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Background: Acinetobacter baumanni infection is a leading cause of morbidity and mortality in the Intensive Care Unit (ICU). Early recognition of patients at risk for infection allows early proper treatment and is associated with improved outcomes. This study aimed to construct an innovative Machine Learning (ML) based prediction tool for Acinetobacter baumanni infection, among patients in the ICU, and to examine its robustness and predictive power., Methods: For model development and internal validation, we used The Medical Information Mart for Intensive Care database (MIMIC) III data from 19,690 consecutive adult patients admitted between 2001 and 2012 at a Boston tertiary center ICU. For external validation, we used a different dataset from Rabin Medical Center (RMC, Israeli tertiary center) ICU, of 1,700 patients admitted between 2017 and 2021. After training on MIMIC cohorts, we adapted the algorithm from MIMIC to RMC and evaluated its discriminating power in terms of Area Under the Receiver Operating Curve (AUROC), sensitivity, specificity, Negative Predictive Value and Positive Predictive Value., Results: The prediction model achieved AUROC = 0.624 (95% CI 0.604-0.647). The most significant predictors were (i) physiological parameters of cardio-respiratory function, such as carbon dioxide (CO2) levels and respiratory rate, (ii) metabolic disturbances such as lactate and acidosis (pH) and (iii) past administration of antibiotics., Conclusions: Infection with Acinetobacter baumanni is more likely to occur in patients with respiratory failure and higher lactate levels, as well as patients who have used larger amounts of antibiotics. The accuracy of Acinetobacter prediction may be enhanced by future studies., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Neuman et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Adequate pain control following lung transplantation (LTx) surgery is paramount. Thoracic epidural analgesia (TEA) is the gold standard; however, the potential use of extracorporeal membrane oxygenation (ECMO) and consequent anticoagulation therapy raises safety concerns, prompting clinicians to seek safer alternatives. The utility of thoracic wall blocks in general thoracic surgery is well established; however, their role in the context of LTx has been poorly investigated., Methods: In this retrospective exploratory study, we assessed the effect of adding a superficial parasternal intercostal plane (sPIP) block and serratus anterior plane (SAP) block to standard anesthetic and analgesic care on tracheal extubation rates, pain scores and opioid consumption until 72 hours postoperatively in LTx., Results: Sixty patients were included in the analysis; 35 received the standard anesthetic and analgesic care (control group), and 25 received sPIP and SAP blocks in addition to the standard anesthetic and analgesic care (intervention group). We observed higher tracheal extubation rates in the intervention group at 8 hours postoperatively (16.0% vs 0.0%, p=0.03). This was also shown after adjusting for known prognostic factors (OR 1.18; 95% CI 1.04 to 1.33, p=0.02). Furthermore, we noted a lower opioid consumption measured by morphine milligram equivalents at 24 hours in the intervention group (median 405 (IQR 300-490) vs 266 (IQR 168-366), p=0.02). This was also found after adjusting for known prognostic factors (β -118; 95% CI -221 to 14, p=0.03)., Conclusion: sPIP and SAP blocks are safe regional analgesic techniques in LTx involving ECMO and clamshell incision. They are associated with faster tracheal extubation and lower opioid consumption. These techniques should be considered when TEA is not appropriate. Further high-quality studies are warranted to confirm these findings., Competing Interests: Competing interests: None declared., (© American Society of Regional Anesthesia & Pain Medicine 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: This case involved a 67-year-old female who was admitted for general anesthesia for a mitral clip procedure. Following anesthesia induction, the patient underwent an uneventful orotracheal intubation. Shortly afterwards she developed an ongoing respiratory failure, accompanied by neck and chest subcutaneous emphysema. Upon workup, she was found to have a 6 cm long tracheal laceration on the posterior side. Emergency repair surgery was performed using an extracorporeal membrane oxygenator (ECMO). The patient passed away 11 days later from sepsis. The aim of this case report is to increase awareness of this rare intubation complication, and explore the best approach to prevent, diagnose and treat tracheal injuries during endotracheal intubation.

Background: Intraoperative pain is a possible complication of neuraxial anaesthesia for caesarean delivery. There is little information available about its incidence, risk factors and physician perception., Methods: Parturients undergoing spinal anaesthesia for elective caesarean delivery were enrolled. Before surgery, parturients were asked about preoperative anxiety on a verbal numerical scale (VNS), anticipated analgesic requirement, postoperative pain levels, Spielberger STATE-TRAIT inventory index, Pain Catastrophizing Scale. After surgery, parturients were asked to answer questions (intraoperative VNS pain). The anaesthesiologist and obstetrician were asked to fill out a questionnaire asking about perceived intraoperative pain. Influence of preoperative anxiety on intraoperative pain (yes/no) was assessed using logistic regression. Mc Fadden's R 2 was calculated. The agreement in physician perception of intraoperative pain with reported pain by the parturient was examined by calculating Cohen's kappa and 95% Confidence Intervals (CI)., Results: We included 193 parturients in our analysis. Incidence of intraoperative pain was 11.9%. Median intraoperative VNS pain of parturients with pain was 4.0 (1st quartile 4.0; 3rd quartile 9.0). Preoperative anxiety was not a good predictor of intraoperative pain (p-value of β-coefficient = 0.43, Mc Fadden's R 2  = 0.01). Including further preoperative variables did not result in a good prediction model. Cohen's kappa between reported pain by parturient and by the obstetrician was 0.21 (95% CI: 0.01, 0.41) and by the anaesthesiologist was 0.3 (95% CI: 0.12, 0.48)., Conclusions: We found a substantial incidence (11.9%) of intraoperative pain during caesarean delivery. Preoperative anxiety did not predict intraoperative pain. Physicians did not accurately identify parturients' intraoperative pain., Significance: Intraoperative pain occurred in 11.9% and severe intraoperative pain occurred in 1.11% of parturients undergoing elective caesarean delivery under spinal anaesthesia. We did not find any preoperative variables that could reliably predict intraoperative pain. Obstetricians and anaesthesiologists underestimated the incidence of intraoperative pain in our cohort and thus, more attention must be put to parturients' pain., (© 2021 The Authors. European Journal of Pain published by John Wiley & Sons Ltd on behalf of European Pain Federation - EFIC ®.)

