Causal connection between detoxification enzyme activity and consumption of a toxic plant compound

Insect herbivores can increase their detoxification activities against a particular plant poison in response to prolonged ingestion of the same compound. For example, larval tobacco hornworms (Manduca sexta) experience a dramatic increase in cytochrome P450 activity against nicotine after ingesting nicotine. While it is generally assumed that this induction process permits increased consumption of toxic plant tissues, we are not aware of any direct experimental support for this assumption. Using a two-tiered approach, we examined the functional significance of P450 induction to M. sexta larvae ingesting a toxic but non-deterrent concentration of nicotine. First, we related the time-course of P450 induction in midgut microsomes to changes in nicotine consumption. When offered a nicotine diet, larvae failed to show a significant increase in consumption before 36 h, which was coincident with the time-course of the induction of midgut P450 activities against aldrin and nicotine. Second, we determined whether inhibiting the induced P450 activities affected nicotine consumption. We found that the increase in nicotine consumption following the induction of nicotine metabolism could be strongly inhibited by treatment with piperonyl butoxide, which by itself did not inhibit consumption. These results provide direct evidence for a causal connection between P450-mediated detoxification activity and consumption of a toxic plant compound.Abbreviation PB piperonyl butoxide     

Reductive detoxification of acrolein as a potential role for aldehyde reductase (AKR1A) in mammals

Aldehyde reductase (AKR1A), a member of the aldo–keto reductase superfamily, suppresses diabetic complications via a reduction in metabolic intermediates; it also plays a role in ascorbic acid biosynthesis in mice. Because primates cannot synthesize ascorbic acid, a principle role of AKR1A appears to be the reductive detoxification of aldehydes. In this study, we isolated and immortalized mouse embryonic fibroblasts (MEFs) from wild-type (WT) and human Akr1a-transgenic (Tg) mice and used them to investigate the potential roles of AKR1A under culture conditions. Tg MEFs showed higher methylglyoxal- and acrolein-reducing activities than WT MEFs and also were more resistant to cytotoxicity. Enzymatic analyses of purified rat AKR1A showed that the efficiency of the acrolein reduction was about 20% that of glyceraldehyde. Ascorbic acid levels were quite low in the MEFs, and while the administration of ascorbic acid to the cells increased the intracellular levels of ascorbic acid, it had no affect on the resistance to acrolein. Endoplasmic reticulum stress and protein carbonylation induced by acrolein treatment were less evident in Tg MEFs than in WT MEFs. These data collectively indicate that one of the principle roles of AKR1A in primates is the reductive detoxification of aldehydes, notably acrolein, and protection from its detrimental effects.     

Diet-induced food preference by Manduca sexta larvae: acceptable non-host plants elicit a stronger induction than host plants

Diet-induced changes in food preference by fifth instar larvae of the tobacco hornworm, Manduca sexta (Johan.) (Lepidoptera, Sphingidae), were examined. Two groups of larvae with different diet experience were used: larvae reared on a host or on an acceptable non-host plant species. Each group of larvae was offered a choice between leaf discs from each rearing plant species (2-plant choice test) and food preference was measured as the consumption of one plant species relative to that of the other plant species. Diet-induced changes in preferences were larger with the host versus acceptable non-host plant pairs Solanum pseudocapsicum (L.) versus Raphanus sativus (L.), Lycopersicon esculentum (Mill.) versus Vigna sinensis (Savi), and Datura innoxia (L.) versus V. sinensis than with the host versus host plant pairs L. esculentum versus Capsicum annuum (L.) and L. esculentum versus D. innoxia. To examine how much the food preference had been altered for each test plant species alone, two other groups of larvae were offered a choice between leaf discs from a single plant species and filter paper discs laced with distilled water (1-plant choice test). Larvae preferred the dietary plant species more strongly than the non-dietary plant species in tests using the following plant species: for C. annuum with C. annuum and L. esculentum as diets, for V. sinensis with V. sinensis and L. esculentum or D. innoxia as diets, and for R. sativus with R. sativus and S. pseudocapsicum as diets. The preference for the hosts L. esculentum and D. innoxia did not change significantly after rearing larvae on different hosts or on an acceptable non-host. Thus, diet-induction by M. sexta larvae results in an enhancement of preference for the dietary plant species which is much stronger with acceptable non-hosts than with hosts.     

