Long-term outcome of the unrestricted use of everolimus-eluting stents compared to sirolimus-eluting stents and paclitaxel-eluting stents in diabetic patients: The Bern–Rotterdam diabetes cohort study

BACKGROUND Newer generation everolimus-eluting stents (EES) improve clinical outcome compared to early generation sirolimus-eluting stents (SES) and paclitaxel-eluting stents (PES). We investigated whether the advantage in safety and efficacy also holds among the high-risk population of diabetic patients during long-term follow-up. METHODS Between 2002 and 2009, a total of 1963 consecutive diabetic patients treated with the unrestricted use of EES (n=804), SES (n=612) and PES (n=547) were followed throughout three years for the occurrence of cardiac events at two academic institutions. The primary end point was the occurrence of definite stent thrombosis. RESULTS The primary outcome occurred in 1.0% of EES, 3.7% of SES and 3.8% of PES treated patients ([EES vs. SES] adjusted HR=0.58, 95% CI 0.39-0.88; [EES vs. PES] adjusted HR=0.29, 95% CI 0.13-0.67). Similarly, patients treated with EES had a lower risk of target-lesion revascularization (TLR) compared to patients treated with SES and PES ([EES vs. SES], 5.6% vs. 11.5%, adjusted HR=0.68, 95% CI: 0.55-0.83; [EES vs. PES], 5.6% vs. 11.3%, adjusted HR=0.51, 95% CI: 0.33-0.77). There were no differences in other safety end points, such as all-cause mortality, cardiac mortality, myocardial infarction (MI) and MACE. CONCLUSION In diabetic patients, the unrestricted use of EES appears to be associated with improved outcomes, specifically a significant decrease in the need for TLR and ST compared to early generation SES and PES throughout 3-year follow-up.

Long-term outcomes of patients receiving zotarolimus-eluting stents in ST elevation myocardial infarction, non-ST elevation acute coronary syndrome, and stable angina: data from the Resolute program

BACKGROUND Outcome data are limited in patients with ST-segment elevation acute myocardial infarction (STEMI) or other acute coronary syndromes (ACSs) who receive a drug-eluting stent (DES). Data suggest that first generation DES is associated with an increased risk of stent thrombosis when used in STEMI. Whether this observation persists with newer generation DES is unknown. The study objective was to analyze the two-year safety and effectiveness of Resolute™ zotarolimus-eluting stents (R-ZESs) implanted for STEMI, ACS without ST segment elevation (non-STEACS), and stable angina (SA). METHODS Data from the Resolute program (Resolute All Comers and Resolute International) were pooled and patients with R-ZES implantation were categorized by indication: STEMI (n=335), non-STEACS (n=1416), and SA (n=1260). RESULTS Mean age was 59.8±11.3 years (STEMI), 63.8±11.6 (non-STEACS), and 64.9±10.1 (SA). Fewer STEMI patients had diabetes (19.1% vs. 28.5% vs. 29.2%; P<0.001), prior MI (11.3% vs. 27.2% vs. 29.4%; P<0.001), or previous revascularization (11.3% vs. 27.9% vs. 37.6%; P<0.001). Two-year definite/probable stent thrombosis occurred in 2.4% (STEMI), 1.2% (non-STEACS) and 1.1% (SA) of patients with late/very late stent thrombosis (days 31-720) rates of 0.6% (STEMI and non-STEACS) and 0.4% (SA) (P=NS). The two-year mortality rate was 2.1% (STEMI), 4.8% (non-STEACS) and 3.7% (SA) (P=NS). Death or target vessel re-infarction occurred in 3.9% (STEMI), 8.7% (non-STEACS) and 7.3% (SA) (P=0.012). CONCLUSION R-ZES in STEMI and in other clinical presentations is effective and safe. Long term outcomes are favorable with an extremely rare incidence of late and very late stent thrombosis following R-ZES implantation across indications.

