[Contribution to the development of the union between the manubrium of the malleus and the tympanic membrane in human fetus]. / Contribución al desarrollo de la unión del mango del martillo y la membrana del tímpano en fetos humanos.

The development of the union between the manubrium of the malleus and the tympanic membrane was studied in human embryo. For that purpose 25 temporal bones of human foetus, aged from 36 days (14 mm) to 29 weeks (270 mm) were analyzed. Samples were fixed in a 10% formaldehyde solution, decalcified with 2% nitric acid, embedded in Paraplast, cut in sections of 7 microns thick and stained with Martin's trichrome method. During the development a pseudojoint between the malleus and the tympanic membrane several stages were seen. In the first stage, the manubrium was adhered to membrane mesenchyme of primitive tympanum, in the second one this mesenchyme was loose and there appeared capillars in it, in the third one there were collagen fibers in a radial disposition, and in the forth stage, the hollow in the meatal plug gave independence to the tympanic membrane for the external acoustic meatus. The distal portion of the manubrium, included in tympanic membrane, has remained with a cartilaginous structure during end of the time it has been studied by us. The collagen fibers at the level of the umbo surround the manubrium, while the rest of it remains in its anterior edge.

Contribución al desarrollo de la unión del mango del martillo y la membrana del tímpano en fetos humanos / Contribution to the development of the union between the manubrium of the malleus and the tympanic membrane in human fetus

Se ha estudiado el desarrollo de la unión entre el mango del martillo y la membrana del tímpano en embriones y fetos humanos.Para ello se han utilizado 25 huesos temporales de fetos humanos, cuyas edades cronológicas van desde los 36 días (14 mm) hasta las 29 semanas (270 mm). Las muestras fueron fijadas en formol al 10 por ciento, descalcificadas con ácido nítrico al 2 por ciento, incluidas en parafina, cortadas en serie a 7 µm, y teñidas con el método Tricrómico de Martins. Se han identificado varias etapas en el desarrollo de la pseudoarticulación entre el martillo y la membrana del tímpano. En la primera el mango se adhiere íntimamente al mesénquima de la membrana del tímpano primitiva, en la segunda dicho mesénquima se hace laxo y en él aparecen capilares, en la tercera aparecen fibras de colágeno de disposición radial, y en la cuarta el ahuecamiento del tapón meatal da independencia a la membrana del tímpano del conducto auditivo externo. La porción distal del mango, incluida en la membrana del tímpano, ha permanecido con una estructura cartilaginosa durante todo el tiempo estudiado. Las fibras de colágeno a nivel del umbo rodean al extremo terminal del mango mientras que en el resto de él lo hacen solamente en su arista anterior (AU)

The development of the union between the manubrium of the malleus and the tympanic membrane was studied in human embryo. For that purpose 25 temporal bones of human foetus, aged from 36 days (14 mm) to 29 weeks (270 mm) were analyzed. Samples were fixed in a 10% formaldehyde solution, decalcified with 2% nitric acid, embedded in Paraplast, cut in sections of 7 microns thick and stained with Martin's trichrome method. During the development a pseudojoint between the malleus and the tympanic membrane several stages were seen. In the first stage, the manubrium was adhered to membrane mesenchyme of primitive tympanum, in the second one this mesenchyme was loose and there appeared capillars in it, in the third one there were collagen fibers in a radial disposition, and in the forth stage, the hollow in the meatal plug gave independence to the tympanic membrane for the external acoustic meatus. The distal portion of the manubrium, included in tympanic membrane, has remained with a cartilaginous structure during end of the time it has been studied by us. The collagen fibers at the level of the umbo surround the manubrium, while the rest of it remains in its anterior edge (AU)

Structure and expression of the human Na,K-ATPase beta 2-subunit gene.

We cloned and characterized the human Na,K-ATPase beta 2-subunit gene. The gene encompasses over 8 kb at chromosome 17 in the human genome and is composed of seven exons. Primer extension analysis identified a major transcription initiation site 529 bases upstream of the translation start site. The 5'-flanking region of the gene harbors a potential TATA sequence, located 94 bases upstream of the transcription initiation site and a number of potential promoter and regulatory elements, among them a Sp1 site, at position -120. A functional Sp1 site has also been found in the rat Na,K-ATPase beta 2-subunit gene (Kawakami, K., Watanabe, Y., Araki, M., Nagano, K., 1993). Sp1 binds to the adhesion molecule on glia regulatory element that functions as a positive transcription regulatory element in astrocytes. (J. Neurosci. Res. 35, 138-146). Putative AATAAA and TG sequences were found at positions 7018 and 7068, respectively. These signals delimit the origin of the the poly(A) tail and mark the end of the sequence that completes the 3'-UT downstream sequence of the human cDNA. An Alu repetitive sequence is located between positions 5961 and 6274. The gene is expressed as a single mRNA species, of 3.36 kb, which is present in cerebrum, cerebellum, kidney and heart, being more abundant in neural tissues. Structural analyses of this and other of the P-type ATPase beta subunit genes reveal that they evolved from a common ancestor.

