Leveraging the Full Continuum of Care to Prevent Opioid Use Disorder.

Substance use disorder prevention programs are most effective when matched appropriately to the baseline risk of the population. Individuals who misuse opioids often have unique risk profiles different from those who use other substances such as alcohol or cannabis. However, most substance use prevention programs are geared toward universal audiences, neglecting key inflection points along the continuum of care. The HEAL Prevention Cooperative (HPC) is a unique cohort of research projects that represents a continuum of care, from community-level universal prevention to indicated prevention among older adolescents and young adults who are currently misusing opioids or other substances. This paper describes the theoretical basis for addressing opioid misuse and opioid use disorder across the prevention continuum, using examples from research projects in the HPC.

Synthesis and Characterization of Ligand-Stabilized Silver Nanoparticles and Comparative Antibacterial Activity against E. coli.

Silver is a well-established antimicrobial agent. Conjugation of organic ligands with silver nanoparticles has been shown to create antimicrobial nanoparticles with improved pharmacodynamic properties and reduced toxicity. Twelve novel organic ligand functionalized silver nanoparticles (AgNPs) were prepared via a light-controlled reaction with derivatives of benzothiazole, benzoxazine, quinazolinone, 2-butyne-1,4-diol, 3-butyne-1-ol, and heptane-1,7-dioic. UV-vis, Fourier-transform infrared (FTIR) spectroscopy, and energy-dispersive X-ray (EDAX) analysis were used to confirm the successful formation of ligand-functionalized nanoparticles. Dynamic light scattering (DLS) revealed mean nanoparticle diameters between 25 and 278 nm. Spherical and nanotube-like morphologies were observed using transmission electron microscopy (TEM) and scanning electron microscopy (SEM). Seven of the twelve nanoparticles exhibited strong antimicrobial activity and five of the twelve demonstrated significant antibacterial capabilities against E. coli in a zone-of-inhibition assay. The synthesis of functionalized silver nanoparticles such as the twelve presented is critical for the further development of silver-nanoconjugated antibacterial agents.

Investigation of copper oxidation states in plasmonic nanomaterials by XAS and Raman spectroscopy.

Plasmonic core-shell-isolated nanoparticles are promising nanoplatforms for photocatalysis and for low detection analysis. This paper describes the characterization of a 2,2'-bipyridine phosphonate functionalized Ag@TiO2 nanocomposite which complexes copper ions by enhanced Raman spectroscopy and X-ray absorption (XANES and EXAFS). We distinguished Cu(i) from Cu(ii) complexes using shell-isolated nanoparticle enhanced Raman (SHINERS) combined with XAS spectroscopy.

Functionalized core-shell Ag@TiO2 nanoparticles for enhanced Raman spectroscopy: a sensitive detection method for Cu(ii) ions.

This paper demonstrates the use of surface plasmon resonance of core-shell Ag@TiO2 particles in SHINERS experiments. A copper(ii) complex grafted onto Ag@TiO2 surface was probed by Raman spectroscopy using resonance excitation profiles vs. excitation wavelengths (514, 633 and 785 nm) to tune the Raman signals. Enhancement factors of the SHINERS assembly have been estimated and compared to the SERS effect of unmodified silver NPs colloidal dispersions. Finally, the grafting of the copper(ii) complex onto Ag@TiO2 was advantageously compared to the grafting onto Ag@SiO2 shell.

Natural products show diverse mechanisms of action against Clostridium difficile.

AIMS: To investigate the mechanisms of action of natural products with bactericidal (cinnamon root powder, peppermint oil, trans-cinnamaldehyde, menthol and zingerone) or bacteriostatic (fresh garlic bulb extract, garlic clove powder, Leptospermum honey and allicin) activity against two Clostridium difficile strains. METHODS AND RESULTS: Bactericidal products significantly reduced intracellular ATP after 1 h (P ≤ 0·01), quantified using the BacTiter-Glo reagent, and damaged the cell membrane, shown by the leakage of both 260-nm-absorbing materials and protein, and the uptake of propidium iodide. Bacteriolysis was not observed, determined by measuring optical density of treated cell suspensions at 620-nm. The effect of three bacteriostatic products on protein synthesis was quantified using an Escherichia coli S30 extract system, with Leptospermum honey (16% w/v) showing significant inhibition (P < 0·01). Lastly, no products showed elevated minimum inhibitory concentrations against antimicrobial-resistant C. difficile, determined by broth microdilution. CONCLUSIONS: Cytoplasmic membrane damage was identified as a mechanism of action that may contribute to the activity of several natural products against C. difficile. SIGNIFICANCE AND IMPACT OF THE STUDY: This study describes the possible mechanisms of action of natural products against C. difficile, yet the efficacy in vivo to be determined.