Cholesteatoma surgery in children: long-term results of the inside-out technique

OBJECTIVE To present the anatomical and functional results of the inside-out technique applied in pediatric cholestetaoma surgery and to evaluate functionality with good hearing results against radicality with lower recurrence rate. METHODS Retrospective analysis and evaluation of the postoperative outcome in a consecutive series of 126 children or 130 ears operated between 1992 and 2008. With the inside-out technique, cholesteatoma is eradicated from the epitympanum toward the mastoid and, as a single stage procedure, functional reconstruction of the middle ear is achieved by tympanoossiculoplasty. RESULTS In 89.2% of all cases, the ear was dry postoperatively. 80.9% of the ears reached a postoperative air-bone gap of 30 dB or less and the median air conduction hearing threshold was 29 dB; in 60.9% of all cases, hearing was postoperatively improved. The recurrence rate was 16.2% in a mean postoperative follow-up 8.5 years. Altogether, 48 ears (36.9%) underwent revision surgery. The complication rate was 3.1% and involved only minor complications. CONCLUSION The inside-out technique allows a safe removal of cholesteatoma from the epitympanum toward the mastoid with a single-stage reconstruction of the ossicular chain. For this reason we support our individual approach, which allows creation of the smallest possible cavity for the size of the cholesteatoma. Our results confirm that the inside-out technique is effective in the treatment of pediatric cholesteatoma.

Non-Hebbian Long-Term Potentiation of Inhibitory Synapses in the Thalamus

The thalamus integrates and transmits sensory information to the neocortex. The activity of thalamocortical relay (TC) cells is modulated by specific inhibitory circuits. Although this inhibition plays a crucial role in regulating thalamic activity, little is known about long-term changes in synaptic strength at these inhibitory synapses. Therefore, we studied long-term plasticity of inhibitory inputs to TC cells in the posterior medial nucleus of the thalamus by combining patch-clamp recordings with two-photon fluorescence microscopy in rat brain slices. We found that specific activity patterns in the postsynaptic TC cell induced inhibitory long-term potentiation (iLTP). This iLTP was non-Hebbian because it did not depend on the timing between presynaptic and postsynaptic activity, but it could be induced by postsynaptic burst activity alone. iLTP required postsynaptic dendritic Ca2+ influx evoked by low-threshold Ca2+ spikes. In contrast, tonic postsynaptic spiking from a depolarized membrane potential (−50 mV), which suppressed these low-threshold Ca2+ spikes, induced no plasticity. The postsynaptic dendritic Ca2+ increase triggered the synthesis of nitric oxide that retrogradely activated presynaptic guanylyl cyclase, resulting in the presynaptic expression of iLTP. The dependence of iLTP on the membrane potential and therefore on the postsynaptic discharge mode suggests that this form of iLTP might occur during sleep, when TC cells discharge in bursts. Therefore, iLTP might be involved in sleep state-dependent modulation of thalamic information processing and thalamic oscillations.

The ubiquitin-proteasome system is necessary for long-term synaptic depression in Aplysia.

The neuropeptide Phe-Met-Arg-Phe-NH(2) (FMRFa) can induce transcription-dependent long-term synaptic depression (LTD) in Aplysia sensorimotor synapses. We investigated the role of the ubiquitin-proteasome system and the regulation of one of its components, ubiquitin C-terminal hydrolase (ap-uch), in LTD. LTD was sensitive to presynaptic inhibition of the proteasome and was associated with upregulation of ap-uch mRNA and protein. This upregulation appeared to be mediated by CREB2, which is generally regarded as a transcription repressor. Binding of CREB2 to the promoter region of ap-uch was accompanied by histone hyperacetylation, suggesting that CREB2 cannot only inhibit but also promote gene expression. CREB2 was phosphorylated after FMRFa, and blocking phospho-CREB2 blocked LTD. In addition to changes in the expression of ap-uch, the synaptic vesicle-associated protein synapsin was downregulated in LTD in a proteasome-dependent manner. These results suggest that proteasome-mediated protein degradation is engaged in LTD and that CREB2 may act as a transcription activator under certain conditions.

TGF-beta1 in Aplysia: role in long-term changes in the excitability of sensory neurons and distribution of TbetaR-II-like immunoreactivity.

Exogenous recombinant human transforming growth factor beta-1 (TGF-beta1) induced long-term facilitation of Aplysia sensory-motor synapses. In addition, 5-HT-induced facilitation was blocked by application of a soluble fragment of the extracellular portion of the TGF-beta1 type II receptor (TbetaR-II), which presumably acted by scavenging an endogenous TGF-beta1-like molecule. Because TbetaR-II is essential for transmembrane signaling by TGF-beta, we sought to determine whether Aplysia tissues contained TbetaR-II and specifically, whether neurons expressed the receptor. Western blot analysis of Aplysia tissue extracts demonstrated the presence of a TbetaR-II-immunoreactive protein in several tissue types. The expression and distribution of TbetaR-II-immunoreactive proteins in the central nervous system was examined by immunohistochemistry to elucidate sites that may be responsive to TGF-beta1 and thus may play a role in synaptic plasticity. Sensory neurons in the ventral-caudal cluster of the pleural ganglion were immunoreactive for TbetaR-II, as well as many neurons in the pedal, abdominal, buccal, and cerebral ganglia. Sensory neurons cultured in isolation and cocultured sensory and motor neurons were also immunoreactive. TGF-beta1 affected the biophysical properties of cultured sensory neurons, inducing an increase of excitability that persisted for at least 48 hr. Furthermore, exposure to TGF-beta1 resulted in a reduction in the firing threshold of sensory neurons. These results provide further support for the hypothesis that TGF-beta1 plays a role in long-term synaptic plasticity in Aplysia.