Environmental tobacco smoke in the workplace: markers of exposure, polymorphic enzymes and implications for disease state

Studies focusing directly on tobacco smoke have tended to center on the differences in effect between smokers and non-smokers and many hundreds of such studies have been performed. In this review, we examine the current literature specifically concerning workplace exposure to environmental tobacco smoke (ETS) and its impact on individuals, particularly non-smokers and never smokers. The paper deals with quantifying and minimizing ETS exposures in a working environment, the effect of polymorphisms and other genetic factors that influence health outcomes after exposure to ETS and the association of occupational ETS exposure to disease-specific biomarkers.     

Specializations and limitations in the utilization of food resources by the carp,Cyprinus carpio: a study of oral food processing

Synopsis The wide variety of aquatic food is considered to be instrumental for the diversification in fish species. Yet their abilities and inabilities of handling food are poorly known. For these reasons the food processing and feeding repertoire of the adult carp, Cyprinus carpio, fed on a variety of food types, were analyzed by light and X-ray cinematography of the head parts and by electromyography of the head and body muscles during feeding. Nine stereotyped movement patterns (particulate intake, gulping, rinsing, spitting, selective retention of food, transport, crushing, grinding and deglutition) compose the feeding process, their sequence and frequency were adjusted to the type of food. Following quantitative morphological analysis at macroscopic, light- and electronmicroscopical level, the relations between the functioning and architecture of the feeding apparatus were established. The structure and dimensions of the mouth opening, the protrusible upper jaw, the slit-shaped pharyngeal cavity, the palatal and postlingual organ, the branchial sieve, the pharyngeal masticatory apparatus and the distribution of taste buds, mucous cells and muscle fibers along the oropharyngeal surface were the directive structural characters used for estimating the abilities in food processing. The specializations for utilizing food items and its limitations, derived from structural and functional data, are compared with diet data found in the literature in order to evaluate the relative position of the carp in competition for food in the aquatic environment. It is established that the ‘omnivorous’ carp is specialized in effective handling of several categories of aquatic food, even when these are mixed with non-food (bottom invertebrates <4% SL in diameter) since the palatal organ enables the carp to separate food from non-food. This includes very hard-skinned food items, processed with the powerful pharyngeal jaws of the fish, and to a lesser extent zooplankton (>250 μm). The carp is at the same time very limited in processing long and struggling prey (e.g. fish) as well as vegetable matter, due to the lack of oral teeth and the specialized morphology of its pharyngeal chewing apparatus. These feeding abilities agree with diet data from literature. The reported herbivorism of carp illustrates its opportunism in feeding behaviour. Specialization in feeding is discussed and the necessity to take into account the total series of post-capture feeding actions for a more complete view on trophic specialization. Food intake and the intra-oral food processing of carp are bound to the structures of its sensory, central processing and effector apparatus and to the plasticity in their functioning. These together determine its feeding efficiency in exploiting the available aquatic food resources. Next to ethological and ecological studies functional morphology is another important tool to explain the trophic interactions of fish.     

