A novel pan-negative-gating modulator of KCa2/3 channels, fluoro-di-benzoate, RA-2, inhibits endothelium-derived hyperpolarization-type relaxation in coronary artery and produces bradycardia in vivo.

Small/intermediate conductance KCa channels (KCa2/3) are Ca(2+)/calmodulin regulated K(+) channels that produce membrane hyperpolarization and shape neurologic, epithelial, cardiovascular, and immunologic functions. Moreover, they emerged as therapeutic targets to treat cardiovascular disease, chronic inflammation, and some cancers. Here, we aimed to generate a new pharmacophore for negative-gating modulation of KCa2/3 channels. We synthesized a series of mono- and dibenzoates and identified three dibenzoates [1,3-phenylenebis(methylene) bis(3-fluoro-4-hydroxybenzoate) (RA-2), 1,2-phenylenebis(methylene) bis(3-fluoro-4-hydroxybenzoate), and 1,4-phenylenebis(methylene) bis(3-fluoro-4-hydroxybenzoate)] with inhibitory efficacy as determined by patch clamp. Among them, RA-2 was the most drug-like and inhibited human KCa3.1 with an IC50 of 17 nM and all three human KCa2 subtypes with similar potencies. RA-2 at 100 nM right-shifted the KCa3.1 concentration-response curve for Ca(2+) activation. The positive-gating modulator naphtho[1,2-d]thiazol-2-ylamine (SKA-31) reversed channel inhibition at nanomolar RA-2 concentrations. RA-2 had no considerable blocking effects on distantly related large-conductance KCa1.1, Kv1.2/1.3, Kv7.4, hERG, or inwardly rectifying K(+) channels. In isometric myography on porcine coronary arteries, RA-2 inhibited bradykinin-induced endothelium-derived hyperpolarization (EDH)-type relaxation in U46619-precontracted rings. Blood pressure telemetry in mice showed that intraperitoneal application of RA-2 (≤100 mg/kg) did not increase blood pressure or cause gross behavioral deficits. However, RA-2 decreased heart rate by ≈145 beats per minute, which was not seen in KCa3.1(-/-) mice. In conclusion, we identified the KCa2/3-negative-gating modulator, RA-2, as a new pharmacophore with nanomolar potency. RA-2 may be of use to generate structurally new types of negative-gating modulators that could help to define the physiologic and pathomechanistic roles of KCa2/3 in the vasculature, central nervous system, and during inflammation in vivo.

Prediction of death in less than 60 minutes after withdrawal of cardiorespiratory support in potential organ donors after circulatory death.

BACKGROUND: Given the stable number of potential organ donors after brain death, donors after circulatory death have been an increasing source of organs procured for transplant. Among the most important considerations for donation after circulatory death (DCD) is the prediction that death will occur within a reasonable period of time after the withdrawal of cardiorespiratory support (WCRS). Accurate prediction of time to death is necessary for the procurement process. We aimed to develop simple predictive rules for death in less than 60 min and test the accuracy of these rules in a pool of potential DCD donors. METHODS: A multicenter prospective longitudinal cohort design of DCD eligible patients (n=318), with the primary binary outcome being death in less than 60 min after withdrawal of cardiorespiratory support conducted in 28 accredited intensive care units (ICUs) in Australia. We used a random split-half method to produce two samples, first to develop the predictive classification rules and then to estimate accuracy in an independent sample. RESULTS: The best classification model used only three simple classification rules to produce an overall efficiency of 0.79 (0.72-0.85), sensitivity of 0.82 (0.73-0.90), and a positive predictive value of 0.80 (0.70-0.87) in the independent sample. Using only intensive care unit specialist prediction (a single classification rule) produced comparable efficiency 0.80 (0.73-0.86), sensitivity 0.87 (0.78-0.93), and positive predictive value 0.78 (0.68-0.86). CONCLUSION: This best predictive model missed only 18% of all potential donors. A positive prediction would be incorrect on only 20% of occasions, meaning there is an acceptable level of lost opportunity costs involved in the unnecessary assembly of transplantation teams and theatres.

Exploring Australian intensive care physicians clinical judgement during Donation after Cardiac Death: an exploratory qualitative study.

