Endothelium-derived relaxing factor release associated with increased endothelial cell inositol trisphosphate and intracellular calcium

The release of eicosanoids and endothelium-derived relaxing factor (EDRF) from endothelial cells is thought to involve a calcium-dependent step. Using cultured bovine aortic endothelial cells as a model system, we have examined the relation between agonist-induced changes in inositol polyphosphates and calcium levels within the endothelial cells and extracellular calcium on EDRF release. In a superfusion-cascade system, EDRF was detected by the relaxation of a rabbit aortic ring without endothelium suspended beneath a column of cultured endothelial cells. Endothelial cell stimulation by bradykinin or melittin induced dose-dependent relaxation of the bioassay ring. In addition, bradykinin and melittin stimulated an increase in intracellular calcium concentration in fura-2 loaded endothelial cells and an increase in inositol 1,4,5-trisphosphate (Ins[1,4,5]P3) in cells prelabeled with 3H-myoinositol. Bradykinin stimulation produced transient increases in Ins(1,4,5)P3, fura-2 fluorescence and transient EDRF release. Melittin stimulation induced more prolonged release of EDRF from the endothelial cell column, which was correlated with sustained increases in the fura-2 signal and the level of Ins(1,4,5)P3. Omission of calcium from the cell superfusate attenuated, but did not eliminate, bradykinin-induced EDRF release and the calcium transient, whereas the melittin-induced responses were only slightly attenuated. Endothelial cells clearly demonstrate receptor-activation of phospholipase C and release of sequestered calcium from subcellular sites in response to Ins(1,4,5)P3. These results imply that EDRF release is correlated with increased intracellular calcium levels seen in the absence of extracellular calcium. However, sustained release of EDRF does require influx of extracellular calcium via an undefined mechanism.     

30 years cardiac pacemaker therapy: a status evaluation

Antibradycardiac pacemaker therapy has become established as one of the most effective forms of cardiological therapy for the indications AV-block, sick sinus syndrome, bradyarrhythmia, and hypersensitive carotid sinus. About 220,000 systems are implanted per year worldwide, about 32,000 in West Germany. Of the pacing modes, the fixed-rate ventricular single chamber systems (VVI) dominate with a share of almost 90%. Prognostic importance: For AV-block, the improvement of the prognosis by pacemaker therapy is unquestionable, since it increases the cumulative survival rates to 81% and 95% after 1 year and 50% to 65% after 5 years. For sick sinus syndrome, VVI-pacing proves to be a symptomatic measure, no prognostic importance can be proven. It is not conclusively clarified at present whether physiological pacing modes (AAI, DDD) have any such importance. Pacemaker therapy also has no prognostic importance for bradyarrhythmia. Hemodynamic importance: Numerous hemodynamic studies show that fixed-rate VVI-pacing fails to produce a long-term hemodynamic improvement for either an AV-block or a sick sinus syndrome. In sick sinus syndrome hemodynamic improvement can only be achieved by physiological pacing modes (AAI, DVI, DDD), whereby the increase in cardiac output is between 11% to 30%. For AV-block a long-term hemodynamic improvement can only be obtained by atrial triggered pacing modes (VAT, VDD, DDD); this is higher than the values of fixed-rate VVI-pacing by 7-25% at rest, or 10-40% under exercise. Similar results with improvements of the exercise hemodynamics between 22% and 66% are reported for rate-modulated single-chamber pacing (VVIR) for AV-block. Future trends: In the fourth decade of pacemaker therapy, developments point toward the "smart pacemaker", toward rate-modulated systems with combinations of parameters, toward rate-modulated dual-chamber systems and universal antibradycardiac and antitachycardiac systems.     

Use of rFVIIa for critical bleeding in cardiac surgery: dose variation and patient outcomes

Background and Objectives Recombinant activated factor VIIa (rFVIIa) is increasingly being used in non‐haemophiliac patients for the treatment of severe bleeding refractory to standard interventions. Optimal dosing regimens remain debated in cardiac surgery. Therefore, this study investigated the use of different rFVIIa dosing practices on response to bleeding and patient outcomes in cardiac surgery patients using data from the Haemostasis Registry. Methods Data were extracted from the Haemostasis Registry that records cases of off‐licence rFVIIa use in participating institutions. Univariate analyses compared patients receiving ≤40 μg/kg, 41–60 μg/kg, 61–80 μg/kg, 81–100 μg/kg and >100 μg/kg of rFVIIa on key parameters. Logistic regression models investigated the relationship between independent variables and 28‐day mortality. Results Complete data was available on 804 cardiac surgery patients who received rFVIIa. Of these, 42 (5·2%) were treated with doses ≤40 μg/kg, while the dose group containing the most patients was 81–100 μg/kg (368, 45·77%). Results demonstrated no significant differences in the rate of thromboembolic adverse events, response to bleeding or 28‐day mortality. Conclusions These findings raise the important question of whether lower doses of rFVIIa may be as effective as higher doses in the treatment of severe bleeding in cardiac surgery patients.     

