Physiological Stress Integrates Resistance to Rattlesnake Venom and the Onset of Risky Foraging in California Ground Squirrels.

Using venom for predation often leads to the evolution of resistance in prey. Understanding individual variation in venom resistance is key to unlocking basic mechanisms by which antagonistic coevolution can sustain variation in traits under selection. For prey, the opposing challenges of predator avoidance and resource acquisition often lead to correlated levels of risk and reward, which in turn can favor suites of integrated morphological, physiological and behavioral traits. We investigate the relationship between risk-sensitive behaviors, physiological resistance to rattlesnake venom, and stress in a population of California ground squirrels. For the same individuals, we quantified foraging decisions in the presence of snake predators, fecal corticosterone metabolites (a measure of "stress"), and blood serum inhibition of venom enzymatic activity (a measure of venom resistance). Individual responses to snakes were repeatable for three measures of risk-sensitive behavior, indicating that some individuals were consistently risk-averse whereas others were risk tolerant. Venom resistance was lower in squirrels with higher glucocorticoid levels and poorer body condition. Whereas resistance failed to predict proximity to and interactions with snake predators, individuals with higher glucocorticoid levels and in lower body condition waited the longest to feed when near a snake. We compared alternative structural equation models to evaluate alternative hypotheses for the relationships among stress, venom resistance, and behavior. We found support for stress as a shared physiological correlate that independently lowers venom resistance and leads to squirrels that wait longer to feed in the presence of a snake, whereas we did not find evidence that resistance directly facilitates latency to forage. Our findings suggest that stress may help less-resistant squirrels avoid a deadly snakebite, but also reduces feeding opportunities. The combined lethal and non-lethal effects of stressors in predator-prey interactions simultaneously impact multiple key traits in this system, making environmental stress a potential contributor to geographic variation in trait expression of toxic predators and resistant prey.

Determination of thiorphan, a racecadotril metabolite, in human plasma by LC-MS/MS and its application to a bioequivalence study in Chinese subjects
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OBJECTIVE: The objective of this study was to use LC-MS/MS to compare the pharmacodynamic properties and bioequivalence of two 200-mg formulations of racecadotril: suspension formulation (test) and granule formulation (reference) in healthy Chinese subjects. MATERIALS AND METHODS: A single-dose, randomized, two-period crossover study was conducted in fasted healthy Chinese subjects, who received a single oral dose of the test or reference formulation, followed by a 7-day washout period and administration of the alternate formulation. RESULTS: The rapid and highly sensitive LC-MS/MS method exhibited a reasonable linearity range (2.324 - 952.000 ng/mL) and high sensitivity (LLOQ of 2.324 ng/mL). The within- and between-run precision, accuracy, and stability results were within the acceptable limits, and no matrix effect was observed. The 90% CI of the ratio of geometric means for AUC0-t, AUC0-∞, and Cmax were 88.1 - 102.3%, 87.9 - 101.5% and 99.5 - 113%, respectively, which met the regulatory criteria for bioequivalence. CONCLUSION: The method is suitable for quantification of thiorphan in human plasma. In addition, the results indicated that the test and reference formulations were bioequivalent in terms of both rate and extent of absorption.

An MMP/TIMP ratio scoring system as a potential predictive marker of diabetic foot ulcer healing.

OBJECTIVES: The mechanism of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in diabetic foot ulcers (DFUs) is unclear. The purpose of this study was to describe changes in MMP-1, MMP-9, and TIMP-1 levels during DFU healing, and to search for any correlation in the changes in MMP levels with wound healing, in order to find possible predictors of healing. METHODS: Patients with a DFU were recruited and placed into two groups, according to the degree of wound healing: 'good healers' and 'poor healers'. Levels of MMP-1, MMP-9, and TIMP-1 were analysed by ELISA (enzyme-linked immunosorbent assay). RESULTS: A total of 22 patients participated in the study. The MMP-1 level was significantly higher at weeks zero (W0) and 12 (W12) in 'good healers' than in 'poor healers' (p=0.045 and 0.008, respectively). In contrast, the MMP-9 level was significantly lower in 'good healers' than in 'poor healers' at W0, W4, and W12 (p=0.001, 0.001 and 0.028, respectively). Receiver operator curve (ROC) analysis of the MMP-9 level, MMP-1/TIMP-1 ratio, and MMP-9/TIMP-1 ratio at W0 provided cut-off levels of 0.38, 0.056, and 9.06, respectively, which were best predictive of a reduction in wound area at W4 ('good healers' versus 'poor healers'; thereby predicting wound healing condition at W12) with a sensitivity of 81.8%, 81.8%, and 90.9%, and a specificity of 64.6%, 55%, and 64.6%, respectively. CONCLUSION: A 'poor healing scoring system' is therefore proposed that could be determined on patient admission, which has the potential to be used clinically as a predictor of healing, thus allowing an appropriate treatment plan to be developed.