BACKGROUND: Donation after Cardiac Death (DCD) is one possible solution to the world wide organ shortage. Intensive care physicians are central to DCD becoming successful since they are responsible for making the clinical judgements and decisions associated with DCD. Yet international evidence shows health care professionals have not embraced DCD and are often reluctant to consider it as an option for patients. PURPOSE: To explore intensive care physicians' clinical judgements when selecting a suitable DCD candidate. METHODS: Using interpretative exploratory methods six intensive care physicians were interviewed from three hospital sites in Australia. Following verbatim transcription, data was subjected to thematic analysis. FINDINGS: Three distinct themes emerged. Reducing harm and increasing benefit was a major focus of intensive care physicians during determination of DCD. There was an acceptance of DCD if there was clear evidence that donation was what the patient and family wanted. Characteristics of a defensible decision reflected the characteristics of sequencing, separation and isolation, timing, consensus and collaboration, trust and communication to ensure that judgements were robust and defensible. The final theme revealed the importance of minimising uncertainty and discomfort when predicting length of survival following withdrawal of life-sustaining treatment. CONCLUSION: DCD decisions are made within an environment of uncertainty due to the imprecision associated with predicting time of death. Lack of certainty contributed to the cautious and collaborative strategies used by intensive care physicians when dealing with patients, family members and colleagues around end-of-life decisions, initiation of withdrawal of life-sustaining treatment and the discussion about DCD. This study recommends that nationally consistent policies are urgently needed to increase the degree of certainty for intensive care staff concerning the DCD processes.

Prediction of death in less than 60 minutes following withdrawal of cardiorespiratory support in ICUs.

OBJECTIVES: Half of all ICU patients die within 60 minutes of withdrawal of cardiorespiratory support. Prediction of which patients die before and after 60 minutes would allow changes in service organization to improve patient palliation, family grieving, and allocation of ICU beds. This study tested various predictors of death within 60 minutes and explored which clinical variables ICU specialists used to make their prediction. DESIGN AND SETTINGS: Prospective longitudinal cohort design (n = 765) of consecutive adult patients having withdrawal of cardiorespiratory support, in 28 ICUs in Australia. Primary outcome was death within 60 minutes following withdrawal of cardiorespiratory support. A random split-half method was used to make two independent samples for development and testing of the predictive indices. The secondary outcome was ICU Specialist prediction of death within 60 minutes. MEASUREMENTS AND MAIN RESULTS: Death within 60 minutes of withdrawal of cardiorespiratory support occurred in 377 (49.3%). ICU specialist opinion was the best individual predictor, with an unadjusted odds ratio of 15.42 (95% CI, 9.33-25.49) and an adjusted odds ratio of 8.44 (4.30-16.58). A predictive index incorporating the ICU specialist opinion and clinical variables had an area under the curve of 0.89 (0.86-0.92) and 0.84 (0.80-0.88) in the development and test sets, respectively; and a second index using only clinical variables had an area under the curve of 0.86 (0.82-0.89) and 0.78 (0.73-0.83). The ICU specialist prediction of death within 60 minutes was independently associated with five clinical variables: pH, Glasgow Coma Scale, spontaneous respiratory rate, positive end-expiratory pressure, and systolic blood pressure. CONCLUSION: ICU specialist opinion is probably the current clinical standard for predicting death within 60 minutes of withdrawal of cardiorespiratory support. This approach is supported by this study, although predictive indices restricted to clinical variables are only marginally inferior. Either approach has a clinically useful level of prediction that would allow ICU service organization to be modified to improve care for patients and families and use ICU beds more efficiently.

Gold Digger or Video Girl: the salience of an emerging hip-hop sexual script.

Concerns have been expressed in the common discourse and scholarly literature about the negative influence of Hip-Hop on its young listeners' ideas about sex and sexuality. Most of the scholarly literature has focused on the impact of this urban, Black media on young African American girls' sexual self-concept and behaviours. In response to this discourse, Stephens and Phillips (2003) proposed a Hip-Hop sexual scripting model that theorises about specific sexual scripts for young African American women. Their model includes eight different sexual scripts including the Gold Digger script. The present study proposes a ninth emerging script - the Video Girl. Participants were 18 female African American college students, between the ages of 18 and 30 years old from a large urban public university in the Southwest USA. Using q-methodology the present study found support for the existence of a Video Girl script. In addition, the data indicates that this script is distinct but closely related to Stephens and Phillips' Gold Digger script. These findings support their theory by suggesting that Hip-Hop sexual scripts are salient and hold real meaning for this sample.

Prediction of death after withdrawal of life-sustaining treatments.