A mysterious journey of a cardiac pacemaker

A permanent bipolar cardiac pacemaker was inserted in the abdomen of a child with Down's syndrome, due to complete heart block following repair of atrioventricular septal defect. This report describes an unusual and unreported complication of the pacemaker getting extruded through the rectum without dysfunction. Implications for early diagnosis and appropriate management and steps to prevent such episodes in future are discussed.     

Calsequestrin determines the functional size and stability of cardiac intracellular calcium stores: Mechanism for hereditary arrhythmia

Calsequestrin is a high-capacity Ca-binding protein expressed inside the sarcoplasmic reticulum (SR), an intracellular Ca release and storage organelle in muscle. Mutations in the cardiac calsequestrin gene (CSQ2) have been linked to arrhythmias and sudden death. We have used Ca-imaging and patch-clamp methods in combination with adenoviral gene transfer strategies to explore the function of CSQ2 in adult rat heart cells. By increasing or decreasing CSQ2 levels, we showed that CSQ2 not only determines the Ca storage capacity of the SR but also positively controls the amount of Ca released from this organelle during excitation-contraction coupling. CSQ2 controls Ca release by prolonging the duration of Ca fluxes through the SR Ca-release sites. In addition, the dynamics of functional restitution of Ca-release sites after Ca discharge were prolonged when CSQ2 levels were elevated and accelerated in the presence of lowered CSQ2 protein levels. Furthermore, profound disturbances in rhythmic Ca transients in myocytes undergoing periodic electrical stimulation were observed when CSQ2 levels were reduced. We conclude that CSQ2 is a key determinant of the functional size and stability of SR Ca stores in cardiac muscle. CSQ2 appears to exert its effects by influencing the local luminal Ca concentration-dependent gating of the Ca-release channels and by acting as both a reservoir and a sink for Ca in SR. The abnormal restitution of Ca-release channels in the presence of reduced CSQ2 levels provides a plausible explanation for ventricular arrhythmia associated with mutations of CSQ2.     

Stimulus-response coupling in sponge cell aggregation: Evidence for calcium as an intracellular messenger

Aggregation of dissociated sponge cells has been proposed as a model for cell-cell recognition mediated by a specific proteoglycan aggregation factor (Microciona aggregation factor). To test whether sponge cells undergo stimulus-response coupling in which intracellular Ca is a messenger, aggregation of mechanically dissociated cells was studied. Changes in light transmission through cell suspensions paralleled aggregation as judged by microscopy. In the presence, but not absence, of Ca (>5 mM) partially purified Microciona aggregation factor aggregated both living and glutaraldehyde-fixed cells. Evidence for a messenger role of Ca was the following: (i) Addition of Ca to Ca-depleted cells induced aggregation that varied with [Ca]. (ii) Addition of Ca ionophores (A23187 and ionomycin) caused aggregation that varied with [Ca] and far exceeded that provoked by Ca alone. Glutaraldehyde-fixed cells did not respond to ionophores with or without Ca. (iii) Calcium antagonists inhibited aggregation. These included inhibitors of the Ca-calmodulin complex (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride and 1-[bis(p-chlorophenyl)methyl]-3-[2,4-dichloro-beta-(2,4-dichlorobenzyloxyl)phenylethyl]imidazolinium chloride), Ca channel blockers (La, Co, Cd, and verapamil), and three nonsteroidal anti-inflammatory agents (indomethacin, ibuprofen, and piroxicam). Results indicated not only that early events of sponge aggregation can be quantified by continuous recording but that aggregation is not simply due to passive agglutination of inert cells by an extracellular proteoglycan. Rather, sponge cells recognize surface ligands to which they respond by Ca-dependent stimulus-response coupling.     

Stabilization of cardiac ryanodine receptor prevents intracellular calcium leak and arrhythmias

Catecholaminergic polymorphic ventricular tachycardia is a form of exercise-induced sudden cardiac death that has been linked to mutations in the cardiac Ca2+ release channel/ryanodine receptor (RyR2) located on the sarcoplasmic reticulum (SR). We have shown that catecholaminergic polymorphic ventricular tachycardia-linked RyR2 mutations significantly decrease the binding affinity for calstabin-2 (FKBP12.6), a subunit that stabilizes the closed state of the channel. We have proposed that RyR2-mediated diastolic SR Ca2+ leak triggers ventricular tachycardia (VT) and sudden cardiac death. In calstabin-2-deficient mice, we have now documented diastolic SR Ca2+ leak, monophasic action potential alternans, and bidirectional VT. Calstabin-deficient cardiomyocytes exhibited SR Ca2+ leak-induced aberrant transient inward currents in diastole consistent with delayed after-depolarizations. The 1,4-benzothiazepine JTV519, which increases the binding affinity of calstabin-2 for RyR2, inhibited the diastolic SR Ca2+ leak, monophasic action potential alternans and triggered arrhythmias. Our data suggest that calstabin-2 deficiency is as a critical mediator of triggers that initiate cardiac arrhythmias.     

