Anoctamin-1 Cl(-) channels in nociception: activation by an N-aroylaminothiazole and capsaicin and inhibition by T16A[inh]-A01.

BACKGROUND: Anoctamin 1 (ANO1 or TMEM16A) Ca(2+)-gated Cl(-) channels of nociceptor neurons are emerging as important molecular components of peripheral pain transduction. At physiological intracellular Cl(-) concentrations ([Cl(-)]i) sensory neuronal Cl(-) channels are excitatory. The ability of sensory neuronal ANO1 to trigger action potentials and subsequent nocifensive (pain) responses were examined by direct activation with an N-aroylaminothiazole. ANO1 channels are also activated by intracellular Ca(2+) ([Ca(2+)]i) from sensory neuronal TRPV1 (transient-receptor-potential vallinoid 1) ion channels and other noxicant receptors. Thus, sensory neuronal ANO1 can facilitate TRPV1 triggering of action potentials, resulting in enhanced nociception. This was investigated by reducing ANO1 facilitation of TRPV1 effects with: (1) T16A[inh]-A01 ANO1-inhibitor reagent at physiological [Cl(-)]i and (2) by lowering sensory neuronal [Cl(-)]i to switch ANO1 to be inhibitory. RESULTS: ANO1 effects on action potential firing of mouse dorsal root ganglia (DRG) neurons in vitro and mouse nocifensive behaviors in vivo were examined with an N-aroylaminothiazole ANO1-activator (E-act), a TRPV1-activator (capsaicin) and an ANO1-inhibitor (T16A[inh]-A01). At physiological [Cl(-)]i (40 mM), E-act (10 µM) increased current sizes (in voltage-clamp) and action potential firing (in current-clamp) recorded in DRG neurons using whole-cell electrophysiology. To not disrupt TRPV1 carried-Ca(2+) activation of ANO1 in DRG neurons, ANO1 modulation of capsaicin-induced action potentials was measured by perforated-patch (Amphotericin-B) current-clamp technique. Subsequently, at physiological [Cl(-)]i, capsaicin (15 µM)-induced action potential firing was diminished by co-application with T16A[inh]-A01 (20 µM). Under conditions of low [Cl(-)]i (10 mM), ANO1 actions were reversed. Specifically, E-act did not trigger action potentials; however, capsaicin-induced action potential firing was inhibited by co-application of E-act, but was unaffected by co-application of T16A[inh]-A01. Nocifensive responses of mice hind paws were dramatically induced by subcutaneous injections of E-act (5 mM) or capsaicin (50 µM). The nocifensive responses were attenuated by co-injection with T16A[inh]-A01 (1.3 mM). CONCLUSIONS: An ANO1-activator (E-act) induced [Cl(-)]i-dependent sensory neuronal action potentials and mouse nocifensive behaviors; thus, direct ANO1 activation can induce pain perception. ANO1-inhibition attenuated capsaicin-triggering of action potentials and capsaicin-induced nocifensive behaviors. These results indicate ANO1 channels are involved with TRPV1 actions in sensory neurons and inhibition of ANO1 could be a novel means of inducing analgesia.

A sensory neuronal ion channel essential for airway inflammation and hyperreactivity in asthma.

Asthma is an inflammatory disorder caused by airway exposures to allergens and chemical irritants. Studies focusing on immune, smooth muscle, and airway epithelial function revealed many aspects of the disease mechanism of asthma. However, the limited efficacies of immune-directed therapies suggest the involvement of additional mechanisms in asthmatic airway inflammation. TRPA1 is an irritant-sensing ion channel expressed in airway chemosensory nerves. TRPA1-activating stimuli such as cigarette smoke, chlorine, aldehydes, and scents are among the most prevalent triggers of asthma. Endogenous TRPA1 agonists, including reactive oxygen species and lipid peroxidation products, are potent drivers of allergen-induced airway inflammation in asthma. Here, we examined the role of TRPA1 in allergic asthma in the murine ovalbumin model. Strikingly, genetic ablation of TRPA1 inhibited allergen-induced leukocyte infiltration in the airways, reduced cytokine and mucus production, and almost completely abolished airway hyperreactivity to contractile stimuli. This phenotype is recapitulated by treatment of wild-type mice with HC-030031, a TRPA1 antagonist. HC-030031, when administered during airway allergen challenge, inhibited eosinophil infiltration and prevented the development of airway hyperreactivity. Trpa1(-/-) mice displayed deficiencies in chemically and allergen-induced neuropeptide release in the airways, providing a potential explanation for the impaired inflammatory response. Our data suggest that TRPA1 is a key integrator of interactions between the immune and nervous systems in the airways, driving asthmatic airway inflammation following inhaled allergen challenge. TRPA1 may represent a promising pharmacological target for the treatment of asthma and other allergic inflammatory conditions.