OBJECTIVE: To assess the predictive value of respiratory and haemodynamic variables and opinion of the intensivist for determining how soon death occurs after withdrawal of life-sustaining treatments (WLST). DESIGN: Multicentre prospective observational study. PARTICIPANTS AND SETTING: 83 consecutive adult intensive care patients at John Hunter and Calvary Mater Hospitals, Newcastle, New South Wales, for whom a decision was made to withdraw life-sustaining treatment between March 2007 and March 2008. MAIN OUTCOME MEASURES: Data were collected before initiation of palliation. Primary outcome was to recognise in a multivariate analysis the parameters associated with a time to death < or = 60 minutes after WLST. RESULTS: 81 patients underwent WLST: 79 died, and two survived to be discharged from hospital. Thirty-six patients (45%) died within 60 minutes of WLST, and 45 (55%) survived 60 minutes or longer. Mean ICU stay before WLST was 4.8 days (range, 1-85 days). Mean time from WLST to death was 6:31 h (range, 1 minute to 31 days). A modified University of Wisconsin assessment tool showed no statistical association with the time from WLST to death (P = 0.09). The adapted United Network for Organ Sharing tool, systolic blood pressure, APACHE II score, ventilatory dependence, oxygen disruption, Glasgow Coma Scale (GCS) score and staff specialist opinion all showed a statistically significant association with time from WLST to death (P < 0.05). CONCLUSIONS: It is possible to predict the time from WLST to death accurately using a tool that combines GCS, respiratory and haemodynamic parameters and intensivist opinion. These results require validation in a large multicentre study.

Misoprostol impairs female reproductive tract innate immunity against Clostridium sordellii.

Fatal cases of acute shock complicating Clostridium sordellii endometritis following medical abortion with mifepristone (also known as RU-486) used with misoprostol were reported. The pathogenesis of this unexpected complication remains enigmatic. Misoprostol is a pharmacomimetic of PGE(2), an endogenous suppressor of innate immunity. Clinical C. sordellii infections were associated with intravaginal misoprostol administration, suggesting that high misoprostol concentrations within the uterus impair immune responses against C. sordellii. We modeled C. sordellii endometritis in rats to test this hypothesis. The intrauterine but not the intragastric delivery of misoprostol significantly worsened mortality from C. sordellii uterine infection, and impaired bacterial clearance in vivo. Misoprostol also reduced TNF-alpha production within the uterus during infection. The intrauterine injection of misoprostol did not enhance mortality from infection by the vaginal commensal bacterium Lactobacillus crispatus. In vitro, misoprostol suppressed macrophage TNF-alpha and chemokine generation following C. sordellii or peptidoglycan challenge, impaired leukocyte phagocytosis of C. sordellii, and inhibited uterine epithelial cell human beta-defensin expression. These immunosuppressive effects of misoprostol, which were not shared by mifepristone, correlated with the activation of the G(s) protein-coupled E prostanoid (EP) receptors EP2 and EP4 (macrophages) or EP4 alone (uterine epithelial cells). Our data provide a novel explanation for postabortion sepsis leading to death and also suggest that PGE(2), in which production is exaggerated within the reproductive tract during pregnancy, might be an important causal determinant in the pathogenesis of more common infections of the gravid uterus.

Synthetic prostacyclin analogs differentially regulate macrophage function via distinct analog-receptor binding specificities.

PGI(2) (prostacyclin) is a lipid mediator with vasodilatory and antithrombotic effects used in the treatment of vasoconstrictive/ischemic diseases including pulmonary artery hypertension. However, emerging research supports a role for PGs, including PGI(2), in the regulation of both innate and acquired immunity. As PGI(2) is unstable, we sought to define the effects of various PGI(2) analogs on resident alveolar macrophage (AM) and peritoneal macrophage (PM) innate immune functions. The effects of iloprost, carbaprostacyclin, and treprostinil on the regulation of phagocytosis, bacterial killing, and inflammatory mediator production were determined in both macrophage populations from rats. Iloprost failed to suppress AM functions to the same degree that it did in PMs, a characteristic shared by carbaprostacyclin. This difference reflected greater expression of the G(alphas) protein-coupled I prostanoid receptor and greater cAMP generation in PMs than AMs. Treprostinil inhibited phagocytosis, bacterial killing, and cytokine generation in AMs to a much greater degree than the other PGI(2) analogs and more closely resembled the effects of PGE(2). Studies with the E prostanoid (EP) 2 receptor antagonist AH-6809 and EP2-null macrophages indicated that this was due in part to the previously unknown ability of treprostinil to stimulate the EP2 receptor. The present investigation for the first time identifies differences in immunoregulatory properties of clinically administered PGI(2) analogs. These studies are the first to explore the capacity of PGI(2) to regulate bacterial killing and phagocytosis in macrophages, and our findings may hold important consequences regarding the risk of infection for patients receiving such agents.