Endothelin-1 Induces Contraction of Female Rat Internal Pudendal and Clitoral Arteries through ET(A) Receptor and Rho-Kinase Activation

Introduction. Endothelin-1 (ET-1), a potent vasoconstrictor peptide, acts mainly through the Gprotein-coupled ET(A) receptor (ET(A)R). Increased vascular ET-1 production and constrictor sensitivity have been observed in various cardiovascular diseases, including hypertension, as well as erectile dysfunction. The internal pudendal artery (IPA) supplies blood to the vagina and clitoris. Inadequate blood flow through the IPA may lead to insufficient vaginal engorgement and clitoral tumescence. Aim. Characterize the effects of ET-1 on the IPA and clitoral artery (CA). Methods. IPA and CA from female Sprague Dawley rats (225-250 g) were mounted in myograph chambers. Arterial segments were submitted to increasing concentrations of ET-1 (10-10-10-6 M). Segments were incubated with the ET(A)R antagonist, atrasentan (10-8 M) or the Rho-kinase inhibitor, Y-27632 (10-6 M) 30 minutes prior to agonist exposure. All E(max) values are expressed as % KCl-induced maximal contraction. ET(A)R, RhoA, and Rho-kinase expression from IPA was evaluated by Western blot. mRNA of preproET-1, ET(A)R, ET(B)R, RhoA, and Rho-kinase were measured by real time PCR. Main Outcome Measures. ET-1 constrictor sensitivity in IPA and CA, protein expression and messenger RNA levels of ET-1-mediated constriction components. Results. ET-1 concentration-dependently contracted IPA (% Contraction and pD2, respectively: 156 +/- 18, 8.2 +/- 0.1) and CA (163 +/- 12, 8.8 +/- 0.08), while ET(A)R antagonism reduced ET-1-mediated contraction (IPA: 104 +/- 23, 6.4 +/- 0.2; CA: 112 +/- 17, 6.6 +/- 0.08). Pretreatment with Y-27632 significantly shifted ET-1 pD2 in IPA (108 +/- 24, 7.9 +/- 0.1) and CA (147 +/- 58 and 8.0 +/- 0.25). Protein expression of ET(A)R, ET(B)R, RhoA, and Rho-kinase were detected in IPA. IPA and CA contained preproET-1, ET(A)R, ET(B)R, RhoA, and Rho-kinase message. Conclusion. We observed that the IPA and CA are sensitive to ET-1, signaling through the ET(A)R and Rho-kinase pathway. These data indicate that ET-1 may play a role in vaginal and clitoral blood flow and may be important in pathologies where ET-1 levels are elevated. Allahdadi KJ, Hannan JL, Tostes RC, and Webb RC. Endothelin-1 induces contraction of female rat internal pudendal and clitoral arteries through ETA receptor and Rho-kinase activation. J Sex Med 2010;7:2096-2103.

Is the Full Version of the AUDIT Really Necessary? Study of the Validity and Internal Construct of Its Abbreviated Versions

Background: This study was aimed at assessing the psychometric qualities of the abbreviated versions of the Alcohol Use Disorders Identification Test (AUDIT-3, AUDIT-4, AUDIT-C, AUDIT-PC, AUDIT-QF, FAST, and Five-Shot) and at comparing them to the 10-item AUDIT and the CAGE in 2 samples of Brazilian adults. Methods: The validity and internal consistency of the scales were assessed in a sample of 530 subjects attended at an emergency department and at a Psychosocial Care Center for Alcohol and Drugs. The Structured Clinical Interview for DSM-IV was used as the diagnostic comparative measure for the predictive validity assessment. The concurrent validity between the scales was analyzed by means of Pearson`s correlation coefficient. Results: The assessment of the predictive validity of the abbreviated versions showed high sensitivity (of 0.78 to 0.96) and specificity (of 0.74 to 0.94) indices, with areas under the curve as elevated as those of the AUDIT (0.89 and 0.92 to screen for abuse and 0.93 and 0.95 in the screening of dependence). The CAGE presented lower indices: 0.81 for abuse and 0.87 for dependence. The analysis of the internal consistency of the AUDIT and its versions exhibited Cronbach`s alpha coefficients between 0.83 and 0.94, while the coefficient for the CAGE was 0.78. Significant correlations were found between the 10-item AUDIT and its versions, ranging from 0.91 to 0.99. Again, the results for the CAGE were satisfactory (0.77), although inferior to the other instruments. Conclusions: The results obtained in this study confirm the validity of the abbreviated versions of the AUDIT for the screening of alcohol use disorders and show that their psychometric properties are as satisfactory as those of the 10-item AUDIT and the CAGE.