The OXE receptor is a GPCR activated by eicosanoids produced by the action of 5-lipoxygenase. We previously found that this membrane receptor participates in the regulation of cAMP-dependent and -independent steroidogenesis in human H295R adrenocortical carcinoma cells. In this study we analyzed the effects of the OXE receptor physiological activator 5-oxo-ETE on the growth and migration of H259R cells. While 5-oxo-ETE did not affect the growth of H295R cells, overexpression of OXE receptor caused an increase in cell proliferation, which was further increased by 5-oxo-ETE and blocked by 5-lipoxygenase inhibition. 5-oxo-ETE increased the migratory capacity of H295R cells in wound healing assays, but it did not induce the production of metalloproteases MMP-1, MMP-2, MMP-9 and MMP-10. The pro-migratory effect of 5-oxo-ETE was reduced by pharmacological inhibition of the MEK/ERK1/2, p38 and PKC pathways. 5-oxo-ETE caused significant activation of ERK and p38. ERK activation by the eicosanoid was reduced by the "pan" PKC inhibitor GF109203X but not by the classical PKC inhibitor Gö6976, suggesting the involvement of novel PKCs in this effect. Although H295R cells display detectable phosphorylation of Ser299 in PKCδ, a readout for the activation of this novel PKC, treatment with 5-oxo-ETE per se was unable to induce additional PKCδ activation. Our results revealed signaling effectors activated by 5-oxo-ETE in H295R cells and may have significant implications for our understanding of OXE receptor in adrenocortical cell pathophysiology., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Hormone-regulated steroidogenesis and StAR protein induction involve the action of lipoxygenated products. The products of 5-lipoxygenase act on inflammation and immunity by stimulation of a membrane receptor called OxeR1. The presence of OxeR1 in other systems has not been described up to date and little is known about its mechanism of action and other functions. In this context, the aim of this study was the identification and characterization of OxeR1 as a mediator of cAMP-dependent and independent pathways. Overexpression of OxeR1 in MA-10 Leydig cells increased cAMP-dependent progesterone production. Angiotensin II and cAMP stimulation of adrenocortical human H295R cells produced an increase in StAR protein induction and steroidogenesis in cells overexpressing OxeR1 as compared to mock-transfected cells. Additionally, activation of OxeR1 caused a time-dependent increase in ERK1/2 phosphorylation. In summary, membrane receptor OxeR1 is involved in StAR protein induction and activation of steroidogenesis triggered by cAMP or angiotensin II, acting, at least in part, through ERK1/2 activation., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

In rat isolated mesenteric beds that were contracted with NA as an in vitro model of the vascular adrenergic hyperactivity that usually precedes the onset of primary hypertension, the oral administration (3 daily doses) of either 10 mg/kg genistein or 20 mg/kg daidzein potentiated the anandamide-induced reduction of contractility to NA in female but not in male rats. Oral treatment with phytoestrogens also restored the vascular effects of anandamide as well as the mesenteric content of calcitonin gene-related peptide (CGRP) that were reduced after ovariectomy. The enhancement of anandamide effects caused by phytoestrogens was prevented by the concomitant administration of the estrogen receptor antagonist fulvestrant (2.5 mg/kg, s.c., 3 daily doses). It is concluded that, in the vasculature of female rats, phytoestrogens produced an estrogen-receptor-dependent enhancement of the anandamide-vascular actions that involves the modulation of CGRP levels and appears to be relevant whenever an adrenergic hyperactivity occurs.

It is known that ERK1/2 and MEK1/2 participate in the regulation of Star gene transcription. However, their role in StAR protein post-transcriptional regulation is not described yet. In this study we analyzed the relationship between the MAPK cascade and StAR protein phosphorylation and function. We have demonstrated that (a) steroidogenesis in MA-10 Leydig cells depends on the specific of ERK1/2 activation at the mitochondria; (b) ERK1/2 phosphorylation is driven by mitochondrial PKA and constitutive MEK1/2 in this organelle; (c) active ERK1/2 interacts with StAR protein, leads to StAR protein phosphorylation at Ser(232) only in the presence of cholesterol; (d) directed mutagenesis of Ser(232) (S232A) inhibited in vitro StAR protein phosphorylation by ERK1; (e) transient transfection of MA-10 cells with StAR S232A cDNA markedly reduced the yield of progesterone production. We show that StAR protein is a substrate of ERK1/2, and that mitochondrial ERK1/2 is part of a multimeric complex that regulates cholesterol transport.

