Hydrogen Inhalation Reduces Lung Inflammation and Blood Pressure in the Experimental Model of Pulmonary Hypertension in Rats.

Hydrogen has been shown to exhibit selective antioxidant properties against hydroxyl radicals, and exerts antioxidant and anti-inflammatory effects. The monocrotaline-induced model of pulmonary hypertension is suitable for studying substances with antioxidant activity because oxidative stress is induced by monocrotaline. On day 1, male Wistar rats were subcutaneously injected with a water-alcohol solution of monocrotaline or a control with an only water-alcohol solution. One group of monocrotaline-injected animals was placed in a plastic box that was constantly ventilated with atmospheric air containing 4% of molecular hydrogen, and the two groups of rats, injected with monocrotaline or vehicle, were placed in boxes ventilated with atmospheric air. After 21 days, hemodynamic parameters were measured under urethane narcosis. The results showed that, although hydrogen inhalation had no effect on the main markers of pulmonary hypertension induced by monocrotaline injection, there was a reduction in systemic blood pressure due to its systolic component, and a decrease in TGF-ß expression, as well as a reduction in tryptase-containing mast cells.

Validity of Coronavirus Disease 2019 International Classification of Diseases, Tenth Revision in the Urgent Care Setting and Impact on Antibiotic Prescribing Rates.

We validated  different coronavirus disease 2019 (COVID-19) International Classification of Diseases, Tenth Edition (ICD-10) encounter definitions across 2 urgent care clinics. Sensitivity of definitions varied throughout the pandemic. Inclusion of COVID-19 and COVID-19-like illness (CLI) ICD-10s rendered highest sensitivity but lowest specificity. Antibiotic prescribing rates were low for COVID-19 ICD-10 encounters, increasing with CLI ICD-10 encounters.

The cascade of care in testing and treatment of latent tuberculosis infection in liver transplant candidates.

BACKGROUND: Testing and treatment for latent tuberculosis infection (LTBI) can mitigate risk of active tuberculosis (TB) post-liver transplant (LT). Testing and treatment completion rates have been reported low in this population. Our study aims to quantify the proportion of LT candidates who completed LTBI care cascade in our center. METHODS: A retrospective chart review was conducted on LT candidates from 2012 to 2021. Primary outcome was the proportion of patients who completed each cascade stage. Secondary outcome was an analysis of factors associated with positive and indeterminate LTBI testing. RESULTS: Of the 273 LT candidates, 265 (97.1%) were referred to transplant infectious disease (TID), 264 (96.7%) had orders for interferon-gamma release assay (IGRA), 262 (96%) underwent TID evaluation, and 259 (94.9%) completed IGRA. Twenty had LTBI, and 18 were treatment naïve and recommended for treatment. Of the 18, 15 (83.3%) agreed to therapy, 14 (77.8%) initiated treatment, and 12 (66.7%) completed treatment. No posttransplant TB reactivation occurred. Patients born in Asia, previous incarceration, past military service, and granuloma findings on chest imaging were likely to have positive IGRA (p < .05). Older age and travel to TB-endemic countries were likely to have indeterminate IGRA (p < .05). Indeterminate IGRAs were more common in QuantiFERON (QTF)-Gold Plus TB (15.3%) versus QTF-Gold TB (9.3%, p < .001). CONCLUSIONS: High rates of LTBI testing and treatment initiation and completion can be attributed to a standardized process that includes TID evaluation. Future studies in larger cohort are needed to better understand factors that can optimize the completion rates of LTBI treatment in LT candidates.

Shorter durations of antibiotic therapy in organ transplant.

