The heterodimeric hypoxia-inducible factor (HIF)-1 is a master transcriptional regulator of oxygen homeostasis and a possible target for gene therapy of ischemic disease. Although the role of oxygen ...concentration in HIF-1a protein stabilization is well established, it is less clear whether and how oxygen-regulated mechanisms contribute to HIF-1a protein modifications, nuclear translocation, heterodimerization with the b-subunit, recruitment of cofactors, and gene trans-activation. Because the HIF-1a protein is proteolytically degraded under normoxic conditions, we established two HeLa Tet-Off cell lines (HT42 and HT43), which inducibly overexpress high levels of HIF-1a under normoxic conditions, allowing to distinguish hypoxia-dependent from hypoxia-independent activation mechanisms. Using these cells, we found that normoxically induced HIF-1a is localized to the nucleus, binds DNA, and trans-activates reporter and endogenous target genes. The levels of p53 expression remained unaffected. The MAP kinase inhibitor PD98059 attenuated HIF-1a protein modifications and trans-activation ability but not protein stabilization and DNA-binding activity. Because overexpressed HIF-1a is fully localized to the nucleus but displays only partial DNA-binding and trans-activation activity, mitogen-activated protein kinase-dependent phosphorylation might be required for full HIF-1 activation. HIF-1a protein was also overexpressed in vivo, following the transplantation of HT42 cells into nude mice, demonstrating the feasibility of HIF-1a gene transfer.
OBJECTIVETo determine the influence of changes in acute physiology scores (APS) and other patient characteristics on predicting intensive care unit (ICU) readmission.
DESIGNSecondary analysis of a ...prospective cohort study.
SETTINGSingle large university medical intensive care unit.
PATIENTSA total of 4,684 consecutive admissions from Janu-ary 1, 1994, to April 1, 1998, to the medical ICU.
INTERVENTIONSNone.
MEASUREMENTS AND MAIN RESULTS The independent influence of patient characteristics, including daily APS, admission diagnosis, treatment status, and admission location, on ICU readmission was evaluated using logistic regression. After accounting for first ICU admission deaths, 3,310 patients were “at-risk” for ICU readmission and 317 were readmitted (9.6%). Hospital mortality was five times higher (43% vs. 8%;p < .0001), and length of stay was two times longer (16 ± 16 vs. 32 ± 28 days;p < .001) in readmitted patients. Mean discharge APS was significantly higher in the readmitted group compared with the not readmitted group (43 ± 19 vs. 34 ± 18;p > .01). Significant independent predictors of ICU readmission included discharge APS >40 (odds ratio OR 2.1; 95% confidence interval CI 1.6–2.7;p < .0001), admission to the ICU from a general medicine ward (Floor) (OR 1.9; 95% CI 1.4–2.6;p < .0001), and transfer to the ICU from other hospital (Transfer) (OR 1.7; 95% CI 1.3–2.3;p < .01). The overall model calibration and discrimination were (H-L χ2 = 3.8, df = 8;p = .85) and (receiver operating characteristic 0.67), respectively.
CONCLUSIONSPatients readmitted to medical ICUs have significantly higher hospital lengths of stay and mortality. ICU readmissions may be more common among patients who respond poorly to treatment as measured by increased severity of illness at first ICU discharge and failure of prior therapy at another hospital or on a general medicine unit. Tertiary care ICUs may have higher than expected readmission rates and mortalities, even when accounting for severity of illness, if they care for significant numbers of transferred patients.
Key points
Hyperexcitability and hypersynchrony of neuronal networks are thought to be linked to the generation of epileptic activity in both humans and animal models.
Here we show that human ...epileptic postoperative neocortical tissue is able to generate two different types of synchronies in vitro.
Epileptiform bursts occurred only in slices derived from epileptic patients and were hypersynchronous events characterized by high levels of excitability.
Spontaneous population activity emerged in both epileptic and non‐epileptic tissue, with a significantly lower degree of excitability and synchrony, and could not be linked to epilepsy.
These results help us to understand better the role of excitatory and inhibitory neuronal circuits in the generation of population events, and to define the subtle border between physiological and pathological synchronies.