Bifurcation and singularity analysis of a molecular network for the induction of long-term memory.

Withdrawal reflexes of the mollusk Aplysia exhibit sensitization, a simple form of long-term memory (LTM). Sensitization is due, in part, to long-term facilitation (LTF) of sensorimotor neuron synapses. LTF is induced by the modulatory actions of serotonin (5-HT). Pettigrew et al. developed a computational model of the nonlinear intracellular signaling and gene network that underlies the induction of 5-HT-induced LTF. The model simulated empirical observations that repeated applications of 5-HT induce persistent activation of protein kinase A (PKA) and that this persistent activation requires a suprathreshold exposure of 5-HT. This study extends the analysis of the Pettigrew model by applying bifurcation analysis, singularity theory, and numerical simulation. Using singularity theory, classification diagrams of parameter space were constructed, identifying regions with qualitatively different steady-state behaviors. The graphical representation of these regions illustrates the robustness of these regions to changes in model parameters. Because persistent protein kinase A (PKA) activity correlates with Aplysia LTM, the analysis focuses on a positive feedback loop in the model that tends to maintain PKA activity. In this loop, PKA phosphorylates a transcription factor (TF-1), thereby increasing the expression of an ubiquitin hydrolase (Ap-Uch). Ap-Uch then acts to increase PKA activity, closing the loop. This positive feedback loop manifests multiple, coexisting steady states, or multiplicity, which provides a mechanism for a bistable switch in PKA activity. After the removal of 5-HT, the PKA activity either returns to its basal level (reversible switch) or remains at a high level (irreversible switch). Such an irreversible switch might be a mechanism that contributes to the persistence of LTM. The classification diagrams also identify parameters and processes that might be manipulated, perhaps pharmacologically, to enhance the induction of memory. Rational drug design, to affect complex processes such as memory formation, can benefit from this type of analysis.

A model of the roles of essential kinases in the induction and expression of late long-term potentiation.

The induction of late long-term potentiation (L-LTP) involves complex interactions among second-messenger cascades. To gain insights into these interactions, a mathematical model was developed for L-LTP induction in the CA1 region of the hippocampus. The differential equation-based model represents actions of protein kinase A (PKA), MAP kinase (MAPK), and CaM kinase II (CAMKII) in the vicinity of the synapse, and activation of transcription by CaM kinase IV (CAMKIV) and MAPK. L-LTP is represented by increases in a synaptic weight. Simulations suggest that steep, supralinear stimulus-response relationships between stimuli (e.g., elevations in [Ca(2+)]) and kinase activation are essential for translating brief stimuli into long-lasting gene activation and synaptic weight increases. Convergence of multiple kinase activities to induce L-LTP helps to generate a threshold whereby the amount of L-LTP varies steeply with the number of brief (tetanic) electrical stimuli. The model simulates tetanic, -burst, pairing-induced, and chemical L-LTP, as well as L-LTP due to synaptic tagging. The model also simulates inhibition of L-LTP by inhibition of MAPK, CAMKII, PKA, or CAMKIV. The model predicts results of experiments to delineate mechanisms underlying L-LTP induction and expression. For example, the cAMP antagonist RpcAMPs, which inhibits L-LTP induction, is predicted to inhibit ERK activation. The model also appears useful to clarify similarities and differences between hippocampal L-LTP and long-term synaptic strengthening in other systems.

Intra-hippocampal administration of the VEGF receptor blocker PTK787/ZK222584 impairs long-term memory.

