A differential scanning calorimetric study of Newcastle disease virus: identification of proteins involved in thermal transitions

The spatial and temporal pattern of expression of enhancer trap lines reporting on the wingless (wg) and engrailed (en) genes was characterized in the adult antenna of Drosophila melanogaster. The time courses of expression seen for wg and en, although different from each other, reveal a complex well-controlled pattern of temporal expression, providing evidence that regulatory mechanisms are preserved throughout the life span of the adult fly. Altering the life span demonstrates that the temporal patterns of expression of both wg and en are linked to life span. These studies suggest that the expression of wg and en in the adult antenna is controlled by age-dependent mechanisms.     

Effect of thrombolytic therapy on platelet expression and plasma concentration of PECAM-1 (CD31) in patients with acute myocardial infarction

OBJECTIVE: Cholelithiasis is a common problem in hospitals of the Peruvian Andes; however, its prevalence in Andean communities is unknown. To estimate the prevalence of gallstone disease in this locale, we conducted a cross-sectional community study in three high-altitude Peruvian rural villages (i.e., > 3000 m above sea level). METHODS: We examined 911 volunteers > 15 yr of age from three villages for gallstone disease by history and ultrasonography. Risk factors for gallstone disease were examined in 382 volunteers from one village. RESULTS: The age-adjusted prevalence of gallstone disease ranged from 4-10% in men and from 18-20% in women. Women had significantly higher age-adjusted prevalence rates than did men. The prevalence of gallstone disease increased significantly with age and decreased significantly with alcohol consumption. Although not statistically significant, we found a positive association between gallstone disease and body mass index. CONCLUSION: The results of this study indicate that gallstone disease, commonly perceived as a disease of the developed world, is also a common problem in high-altitude Peruvian communities.     

Developmentally regulated expression of persyn, a member of the synuclein family, in skin

Synucleins constitute a group of unique, evolutionarily conserved proteins that are expressed predominantly in neurons of the central and peripheral nervous system. Although the normal cellular functions of synucleins are not clear, these proteins have been implicated in various neurodegenerative conditions in humans. We found that persyn, a recently characterized member of the synuclein family, is expressed not only in the nervous system but also in the stratum granulosum of the epidermis of neonatal and adult mice. This finding together with our recent observations that persyn influences neurofilament network integrity in sensory neurons raises the possibility that persyn in skin could be involved in modulation of the keratin network.     

Identification of seven Treponema species in health- and disease-associated dental plaque by nested PCR

Species-specific nested PCR was used to detect Treponema amylovorum, Treponema denticola, Treponema maltophilum, Treponema medium, Treponema pectinovorum, Treponema socranskii, and Treponema vincentii in dental plaque. Subjects with periodontitis harbored all species, but T. pectinovorum and T. vincentii were not found in plaque from disease-free subjects.     

Generation of radiation-induced deletion complexes in the mouse genome using embryonic stem cells

As the genetic and physical mapping stage of the Human Genome Project nears completion, the focus is shifting toward the development of technologies for high-throughput analysis of gene function. Whereas DNA sequencing will enable the assignment of presumed function to a large number of genes in mice and humans, it is clear that the great majority of genes will have to be evaluated in vivo to accurately assess their role in a complex organism. While gene targeting in mouse embryonic stem (ES) cells is the current method of choice for the characterization of gene function in mice, it remains relatively labor intensive and lacks the throughput required for analysis of genome function on a large scale. Alternative methods of efficient mutagenesis will clearly be required for this task. Chromosomal deletions are powerful tools in the genetic analysis of complex genomes, enabling the systematic identification and localization of functional units along defined chromosomal regions. Not only are deletions useful for the identification of genetic functions, but they serve as mapping reagents for existing mutations or traits. While their use has been an essential tool in Drosophila genetics, classical mutagenesis in mice has been logistically impractical for generating deletions. We have previously described an efficient method for generating radiation-induced deletion complexes at defined regions in the genome using ES cells. In this article, we detail the methodological aspects of this technology and describe the applications of chromosomal deletions for characterizing gene function in ways that make optimal use of the information generated by the first stage of the Genome Project.     