Serum asunaprevir concentrations showing correlation with the extent of liver fibrosis as a factor inducing liver injuries in patients with genotype-1b hepatitis C virus receiving daclatasvir plus asunaprevir therapy.

AIMS: Liver injury can occur during antiviral therapies with direct-acting antivirals (DAAs), potentially necessitating discontinuation of the therapies, with consequent worsening of the sustained viral response (SVR) rates, in patients with hepatitis C virus (HCV). To clarify the mechanisms involved in serum transaminase level elevation, we performed a retrospective evaluation of the serum concentrations of daclatasvir and asunaprevir, both classified as DAAs, in patients receiving treatment with a combination of the two drugs. METHODS: Subjects were 278 Japanese patients with genotype-1b HCV who received daclatasvir plus asunaprevir therapy for more than 4 weeks. Serum concentrations of both the DAAs were measured at 4 weeks after the initiation of therapy. RESULT: Liver injuries including serum AST and/or ALT level elevation to 150 U/L or over were found in 34 patients (12.2%). Multivariate logistic regression analysis identified serum asunaprevir concentrations as being significantly associated with developing liver injury, with an odds ratio of 1.046 (95% confidence interval 1.011-1.082, p<0.05). Serum asunaprevir concentrations showed correlation with the extent of liver fibrosis, estimated by peripheral platelets counts and serum albumin levels and baseline and FIB4 index and serum Mac-2 binding protein glycosylation isomer (M2BPGi) levels at 4 weeks of the therapy; the concentrations were significantly higher among patients showing 3.0 or more of M2BPGi levels than among those with the levels less than 3.0; on the other hand, no such correlation/difference was found in serum daclatasvir concentrations. CONCLUSION: High serum concentrations of serum asunaprevir, which were associated with the extent of liver fibrosis, appear to provoke the occurrence of liver injury in patients with genotype-1b HCV receiving combined daclatasvir plus asunaprevir therapy.

Serum α1-proteinase inhibitor concentrations in dogs with exocrine pancreatic disease, chronic hepatitis or proteinuric chronic kidney disease.

Serum canine α1-proteinase inhibitor (cα1-PI) concentrations were evaluated in dogs with pancreatitis (n=24), exocrine pancreatic insufficiency (EPI; n=29), chronic hepatitis (CH; n=11) or proteinuric chronic kidney disease (CKD-P; n=61) to determine whether systemic proteinase/proteinase-inhibitor balance is altered in these conditions. Dogs with CKD-P had significantly lower cα1-PI concentrations than dogs with pancreatitis, EPI or CH; 16% of dogs with CKD-P had serum cα1-PI concentrations below the reference interval. Serum and urine cα1-PI concentrations were inversely correlated in dogs with CKD-P, but not in dogs with CH. This suggests that renal loss of cα1-PI contributes to decreased serum concentrations in dogs with CKD-P, while hepatic cα1-PI synthesis with CH either is not compromised or is counterbalanced by extrahepatic production.

Auxiliary activation of the complement system and its importance for the pathophysiology of clinical conditions.

Activation and regulation of the cascade systems of the blood (the complement system, the coagulation/contact activation/kallikrein system, and the fibrinolytic system) occurs via activation of zymogen molecules to specific active proteolytic enzymes. Despite the fact that the generated proteases are all present together in the blood, under physiological conditions, the activity of the generated proteases is controlled by endogenous protease inhibitors. Consequently, there is remarkable little crosstalk between the different systems in the fluid phase. This concept review article aims at identifying and describing conditions where the strict system-related control is circumvented. These include clinical settings where massive amounts of proteolytic enzymes are released from tissues, e.g., during pancreatitis or post-traumatic tissue damage, resulting in consumption of the natural substrates of the specific proteases and the available protease inhibitor. Another example of cascade system dysregulation is disseminated intravascular coagulation, with canonical activation of all cascade systems of the blood, also leading to specific substrate and protease inhibitor elimination. The present review explains basic concepts in protease biochemistry of importance to understand clinical conditions with extensive protease activation.