In silico, in vitro and in vivo analysis of putative virulence factors identified in large clostridial toxin-negative, binary toxin- producing C. difficile strains.

The relevance of large clostridial toxin-negative, binary toxin-producing (A-B-CDT+) Clostridium difficile strains in human infection is still controversial. In this study, we investigated putative virulence traits that may contribute to the role of A-B-CDT+C. difficile strains in idiopathic diarrhea. Phenotypic assays were conducted on 148 strains of C. difficile comprising 10 different A-B-CDT+C. difficile ribotypes (RTs): 033, 238, 239, 288, 585, 586, QX143, QX444, QX521 and QX629. A subset of these isolates (n = 53) was whole-genome sequenced to identify genetic loci associated with virulence and survival. Motility studies showed that with the exception of RT 239 all RTs tested were non-motile. C. difficile RTs 033 and 288 had deletions in the F2 and F3 regions of their flagella operon while the F2 region was absent from strains of RTs 238, 585, 586, QX143, QX444, QX521 and QX629. The flagellin and flagella cap genes, fliC and fliD, respectively, involved in adherence and host colonization, were conserved in all strains, including reference strains. All A-B-CDT+C. difficile strains produced at least three extracellular enzymes (deoxyribonuclease, esterase and mucinase) indicating that these are important extracellular proteins. The toxicity of A-B-CDT+C. difficile strains in Vero cells was confirmed, however, pathogenicity was not demonstrated in a mouse model of disease. Despite successful colonization by most strains, there was no evidence of disease in mice. This study provides the first in-depth analysis of A-B-CDT+C. difficile strains and contributes to the current limited knowledge of these strains as a cause of C. difficile infection.

Effects of human rhinovirus on epithelial barrier integrity and function in children with asthma.

BACKGROUND: Bronchial epithelial tight junctions (TJ) have been extensively assessed in healthy airway epithelium. However, no studies have yet assessed the effect of human rhinovirus (HRV) infection on the expression and resultant barrier function in epithelial tight junctions (TJ) in childhood asthma. OBJECTIVES: To investigate the impact of HRV infection on airway epithelial TJ expression and barrier function in airway epithelial cells (AECs) of children with and without asthma. Furthermore, to test the hypothesis that barrier integrity and function is compromised to a greater extent by HRV in AECs from asthmatic children. METHODS: Primary AECs were obtained from children with and without asthma, differentiated into air-liquid interface (ALI) cultures and infected with rhinovirus. Expression of claudin-1, occludin and zonula occluden-1 (ZO-1) was assessed via qPCR, immunocytochemistry (ICC), in-cell western (ICW) and confocal microscopy. Barrier function was assessed by transepithelial electrical resistance (TER; RT ) and permeability to fluorescent dextran. RESULTS: Basal TJ gene expression of claudin-1 and occludin was significantly upregulated in asthmatic children compared to non-asthmatics; however, no difference was seen with ZO-1. Interestingly, claudin-1, occludin and ZO-1 protein expression was significantly reduced in AEC of asthmatic children compared to non-asthmatic controls suggesting possible post-transcriptional inherent differences. HRV infection resulted in a transient dissociation of TJ and airway barrier integrity in non-asthmatic children. Although similar dissociation of TJ was observed in asthmatic children, a significant and sustained reduction in TJ expression concurrent with both a significant decrease in TER and an increase in permeability in asthmatic children was observed. CONCLUSION: This study demonstrates novel intrinsic differences in TJ gene and protein expression between AEC of children with and without asthma. Furthermore, it correlates directly the relationship between HRV infection and the resultant dissociation of epithelial TJ that causes a continued altered barrier function in children with asthma.

Synthesis of a Reactive Oxygen Species-Responsive Doxorubicin Derivative.

A heterobifunctional reactive oxygen species (ROS)-responsive linker for directed drug assembly onto and delivery from a quantum dot (QD) nanoparticle carrier was synthesized and coupled to doxorubicin using N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC)/sulfo⁻NHS coupling. The doxorubicin conjugate was characterized using ¹H NMR and LC-MS and subsequently reacted under conditions of ROS formation (Cu2+/H2O2) resulting in successful and rapid thioacetal oxidative cleavage, which was monitored using ¹H NMR.

