Preliminary validation of the Diversion Risk Scale (DRS): Using environmental and individual factors to predict opioid-related diversion events.

The standard of care calls for the assessment of patients with chronic pain prior to the initiation of opioids, with one part of this assessment including assessment of the risk of misuse of medications. However, traditional opioid risk assessment tools focus almost entirely on individual factors and on the risk of misuse and addiction to opioids. Diversion of opioid medications has been found to be not uncommon, but to date, there have been no assessment tools specifically designed to assess the risk of diversion. In this study, we developed a measure designed specifically to assess the risk of an opioid medication ending up in the hands of someone other than the chronic pain patient to whom they were prescribed. A 15-item measure, the Diversion Risk Scale, was created and administered to 85 patients at a chronic pain practice. Results found that the measure had acceptable predictive validity. It was moderately correlated with traditional opioid risk assessment tools and showed improved ability to predict specific indicators of diversion. Diversion has been an understudied phenomenon, and the clinical value of an assessment tool that can help predict diversion in the chronic pain population is discussed.

Tapentadol is the least common opioid found in admission urine drug-test results at an intensive outpatient opioid-use disorder treat-ment program in Ohio: A brief report.

BACKGROUND: Previous data suggest that tapentadol, an atypical opioid with a putative dual mechanism of action, has relatively low rates of abuse. A better understanding of the rates of abuse among different prescription opioids may help clinicians when considering their potential risks and benefits. The results of urine drug tests (UDTs) may provide a unique opportunity to help answer this question. METHOD: To investigate different rates of prescription-opioid abuse in this retrospective study, we examined urine drug test results from patients seeking treatment at four facilities of an opioid-use-disorder (OUD) treatment program in Ohio. Urine specimens were collected on admission, one from each patient, in the regular course of care. The opioids reviewed in the present study were tapentadol, hydrocodone, oxycodone, hydromorphone, oxymorphone, and tramadol. Drug dispensing data, including morphine-milligram equivalents (MME) dispensed, were examined to adjust for the relative prevalence of each opioid being examined. RESULTS: Data from 4,162 patients were examined. Tapentadol was the least common finding in UDT results in this cohort and remained so after adjusting for drug availability. The percentage of specimens positive for a given opioid ranged from 0.12 percent (tapentadol) to 7.04 percent (oxycodone). The availability and MME adjustments resulted in a change of rank order, with tapentadol remaining the lowest but tramadol replacing oxycodone as the prescription opioid with the highest rate of abuse. CONCLUSIONS: In this sample of UDT results from patients seeking treatment at an OUD program in Ohio, tapentadol was the least frequent finding among the opioids examined, and this remained true when adjusting for dispensing data. Factors potentially contributing to this difference may include pharmacological properties unique to tapentadol. Several important limitations notwithstanding, these findings are consistent with previous real-world evidence and warrant an ongoing line of inquiry.

Injectable Capsaicin for the Management of Pain Due to Osteoarthritis.

Capsaicin is a potent agonist of the TRPV1 channel, a transduction channel that is highly expressed in nociceptive fibers (pain fibers) throughout the peripheral nervous system. Given the importance of TRPV1 as one of several transduction channels in nociceptive fibers, much research has been focused on the potential therapeutic benefits of using TRPV1 antagonists for the management of pain. However, an antagonist has two limitations. First, an antagonist in principle generally only affects one receptor. Secondly, most antagonists must have an ongoing presence on the receptor to have an effect. Capsaicin overcomes both liabilities by disrupting peripheral terminals of nociceptive fibers that express TRPV1, and thereby affects all of the potential means of activating that pain fiber (not just TRPV1 function). This disruptive effect is dependent on the dose and can occur within minutes. Thus, unlike a typical receptor antagonist, continued bioavailability at the level of the receptor is not necessary. By disrupting the entire terminal of the TRPV1-expressing nociceptive fiber, capsaicin blocks all the activation mechanisms within that fiber, and not just TRPV1 function. Topical capsaicin, an FDA approved treatment for neuropathic pain, addresses pain from abnormal nociceptor activity in the superficial layers of the skin. Effects after a single administration are evident over a period of weeks to months, but in time are fully reversible. This review focuses on the rationale for using capsaicin by injection for painful conditions such as osteoarthritis (OA) and provides an update on studies completed to date.