The studies presented herein were designed to investigate the effect of mouse epidermal growth factor (mEGF) on arachidonic acid (AA) release in a clonal strain of cultured murine Leydig cells (designed MA-10). In MA-10 cells, mEGF promotes AA release and metabolism to lipoxygenated products to induce the steroidogenic acute regulatory (StAR) protein. However, the mechanism by which mEGF releases AA in these cells is not totally elucidated. We show that mEGF produces an increment in the mitochondrial AA content in a short-term incubation (30 min). This AA is released by the action of a mitochondrial acyl-CoA thioesterase (Acot2), as demonstrated in experiments in which Acot2 was down or overexpressed. This AA in turn regulates the StAR protein expression, indirect evidence of its metabolism to lipoxygenated products. We also show that mEGF induces the expression (mRNA and protein) of Acot2 and an acyl-CoA synthetase that provides the substrate, arachidonyl-CoA, to Acot2. This effect is also observed in another steroidogenic cell line, the adrenocortical Y1 cells. Taken together, our results show that: 1) mEGF can induce the generation of AA in a specific compartment of the cells, i.e. the mitochondria; 2) mEGF can up-regulate acyl-CoA synthetase and Acot2 mRNA and protein levels; and 3) mEGF-stimulated intramitochondrial AA release leads to StAR protein induction.

ERK1/2 is known to be involved in hormone-stimulated steroid synthesis, but its exact roles and the underlying mechanisms remain elusive. Both ERK1/2 phosphorylation and steroidogenesis may be triggered by cAMP/cAMP-dependent protein kinase (PKA)-dependent and-independent mechanisms; however, ERK1/2 activation by cAMP results in a maximal steroidogenic rate, whereas canonical activation by epidermal growth factor (EGF) does not. We demonstrate herein by Western blot analysis and confocal studies that temporal mitochondrial ERK1/2 activation is obligatory for PKA-mediated steroidogenesis in the Leydig-transformed MA-10 cell line. PKA activity leads to the phosphorylation of a constitutive mitochondrial MEK1/2 pool with a lower effect in cytosolic MEKs, while EGF allows predominant cytosolic MEK activation and nuclear pERK1/2 localization. These results would explain why PKA favors a more durable ERK1/2 activation in mitochondria than does EGF. By means of ex vivo experiments, we showed that mitochondrial maximal steroidogenesis occurred as a result of the mutual action of steroidogenic acute regulatory (StAR) protein -a key regulatory component in steroid biosynthesis-, active ERK1/2 and PKA. Our results indicate that there is an interaction between mitochondrial StAR and ERK1/2, involving a D domain with sequential basic-hydrophobic motifs similar to ERK substrates. As a result of this binding and only in the presence of cholesterol, ERK1/2 phosphorylates StAR at Ser(232). Directed mutagenesis of Ser(232) to a non-phosphorylable amino acid such as Ala (StAR S232A) inhibited in vitro StAR phosphorylation by active ERK1/2. Transient transfection of MA-10 cells with StAR S232A markedly reduced the yield of progesterone production. In summary, here we show that StAR is a novel substrate of ERK1/2, and that mitochondrial ERK1/2 is part of a multimeric protein kinase complex that regulates cholesterol transport. The role of MAPKs in mitochondrial function is underlined.

The aim of the present study was to determine whether the transient receptor potential vanilloid (TRPV1) receptor protein as well as the calcitonin gene-related peptide (CGRP) content could be enhanced after the i.p. administration of 5 mg/kg lipopolysaccharide (LPS) to Sprague-Dawley rats. In tongue tissue, used as a representative model of TRPV1 receptors expression, there was a significant increase in the abundance of TRPV1 receptor protein 6 h after LPS administration. In mesenteric arteries, the density of the CGRP-positive nerves as well as the release of CGRP induced by 10 microM anandamide was also significantly increased 6 h after LPS administration. The relaxant responses induced by anandamide in mesenteric beds isolated from either untreated or LPS-treated rats were abolished after a 2 h exposure to 10 microM capsaicin. Moreover, anandamide-induced relaxations of untreated mesenteries were potentiated by the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA, 0.1 microM), but not by its inactive analogue 4alpha-phorbol (0.1 microM). The potentiation of anandamide effects caused by the PKC activator was accompanied by a significant increase in the overflow of CGRP induced by anandamide in the untreated rats. It is proposed that the overexpression of the TRPV1 receptors and the increased content of CGRP could contribute to the enhancement of anandamide effects during the endotoxemic shock. An eventual phosphorylation event linked to the overflow of CGRP could also participate in the enhanced relaxation caused by anandamide in endotoxemia.

