouse.

The protein tyrosine phosphatases (PTPs) IA-2 and phogrin (IA-2beta) are major autoantigens in type 1 diabetes that possess common serological epitopes in their COOH termini. The epitopes recognized by the T-cells that cause the disease, however, remain to be defined. Eight phogrin-specific T-cell clones were generated from NOD mice, and their epitopes were mapped. The mapping was performed initially with recombinant gluthathione S-transferase-phogrin COOH deletion constructs and ultimately with overlapping synthetic peptides. Two dominant epitopes were identified: one (aa 629-649) immediately adjacent to the transmembrane domain (aa 604-628) and the second (aa 755-777) lying in the NH(2)-terminal region of the conserved PTP domain. T-cells that are specific to either of these peptides and that could destroy islet tissue in vivo though spontaneous T-cell proliferative responses were observed in prediabetic female NOD splenocytes only to the aa 755-777 epitope. In NOD female mice immunized with the epitope peptide, intramolecular determinant spreading occurred from the aa 629-649 epitope to the aa 755-777 epitope but not in the opposite direction. We concluded that the initial T-cell response to phogrin is restricted to a small number of dominant peptides and that it subsequently spreads to other regions of the molecule, including those containing the major humoral epitopes that are highly conserved between IA-2 and phogrin.

Humoral autoreactivity to an alternatively spliced variant of ICA512/IA-2 in Type I diabetes.

AIMS/HYPOTHESIS: The receptor tyrosine phosphatase like-protein ICA512/IA-2 occurs as a proteolytically-processed 65,000 Mr type 1 transmembrane glycoprotein in beta cells and is a major autoantigen of Type I (insulin-dependent) diabetes mellitus. We investigated whether alternative splicing could affect humoral autoreactivity to the molecule. METHODS: Genomic and cDNA sequence analysis showed the presence of a ICA512 variant in islets and lymphoid tissues with an in-frame deletion of exon 13 which produces a secreted form lacking aa 557-629 including the transmembrane domain (aa 577 to 600). The alternatively spliced protein is detectable by western blotting in normal islets and translated into a protein that is processed to a series of soluble forms of 25,000-35,000 Mr Radioimmuno-precipitation assays for anti-ICA512 autoantibodies were developed with the widely used ICA512.bdc construct (which has exon 13 deleted) and a series of full-length and modified ICA512/IA-2 molecules. RESULTS: The assays showed that ICA512.bdc and ICA512604-979 gave the best discrimination between diabetic and control sera. With ICA512604-979 a somewhat greater proportion of patients expressing antibodies were detected than with ICA512.bdc in the groups studied (70.5 % vs 63.2 % of prediabetic/new-onset and 25.0 vs 13.9% in patients with diabetes > 20 years). Conversely, a small proportion (3 % recent-onset and 6% > 20 years) had antibodies to ICA512.bdc but not ICA512(604-979). CONCLUSION/INTERPRETATION: Important epitopes lie within the exon 13 region and others can be generated by the alternative splicing. As the deltaexon 13 variant is probably secreted by the beta cell, it could be recognized by the cellular and humoral arm of the immune system in the absence of cellular damage.

Early expression of antiinsulin autoantibodies of humans and the NOD mouse: evidence for early determination of subsequent diabetes.

With the development of an insulin autoantibody (IAA) assay performed in 96-well filtration plates, we have evaluated prospectively the development of IAA in NOD mice (from 4 weeks of age) and children (from 7 to 10 months of age) at genetic risk for the development of type 1 diabetes. NOD mice had heterogeneous expression of IAA despite being inbred. IAA reached a peak between 8 and 16 weeks and then declined. IAA expression by NOD mice at 8 weeks of age was strongly associated with early development of diabetes, which occurred at 16-18 weeks of age (NOD mice IAA(+) at 8 weeks: 83% (5/6) diabetic by 18 weeks versus 11% (1/9) of IAA negative at 8 weeks; P <.01). In man, IAA was frequently present as early as 9 months of age, the first sampling time. Of five children found to have persistent IAA before 1 year of age, four have progressed to diabetes (all before 3.5 years of age) and the fifth is currently less than age 2. Of the 929 children not expressing persistent IAA before age 1, only one has progressed to diabetes to date (age onset 3), and this child expressed IAA at his second visit (age 1.1). In new onset patients, the highest levels of IAA correlated with an earlier age of diabetes onset. Our data suggest that the program for developing diabetes of NOD mice and humans is relatively "fixed" early in life and, for NOD mice, a high risk of early development of diabetes is often determined by 8 weeks of age. With such early determination of high risk of progression to diabetes, immunologic therapies in humans may need to be tested in children before the development of IAA for maximal efficacy.

Cellular immune response to phogrin in the NOD mouse: cloned T-cells cause destruction of islet transplants.