Number Of Clinical Trial Study Sites Impacts Observed Treatment Effect Size: An Analysis Of Randomized Controlled Trials Of Opioids For Chronic Pain.

BACKGROUND: Many aspects of study conduct impact the observed effect size of treatment. Data were utilized from a recently published meta-analysis of randomized, double-blind, placebo-controlled, clinical trials performed for the United States Food and Drug Administration (FDA) approval of full mu-agonist opioids for the treatment of chronic pain. METHODS: The number of study sites in each clinical trial and standardized effect size (SES) were extracted and computed. Standardized effect size was plotted against number of sites, and a two-piece linear model was fit to the plot. Ten studies were included. RESULTS: The SES decreased linearly by 0.13 units for every 10 sites (p=0.037), from 0.75 to 0.36, until an inflection point of 60 sites, after which SES did not decline further. The total number of subjects required for 90% power to discriminate drug from placebo increased from 78 to 336 subjects going from 30 to 60 sites. CONCLUSION: Results showed that the number of sites was a source of loss of assay sensitivity in clinical trials, which may contribute to the well-known problem of failure to successfully transition from Phase 2 to Phase 3 clinical development. Potential solutions include minimizing the number of sites, more rigorous and validated training, central statistical monitoring with rapid correction of performance issues, and more rigorous subject and site selection.

Evaluation of the oral human abuse potential of Oxycodone DETERx® formulation (Xtampza® ER).

OBJECTIVE: To further characterize the human abuse potential and pharmacokinetics (PK) of Oxycodone DETERx (Xtampza® ER) after intact and chewed oral administration. DESIGN: Randomized, double-blind, triple-dummy, active- and placebo-controlled, single-dose, six-period, crossover comparison study. SETTING: Clinical research unit. SUBJECTS: Adult, nondependent recreational opioid users who liked the effects of crushed immediate-release (IR) oxycodone in solution and were able to differentiate the effects from placebo solution. INTERVENTIONS: Oral administration of intact Oxycodone DETERx (fasted and fed), chewed Oxycodone DETERx (fasted and fed), crushed IR oxycodone (fasted), and placebo (fed). MAIN OUTCOME MEASURES: Subject ratings (100-point visual analog scales) of Drug Liking (primary measure) and Take Drug Again (key secondary measure). RESULTS: The pharmacodynamic (PD) analysis included 52 subjects who completed the study; the PK analysis included 71 subjects. Compared with crushed IR oxycodone fasted, the least-squares mean maximum effect (Emax) was statistically significant (p < 0.01) for Drug Liking and Take Drug Again, respectively, for chewed Oxycodone DETERx fasted (LS mean difference ± standard error of the mean: 13.1 ± 2.2 and 10.0 ± 3.2 points) and fed (10.9 ± 2.2 and 9.7 ± 3.3 points) and intact Oxycodone DETERx fasted (12.2 ± 2.2 and 9.3 ± 3.3 points) and fed (10.3 ± 2.2 and 9.2 ± 3.3 points). Results were consistent for other PD measures (Good Effects, Feeling High). Chewed Oxycodone DETERx fasted and fed treatments were bioequivalent to the respective intact treatments based on PK parameters. CONCLUSIONS: This study showed that when chewed or swallowed intact, under fasted or fed conditions, Oxycodone DETERx had statistically significantly lower abuse potential via the oral route compared with IR oxycodone.

Analgesic efficacy, safety, and tolerability of a long-acting abuse-deterrent formulation of oxycodone for moderate-to-severe chronic low back pain in subjects successfully switched from immediate-release oxycodone.