西花蓟马或二斑叶螨为害的菜豆对两者间后取食者体内保护酶和解毒酶活性的影响

【目的】害虫取食后会导致植物的防御反应;取食同种植物的不同种害虫其生理适应性可能不同。本研究旨在阐明一种害虫取食后的植物对后取食的另一种害虫虫体酶活性的影响。【方法】采用生化分析法研究了西花蓟马Frankliniella occidentalis 2龄若虫或二斑叶螨Tetranychus urticae雌成螨为害后的菜豆对后取食不同时间下的二斑叶螨第2若螨和雌成螨及西花蓟马2龄若虫和雌成虫体内保护酶[过氧化物酶(POD)、过氧化氢酶(CAT)和超氧化物歧化酶(SOD)]和解毒酶[多功能氧化酶(MFO)、羧酸酯酶(Car E)、谷胱甘肽-S-转移酶(GSTs)和乙酰胆碱酯酶(Ach E)]活性的影响。【结果】二斑叶螨雌成螨取食西花蓟马为害菜豆5 h时,POD,CAT和SOD这3种保护酶的活性均明显高于取食健康植株(P0.01),取食蓟马为害的菜豆18 h时只有POD活性低于取食健康植株(P0.01);二斑叶螨第2若螨取食蓟马为害菜豆后只有CAT活性在5 h时明显高于取食健康植株(P0.01)。二斑叶螨雌成螨取食蓟马为害菜豆5 h时,解毒酶MFO和Car E的活性受到抑制(P0.01),但GSTs和Ach E活性均比取食健康菜豆高(P0.01);取食18 h时,只有MFO的活性变化与5 h不同。第2若螨取食蓟马为害菜豆只有MFO的活性不论在5 h还是18 h时明显高于取食健康菜豆(P0.01)。西花蓟马取食二斑叶螨雌成螨为害菜豆时,雌成虫体内的3种保护酶的活性不论取食5 h还是18 h均明显升高(P0.01),但2龄若虫只有CAT活性在这两个时间段下明显升高(P0.01)。西花蓟马雌成虫取食螨害菜豆5 h时,解毒酶活性的变化同二斑叶螨雌成螨,取食18 h时只有GSTs活性变化状态与取食5 h不同;西花蓟马2龄若虫取食螨害菜豆时除GSTs外解毒酶的活性变化在这两个时间下正好相反。【结论】西花蓟马或二斑叶螨为害的菜豆能诱导后取食的二斑叶螨和西花蓟马产生一系列应激生化反应,且两种害虫成虫体内所有的保护酶和解毒酶活性在5 h时变化状态相同,但在其余时间和虫态下酶活性的变化状态不同。说明这两种昆虫体内保护酶和解毒酶活性的变化既有共性,又与害虫种类、虫(螨)态、取食时间相关,两种害虫对虫害植物的生理适应性不完全相同。     

Lisosan G, a powder of grain, does not interfer with the drug metabolizing enzymes and has a protective role on carbon tetrachloride-induced hepatotoxicity

Lisosan G is a powder of grain registered as an alimentary integrator. The treatment of rats for 4 days with 0.5 g Lisosan G/kg had no effect on various drug metabolizing enzymes. Experiments in vitro showed that Lisosan G had radical scavenger activity. A confirmation of the antioxidative property of Lisosan G was also confirmed when it was administered in vivo to carbon tetrachloride (CCl₄)-intoxicated rats. The toxicity caused by CCl₄-treatment of rats was restored to the control levels when the rats were given Lisosan G for 4 days before CCl₄. Lisosan G thus does not interfer with drug metabolizing system but has antioxidant properties and protects against CCl₄-induced hepatotoxicity.     

Pulicaria crispa mitigates gastric ulcer induced by ethanol in rats: role of treatment and auto healing

Context: Stomach ulcers are the common gastrointestinal disorders worldwide.

Objective: This study aimed to investigate the therapeutic impact of Pulicaria crispa aerial parts ethanol extract against gastric ulcer in rats.

Materials and methods: Ulcer was induced by one oral dose of ethanol (0.5?ml/100g body weight) on 24?hours empty stomach, then the plant extract (500?mg/kg b.wt.) was orally administered daily for one week. Ranitidine (100?mg/kg b.wt.); as a reference drug was evaluated. Stomach acidity and volume, as well as lesion counts were measured. Levels of malondialdehyde (MDA), glutathione (GSH) and superoxide dismutase (SOD) were estimated. Assay of different marker enzymes; succinate dehydrogenase (SDH), lactate dehydrogenase (LDH), glucose-6-phosphatase (G-6-Pase), acid phosphatase (AP) and 5′-nucleotidase (5′NT) were determined. Interlukin-10 (IL-10), intracellular adhesion molecule-1 (ICAM-1) and tumor necrosis factor alpha (TNF-α) were also determined. Stomach histopathological assessment was detected.