The ability of nonobese diabetic (NOD) mice to mount a cellular immune response to the secretory granule protein tyrosine phosphatase (PTP), phogrin was evaluated by immunization of 8- to 12-week-old animals with recombinant phogrin in complete Freund's adjuvant. Draining lymph nodes displayed a robust proliferative response to the protein, as did derived T-cell lines and clones. Ten clones obtained by limiting dilution were all CD4+ and of a T-helper-1-like phenotype, but showed variation in their Vbeta usage. Of the 10 clones, 3 responded to endogenous antigens in rat islets. Two of these caused the destruction of rat islets that had been transplanted under the kidney capsule of streptozotocin-treated NOD scid mice without affecting adjacent thyroid implants. The results demonstrate the feasibility of generating antigen-specific diabetes-inducing CD4+ cells by direct immunization of NOD mice and their potential use for further studies of the antigenic epitopes in the PTP family members. The conclusion, based on serological studies, that PTP members do not play a role in the pathogenesis of type 1 diabetes in rodent models needs reevaluation in light of these findings.

Early recognition of pregnancy by the maternal immune system.

PROBLEM: Immunologic recognition of pregnancy is important for normal gestation. Successful pregnancy is characterized by a Th2 dominance, whereas there is a Th1 dominance in failed pregnancies. We assume that a signal given by the fertilized ovum induces a Th2 shift, necessary for a normal outcome. In vitro fertilization provides a tool for testing this possibility. METHOD OF STUDY: Phytohemagglutinin-stimulated lymphocytes were incubated for 48 hr in the presence of culture media from in vitro fertilized eggs, as well as in follicular fluid (FF) and control supernatants. Total RNA was isolated from the lymphocytes by the guanidine-isothiocyanate method and interleukin (IL)-10 mRNA expression was detected by reverse transcription-polymerase chain reaction. RESULTS: Ten percent of the activated lymphocytes incubated with FF expressed IL-10 mRNA, whereas 88% of the lymphocytes activated with supernatants of sperm + oocytes gave a positive signal. Significantly (P < 0.05) fewer (50%) lymphocytes stimulated in the presence of control supernatants also expressed mRNA for IL-10. In supernatants of activated lymphocytes incubated with the culture medium of spermia + oocytes, the concentration of IL-10 was significantly higher than in the lymphocytes incubated with FF. CONCLUSION: These data suggest that the presence of the fertilized ovum induces a Th2 shift, which enables pregnancy to proceed to term.

The immunological pregnancy protective effect of progesterone is manifested via controlling cytokine production.

PROBLEM: This study was aimed at investigating the involvement of an altered cytokine pattern in the immunomodulatory and anti-abortive effects of a progesterone-induced immunomodulatory protein (PIBF). METHOD: PIBF expression on lymphocytes of healthy pregnant women and from women at risk for premature pregnancy termination was determined. In sera of the same women TNF alpha was quantified by a bioassay using L929 cells. NK activity was determined by a single cell cytotoxicity assay. Cytokine production of the lymphocytes or murine spleen cells was measured by ELISA or detected by immunocytochemistry. In pregnant mice endogenous PIBF activity was neutralized by anti-PIBF IgG. RESULTS: Sera of women at risk for premature pregnancy termination contained significantly higher concentrations of TNF alpha than those from healthy pregnant women and PIBF expression on the lymphocytes was inversely related to serum concentration of TNF alpha. Increased NK activity of lymphocytes after neutralization of endogenous PIBF activity is corrected by anti-IL 2 treatment and PIBF inhibits IL 12 expression on activated lymphocytes. PIBF increases IL-10 production by activated spleen cells. In pregnant mice, neutralization of endogenous PIBF activity by specific antibody results in increased resorption rate and reduced splenic IL-10 production. CONCLUSIONS: Our data allow the assumption that via blocking IL-12 production PIBF inhibits NK activation with a concomitant reduction of TNF alpha levels. Disturbances in this system might lead to the expression of the known synergistic effect of IL-12 and TNF alpha, resulting in a Th 1 type cytokine dominance and pregnancy termination.

(-)Deprenyl and (-)1-phenyl-2-propylaminopentane, [(-)PPAP], act primarily as potent stimulants of action potential-transmitter release coupling in the catecholaminergic neurons.

The activity of the catecholaminergic neurons in the rat brain is enhanced significantly 30 min after the subcutaneous injection of very small doses of (-)deprenyl (threshold doses: 0.01 mg/kg for noradrenergic neurons and 0.025 mg/kg for dopaminergic neurons). As a catecholaminergic activity enhancer (CAE) substance (-)deprenyl is about ten times more potent than its parent compound, (-)methamphetamine. While the (+)methamphetamine is 3-5 times more potent than (-)methamphetammine in releasing catecholamines, the (-)methamphetamine is the more potent CAE substance. The mechanism of the CAE effect of (-)deprenyl and (-)PPAP, a deprenyl-derived substance devoid of MAO inhibitory potency, was studied in rats by measuring: a) the release of catecholamines from striatum, substantia nigra, tuberculum olfactorium and locus coeruleus; b) the stimulation induced release of 3H-noradrenaline from the isolated brain stem; and c) the antagonistic effect against tetrabenazine-induced depression of learning in the shuttle box. The CAE effect was found to be unrelated: a) to the inhibition of MAO activity; b) to the inhibition of presynaptic catecholamine receptors; c) to the inhibition of the uptake of catecholamines; and d) to the release of catecholamines. It was concluded that (-)deprenyl and (-)PPAP act primarily as potent stimulants of action potential-transmitter release coupling in the catecholaminergic neurons of the brain. We show that both (-)deprenyl and (-)PPAP enhance the inward Ca2+ current in sino-auricular fibers of the frog heart. (-)PPAP was much more potent than either (+)PPAP or (-)deprenyl in this test.