Multiplex semi-nested RT-PCR with exogenous internal control for simultaneous detection of bovine coronavirus and group A rotavirus

Neonatal calf diarrhea is a multi-etiology syndrome of cattle and direct detection of the two major agents of the syndrome, group A rotavirus and Bovine coronavirus (BCoV) is hampered by their fastidious growth in cell culture. This study aimed at developing a multiplex semi-nested RT-PCR for simultaneous detection of BCoV (N gene) and group A rotavirus (VP1 gene) with the addition of an internal control (mRNA ND5). The assay was tested in 75 bovine feces samples tested previously for rotavirus using PAGE and for BCoV using nested RT-PCR targeted to RdRp gene. Agreement with reference tests was optimal for BCoV (kappa = 0.833) and substantial for rotavirus detection (kappa = 0.648). the internal control, ND5 mRNA, was detected successfully in all reactions. Results demonstrated that this multiplex semi-nested RT-PCR was effective in the detection of BCoV and rotavirus, with high sensitivity and specificity for simultaneous detection of both viruses at a lower cost, providing an important tool for studies on the etiology of diarrhea in cattle. (C) 2010 Elsevier B.V. All rights reserved.

GAPD and tubulin are suitable internal controls for qPCR analysis of oral squamous cell carcinoma cell lines

The selection of housekeeping genes is critical for gene expression studies. To address this issue, four candidate housekeeping genes, including several commonly used ones, were investigated in oral squamous cell carcinoma cell lines. A simple quantitative RT-PCR approach was employed by comparing relative expression of the four candidate genes within two cancerous cell tines (HN6 and HN31) and one noncancerous cell tine (HaCaT) treated or not with EGF and TGF-beta 1. Data were analyzed using ANOVA followed by the NormFinder software program. On this basis, stability of the candidate housekeeping genes was ranked and non statistical differences were found using ANOVA test. On the other hand, the NormFinder was able to show that GAPD and TUBB presented the less variable results, representing appropriated housekeeping genes for the samples and conditions anatyzed. In conclusion, this study suggests that the GAPD and the TUBB represent adequate normalizers for gene profiting studies in OSCC cell. tines, covering, respectively, high and low expression levels genes. (C) 2008 Elsevier Ltd. All rights reserved.

Effects of Orthopedic Rapid Maxillary Expansion on Internal Nasal Dimensions in Children With Cleft Lip and Palate Assessed by Acoustic Rhinometry

The objective of the current study was to characterize the internal nasal dimensions of children with repaired cleft lip and palate and transverse maxillary deficiency, using acoustic rhinometry and analyze the changes caused by rapid maxillary expansion (RME). A convenience sampling of 19 cleft lip and palate individuals, aged 14 to 18 years, of both sexes, previously submitted to primary surgeries and referred for RME were analyzed prospectively at the Hospital for Rehabilitation of Craniofacial Anomalies, University of Sao Paulo, Bauru, Sao Paulo, Brazil. All patients underwent acoustic rhinometry before installation of the expansor and at 30 and 180 days after the active expansion phase. Nasal cross-sectional areas and volumes corresponding to the nasal valve (CSA(1) and V(1)) and the turbinates (CSA(2), CSA(3), and V(2)) regions were determined before and after nasal decongestion. Rapid maxillary expansion led to a statistically significant increase (P < 0.05) in mean CSA(1), CSA(2), V(1), and V(2) (without nasal decongestion) and in CSA(1) and V(1) (with decongestion) in the group as a whole. Individual data analysis showed that 58% of the patients responded positively to RME, with an average increase in CSA(1) of 26% (with decongestion), whereas 37% of the patients had no significant change. Only 1 patient (5%) showed a decrease. The findings contribute toward the characterization of nasal deformities determined by the cleft and demonstrate the positive effect RME had on nasal morphophysiology in a significant number of the patients who underwent this procedure.

