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121.
J?Craig CohenEmail author Lennart?KA?Lundblad Jason?HT?Bates Michael?Levitzky Janet?E?Larson 《BMC genetics》2004,5(1):21
Background
Cystic Fibrosis is a pleiotropic disease in humans with primary morbidity and mortality associated with a lung disease phenotype. However, knockout in the mouse of cftr, the gene whose mutant alleles are responsible for cystic fibrosis, has previously failed to produce a readily, quantifiable lung phenotype. 相似文献122.
123.
Larson SD Plopper CG Baker G Tarkington BK Decile KC Pinkerton K Mansoor JK Hyde DM Schelegle ES 《American journal of physiology. Lung cellular and molecular physiology》2004,287(2):L286-L295
Mucous cell hypersecretion and increased neuropeptide production play a role in the exacerbation of symptoms associated with asthma. The source of these neuropeptides have been confined to the contributions of small afferent nerves or possibly neuroendocrine cells. We tested the hypothesis that repeated exposure to allergen would alter the sources and abundance of neuropeptides in airways. Right middle lobes from rats (8 wk old) exposed to 2.5% ovalbumin (OVA) for five episodes (30 min each) or filtered air were inflation fixed with paraformaldehyde. The lobes were dissected to expose the airway tree, permeabilized with DMSO, and incubated in antibody to rat calcitonin gene-related peptide (CGRP), followed with a fluorochrome-labeled second antibody. CGRP-positive structures were imaged via confocal microscopy. Airways were later embedded in plastic and sectioned for cell identification. In animals challenged with OVA, CGRP-positive cells, not neuroendocrine or neuronal in origin (confirmed by a lack of protein gene product 9.5 signal), were recorded along the axial path. In section, this fluorescent signal was localized to granules within epithelial cells. Alcian blue/periodic acid-Schiff staining of these same sections positively identify these cells as mucous cells. Mucous cells of animals not challenged with OVA were not positive for CGRP. We conclude that episodic allergen exposure results in the accumulation of CGRP within mucous cells, creating a new source for the release of this neuropeptide within the airway. 相似文献
124.
Alpha4beta1 integrin affinity changes govern cell adhesion 总被引:3,自引:0,他引:3
Chigaev A Zwartz G Graves SW Dwyer DC Tsuji H Foutz TD Edwards BS Prossnitz ER Larson RS Sklar LA 《The Journal of biological chemistry》2003,278(40):38174-38182
Integrin alpha4beta1 is a receptor for vascular cell adhesion molecule-1 and fibronectin. It is important in lymphopoiesis, inflammatory recruitment of leukocytes, and other situations that require cell adhesion to the vascular endothelium. The avidity of the cells expressing alpha4beta1 integrin can be rapidly changed by chemokines and chemoattractants. Different mechanisms, including changes in the number of interacting molecules due to the alteration of the receptor topology or changes in the affinity of the individual bonds, have been proposed to explain the nature of these fast changes in avidity. Recently, we described a fluorescent LDV-containing small molecule, which we used to monitor the affinity changes on live cells in real time (Chigaev, A., Blenc, A. M., Braaten, J. V., Kumaraswamy, N., Kepley, C. L., Andrews, R. P., Oliver, J. M., Edwards, B. S., Prossnitz, E. R., Larson, R. S. et al. (2001) J. Biol. Chem. 276, 48670-48678). Here we show that the affinity of the small molecule probe as well as the native ligand vascular cell adhesion molecule-1 varies in parallel when the integrin is modulated with divalent cations and that the affinity modulation leads to the changes in cell avidity. Using formyl peptide receptor-transfected U937 cells, we further show that the time course of avidity changes in response to the receptor activation coincides with the time course of the affinity changes. Taken together, these data are consistent with the idea that affinity regulation is a major factor that governs the avidity of cell adhesion mediated by the alpha4 integrin. 相似文献
125.
