Administration of a CXC Chemokine Receptor 2 (CXCR2) Antagonist,SCH527123, Together with Oseltamivir Suppresses NETosis and Protects Mice from Lethal Influenza and Piglets from Swine-Influenza Infection

Excessive neutrophil influx, their released neutrophil extracellular traps (NETs), and extracellular histones are associated with disease severity in influenza-infected patients. Neutrophil chemokine receptor CXC chemokine receptor 2 (CXCR2) is a critical target for suppressing neutrophilic inflammation. Herein, temporal dynamics of neutrophil activity and NETosis were investigated to determine the optimal timing of treatment with the CXCR2 antagonist, SCH527123 (2-hydroxy-N,N-dimethyl-3-[2-([(R)-1-(5-methyl-furan-2-yl)-propyl]amino)-3,4-dioxo-cyclobut-1-enylamino]-benzamide), and its efficacy together with antiviral agent, oseltamivir, was tested in murine and piglet influenza-pneumonia models. SCH527123 plus oseltamivir markedly improved survival of mice infected with lethal influenza, and diminished lung pathology in swine-influenza–infected piglets. Mechanistically, addition of SCH527123 in the combination treatment attenuated neutrophil influx, NETosis, in both mice and piglets. Furthermore, neutrophils isolated from influenza-infected mice showed greater susceptibility to NETotic death when stimulated with a CXCR2 ligand, IL-8. In addition, CXCR2 stimulation induced nuclear translocation of neutrophil elastase, and enhanced citrullination of histones that triggers chromatin decondensation during NET formation. Studies on temporal dynamics of neutrophils and NETs during influenza thus provide important insights into the optimal timing of CXCR2 antagonist treatment for attenuating neutrophil-mediated lung pathology. These findings reveal that pharmacologic treatment with CXCR2 antagonist together with an antiviral agent could significantly ameliorate morbidity and mortality in virulent and sublethal influenza infections.

Influenza virus infections during pandemic outbreaks and yearly seasonal epidemics cause significant morbidity and mortality rates globally.¹ Seasonal influenza-associated deaths have increased in recent years, with an estimated of >600,000 fatalities annually.² A significant proportion of hospitalized patients with influenza develop complications of acute respiratory distress syndrome, characterized by widespread alveolar-capillary injury, inflammation, edema, and parenchymal hemorrhage.3, 4, 5, 6, 7, 8 These pathologic manifestations are driven by virus-inflicted cytopathic effects as well as exaggerated host immune responses.9, 10, 11 Vaccination is the logical choice for controlling the virus. However, because of unrelenting emergence of new strains and their mutative ability, vaccination presents a major challenge during influenza outbreaks.^12,13 In such cases, treatment primarily depends on antiviral therapy. Administration of antiviral drugs may not always be effective, as considerable lung pathology is mediated by exaggerated host-immune responses in addition to virus-inflicted cytotoxicity.14, 15, 16, 17Previously, we established that massive neutrophil influx, their induced neutrophil extracellular traps (NETs), and extracellular histones (ECHs) aggravate pulmonary pathology in severe influenza.18, 19, 20, 21, 22, 23 Aberrant neutrophil activity and accumulation of NETs are also documented in patients with severe influenza.^24,25 Neutrophils are recruited to the site of injury/infection via chemokine signaling, mediated through chemokine receptors. Among various chemokine receptors, CXC chemokine receptor 2 (CXCR2) plays a critical role in modulating neutrophil functionality during influenza.²⁶ Numerous clinical studies have also tested CXCR2 antagonists for their efficacy in reducing inflammation and organ injuries in acute and chronic diseases.27, 28, 29, 30, 31 Recently, human phase 2 trials evaluated the safety and efficacy of a CXCR2 antagonist, danirixin, alone or in combination with oseltamivir in influenza-infected patients.^27,28 Although administration of danirixin was found to be safe and well-tolerated, no differences in the clinical scores were observed between patients given oseltamivir alone and those given danirixin plus oseltamivir.²⁷ Furthermore, there was inconsistency in neutrophil numbers among different treatment groups. This inconsistency may be attributed to the absence of rational determination of the optimal timing and dosing of danirixin, to achieve the fine balance of suppressing excessive neutrophil influx, without compromising the beneficial host immunity by neutrophils. Moreover, the underlying mechanistic roles of targeting CXCR2 and its pathogenic association with influenza pneumonia have not been established.NETs are large extracellular web-like chromatin strands that were initially proposed to have a defense mechanism against invading pathogens.³² However, excessive release of NETs aggravates tissue injury and death, as reported in several disease conditions.33, 34, 35, 36 NETosis is regulated by various granule and nuclear proteins.³⁷ Myeloperoxidase (MPO) and neutrophil elastase (NEs) are released from azurophilic granules, anchor chromatin scaffolds in NETs, and mediate histone degradation during NETosis.³⁸ We reported earlier that blocking MPO decreases NETs, but signaling mechanisms in influenza-induced NETosis remain unclear.^17,18 Herein, we evaluated the therapeutic efficacy of a CXCR2 antagonist, SCH527123 (2-hydroxy-N,N-dimethyl-3-[2-([(R)-1-(5-methyl-furan-2-yl)-propyl]amino)-3,4-dioxo-cyclobut-1-enylamino]-benzamide) alone or in combination with antiviral agent, oseltamivir (which inhibits viral neuraminidase and prevents progeny virus release from infected cells), in models of lethal influenza-infected mice and sublethal swine influenza-infected piglets. SCH527123 plus oseltamivir significantly improved survival in lethal influenza-challenged mice, and attenuated lung pathology in swine influenza-infected piglets. Thus, SCH5277123 plus oseltamivir represents a promising combination treatment against influenza pneumonia.     