Interactions of aliskiren, a direct renin inhibitor, with antiepileptic drugs in the test of maximal electroshock in mice.

Experimental studies showed that certain angiotensin-converting enzyme inhibitors and angiotensin AT1 receptor antagonists can decrease seizure severity in rodents. Additionally, some of these blockers of the renin-angiotensin system have been documented to enhance the anticonvulsant activity of antiepileptic drugs against maximal electroshock-induced seizures. The aim of the current study was to investigate the effect of aliskiren, a direct renin inhibitor and a novel antihypertensive drug, on the protective action of numerous antiepileptic drugs (carbamazepine, valproate, clonazepam, phenobarbital, oxcarbazepine, lamotrigine, topiramate and pregabalin) in the test of maximal electroshock in mice. The examined drugs were administered intraperitoneally. Aliskiren up to a dose of 75mg/kg did not affect the threshold for electroconvulsions, however, aliskiren (75mg/kg) enhanced the anticonvulsant action of clonazepam and valproate. Following aliskiren treatment, a higher brain concentration of valproate was noted, suggesting a pharmacokinetic interaction. In the rota-rod test, the concomitant treatment with aliskiren (50 or 75mg/kg) and clonazepam (22.6mg/kg) impaired motor coordination while clonazepam (22.6mg/kg) alone showed strong tendency towards this impairment. The combination of aliskiren (75mg/kg) with phenobarbital (25.5mg/kg) caused long-term memory deficits in the passive avoidance task. This study shows that there are no negative interactions between aliskiren and the examined antiepileptic drugs as concerns their anticonvulsant activity. Aliskiren even potentiated the anticonvulsant action of clonazepam and valproate against maximal electroshock. The impact of aliskiren alone on seizure activity or on the anticonvulsant and adverse activity of antiepileptic drugs needs further evaluation in other animal models of seizures.

In vitro investigations on extracellular proteins secreted by Aphanomyces invadans, the causative agent of epizootic ulcerative syndrome.

BACKGROUND: Proteases produced by many microorganisms, including oomycetes, are crucial for their growth and development. They may also play a critical role in disease manifestation. Epizootic ulcerative syndrome is one of the most destructive fish diseases known. It is caused by the oomycete Aphanomyces invadans and leads to mass mortalities of cultured and wild fish in many countries. The areas of concern are Australia, China, Japan, South and Southeast Asian countries and the USA. Extracellular proteases produced by this oomycete are believed to trigger EUS pathogenesis in fish. To address this activity, we collected the extracellular products (ECP) of A. invadans and identified the secreted proteins using SDS-PAGE and mass spectrometery. A. invadans was cultivated in liquid Glucose-Peptone-Yeats media. The culture media was ultra-filtered through 10 kDa filters and analysed using SDS-PAGE. Three prominent protein bands from the SDS gel were excised and identified by mass spectrometery. Furthermore, we assessed their proteolytic effect on casein and immunoglobulin M (IgM) of rainbow trout (Oncorhynchus mykiss) and giant gourami (Osphronemus goramy). Antiprotease activity of the fish serum was also investigated. RESULTS: BLASTp analysis revealed that the prominent secreted proteins were proteases, mainly of the serine and cysteine types. Proteins containing fascin-like domain and bromodomain were also identified. We could demonstrate that the secreted proteases showed proteolytic activity against the casein and the IgM of both fish species. The anti-protease activity experiment showed that the percent inhibition of the common carp serum was 94.2% while that of rainbow trout and giant gourami serum was 7.7 and 12.9%, respectively. CONCLUSIONS: The identified proteases, especially serine proteases, could be the potential virulence factors in A. invadans and, hence, are candidates for further functional and host-pathogen interaction studies. The role of identified structural proteins in A. invadans also needs to be investigated further.

Serum alarm antiproteases in systemic sclerosis patients.