PURPOSE OF REVIEW: Recent evidence supports shorter courses of antibiotics for several common infections and prophylactic indications. Unfortunately, solid organ transplant patients are often underrepresented or excluded from these studies. As a result, prolonged antibiotic durations are often used in clinical practice despite a lack of demonstrable benefit. This paper reviews recent publications addressing antibiotic duration of therapy in SOT recipients. RECENT FINDINGS: Although largely limited to observational studies, longer courses of antibiotics for surgical prophylaxis, urinary tract infections, and bloodstream infections have not demonstrated benefit compared to shorter courses. In some instances, longer courses of therapy have been associated with harm (i.e., adverse drug events and development of resistance). SUMMARY: Although the data remains limited, findings from retrospective studies evaluating shorter courses of antibiotics in SOT patients is encouraging. More robust research is desperately needed to define the optimal duration of antibiotics for common infections in SOT patients.

Real-world assessment of the clinical utility of whole body 18F-FDG PET/CT in the diagnosis of infection.

Few studies have aimed to capture the full spectrum of 18fluorodeoxyglucose-positron emission tomography/computed tomography (18F-FDG PET/CT) use for evaluation of infections in a real-world context. We performed a retrospective chart review of hospitalized patients who underwent 18F-FDG PET/CT for the workup of infection between April, 2013 and September, 2019. The clinical indications for and impact of 18F-FDG PET/CT on diagnostic and antimicrobial management were evaluated across different infectious indications. Sixty-one patients met the inclusion criteria. The most common indication was identifying a source of a known infection (46%), followed by fever of unknown etiology (FUE)/fever of unknown origin (FUO) (38%), and other (16%). 18F-FDG PET/CT was determined to have had a diagnostic or management clinical impact for a total of 22 patients (36%) including 12/28 (43%) of patients with known infection, 7/23 (30%) of patients with FUE/FUO, and 3/10 (30%) of patients with other indications. 18F-FDG PET/CT confirmed suspected prosthetic endovascular infection for 6/16 (38%) patients. In this study,18F-FDG PET/CT led to a clinical impact on diagnostic and treatment management of hospitalized patients across a variety of syndromes and particularly for source identification in the setting of known infection.

Dimensionality Mediated Highly Repeatable and Fast Transformation of Coordination Polymer Single Crystals for All-Optical Data Processing.

A family of coordination polymers (CPs) based on dynamic structural elements are of great fundamental and commercial interest addressing modern problems in controlled molecular separation, catalysis, and even data processing. Herein, the endurance and fast structural dynamics of such materials at ambient conditions are still a fundamental challenge. Here, we report on the design of a series of Cu-based CPs [Cu(bImB)Cl2] and [Cu(bImB)2Cl2] with flexible ligand bImB (1,4-bis(imidazol-1-yl)butane) packed into one- and two-dimensional (1D, 2D) structures demonstrating dimensionality mediated flexibility and reversible structural transformations. Using the laser pulses as a fast source of activation energy, we initiate CP heating followed by anisotropic thermal expansion and 0.2-0.8% volume changes with the record transformation rates from 2220 to 1640 s-1 for 1D and 2D CPs, respectively. The endurance over 103 cycles of structural transformations, achieved for the CPs at ambient conditions, allows demonstrating optical fiber integrated all-optical data processing.

Sex-Dependent Protective Effect of Combined Application of Solubilized Ubiquinol and Selenium on Monocrotaline-Induced Pulmonary Hypertension in Wistar Rats.

Ubiquinol exhibits anti-inflammatory and antioxidant properties. Selenium is a part of a number of antioxidant enzymes. The monocrotaline inducible model of pulmonary hypertension used in this study includes pathological links that may act as an application for the use of ubiquinol with high bioavailability and selenium metabolic products. On day 1, male and female rats were subcutaneously injected with a water-alcohol solution of monocrotaline or only water-alcohol solution. On days 7 and 14, some animals were intravenously injected with either ubiquinol's vehicle or solubilized ubiquinol, or orally with selenium powder daily, starting from day 7, or received both ubiquinol + selenium. Magnetic resonance imaging of the lungs was performed on day 20. Hemodynamic parameters and morphometry were measured on day 22. An increased right ventricle systolic pressure in relation to control was demonstrated in all groups of animals of both sexes, except the group of males receiving the combination of ubiquinol + selenium. The relative mass of the right ventricle did not differ from the control in all groups of males and females receiving either ubiquinol alone or the combination. Magnetic resonance imaging revealed impaired perfusion in almost all animals examined, but pulmonary fibrosis developed in only half of the animals in the ubiquinol group. Intravenous administration of ubiquinol has a protective effect on monocrotaline-induced pulmonary hypertension development resulting in reduced right ventricle hypertrophy, and lung mass. Ubiquinol + selenium administration resulted in a less severe increase in the right ventricle systolic pressure in male rats but not in females 3 weeks after the start of the experiment. This sex-dependent effect was not observed in the influence of ubiquinol alone.