Impaired airway epithelial cell responses from children with asthma to rhinoviral infection.

BACKGROUND: The airway epithelium forms an effective immune and physical barrier that is essential for protecting the lung from potentially harmful inhaled stimuli including viruses. Human rhinovirus (HRV) infection is a known trigger of asthma exacerbations, although the mechanism by which this occurs is not fully understood. OBJECTIVE: To explore the relationship between apoptotic, innate immune and inflammatory responses to HRV infection in airway epithelial cells (AECs) obtained from children with asthma and non-asthmatic controls. In addition, to test the hypothesis that aberrant repair of epithelium from asthmatics is further dysregulated by HRV infection. METHODS: Airway epithelial brushings were obtained from 39 asthmatic and 36 non-asthmatic children. Primary cultures were established and exposed to HRV1b and HRV14. Virus receptor number, virus replication and progeny release were determined. Epithelial cell apoptosis, IFN-ß production, inflammatory cytokine release and epithelial wound repair and proliferation were also measured. RESULTS: Virus proliferation and release was greater in airway epithelial cells from asthmatics but this was not related to the number of virus receptors. In epithelial cells from asthmatic children, virus infection dampened apoptosis, reduced IFN-ß production and increased inflammatory cytokine production. HRV1b infection also inhibited wound repair capacity of epithelial cells isolated from non-asthmatic children and exaggerated the defective repair response seen in epithelial cells from asthmatics. Addition of IFN-ß restored apoptosis, suppressed virus replication and improved repair of airway epithelial cells from asthmatics but did not reduce inflammatory cytokine production. CONCLUSIONS: Collectively, HRV infection delays repair and inhibits apoptotic processes in epithelial cells from non-asthmatic and asthmatic children. The delayed repair is further exaggerated in cells from asthmatic children and is only partially reversed by exogenous IFN-ß.

A strategy for structuring and reporting a read-across prediction of toxicity.

Category formation, grouping and read across methods are broadly applicable in toxicological assessments and may be used to fill data gaps for chemical safety assessment and regulatory decisions. In order to facilitate a transparent and systematic approach to aid regulatory acceptance, a strategy to evaluate chemical category membership, to support the use of read-across predictions that may be used to fill data gaps for regulatory decisions is proposed. There are two major aspects of any read-across exercise, namely assessing similarity and uncertainty. While there can be an over-arching rationale for grouping organic substances based on molecular structure and chemical properties, these similarities alone are generally not sufficient to justify a read-across prediction. Further scientific justification is normally required to justify the chemical grouping, typically including considerations of bioavailability, metabolism and biological/mechanistic plausibility. Sources of uncertainty include a variety of elements which are typically divided into two main issues: the uncertainty associated firstly with the similarity justification and secondly the completeness of the read-across argument. This article focuses on chronic toxicity, whilst acknowledging the approaches are applicable to all endpoints. Templates, developed from work to prepare for the application of new toxicological data to read-across assessment, are presented. These templates act as proposals to assist in assessing similarity in the context of chemistry, toxicokinetics and toxicodynamics as well as to guide the systematic characterisation of uncertainty both in the context of the similarity rationale, the read across data and overall approach and conclusion. Lastly, a workflow for reporting a read-across prediction is suggested.

Threat-related learning relies on distinct dorsal prefrontal cortex network connectivity.

Conditioned changes in the emotional response to threat (e.g. aversive unconditioned stimulus; UCS) are mediated in part by the prefrontal cortex (PFC). Unpredictable threats elicit large emotional responses, while the response is diminished when the threat is predictable. A better understanding of how PFC connectivity to other brain regions varies with threat predictability would provide important insights into the neural processes that mediate conditioned diminution of the emotional response to threat. The present study examined brain connectivity during predictable and unpredictable threat exposure using a fear conditioning paradigm (previously published in Wood et al., 2012) in which unconditioned functional magnetic resonance imaging data were reanalyzed to assess effective connectivity. Granger causality analysis was performed using the time series data from 15 activated regions of interest after hemodynamic deconvolution, to determine regional effective connectivity. In addition, connectivity path weights were correlated with trait anxiety measures to assess the relationship between negative affect and brain connectivity. Results indicate the dorsomedial PFC (dmPFC) serves as a neural hub that influences activity in other brain regions when threats are unpredictable. In contrast, the dorsolateral PFC (dlPFC) serves as a neural hub that influences the activity of other brain regions when threats are predictable. These findings are consistent with the view that the dmPFC coordinates brain activity to take action, perhaps in a reactive manner, when an unpredicted threat is encountered, while the dlPFC coordinates brain regions to take action, in what may be a more proactive manner, to respond to predictable threats. Further, dlPFC connectivity to other brain regions (e.g. ventromedial PFC, amygdala, and insula) varied with negative affect (i.e. trait anxiety) when the UCS was predictable, suggesting that stronger connectivity may be required for emotion regulation in individuals with higher levels of negative affect.