OBJECTIVES: This post hoc analysis of data from a randomized, double-blind, placebo-controlled, enriched-enrollment randomized-withdrawal Phase III study evaluated the safety, tolerability, and analgesic efficacy of Oxycodone DETERx extended-release (ER), abuse-deterrent capsules (Xtampza® ER) in subjects with chronic low back pain who were successfully transitioned from immediate-release (IR) oxycodone. METHODS: Continuous outcomes were analyzed using a mixed-model repeated-measures approach; binomial outcomes were analyzed using chi-squared; and time-to-event outcomes using Kaplan-Meier analyses. RESULTS: A total of 110 subjects previously prescribed IR oxycodone entered the Open-label Titration Phase. Forty-four subjects were randomized to Oxycodone DETERx (n=22) or placebo (n=22) in the 12-week Double-blind Maintenance Phase. Efficacy results in this subgroup showed a statistically significant difference between Oxycodone DETERx and placebo in average pain intensity scores from Randomization Baseline to Week 12 (least squares mean [± standard error], -1.88 [0.70]; P=0.0078). Additional efficacy results indicated that Oxycodone DETERx vs placebo was associated with a statistically significant benefit in durability of effect from Week 2 through Week 12 (P<0.01), numbers of subjects with a ≥30% (n [%] 10 [45.5%] vs 0 [0%]; P=0.0004) and ≥50% (10 [45.5%] vs 0 [0%]; P=0.0004) improvement in pain intensity, longer time-to-exit (P=0.0014), a greater number of subjects who completed the study (14 [63.6%] vs 4 [18.2%]), and less rescue medication use (acetaminophen; mean [SD], 163.5 [337.8] mg) vs 216.2 [377.3] mg). Adverse event profiles were consistent with opioid class effects and results from the original study; Oxycodone DETERx was well tolerated in subjects previously treated with short-acting oxycodone. CONCLUSIONS: Oxycodone DETERx resulted in clinically meaningful and statistically significant efficacy in subjects with chronic low back pain who were previously prescribed IR oxycodone and were successfully switched to ER Oxycodone DETERx.

Efficacy of opioids versus placebo in chronic pain: a systematic review and meta-analysis of enriched enrollment randomized withdrawal trials.

INTRODUCTION: Opioids have been used for millennia for the treatment of pain. However, the long-term efficacy of opioids to treat chronic non-cancer pain continues to be debated. To evaluate opioids' efficacy in chronic non-cancer pain, we performed a meta-analysis of published clinical trials for µ-opioid receptor agonists performed for US Food and Drug Administration approval. METHODS: MEDLINE and Cochrane trial register were searched for enriched enrollment randomized withdrawal studies (before June 2016). Selection criteria included: adults, ≥10 subjects per arm, any chronic pain condition, double-blind treatment period lasting ≥12 weeks, and all µ-agonist opioids approved in the USA. RESULTS: Fifteen studies met criteria. Opioid efficacy was statistically significant (p<0.001) versus placebo for pain intensity (standardized mean difference: -0.416), ≥30% and ≥50% improvement in pain (risk difference: 0.166 and 0.137), patient global impression of change (0.163), and patient global assessment of study medication (0.194). There were minor benefits on physical function and no effect on mental function. CONCLUSION: Opioids are efficacious in the treatment of chronic non-cancer pain for up to 3 months in randomized controlled trials. This should be considered, alongside data on opioid safety, in the use of opioids for the treatment of chronic pain.

Exploring the Interplay between Rescue Drugs, Data Imputation, and Study Outcomes: Conceptual Review and Qualitative Analysis of an Acute Pain Data Set.

In placebo-controlled acute surgical pain studies, provisions must be made for study subjects to receive adequate analgesic therapy. As such, most protocols allow study subjects to receive a pre-specified regimen of open-label analgesic drugs (rescue drugs) as needed. The selection of an appropriate rescue regimen is a critical experimental design choice. We hypothesized that a rescue regimen that is too liberal could lead to all study arms receiving similar levels of pain relief (thereby confounding experimental results), while a regimen that is too stringent could lead to a high subject dropout rate (giving rise to a preponderance of missing data). Despite the importance of rescue regimen as a study design feature, there exist no published review articles or meta-analysis focusing on the impact of rescue therapy on experimental outcomes. Therefore, when selecting a rescue regimen, researchers must rely on clinical factors (what analgesics do patients usually receive in similar surgical scenarios) and/or anecdotal evidence. In the following article, we attempt to bridge this gap by reviewing and discussing the experimental impacts of rescue therapy on a common acute surgical pain population: first metatarsal bunionectomy. The function of this analysis is to (1) create a framework for discussion and future exploration of rescue as a methodological study design feature, (2) discuss the interplay between data imputation techniques and rescue drugs, and (3) inform the readership regarding the impact of data imputation techniques on the validity of study conclusions. Our findings indicate that liberal rescue may degrade assay sensitivity, while stringent rescue may lead to unacceptably high dropout rates.