Results: Gastric ulcer showed drastic changes in oxidative stress, cell organelles and inflammatory markers. These biomarkers served as good tools to identify the presence of gastric ulcer. Treatment with P. crispa recorded amelioration in most parameters exceeding the auto healing effect.

Conclusion: Healing potency of P. crispa is possibly related to its content of glycosides, coumarins, flavonoids, tannins, sterols and triterpenes.     

Oxidative stress induced by intermittent exposure at a simulated altitude of 4000 m decreases mitochondrial superoxide dismutase content in soleus muscle of rats

The effects were examined of 6-month intermittent hypobaric (4000 m) exposure on the antioxidant enzyme systems in soleus and tibialis muscles of rats. At the end of the 6-month experimental exposure, the six rats in both the exposed group and the control group were sacrificed. Immunoreactive mitochondrial superoxide dismutase (Mn-SOD) contents were measured as well as the activities of antioxidant enzymes [Mn-SOD, cytosolic SOD (Cu,Zn-SOD), catalase (CAT), and glutathione peroxidase (GPX)]. Thiobarbituric acid-reactive substances (TBARS) were also determined as an indicator of lipid peroxidation. The high altitude exposure resulted in a marked increase in TBARS content in soleus muscle, suggesting increased levels of oxygen free radicals. Conversely, significant decreases in both Mn-SOD content and activity in solens muscle were oted affer exposure. Such trends were not noticed in tibialis muscle. On the other hand, no significant changes in Cu,Zn-SOD, CAT, or GPX were observed in either muscle. These results suggested that the increases in lipid peroxidation were most probably a result of decreased Mn-SOD function which was more depressed in oxidative than in glycolytic muscle.     

Solubilization of low-rank coal by Trichoderma atroviride: Evidence for the involvement of hydrolytic and oxidative enzymes by using 14C-labelled lignite

The deuteromycete Trichoderma atroviride is able to solubilize lignite in dependence on a given carbon source for growth. When cultivated on media containing glutamate, this mold excreted a set of different enzymes with hydrolytic activity. Addition of lignite to the growth media induced the synthesis of extracellular lignite-specific esterase activity but no evidence has been provided for its direct involvement in the process of lignite solubilization. Hence, the basic capability of T. atroviride enzymes to degrade a variety of ester and ether bonds at the surface or within the bulky lignite structure was tested using coal following its direct labelling with ¹⁴C-alkyl iodide. The participation of hydrolytic and oxidative enzymes in lignite degradation was assessed by measuring the release of ¹⁴C radioactivity from selectively alkylated carboxylic and phenolic OH groups. T. atroviride cleaved both carboxylic esters using esterases and the phenolic ether bonds by using oxidative enzymes, most likely laccases. Journal of Industrial Microbiology & Biotechnology (2002) 28, 207–212 DOI: 10.1038/sj/jim/7000232 Received 05 July 2001/ Accepted in revised form 08 December 2001     

The intensity of interference varies with food density: support for behaviour-based models of interference

Dolman (1995) measured the intake rates of snow buntings feeding on seed patches at difference bird densities, for each of two different seed densities. Interference occurred in the low food density treatment, with intake rate declining at high bird densities, but did not occur in the high food density treatment. Dolman states that existing models of the interference process assume that the intensity of interference is independent of food density. While this statement is true for the model of Hassell and Varley (1969), we show that behaviour-based models of interference do not assume that interference is independent of food density. We examine two simple analytic behaviour-based models of interference and show that, in agreement with Dolman's observations, intensity of interference is predicted to decrease with increasing food density. Dolman plotted log (intake rate) against log (bird density). Rather than obtaining a linear relationship predicted by the Hassell and Varley model, the results from the low seed density treatment indicated a curvilinear relationship consistent with those produced by our behaviour-based models. These results provide support for the use of behaviour-based models of interference over the model of Hassell and Varley.     