Interictal activity is a hallmark of epilepsy diagnostics and is linked to neuronal hypersynchrony. Little is known about perturbations in human epileptic neocortical microcircuits, and their role in generating pathological synchronies. To explore hyperexcitability of the human epileptic network, and its contribution to convulsive activity, we investigated an in vitro model of synchronous burst activity spontaneously occurring in postoperative tissue slices derived from patients with or without preoperative clinical and electrographic manifestations of epileptic activity. Human neocortical slices generated two types of synchronies. Interictal‐like discharges (classified as epileptiform events) emerged only in epileptic samples, and were hypersynchronous bursts characterized by considerably elevated levels of excitation. Synchronous population activity was initiated in both epileptic and non‐epileptic tissue, with a significantly lower degree of excitability and synchrony, and could not be linked to epilepsy. However, in pharmacoresistant epileptic tissue, a higher percentage of slices exhibited population activity, with higher local field potential gradient amplitudes. More intracellularly recorded neurons received depolarizing synaptic potentials, discharging more reliably during the events. Light and electron microscopic examinations showed slightly lower neuron densities and higher densities of excitatory synapses in the human epileptic neocortex. Our data suggest that human neocortical microcircuits retain their functionality and plasticity in vitro, and can generate two significantly different synchronies. We propose that population bursts might not be pathological events while interictal‐like discharges may reflect the epileptogenicity of the human cortex. Our results show that hyperexcitability characterizes the human epileptic neocortical network, and that it is closely related to the emergence of synchronies.
Key points
Hyperexcitability and hypersynchrony of neuronal networks are thought to be linked to the generation of epileptic activity in both humans and animal models.
Here we show that human epileptic postoperative neocortical tissue is able to generate two different types of synchronies in vitro.
Epileptiform bursts occurred only in slices derived from epileptic patients and were hypersynchronous events characterized by high levels of excitability.
Spontaneous population activity emerged in both epileptic and non‐epileptic tissue, with a significantly lower degree of excitability and synchrony, and could not be linked to epilepsy.
These results help us to understand better the role of excitatory and inhibitory neuronal circuits in the generation of population events, and to define the subtle border between physiological and pathological synchronies.
Key points
•Initiation of pathological synchronous events such as epileptic spikes and seizures is linked to the hyperexcitability of the neuronal network in both humans and animals.
•In the present ...study, we show that epileptiform interictal‐like spikes and seizures emerged in human neocortical slices by blocking GABAA receptors, following the disappearance of the spontaneously occurring synchronous population activity.
•Large variability of temporally and spatially simple and complex spikes was generated by tissue from epileptic patients, whereas only simple events appeared in samples from non‐epileptic patients.
•Physiological population activity was associated with a moderate level of principal cell and interneuron firing, with a slight dominance of excitatory neuronal activity, whereas epileptiform events were mainly initiated by the synchronous and intense discharge of inhibitory cells.
•These results help us to understand the role of excitatory and inhibitory neurons in synchrony‐generating mechanisms, in both epileptic and non‐epileptic conditions.
Understanding the role of different neuron types in synchrony generation is crucial for developing new therapies aiming to prevent hypersynchronous events such as epileptic seizures. Paroxysmal activity was linked to hyperexcitability and to bursting behaviour of pyramidal cells in animals. Human data suggested a leading role of either principal cells or interneurons, depending on the seizure morphology. In the present study, we aimed to uncover the role of excitatory and inhibitory processes in synchrony generation by analysing the activity of clustered single neurons during physiological and epileptiform synchronies in human neocortical slices. Spontaneous population activity was detected with a 24‐channel laminar microelectrode in tissue derived from patients with or without preoperative clinical manifestations of epilepsy. This population activity disappeared by blocking GABAA receptors, and several variations of spatially and temporally simple or complex interictal‐like spikes emerged in epileptic tissue, whereas peritumoural slices generated only simple spikes. Around one‐half of the clustered neurons participated with an elevated firing rate in physiological synchronies with a slight dominance of excitatory cells. By contrast, more than 90% of the neurons contributed to interictal‐like spikes and seizures, and an intense and synchronous discharge of inhibitory neurons was associated with the start of these events. Intrinsically bursting principal cells fired later than other neurons. Our data suggest that a balanced excitation and inhibition characterized physiological synchronies, whereas disinhibition‐induced epileptiform events were initiated mainly by non‐synaptically synchronized inhibitory neurons. Our results further highlight the differences between humans and animal models, and between in vivo and (pharmacologically manipulated) in vitro conditions.
Key points
•Initiation of pathological synchronous events such as epileptic spikes and seizures is linked to the hyperexcitability of the neuronal network in both humans and animals.
•In the present study, we show that epileptiform interictal‐like spikes and seizures emerged in human neocortical slices by blocking GABAA receptors, following the disappearance of the spontaneously occurring synchronous population activity.
•Large variability of temporally and spatially simple and complex spikes was generated by tissue from epileptic patients, whereas only simple events appeared in samples from non‐epileptic patients.
•Physiological population activity was associated with a moderate level of principal cell and interneuron firing, with a slight dominance of excitatory neuronal activity, whereas epileptiform events were mainly initiated by the synchronous and intense discharge of inhibitory cells.
•These results help us to understand the role of excitatory and inhibitory neurons in synchrony‐generating mechanisms, in both epileptic and non‐epileptic conditions.