TRPA1 is a major oxidant sensor in murine airway sensory neurons.

Sensory neurons in the airways are finely tuned to respond to reactive chemicals threatening airway function and integrity. Nasal trigeminal nerve endings are particularly sensitive to oxidants formed in polluted air and during oxidative stress as well as to chlorine, which is frequently released in industrial and domestic accidents. Oxidant activation of airway neurons induces respiratory depression, nasal obstruction, sneezing, cough, and pain. While normally protective, chemosensory airway reflexes can provoke severe complications in patients affected by inflammatory airway conditions like rhinitis and asthma. Here, we showed that both hypochlorite, the oxidizing mediator of chlorine, and hydrogen peroxide, a reactive oxygen species, activated Ca(2+) influx and membrane currents in an oxidant-sensitive subpopulation of chemosensory neurons. These responses were absent in neurons from mice lacking TRPA1, an ion channel of the transient receptor potential (TRP) gene family. TRPA1 channels were strongly activated by hypochlorite and hydrogen peroxide in primary sensory neurons and heterologous cells. In tests of respiratory function, Trpa1(-/-) mice displayed profound deficiencies in hypochlorite- and hydrogen peroxide-induced respiratory depression as well as decreased oxidant-induced pain behavior. Our results indicate that TRPA1 is an oxidant sensor in sensory neurons, initiating neuronal excitation and subsequent physiological responses in vitro and in vivo.

Transient receptor potential ankyrin 1 antagonists block the noxious effects of toxic industrial isocyanates and tear gases.

The release of methyl isocyanate in Bhopal, India, caused the worst industrial accident in history. Exposures to industrial isocyanates induce lacrimation, pain, airway irritation, and edema. Similar responses are elicited by chemicals used as tear gases. Despite frequent exposures, the biological targets of isocyanates and tear gases in vivo have not been identified, precluding the development of effective countermeasures. We use Ca(2+) imaging and electrophysiology to show that the noxious effects of isocyanates and those of all major tear gas agents are caused by activation of Ca(2+) influx and membrane currents in mustard oil-sensitive sensory neurons. These responses are mediated by transient receptor potential ankyrin 1 (TRPA1), an ion channel serving as a detector for reactive chemicals. In mice, genetic ablation or pharmacological inhibition of TRPA1 dramatically reduces isocyanate- and tear gas-induced nocifensive behavior after both ocular and cutaneous exposures. We conclude that isocyanates and tear gas agents target the same neuronal receptor, TRPA1. Treatment with TRPA1 antagonists may prevent and alleviate chemical irritation of the eyes, skin, and airways and reduce the adverse health effects of exposures to a wide range of toxic noxious chemicals.

Reduced Z-axis coverage multidetector CT angiography for suspected acute pulmonary embolism could decrease dose and maintain diagnostic accuracy.

Multidetector computed tomographic angiography (MDCTA) is the method of choice for evaluation of suspected acute pulmonary embolism (PE) in most patients because it is accurate and widely available. The use of computed tomography, including MDCTA for PE, has risen dramatically over the last several years with an attendant rise in radiation exposure. Many methods currently employed to reduce radiation dose may affect image quality and potentially affect diagnostic accuracy. Reducing Z-axis coverage would decrease radiation dose without any effect on image quality. This study was performed to assess the effect on the accuracy of MDCTA for suspected acute PE if the Z-axis coverage was reduced to the anatomic range from the top of the aortic arch through the heart. Two hundred ninety-five examinations were performed on a 64-detector-row MDCT and interpreted as positive for PE from July 2005 to February 2008. When the anatomic range of these data sets were retrospectively reduced and reinterpreted for PE, no case was interpreted as negative for PE. The Z-axis coverage was reduced by 37%. In the interest of keeping radiation doses as low as reasonably achievable, further research in this area is warranted.

Incorporation of a formalized emergency radiology curriculum to facilitate population of a MIRC-based digital teaching file.