Fitness of Enterohemorrhagic Escherichia coli (EHEC)/Enteroaggregative E. coli O104:H4 in Comparison to That of EHEC O157: Survival Studies in Food and In Vitro.

In 2011, one of the world's largest outbreaks of hemolytic-uremic syndrome (HUS) occurred, caused by a rare Escherichia coli serotype, O104:H4, that shared the virulence profiles of Shiga toxin-producing E. coli (STEC)/enterohemorrhagic E. coli (EHEC) and enteroaggregative E. coli (EAEC). The persistence and fitness factors of the highly virulent EHEC/EAEC O104:H4 strain, grown either in food or in vitro, were compared with those of E. coli O157 outbreak-associated strains. The log reduction rates of the different EHEC strains during the maturation of fermented sausages were not significantly different. Both the O157:NM and O104:H4 serotypes could be shown by qualitative enrichment to be present after 60 days of sausage storage. Moreover, the EHEC/EAEC O104:H4 strain appeared to be more viable than E. coli O157:H7 under conditions of decreased pH and in the presence of sodium nitrite. Analysis of specific EHEC strains in experiments with an EHEC inoculation cocktail showed a dominance of EHEC/EAEC O104:H4, which could be isolated from fermented sausages for 60 days. Inhibitory activities of EHEC/EAEC O104:H4 toward several E. coli strains, including serotype O157 strains, could be determined. Our study suggests that EHEC/EAEC O104:H4 is well adapted to the multiple adverse conditions occurring in fermented raw sausages. Therefore, it is strongly recommended that STEC strain cocktails composed of several serotypes, instead of E. coli O157:H7 alone, be used in food risk assessments. The enhanced persistence of EHEC/EAEC O104:H4 as a result of its robustness, as well as the production of bacteriocins, may account for its extraordinary virulence potential. IMPORTANCE: In 2011, a severe outbreak caused by an EHEC/EAEC serovar O104:H4 strain led to many HUS sequelae. In this study, the persistence of the O104:H4 strain was compared with those of other outbreak-relevant STEC strains under conditions of fermented raw sausage production. Both O157:NM and O104:H4 strains could survive longer during the production of fermented sausages than E. coli O157:H7 strains. E. coli O104:H4 was also shown to be well adapted to the multiple adverse conditions encountered in fermented sausages, and the secretion of a bacteriocin may explain the competitive advantage of this strain in an EHEC strain cocktail. Consequently, this study strongly suggests that enhanced survival and persistence, and the presumptive production of a bacteriocin, may explain the increased virulence of the O104:H4 outbreak strain. Furthermore, this strain appears to be capable of surviving in a meat product, suggesting that meat should not be excluded as a source of potential E. coli O104:H4 infection.

Ethanol Production by Selected Intestinal Microorganisms and Lactic Acid Bacteria Growing under Different Nutritional Conditions.