Clostridium difficile infection diagnosis in a paediatric population: comparison of methodologies.

The increasing incidence of Clostridium difficile infection (CDI) in paediatric hospitalised populations, combined with the emergence of hypervirulent strains, community-acquired CDI and the need for prompt treatment and infection control, makes the rapid, accurate diagnosis of CDI crucial. We validated commonly used C. difficile diagnostic tests in a paediatric hospital population. From October 2011 to January 2012, 150 consecutive stools were collected from 75 patients at a tertiary paediatric hospital in Perth, Western Australia. Stools were tested using: C. Diff Quik Chek Complete, Illumigene C. difficile, GeneOhm Cdiff, cycloserine cefoxitin fructose agar (CCFA) culture, and cell culture cytotoxin neutralisation assay (CCNA). The reference standard was growth on CCFA or Cdiff Chromagar and PCR on isolates to detect tcdA, tcdB, cdtA, and cdtB. Isolates were PCR ribotyped. The prevalence of CDI was high (43 % of patients). Quik Chek Complete glutamate dehydrogenase (GDH) demonstrated a low negative predictive value (NPV) (93 %). Both CCNA and Quik Chek Complete toxin A/B had poor sensitivity (33 % and 29 % respectively). Molecular methods both had 89 % sensitivity. Algorithms using GDH + Illumigene or GeneOhm reduced the sensitivity to 85 % and 83 % respectively. Ribotype UK014/20 predominated. GDH NPV and GeneOhm and Illumigene sensitivities were reduced compared with adult studies. Quik Chek Complete and CCNA cannot reliably detect toxigenic CDI. A GDH first algorithm showed reduced sensitivity. In a high prevalence paediatric population, molecular methods alone are recommended over the use of GDH algorithm or culture and CCNA, as they demonstrate the best test performance characteristics.

Considerations for determining the performance of ultraviolet light emitting diode fluid disinfection systems.

The International Ultraviolet Association (IUVA) Task Force was formed to develop guidelines regarding testing and reporting on performance of UV LED water disinfection systems. The goal was to provide clarity in a guidance document on measuring system performance across the global UV LED water disinfection system market. A review of current performance measurement protocols for mercury lamp based systems shows that the common elements of UV LED system performance measurement protocols should be as follows: specified standard for the amount of pathogen reduction required, the requirement that the validation testing be conducted by a competent facility, and that the system be continually monitored by UV sensors while in use to verify system performance unless pathogen reduction is not claimed. UV LEDs have selectable peak wavelengths, as opposed to mercury lamps that have fixed emission wavelength values. As a result of this difference, the following changes to protocols used to test mercury lamp systems are recommended. First, the use of disinfection benchmarks other than 254 nm dose, such as direct inactivation values, dose benchmarks referenced to 254 nm, and/or dose benchmarks at the UV LED emission wavelength that give the same inactivation as the original 254 nm UV dose benchmark. Second, the use of 254 nm UV water transmittance values as a placeholder, rather than an assumed correct value, for systems under test with LED wavelengths >250 nm and water transmittance values ≥87%. More research is needed for lower wavelengths and UVTs. Third, the recommendation that germicidal response UV sensors be used in UV LED based systems to ensure that the validated disinfection is delivered. Finally, additional LED-specific considerations were also noted. UV LEDs are also instant-on devices, making them ideal light sources for systems operated intermittently. Performance testing of systems operated intermittently should include a test to insure that pathogens do not migrate past the UV LEDs while the LEDs are off. UV LED devices have recognized protocols for determining the lifetime of the devices, as well as for measuring other device properties. Caution should be exercised in using these lifetime values for devices in UV disinfection systems, since the thermal environment of the devices may be different for protocol testing and disinfection system operation. PRACTITIONER POINTS: Validation of UVC LED fluid disinfection is necessary for point of use, point of entry, and municipal applications. The emission spectrum properties, considerations and measurements for output over lifetime, and unique system design considerations of UVC LEDs as light sources are factors that must be considered when evaluating fluid disinfection performance. The instant-on operation, system geometry, validation benchmarks, system sensing, water transmittance, and fouling must also be considered for UVC LED devices when evaluating fluid disinfection performance.