The true utility of quality measurement lies in its ability to inspire quality improvement, with resultant enhancements in the processes and outcomes of care. Because quality measurement is ...expensive, it is difficult to justify using measures that are not likely to lead to important improvements in health. Many current measures of chronic disease technical quality, however, have one or more pitfalls that prevent them from motivating quality improvement reactions. These pitfalls include that: (1) measured processes of care lack strong links to outcomes; (2) actionable processes of care are not measured; (3) measures do not target those at highest risk; (4) measures do not allow for patient exceptions; and (5) intermediate outcome measures are not severity adjusted. To exemplify recent advancements and current pitfalls in chronic disease quality measurement, we examine the evolution of quality measures for diabetes mellitus and discuss the limitations of many currently used diabetes mellitus care measures. We then propose more clinically meaningful "tightly linked" measures that examine clinical processes directly linked to outcomes, target populations with specific diagnoses or intermediate disease outcomes that contribute to risk for poor downstream health outcomes, and explicitly incorporate exceptions. We believe that using more tightly linked measures in quality assessment will identify important quality of care problems and is more likely to produce improved outcomes for those with chronic diseases.
Cav1.3 L-type Ca
2+
channels (LTCCs) in cochlear inner hair cells (IHCs) are essential for hearing as they convert sound-induced graded receptor potentials into tonic postsynaptic glutamate release. ...To enable fast and indefatigable presynaptic Ca
2+
signaling, IHC Cav1.3 channels exhibit a negative activation voltage range and uniquely slow inactivation kinetics. Interaction with CaM-like Ca
2+
-binding proteins inhibits Ca
2+
-dependent inactivation, while the mechanisms underlying slow voltage-dependent inactivation (VDI) are not completely understood. Here we studied if the complex formation of Cav1.3 LTCCs with the presynaptic active zone proteins RIM2α and RIM-binding protein 2 (RBP2) can stabilize slow VDI. We detected both RIM2α and RBP isoforms in adult mouse IHCs, where they co-localized with Cav1.3 and synaptic ribbons. Using whole-cell patch-clamp recordings (tsA-201 cells), we assessed their effect on the VDI of the C-terminal full-length Cav1.3 (Cav1.3
L
) and a short splice variant (Cav1.3
42A
) that lacks the C-terminal RBP2 interaction site. When co-expressed with the auxiliary β3 subunit, RIM2α alone (Cav1.3
42A
) or RIM2α/RBP2 (Cav1.3
L
) reduced Cav1.3 VDI to a similar extent as observed in IHCs. Membrane-anchored β2 variants (β2a, β2e) that inhibit inactivation on their own allowed no further modulation of inactivation kinetics by RIM2α/RBP2. Moreover, association with RIM2α and/or RBP2 consolidated the negative Cav1.3 voltage operating range by shifting the channel’s activation threshold toward more hyperpolarized potentials. Taken together, the association with “slow” β subunits (β2a, β2e) or presynaptic scaffolding proteins such as RIM2α and RBP2 stabilizes physiological gating properties of IHC Cav1.3 LTCCs in a splice variant-dependent manner ensuring proper IHC function.
PURPOSE: Although infections associated with indwelling urinary catheters are common, costly, and morbid, the use of these catheters is unnecessary in more than one-third of patients. We sought to ...assess whether attending physicians, medical residents, and medical students are aware if their hospitalized patients have an indwelling urinary catheter, and whether physician awareness is associated with appropriate use of these catheters.
METHODS: The physicians and medical students responsible for patients admitted to the medical services at four university-affiliated hospitals were given a list of the patients on their service. For each patient, the provider was asked: “As of yesterday afternoon, did this patient have an indwelling urethral catheter?” Respondents’ answers were compared with the results of examining the patient.
RESULTS: Among 288 physicians and students on 56 medical teams, 256 (89%) completed the survey. Of 469 patients, 117 (25%) had an indwelling catheter. There were a total of 319 provider-patient observations among these 117 patients. Overall, providers were unaware of catheterization for 88 (28%) of the 319 provider-patient observations. Unawareness rates by level of training were 21% for students, 22% for interns, 27% for residents, and 38% for attending physicians (
P = 0.06). Catheter use was inappropriate in 36 (31%) of the 117 patients with a catheter. Providers were unaware of catheter use for 44 (41%) of the 108 provider-patient observations of patients who were inappropriately catheterized. Catheterization was more likely to be appropriate if respondents were aware of the catheter (odds ratio = 3.7; 95% confidence interval, 2.1 to 6.7,
P <0.001).
CONCLUSION: Physicians are commonly unaware that their patients have an indwelling urinary catheter. Inappropriate catheters are more often “forgotten” than appropriate ones. System-wide interventions aimed at discontinuing unnecessary catheterization seem warranted.