Synergistic interactions of Ferulic acid with Ascorbic acid: Its cardioprotective role during isoproterenol induced myocardial infarction in rats

Studies on the lipid peroxidation and antioxidant changes and their significance during myocardial injury have provided a new insight into the pathogenesis of heart disease. The heart failure subsequent to myocardial infarction may be associated with an antioxidant deficit as well as increased myocardial oxidative stress. The present study was designed to evaluate the effect of the combination of ferulic acid and ascorbic acid on antioxidant defense system and lipid peroxidation against isoproterenol (ISO)-induced myocardial infarction in rats. Induction of rats with isoproterenol (150 mg/kg body weight daily, i.p.) for 2 days resulted in a marked elevation in lipid peroxidation, serum marker enzymes (LDH, CPK, GOT, and GPT), and a significant decrease in activities of endogenous antioxidants (SOD, GPx, GST, CAT, and GSH). Pre-co-treatment with the combination of ferulic acid (20 mg/kg body weight/day) and ascorbic acid (80 mg/kg body weight/day) orally for 6 days, significantly attenuated these changes when compared to the individual treatment groups. Histopathological observations were also in correlation with the biochemical parameters. Thus, ferulic acid and ascorbic acid significantly counteracted the pronounced oxidative stress effect of ISO by the inhibition of lipid peroxidation, restoration of antioxidant status, and myocardial marker enzymes levels. In conclusion, these findings indicate the synergistic protective effect of ferulic acid and ascorbic acid on lipid peroxidation and antioxidant defense system during ISO-induced myocardial infarction and associated oxidative stress in rats.     

Declining acceptability of a food plant for the polyphagous grasshopper Schistocerca americana: the role of food aversion learning

ABSTRACT Successive contacts with a food that was initially acceptable to the acridid Schistocerca americana (Drury) (Orthoptera: Acrididae) were monitored. It was found that acceptability, as measured by meal length, declined with experience until the food, spinach, was completely rejected. Reduced acceptability of spinach was maintained even after feeding for three meals on a fully acceptable and suitable food, a period of 2–3 h. Experiments with spinach-filled capsules placed in the gut suggested that the decline in acceptability was not due to direct feedback from the gut or haemolymph on the sensory or feeding control systems. Food aversion learning is indicated as the mechanism underlying the decline in acceptability.     

Formation of albumin dimers induced by exposure to peroxides in human plasma: a possible biomarker for oxidative stress

Human serum albumin (HSA) has one free thiol residue at Cys-34 that is likely oxidized by various reactive oxygen species (ROS). We attempted to identify the oxidation product of Cys-34 of HSA following exposure of plasma to ROS. Oxidation induced by tert-butyl hydroperoxide (t-BuOOH) of this free cysteine residue in HSA was observed in detail. Analysis of oxidized albumin in a partially purified fraction obtained by affinity column chromatography clearly revealed the formation of albumin disulfide dimers following t-BuOOH exposure. Albumin disulfide dimer formation was observed in normal plasma following treatment with various peroxides, as well as in untreated plasma from patients on hemodialysis using SDS-PAGE and Western blot analysis. The present results indicate that albumin dimers are oxidative products derived from peroxides, and that their presence in plasma might be a marker of oxidative stress as secondary metabolites of peroxidation.     

