Reliability of multiple frequency bioelectrical impedance analysis: An intermachine comparison

The technical reliability (i.e., interinstrument and interoperator reliability) of three SEAC-swept frequency bioimpedance monitors was assessed for both errors of measurement and associated analyses. In addition, intraoperator and intrainstrument variability was evaluated for repeat measures over a 4-hour period. The measured impedance values from a range of resistance-capacitance circuits were accurate to within 3% of theoretical values over a range of 50-800 ohms. Similarly, phase was measured over the range 1 degrees-19 degrees with a maximum deviation of 1.3 degrees from the theoretical value. The extrapolated impedance at zero frequency was equally well determined (+/-3%). However, the accuracy of the extrapolated value at infinite frequency was decreased, particularly at impedances below 50 ohms (approaching the lower limit of the measurement range of the instrument). The interinstrument/operator variation for whole body measurements were recorded on human volunteers with biases of less than +/-1% for measured impedance values and less than 3% for phase. The variation in the extrapolated values of impedance at zero and infinite frequencies included variations due to operator choice of the analysis parameters but was still less than +/-0.5%. (C) 1997 Wiley-Liss, Inc.

Changes in purines concentration in the cerebrospinal fluid of patients experiencing pain: A case-control study

This study analyzes the relationship between extracellular purines and pain perception in humans. Cerebrospinal fluid (CSF) levels of purines and their metabolites were compared between patients displaying acute and/or chronic pain syndromes and control subjects. The CSF levels of IMP, inosine, guanosine and uric acid were significantly increased in the chronic pain group and correlated with pain severity (P<0.05). Patients displaying both chronic and acute pain presented similar changes in the CSF purines concentration (P<0.05). However, in the acute pain group, only CSF inosine and uric acid levels were significantly increased (P<0.05). These findings suggest that purines, in special inosine, guanosine and uric acid, are associated with the spinal mechanisms underlying nociception. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Ratio of Metalloproteinase 2 to Tissue Inhibitor of Metalloproteinase 2 in Medullary Thyroid Carcinoma

Objectives: To develop an index for the ratio of metalloproteinase 2 (MMP-2) to its tissue inhibitor (TIMP-2) in immunostained medullary thyroid carcinoma specimens and to correlate it with clinical and pathologic prognostic factors. Metalloproteinases, enzymes related to the degradation of the extracellular matrix, take part in carcinogenesis and have been associated with the prognosis of neoplasias. Nevertheless, medullary carcinoma is rarely considered in research analysis. Researchers tend to favor the ratio of enzymes to their inhibitors over the absolute concentrations of these enzymes. Design: Retrospective study of surgical samples. Setting: Head and Neck Surgery and Endocrinology Departments, Universidade de Sao Paulo Medical School Hospital. Patients: Surgical specimens from 33 patients who had been observed for a mean of 76.8 months (range, 4-201 months) were immunohistochemically stained for MMP-2 and TIMP-2. Only patients whose clinical and pathologic data were complete and whose specimens were preserved were included in the study. Main Outcome Measures: The ratio between the expressions of MMP-2 and TIMP-2 was based on a staining index (immunostaining extent and intensity) of each of the markers. Results: Proportionally large expressions of TIMP-2 over MMP-2 correlated with low occurrences of positive findings on initial cervical examination for the presence of thyroid nodules and/or lymphadenopathy (P = .02) and cervical lymph node metastases (P < .001), conditions correlated with prognosis. A correlation with cure at the end of follow-up (P = .01) was also observed. (P < .05 was considered statistically significant.) Conclusion: The ratio of MMP-2 to TIMP-2 expression is an additional and novel prognostic predictor of the outcome of medullary carcinoma treated surgically.