Teaching files are integral to radiological training. Digital Imaging and Communication in Medicine compatible digital radiological data and technological advances have made digital teaching files a desirable way to preserve and share representative and/or unusual cases for training purposes. The Medical Imaging Resource Community (MIRC) system developed by the Radiological Society of North America (RSNA) is a robust multi-platform digital teaching file implementation that is freely available. An emergency radiology training curriculum developed by the American Society of Emergency Radiology (ASER) was incorporated to determine if such an approach might facilitate the entry, maintenance, and cataloguing of interesting cases. The RSNA MIRC software was obtained from the main MIRC website and installed. A coding system was developed based on the outline form of the ASER curriculum. Weekly reports were generated tallying the number of cases in each category of the curriculum. Resident participation in the entry and maintenance of cases markedly increased after incorporation of the ASER curriculum. The coding schema facilitated progress assessment. Ultimately, 454 total cases were entered into the MIRC database, representing at least 42% of the subcategories within the ASER curriculum (161 out of 376). The incorporation of the ASER emergency radiology curriculum greatly facilitated the location, cataloguing, tracking, and maintenance of representative cases and served as an effective means by which to unify the efforts of the department to develop a comprehensive teaching resource within this subspecialty. This approach and format will be extended to other educational curricula in other radiological subspecialties.

Biological activities of a new crotamine-like peptide from Crotalus oreganus helleri on C2C12 and CHO cell lines, and ultrastructural changes on motor endplate and striated muscle.

Crotamine and crotamine-like peptides are non-enzymatic polypeptides, belonging to the family of myotoxins, which are found in high concentration in the venom of the Crotalus genus. Helleramine was isolated and purified from the venom of the Southern Pacific rattlesnake, Crotalus oreganus helleri. This peptide had a similar, but unique, identity to crotamine and crotamine-like proteins isolated from other rattlesnakes species. The variability of crotamine-like protein amino acid sequences may allow different toxic effects on biological targets or optimize the action against the same target of different prey. Helleramine was capable of increasing intracellular Ca2+ in Chinese Hamster Ovary (CHO) cell line. It inhibited cell migration as well as cell viability (IC50 = 11.44 µM) of C2C12, immortalized skeletal myoblasts, in a concentration dependent manner, and promoted early apoptosis and cell death under our experimental conditions. Skeletal muscle harvested from mice 24 h after helleramine injection showed contracted myofibrils and profound vacuolization that enlarged the subsarcolemmal space, along with loss of plasmatic and basal membrane integrity. The effects of helleramine provide further insights and evidence of myotoxic activities of crotamine-like peptides and their possible role in crotalid envenomings.

Cyclophosphamide dose adjustment based on body weight and albuminemia in elderly patients treated with R-mini-CHOP.

BACKGROUND: Diffuse large B-cell lymphoma (DLBCL) is the most common lymphoma in elderly patients, and R-CHOP chemotherapy is the standard treatment protocol for DLBCL. Elderly patients (often defined as 75 years of age) are treated with anticancer drugs with precaution; however, the pharmacokinetics and pharmacodynamics (PK and PD) of these agents have not been thoroughly investigated in this population. In this study, we investigated the PK of cyclophosphamide (CP) and doxorubicin (DOXO) in elderly patients in order to verify if there is an influence of age on the PK of these anticancer drugs. MATERIALS AND METHODS: This is a prospective multi-center clinical trial investigating the PK of CP and DOXO in elderly and very elderly patients with DLBCL treated by R-mini-CHOP regimen. Dose levels were 25 mg/m2, 0.7-1.4 mg/m2, 750 mg/m2, and 375 mg/m2 for DOXO, Vincristine (VCR), CP, and Rituximab, respectively. For PK analysis, 7 time point samples were collected over 48 h post-administration on cycle 3. CP and VCR plasma concentrations were measured using UPLC-MS/MS validated method. DOX plasma concentrations were measured using UPLC coupled with fluorescence detection-validated method. PK-POP modeling has been performed with a non-linear mixed-effect model program (Monolix). RESULTS: 31 patients (15 males and 16 females), 75 to 96 years old, were treated with R-miniCHOP protocol. Among them, 19 patients were treated with VCR. A one-compartment (1cpt) open model with linear elimination adequately described CP concentration-time courses. The population PK parameters for CP were: CL = 3.58 L/h, Vmale = 32.2 L, and Vfemale = 28.7 L. Body weight (BW), albuminemia, and gender demonstrated a significant impact on CP PK. A 2-compartment (2cpt) open model with linear elimination best described DOXO concentration-time courses. The population PK parameters for DOXO obtained for the structural model were: CL = 51.1 L/h, Q = 49.6 L/h, V1 = 29.4 L, V2 = 1,130 L (clearances: CL, Q, volumes of distribution: V1, V2). The main covariate effects on DOXO PK were related to gender, BW, and VCR administration. VCR increases DOXO V1 from 29.4 L to 57.5 L (p = 0.02). No hematologic and cardiac grade 3 or 4 toxicity were recorded. CONCLUSIONS: Usually, in the absence of specific data, the majority of the physicians empirically reduce anticancer drug dose in the elderly patients (Tourani in J Geriatr Oncol 3(1): 41-48, 2012), or even does not treat these very-old patients. A better knowledge of the pharmacokinetics in very-old patients should allow a better dose adjustment based on the most significant physiological factors that modify the pharmacokinetic parameters. In this study, no serious toxicity was observed in these very elderly patients (84.1 years). This indicates that dose adjustment of chemotherapies should not only be based on age and creatinine clearance, but also, based upon appropriate physiological and biological data. Our findings indicate that, CP dose adjustment should be done according to serum albumin levels and patients BW and gender.