The activation of the rate-limiting step in steroid biosynthesis, that is the transport of cholesterol into the mitochondria, is dependent on PKA-mediated events triggered by hormones like ACTH and LH. Two of such events are the protein tyrosine dephosphorylation mediated by protein tyrosine phosphatases (PTPs) and the release of arachidonic acid (AA) mediated by two enzymes, ACS4 (acyl-CoA synthetase 4) and Acot2 (mitochondrial thioesterase). ACTH and LH regulate the activity of PTPs and Acot2 and promote the induction of ACS4. Here we analyzed the involvement of PTPs on the expression of ACS4. We found that two PTP inhibitors, acting through different mechanisms, are both able to abrogate the hormonal effect on ACS4 induction. PTP inhibitors also reduce the effect of cAMP on steroidogenesis and on the level of StAR protein, which facilitates the access of cholesterol into the mitochondria. Moreover, our results indicate that exogenous AA is able to overcome the inhibition produced by PTP inhibitors on StAR protein level and steroidogenesis. Then, here we describe a link between PTP activity and AA release, since ACS4 induction is under the control of PTP activity, being a key event for AA release, StAR induction and steroidogenesis.

We have described that, in adrenal and Leydig cells, the hormonal regulation of free arachidonic acid (AA) concentration is mediated by the concerted action of two enzymes: an acyl-CoA thioesterase (MTE-I or ARTISt) and an acyl-CoA synthetase (ACS4). In this study we analyzed the potential regulation of these proteins by hormonal action in steroidogenic cells. We demonstrated that ACS4 is rapidly induced by adrenocorticotropin (ACTH) and cAMP in Y1 adrenocortical cells. The hormone and its second messenger increased ACS4 protein levels in a time and concentration dependent way. Maximal concentration of ACTH (10 mIU/ml) produced a significant effect after 15 min of treatment and exerted the highest increase (3-fold) after 30 min. Moreover, (35)S-methionine incorporation showed that the increase in ACS4 protein levels is due to an increase in the de novo synthesis of the protein. On the contrary MTE-I protein levels in Y1 and MA-10 cells did not change after steroidogenic stimuli. In contrast with the effect observed on protein levels, stimulation of both cell lines did not change ACS4 RNA levels during the first hour of treatment, indicating that the effect of both stimuli is exerted at the level of ACS4 protein synthesis.StAR protein induction has a key role on the activation of steroidogenesis since this protein increases the rate of the limiting step of the whole process. In agreement with the fact that the inhibition of ACS4 activity by triacsin C blocks cAMP-stimulated progesterone production by MA-10 Leydig cells, here we demonstrated that ACS4 inhibition also reduces StAR protein levels. Moreover, exogenous AA was able to overcome the effect of triacsin C on both events, StAR induction and steroidogenesis. These results were confirmed by experiments using ACS4-targeted siRNA which result in a reduction in both ACS4 and StAR protein levels. The concomitant decrease in steroid production was overcome by the addition of AA to the knocked-out cells. In summary, this study suggests that in adrenal and Leydig cells the hormonal action prompts the synthesis of a labile protein, ACS4, which activity is involved in the regulation of AA release and is essential for steroidogenesis and StAR protein induction.

Background: It has been suggested that exercise-induced bronchoconstriction may involve oxidative stress. Strenuous exercise promotes free radical production, which can lead to many of the pathophysiologic changes associated with asthma, including bronchoconstriction, mucus secretion, and microvascular leakage. Lycopene has been shown to have high antioxidative activity., Objective: To evaluate the effect of lycopene supplementation on airway hyperreactivity and inflammation in young athletes who complain of difficulty in breathing related to physical exertion., Methods: Nineteen young athletes with exercise-related difficulty in breathing visited the exercise laboratory 3 times. During the first visit, participants underwent a baseline evaluation of exercise-induced bronchoconstriction. Daily for 1 week before each of the 2 subsequent visits, participants ingested 30 mg of lycopene (a natural antioxidant) or placebo (in randomized order, double-blind). A 2-week washout period was given between each visit. During each visit, lung functions were evaluated before and after an 8-minute run on the treadmill (85% of the predicted maximal heart rate)., Results: There was no difference in the mean+/-SD decrease in forced expiratory volume in 1 second after exercise during lycopene treatment compared with placebo treatment (11.8%+/-12.5% and 11.0%+/-11.6%). In addition, there was no apparent division into responders and nonresponders., Conclusion: A daily dose of lycopene for 1 week does not affect lung function after exercise and may not provide any protective effect against clinical difficulty in breathing in young athletes.

The ACTH signaling pathway includes both PKA activation as well as PKA-dependent tyrosine phosphatase activation. In addition, the action of this hormone also includes the regulation of the intracellular levels of arachidonic acid (AA) by the concerted action of two enzymes: an acylCoA-thioesterase and an acyl-CoA-synthetase (ACS4). This work describes the production and characterization of a specific ACS4 antibody, which was used to analyze the effect of ACTH on ACS4 protein level in Y1 adrenocortical cells and the putative relationship between tyrosine phosphatases and ACS4. The antiserum was obtained from rabbits immunized with the recombinant ACS4. This immunogen was produced in bacteria and eluted from an acrylamide gel after SDS-PAGE separation of a partially purified bacteria lysate. When used in Western blot analysis, the antibody obtained specifically recognized only one protein of the molecular mass corresponding to ACS4, in Y1 cells and in several rat tissues. Using the antibody described here, we analyzed the effect of ACTH stimulation on ACS4 protein level. The hormone produced an increase of this acyl-CoA synthetase in Y1 adrenocortical cells. Moreover, this effect was mimicked by cAMP and partially reduced by a tyrosine phosphatase inhibitor. We propose that ACTH regulates ACS4 protein levels through a PKA-dependent mechanism that could involve also PTP activity.