Endoreduplication of human smooth muscle cells induced by 2-methoxyestradiol: a role for cyclin-dependent kinase 2

Endoreduplication has been suggested to contribute to the development of hypertrophy of smooth muscle cells (SMCs) in hypertension. However, endoreduplication in vascular SMCs and the underlying molecular mechanisms are not clear. Treatment of human SMCs with 10 microM 2-methoxyestradiol (2-ME) for 24 h induces accumulation of cells with > or =4N DNA content, and some polyploid/aneuploid cells actively synthesize their DNA, suggesting the occurrence of endoreduplication. In addition, 2-ME treatment upregulates the expression of cyclin-dependent kinase 2 (Cdk2). The present study was designed to characterize endoreduplication of human SMCs and explore the potential roles of Cdk2 in endoreduplication induced by 2-ME. Treatment with 2-ME (10 microM) for 2-4 days not only caused increases in >4N cells and their reentry into S phase but also induced overduplication of chromosomes. Furthermore, 2-ME increased the kinase activity of Cdk2 and its interaction with cyclin E. Inducible overexpression of dominant-negative Cdk2 in human SMCs inhibited both DNA synthesis of >4N cells and the accumulation of >4N cells induced by 2-ME. We conclude that 2-ME induces endoreduplication of human SMCs and Cdk2 plays an important role in endoreduplication in response to 2-ME.     

Influence of selenium on glutathione and some associated enzymes in rats with mammary tumor induced by 7,12-Dimethylbenz(a)anthracene

A recent finding in epidemiological and laboratory studies suggests that the ratio of selenium to glutathione is lower in breast cancer subjects than its control counterparts. Selenium, an antioxidant and anticarcinogen, can modify the status of glutathione and some associated enzymes by blocking peroxidation of lipids in membranes of cancer subjects. Studies were conducted using female albino rats of Wistar strain bearing mammary tumor induced by 7,12-dimethylbenz(a) anthracene to assess the biological role of selenium on some antioxidant enzymes associated with the maintenance of glutathione status. For induction of mammary tumor, 25 mg DMBA in a 1 ml emulsion of sunflower oil and physiological saline was injected subcutaneously to each rat. One group in each of control and tumor bearing rats, were fed 5 mg sodium selenite/kg diet from the day of tumor induction for 24 weeks. Increase in the reduced glutathione concentration was preceded by significant increase in the oxidized glutathione as well as in the activities of -glutamylcysteine synthetase, glutathione peroxidase, glutathione reductase, glutathione S-transferase, and glucose-6-phosphate dehydrogenase by selenium administration in rats bearing tumor. However, selenium administration to rats bearing tumor decreased the activity of -glutamyl transpeptidase. These observations clearly demonstrate the influence of dietary selenium supplementation in correcting abnormal changes in glutathione turnover and some associated enzymes in tumor induced rats.     

Hyperglycemia after protein ingestion concurrent with injection of a GLP-1 receptor agonist in rats: a possible role for dietary peptides

Protein ingestion after injection of the glucagon-like peptide-1 receptor agonist Exendin-4 (Ex-4) causes hyperglycemia in rats. The objectives of this study were to determine the components of protein digestion responsible for this effect and to associate it with changes in the concentrations of other metabolites and hormones. Two experiments were conducted. In the first experiment, food-deprived rats were gavaged with intact whey (WP) or albumin protein, their hydrolysates, amino acid mixtures (1 g/2.5 ml), or water 5 min after injection of either PBS or Ex-4 (0.5 microg/rat). Tail vein blood was analyzed for glucose over 2 h. In the second experiment, food-deprived rats were gavaged with WP with or without Ex-4. Groups of conscious rats were killed by decapitation either before, or at selected times after gavage. Plasma concentrations of glucose, amino acids, free fatty acids (FFA), glycerol, insulin, glucagon, and leptin were measured. In experiment 1, blood glucose was higher when intact proteins and protein hydrolysates, but not amino acid mixtures, were given with than without Ex-4 (P < 0.05). In experiment 2, concentrations of glucose, FFA, and the ratio of tyrosine to branched-chain amino acid were higher (P < 0.01), but leptin and essential amino acid concentrations were lower (P < 0.05), and insulin, glucagon, and glycerol were similar when WP was given with or without Ex-4. We conclude that the hyperglycemia caused by the administration of Ex-4 concurrently with dietary protein arises from the action of peptides released during digestion and their interaction with Ex-4 in the regulation of glucose, fatty acid, and amino acid metabolism.     