We have examined the metal ion-binding domains from rat alpha and beta parvalbumin. We find that the CD-EF fragments differ markedly in their tendency to self-associate. Whereas Ca(2+)-free alpha CD-EF is monomeric, the Ca(2+)-free beta peptide dimerizes weakly (K(2) = 2400 +/- 200 M(-1)). In buffer containing 1.0 mM Ca(2+), the apparent dimerization constant for beta CD-EF (191,000 +/- 29,000 M(-1)) is more than 50 times that of alpha (3400 +/- 200 M(-1)). Alpha CD-EF binds two Ca(2+) with positive cooperativity. Titration calorimetry data afford binding constants of 3.7(0.1) x 10(3) M(-1) and 8.6(0.2) x 10(4) M(-1). Beta CD-EF also binds two Ca(2+) cooperatively but with lower affinity. Equilibrium dialysis yields Adair constants of 4.2(0.1) x 10(3) and 6.1(0.2) x 10(3) M(-1). Significantly, the difference in Ca(2+) affinity is substantially smaller than that observed for the full-length proteins-suggesting that the AB domain can modulate divalent ion affinity. Analysis of beta calorimetry data requires explicit consideration of the self-association behavior. Data collected at low CD-EF concentration are consistent with preferential occupation of the EF site, dimerization of singly bound monomers, and cooperative filling of the CD sites. At higher concentrations, apo-protein dimerization can apparently precede cooperative occupation of the EF sites. In the presence of Ca(2+), alpha CD-EF exhibits higher thermal stability, consistent with its higher Ca(2+) affinity. However, the beta melting temperature shows greater concentration dependence, consistent with its greater tendency to dimerize. Neither fragment exhibits a sigmoidal melting curve in the Ca(2+)-free state, suggesting that the apo-peptides are disordered. 相似文献
126.
Summers TR Matter JM McKay JM Ronan PJ Larson ET Renner KJ Summers CH 《Hormones and behavior》2003,43(1):245-253
Central serotonin (5-HT) is activated during stressful situations and aggressive interactions in a number of species. Glucocorticoids secreted peripherally during stressful events feed back on central systems and may affect 5-HT mediation of stress-induced behavioral events. To test the neuromodulatory effect of stress hormone secretion, serotonin overflow was measured from the hippocampus of the lizard Anolis carolinensis. Microdialysis was used to collect repeated samples from anesthetized lizards, with perfusate measured by HPLC with electrochemical analysis. Following initially high levels of 5-HT, concentrations stabilized to basal levels after approximately 2 h. Intracortical infusion of 200 ng/ml corticosterone evoked transient increases in 5-HT release of approximately 400%. The effect of corticosterone on 5-HT overflow appears to be dose dependent as 20 ng/ml stimulated an increase of 200%, whereas 2 ng/ml stimulated a 50% increase. Administration of 0.1 and 1 ng/ml GABA via the dialysis probe significantly inhibited 5-HT overflow by 20 and 40%, respectively. The duration of GABA inhibition is greater than the stimulatory response for glucocorticoids. Short-lived glucocorticoid stimulation of 5-HT release suggests a possible mechanism for endocrine mediation of continuously changing social behavioral events. 相似文献
127.
Technical Advance: a novel technique for the sensitive quantification of acyl CoA esters from plant tissues 总被引:9,自引:0,他引:9
We report a novel, highly sensitive and selective method for the extraction and quantification of acyl CoA esters from plant tissues. The method detects acyl CoA esters with acyl chain lengths from C4 to C20 down to concentrations as low as 6 fmol in extracts. Acyl CoA esters from standard solutions or plant extracts were derived to their fluorescent acyl etheno CoA esters in the presence of chloroacetaldehyde, separated by ion-paired reversed-phase high-performance liquid chromatography, and detected fluorometrically. This derivitization procedure circumvents the selectivity problems associated with previously published enzymatic methods, and methods that rely on acyl chain or thiol group modification for acyl CoA ester detection. The formation of acyl etheno CoA esters was verified by mass spectrometry, which was also used to identify unknown peaks from chromatograms of plant extracts. Using this method, we report the composition and concentration of the acyl CoA pool during lipid synthesis in maturing Brassica napus seeds and during storage lipid breakdown in 2-day-old Arabidopsis thaliana seedlings. The concentrations measured were in the 3--6 microM range for both tissue types. We also demonstrate the utility of acyl CoA profiling in a transgenic B. napus line that has high levels of lauric acid. To our knowledge, this is the first time that reliable estimates of acyl CoA ester concentrations have been made for higher plants, and the ability to profile these metabolites provides a valuable new tool for the investigation of gene function. 相似文献
128.