Arachidonic acid is not freely stored in the cells. A number of different pathways for the mobilization of this compound have been proposed, including a novel mechanism that involves the release of arachidonic acid from arachidonoyl-CoA by a thioesterase with substrate specificity for very-long-chain fatty acids. In rat heart, the acyl-CoA thioesterase activity can be regulated by a mechanism that involves beta-adrenoceptors. In this paper we demonstrate that beta-adrenergic agonists also regulate the acyl-CoA thioesterase mRNA levels. Isoproterenol (10(-7)M)-a concentration known to exert physiological responses-increases in a time-dependent manner the acyl-CoA thioesterase mRNA levels, an effect blocked by a specific beta-adrenoceptor antagonist. In addition, our results show that cAMP is involved in this process. The acyl-CoA thioesterase mRNA levels are also increased by fasting, but not by di(2-ethylhexyl)phthalate, a peroxisome proliferator. These results may suggest the existence of a beta-adrenoceptor-activated regulatory pathway for arachidonic acid release in cardiac tissue.

We have recently described an acyl-CoA thioesterase specific for very-long-chain fatty acids, named ARTISt, that regulates steroidogenesis through the release of arachidonic acid in adrenal zona fasciculata cells. In this paper we demonstrate the presence of the protein as a 43 kDa band and its mRNA in cardiac tissue. The activity of the protein was measured using an heterologous cell-free assay in which it is recombined with adrenal microsomes and mitochondria to activate mitochondrial steroidogenesis. Isoproterenol and phenylephrine activate the enzyme in a dose-dependent manner (10(-10)-10(-6) M). Both propranolol (10(-5) M) and prazosin (10(-5) M) block the action of isoproterenol and phenylephrine respectively. Antipeptide antibodies against the serine lipase motif of the protein and the Cys residue present in the catalytic domain also block the activity of the protein. Taken together, our results confirm the presence of ARTISt in heart and provide evidence for a catecholamine-activated regulatory pathway of the enzyme in that tissue.

Background: The unicellular alga Dunaliella bardawil was previously shown to contain very high concentrations of beta-carotene composed of equal amounts of the all-trans and 9-cis stereoisomers which differ in their physicochemical features and antioxidative activity. Due to the controversy regarding the beneficial effect of antioxidants on asthma, the acute effects of beta-carotene of Dunaliella was assessed on airway hyperreactivity in patients with exercise-induced asthma (EIA)., Methods: Thirty-eight patients with EIA participated in our study to verify the antioxidative effect. The test was based on the following sequence: baseline pulmonary function, 7 minutes exercise session on a motorized treadmill, 8 minutes rest, 1-week oral random, double-blind supplementation of placebo or 64 mg/day beta-carotene, pulmonary functions at rest, 7 minutes exercise session, 8 minutes rest and again pulmonary functions., Results: All patients given placebo showed a significant postexercise reduction of more than 15% in their forced expiratory volume in one second (FEV1). Of the 38 patients who received a daily dose of 64 mg of beta-carotene for 1 week, 20 (53%) were protected against EIA., Conclusions: Our results indicate that a daily dose of Dunaliella beta-carotene exerts a protective effect against EIA in some patients most probably through in vivo antioxidative effect.

We have previously reported the purification of a phosphoprotein (p43) intermediary in steroid synthesis from adrenal zona fasciculata [Paz C., Dada, L. A., Cornejo Maciel, M. F., Mele, P. G., Cymeryng, C. B., Neuman, I., Mendez, C. F., Finkielstein, C. V., Solano, A. R., Park, M., Fischer, W. H., Towbin, H., Scartazzini, R. & Podestá, E. J. (1994) Eur J. Biochem. 224, 709-716]. Here, we describe the cloning and sequencing of a cDNA encoding p43 as well as the hormonal regulation of the p43 transcript. The protein resulted homologous to a very recently described mitochondrial peroxisome-proliferator-induced very-long-chain acyl-CoA thioesterase (MTE-I). The deduced amino acid sequence of the protein shows consensus sites for phosphorylation by different protein kinases, and a lipase serine motif. Antibodies raised against a synthetic peptide that includes the lipase serine motif and against the N-terminal region of p43 block the action of the protein. The transcript of p43 was detected in ovary of pseudopregnant rats, rat adrenal zona fasciculata and glomerulosa, mouse Leydig tumor cell line (MA-10), rat brain and human placenta. Inhibition of adrenocorticotropin hormone (ACTH) release and steroid synthesis by dexamethasone produced a dose-dependent decrease in the abundance of the adrenal transcript. The transcript was induced by in vivo stimulation of the adrenals with ACTH. The effect had a rapid onset (5 min), reached maximal stimulation (62%) at 15 min, and returned to basal levels at 30 min. The effect of ACTH on the p43 transcript was inhibited by actinomycin D and enhanced by cycloheximide. Our results provide the first evidence linking acyl-CoA thioesterases with very-long-chain specificities, and a protein intermediary in steroid synthesis, thereby supporting a regulatory role for acyl-CoA thioesterases in steroidogenic tissues.

