In vitro development and preservation of specific features of collecting duct epithelial cells from embryonic rabbit kidney are regulated by the electrolyte environment

During kidney development the embryonic collecting duct (CD) epithelium changes its function. The capability for nephron induction is lost and the epithelium develops into functional principal (P) and intercalated (IC) cells. Aldosterone is able to modulate this differentiation. Consequently we investigated whether increased concentrations of extracellular NaCl or Na gluconate may also have an influence on the development of individual CD cell features. Embryonic CD epithelia were isolated from neonatal rabbit kidneys, placed on tissue carriers and cultured in gradient containers, which were constantly perfused with medium for 13 days. Isotonic culture conditions could be mimicked, when on both the luminal and basal side standard Iscove's Modified Dulbecco's Medium (IMDM) was used. In another set of experiments, gradient culture was performed. Standard IMDM was applied on the basal side and IMDM supplemented with 12 mM NaCl and 17 mM Na gluconate on the luminal side. This adaptation of IMDM led to the same Na concentrations as found in the serum of neonatal rabbits. The development of CD cell features was monitored by cellular markers such as the monoclonal antibodies (Mabs) 703 and 503 recognizing P and IC cell features respectively. Epithelia cultured under isotonic conditions showed less than 5% Mab 703- and 503-immunopositive cells. In contrast, epithelia cultured in a luminal-basal medium gradient revealed more than 80% positive cells. Immunoreactivity started to develop after a long lag period of 4 days, then increased continuously during the following 5 days and reached a maximum at day 14. When the medium gradient was then changed to an isotonic environment for another 5 days immunoreactivity for Mab 703 remained stable, while the number of Mab 503-positive cells was found to be decreased to 10%. Thus, the extra-cellular electrolyte environment not only induces but also preserves individual cell features.     

The effect of calcipotriol on lesional fibroblasts from patients with active morphoea

We examined the responsiveness of cultured dermal fibroblasts from biopsies of 6 patients with active morphoea and a similar number of matched controls to the cell proliferation inhibition activity of calcipotriol. Cultured fibroblasts from controls showed no significant response to calcipotriol (at concentrations of 1 x 10(-8) to 1 x 10(-4) M). However, calcipotriol did inhibit the proliferation response of morphoea fibroblasts at all concentrations when compared with controls. There was 4- to 20-fold inhibition in 2 of the morphoea patients when compared with control samples. Four other morphoea samples showed inhibition but to a lesser extent compared with controls.     

Lack of evidence for the transmission of JC polyomavirus between human populations

Human polyomavirus JC virus (JCV), the causative agent of progressive multifocal leukoencephalopathy, is ubiquitous in humans, infecting children asymptomatically then persisting in renal tissue. Since JCV DNA can readily be detected from urine, it should be a useful tool with which to study the mode of virus transmission in humans. Based on this notion, we examined the extent to which JCV was transmitted from the American to Japanese populations in Okinawa Island, Japan. (A population of about 50 000 American soldiers and families have been stationed in Okinawa since 1945.) Four JCV types (A to D) were identified in American populations in U.S.A., whereas only type B was prevalent in elder Japanese in Okinawa who had reached adulthood by 1945. Thus, types A, C, and D served as indicators of the transmission of JCV from American to Japanese populations. We then examined whether types A, C, and D were detectable in Japanese in Okinawa aged 30-50 years who may have been in contact with Americans during childhood. However, all the 125 isolates from the younger Japanese population were type B without exception. From this finding, we concluded that JCV is rarely transmitted between human populations.     

Nitric oxide-synthesising neurons in the central subnucleus of the nucleus tractus solitarius provide a major innervation of the rostral nucleus ambiguus in the rabbit

We describe an intramedullary nitric oxide synthase (NOS) neural pathway that projects from the nucleus tractus solitarius (NTS) to the rostral nucleus ambiguus (NA) in the rabbit. With the use of NADPH diaphorase histochemistry and NOS immunohistochemistry, a compact group of NOS-positive perikarya was identified in the central subnucleus of the NTS dorsomedial to the tractus solitarius and rostral to the obex. A dense network of NOS terminals was seen in the rostral NA. We investigated whether NOS terminals in the NA derive from NOS perikarya in the central NTS and whether the central NOS pathway links esophageal afferents and efferents. In some rabbits, the central NTS was unilaterally lesioned. In others, Phaseolus vulgaris-leucoagglutinin (PHA-L) was injected into the central NTS, or cholera toxin-gold was injected into the NA, or cholera toxin-horseradish peroxidase (HRP) was injected into the wall of the esophagus. The medulla was subsequently processed to demonstrate PHA-L, cholera toxin-gold, HRP, and NOS reactivity. Seven days after the NTS lesion, we observed a marked decrease in the density of NOS terminals in the ipsilateral NA. After injection of PHA-L into the central NTS, a dense group of PHA-L fibres was seen in the rostral NA, principally ipsilaterally. Afferent fibres from the esophagus were found around the NOS cell bodies in the central NTS, and many of these NOS neurons were double labeled with cholera toxin-gold after injection of this tracer into the NA. NOS terminals were found around NA neurons that were retrogradely labelled from the esophagus. We conclude that the NOS neurons in the central NTS act as interneurons in a central pathway connecting esophageal afferents and efferents.     