Expression of the beta-isoforms of Na,K-ATPase in the renal cortex of rats.

The aim of the present study was to assess the presence of beta1- and beta2-isoforms of the beta-subunit of Na,K-ATPase in the rat renal cortex. This has been accomplished by immunohistochemistry and Western blotting using isoform-specific antisera. Western blot of brain extract, used as positive control, revealed the bands corresponding to beta1- and beta2-glycosylated peptides, with a molecular weight (MW) of approximately 50-60 that, after exhaustive treatment with N-endoglycosidase-F, migrated to the MW corresponding to the core peptides (approximately 35). In the renal cortex, Western blot revealed the bands corresponding to beta1. After deglycosylation of the samples, the bands hybridizing with the anti-beta1-antibodies moved to the MW corresponding to a partially deglycosylated form and the core peptide. Bands with a MW of approximately 50-60 hybridized with anti-beta2, although digestion with endoglycosidase failed to move the band towards a lower MW. Immunohistochemistry revealed the presence of beta1- but not beta2-isoform. Northern blot for total mRNA showed strong signals for beta1 in renal cortex, the mRNA for the beta2-isoform being undetectable. In conclusion, only mRNA and glycopeptide of the beta1-isoform seem to be present in renal cortex of adult control rats.

[Frequency of drug adverse reactions among hospitalized patients]. / Frecuencia de reacciones adversas a fármacos en pacientes hospitalizados.

This article describes the frequency of adverse reactions to drugs in a sample of hospitalized patients in the internal medicine ward seen during a year's term. Of 61 medical charts, we found 8 patients with adverse reactions to drugs during their hospital stay and another 4 patients hospitalized due to adverse reactions to drugs. The majority of the adverse reactions were of moderate degree (75%) and were related to drugs of cardiovascular action (58%). The frequency of reactions in hospitalized patients (13%) is comparable with the results obtained from other hospitals. Yet, the real magnitude of the problem is probably greater since the source of information (hospital charts) the totality of the clinical manifestations are not registered.

[Production of specific antibodies against isoforms of the beta subunit of human Na,K-ATPase]. / Generación de anticuerpos específicos contra las isoformas de la subunidad beta de la Na,K-ATPasa humana.

The study of the differential and functional tissular distribution of the isoforms of Na,K-ATPase, an enzyme located on the plasma membrane of animal cells, must be performed with the aid of antibodies capable of specifically recognising beta 1 and beta 2 subunits. In order to facilitate the study of the functions and distribution of such isoforms, proteins were generated using pET series plasmid construction and transformation in E coli bacteria, and their production was promoted with IPTG, these being the antigens that induce antibodies specific for each isoform. The antisera thus obtained were used in Western Blot assays with tissue microsomes from different animal species; specifically, beta 1 and beta 2 isoforms of Na,K-ATPase could be recognised. A panel of polyclonal antibodies was obtained which proved capable of recognising these isoforms in different animal species, as well as their deglycosylated isoform. Such antibodies can be used as an efficient tool for functional and tissue distribution studies of the two isoforms.

Expression of the beta-subunit isoforms of the Na,K-ATPase in rat embryo tissues, inner ear and choroid plexus.

We report evidence of the apical localization of the two Na,K-ATPase beta-subunit isoforms in cells of the inner ear and of the choroid plexus of the rat. To this end, we generated isoform-specific antisera against the human Na,K-ATPase beta 1 and beta 2 subunits. These polyclonal rabbit antisera were raised against truncated beta-isoform proteins that were made in E coli with pET expression vectors. Deglycosylation of the native antigen with N-endoglycosidase F shows four bands in the beta 1 isoform and five bands in the beta 2 isoform immunoblots. In E15 rat embryos, the beta 1 isoform was detected in brain, heart and kidney and the beta 2 isoform only in brain. While beta-subunit mRNA expression (Watts AG, Sanchez-Watts G, Emanuel JR, Levenson R (1991) Proc Natl Acad Sci USA 88, 7425-7429), immunoblotting and enzymatic activity have been determined (Zlokovic BV< mackic JB, Wang L, McComb JG, McDonough A (1993) J Biol Chem 268, 8019-8025), very little is known about the specific localization of each beta-isoform in the epithelia of choroid plexus and inner ear. Immunocytochemical preparations of 15-day-old whole rat embryos and adult rat brain showed an enhanced staining for the beta 1 and beta 2 isoforms in the apical membrane of the ampullary crests of the inner ear's semicircular ducts and in the cuboidal cells of the choroid plexus.