Synthesis of a 2,2'-bipyridyl functionalized oligovinylene-phenylene using Heck and Horner-Wadsworth-Emmons reactions and X-ray crystal structure of E-(4-(4-bromostyryl)phenyl)(methyl)sulfane.

The synthesis of a new 2,2'-bipyridyl functionalized oligovinylenephenylene (OVP-5) containing a methyl protected thiol using Heck coupling and the Horner-Wadsworth-Emmons reaction and is described. A key step involving a diisopropylcarbodiimide promoted dehydration of a stable ß-hydroxyphosphonate intermediate was identified. The structure of precursor E-(4-(4-bromostyryl)phenyl)(methyl)sulfane was determined using X-ray crystallography.

Cobalt(III)- and rhodium(III)-porphyrin complexes for the effective degradation of fentanyl: Biological inactivation and mechanistic insights.

Cobalt(III) and rhodium(III) complexes containing the water-soluble porphyrin ligand meso-tri(4-sulfonatophenyl)mono(4-carboxyphenyl)porphine (C1S3TPP), [Rh(C1S3TPP)]Nax•nH2O (1) and [Co(C1S3TPP)]Nax•nH2O (2) were prepared from the direct reaction of free porphyrin and metal chloride salts in refluxing MeOH/DMF or EtOH/H2O. Compounds 1 and 2 were characterized using UV-vis and 1H NMR spectroscopies, and high-resolution mass spectrometry. Cell culture based assays of opioid receptor activation showed that while the rhodium complex reduced fentanyl opioid activity 113-fold to an IC50 value of 1.7 µM, the cobalt complex reduced fentanyl activity by 160-fold to an IC50 value of 2.4 µM. An oxidative mechanism for fentanyl breakdown is proposed.

Oxidation of trimethoprim by ferrate(VI): kinetics, products, and antibacterial activity.

Kinetics, stoichiometry, and products of the oxidation of trimethoprim (TMP), one of the most commonly detected antibacterial agents in surface waters and municipal wastewaters, by ferrate(VI) (Fe(VI)) were determined. The pH dependent second-order rate constants of the reactions of Fe(VI) with TMP were examined using acid-base properties of Fe(VI) and TMP. The kinetics of reactions of diaminopyrimidine (DAP) and trimethoxytoluene (TMT) with Fe(VI) were also determined to understand the reactivity of Fe(VI) with TMP. Oxidation products of the reactions of Fe(VI) with TMP and DAP were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Reaction pathways of oxidation of TMP by Fe(VI) are proposed to demonstrate the cleavage of the TMP molecule to ultimately result in 3,4,5,-trimethoxybenzaldehyde and 2,4-dinitropyrimidine as among the final identified products. The oxidized products mixture exhibited no antibacterial activity against E. coli after complete consumption of TMP. Removal of TMP in the secondary effluent by Fe(VI) was achieved.

Poly[aqua(µ-vinyl-phospho-nato)cadmium].

The title compound, [Cd(C(2)H(3)O(3)P)(H(2)O)](n), was obtained from vinyl-phospho-nic acid and cadmium nitrate. The vinyl groups project into the inter-lamellar space and the structure is held together via van der Waals forces. The Cd(2+) ion is six-coordinate and the geometry is best described as distorted octa-hedral, with O-Cd-O angles falling within the range 61.72 (13)-101.82 (14)°. Five of the coordinated oxygen atoms originate from the phospho-nate group and the sixth from a bound water molecule. Cd-O distances lie between 2.220 (3) and 2.394 (2) Å. The water mol-ecule is hydrogen bonded to a phospho-nate oxygen atom.

Sex-related Differences in Stress Reactivity and Cingulum White Matter.