Background: Allergic rhinitis and nasal polyposis are common nasal diseases, but the available treatment modalities have only limited success., Objective: To assess the therapeutic effect of low-energy narrow-band red light phototherapy on nasal clinical symptoms of allergic rhinitis and nasal polyposis., Methods: In a double-blind randomized prospective study, 50 patients with allergic rhinitis and 10 with nasal polyposis received intranasal illumination at 660 nm for 4.4 minutes three times a day for 14 days (total dose 6 joules per day). Twenty-nine rhinitic patients and one patient with polyposis received equivalent sham illumination as placebo. Evaluation was based on symptom scores and a clinical assessment that included pre-treatment and post-treatment videotaped rigid and flexible nasendoscopy., Results: Following treatment, improvement of symptoms was reported by 72% of the allergic rhinitis patients and objective improvement was endoscopically demonstrated in 70% of them as compared with 24% and 3%, respectively, in the placebo group. These differences were significant. No improvement was obtained in any of the patients with polyposis., Conclusions: Allergic rhinitis, if uncomplicated by polyps or chronic sinusitis, can be effectively treated by narrow-band red light illumination of the nasal mucosa at 660 nm, with marked alleviation of clinical symptoms. Whenever possible, candidates for phototherapy should be selected by endoscopic examination.

Objective: To determine if vitamin C (ascorbic acid) has a protective effect on the hyperreactive airways of patients with exercise-induced asthma (EIA)., Design: All the patients underwent pulmonary function tests at rest, before and 1 hour after receiving 2 g of oral ascorbic acid. They were then randomly assigned in a double-blind manner to receive 2 g of ascorbic acid or a placebo 1 hour before a 7-minute exercise session on a treadmill. Pulmonary function tests were performed after an 8-minute rest. This procedure was repeated 1 week later, with each patient receiving the alternative medication., Setting: A university hospital., Participants: Twenty patients with asthma (13 males and 7 females), with ages ranging from 7 to 28 years (mean, 13.8 years). All patients who had a decline of at least 15% in their forced expiratory volume in 1 second after a standard exercise test on a motorized treadmill received a diagnosis of EIA. MAIN-OUTCOME MEASURES: All patients were advised to stop using their regular asthma medication or bronchodilator 12 hours before the test. Pulmonary function tests were performed in the same ambient conditions on all patients., Results: All patients received a diagnosis of EIA. Ascorbic acid administration did not change the results of pulmonary functions at rest after 1 hour. In 9 patients, a protective effect on exercise-induced hyperreactive airways was documented. Four of 5 patients who received ascorbic acid and documented a protective effect on EIA continued to receive ascorbic acid, 0.5 g/d, for 2 more weeks with the same protective effect., Conclusions: The efficacy of vitamin C in preventing EIA cannot be predicted. However, vitamin C may have a protective effect on airway hyperreactivity in some patients with EIA.

Evidence has been introduced linking the lipoxygenase products and steroidogenesis in Leydig cells, thereby supporting that this pathway may be a common event in the hormonal control of steroid synthesis. On the other hand, it has also been reported that lipoxygenase products of arachidonic acid (AA) may not be involved in Leydig cells steroidogenesis. In this paper, we investigated the effects of PLA2 and lipoxygenase pathway inhibitors on steroidogenesis in rat testis Leydig cells. The effects of two structurally unrelated PLA2 inhibitors (4-bromophenacyl bromide (BPB) and quinacrine) were determined. BPB blocked the LH- and Bt2cAMP-stimulated testosterone production but had no effect on 22(4)-OH-cholesterol conversion to testosterone. Quinacrine caused a dose-dependent inhibition of LH- and Bt2cAMP-induced steroidogenesis. The effects of different lipoxygenase pathway inhibitors (nordihydroguaiaretic acid (NDGA), 5,8,11,14-eicosatetraynoic acid (ETYA), caffeic acid and esculetin) have also been determined. Both NDGA and ETYA inhibited LH- and Bt2cAMP-stimulated steroid synthesis in a dose-related manner. Furthermore caffeic acid and esculetin also blocked the LH-stimulated testosterone production. Moreover, exogenous AA induced a dose-dependent increase of testosterone secretion which was inhibited by NDGA. Our results strongly support the previous concept that the lipoxygenase pathway is involved in the mechanism of action of LH on testis Leydig cells.

We have previously isolated and partially-sequenced a soluble phosphoprotein (p43) that acts as intermediary in the stimulation of steroid synthesis. In this report we have used synthetic peptides whose sequences match those obtained from p43 to generate antipeptide antibodies and show that these antibodies bind to purified p43 protein as determined by immunoblot analysis. The presence of p43 was detected by Western blot in both steroidogenic and non-steroidogenic tissues. One of the antibodies was also used to purify p43 on immunoaffinity chromatography columns. Proteins eluting from affinity columns produce a twelve-fold stimulation of progesterone synthesis. This effect was blocked by the use of an inhibitor of phospholipase A2. These results suggest the involvement of p43 in transducing the adrenocorticotropin signal to mitochondria in zona fasciculata cells. We also describe a partial cDNA clone with a predicted amino acid sequence that matches the sequences of the internal peptides of p43.