Alarm antiproteases, i.e. secretory leukocyte protease inhibitor ad elafin, are key mediators in innate immune response and integrate innate and adaptive immunity systems. The aim of the study was to assess clinical significance of serum levels of alarm antiproteases, elafin and secretory leukocyte protease inhibitor (SLPI) in patients with systemic sclerosis (SSc). Twenty-eight patients with SSc, 25 patients with rheumatoid arthritis (RA) and 22 healthy controls were recruited. Serum elafin and SLPI levels were examined using enzyme-linked immunosorbent assay (ELISA). The patients with SSc had significantly increased serum levels of SLPI in comparison with the RA patients and the healthy controls (p<0.01), and the RA patients presented significantly higher serum levels of elafin in comparison with the controls (p=0.003). In the SSc subgroup serum SLPI level negatively correlated with diffusing capacity of the lung for carbon monoxide (DLCO) (r=-0.41, p=0.03) and total lung capacity (r=-0.42, p=0.03). Both alarm antiproteases, elafin and SLPI could be potentially implicated in the pathogenesis of SSc and SLPI may be considered a candidate for serum biomarker of lung involvement in SSc.

Asunaprevir: An HCV Protease Inhibitor With Preferential Liver Distribution.

Asunaprevir is an inhibitor of the hepatitis C virus (HCV) NS3/4A protease, demonstrating efficacy in clinical studies in patients infected with HCV genotype 1 or 4, with either peginterferon/ribavirin or combinations of direct-acting antivirals. Because of preferential distribution of asunaprevir to the liver via organic anion-transporting polypeptide (OATP)-mediated transport, asunaprevir demonstrates high apparent oral clearance and very low plasma concentrations. Asunaprevir plasma concentrations are markedly increased by single-dose rifampin (an OATP inhibitor) and in subjects with moderate to severe hepatic impairment. In addition, modestly higher plasma concentrations of asunaprevir have been noted in subjects infected with HCV relative to healthy subjects and in Asian subjects relative to whites. At the marketed dose, infrequent hepatic transaminase abnormalities were poorly predicted by plasma concentrations. For a compound with these characteristics, hepatic concentrations may have provided an improved understanding of the in vivo pharmacokinetic and pharmacodynamic data to support decision making during development.

Isolation and characterization of a novel metalloprotease inhibitor from Bothrops alternatus snake serum.

Resistance of snakes and some other animals to snake envenomation has been attributed to soluble factors present in their tissues. Here we report the isolation of a novel metalloprotease inhibitor from Bothrops alternatus snake serum (named BaltMPI) with high purity, using a four-step chromatographic method. BaltMPI has molecular weights of 60.5 and 42.4kDa, as determined by SDS-PAGE and mass spectrometry, respectively, and pI=5.27. The first 60 amino acids from the N-terminal region of BaltMPI, determined by Edman's degradation, showed high homology (97%) with the snake venom metalloprotease inhibitor (SVMPI) BJ46a and other SVMPIs (78-82%). The chromatographic fractions and purified BaltMPI exhibited anti-hemorrhagic activity against Batroxase and BjussuMP-I. BaltMPI was stable over wide ranges of pH (1, 5, 8, and 9) and temperature (-80, -20, 4, 60, and 100°C), and suppressed the fibrinogenolytic, fibrinolytic, and azocaseinolytic activities of Batroxase. BaltMPI specifically inhibited the activity of metalloproteases, without affecting the activity of serine proteases. Together, our results suggest that BaltMPI and other SVMPIs are promising molecules for the treatment of snake envenomation, in particular that caused by Bothrops sp.

Development and validation of a UPLC-MS/MS method for simultaneous determination of fotagliptin and its two major metabolites in human plasma and urine.

AIM: Fotagliptin is a novel dipeptidyl peptidase IV inhibitor under clinical development for the treatment of Type II diabetes mellitus. The objective of this study was to develop and validate a specific and sensitive ultra-performance liquid chromatography (UPLC)-MS/MS method for simultaneous determination of fotagliptin and its two major metabolites in human plasma and urine. Methodology & results: After being pretreated using an automatized procedure, the plasma and urine samples were separated and detected using a UPLC-ESI-MS/MS method, which was validated following the international guidelines. CONCLUSION: A selective and sensitive UPLC-MS/MS method was first developed and validated for quantifying fotagliptin and its metabolite in human plasma and urine. The method was successfully applied to support the clinical study of fotagliptin in Chinese healthy subjects.