Odorant-induced currents in intact patches from rat olfactory receptor neurons: Theory and experiment

Odorant-induced currents in mammalian olfactory receptor neurons have proved difficult to obtain reliably using conventional whole-cell recording. By using a mathematical model of the electrical circuit of the patch and rest-of-cell, we demonstrate how cell-attached patch measurements can be used to quantitatively analyze responses to odorants or a high (100 mM) K+ solution. High K+ induced an immediate current flux from cell to pipette, which was modeled as a depolarization of similar to 52 mV, close to that expected from the Nernst equation (56 mV), and no change in the patch conductance. By contrast, a cocktail of cAMP-stimulating odorants induced a current flux from pipette into cell following a significant (4-10 s) delay. This was modeled as an average patch conductance increase of 36 pS and a depolarization of 13 mV, Odorant-induced single channels had a conductance of 16 pS. In cells bathed with no Mg2+ and 0.25 mM Ca2+, odorants induced a current flow from cell to pipette, which was modeled as a patch conductance increase of similar to 115 pS and depolarization of similar to 32 mV, All these results are consistent with cAMP-gated cation channels dominating the odorant response, This approach, which provides useful estimates of odorant-induced voltage and conductance changes, is applicable to similar measurements in any small cells.

The glycine receptor

The inhibitory glycine receptor (GlyR) is a member of the ligand-gated ion channel receptor superfamily. The GlyR comprises a pentameric complex that forms a chloride-selective transmembrane channel, which is predominantly expressed in the spinal cord and brain stem. We review the pharmacological and physiological properties of the GlyR and relate this information to more recent insights that have been obtained through the cloning and recombinant expression of the GlyR subunits. We also discuss insights into our understanding of GlyR structure and function that have been obtained by the genetic characterisation of various heritable disorders of glycinergic neurotransmission. (C) 1997 Elsevier Science Inc.

Solution structure of the sodium channel antagonist conotoxin GS: A new molecular caliper for probing sodium channel geometry

Background: The venoms of Conus snails contain small, disulfide-rich inhibitors of voltage-dependent sodium channels. Conotoxin GS is a 34-residue polypeptide isolated from Conus geographus that interacts with the extracellular entrance of skeletal muscle sodium channels to prevent sodium ion conduction. Although conotoxin GS binds competitively with mu conotoxin GIIIA to the sodium channel surface, the two toxin types have little sequence identity with one another, and conotoxin GS has a four-loop structural framework rather than the characteristic three-loop mu-conotoxin framework. The structural study of conotoxin GS will form the basis for establishing a structure-activity relationship and understanding its interaction with the pore region of sodium channels. Results: The three-dimensional structure of conotoxin GS was determined using two-dimensional NMR spectroscopy. The protein exhibits a compact fold incorporating a beta hairpin and several turns. An unusual feature of conotoxin GS is the exceptionally high proportion (100%) of cis-imide bond geometry for the three proline or hydroxyproline residues. The structure of conotoxin GS bears little resemblance to the three-loop mu conotoxins, consistent with the low sequence identity between the two toxin types and their different structural framework. However, the tertiary structure and cystine-knot motif formed by the three disulfide bonds is similar to that present in several other polypeptide ion channel inhibitors. Conclusions: This is the first three-dimensional structure of a 'four-loop' sodium channel inhibitor, and it represents a valuable new structural probe for the pore region of voltage-dependent sodium channels. The distribution of amino acid sidechains in the structure creates several polar and charged patches, and comparison with the mu conotoxins provides a basis for determining the binding surface of the conotoxin GS polypeptide.

Hypertonic saline reduces metalloproteinase expression in liver during pancreatitis

P>We recently demonstrated that hypertonic saline reduces inflammation and mortality in acute pancreatitis. The present study investigated the effects of hypertonic saline in metalloproteinase (MMP) regulation and pancreatitis-associated hepatic injury. Wistar rats were divided into four groups: (i) control, not subjected to insult or treatment; (ii) no treatment (NT), induction of pancreatitis (retrograde infusion of 2.5% sodium taurocholate (1.0 mL/kg)), but no further treatment; (iii) normal saline (NS), induction of pancreatitis and treatment with normal saline (0.9% NaCl, 34 mL/kg, i.v. bolus, 1 h after the induction of pancreatitis); and (iv) hypertonic saline (HS), induction of pancreatitis and treatment with hypertonic saline (7.5% NaCl, 4 mL/kg administered over a period of 5 min, 1 h after the induction of pancreatitis). In all four groups, 4, 12 and 24 h after the induction of pancreatitis, liver tissue samples were assayed to determine levels of MMP-2, MMP-9, 47 kDa heat shock protein (HSP47) and collagen (Type I and III). Compared with the control group, MMP-9 expression and activity was increased twofold in the NS and NT groups 4 and 12 h after the induction of pancreatitis, but remained at basal levels in the HS group. In contrast, MMP-2 expression was increased twofold 12 h after the induction of pancreatitis only in the NS group, whereas the expression of HSP47 was increased 4 h after the induction of pancreatitis in the NS and NT groups. Greater extracellular matrix remodelling occurred in the NS and NT groups compared with the HS group, probably as a result of the hepatic wound-healing response to repeated injury. However, the collagen content in hepatic tissue remained at basal levels in the HS group. In conclusion, the results of the present study indicate that hypertonic saline is hepatoprotective and reduces hepatic remodelling, maintaining the integrity of the hepatic extracellular matrix during pancreatitis. Hypertonic saline-mediated regulation of MMP expression may have clinical relevance in pancreatitis-associated liver injury.