Anatomical complications of epidural anesthesia: A comprehensive review

Epidural anesthesia is a versatile technique widely used in treating lumbar spinal pain syndromes. Complications during these procedures can arise either from needle placement or from administration of medication. Potential risks include infection, hematoma, intravascular or subdural injections of medication, direct nerve trauma, air embolism, entry into a disc space, urinary retention, radiation exposure, and hypersensitivity reactions. The objective of this article is to review the complications of lumbar epidural injections and discuss the potential pitfalls related to these procedures. We searched Medline comprehensively for relevant case reports, clinical trials, and review articles. Complications from lumbar epidural injections are extremely rare. Most if not all of them can be avoided by careful techniques with accurate needle placement, sterile precautions, and a thorough understanding of the relevant anatomy and contrast patterns on fluoroscopic imaging. Clin. Anat. 30:342–346, 2017. © 2017 Wiley Periodicals, Inc.     

Protective effect of MOTILIPERM in varicocele-induced oxidative injury in rat testis by activating phosphorylated inositol requiring kinase 1α (p-IRE1α) and phosphorylated c-Jun N-terminal kinase (p-JNK) pathways

Context: MOTILIPERM was prepared as a mixture of extracts of three medicinal herbs [roots of Morinda officinalis How (Rubiaceae), outer scales of Allium cepa L. (Liliaceae) and seeds of Cuscuta chinensis Lamark (Convolvulaceae)].

Objective: To investigate the role of reactive oxygen species (ROS)-based endoplasmic reticulum (ER) stress in a rat model of varicocele and the therapeutic efficacy of MOTILIPERM in this model.

Materials and methods: Sixty male rats were divided into five experimental groups: a normal control group (CTR?+?vehicle), a control group administered MOTILIPERM 200?mg/kg (CTR?+?M 200), a varicocele-induced control group (VC?+?vehicle) and two varicocele-induced groups administered MOTILIPERM 100 (VC?+?M 100) or 200 (VC?+?M 200) mg/kg for 4 weeks. Testis weights were recorded and serums were assayed for hormone concentrations. Tissues were subjected to semen analysis, histopathology, analyses of ER response protein expression levels and oxidative stress were assessed by measuring ROS, reactive nitrogen species (RNS), malondialdehyde (MDA) level and ratios of total glutathione (GSH)/oxidized GSH (GSSG).

