Multipotent capacity of immortalized human bronchial epithelial cells

While the adult murine lung utilizes multiple compartmentally restricted progenitor cells during homeostasis and repair, much less is known about the progenitor cells from the human lung. Translating the murine stem cell model to humans is hindered by anatomical differences between species. Here we show that human bronchial epithelial cells (HBECs) display characteristics of multipotent stem cells of the lung. These HBECs express markers indicative of several epithelial types of the adult lung when experimentally tested in cell culture. When cultured in three different three-dimensional (3D) systems, subtle changes in the microenvironment result in unique responses including the ability of HBECs to differentiate into multiple central and peripheral lung cell types. These new findings indicate that the adult human lung contains a multipotent progenitor cell whose differentiation potential is primarily dictated by the microenvironment. The HBEC system is not only important in understanding mechanisms for specific cell lineage differentiation, but also for examining changes that correlate with human lung diseases including lung cancer.     

Structural requirements for the interaction of human IgA with the human polymeric Ig receptor

Transport of polymeric IgA onto mucosal surfaces to become secretory IgA is mediated by the polymeric Ig receptor (pIgR). To study the interaction of human dimeric IgA (dIgA) (the predominant form of IgA polymer) with the human pIgR (hpIgR), we generated recombinant wild-type dIgA1 and dIgA2m(1) and various mutant dIgA1 and analyzed their interaction with a recombinant human secretory component and membrane-expressed hpIgR. We found that wild-type dIgA1 and dIgA2m(1) bound to recombinant human secretory component with similar affinity and were transcytosed by the hpIgR to the same extent. Mutation of the IgA Calpha2 domain residue Cys311 to Ser reduced binding to hpIgR, possibly through disruption of noncovalent interactions between the Calpha2 domain and domain 5 of the receptor. Within the Calpha3 domain of IgA1, we found that combined mutation of residues Phe411, Val413, and Thr414, which lie close to residues previously implicated in hpIgR binding, abolished interaction with the receptor. Mutation of residue Lys377, located very close to this same region, perturbed receptor interaction. In addition, 4 aa (Pro440-Phe443), which lie on a loop at the domain interface and form part of the binding site for human FcalphaRI, appear to contribute to hpIgR binding. Lastly, use of a monomeric IgA1 mutant lacking the tailpiece revealed that the tailpiece does not occlude hpIgR-binding residues in IgA1 monomers. This directed mutagenesis approach has thus identified motifs lying principally across the upper surface of the Calpha3 domain (i.e., that closest to Calpha2) critical for human pIgR binding and transcytosis.     

Molecular evolution of voltage-sensitive ion channel genes: on the origins of electrical excitability

We have analyzed nucleic acid and amino acid sequence alignments of a variety of voltage-sensitive ion channels, using several methods for phylogenetic tree reconstruction. Ancient duplications within this family gave rise to three distantly related groups, one consisting of the Na+ and Ca++ channels, another the K+ channels, and a third including the cyclic nucleotide-binding channels. A series of gene duplications produced at least seven mammalian homologues of the Drosophila Shaker K+ channel; clones of only three of these genes are available from all three mammalian species examined (mouse, rat, and human), pointing to specific genes that have yet to be recovered in one or another of these species. The Shaw-related K+ channels and the Na+ channel family have also undergone considerable expansion in mammals, relative to flies. These expansions presumably reflect the needs of the high degree of physiological and neuronal complexity of mammals. Analysis of the separate domains of the four-domain channels (Ca++ and Na+) supports their having evolved by two sequential gene duplications and implies the historical existence of a functional two-domain channel.      

Facies development of a Late Ordovician mixed carbonate-siliciclastic ramp proximal to the developing Taconic orogen: Lourdes Formation, Newfoundland, Canada

The Upper Ordovician (late Whiterockian to Mohawkian) Lourdes Formation represents a narrow (tens of kilometers), short-lived [∼5–7 million years (my)], open-ocean (high-energy) mixed siliciclastic-carbonate ramp that onlapped allochthonous strata along the orogen side of the local Taconic foreland basin. Platform development followed a 6–8 my hiatus during which weathering had concentrated chemically mature siliciclastics that were admixed with initial carbonate sediments. A cross-platform facies gradient contains paleokarst and peritidal carbonates and sandstones, shallow-ramp carbonate bioherms and skeletal shoals, and deeper ramp calcareous shales. Transgressive systems tracts are marked by ramp-wide sheets and shoals of skeletal grainstone and low accumulation rates, and highstand systems tracts are marked by significant admixture and interbedding of siliciclastics with cross-ramp carbonate facies. Platform demise coincides with increased siliciclastic input, which is likely tectonically influenced. The Lourdes platform is equivalent to epicontinental foreland ramps along eastern Laurentia, but its narrower width precluded formation of oceanographically restricted platform-interior facies.     