Facies architecture of the CRP-3 drillhole, Victoria Land Basin, Antarctica

The Cenozoic Victoria Land Basin (VLB) stratigraphic section penetrated by CRP-3 is mostly of Early Oligocene age. It contains an array of lithofacies comprising fine-grained mudrocks, interlaminated and interbedded mudrocks/sandstones, mud-rich and mud-poor sandstones, conglomerates and diametites that are together interpreted as the products of shallow marine to possibly non-marine environments of deposition, affected by the periodic advance and retreat of tidewater glaciers. This lithofacies assemblage can be readily rationalised using the facies scheme designed originally for CRP-2/2A, and published previously. The uppermost 330 metres below sea floor (mbsf) shows a cyclical arrangement of lithofacies also similar to that recognised throughout CRP-2/2A, and interpreted to reflect cyclical variations in relative sea-level driven by ice volume fluctuations ("Motif A"). Between 330 and 480 mbsf, a series of less clearly cyclical units, generally fining-upward but nonetheless incorporating a significant subset of the facies assemblage, has been identified and noted in the Initial Report as "Motif B. Below 480 mbsf, the section is arranged into a repetitive succession of fining-upward units, each of which comprises dolerite clast conglomerate at the base passing upward into relatively thick intervals of sandstones. The cycles present down 480 mbsf are defined as sequences, each interpreted to record cyclical variation of relative sea-level. The thickness distribution of sequences in CRP-3 provides some insights into the geological variables controlling sediment accumulation in the Early Oligocene section. The uppermost part of the section in CRP-3 comprises two or three thick, complete sequences that show a broadly symmetrical arrangement of lithofacies (similar to Sequences 9-11 in CRP-2/2A). This suggests a period of relatively rapid tectonic subsidence, which allowed preservation of the complete facies cycle. Below Sequence 3, however, is a considerable interval of thin, incomplete and erosionally truncated sequences (4-23), which incorporates both the remainder of Motif A sequences and all Motif B sequences recognised. The thinner and more truncated sequences suggest sediment accumulation under conditions of reduced accommodation, and given the lack of evidence for glacial conditions (see Powell et al., this volume) tends to argue for a period of reduced tectonic subsidence. The section below 480 mbsf consists of a series of fining-upward, conglomerate to sandstone intervals which cannot be readily interpreted in terms of relative sea-level change. A relatively mudrock-rich interval above the basal conglomerate/breccia (782-762 mbsf) may record initial flooding of the basin during early rift subsidence. The lithostratigraphy summarised above has been linked to seismic reflection data using depth conversion techniques (Henrys et al., this volume). The three uppermost reflectors ("o", "p" and "q") correlate to the package of thick sequences 1-3, and several deeper reflectors can also be correlated to sequence boundaries. The package of thick Sequences 1-3 shows a sheet-like cross-sectional geometry on seismic reflection lines, unlike the similar package recognised in CRP-2/2A.

A critical comparison of the external and internal boron requirements for contrasting species in boron-buffered solution culture

Despite reports that boron (B) requirements differ among plant species there is a shortage of critical evidence to demonstrate unequivocally whether species differ in internal or external B requirements or both. The present research was conducted to establish the external and internal B requirements of three contrasting species, a woody dicot (marri), an herbaceous dicot (sunflower) and a monocot (wheat) using B-buffered solution culture. Boron-buffered solution culture provided satisfactory control of external B concentrations ranging from 0.04 to 30 muM throughout the 20- (sunflower and wheat) or 40-day (marri) growth period. At low external B concentrations (less than or equal to 0.13 muM), the growth of marri and sunflower was severely depressed but by contrast the vegetative growth of wheat plants was satisfactory and free of B deficiency symptoms. Marri and sunflower plants achieved total maximum shoot growth at greater than or equal to1.2 muM B in solutions while wheat plants did so at greater than or equal to 0.6 muM B. The critical B concentrations (mg kg(-1) dry matter) in the youngest open leaf blades of marri, sunflower and wheat plants were 17.9, 19.7 and 1.2 on 20, 10 and 10 days after transplanting (DAT), respectively. Lower internal and external B requirements of wheat were matched by a lower uptake rate of B compared to marri and sunflower.