Results: MOTILIPERM treatment of varicocele-induced groups significantly increased left testis weight, testosterone level, sperm motility, count and spermatogenic cell density. ER-response protein expression levels were dose-dependently decreased in VC?+?M 200 group compared with VC?+?vehicle group. MOTILIPERM treatment also decreased MDA and ROS/RNS level but increased GSH/GSSG ratio.

Discussion and conclusions: This study suggests that ROS-related ER stress may play a major role in varicocele-induced infertility and MOTILIPERM, a novel compound targeting ROS-based ER stress, may be therapeutically useful in treatment of varicocele, or as a supplement for the treatment of infertility.     

Global Childhood Unintentional Injury Study: Multisite Surveillance Data

Objectives. We aimed to analyze the epidemiology of childhood unintentional injuries presenting to hospitals in 5 select sites in low- and middle-income countries (LMICs) (Bangladesh, Colombia, Egypt, Malaysia, and Pakistan).Methods. We collected standardized data from children ages 0 to 12 years at participating emergency departments (EDs) in 2007. Statistical analyses were conducted to compare the characteristics of these injuries and to explore the determinants of injury outcomes.Results. Among 2686 injured children, falls (50.4%) and road traffic injuries (16.4%) were the most common, affecting boys more often (64.7%). Home injuries were more common among younger children (average 5.41 vs 7.06 years) and girls (38.2% vs 31.7%). Following an ED visit, 24% of injured children were admitted to the hospital, and 6 died. Injury outcomes were associated with risk factors, such as age and sex, to varying extents.Conclusions. Standardized ED surveillance revealed unintentional injuries are a threat to child health. The majority of events took place inside the home, challenging traditional concepts of children’s safety and underscoring the need for intensified context-appropriate injury prevention.Injuries cause upwards of 5 million deaths each year, of which unintentional injuries account for approximately 80% (3.9 million).1 Unintentional injuries kill approximately 830 000 children every year, and more than 95% of child injury deaths (both intentional and unintentional) occur in low- and middle-income countries (LMICs).2 The 5 most common unintentional injuries reported by the World Health Organization (WHO) are road traffic injuries (RTIs), falls, burns, drowning, and poisoning.2 Global aggregate data reveal that the burden of childhood unintentional injuries is highest in South-East Asia and Africa, with a cause-specific mortality of 64 and 55 children per 100 000 population, respectively.1,2 The disproportionate share of the burden of unintentional childhood injury in LMICs results from a number of factors. First, children are more susceptible to injuries because of their curious nature, which, combined with their low capacity to assess and comprehend risks, and a general lack of safe play spaces in many LMICs, puts them at high risk.3 Second, global economic trends have uprooted communities and traditional forms of subsistence, necessitating persons from rural areas moving to urban slums and at times forcing children into labor, some in hazardous conditions, to contribute to a household’s income. For instance, it has become increasingly dangerous for children to share the road with the ever-growing number of motor vehicles.2,3 In this light, unintentional injuries not only affect children themselves, leaving them temporarily hospitalized or with short- or long-term disabilities, but also adversely affect their families and society as a whole.2,4–6The WHO has called for better data collection on child injury and its determinants for the purposes of national research and investment priority settings, as well as the targeting of high-risk groups.2 Reductions in child injury mortality have been observed in several high-income countries (HICs) as a result of the implementation of evidence-based programs.2,7,8 Likewise, a number of studies have provided reliable information to characterize the pattern of injuries in HICs.9,10 Despite the high burden of unintentional injuries in LMICs, there are few studies that provide standardized data from multiple sites. In the absence of reliable national population data, hospital-based data are an important source of injury information, particularly for children.11,12 In response, the Global Childhood Unintentional Injury Surveillance (GCUIS) study was initiated to collect standardized child injury data from emergency departments (EDs) at 5 sites: Bangladesh, Colombia, Egypt, Malaysia, and Pakistan.4,13 The objectives of the GCUIS study were (1) to determine the epidemiology of 5 major childhood unintentional injuries in 5 EDs in urban LMIC sites; (2) to explore potential risk factors and determinants of injury severity and outcomes, based on the injury records in the GCUIS study; and (3) to briefly summarize the characteristics of injuries sustained in homes, based on GCUIS data.6 In 2009, partial data from GCUIS were analyzed to report the initial pattern of injuries in 4 sites.13 This article adds data from an additional site and further analyses; therefore, it provides more insights into key unintentional injuries.     