Testicular cooling associated with testicular torsion and its detection by infrared thermography: an experimental study in sheep.

PURPOSE: We determined whether experimental testicular torsion results in gonadal cooling and whether testicular temperature changes can be detected by infrared thermography. MATERIALS AND METHODS: A nonblinded, randomized, controlled trial was done in 6 anesthetized sheep. Thermocouple probes recorded testicular temperature every 15 minutes for 6 hours after experimental side 720-degree medial testicular torsion with orchiopexy or control side sham procedure with orchiopexy and for 75 minutes after procedure reduction. Color Duplex ultrasound was done to control the experimental assignment. Mean hemiscrotal infrared thermography temperatures were calculated and nonparametric repeated measures analysis was performed to determine whether there were significant changes in temperature as a function of the experimental condition and time. RESULTS: Testicular torsion resulted in significant testicular cooling by probe and infrared thermography (p <0.05 and <0.0001, respectively), which was promptly reversed upon the reduction of experimental torsion. Two hours after experimental torsion the median temperature difference (control side minus torsion side) was 2.5C for the probe and 1.7C for infrared thermography. CONCLUSIONS: Experimental testicular torsion resulted in significant gonadal cooling that was detectable by infrared thermography of the hemiscrotum. The applicability of these findings to the clinical setting remains to be determined.

Feasibility of using near-infrared spectroscopy to diagnose testicular torsion: an experimental study in sheep.

STUDY OBJECTIVE: To assess whether near-infrared spectroscopy can detect testicular hypoxia in a sheep model of testicular torsion within 6 hours of experimental torsion. METHODS: This was a randomized, controlled, nonblinded study. Trans-scrotal, near-infrared, spectroscopy-derived testicular tissue saturation of oxygen values were obtained from the posterior hemiscrota of 6 anesthetized sheep at baseline and every 15 minutes for 6 hours after either experimental-side, 720-degree, unilateral, medial testicular torsion and orchidopexy or control-side sham procedure with orchidopexy and then for 75 minutes after reduction of torsion and pexy. Color Doppler ultrasonography was performed every 30 minutes to confirm loss of vascular flow on the experimental side, return of flow after torsion reduction, and preserved flow on the control side. RESULTS: Near infrared spectroscopy detected a prompt, sustained reduction in testicular tissue saturation of oxygen after experimental torsion. Further, it documented a rapid return of these values to pretorsion levels after reduction of torsion. Experimental-side testicular tissue saturation of oxygen fell from a median value of 59% (interquartile range [IQR] 57% to 69%) at baseline to 14% (IQR 11% to 29%) at 2.5 hours of torsion, and postreduction values were approximately 70%. Control-side testicular tissue saturation of oxygen values increased from a median value of 67% (IQR 59% to 68%) at baseline to 77% (IQR 77% to 94%) at 2.5 hours and remained at approximately 80% for the entire protocol. The difference in median testicular tissue saturation of oxygen between experimental and control sides, using the Friedman test, was found to be significant (P=.017). CONCLUSION: This study demonstrates the feasibility, in a sheep model, of using near-infrared spectroscopy for the noninvasive diagnosis of testicular torsion and for quantification of reperfusion after torsion reduction. The applicability of these findings, from an animal model using complete torsion, to the clinical setting remains to be established.