Prevention of vertical HIV transmission with zidovudine: projected impact of HIV testing and prenatal care

We sought to estimate the impact of maternal HIV testing and prenatal care on the potential to reduce vertical transmission through zidovudine (AZT) use by HIV-infected mothers. We evaluated the prepartum maternal HIV diagnosis rate, prenatal care, disease stage, and vertical transmission rate (from a two-part mixture model) using New York State Medicaid and vital statistics data for HIV-infected mothers and their singletons in 1985-90. We used published data to estimate the effect of AZT on vertical transmission and expert input to define other parameters for the model. Our HIV-infected (N = 1514) had a vertical transmission rate of 27.0%. HIV was diagnosed prepartum for 39.5% of women in 1990. Transmission would have been 22.2% if AZT had been taken only by the subset of women diagnosed prepartum with HIV and receiving prenatal care by 34 weeks gestation (86.7%). Transmission would have dropped to 11.2% if all women had been diagnosed prepartum with HIV and received adequate prenatal care. The observed deficiencies in prenatal care and maternal HIV diagnosis rates in this Medicaid population-based cohort must be addressed to realize the promise of AZT to reduce vertical transmission.     

Large scale identification of genes involved in cell surface biosynthesis and architecture in Saccharomyces cerevisiae

The sequenced yeast genome offers a unique resource for the analysis of eukaryotic cell function and enables genome-wide screens for genes involved in cellular processes. We have identified genes involved in cell surface assembly by screening transposon-mutagenized cells for altered sensitivity to calcofluor white, followed by supplementary screens to further characterize mutant phenotypes. The mutated genes were directly retrieved from genomic DNA and then matched uniquely to a gene in the yeast genome database. Eighty-two genes with apparent perturbation of the cell surface were identified, with mutations in 65 of them displaying at least one further cell surface phenotype in addition to their modified sensitivity to calcofluor. Fifty of these genes were previously known, 17 encoded proteins whose function could be anticipated through sequence homology or previously recognized phenotypes and 15 genes had no previously known phenotype.     

Electrolyte environment modulates differentiation in embryonic renal collecting duct epithelia

The influence of electrolytes on the development of renal principal and intercalated collecting duct cells is unknown. Consequently embryonic collecting duct epithelia were exposed to different electrolyte concentrations, and their degree of differentiation was registered by immunohistochemical methods. Embryonic collecting duct epithelia were isolated from neonatal rabbit kidneys and placed on tissue carriers. The apical urine and the basal serum compartments were simulated in a gradient culture container. The two sides of the epithelium were each constantly superfused with medium for 13 days. In controls the medium on both apical and basal side was standard Iscove's modified Dulbecco's Medium (IMDM) with 112 mmol/l Na+ and 85 mmol/l Cl-. In experimental series the NaCl concentration at the basal side of the epithelium was increased up to 137 mmol/l Na+ and 99 mmol/l Cl- as found in the serum of neonatal rabbits. Light microscopy revealed morphologically faultless epithelia following gradient perfusion culture in standard and NaCl-adapted IMDM. The development of principal and intercalated cell features was monitored with the monoclonal antibodies 703, 503, PCD9, and peanut lectin. Cells immunopositive for monoclonal antibody 703, for example, increased from less than 10% in controls to more than 80% in NaCl-adapted IMDM. It is a new finding that the development of collecting duct cell features is influenced by the extracellular electrolyte environment.     

Changes in muscle morphology, electromyographic activity, and force production characteristics during progressive strength training in young and older men

Effects of a 10-week progressive strength training program composed of a mixture of exercises for increasing muscle mass, maximal peak force, and explosive strength (rapid force production) were examined in 8 young (YM) (29+/-5 yrs) and 10 old (OM) (61+/-4 yrs) men. Electromyographic activity, maximal bilateral isometric peak force, and maximal rate of force development (RFD) of the knee extensors, muscle cross-sectional area (CSA) of the quadriceps femoris (QF), muscle fiber proportion, and fiber areas of types I, IIa, IIb, and IIab of the vastus lateralis were evaluated. Maximal and explosive strength values remained unaltered in both groups during a 3-week control period with no training preceding the strength training. After the 10-week training period, maximal isometric peak force increased from 1311+/-123 N by 15.6% (p <.05) in YM and from 976+/-168 N by 16.5% (p <.01) in OM. The pretraining RFD values of 4049+/-791 N*s(-1) in YM and 2526+/-1197 N*s(-1) in OM remained unaltered. Both groups showed significant increases (p < .05) in the averaged maximum IEMGs of the vastus muscles. The CSA of the QF increased from 90.3+/-7.9 cm2 in YM by 12.2% (p <.05) and from 74.7+/-7.8 cm2 in OM by 8.5% (p <.001). No changes occurred in the muscle fiber distribution of type I during the training, whereas the proportion of subtype IIab increased from 2% to 6% (p < .05) in YM and that of type IIb decreased in both YM from 25% to 16% (p < .01) and in OM from 15% to 6% (p < .05). The mean fiber area of type I increased after the 10-week training in YM (p < .001) and OM (p < .05) as well as that of type IIa in both YM (p < .01) and OM (p < .01). The individual percentage values for type I fibers were inversely correlated with the individual changes recorded during the training in the muscle CSA of the QF (r=-.56, p < .05). The present results suggest that both neural adaptations and the capacity of the skeletal muscle to undergo training-induced hypertrophy even in older people explain the gains observed in maximal force in older men, while rapid force production capacity recorded during the isometric knee extension action remained unaltered during the present mixed strength training program.