To gain some specific insight into the roles microorganisms might play in non-alcoholic fatty liver disease (NAFLD), some intestinal and lactic acid bacteria and one yeast (Anaerostipes caccae, Bacteroides thetaiotaomicron, Bifidobacterium longum, Enterococcus fecalis, Escherichia coli, Lactobacillus acidophilus, Lactobacillus fermentum, Lactobacillus plantarum, Weissella confusa, Saccharomyces cerevisiae) were characterized by high performance liquid chromatography for production of ethanol when grown on different carbohydrates: hexoses (glucose and fructose), pentoses (arabinose and ribose), disaccharides (lactose and lactulose), and inulin. Highest amounts of ethanol were produced by S. cerevisiae, L. fermentum, and W. confusa on glucose and by S. cerevisiae and W. confusa on fructose. Due to mannitol-dehydrogenase expressed in L. fermentum, ethanol production on fructose was significantly (P < 0.05) reduced. Pyruvate and citrate, two potential electron acceptors for regeneration of NAD(+)/NADP(+), drastically reduced ethanol production with acetate produced instead in L. fermentum grown on glucose and W. confusa grown on glucose and fructose, respectively. In fecal slurries prepared from feces of four overweight volunteers, ethanol was found to be produced upon addition of fructose. Addition of A. caccae, L. acidophilus, L. fermentum, as well as citrate and pyruvate, respectively, abolished ethanol production. However, addition of W. confusa resulted in significantly (P < 0.05) increased production of ethanol. These results indicate that microorganisms like W. confusa, a hetero-fermentative, mannitol-dehydrogenase negative lactic acid bacterium, may promote NAFLD through ethanol produced from sugar fermentation, while other intestinal bacteria and homo- and hetero-fermentative but mannitol-dehydrogenase positive lactic acid bacteria may not promote NAFLD. Also, our studies indicate that dietary factors interfering with gastrointestinal microbiota and microbial metabolism may be important in preventing or promoting NAFLD.

Influence of fermentation on glucosinolates and glucobrassicin degradation products in sauerkraut.

A systematic investigation was carried out on the influence of fermentation on glucosinolates and their degradation products from fresh raw cabbage, throughout fermentation at 20 °C and storage at 4 °C. Glucosinolates were degraded dramatically between Day 2 and 5 of fermentation and by Day 7 there was no detectable amount of glucosinolates left. Fermentation led to formation of potential bioactive compounds ascorbigen (13.0 µmol/100 g FW) and indole-3-carbinol (4.52 µmol/100g FW) with their higher concentrations from Day 5 to Day 9. However, during storage indole-3-carbinol slowly degraded to 0.68 µmol/100 g FW, while ascorbigen was relatively stable from Week 4 until Week 8 at 6.78 µmol/100 g FW. In contrast, the content of indole-3-acetonitrile decreased rapidly during fermentation from 3.6 to 0.14 µmol/100 g FW. The results imply a maximum of health beneficial compounds after fermentation (7-9 days) in contrast to raw cabbage or stored sauerkraut.

Endogenous opioid activity in the anterior cingulate cortex is required for relief of pain.

Pain is aversive, and its relief elicits reward mediated by dopaminergic signaling in the nucleus accumbens (NAc), a part of the mesolimbic reward motivation pathway. How the reward pathway is engaged by pain-relieving treatments is not known. Endogenous opioid signaling in the anterior cingulate cortex (ACC), an area encoding pain aversiveness, contributes to pain modulation. We examined whether endogenous ACC opioid neurotransmission is required for relief of pain and subsequent downstream activation of NAc dopamine signaling. Conditioned place preference (CPP) and in vivo microdialysis were used to assess negative reinforcement and NAc dopaminergic transmission. In rats with postsurgical or neuropathic pain, blockade of opioid signaling in the rostral ACC (rACC) inhibited CPP and NAc dopamine release resulting from non-opioid pain-relieving treatments, including peripheral nerve block or spinal clonidine, an α2-adrenergic agonist. Conversely, pharmacological activation of rACC opioid receptors of injured, but not pain-free, animals was sufficient to stimulate dopamine release in the NAc and produce CPP. In neuropathic, but not sham-operated, rats, systemic doses of morphine that did not affect withdrawal thresholds elicited CPP and NAc dopamine release, effects that were prevented by blockade of ACC opioid receptors. The data provide a neural explanation for the preferential effects of opioids on pain affect and demonstrate that engagement of NAc dopaminergic transmission by non-opioid pain-relieving treatments depends on upstream ACC opioid circuits. Endogenous opioid signaling in the ACC appears to be both necessary and sufficient for relief of pain aversiveness.

The genus Weissella: taxonomy, ecology and biotechnological potential.