Comparison of the effect of asphyxia, hypoxia, and acidosis on intestinal blood flow and O2 uptake in newborn piglets

The aim of our study was to compare the effects of asphyxia and the two components of asphyxia, i.e. hypoxia and acidosis, on intestinal blood flow and oxygen consumption in anesthetized newborn piglets less than 3 days old. The first series of experiments, consisting of four groups of piglets, showed that blood flow to the proximal and distal small intestine and colon (as determined by the microsphere technique) significantly decreased after piglets were subjected to a sustained hypoxic hypoxemia (PaO2 50% of control) or asphyxia (acidosis plus hypoxia) for 90 min. A sustained acidosis (arterial pH = 7.0-7.15 for 90 min), however, decreased blood flow only to the proximal small intestine, and sham operation did not significantly alter any intestinal blood flow. All animals subjected to asphyxia and two of five of the animals subjected to hypoxia alone in this series, produced gross and microscopic intestinal lesions similar to those seen in human newborn with necrotizing enterocolitis. Acidosis alone, however, did not produce any pathologic lesions. The second series of experiments showed that the 90-min hypoxic hypoxemia decreased blood flow to both the mucosa and muscularis layers of the small intestine. The third series of experiments, consisting of four groups of piglets, determined the effects of 60-min acidosis, hypoxic hypoxemia, asphyxia, or sham operation on venous outflow and oxygen consumption of the isolated in situ terminal ileum. Acidosis or sham operation altered neither ileal blood flow nor oxygen consumption. Hypoxia or asphyxia, however, decreased ileal oxygen consumption without significantly decreasing blood flow.(ABSTRACT TRUNCATED AT 250 WORDS)     

Melanin potentiates gentamicin-induced inhibition of collagen biosynthesis in human skin fibroblasts

One of the recognized side effects of gentamicin is ototoxicity. The mechanism underlying the organ specificity of this side effect of gentamicin has not been fully established. In view of the fact that a number of pharmacologic agents are known to form complexes with melanin and melanin is an abundant constituent of the inner ear tissues, we determined whether gentamicin interacts with melanin and how this process affects the biosynthesis of collagen in cultured human skin fibroblasts. Our results indicate that gentamicin forms complexes with melanin. The amount of gentamicin bound to melanin increases with increasing of initial drug concentration. The Scatchard plot analysis of drug binding to melanin showed that at least two classes of independent binding sites are implicated in gentamicin-melanin complex formation: one class with an association constant K(1) approximately 4 x 10(3) M(-1), and the second class with an association constant K(2) approximately 3 x 10(2) M(-1). The number of total binding sites (n(1)+n(2)) was calculated as about 1.36 micromol gentamicin per 1 mg melanin. We have suggested that prolidase, an enzyme involved in collagen metabolism, may be one of the targets for gentamicin-induced inhibition of collagen biosynthesis. We found that gentamicin-induced inhibition of prolidase activity (IC(50) approximately 100 microM) and collagen biosynthesis (IC(50) approximately 100 microM). At this concentration of gentamicin, DNA biosynthesis in human skin fibroblasts was inhibited only by about 30%. Melanin at 100 microg/ml produced about 25% inhibition of DNA synthesis and about 30% inhibition of prolidase activity, but it had no effect on collagen biosynthesis in cultured fibroblasts. However, the addition of melanin (100 microg/ml) to gentamicin-treated cells (100 microM) augmented the inhibitory action of gentamicin on collagen and DNA biosynthesis and partially reversed its inhibitory effect on prolidase activity. A melanin-induced augmentation of the inhibitory effects of gentamicin on collagen and DNA biosynthesis may explain the mechanism for the organ specificity of gentamicin-induced hearing loss in patients administered this drug.