A number of studies have established a role for vascular endothelial growth factor (VEGF) in angiogenesis. Recent reports have shown that VEGF overexpression in the hippocampus improves learning and memory and is associated with enhanced neurogenesis. PTK787/ZK222584 (PTK/ZK) is a reported inhibitor of VEGFR signaling that is currently being tested for its effects on lung and colon cancer. However, the influence of this drug on cognition has not been examined. In the present study, we questioned if post-training administration of PTK/ZK influences hippocampus-dependent memory. When administered to rats immediately following massed training in the Morris water maze, PTK/ZK impaired spatial memory retention tested 48 h later. This impairment was evidenced by increased latency to the hidden platform and fewer platform crossings. However, this impairment was not associated with a change in neurogenesis during this time frame. PTK/ZK infusion did not reduce VEGFR or AKT phosphorylation, but increased the phosphorylation of ERK. These studies suggest that VEGFR inhibitors such as PTK/ZK may negatively influence cognition.

The ubiquitin-proteasome system is necessary for long-term synaptic depression in Aplysia.

The neuropeptide Phe-Met-Arg-Phe-NH(2) (FMRFa) can induce transcription-dependent long-term synaptic depression (LTD) in Aplysia sensorimotor synapses. We investigated the role of the ubiquitin-proteasome system and the regulation of one of its components, ubiquitin C-terminal hydrolase (ap-uch), in LTD. LTD was sensitive to presynaptic inhibition of the proteasome and was associated with upregulation of ap-uch mRNA and protein. This upregulation appeared to be mediated by CREB2, which is generally regarded as a transcription repressor. Binding of CREB2 to the promoter region of ap-uch was accompanied by histone hyperacetylation, suggesting that CREB2 cannot only inhibit but also promote gene expression. CREB2 was phosphorylated after FMRFa, and blocking phospho-CREB2 blocked LTD. In addition to changes in the expression of ap-uch, the synaptic vesicle-associated protein synapsin was downregulated in LTD in a proteasome-dependent manner. These results suggest that proteasome-mediated protein degradation is engaged in LTD and that CREB2 may act as a transcription activator under certain conditions.

Glutamate iontophoresis induces long-term potentiation in the absence of evoked presynaptic activity

$\rm\underline{L}$ong-$\rm\underline{t}$erm $\rm\underline{p}$otentiation (LTP) is a candidate cellular mechanism underlying mammalian learning and memory. Protocols that induce LTP typically involve afferent stimulation. The experiments described in this dissertation tested the hypothesis that LTP induction does not require presynaptic activity. The significance of this hypothesis is underscored by results suggesting that LTP expression may involve activity-dependent presynaptic changes. An induction protocol using glutamate iontophoresis was developed that reliably induces LTP in hippocampal slices without afferent stimulation (ionto-LTP). Ionto-LTP is induced when excitatory postsynaptic potentials are completely blocked with adenosine and $\rm\underline{t}$etrodo$\rm\underline{t}$o$\rm\underline{x}$in (TTX). These results suggest constraints on the involvement of presynaptic mechanisms and putative retrograde messengers in LTP induction and expression; namely, these processes must function without many forms of activity-dependent presynaptic processes.^ In testing the role of pre-and postsynaptic mechanisms in LTP expression whole-cell recordings were used to examine the frequency and amplitude of $\rm\underline{s}$pontaneous $\rm\underline{e}$xcitatory $\rm\underline{p}$o$\rm\underline{s}$ynaptic $\rm\underline{c}$urrents (sEPSCs) in CA1 pyramidal neurons. sEPSCs where comprised of an equal mixture of TTX insensitive miniature EPSCs and sEPSCs that appeared to result from spontaneous action potentials (i.e., TTX sensitive EPSCs). The detection of all sEPSCs was virtually eliminated by CNQX, suggesting that sEPSCs were glutamate mediated synaptic events. Changes in the amplitude and frequency sEPSCs were examined during the expression of ionto-LTP to obtain new information about the cellular location of mechanisms involved in synaptic plasticity. The findings of this dissertation show that ionto-LTP expression results from increased sEPSC amplitude in the absence of lasting increases in sEPSC frequency. Potentiation of sEPSC amplitude without changes in sEPSC frequency has been previously interpreted to be due to postsynaptic mechanisms. Although this interpretation is supported by findings from peripheral synapses, its application to the central nervous system is unclear. Therefore, alternative mechanisms are also considered in this dissertation. Models based on increased release probability for action potential dependent transmitter release appear insufficient to explain our results. The most straightforward interpretation of the results in this dissertation is that LTP induced by glutamate iontophoresis on dendrites of CA1 pyramidal neurons is mediated by postsynaptic mechanisms. ^