Collagen is reduced and disrupted in human aneurysms and dissections of ascending aorta

In ascending aorta aneurysms, there is an enlargement of the whole vessel, whereas aortic dissections (ADs) are characterized by the cleavage of the wall into 2 sheets at the external half. We searched if alterations in collagen could be related to these diseases. Sections of aortas from 14 case patients with acute dissections, 10 case patients with aneurysms, and 9 control subjects were stained with picrosirius. Slides were analyzed under polarized microscopy to evaluate the structure of collagen fibers. The proportion of collagen was calculated in each half of the medial layer by color detection in a computerized image analysis system. Collagen appearance under polarized light was consistent with collagenolysis. The mean collagen proportions at the inner and outer halves, respectively, were 0.50 +/- 0.13 and 0.40 +/- 0.08 in the control group, 0.20 +/- 0.10 and 0.18 +/- 0.12 in the AD group, and 0.33 +/- 0.12 and 0.19 +/- 0.12 in the aneurysm group. The AD (P < .01) and control (P = .04) groups had less collagen at the external half, no difference was found in the aneurysm group (P = .71). In both halves, there was less collagen in the case patients than in the control subjects (all P < .01), but at the internal half, the decrease was significantly greater in the case patients with aneurysms than in those with dissections (P = .03; at the external half, P = .99). Aortic dissections and aneurysms show a decrease in collagen content that could be related to a weakness of the wall underlying the diseases, but the locations of the decrease differ: in dissections, it is situated mostly at the external portion of the media (site of cleavage), whereas in aneurysms, it is more diffuse, consistent with the global enlargement. (c) 2008 Elsevier Inc. All rights reserved.

Acute and Chronic Effects of Epicardial Radiofrequency Applications Delivered on Epicardial Coronary Arteries

Background-Epicardial coronary injury is by far the most feared complication of epicardial ablation. Little information is available regarding the chronic effects of delivering radiofrequency in the vicinity of large coronary vessels, and the long-term impact of this approach for mapping and ablation on epicardial vessel integrity is poorly understood. Therefore, the aim of this study was to characterize the acute and chronic histopathologic changes produced by in vivo epicardial pulses of radiofrequency ablation on coronary artery of porcine hearts. Methods and Results-Seven pigs underwent a left thoracotomy. The catheter was sutured adjacent to the left anterior descending artery and left circumflex artery, and 20 pulses of radiofrequency energy were applied. Radiofrequency lesions located no more than 1 mm of the vessel were used for this analysis. Three animals were euthanized 20 days (acute phase) after the procedure and 4 animals after 70 days (chronic phase). The following parameters were obtained in each vessel analyzed: (1) internal and external perimeter; (2) vessel wall thickness; (3) tunica media thickness, and (4) tunica intima thickness. The presence of adipose tissue around the coronary arteries, the distance between the artery and the epicardium, and the anatomic relationship of the artery with the coronary vein was also documented for each section. Sixteen of 20 (80%) sections analyzed, showed intimal thickening with a mean of 0.18 +/- 0.14 mm compared with 0.13 +/- 0.16 mm in the acute phase (P=0.331). The mean tunica media thickness was 0.25 +/- 0.10 mm in the chronic phase animals compared with 0.18 +/- 0.03 mm in the acute phase animals (P=0.021). A clear protective effect of pericardial fat and coronary veins was also present. A positive correlation between depth of radiofrequency lesion and the degree of vessel injury expressed as intimal and media thickening (P=0.001) was present. A negative correlation was identified (r = -0.83; P=0.002) between intimal thickening and distance between epicardium and coronary artery. Conclusions-In this porcine model of in vivo epicardial radiofrequency ablation in proximity to coronary arteries leads to acute and chronic histopathologic changes characterized by tunica intima and media thickening, with replacement of smooth muscle cells with extracellular matrix, but no significant stenosis was observed up to 70 days after the ablation. The absence of acute coronary occlusion or injury does not preclude subsequent significant arterial damage, which frequently occurs when epicardial radiofrequency applications are delivered in close vicinity to the vessels. (Circ Arrhythm Electrophysiol. 2011;4:526-531.)