Effect of postnatal development on calcium currents and slow charge movement in mammalian skeletal muscle

Single- (whole-cell patch) and two-electrode voltage-clamp techniques were used to measure transient (Ifast) and sustained (Islow) calcium currents, linear capacitance, and slow, voltage-dependent charge movements in freshly dissociated fibers of the flexor digitorum brevis (FDB) muscle of rats of various postnatal ages. Peak Ifast was largest in FDB fibers of neonatal (1-5 d) rats, having a magnitude in 10 mM external Ca of 1.4 +/- 0.9 pA/pF (mean +/- SD; current normalized by linear fiber capacitance). Peak Ifast was smaller in FDB fibers of older animals, and by approximately 3 wk postnatal, it was so small as to be unmeasurable. By contrast, the magnitudes of Islow and charge movement increased substantially during postnatal development. Peak Islow was 3.6 +/- 2.5 pA/pF in FDB fibers of 1-5-d rats and increased to 16.4 +/- 6.5 pA/pF in 45-50-d-old rats; for these same two age groups, Qmax, the total mobile charge measurable as charge movement, was 6.0 +/- 1.7 and 23.8 +/- 4.0 nC/microF, respectively. As both Islow and charge movement are thought to arise in the transverse-tubular system, linear capacitance normalized by the area of fiber surface was determined as an indirect measure of the membrane area of the t-system relative to that of the fiber surface. This parameter increased from 1.5 +/- 0.2 microF/cm2 in 2-d fibers to 2.9 +/- 0.4 microF/cm2 in 44-d fibers. The increases in peak Islow, Qmax, and normalized linear capacitance all had similar time courses. Although the function of Islow is unknown, the substantial postnatal increase in its magnitude suggests that it plays an important role in the physiology of skeletal muscle.     

An unusual megaspore of uncertain systematic affinity from Lower Cretaceous deposits in south-east England and its biostratigraphic and palaeoenvironmental significance

An unusual megaspore has been recorded from beds within the Weald Clay Group of south-east England. Described as Clockhousea capelensis gen. et sp. nov., it is characterised by having a thick outer layer of exine consisting of closely packed columnar to clavate elements with constricted bases attached to a perforated inner layer and a sculpture that ranges from having the appearance of a negative reticulum through closely spaced verrucae to a mixture of verrucate and essentially baculate elements, all of which are surface manifestations and extensions of the underlying structure. These characters do not readily indicate a systematic relationship with any known heterosporous plant genus or family. The localised occurrence and relative abundance of the spores in a few beds suggest that some of the parent plants grew close to water bodies where they were deposited and preserved. Their recovery from sediments of late Hauterivian–early Barremian age indicates that the species has potential as a biostratigraphic marker in the upper Wealden succession of southern England and perhaps elsewhere.     

The role of glutamate signaling in incentive salience: second‐by‐second glutamate recordings in awake Sprague‐Dawley rats

The attribution of incentive salience to reward‐predictive stimuli has been shown to be associated with substance abuse‐like behavior such as increased drug taking. Evidence suggests that glutamate neurotransmission and sequential N‐methyl‐D‐aspartate (NMDA) activation are involved in the attribution of incentive salience. Here, we further explore the role of second‐by‐second glutamate neurotransmission in the attribution of incentive salience to reward‐predictive stimuli by measuring sign‐tracking behavior during a Pavlovian conditioned approach procedure using ceramic‐based microelectrode arrays configured for sensitive measures of extracellular glutamate in awake behaving Sprague‐Dawley rats. Specifically, we show that there is an increase in extracellular glutamate levels in the prelimbic cortex (PrL) and the nucleus accumbens core (NAcC) during sign‐tracking behavior to a food‐predictive conditioned stimulus (CS+) compared to the presentation of a non‐predictive conditioned stimulus (CS?). Furthermore, the results indicate greater increases in extracellular glutamate levels in the PrL compared to NAcC in response to the CS+, including differences in glutamate release and signal decay. Taken together, the present research suggests that there is differential glutamate signaling in the NAcC and PrL during sign‐tracking behavior to a food‐predictive CS+.