Abnormal liver findings as the first manifestation in a patient with brucellosis : Case report.

In January 2019, a 30-year-old woman admitted to our inpatient department presented with undulating fever, pain in several joints, and significantly elevated liver enzymes and lactate dehydrogenase. After extended examination, infection with Brucella melitensis with liver, musculoskeletal, and pulmonary involvement was diagnosed and treated. Diagnosis was based on clinical examination, laboratory findings including seroconversion as a proof of immune response, magnetic resonance imaging, three-phase bone scintigraphy, and F­18 FDG-PET (F-18 Flourdeoxyglucose positron emission tomography) illustrating the bone involvement and its normalization upon treatment. After treatment the patient showed a remarkable improvement of clinical symptoms within a short period. The patient remained symptom free and polymerase chain reaction (PCR) testing for brucellosis was negative, even at the follow-up examination 12 months after the end of the antibiotic therapy. The family members were also examined due to the similar travel history, and by this, brucellosis was also diagnosed in her husband but not in her children.

IGFBP-4 Proteolysis by PAPP-A in a Primary Culture of Rat Neonatal Cardiomyocytes under Normal and Hypertrophic Conditions.

Cardiovascular diseases (CVD) are among the leading causes of death and disability worldwide. Pregnancy-associated plasma protein-A (PAPP-A) is a matrix metalloprotease localized on the cell surface. One of the substrates that PAPP-A cleaves is the insulin-like growth factor binding protein-4 (IGFBP-4), a member of the family of proteins that bind insulin-like growth factor (IGF). Proteolysis of IGFBP-4 by PAPP-A occurs at a specific site resulting in formation of two proteolytic fragments - N-terminal IGFBP-4 (NT-IGFBP-4) and C-terminal IGFBP-4 (CT-IGFBP-4), and leads to the release of IGF activating various cellular processes including migration, proliferation, and cell growth. Increased levels of the proteolytic IGFBP-4 fragments correlate with the development of CVD complications and increased risk of death in patients with the coronary heart disease, acute coronary syndrome, and heart failure. However, there is no direct evidence that PAPP-A specifically cleaves IGFBP-4 in the cardiac tissue under normal and pathological conditions. In the present study, using a primary culture of rat neonatal cardiomyocytes as a model, we have demonstrated that: 1) proteolysis of IGFBP-4 by PAPP-A occurs in the conditioned medium of cardiomyocytes, 2) PAPP-A-specific IGFBP-4 proteolysis is increased when cardiomyocytes are transformed to a hypertrophic state. Thus, it can be assumed that the enhancement of IGFBP-4 cleavage by PAPP-A and hypertrophic changes in cardiomyocytes accompanying CVD are interrelated, and PAPP-A appears to be one of the activators of the IGF-dependent processes in normal and hypertrophic-state cardiomyocytes.

Desmin Interacts Directly with Mitochondria.

