Gene Copy Number Quantification of SHOX , VAMP7 , and SRY for the Detection of Sex Chromosome Aneuploidies in Neonates

Aims: To explore the feasibility of detecting sex chromosome aneuploidies (SCAs) by means of gene copy number quantification of short stature homeobox (SHOX), vesicle-associated membrane protein 7 (VAMP7), and SRY in newborns. Materials and Methods: Gene doses of SHOX, VAMP7, and SRY were determined by quantitative polymerase chain reaction (qPCR) using DNA obtained from dried blood samples from newborns. Relative quantification values were obtained. An aneuploidy profile was established according to cutoff values. Samples with ≥2 gene doses (out of range) were reanalyzed, and those with aneuploidy profiles were confirmed by karyotyping. Sensitivity, specificity, and positive and negative predictive values were obtained. Results: A total of 10,033 samples were collected (4945 females and 5088 males). Of 244 (2.43%) samples with ≥2 gene doses that were retested, 20 cases were confirmed. The overall incidence of SCAs was 1 in 500 live newborns. There were six cases of Turner syndrome (1/824), 3 cases of XXX (1/1648), 7 cases of Klinefelter syndrome (1/726), and 4 cases of of XYY (1/1272). The sensitivity was 0.952 (95.24%), specificity of 0.975 (97.56%), positive predictive value of 0.909 (90.91%), and negative predictive value of 0.987 (98.77%). Conclusions: Gene copy number analyses of VAMP7, SHOX, and SRY genes by qPCR from blood samples spotted onto filter paper is a highly reliable method for the early detection of male and female SCAs

[Ca2+]i changes in guinea pig tracheal smooth muscle cells in culture: effects of Na+ and ouabain

The objective of this work was to confirm that the contractile effects of ouabain and Na+-free solutions in guinea pig tracheal rings are associated with increments in the cytosolic free Ca2+ concentration ([Ca2+]i) in cultured tracheal smooth muscle (TSM) cells. Cultured cells were a-actin positive. Histamine (50 mM) and Na+ -free solution elicited a transient increase in [Ca2+]I, while the responses to thapsigargin (1 µM) and ouabain (1 mM) were long lasting. However, carbachol (10, 200, and 500 mM) and high K+-solution produced no effect on [Ca2+]i, suggesting that cultured guinea pig TSM cells display a phenotype change but maintain some of the tracheal rings physiological properties. The transient rise in [Ca2+]i in response to the ce of extracellular Na+ and the effect of ouabain may indicate the participation of the Na+/Ca2+ exchanger (NCX) in the regulation of [Ca2+]i.El objetivo del presente trabajo es el de confirmar que los efectos contráctiles producidos por la ouabaína y la ausencia de Na+ en la solución en anillos de traquea de cobayo, se asocian a incrementos en la concentración libre de Ca2+ citosólico ([Ca2+]i) en células cultivadas de músculo liso traqueal de cobayo. Las células usadas en los experimentos eran positivas para a-actina de músculo liso. La histamina (50 µM) y las soluciones libres de Na+ producen un incremento transitorio en la [Ca2+]i mientras que la thapsigargina (1 µM) y la ouabaína (1 mM) dan lugar a incremento de larga duración. Sin embargo, el carbacol (10, 200 y 500 µM) y soluciones con alta concentración de K+ no producen cambios en la [Ca2+]i . Estos resultados sugieren que, en nuestras condiciones experimentales, las células cultivadas de músculo liso traqueal presentan cambios en su fenotipo, pero conservan algunas de las propiedades fisiológicas observadas en los anillos traqueales. El incremento transitorio de la [Ca2+]i producido por la ausencia de Na+ extracelular y el efecto de la ouabaína sugieren la participación del intercambiador Na+/Ca2+ (NCX) en la regulación de la [Ca2+]i