Myeloid cell factor-1 is a critical survival factor for multiple myeloma

Multiple myeloma (MM) is characterized by the accumulation of malignant plasma cells in the bone marrow caused primarily by failure of normal homeostatic mechanisms to prevent the expansion of postgerminal center plasma cells. We have examined the molecular mechanisms that promote the survival of MM cells and have identified a key role for myeloid cell factor-1 (Mcl-1), an antiapoptotic member of the Bcl-2 family. These experiments were initiated by the observation that MM cells were exquisitely sensitive to culture in the presence of actinomycin D: caspase activation occurred within 3 hours of treatment and cells were not protected by interleukin-6, the main MM cell growth and survival factor. Actinomycin D-induced apoptosis was blocked by proteasome inhibitors, suggesting that a labile protein was required for MM cell survival. Further analysis demonstrated that Mcl-1 was likely to be the labile factor governing MM cell survival. Mcl-1 protein levels decreased rapidly after culture in the presence of actinomycin D in concordance with effector caspase activation, but addition of proteasome inhibitors reversed the loss of Mcl-1 and maintained cell viability. The levels of other antiapoptotic proteins, including Bcl-2 and members of the inhibitors-of-apoptosis family, were unaffected by these interventions. Furthermore, Mcl-1 antisense oligonucleotides caused a rapid down-regulation of Mcl-1 protein levels and the coincident induction of apoptosis, whereas overexpression of Mcl-1 delayed actinomycin D-induced apoptosis with kinetics that correlated with expression levels of Mcl-1. These data indicate that Mcl-1 expression is required for the survival of MM cells and may represent an important target for future therapeutics.     

Testosterone induces an intracellular calcium increase by a nongenomic mechanism in cultured rat cardiac myocytes

Androgens are associated with important effects on the heart, such as hypertrophy or apoptosis. These responses involve the intracellular androgen receptor. However, the mechanisms of how androgens activate several membrane signaling pathways are not fully elucidated. We have investigated the effect of testosterone on intracellular calcium in cultured rat cardiac myocytes. Using fluo3-AM and epifluorescence microscopy, we found that exposure to testosterone rapidly (1-7 min) led to an increase of intracellular Ca2+, an effect that persisted in the absence of external Ca2+. Immunocytochemical analysis showed that these effects occurred before translocation of the intracellular androgen receptor to the perinuclear zone. Pretreatment of the cells with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethylester and thapsigargin blocked this response, suggesting the involvement of internal Ca2+ stores. U-73122, an inhibitor of phospholipase C, and xestospongin C, an inhibitor of inositol 1,4,5-trisphosphate receptor, abolished the Ca2+ signal. The rise in intracellular Ca2+ was not inhibited by cyproterone, an antagonist of intracellular androgen receptor. Moreover, the cell impermeant testosterone-BSA complex also produced the Ca2+ signal, indicating its origin in the plasma membrane. This effect was observed in cultured neonatal and adult rat cardiac myocytes. Pertussis toxin and the adenoviral transduction of beta- adrenergic receptor kinase carboxy terminal peptide, a peptide inhibitor of betagamma-subunits of G protein, abolished the testosterone-induced Ca2+ release. In summary, this is the first study of rapid, nongenomic intracellular Ca2+ signaling of testosterone in cardiac myocytes. Using various inhibitors and testosterone-BSA complex, the mechanism for the rapid, testosterone-induced increase in intracellular Ca2+ is through activation of a plasma membrane receptor associated with a Pertussis toxin-sensitive G protein-phospholipase C/inositol 1,4,5-trisphosphate signaling pathway.     

心脏起搏器更换原因的临床分析

我院自1981年5月至2004年7月,共有159例病人接受了起搏器和/或电极导线更换手术,累计达196例次.现将更换原因报告如下.     

Calcium and cellular adhesion mechanism--involvement of intracellular calcium to cell adhesion

Cell to cell and cell to extracellular matrix adhesion play an essential role in embryogenesis, differentiation, morphological development and disease processes. Adhesion processes are affected by two reverse directional signaling: expression of adhesion molecules is regulated by inside-out signaling and the molecules transduce extracellular information into cytoplasm by "outside-in" signaling. Cadherins are a family of Ca(2+)-dependent cell-cell adhesion molecules that are important for the mutual association of vertebrate cells. During cell differentiation, for example, the amount or the nature of the cell-surface cadherins and other adhesion molecules change, affecting many aspects of cell-cell adhesion and cell migration.