-->Low voltage-activated Cav1.3 L-type Ca
2+
-channels are key regulators of neuronal excitability controlling neuronal development and different types of learning and memory. Their physiological ...functions are enabled by their negative activation voltage-range, which allows Cav1.3 to be active at subthreshold voltages. Alternative splicing in the C-terminus of their pore-forming α1-subunits gives rise to C-terminal long (Cav1.3
L
) and short (Cav1.3
S
) splice variants allowing Cav1.3
S
to activate at even more negative voltages than Cav1.3
L
. We discovered that inclusion of exons 8b, 11, and 32 in Cav1.3
S
further shifts activation (-3 to -4 mV) and inactivation (-4 to -6 mV) to more negative voltages as revealed by functional characterization in tsA-201 cells. We found transcripts of these exons in mouse chromaffin cells, the cochlea, and the brain. Our data further suggest that Cav1.3-containing exons 11 and 32 constitute a significant part of native channels in the brain. We therefore investigated the effect of these splice variants on human disease variants. Splicing did not prevent the gating defects of the previously reported human pathogenic variant S652L, which further shifted the voltage-dependence of activation of exon 11-containing channels by more than -12 mV. In contrast, we found no evidence for gating changes of the CACNA1D missense variant R498L, located in exon 11, which has recently been identified in a patient with an epileptic syndrome.
Our data demonstrate that alternative splicing outside the C-terminus involving exons 11 and 32 contributes to channel fine-tuning by stabilizing negative activation and inactivation gating properties of wild-type and mutant Cav1.3 channels.
-->Low voltage-activated Cav1.3 L-type Ca
-channels are key regulators of neuronal excitability controlling neuronal development and different types of learning and memory. Their physiological ...functions are enabled by their negative activation voltage-range, which allows Cav1.3 to be active at subthreshold voltages. Alternative splicing in the C-terminus of their pore-forming α1-subunits gives rise to C-terminal long (Cav1.3
) and short (Cav1.3
) splice variants allowing Cav1.3
to activate at even more negative voltages than Cav1.3
. We discovered that inclusion of exons 8b, 11, and 32 in Cav1.3
further shifts activation (-3 to -4 mV) and inactivation (-4 to -6 mV) to more negative voltages as revealed by functional characterization in tsA-201 cells. We found transcripts of these exons in mouse chromaffin cells, the cochlea, and the brain. Our data further suggest that Cav1.3-containing exons 11 and 32 constitute a significant part of native channels in the brain. We therefore investigated the effect of these splice variants on human disease variants. Splicing did not prevent the gating defects of the previously reported human pathogenic variant S652L, which further shifted the voltage-dependence of activation of exon 11-containing channels by more than -12 mV. In contrast, we found no evidence for gating changes of the
missense variant R498L, located in exon 11, which has recently been identified in a patient with an epileptic syndrome. Our data demonstrate that alternative splicing outside the C-terminus involving exons 11 and 32 contributes to channel fine-tuning by stabilizing negative activation and inactivation gating properties of wild-type and mutant Cav1.3 channels.
The goal of this study was to determine whether outcomes of nonemergent coronary artery bypass grafting (CABG) differed between low- and high-volume hospitals in patients at different levels of ...surgical risk.
Regionalizing all CABG surgeries from low- to high-volume hospitals could improve surgical outcomes but reduce patient access and choice. "Targeted" regionalization could be a reasonable alternative, however, if subgroups of patients that would clearly benefit from care at high-volume hospitals could be identified.
We assessed outcomes of CABG at 56 U.S. hospitals using 1997 administrative and clinical data from Solucient EXPLORE, a national outcomes benchmarking database. Predicted in-hospital mortality rates for subjects were calculated using a logistic regression model, and subjects were classified into five groups based on surgical risk: minimal (< 0.5%), low (0.5% to 2%), moderate (2% to 5%), high (5% to 20%), and severe (> or =20%). We assessed differences in in-hospital mortality, hospital costs and length of stay between low- and high-volume facilities (defined as > or =200 annual cases) in each of the five risk groups.
A total of 2,029 subjects who underwent CABG at 25 low-volume hospitals and 11,615 subjects who underwent CABG at 31 high-volume hospitals were identified. Significant differences in in-hospital mortality were seen between low- and high-volume facilities in subjects at moderate (5.3% vs. 2.2%; p = 0.007) and high risk (22.6% vs. 11.9%; p = 0.0026) but not in those at minimal, low or severe risk. Hospital costs and lengths of stay were similar across each of the five risk groups. Based on these results, targeted regionalization of subjects at moderate risk or higher to high-volume hospitals would have resulted in an estimated 370 transfers and avoided 16 deaths; in contrast, full regionalization would have led to 2,029 transfers and avoided 20 deaths.
Targeted regionalization might be a feasible strategy for balancing the clinical benefits of regionalization with patients' desires for choice and access.
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