Validation of a panel of ADAMTS13 assays for diagnosis of thrombotic thrombocytopenic purpura: activity, functional inhibitor, and autoantibody test.

INTRODUCTION: Quantitation of ADAMTS13 activity, functional inhibitors, and autoantibodies is crucial in diagnosis and management of thrombotic thrombocytopenic purpura. We compared and optimized commercial assay kits and validated a testing panel. METHODS: Citrated plasma specimens from healthy volunteers and residual samples submitted for clinical testing were used in the study. Commercially available ADAMTS13 activity assays including ACTIFLUOR(™) ADAMTS13 (Sekisui Diagnostics, Stamford, CT, USA), LIFECODES ATS-13 (Gen-Probe Inc., San Diego, CA, USA), and TECHNOZYM(®) ADAMTS-13 (Technoclone, Vienna, Austria) were evaluated. Functional inhibitor assays were performed using internally developed mixing protocols. Two autoantibody assays were also evaluated: IMUBIND(®) (Sekisui Diagnostics) and TECHNOZYM(®) ADAMTS-13 INH ELISA kits (Technoclone). RESULTS: A laboratory-developed assay using ACTIFLUOR(™) reagents showed best agreement with the reference method, and full validation showed a reportable range of 5% (LLOQ) to 114% with a reference interval of ≥68%. Both intra- and interassay coefficients of variation were <10%. Inhibitor assays performed with the kits showed 95% overall agreement with the reference method. A modification of the TECHNOZYM(®) autoantibody assay showed 85% overall agreement with the reference method with imprecision approximately 20%. CONCLUSION: ADAMTS13 activity and inhibitor tests using ACTIFLUOR(™) reagents and modified TECHNOZYM(®) autoantibody ELISA showed superior performance compared to the other kits for clinical use in this study.

Metabolism and Disposition of the Hepatitis C Protease Inhibitor Paritaprevir in Humans.

Paritaprevir (also known as ABT-450), a potent NS3-4A serine protease inhibitor [identified by AbbVie (North Chicago, IL) and Enanta Pharmaceuticals (Watertown, MA)] of the hepatitis C virus (HCV), has been developed in combination with ombitasvir and dasabuvir in a three-direct-acting antiviral agent (DAA) oral regimen for the treatment of patients infected with HCV genotype 1. This article describes the mass balance, metabolism, and disposition of paritaprevir in humans. After the administration of a single 200-mg oral dose of [(14)C]paritaprevir coadministered with 100 mg of ritonavir to four male healthy volunteers, the mean total percentage of the administered radioactive dose recovered was 96.5%, with recovery in individual subjects ranging from 96.0% to 96.9%. Radioactivity derived from [(14)C]paritaprevir was primarily eliminated in feces (87.8% of the dose). Radioactivity recovered in urine accounted for 8.8% of the dose. The biotransformation of paritaprevir in humans involves: 1) P450-mediated oxidation on the olefinic linker, the phenanthridine group, the methylpyrazinyl group, or combinations thereof; and 2) amide hydrolysis at the acyl cyclopropane-sulfonamide moiety and the pyrazine-2-carboxamide moiety. Paritaprevir was the major component in plasma [90.1% of total radioactivity in plasma, AUC from time 0 to 12 hours (AUC0-12hours) pool]. Five minor metabolites were identified in plasma, including the metabolites M2, M29, M3, M13, and M6; none of the metabolites accounted for greater than 10% of the total radioactivity. Paritaprevir was primarily eliminated through the biliary-fecal route followed by microflora-mediated sulfonamide hydrolysis to M29 as a major component in feces (approximately 60% of dose). In summary, the biotransformation and clearance pathways of paritaprevir were characterized, and the structures of metabolites in circulation and excreta were elucidated.

Protective effects of elafin against adult asthma.