It is well accepted that protein(s) with a short half-life are required in the pathway leading to steroid synthesis following stimulation by trophic hormones. A correlation between the disappearance of several proteins in different subcellular compartments and the inhibition of steroid synthesis produced by cycloheximide (CHx) has also been shown. In the present report we describe the effect of CHx in the stimulation of steroid synthesis using a cell-free assay. Mitochondrial progesterone (P4) production was studied by recombination of the different subcellular fractions of adrenal zona fasciculata and determined by radioimmunoassay. Soluble factors from ACTH-treated adrenals produced a four-fold stimulation of mitochondrial steroidogenesis (3.0 +/- 0.6 vs. 13.3 +/- 0.5 ng P4/tube for control and ACTH-treated adrenals respectively). Mitochondria obtained from CHx-ACTH-treated adrenals fail to respond to soluble ACTH-dependent factors. A permeable analogue of cholesterol (22(R)-OH cholesterol) could overcome the inhibition imposed by CHx, confirming the role of mitochondrial proteins in intramitochondrial cholesterol transport. The treatment of the adrenals with CHx 10 minutes before ACTH administration abolished also the stimulation induced by the cytosol on control mitochondria (2.6 +/- 0.5 vs. 13.0 +/- 1.0 ng P4/tube for CHx-ACTH-treated cytosol vs. ACTH-treated cytosol). Arachidonic acid (AA) added to CHx-ACTH-treated cytosol subdued this inhibition (10.3 +/- 1.2 ng P4/tube). CHx treatment had no effect on the stimulation by ACTH of the cAMP-dependent protein kinase. These results indicate the involvement of a cycloheximide-sensitive protein in the release of AA in adrenal steroidogenesis.

In previous reports we have demonstrated the presence of a soluble factor that responds to cAMP signals to induce steroid synthesis in adrenocortical tissue. Here, we describe the purification of this factor from adrenal zona fasciculata cells by using a five-step procedure that includes DEAE-cellulose, gel filtration, Mono Q HPLC and Superose HPLC, and elution of the protein from SDS/PAGE. This procedure results in the purification to homogeneity of a protein of 43-kDa that retains the capacity to stimulate steroid synthesis in an in vitro recombination assay. This activity is inhibited by the use of phospholipase A2 inhibitors. Antipeptide antibodies against the N-terminal region recognize p43 as a double band on SDS/PAGE that resolves in different spots on two-dimensional gel electrophoresis. Adrenocorticotropin treatment of adrenal glands results in the appearance of multiple spots that migrated towards a lower pH compared to controls, suggesting the presence of phosphorylated and dephosphorylated forms of p43. Sequencing of the N-terminal region and internal peptides reveals no significant similarities with other proteins, suggesting that p43 is a novel protein. We conclude from our data that the isolated protein (p43) is a novel, soluble protein that acts as intermediary in adrenocorticotropin-induced stimulation of arachidonic acid release and steroid synthesis.

At the First Medical Clinic of the Faculty Hospital Prague Motol the authors detected during a routine gastroscopic examination of a 71-year-old female patient the presence of a gastric bezoar. The examination was indicated on account of dyspeptic complaints. With regard to the size of the bezoar (50 x 20 mm) the authors decided for mechanical lithotripsy. It proved possible to reduce the concrement during the first session and during subsequent sessions it was broken down to small size and the fragments were eliminated per vias naturales. Several fragments were extracted. Chemical analysis provided evidence of the origin of the concrement in the biliary pathways--the concrement was pure cholesterol. ERCP did not reveal an artificial communication between the biliary system and the stomach or duodenum. It did not prove possible to visualize the gallbladder by ultrasonography. The selected therapeutic procedure was successful.

1. Exposure of catfish, Ictalurus nebulosus, to sublethal concentrations of cadmium deteriorates electro-orientation performance. 2. Cadmium, at a concentration of 40 micrograms/l, doubles the behavioural threshold for electric stimuli within 48 hr of exposure; both prolonged exposure and higher concentrations result in higher thresholds. The effect is reversible. 3. Electro-orientation performance can be used to monitor the quality of surface water.

Luteinizing hormone (LH) and human chorionic gonadotrophin (hCG) receptors are coupled to intracellular effector systems, most notably adenylate cyclase, through guanyl nucleotide-binding proteins or G-proteins. The molecular mechanism involved in the dynamic coupling of the LH/hCG receptor however, are not known. It has been postulated that receptor aggregation at the molecular level plays a critical role in this process. There have been attempts to understand the receptor association and dissociation phenomena at the molecular level. One of them involves the participation of the major histocompatibility complex (MHC) class I antigen in the mechanism of receptor activation and/or expression. One molecular basis for these mechanisms consists of a physical interaction between MHC proteins and receptors to form "compound receptors" able to transfer a hormonal signal to the cell. Using a photo-reactive probe we demonstrated that the LH/hCG receptors and the class I antigens are closely associated in the membrane. Thus, it is possible to form covalent complexes of hCG and class I antigens through the binding of the hormone to specific receptors. These findings imply that LH/hCG receptors and the MHC class I antigens may interact at the level of the plasma membrane in the mechanism of LH action. We also performed experiments using a single cell and limiting stimulation to a patch of membrane. The results stimulating the cell in a localized area suggested that even if all components are entirely free to float there is a constraint in the localization of the receptor, G-protein, and/or the effector, supporting the constraint dissociation model. Within a limited area subunits could dissociate, but they would not be free to diffuse throughout the membrane. Moreover the concept of compartmentalization that has been utilized to explain some inconsistencies in second-messenger action now can be proved by experimental design.