A dominant plant of the California grasslands, purple needlegrass [Nassella pulchra (Hitchc.) Barkworth] is an important revegetation species in its native range. The amplified fragment length polymorphism (AFLP) method was used to elucidate mode of reproduction and nucleotide variation among 11 natural populations and three selected natural germplasm releases of N. pulchra. A total of 12 co-dominant AFLPs, informative within eight populations, failed to reveal any heterozygous individuals, indicating very high selfing rates (S(H)=1). Estimates of nucleotide diversity within populations ranged from 0 to 0.00069 (0.00035 average), whereas the total nucleotide divergence among populations ranged from 0.00107 to 0.00382 (0.00247 average). Measures of population differentiation (GS) in terms of Shannon-Weaver diversity values and estimated nucleotide substitutions were 0.90 and 0.86, respectively. Although some of the sample populations contained a mixture of true breeding genotypes, most populations could be distinguished unambiguously. Moreover, geographical distance between the natural source populations was significantly correlated with genetic distance (r = 0.60) among the corresponding sample populations. Results indicate that inbreeding, combined with founder effects and/or selection, has contributed to the differentiation of N. pulchra populations. Foundation seed populations of the selected natural germplasm releases were genetically well defined and most similar to natural seed collected near the corresponding source populations. Thus, these commercial germplasm sources will be made practically available and useful for conservation plantings within the intended areas of utilization. 相似文献
129.
130.
Macey JR Strasburg JL Brisson JA Vredenburg VT Jennings M Larson A 《Molecular phylogenetics and evolution》2001,19(1):131-143
Phylogenetic relationships among frogs of the genus Rana from western North America are investigated using 2013 aligned bases of mitochondrial DNA sequence from the genes encoding ND1 (subunit one of NADH dehydrogenase), tRNA(Ile), tRNA(Gln), tRNA(Met), ND2, tRNA(Trp), tRNA(Ala), tRNA(Asn), tRNA(Cys), tRNA(Tyr), and COI (subunit I of cytochrome c oxidase), plus the origin for light-strand replication (O(L)) between the tRNA(Asn) and tRNA(Cys) genes. The aligned sequences contain 401 phylogenetically informative characters. A well-resolved phylogenetic hypothesis in which the Rana boylii species group (R. aurora, R. boylii, R. cascadae, R. muscosa, and R. pretiosa) is monophyletic is obtained. Molecular sequence divergence suggests that the R. boylii species group is approximately 8 million years old. The traditional hypothesis showing monophyly of the yellow-legged frogs (R. boylii and R. muscosa) is statistically rejected in favor of a hypothesis in which R. aurora, R. cascadae, and R. muscosa form a clade. Reanalyses of published nuclear ribosomal DNA restriction-site data and allozymic data support a monophyletic R. boylii group, but do not effectively resolve relationships among species within this group. Eight populations of R. muscosa form two major clades separated by a biogeographic break in the Sierra Nevada of California. This biogeographic break is broadly concordant with breaks found in four other amphibian and reptilian taxa. The two major clades within R. muscosa are estimated to have diverged approximately 2.2 million years before present. Each of these major clades contains two subgroups showing approximately 1.5 million years divergence, implicating climatic effects of Pleistocene glaciation in vicariance. The four distinct subgroups of R. muscosa separated by at least 1.4 million years of evolutionary divergence are suggested as potential units for conservation. 相似文献