Exploring the role of large predators in marsh food webs: evidence for a behaviorally-mediated trophic cascade

The influence of large predators on lower trophic levels in oligotrophic, structurally complex, and frequently disturbed aquatic environments is generally thought to be limited. We looked for effects of large predators in two semi-permanent, spikerush-dominated marshes by excluding large fish (>12 mm body depth) and similarly sized herpetofauna from 1 m² cages (exclosures) for 2 weeks. The exclosures allowed for colonization by intermediate (in size and trophic position) consumers, such as small fish, shrimp, and crayfish. Exclosures were compared to control cages that allowed large fish to move freely in and out. At the end of the experiment, intermediate-consumer densities were higher in exclosures than in controls at both sites. Decapod crustaceans, especially the riverine grass shrimp (Palaemonetes paludosus), accounted for the majority of the response. Effects of large fish on shrimp were generally consistent across sites, but per capita effects were sensitive to estimates of predator density. Densities of intermediate consumers in our exclosures were similar to marsh densities, while the open controls had lower densities. This suggests that these animals avoided our experimental controls because they were risky relative to the surrounding environment, while the exclosures were neither avoided nor preferred. Although illuminating about the dynamics of open-cage experiments, this finding does not influence the main results of the study. Small primary consumers (mostly small snails, amphipods, and midges) living on floating periphyton mats and in flocculent detritus (“floc”) were less abundant in the exclosures, indicative of a trophic cascade. Periphyton mat characteristics (i.e., biomass, chlorophyll a, TP) were not clearly or consistently affected by the exclosure, but TP in the floc was lower in exclosures. The collective cascading effects of large predators were consistent at both sites despite differences in drought frequency, stem density, and productivity.     

Lipoapoptosis induced by saturated free fatty acids stimulates monocyte migration: a novel role for Pannexin1 in liver cells

Recruitment of monocytes in the liver is a key pathogenic feature of hepatic inflammation in nonalcoholic steatohepatitis (NASH), but the mechanisms involved are poorly understood. Here, we studied migration of human monocytes in response to supernatants obtained from liver cells after inducing lipoapoptosis with saturated free fatty acids (FFA). Lipoapoptotic supernatants stimulated monocyte migration with the magnitude similar to a monocyte chemoattractant protein, CCL2 (MCP-1). Inhibition of c-Jun NH₂-terminal kinase (JNK) in liver cells with SP600125 blocked migration of monocytes in a dose-dependent manner, indicating that JNK stimulates release of chemoattractants in lipoapoptosis. Notably, treatment of supernatants with Apyrase to remove ATP potently inhibited migration of THP-1 monocytes and partially blocked migration of primary human monocytes. Inhibition of the CCL2 receptor (CCR2) on THP-1 monocytes with RS102895, a specific CCR2 inhibitor, did not block migration induced by lipoapoptotic supernatants. Consistent with these findings, lipoapoptosis stimulated pathophysiological extracellular ATP (eATP) release that increased supernatant eATP concentration from 5 to ~60 nM. Importantly, inhibition of Panx1 expression in liver cells with short hairpin RNA (shRNA) decreased supernatant eATP concentration and inhibited monocyte migration, indicating that monocyte migration is mediated in part by Panx1-dependent eATP release. Moreover, JNK inhibition decreased supernatant eATP concentration and inhibited Pannexin1 activation, as determined by YoPro-1 uptake in liver cells in a dose-dependent manner. These results suggest that JNK regulates activation of Panx1 channels, and provide evidence that Pannexin1-dependent pathophysiological eATP release in lipoapoptosis is capable of stimulating migration of human monocytes, and may participate in the recruitment of monocytes in chronic liver injury induced by saturated FFA.

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