Desmin intermediate filaments (IFs) play an important role in maintaining the structural and functional integrity of muscle cells. They connect contractile myofibrils to plasma membrane, nuclei, and mitochondria. Disturbance of their network due to desmin mutations or deficiency leads to an infringement of myofibril organization and to a deterioration of mitochondrial distribution, morphology, and functions. The nature of the interaction of desmin IFs with mitochondria is not clear. To elucidate the possibility that desmin can directly bind to mitochondria, we have undertaken the study of their interaction in vitro. Using desmin mutant Des(Y122L) that forms unit-length filaments (ULFs) but is incapable of forming long filaments and, therefore, could be effectively separated from mitochondria by centrifugation through sucrose gradient, we probed the interaction of recombinant human desmin with mitochondria isolated from rat liver. Our data show that desmin can directly bind to mitochondria, and this binding depends on its N-terminal domain. We have found that mitochondrial cysteine protease can disrupt this interaction by cleavage of desmin at its N-terminus.

Detection of Neprilysin-Derived BNP Fragments in the Circulation: Possible Insights for Targeted Neprilysin Inhibition Therapy for Heart Failure.

BACKGROUND: Entresto™ is a new heart failure (HF) therapy that includes the neprilysin (NEP) inhibitor sacubitril. One of the NEP substrates is B-type natriuretic peptide (BNP); its augmentation by NEP inhibition is considered as a possible mechanism for the positive effects of Entresto. We hypothesized that the circulating products of BNP proteolysis by NEP might reflect NEP impact on the metabolism of active BNP. We suggest that NEP-based BNP cleavage at position 17-18 results in BNP ring opening and formation of a novel epitope with C-terminal Arg-17 (BNP-neo17 form). In this study, we use a specific immunoassay to explore BNP-neo17 in a rat model and HF patient plasma. METHODS: We injected BNP into rats, with or without NEP inhibition with sacubitril. BNP-neo17 in plasma samples at different time points was measured with a specific immunoassay with neglectable cross-reactivity to intact forms. BNP-neo17 and total BNP were measured in EDTA plasma samples of HF patients. RESULTS: BNP-neo17 generation in rat circulation was prevented by NEP inhibition. The maximum 13.2-fold difference in BNP-neo17 concentrations with and without sacubitril was observed at 2 min after injection. BNP-neo17 concentrations in 32 HF patient EDTA plasma samples ranged from 0 to 37 pg/mL (median, 5.4; interquartile range, 0-9.1). BNP-neo17/total BNP had no correlation with total BNP concentration (with r = -0.175, P = 0.680) and showed variability among individuals. CONCLUSIONS: BNP-neo17 formation is NEP dependent. Considering that BNP-neo17 is generated from the active form of BNP by NEP, we speculate that BNP-neo17 may reflect both the NEP activity and natriuretic potential and serve for HF therapy guidance.

Deceptive but Not Honest Manipulative Actions Are Associated with Increased Interaction between Middle and Inferior Frontal gyri.

The prefrontal cortex is believed to be responsible for execution of deceptive behavior and its involvement is associated with greater cognitive efforts. It is also generally assumed that deception is associated with the inhibition of default honest actions. However, the precise neurophysiological mechanisms underlying this process remain largely unknown. The present study was aimed to use functional magnetic resonance imaging to reveal the underlying functional integration within the prefrontal cortex during the task which requires that subjects to deliberately mislead an opponent through the sequential execution of deceptive and honest claims. To address this issue, we performed psychophysiological interaction (PPI) analysis, which allows for statistical assessment of changes in functional relationships between active brain areas in changing psychological contexts. As a result the whole brain PPI-analysis established that both manipulative honest and deceptive claiming were associated with an increase in connectivity between the left middle frontal gyrus and right temporo-parietal junction (rTPJ). Taking into account the role played by rTPJ in processes associated with the theory of mind the revealed data can reflect possible influence of socio-cognitive context on the process of selecting manipulative claiming regardless their honest or deceptive nature. Direct comparison between deceptive and honest claims revealed pattern enhancement of coupling between the left middle frontal gyrus and the left inferior frontal gyrus. This finding provided evidence that the execution of deception relies to a greater extent on higher-order hierarchically-organized brain mechanisms of executive control required to select between two competing deceptive or honest task sets.