Recurrent gains and losses of large (84-109 bp) repeats in the rDNA internal transcribed spacer 2 (ITS2) of rhipicephaline ticks

We studied the internal transcribed spacer 2 (ITS2) in twenty-two spp. of ticks from the subfamily Rhipicephalinae. A 104-109 base pair (bp) region was Imperfectly repeated In most ticks studied. Mapping the number of repeat copies on to a phylogeny from the ITS2 showed that there have been many Independent gains and losses of repeats. Comparison of the sequences of the repeat copies Indicated that in most taxa concerted evolution had played little if any role in the evolution of these regions, as the copies clustered by sequence position rather than species, In our putative secondary structure, each repeat copy can fold into a distinct and almost identical stem-loop complex; a gain or loss of a repeat copy apparently does not impair the function of the ITS2 in these ticks.

Low intraspecific variation in the rRNA internal transcribed spacer 2 (ITS2) of the Australian paralysis tick, Ixodes holocyclus

Ixodes holocyclus has a narrow, discontinuous distribution along the east coast of Australia. We studied ticks from 17 localities throughout the geographic range of this tick. The ITS2 of I. holocyclus is 793 bp long. We found nucleotide variation at eight of the 588 nucleotide positions (1.4%) that were compared for all ticks. There were eight different nucleotide sequences. Most sequences were not restricted to a particular geographic region. However, sequences F, G and H, which had an adenine at position 197, were found only in the far north of Queensland - all other ticks had a guanine at this position. The low level of intraspecific variation in this tick (0.7%) contrasts with the sequence divergence between L holocyclus and its close relative, I. cornuatus (13.1 %). These data indicate that L holocyclus does not contain cryptic species despite possible geographic isolation of some populations. We conclude that variation in the ITS2 is likely to be informative about the phylogeny of the group.

Combining the analyses of introgressive hybridisation and linkage mapping to investigate the genetic architecture of population divergence in the lake whitefish (Coregonus clupeaformis, Mitchill)

Adaptation and reproductive isolation, the engines of biological diversity, are still elusive when discussing the genetic bases of speciation. Namely, the number of genes and magnitude of selection acting positively or negatively on genomic traits implicated in speciation is contentious. Here, we describe the first steps of an ongoing research program aimed at understanding the genetic bases of population divergence and reproductive isolation in the lake whitefish (Coregonus clupeaformis). A preliminary linkage map originating from a hybrid cross between dwarf and normal ecotypes is presented, whereby some of the segregating AFLP markers were found to be conserved among natural populations. Maximum-likelihood was used to estimate hybrid indices from non-diagnostic markers at 998 AFLP loci. This allowed identification of the most likely candidate loci that have been under the influence of selection during the natural hybridisation of whitefish originating from different glacial races. As some of these loci could be identified on the linkage map, the possibility that selection of traits in natural populations may eventually be correlated to specific chromosomal regions was demonstrated. The future prospects and potential of these approaches to elucidate the genetic bases of adaptation and reproductive isolation among sympatric ecotypes of lake whitefish is discussed.

Vitamin B12 stalls the 80 S ribosomal complex on the hepatitis C internal ribosome entry site

The effect of cyanocobalamin (CNCbl, vitamin 1312) on hepatitis C virus internal ribosome entry site (HCV IRES)-dependent initiation of translation was studied by ribosomal toeprinting and sucrose gradient centrifugation analysis. These results suggested that CNCbl did not inhibit HCV IRES-dependent translation by a competitive binding mechanism. CNCbl allowed 80 S elongation complex formation on the mRNA, but stalled the initiation at that point, effectively trapping the 80 S ribosomal complexes on the HCV TRES. CNCbl had no effect on cap-dependent mRNA, consistent with the known mRNA specificity of this translational inhibitor. To help elucidate the mechanism, comparative data were collected for the well-characterised translation inhibitors cycloheximide and 5'-guanylyl-imidophosphate, Although CNCbl stalled HCV IRES-dependent translation at approximately the same step in initiation as cycloheximide, the mechanisms of these two inhibitors are distinct. (C) 2002 Elsevier Science Ltd. All rights reserved.

Cross-national comparison of internal migration: issues and measures

Our objectives are to identify the issues that researchers encounter when measuring internal migration in different countries and to propose key indicators that analysts can use to compare internal migration at the 'national' level. We establish the benefits to be gained by a rigorous approach to cross-national comparisons of internal migration and discuss issues that affect such comparisons. We then distinguish four dimensions of internal migration on which countries can be compared and, for each dimension, identify a series of summary measures. We illustrate the issues and measures proposed by comparing migration in Australia and Great Britain.