The prefrontal cortex and limbic system are important components of the neural circuit that underlies stress and anxiety. These brain regions are connected by white matter tracts that support neural communication including the cingulum, uncinate fasciculus, and the fornix/stria-terminalis. Determining the relationship between stress reactivity and these white matter tracts may provide new insight into factors that underlie stress susceptibility and resilience. Therefore, the present study investigated sex differences in the relationship between stress reactivity and generalized fractional anisotropy (GFA) of the white matter tracts that link the prefrontal cortex and limbic system. Diffusion weighted images were collected and deterministic tractography was completed in 104 young adults (55 men, 49 women; mean age = 18.87 SEM = 0.08). Participants also completed self-report questionnaires (e.g., Trait Anxiety) and donated saliva (later assayed for cortisol) before, during, and after the Trier Social Stress Test. Results revealed that stress reactivity (area under the curve increase in cortisol) and GFA of the cingulum bundle varied by sex. Specifically, men demonstrated greater cortisol reactivity and greater GFA within the cingulum than women. Further, an interaction between sex, stress reactivity, and cingulum GFA was observed in which men demonstrated a positive relationship while women demonstrated a negative relationship between GFA and cortisol reactivity. Finally, trait anxiety was positively associated with the GFA of the fornix/stria terminalis - the white matter pathways that connect the hippocampus/amygdala to the hypothalamus. These findings advance our understanding of factors that underlie individual differences in stress reactivity.

Clostridium difficile and One Health.

BACKGROUND: For over four decades, Clostridium difficile has been a significant enteric pathogen of humans. It is associated with the use of antimicrobials that generally disrupt the microbiota of the gastrointestinal tract. Previously, it was thought that C. difficile was primarily a hospital-acquired infection; however, with the emergence of community-associated cases, and whole-genome sequencing suggesting the majority of the hospital C. difficile infection (CDI) cases are genetically distinct from one another, there is compelling evidence that sources/reservoirs of C. difficile outside hospitals play a significant role in the transmission of CDI. OBJECTIVES: To review the 'One Health' aspects of CDI, focusing on how community sources/reservoirs might be acting as a conduit in the transfer of C. difficile between animals and humans. The importance of a One Health approach in managing CDI is discussed. SOURCES: A literature search was performed on PubMed and Web of Science for relevant papers published from 1 January 2000 to 10 July 2019. CONTENT: We present evidence that demonstrates transmission of C. difficile in hospitals from asymptomatic carriers to symptomatic CDI patients. The source of colonization is most probably community reservoirs, such as foods and the environment, where toxigenic C. difficile strains have frequently been isolated. With high-resolution genomic sequencing, the transmission of C. difficile between animals and humans can be demonstrated, despite a clear epidemiological link often being absent. The ways in which C. difficile from animals and humans can disseminate through foods and the environment are discussed, and an interconnected transmission pathway for C. difficile involving food animals, humans and the environment is presented. IMPLICATIONS: Clostridium difficile is a well-established pathogen of both humans and animals that contaminates foods and the environment. To manage CDI, a One Health approach with the collaboration of clinicians, veterinarians, environmentalists and policy-makers is paramount.

Blockade of K(ATP) channels reduces endothelial hyperpolarization and leukocyte recruitment upon reperfusion after hypoxia.

Ischemia/reperfusion injury in renal transplantation leads to slow or initial nonfunction, and predisposes to acute and chronic rejection. In fact, severe ischemia reperfusion injury can significantly reduce graft survival, even with modern immunosuppressive agents. One of the mechanisms by which ischemia/reperfusion causes injury is activation of endothelial cells resulting in inflammation. Although several therapies can be used to prevent leukocyte recruitment to ischemic vessels (e.g. antiadhesion molecule antibodies), there have been no clinical treatments reported that can prevent initial immediate neutrophil recruitment upon reperfusion. Using intravital microscopy, we describe abrogation of immediate neutrophil recruitment to ischemic microvessels by the K(ATP) antagonist glibenclamide (Glyburide). Further, we show that glibenclamide can reduce leukocyte recruitment in vitro under physiologic flow conditions. ATP-regulated potassium channels (K(ATP)) are important in the control of cell membrane polarization. Here we describe profound hyperpolarization of endothelial cells during hypoxia, and the reduction of this hyperpolarization using glibenclamide. These findings suggest that control of endothelial membrane potential during ischemia may be an important therapeutic tool in avoiding ischemia/reperfusion injury, and therefore, enhancing transplant long-term function.