Distribution of transposable elements in neotropical species of Drosophila

The phylogenetic distribution of transposable families, P, gypsy, hobo, I, and mariner has been analyzed in 33 species of 11 groups of neotropical Drosophila and a Drosophilidae species Zygotrica vittimaculosa, using squash blot and dot blot. Genomic DNA of almost all neotropical species tested hybridized with gypsy probe and some species showed a particularly strong hybridization signal, as D. gaucha, D. virilis, and species of flavopilosa group. The hobo element was restricted to melanogaster group and some strains of D. willistoni. Only D. simulans DNA showed hybridization to mariner probe in all species tested and D. simulans and D. melanogaster showed hybridization with I element probe. P element homologous sequence was present in D. melanogaster and all species and strains of the willistoni and saltans groups tested. The presence of at least one P-homologous sequence was detected in Drosophila mediopunctata. This one was the only P-bearing species of all six tested from the tripunctata group. Four different pairs of primers homologous to segments of the canonical sequence of D. melanogaster's P were used to amplify specific sequences from D. mediopunctata DNA, showing the occurrence of seemingly well-conserved P-homologous sequences.     

Ricin A-chain: kinetics, mechanism, and RNA stem-loop inhibitors

Ricin A-chain (RTA) catalyzes the depurination of a single adenine at position 4324 of 28S rRNA in a N-ribohydrolase reaction. The mechanism and specificity for RTA are examined using RNA stem-loop structures of 10-18 nucleotides which contain the required substrate motif, a GAGA tetraloop. At the optimal pH near 4.0, the preferred substrate is a 14-base stem-loop RNA which is hydrolyzed at 219 min-1 with a kcat/Km of 4.5 x 10(5) M-1 s-1 under conditions of steady-state catalysis. Smaller or larger stem-loop RNAs have lower kcat values, but all have Km values of approximately 5 microM. Both the 10- and 18-base substrates have kcat/Km near 10(4) M-1 s-1. Covalent cross-linking of the stem has a small effect on the kinetic parameters. Stem-loop DNA (10 bases) of the same sequence is also a substrate with a kcat/Km of 0.1 that for RNA. Chemical mechanisms for enzymatic RNA depurination reactions include leaving group activation, stabilization of a ribooxocarbenium transition state, a covalent enzyme-ribosyl intermediate, and ionization of the 2'-hydroxyl. A stem-loop RNA with p-nitrophenyl O-riboside at the depurination site is not a substrate, but binds tightly to the enzyme (Ki = 0.34 microM), consistent with a catalytic mechanism of leaving group activation. The substrate activity of stem-loop DNA eliminates ionization of the 2'-hydroxyl as a mechanism. Incorporation of the C-riboside formycin A at the depurination site provides an increased pKa of the adenine analogue at N7. Binding of this analogue (Ki = 9.4 microM) is weaker than substrate which indicates that the altered pKa at this position is not an important feature of transition state recognition. Stem-loop RNA with phenyliminoribitol at the depurination site increases the affinity substantially (Ki = 0.18 microM). The results are consistent with catalysis occurring by leaving group protonation at ring position(s) other than N7 leading to a ribooxocarbenium ion transition state. Small stem-loop RNAs have been identified with substrate activity within an order of magnitude of that reported for intact ribosomes.     

Microvascular injection studies in rubeosis iridis and neovascular glaucoma

Microvascular silicone injection, tissue clearing, and histologic examination were used to demonstrate the origin, distribution, and interconnections of newly formed iris and chamber angle blood vessels in four eyes with rubeosis iridis and neovascular glaucoma associated with diabetic retinopathy and central retinal vessel occlusion. The newly formed iris vessels that formed either a tight, evenly distributed (diabetic) or loose, irregularly distributed (central vessel occlusion) network in the iris originated from the normal iris arteries that were branches of either the major arterial circle or of the perforating branches of the anterior ciliary arteries, and drained into the normal iris and ciliary body veins and occasionally into the paralimbal episcleral veins. These newly formed iris vessels appeared to shunt intravascular fluid from arteries to veins. The newly formed anterior chamber angle vessels that formed tufts and arcades at the trabecular meshwork also originated from the roots of the iris arteries and the ciliary body arteries and connected with the peripheral neovascular iris network. In addition, the circumferentially running angle vessels that coursed within the trabecular meshwork branched into and coursed within a fibrosed Schlemm's canal and into two of its intrascleral collector channels. No open communication between these newly formed vessels and the Schlemm's canal-aqueous outflow system was seen.