Among factors which possibly influence the responses of asthmatic children to exercise, climate has received little attention. This study was performed to determine whether the level of air humidity is a factor to be considered. Twenty asthmatic (extrinsic perennial) girls and boys, 6 to 14 yr of age, with unverified history of exercise-induced asthma (EIA) took part. They rested and exercised in a climatic chamber in dry (25% relative humidity) and humid (90%) sessions at 25 degrees to 26 degrees C. One to three weeks separated the sessions, the order of which was counterbalanced. No changes in pulmonary functions (FVC, FEV 1.0, MMEF, MBC) were found following a sitting period of 60 min in either climate. Five and ten minutes following the treadmill run, however, bronchoconstriction was distinctly more pronounced in the dry than in the humid climate. Exercise heart rate and the subjective rating of effort were not affected by climate. It was concluded that, under the above experimental conditions, EIA is more likely in dry air than in humid air, possibly due to heat loss at the airway mucosa caused by evaporation. High humidity of inspired air could be the reason why EIA is less prevalent in swimming, as compared with other modes of exercise.

Seventeen asthmatic children, nine with and eight without exercise-induced asthma (EIA), and nine control non-asthmatic children were studied in an attempt to discern possible associations between the activity of the kinin system and EIA. Pulmonary function tests and clinical check-up were performed before and after 6 min of free-range running. Concomitant blood tests revealed a consistent elevation of the kallikrein levels following the exercise challenge in all experimental groups. However, only in the EIA positive group did this elevation exceed the normal laboratory range (16 +/- 7.0 mu/ml). Prekallikrein levels both before and after exercise did not exceed in any group the normal laboratory values. The findings thus suggest that provocation of EIA is associated with a certain threshold of kallikrein level below which no such symptoms are observed. EIA may be triggered only when kallikrein levels are in excess.

Astemizole is a nonsedative H1-antagonist. It was used in a randomized double-blind, placebo-controlled in-season trial to assess its efficacy in patients with perennial allergic rhinitis showing seasonal exacerbations. By the last (6th) week of the study, the mean overall symptomatology, as rated by a global composite score, was significantly mitigated in the astemizole-treated group (18 patients) compared with their first week's global composite score (P less than .01). No such improvement was observed in the placebo-treated group (18 patients). Side effects were not appreciable throughout this short-term study. We conclude that astemizole is effective in the treatment of patients with perennial allergic rhinitis showing seasonal enhancement of symptoms.

Recent studies have shown the relevance of air humidity to the provocation of bronchoconstriction by running. The present study was undertaken to ascertain whether the humid air breathed during swimming could explain the protective effect of swimming on the asthmatic. Nine asthmatic children 9--15 years old swam while inspiring dry (15--35% R.H.) or humid (80--90% R.H.) air administered in a random order, a week separating the two sessions. The exercise challenge was an 8-min tethered swim at a metabolic rate (VO2) of 29 ml.kg-1.min-1, minute ventilation (VE) of 34 L.min-1, and a heart rate (HR) of 161 beats.min-1. Ambient air and water temperature were 28 +/- 2 degrees C and 27 +/- 2 degrees C, respectively. Pulmonary functions were tested pre and post swimming. Exercise VE, VO2 and HR were similar under the two conditions. No reduction in any of the pulmonary functions (FVC,FEV1.0,MMEFR,MBC) was found after 5 and 10 minutes following the swimming exercise in either of the conditions. In contrast, a treadmill run of similar metabolic and ventilatory intensity induced bronchoconstriction when room air was dried to 25--30% R.H. It is suggested that, unlike running, swimming is of low asthmogenicity even when inspired air is dried to 25--30% at neutral temperatures.

Thirteen children each exercised for 6 min by running on a treadmill and by tethered swimming, breathing air at room temperature and either 8% or 99% relative humidity continuously. Ventilation, gas exchange and heart rate were closely matched in all four tests in each child, with a mean oxygen consumption of 32.3 +/- 1.7 ml x min-1 x kg-1. The post-exercise fall in FEV1 expressed as a percentage of the baseline FEV1 (delta FEV1) was significantly greater after running compared with swimming breathing either humid or dry air. The delta FEV1 was also related to respiratory heat loss (RHL) calculated from measurements of inspired and expired gas temperature and humidity. At a standardised RHL, the difference between running and swimming was highly significant [delta FEV1 (%) +/- SE = 39 +/- 5 and 28 +/- 4 respectively, p less than 0.01]. These experiments suggest that the type of exercise influences the severity of exercise-induced asthma even under conditions of the same metabolic stress and respiratory heat loss.