Complex cystic breast masses: diagnostic approach and imaging-pathologic correlation.

Complex cystic breast masses demonstrate both anechoic (cystic) and echogenic (solid) components at ultrasonography (US). US is used to identify and characterize such masses and to guide percutaneous biopsy. Numerous pathologic entities may produce complex cystic breast lesions or may be associated with them, and biopsy is usually indicated. Common benign findings include fibrocystic changes, intraductal or intracystic papilloma without atypia, and fibroadenoma. Common atypical findings include atypical ductal hyperplasia, atypical papilloma, atypical lobular hyperplasia, and lobular carcinoma in situ. Malignant findings include ductal carcinoma in situ, infiltrating ductal carcinoma, and infiltrating lobular carcinoma. If the biopsy approach is tailored to the individual patient and if the imaging features are closely correlated with findings at pathologic analysis, US-guided percutaneous biopsy may be used effectively to diagnose and to guide management of complex cystic masses.

Early diagnosis of traumatic thoracic aortic rupture by transesophageal echocardiography.

Traumatic aorta rupture survival depends on early diagnosis requiring aortography. Aortography is the "gold standard" method, but it is time-consuming and may be dangerous in trauma patients with multiple organ injuries. Transesophageal echocardiography is a noninvasive technology that can be performed at the bedside. We report two cases in which transesophageal echocardiography enabled us to make the early diagnosis of thoracic descending aorta rupture.

Diagnosis of traumatic mediastinal hematoma with transesophageal echocardiography.

In patients with blunt chest trauma, early diagnosis of mediastinal hematoma is important, because it could be associated with thoracic vessel injury. Mediastinal hematoma is generally evoked because of a widened mediastinum on chest radiograph, but radiologic diagnosis may lead to excessive angiography being performed. Transesophageal echocardiography (TEE) provides accurate views of the mediastinum and can be rapidly performed at the bedside. Thus, we conducted a prospective study to define TEE signs of mediastinal hematoma. TEE was performed in 22 thoracic trauma patients (trauma group) and in 20 brain-dead patients without thoracic trauma (control group). The positive diagnosis of mediastinal hematoma was made using thoracic surgery or computed tomographic scan. The specificity of TEE was 75 percent and sensitivity was 100 percent. In the trauma group, there was only one false positive but angiography discovered a traumatic aneurysm of the proximal right subclavian artery. No false negative was noted. We described three different TEE signs of mediastinal hematoma: (1) an increased distance between the probe and the aortic wall; (2) a double contour of the aortic wall; and (3) visualization of the ultrasound signal between the aortic wall and the visceral pleura. The distance between the esophageal probe and the aortic wall was the most accurate sign because it could be easily obtained; the threshold value for this distance was 3 mm. TEE appears to be an accurate method to diagnose traumatic mediastinal hematoma.

Sensory detection and responses to toxic gases: mechanisms, health effects, and countermeasures.

The inhalation of reactive gases and vapors can lead to severe damage of the airways and lung, compromising the function of the respiratory system. Exposures to oxidizing, electrophilic, acidic, or basic gases frequently occur in occupational and ambient environments. Corrosive gases and vapors such as chlorine, phosgene, and chloropicrin were used as warfare agents and in terrorist acts. Chemical airway exposures are detected by the olfactory, gustatory, and nociceptive sensory systems that initiate protective physiological and behavioral responses. This review focuses on the role of airway nociceptive sensory neurons in chemical sensing and discusses the recent discovery of neuronal receptors for reactive chemicals. Using physiological, imaging, and genetic approaches, Transient Receptor Potential (TRP) ion channels in sensory neurons were shown to respond to a wide range of noxious chemical stimuli, initiating pain, respiratory depression, cough, glandular secretions, and other protective responses. TRPA1, a TRP ion channel expressed in chemosensory C-fibers, is activated by almost all oxidizing and electrophilic chemicals, including chlorine, acrolein, tear gas agents, and methyl isocyanate, the highly noxious chemical released in the Bhopal disaster. Chemicals likely activate TRPA1 through covalent protein modification. Animal studies using TRPA1 antagonists or TRPA1-deficient mice confirmed the role of TRPA1 in chemically induced respiratory reflexes, pain, and inflammation in vivo. New research shows that sensory neurons are not merely passive sensors of chemical exposures. Sensory channels such as TRPA1 are essential for maintenance of airway inflammation in asthma and may contribute to the progression of airway injury following high-level chemical exposures.