Comparison of the long-term behavioral effects of neonatal exposure to retigabine or phenobarbital in rats.

Anticonvulsant drugs, when given during vulnerable periods of brain development, can have long-lasting consequences on nervous system function. In rats, the second postnatal week approximately corresponds to the late third trimester of gestation/early infancy in humans. Exposure to phenobarbital during this period has been associated with deficits in learning and memory, anxiety-like behavior, and social behavior, among other domains. Phenobarbital is the most common anticonvulsant drug used in neonatology. Several other drugs, such as lamotrigine, phenytoin, and clonazepam, have also been reported to trigger behavioral changes. A new generation anticonvulsant drug, retigabine, has not previously been evaluated for long-term effects on behavior. Retigabine acts as an activator of KCNQ channels, a mechanism that is unique among anticonvulsants. Here, we examined the effects retigabine exposure from postnatal day (P)7 to P14 on behavior in adult rats. We compared these effects with those produced by phenobarbital (as a positive control) and saline (as a negative control). Motor behavior was assessed by using the open field and rotarod, anxiety-like behavior by the open field, elevated plus maze, and light-dark transition task, and learning/memory by the passive avoidance task; social interactions were assessed in same-treatment pairs, and nociceptive sensitivity was assessed via the tail-flick assay. Motor behavior was unaltered by exposure to either drug. We found that retigabine exposure and phenobarbital exposure both induced increased anxiety-like behavior in adult animals. Phenobarbital, but not retigabine, exposure impaired learning and memory. These drugs also differed in their effects on social behavior, with retigabine-exposed animals displaying greater social interaction than phenobarbital-exposed animals. These results indicate that neonatal retigabine induces a subset of behavioral alterations previously described for other anticonvulsant drugs and extend our knowledge of drug-induced behavioral teratogenesis to a new mechanism of anticonvulsant action.

Profile of anticonvulsant action of levetiracetam, tiagabine and phenobarbital against seizures evoked by DMCM (methyl-6,7-dimethoxy-4-ethyl-ß-carboline-3-carboxylate) in neonatal rats.

Levetiracetam (LEV) and tiagabine (TGB) are utilized for the treatment of seizures, including neonatal seizures. However, relatively little is known about the preclinical therapeutic profile of these drugs during brain development. The relative paucity of information regarding these drugs in neonatal animals may be due to their unusual profile of anticonvulsant action in experimental models. LEV and TGB are without effect against seizures in several common screening models (e.g., the maximal electroshock test, maximal pentylenetetrazole seizures), instead showing preferential efficacy against models of partial seizures. We have recently described a method for reliably evoking partial seizures in neonatal animals by systemic administration of the chemoconvulsant, DMCM (Kulick et al., 2014, Eur. J. Pharmacol., doi:10.1016/j.ejphar.2014.06.012). DMCM is a negative allosteric modulator of GABAA receptors, and offers a wide separation between doses required to evoke complex partial as compared to tonic-clonic seizures. Here we used DMCM to evaluate the effect of LEV and TGB against seizures in postnatal day (P) 10 rat pups. We compared the profile of LEV and TGB to that of phenobarbital (PB), the most widely utilized anticonvulsant in neonates. We found that LEV significantly protected against DMCM seizures when administered in doses of 10mg/kg and greater. TGB protected against DMCM-evoked seizures when administered in doses of 1mg/kg or greater. PB protected against DMCM-evoked seizures when administered in doses of 5mg/kg or greater. These data provide preclinical evidence for the efficacy of LEV and TGB in neonates and underscore the utility of DMCM for screening anticonvulsant action in neonatal animals.

Possible role of an error detection mechanism in brain processing of deception: PET-fMRI study.