BACKGROUND: Elafin inhibits serine proteases, such as human neutrophil elastase and proteinase 3, to prevent excessive damage during inflammation. However, the relationship between elafin and asthma is still unclear. Microarray technology was used to evaluate smoking- and asthma-related biomarkers in a discovery-driven manner. We identified candidate genes, e.g., proteinase inhibitor 3 (PI3), related to asthma and smoking from gene expression microarray data sets and evaluated their potential as biomarkers for asthma. METHODS: We used human genome microarray data sets from smoking- and asthma-related gene expression data sets and performed real-time quantitative polymerase chain reaction to measure and validate differences in gene expression. We also recruited adult patients with asthma and age- and sex-matched control patients who were administered a structured questionnaire and evaluated for lung function and plasma elafin levels, which are encoded by the PI3 gene. RESULTS: Six significantly altered candidate genes, PI3, protein kinase C iota, phosphoserine phosphatase, IQ motif-containing GTPase activating protein 1, interleukin 13 receptor α 1, and signal transducing adaptor molecule SH3 domain and ITAM motif 2, were identified from comparisons across the four asthma- and four smoking-related data sets included in this study. An in vitro study of human airway epithelial cells (A549) and a human monocytic cell line (THP-1) demonstrated that PI3 messenger RNA levels were significantly altered by nicotine exposure. Elafin concentration was significantly higher in control patients than in patients with asthma (p < 0.001). The plasma elafin concentration in the highest quartile (≥12.69 ng/mL) was inversely associated with asthma (adjusted odds ratio 0.122 [95% confidence interval, 0.053-0.278]) compared with the lowest quartile (<5.82 ng/mL) after adjusting for age, sex, smoking status, waist-to-hip ratio, percentage predicted forced expiratory volume in 1 second, cockroaches in the home, incense burning, and family history. CONCLUSION: Our study revealed that high elafin levels identified in smoking- and asthma-related microarray data sets and an epidemiologic study significantly reduced the risk of asthma. Further studies of elafin as a potential therapy for asthma are warranted.

Constant pH Molecular Dynamics Reveals pH-Modulated Binding of Two Small-Molecule BACE1 Inhibitors.

Targeting ß-secretase (BACE1) with small-molecule inhibitors offers a promising route for treatment of Alzheimer's disease. However, the intricate pH dependence of BACE1 function and inhibitor efficacy has posed major challenges for structure-based drug design. Here we investigate two structurally similar BACE1 inhibitors that have dramatically different inhibitory activity using continuous constant pH molecular dynamics (CpHMD). At high pH, both inhibitors are stably bound to BACE1; however, within the enzyme active pH range, only the iminopyrimidinone-based inhibitor remains bound, while the aminothiazine-based inhibitor becomes partially dissociated following the loss of hydrogen bonding with the active site and change of the 10s loop conformation. The drastically lower activity of the second inhibitor is due to the protonation of a catalytic aspartate and the lack of a propyne tail. This work demonstrates that CpHMD can be used for screening pH-dependent binding profiles of small-molecule inhibitors, providing a new tool for structure-based drug design and optimization.

An accessible pharmacodynamic transcriptional biomarker for notch target engagement.

γ-Secretase mediates amyloid production in Alzheimer's disease (AD) and oncogenic activity of Notch. γ-Secretase inhibitors (GSIs) are thus of interest for AD and oncology. A peripheral biomarker of Notch activity would aid determination of the therapeutic window and dosing regimen for GSIs, given toxicities associated with chronic Notch inhibition. This study examined the effects of GSI MK-0752 on blood and hair follicle transcriptomes in healthy volunteers. The effects of a structurally diverse GSI on rhesus blood and hair follicles were also compared. Significant dose-related effects of MK-0752 on transcription were observed in hair follicles, but not blood. The GSI biomarker identified in follicles exhibited 100% accuracy in a clinical test cohort, and was regulated in rhesus by a structurally diverse GSI. This study identified a translatable, accessible pharmacodynamic biomarker of GSI target engagement and provides proof of concept of hair follicle RNA as a translatable biomarker source.

Plasma Concentrations of Protein Z and Protein Z-Dependent Protease Inhibitor in Patients With Essential Thrombocythemia.