Electrophysiological characteristics of the Marshall bundle in humans

BACKGROUND Marshall bundles (MBs) are the muscle bundles within the ligament of Marshall. OBJECTIVE This trial sought to the electrophysiological characteristics of the MB and the anatomical connections between MB and left atrium (LA) in patients with persistent atrial fibrillation (AF). METHODS We enrolled 72 patients (male: female 59: 13, age 59.9 +/- 9.4 years) who underwent MB mapping and ablation for AF. MB mapping was done via an endocardial or epicardial approach during sinus rhythm and AF. RESULTS Recordings were successful in 64 of 72 patients (89%). A single connection was noted in 11 of 64 patients between the MB and the coronary sinus (CS) muscle sleeves. The MB recordings showed distinct MB potentials with a proximal-to-distal activation pattern during sinus rhythm. During AF, organized passive activations and dissociated slow MB ectopic activities were commonly observed in this type of connection. Double connections to both CS and LA around left pulmonary veins were noted in 23 of 64 patients (36%). After the ablation of the distal connection, MB recording showed typical double potentials as in single connection. Multiple connections were noted in 30 of 64 patients (47%). During sinus rhythm, the earliest activation was in the middle of the MB. The activation patterns were irregular and variable in each patient. During AF, rapid and fractionated complex activations were noted in all patients of this group. CONCLUSION We documented 3 different types of MB-LA connections. Rapid and fractionated activations were most commonly observed in the MB that had multiple LA connections.

Rapid increases in cytokinin concentration in lateral buds of chickpea (Cicer arietinum L) during release of apical dominance

We examined the role of cytokinins (CKs) in release of apical dominance in lateral buds of chickpea (Cicer arietinum L.). Shoot decapitation or application of CKs (benzyladenine, zeatin or dihydrozeatin) stimulated rapid bud growth. Time-lapse video recording revealed growth initiation within 2 h of application of 200 pmol benzyladenine or within 3 h of decapitation. Endogenous CK content in buds changed little in the first 2 h after shoot decapitation, but significantly increased by 6 h, somewhat later than the initiation of bud growth. The main elevated CK was zeatin riboside, whose content per bud increased 7-fold by 6 h and 25-fold by 24 h. Lesser changes were found in amounts of zeatin and isopentenyl adenine CKs. We have yet to distinguish whether these CKs are imported from the roots via the xylem stream or are synthesised in situ in the buds, but CKs may be part of an endogenous signal involved in lateral bud growth stimulation following shoot decapitation. To our knowledge, this is the first detailed report of CK levels in buds themselves during release of apical dominance.

Coordinated expression of galectin-3 and galectin-3-binding sites in malignant mammary tumors: implications for tumor metastasis