To investigate brain maintenance of deliberate deception the positron emission tomography and the event related functional MRI studies were performed. We used an experimental paradigm that presupposed free choices between equally beneficial deceptive or honest actions. Experimental task simulated the "Cheat" card game which aims to defeat an opponent by sequential deceptive and honest claims. Results of both the PET and the fMRI studies revealed that execution of both deliberately deceptive and honest claims is associated with fronto-parietal brain network comprised of inferior and middle frontal gyri, precentral gyrus (BA 6), caudate nucleus, and inferior parietal lobule. Direct comparison between those claims, balanced in terms of decision making and action outcome (gain and losses), revealed activation of areas specifically associated with deception execution: precentral gyrus (BA 6), caudate nuclei, thalamus and inferior parietal lobule (BA 39/40). The obtained experimental data were discussed in relation to a possible role of an error detection system in processing deliberate deception.

Human pro-B-type natriuretic peptide is processed in the circulation in a rat model.

BACKGROUND: The appearance of B-type natriuretic peptide (BNP) in the blood is ultimately caused by proteolytic processing of its precursor, proBNP. The mechanisms leading to the high plasma concentration of unprocessed proBNP are still poorly understood. The goals of the present study were to examine whether processing of proBNP takes place in the circulation and to evaluate the clearance rate of proBNP and proBNP-derived peptides. METHODS: We studied the processing of human proBNP in the circulation and the clearance rate of proBNP and proBNP-derived peptides (BNP and N-terminal fragment of proBNP, NT-proBNP) in rats by injecting the corresponding peptides and analyzing immunoreactivity at specific time points. Glycosylated and nonglycosylated proBNP and NT-proBNP were used in the experiments. We applied immunoassays, gel filtration, and mass spectrometry (MS) techniques to analyze the circulation-mediated processing of proBNP. RESULTS: ProBNP was effectively processed in the circulation into BNP (1-32) and various truncated BNP forms as confirmed by gel filtration and MS analysis. Glycosylation of proBNP close to the cleavage-site region suppressed its processing in the circulation. The terminal half-life for human glycosylated proBNP was 9.0 (0.5) min compared with 6.4 (0.5) min for BNP. For NT-proBNP, the terminal half-lives were 15.7 (1.4) min and 15.5 (1.3) min for glycosylated and nonglycosylated forms, respectively. CONCLUSIONS: In rats, processing of human proBNP to active BNP occurs in the circulation. The clearance rate of proBNP is quite similar to that of BNP. These observations suggest that peripheral proBNP processing may be an important regulatory step rather than mere degradation.

Extracellular HspBP1 and Hsp72 synergistically activate epidermal growth factor receptor.

BACKGROUND INFORMATION: Heat-inducible Hsp72 is the founding member of the Hsp70 (heat shock proteins of 70 kDa) family of molecular chaperones. It is localized primarily in cytoplasm and nucleus but is also found extracellularly. The source of e-Hsp72 (extracellular Hsp72) is not precisely identified and may not be the same in every situation. A number of studies demonstrated that e-Hsp72 plays an important role in cell survival, tumour rejection and immune response. However, currently little is known about regulation of e-Hsp72 function. In cells, Hsp72 is controlled by co-chaperones. An abundant co-chaperone, HspBP1 (Hsp72-binding protein 1) was found extracellularly in the serum. In the present study we analysed the secretion and function of e-HspBP1 (extracellular HspBP1). RESULTS: A431 human squamous carcinoma cells accumulated Hsp72 and HspBP1 in chromogranin A-positive granules following heat stress or in the presence of U73122, an inhibitor of phospholipase C. Following these treatments, A431 cells also increased the secretion of both proteins into the culture medium. The secreted e-Hsp72 and e-HspBP1 were co-immunoprecipitated from the conditioned medium. Purified recombinant HspBP1 augmented e-Hsp72-mediated phosphorylation of EGFR (epidermal growth factor receptor) and its down-stream targets, ERK1 (extracellular signal-regulated kinase 1) and ERK2 in a concentration-dependent manner. Finally, a HspBP1 N-terminal domain deletion mutant and boiled recombinant HspBP1 did not affect the e-Hsp72-mediated activity. CONCLUSIONS: Heat stress and PLC (phospholipase C) inhibition result in the enhanced secretion of both Hsp72 and HspBP1. In an extracellular environment, the two chaperones interact both physically and functionally, leading to the activation of th EGFR-ERK1/2 signalling pathway. However, the magnitude of EGFR activation depends on the e-HspBP1/e-Hsp72 ratio in the medium. Extracellular chaperone-mediated activation of EGFR can provide a survival advantage to cells under heat shock and other stresses.