Bacteria assigned to the genus Weissella are Gram-positive, catalase-negative, non-endospore forming cells with coccoid or rod-shaped morphology (Collins et al., 1993; Björkroth et al., 2009, 2014) and belong to the group of bacteria generally known as lactic acid bacteria. Phylogenetically, the Weissella belong to the Firmicutes, class Bacilli, order Lactobacillales and family Leuconostocaceae (Collins et al., 1993). They are obligately heterofermentative, producing CO2 from carbohydrate metabolism with either d(-)-, or a mixture of d(-)- and l(+)- lactic acid and acetic acid as major end products from sugar metabolism. To date, there are 19 validly described Weissella species known. Weissella spp. have been isolated from and occur in a wide range of habitats, e.g., on the skin and in the milk and feces of animals, from saliva, breast milk, feces and vagina of humans, from plants and vegetables, as well as from a variety of fermented foods such as European sourdoughs and Asian and African traditional fermented foods. Thus, apart from a perceived technical role of certain Weissella species involved in such traditional fermentations, specific Weissella strains are also receiving attention as potential probiotics, and strain development of particularly W. cibaria strains is receiving attention because of their high probiotic potential for controlling periodontal disease. Moreover, W. confusa and W. cibaria strains are known to produce copius amounts of novel, non-digestible oligosaccharides and extracellular polysaccharides, mainly dextran. These polymers are receiving increased attention for their potential application as prebiotics and for a wide range of industrial applications, predominantly for bakeries and for the production of cereal-based fermented functional beverages. On the detrimental side, strains of certain Weissella species, e.g., of W. viridescens, W. cibaria and W. confusa, are known as opportunistic pathogens involved in human infections while strains of W. ceti have been recently recongnized as etiological agent of "weissellosis," which is a disease affecting farmed rainbow trouts. Bacteria belonging to this species thus are important both from a technological, as well as from a medical point of view, and both aspects should be taken into account in any envisaged biotechnological applications.

Opioid and noradrenergic contributions of tapentadol in experimental neuropathic pain.

Tapentadol is a dual action molecule with mu opioid agonist and norepinephrine (NE) reuptake blocking activity that has recently been introduced for the treatment of moderate to severe pain. The effects of intraperitoneal (i.p.) morphine (10mg/kg), tapentadol (10 or 30 mg/kg) or duloxetine (30 mg/kg), a norepinephrine/serotonin (NE/5HT) reuptake inhibitor, were evaluated in male, Sprague-Dawley rats with spinal nerve ligation (SNL) or sham surgery. Additionally, the effects of these drugs on spinal cerebrospinal fluid (CSF) NE levels were quantified. Response thresholds to von Frey filament stimulation decreased significantly from baseline in SNL, but not sham, operated rats. Duloxetine, tapentadol and morphine produced significant and time-related reversal of tactile hypersensitivity. Duloxetine significantly increased spinal CSF NE levels in both sham and SNL rats and no significant differences were observed in these groups. Tapentadol (10 mg/kg) produced a significant increase in spinal NE levels in SNL, but not in sham, rats. At the higher dose (30 mg/kg), tapentadol produced a significant increase in spinal CSF NE levels in both SNL and sham groups; however, spinal NE levels were elevated for an extended period in the SNL rats. This could be detected 30 min following tapentadol (30 mg/kg) in both sham and SNL groups. Surprisingly, while the dose of morphine studied reversed tactile hypersensitivity in nerve-injured rats, CSF NE levels were significantly reduced in both sham- and SNL rats. The data suggest that tapentadol elicits enhanced elevation in spinal NE levels in a model of experimental neuropathic pain offering a mechanistic correlate to observed clinical efficacy in this pain state.

Metagenomic cellulases highly tolerant towards the presence of ionic liquids--linking thermostability and halotolerance.