The pathological consequences of decreased protein Z (PZ) and/or Z-dependent protease inhibitor (ZPI) levels remain as yet unclear, despite a growing body of evidence which supports their involvement in an increased thrombotic risk. The purpose of the present study was 2-fold: to evaluate plasma concentrations of protein Z and ZPI in patients with essential thrombocythemia (ET) and to determine their significance in thrombotic complications. The median (range) plasma concentrations of PZ in our patients with ET were lower, but not significantly, than in healthy individuals: PZ (1.42 µg/mL, 0.36-3.14 µg/mL vs 1.6 µg/mL, 0.75-2.56 µg/mL, P = .08). On the other hand, the median (range) plasma concentrations of ZPI in the said patients with ET were meaningfully lower than in the reference group: ZPI (3.22 µg/mL, 0.85-6.97 µg/mL vs 4.41 µg/mL, 1.63-7.83 µg/mL, P = .0004). More importantly, the study revealed a statistically significant lower concentration of PZ and ZPI in patients with the presence of the JAK2V617F mutation relative to patients without the mutation, for PZ: 1.38 µg/mL, 0.36-2.6 µg/mL versus 1.63 µg/mL, 0.88-3.14 µg/mL, P = .03, and ZPI 2.89 µg/mL, 0.85-5.91 µg/mL versus 3.61 µg/mL, 1.53-6.97 µg/mL, P = .002. Additionally, significant differences between the concentrations of PZ and ZPI were found in patients with venous thrombotic episodes compared to healthy individuals, for PZ: 1.23 µg/mL, 0.82-1.99 µg/mL versus 1.6 µg/mL, 0.75-2.56 µg/mL, P = .043, and ZPI: 2.42 µg/mL, 0.85-4.21 µg/mL versus 4.41 µg/mL, 1.63-7.83 µg/mL, P < .0001. To recapitulate, our results suggest that the deficiency of PZ may increase tendency to thrombosis in patients with ET.

Immunostimulation by phospholipopeptide biosurfactant from Staphylococcus hominis in Oreochromis mossambicus.

The immunostimulatory effect of phospholipopeptide biosurfactant from Staphylococcus hominis (GenBank Accession No: KJ564272) was assessed with Oreochromis mossambicus. The non-specific (serum lysozyme activity, serum antiprotease activity, serum peroxidase activity and serum bactericidal activity), specific (bacterial agglutination assay) immune responses and disease resistance activity against Aeromonas hydrophila were examined. Fish were intraperitonially injected with water soluble secondary metabolite (biosurfactant) of S. hominis at a dose of 2 mg, 20 mg and 200 mg kg(-1) body weight. Commercial surfactant surfactin (sigma) at 20 mg kg(-1) was used as standard and saline as negative control. All the doses of water soluble biosurfactant tested, significantly enhanced the specific, nonspecific immunity and disease resistance from the day of post administration of phospholipopeptide biosurfactant till the tail of the experimental period. These results clearly indicated that the secondary metabolite isolated from S. hominis stimulates the immunity of finfish thereby could enhance aquaculture production.

Evaluation of the Pharmacokinetics and Renal Excretion of Simeprevir in Subjects with Renal Impairment.

BACKGROUND: Simeprevir is a N3/4 protease inhibitor approved for the treatment of hepatitis C virus (HCV) infection. HCV prevalence is higher in patients with chronic kidney disease compared with the general population; safe and efficacious therapies in renal impairment are needed. OBJECTIVES: To evaluate simeprevir renal excretion in healthy subjects and to compare the simeprevir steady-state pharmacokinetics between subjects with severe renal impairment and healthy subjects. METHODS: In the mass balance study, healthy adults received a single 200-mg dose of (14)C-simeprevir; radioactivity in the urine and feces was quantified until concentrations were <2% of the administered dose and seven or more stools were produced. In the pharmacokinetic study, non-HCV-infected adults with severe renal impairment (estimated glomerular filtration rate ≤29 mL/min/1.73 m(2)) and matched healthy subjects (estimated glomerular filtration rate ≥80 mL/min/1.73 m(2)) received 150 mg simeprevir for 7 days. Pharmacokinetic analysis was performed post-dose on Day 7. RESULTS: (14)C-simeprevir recovery from the urine was low (0.009-0.138% of total dose). The minimum plasma concentration, maximum plasma concentration, and area under the plasma concentration-time curve at 24 h were 71, 34, and 62% higher, respectively, in subjects with severe renal impairment compared with healthy subjects. The mean fraction of simeprevir unbound to protein was <0.0001 (all subjects). Most adverse events were grade I or II; one subject with renal impairment who was receiving fenofibrate presented with grade 3 rhabdomyolysis. CONCLUSIONS: Simeprevir plasma concentrations were mildly elevated in subjects with severe renal impairment. The results suggest that simeprevir may be administered without dose adjustment in patients with renal impairment.