Phosphorylation of histone H2AX in radiation-induced micronuclei.

DNA double-strand breaks are thought to precede the formation of most radiation-induced micronuclei. Phosphorylation of the histone H2AX is an early indicator of DNA double-strand breaks. Here we studied the phosphorylation status of the histone H2AX in micronuclei after exposure of cultured cells to ionizing radiation or treatment with colchicine. In human astrocytoma SF268 cells, after exposure to gamma radiation, the proportion of gamma-H2AX-positive to gamma-H2AX-negative micronuclei increases. The majority of the gamma-H2AX-positive micronuclei are centromere-negative. The number of gamma-H2AX-positive micronuclei continues to increase even 24 h postirradiation when most gamma-H2AX foci in the main nucleus have disappeared. In contrast, in normal human fibroblasts (BJ), the proportion of gamma-H2AX-positive to gamma-H2AX-negative micronuclei remains constant, and the majority of the centromere-negative cells are gamma-H2AX-negative. Treatment of both cell lines with colchicine results in mostly centromere-positive, gamma-H2AX-negative micronuclei. Immunostaining revealed co-localization of MDC1 and ATM with gamma-H2AX foci in both main nuclei and micronuclei; however, other repair proteins, such as Rad50, 53BP1 and Rad17, that co-localized with gamma-H2AX foci in the main nuclei were not found in the micronuclei. Combination of the micronucleus assay with gamma-H2AX immunostaining provides new insights into the mechanisms of the formation and fate of micronuclei.

Cyclization of natural allene oxide in aprotic solvent: formation of the novel oxylipin methyl cis-12-oxo-10-phytoenoate.

Allene oxide, (9Z,11E)-12,13-epoxy-9,11-octadecadienoic acid (12,13-EOD), was prepared by incubation of linoleic acid (13S)-hydroperoxide with flaxseed allene oxide synthase (AOS) and purified (as methyl ester) by low temperature HPLC. Identification of pure 12,13-EOD was substantiated by its UV and (1)H NMR spectra and by GC-MS data for its methanol trapping product. The methyl ester of 12,13-EOD (but not the free carboxylic acid) is slowly cyclized in hexane solution, affording a novel cyclopentenone cis-12-oxo-10-phytoenoic acid. Free carboxylic form of 12,13-EOD does not cyclize due to the exceeding formation of macrolactone (9Z)-12-oxo-9-octadecen-11-olide. The spontaneous cyclization of pure natural allene oxide (12,13-EOD) into cis-cyclopentenone have been observed first time.

Extracellular heat shock protein 70 mediates heat stress-induced epidermal growth factor receptor transactivation in A431 carcinoma cells.

The initial steps of heat stress in A431 cells were previously characterized by ligand-independent EGFR transactivation via an unknown mechanism and concomitant secretion of Hsp70. In this work we demonstrate that the depletion of Hsp70 from the conditioned medium of heated cells abolishes EGFR transactivation indicating that secreted Hsp70 is essential for EGFR transactivation during heat shock. This notion is supported by the findings that purified Hsp70 can induce EGFR transactivation and the activation of EGFR-dependent signaling pathways. Both heat stress and pure Hsp70 stimulate activation of TLR2/4 and their association with EGFR. These results suggest that the secreted Hsp70 mediates the cross-communication of TLR and EGFR signaling systems in A431 cells.