The Role of Neutrophil Extracellular Traps in Post‐Injury Inflammation

Polymorphonuclear (neutrophil) granulocytes (PMNs) are an essential part of the innate immune responses and key instigators and effectors of the underlying pathological mechanisms (endothelial damage, interstitial histolysis, cytokine production, phagocytosis) leading to post-injury inflammation and secondary tissue injury. In 2004, the formation of neutrophil extracellular traps (NETs) was identified as an additional defence mechanism of PMN against microbes. The understanding of complex regulation of neutrophil functions and NET formation is essential for differentiating between healthy and pathological inflammatory response, which frequently determines if patient recovers uneventfully or develops catastrophic complications. Recent discoveries have revealed the potential role of NETs in the pathogenesis of a wide range of non-infectious diseases, including post-injury sterile inflammation. In such conditions, both spontaneous NET formation and impaired NETosis are documented. In this chapter, we review the evidence for the role of NETs in post-injury inflammation, the key molecular and cellular participants in pathological NET formation, the clinical relevance of NETs in post-injury complications and the therapeutic potential of NET inhibition/clearance

Neurobehavioral impairments in ciprofloxacin-treated osteoarthritic adult rats

Background and purpose – Ciprofloxacin (CIP) is a broad-spectrum antibiotic widely used in clinical practice to treat musculoskeletal infections. Fluoroquinoloneinduced neurotoxic adverse events have been reported in a few case reports, all the preclinical studies on its neuropsychiatric side effects involved only healthy animals. This study firstly investigated the behavioral effects of CIP in an osteoarthritis rat model with joint destruction and pain, which can simulate inflammation-associated musculoskeletal pain. Furthermore, effects of CIP on regional brain-derived neurotrophic factor (BDNF) expression were examined given its major contributions to the neuromodulation and plasticity underlying behavior and cognition. Methods – Fourteen days after induction of chronic osteoarthritis, animals were administered vehicle, 33 mg/kg or 100 mg/kg CIP for five days intraperitoneally. Motor activity, behavioral motivation, and psychomotor learning were examined in a reward-based behavioral test (Ambitus) on Day 4 and sensorimotor gating by the prepulse inhibition test on Day 5. Thereafter, the prolonged BDNF mRNA and protein expression levels were measured in the hippocampus and the prefrontal cortex. Results – CIP dose-dependently reduced both locomotion and reward-motivated exploratory activity, accompanied with impaired learning ability. In contrast, there were no significant differences in startle reflex and sensory gating among treatment groups; however, CIP treatment reduced motor activity of the animals in this test, too. These alterations were associated with reduced BDNF mRNA and protein expression levels in the hippocampus but not the prefrontal cortex. Conclusion – This study revealed the detrimental effects of CIP treatment on locomotor activity and motivation/learning ability during osteoarthritic condition, which might be due to, at least partially, deficient hippocampal BDNF expression and ensuing impairments in neural and synaptic plasticity

Compared efficacy of preservation solutions on the outcome of liver transplantation: Meta-analysis

AIM: To compare the effects of the four most commonly used preservation solutions on the outcome of liver transplantations. METHODS: A systematic literature search was performed using MEDLINE, Scopus, EMBASE and the Cochrane Library databases up to January 31(st), 2017. The inclusion criteria were comparative, randomized controlled trials (RCTs) for deceased donor liver (DDL) allografts with adult and pediatric donors using the gold standard University of Wisconsin (UW) solution or histidine-tryptophan-ketoglutarate (HTK), Celsior (CS) and Institut Georges Lopez (IGL-1) solutions. Fifteen RCTs (1830 livers) were included; the primary outcomes were primary non-function (PNF) and one-year post-transplant graft survival (OGS-1). RESULTS: All trials were homogenous with respect to donor and recipient characteristics. There was no statistical difference in the incidence of PNF with the use of UW, HTK, CS and IGL-1 (RR = 0.02, 95%CI: 0.01-0.03, P = 0.356). Comparing OGS-1 also failed to reveal any difference between UW, HTK, CS and IGL-1 (RR = 0.80, 95%CI: 0.80-0.80, P = 0.369). Two trials demonstrated higher PNF levels for UW in comparison with the HTK group, and individual studies described higher rates of biliary complications where HTK and CS were used compared to the UW and IGL-1 solutions. However, the meta-analysis of the data did not prove a statistically significant difference: the UW, CS, HTK and IGL-1 solutions were associated with nearly equivalent outcomes. CONCLUSION: Alternative solutions for UW yield the same degree of safety and effectiveness for the preservation of DDLs, but further well-designed clinical trials are warranted

L-Alpha-glycerylphosphorylcholine can be cytoprotective or cytotoxic in neonatal rat cardiac myocytes