Galectin-3 is a glycan-binding protein that mediates cell-cell and/or cell-extracellular matrix (ECM) interactions. Although galectin-3 is implicated in the progression of various types of cancers, the mechanisms by which galectin-3 enhances metastasis remain unclear. In order to elucidate the role of galectin-3 in the complex multistage process of cancer metastasis, we examined galectin-3 and galectin-3-binding site expression in a series of 82 spontaneous canine mammary tumors (CMT) and two CMT cell lines. Benign CMT tumors exhibited strong nuclear/cytoplasmic galectin-3 immunostaining, whereas malignant CMT tumors and metastases exhibited dramatically decreased galectin-3 expression with the majority of the immunostaining confined to the cytoplasm. Interestingly, intravascular tumor cells overexpressed galectin-3 regardless of their location. CMT-U27 xenografts displayed the same pattern of galectin-3 expression found in spontaneous malignant CMT. In parallel with the downregulation of galectin-3, malignant CMT displayed an overall loss of galectin-3-binding sites in the ECM and focal expression of galectin-3-binding sites mainly detected in intravascular tumor cells and endothelium. Furthermore, loss of galectin-3-binding sites was correlated with the downregulation of GLT25D1, a beta (1-O) galactosyltransferase that modifies collagen, and upregulation of stromal galectin-1. Finally, GLT25D1 mRNA expression was strikingly downregulated in malignant CMT-U27 compared with the benign cell line, and its expression was further de-creased in a galectin-3 knockdown CMT-U27 cell line. We therefore hypothesized that the loss of galectin-3-binding sites in the ECM in conjunction with the overexpression of galectin-3 in specific tumor cell subpopulations are crucial events for the development of mammary tumor metastases.

Prion protein ablation increases cellular aggregation and embolization contributing to mechanisms of metastasis

Cellular Prion Protein (PrP(C)) is a cell surface protein highly expressed in the nervous system, and to a lesser extent in other tissues. PrP(C) binds to the extracellular matrix laminin and vitronectin, to mediate cell adhesion and differentiation. Herein, we investigate how PrP(C) expression modulates the aggressiveness of transformed cells. Mesenchymal embryonic cells (MEC) from wildtype (Prnp(+/+)) and PrP(C)-null (Prnp(0/0)) mice were immortalized and transformed by co-expression of ras and myc. These cells presented similar growth rates and tumor formation in vivo. When injected in the tail vein, PrnP(0/0)raS/myc cells exhibited increased lung colonization compared with Prnp(+/+)ras/myc cells. Additionally, Prnp(0/0)ras/myc cells form more aggregates with blood components than Prnp(+/+)ras/myc cells, facilitating the arrest of Prnp(0/0)ras/myc cells in the lung vasculature. Integrin alpha(v)beta(3) is more expressed and activated in MEC and in transformed Prnp(0/0) cells than in the respective Prnp(+/+) cells. The blocking of integrin alpha(v)beta(3) by RGD peptide reduces lung colonization in transformed Prnp(0/0) cells to similar levels of those presented by transformed Prnp(+/+) cells. Our data indicate that PrP(C) negatively modulates the expression and activation of integrin alpha(v)beta(3) resulting in a more aggressive phenotype. These results indicate that PrP(C) may have main implications in modulating metastasis formation. (C) 2009 UICC

Expression of activated mutants of the human interleukin-3/interleukin-5 granulocyte-macrophage colony-stimulating factor receptor common beta subunit in primary hematopoietic cells induces factor-independent proliferation and differentiation

To date, several activating mutations have been discovered in the common signal-transducing subunit (h beta c) of the receptors for human granulocyte-macrophage colony-stimulating factor, interleukin-3, and interleukin-5. Two of these, Fl Delta and 1374N, result in a 37 amino acid duplication and a single amino acid substitution in the extracellular domain of h beta c, respectively. A third, V449E, results in a single amino acid substitution in the transmembrane domain, Previous studies comparing the activity of these mutants in different hematopoietic cell lines imply that the transmembrane and extracellular mutations act by different mechanisms and suggest the requirement for cell type-specific molecules in signalling. To characterize the ability of these mutant hpc subunits to mediate growth and differentiation of primary cells and hence investigate their oncogenic potential, we have expressed all three mutants in primary murine hematopoietic cells using retroviral transduction. It is shown that, whereas expression of either extracellular hpc mutant confers factor-independent proliferation and differentiation on cells of the neutrophil and monocyte lineages only, expression of the transmembrane mutant does so on these lineages as well as the eosinophil, basophil, megakaryocyte, and erythroid lineages, Factor-independent myeloid precursors expressing the transmembrane mutant display extended proliferation in liquid culture and in some cases yielded immortalized cell lines. (C) 1997 by The American Society of Hematology.