A progesterone-induced protein increases the synthesis of asymmetric antibodies.

The immunological effects of progesterone are mediated by a 34-kDa protein named the progesterone-induced blocking factor (PIBF). PIBF induces increased production of Th2-type cytokines; thus, it might stimulate antibody synthesis by B cells. There is a population of antibodies which, owing to the presence of a mannose-rich oligosaccharide residue on one of the Fab arms of the molecule, possess an asymmetric structure. Due to the asymmetric structure these molecules have no effector functions; however, they might act as blocking antibodies. This study was aimed at investigating the effect of progesterone-dependent immunomodulation on antibody production by B cells, with special emphasis on the synthesis of asymmetric nonprecipitating antibodies. The ratio of asymmetric IgG was significantly higher in supernatants of hybridoma cells cultured in the presence of PIBF than in those cultured in the absence of PIBF. Lymphocytes from healthy pregnant women produce significantly more PIBF than those of women with pathological pregnancies. The present studies revealed a positive relationship between asymmetric antibody content of the sera and PIBF expression on lymphocytes. Blocking of progesterone receptors by RU 486 or neutralizing endogenous PIBF activity by specific antibody significantly reduced the production of asymmetric antibodies in pregnant mice. Effector function of conventional and asymmetric antibodies was compared in a TNF alpha neutralization assay. Purified asymmetric anti-TNF alpha antibodies did not neutralize the cytotoxic effect of TNF alpha on L929 murine fibroblast target cells, whereas conventional anti-TNF alpha antibodies in the same concentration significantly (P < 0.001) reduced cytotoxicity. Our data suggest that PIBF induces increased production of asymmetric antibodies. These antibodies fail to exhibit effector functions and by blocking fetally derived antigens might contribute to protection of the fetus.

The use of an internal mammary artery for creating a systemic-to-pulmonary artery shunt.

We report on two patients who had systemic-to-pulmonary artery shunts created by the use of the internal mammary artery (IMA). The first patients was operated on more then 30 years ago (the case has never been published) and the second one move recently. We give a brief summary of the ten cases of the same operation published so far, and emphasize the usefulness of the IMA that stays open despite the initially poor run-off and is capable of supplying increasing amounts of blood to the growing pulmonary arteries.

[Electrophysiologic study of the anti-arrhythmic mechanism of action of phosphocreatine in acute myocardial ischemia and reperfusion]. / Elektrofiziologicheskoe issledovanie mekhanizmov antiaritmicheskogo deistviia fosfokreatina pri ostroi ishemii i reperfuzii miokarda.

To determine the possible application of exogenous phosphocreatine (ePC) to protect the ischemic myocardium from reperfusion abnormalities in cardiac rhythm, the antiarrhythmic and antifibrillatory activities of the agent were studied in an acute myocardial ischemia model and reperfusion-induced cardiac damage. It was shown that ePC produced a pronounced antifibrillatory effect in acute coronary occlusion and subsequent reperfusion. The agent substantially increased the threshold of electric ventricular fibrillation and the frequency of spontaneous defibrillation. The highest activity was shown by ePC in ischemic myocardial reperfusion. The agent suppressed both rapid inward Na+ current and slow inward Ca2+ current. The effects of ePC on transmembrane ion currents suggest that the agent has a unique electrophysiological mechanism of action involving the properties of Classes I and IV antiarrhythmics, making ePC promising in clinical application in patients with impaired conduction and automatism.

Sensitized immunochemical method for the specific detection of human haemoglobin in dried biological samples.

The authors describe a modified version of counterimmunoelectrophoresis of easy performance and high sensitivity. With this method human haemoglobin can easily and specifically be determined either in fresh or dried biological samples. Glutaraldehyde pretreatment of the samples results in the development of complexes with highly favourable electrophoretic mobility and precipitating capacity. By this chemical modification and use of a double set of samples with varying antigen-antibody proportion, the sensitivity of haemoglobin detection in erythrocyte-containing haemolysate and native blood was 300 ng/ml. The examination is easily performed, and without considerable outlay, even with conventional laboratory facilities. It is equally suitable for the detection of occult colorectal bleeding and for species-specific study of blood stains of unknown origin. The authors succeeded in identifying human haemoglobin even in a 42-month dried blood stain.