L-Alpha-glycerylphosphorylcholine (GPC) is a widely used food supplement. GPC has been shown to exert beneficial effects in several organs; however, the cardiac effects of GPC have yet to be investigated. The aim of the present study was therefore to map out the effects of GPC on cardiac myocytes, with or without ischemia-reperfusion insult. Neonatal rat cardiac myocytes were treated with GPC at 1, 10, 80, and 100 µM concentrations for 15 min, 3 h, or 24 h, respectively. Cell viability by calcein assay and the degree of oxidative stress by DHE (superoxide level) and H2DCF (total ROS accumulation) staining were measured. In separate experiments, cardiomyocytes were pre-treated with the optimal concentration of GPC for 3 h and then cells were exposed to 4 h of simulated ischemia followed by 2 h of reperfusion (SI/R). Cell viability was measured at the end of the SI/R protocol. In normoxic conditions, the 15-min and the 3-h GPC treatment did not affect cell viability, total ROS, and superoxide levels. Under SI/R conditions, the 3-h GPC treatment protected the cardiac myocytes from SI/R-induced cell death and did not alter the level of oxidative stress. The 24-h GPC treatment in normoxic conditions resulted in significant cell death and increased oxidative stress at each concentration. Here we provide the first evidence for the cytoprotective effect of short-term GPC treatment. However, long-term administration of GPC may exert cytotoxicity in a wide concentration range in cardiac myocytes. These results may draw attention to a comprehensive cardiac safety protocol for the testing of GPC

New therapeutic and diagnostic approaches with which to influence mitochondrial dysfunctions

Methane (CH4) was earlier thought to be produced in the gastrointestinal (GI) tract by methanogenic bacterial fermentation, under strictly anaerobic conditions. Humans can be CH4 producers or CH4 nonproducers, depending on the presence of methanogenic strains, age, race and lifestyle. The producer status, or a higher CH4 level in the exhaled breath, is considered to be associated with various GI disorders, such as chronic constipation or lactose malabsorption. Nevertheless, the exclusivity of bacterial CH4 formation was challenged recently when in vitro and in vivo studies revealed the possibility of non-microbial CH4 formation in mitochondria and eukaryote cells, in both plants and animals. The classical CH4 detection method is based on gas chromatography, which has many limitations, and real-time measurement is not possible. We set out to develop a photoacoustic spectroscopy (PAS)-based on-line method with appropriate specificity and sensitivity to describe the process of CH4 emission in rodents and also in human volunteers. Our next objective was to investigate nonbacterial biotic methanogenesis and to shed light on the mechanistic details of the reaction. The initial in vitro studies led to the proposal that a continuous lack of the electron acceptor oxygen will maintain an elevated mitochondrial NADH/NAD+ ratio, causing reductive stress. Electrophilic methyl groups bound to positively-charged nitrogen moieties, such as in phosphatidylcholine molecules, may potentially act as substitute electron acceptors, and in consequence CH4 may be liberated. Thus, priming during hypoxia occurs as a progressive process involving depressed electron transport, the loss of cytochrome c and antioxidants, and the triggering of methane release during the abnormal formation of reactive oxygen species induced by reoxygenation or reperfusion. We therefore hypothesized that the formation and emission of CH4 in mammals may be connected with hypoxic events leading to, or associated with a mitochondrial dysfunction. A further aim was to influence the consequences of mitochondrial dysfunction by the administration of a water-soluble, deacylated derivative of phosphatidylcholine, L-alpha-glycerylphosphorylcholine (GPC). Here, we took into account the earlier in vivo findings that GPC is a centrally-acting cholinergic precursor which increases the tolerance to ischemic tissue damage and is clinically effective in various neurodegenerative diseases. We assumed that GPC treatment could moderate the CH4 generation and also the inflammatory consequences of experimental oxidoreductive stress. In three parallel studies, we determined data on the applicability of a novel PAS-based instrument. As compared with gas chromatography, this allows real-time and dynamic measurements, while the gas-sampling procedure does not demand the use of disposable bags or syringes and operates without chemicals. The use of the instrument is noninvasive, allows the detection of the whole-body gas emission of rodents and is appropriate for human exhaled breath analysis too. Moreover, it is relatively cost-effective due to the application of near-infrared diode lasers. After the development of an appropriately specific and sensitive detection system for CH4, we investigated the functional role of mitochondrial electron transport in biogenesis. We demonstrated that the extent of CH4 generation is significantly increased in rodents exposed to chronic sodium azide challenge or endotoxemia. The phenomenon proved to be independent of the methanogenic flora, since the CH4 emission was also elevated in antibiotic-treated groups. As endotoxemia and specific inhibition of mitochondrial complex IV with sodium azide led to an increased CH4 output, we assumed that mitochondrial distress and the subsequent inflammatory reaction might be common denominators of CH4 biogenesis. The results pointed to a possible role of CH4 as an alarm signal for the development of mitochondrial responses under hypoxic conditions, and accordingly it may be a biomarker of such events. In our studies, the stress-induced CH4 generation, the hepatic microcirculatory reduction and the increased pro-oxidant and inflammatory responses were markedly attenuated by GPC treatment. When the consequence of mesenteric ischemia/reperfusion, or gamma-irradiation of the hippocampus were investigated, it emerged that GPC maintained the decreased ATP content of the liver. In conclusion, the results suggest that PAS-based spectroscopy is an excellent approach for CH4 detection, not only in animal experiments, but also in human investigations. CH4 production in mammals is connected with hypoxic events and is associated with a mitochondrial dysfunction. GPC is protective against the inflammatory consequences of a hypoxic reaction that might involve cellular or mitochondrial CH4 generation