Cellulose is an important renewable resource for the production of bioethanol and other valuable compounds. Several ionic liquids (ILs) have been described to dissolve water-insoluble cellulose and/or wood. Therefore, ILs would provide a suitable reaction medium for the enzymatic hydrolysis of cellulose if cellulases were active and stable in the presence of high IL concentrations. For the discovery of novel bacterial enzymes with elevated stability in ILs, metagenomic libraries from three different hydrolytic communities (i.e. an enrichment culture inoculated with an extract of the shipworm Teredo navalis, a biogas plant sample and elephant faeces) were constructed and screened. Altogether, 14 cellulolytic clones were identified and subsequently assayed in the presence of six different ILs. The most promising enzymes, CelA2, CelA3 (both derived from the biogas plant) and CelA84 (derived from elephant faeces), showed high activities (up to 6.4 U/mg) in the presence of 30% (v/v) ILs. As these enzymes were moderately thermophilic and halotolerant, they retained 40% to 80% relative activity after 34 days in 4 M NaCl, and they were benchmarked with two thermostable enzymes, CelA from Thermotoga maritima and Cel5K from a metagenome library derived from Avachinsky crater in Kamchatka. These enzymes also exhibited high activity (up to 11.1 U/mg) in aqueous IL solutions (30% (v/v)). Some of the enzymes furthermore exhibited remarkable stability in 60% (v/v) IL. After 4 days, CelA3 and Cel5K retained up to 79% and 100% of their activity, respectively. Altogether, the obtained data suggest that IL tolerance appears to correlate with thermophilicity and halotolerance.

Epibacterial community patterns on marine macroalgae are host-specific but temporally variable.

Marine macroalgae are constantly exposed to epibacterial colonizers. The epiphytic bacterial patterns and their temporal and spatial variability on host algae are poorly understood. To investigate the interaction between marine macroalgae and epiphytic bacteria, this study tested if the composition of epibacterial communities on different macroalgae was specific and persisted under varying biotic and abiotic environmental conditions over a 2-year observation time frame. Epibacterial communities on the co-occurring macroalgae Fucus vesiculosus, Gracilaria vermiculophylla and Ulva intestinalis were repeatedly sampled in summer and winter of 2007 and 2008. The epibacterial community composition was analysed by denaturing gradient gel electrophoresis (DGGE) and 16S rRNA gene libraries. Epibacterial community profiles did not only differ significantly at each sampling interval among algal species, but also showed consistent seasonal differences on each algal species at a bacterial phylum level. These compositional patterns re-occurred at the same season of two consecutive years. Within replicates of the same algal species, the composition of bacterial phyla was subject to shifts at the bacterial species level, both within the same season but at different years and between different seasons. However, 7-16% of sequences were identified as species specific to the host alga. These findings demonstrate that marine macroalgae harbour species-specific and temporally adapted epiphytic bacterial biofilms on their surfaces. Since several algal host-specific bacteria were highly similar to other bacteria known to either avoid subsequent colonization by eukaryotic larvae or to exhibit potent antibacterial activities, algal host-specific bacterial associations are expected to play an important role for marine macroalgae.

Protein kinase C activation modulates reversible increase in cortical blood-brain barrier permeability and tight junction protein expression during hypoxia and posthypoxic reoxygenation.

Hypoxia (Hx) is a component of many disease states including stroke. Ischemic stroke occurs when there is a restriction of cerebral blood flow and oxygen to part of the brain. During the ischemic, and subsequent reperfusion phase of stroke, blood-brain barrier (BBB) integrity is lost with tight junction (TJ) protein disruption. However, the mechanisms of Hx and reoxygenation (HR)-induced loss of BBB integrity are not fully understood. We examined the role of protein kinase C (PKC) isozymes in modifying TJ protein expression in a rat model of global Hx. The Hx (6% O(2)) induced increased hippocampal and cortical vascular permeability to 4 and 10 kDa dextran fluorescein isothiocyanate (FITC) and endogenous rat-IgG. Cortical microvessels revealed morphologic changes in nPKC-θ distribution, increased nPKC-θ and aPKC-ζ protein expression, and activation by phosphorylation of nPKC-θ (Thr538) and aPKC-ζ (Thr410) residues after Hx treatment. Claudin-5, occludin, and ZO-1 showed disrupted organization at endothelial cell margins, whereas Western blot analysis showed increased TJ protein expression after Hx. The PKC inhibition with chelerythrine chloride (5 mg/kg intraperitoneally) attenuated Hx-induced hippocampal vascular permeability and claudin-5, PKC (θ and ζ) expression, and phosphorylation. This study supports the hypothesis that nPKC-θ and aPKC-ζ signaling mediates TJ protein disruption resulting in increased BBB permeability.