Human papillomavirus infection and cervical intraepithelial neoplasia in women with renal allografts.

An increased prevalence of cervical cancer has been observed in immunosuppressed women, but controlled studies are rare. Biopsy specimens from 49 women with renal allografts and 69 non-immunosuppressed controls (with no history of cervical intraepithelial neoplasia, vulval warts, or abnormal results of cervical smear tests) were assessed for colposcopic appearance, cytological and histological diagnosis, and the presence of human papillomavirus types 6/11 and 16/18 DNA sequences. At colposcopy 26 (53%) of the women with allografts had cervical abnormalities compared with 20 (29%) of the controls. The prevalence of cervical intraepithelial neoplasia was significantly higher in the women with allografts (24 (49%) compared with 7 (10%]. The overall rate of detection of human papillomavirus DNA did not differ significantly between the two groups. There was however, a significant difference in the rate of detection of human papillomavirus type 16/18 DNA (27% in the women with allografts and 6% in the controls). These data confirm that pathological and virological changes affecting the cervix are significantly increased in immunosuppressed women and emphasise the need for regular colposcopic examination.     

Isolation and characterization of plasma membrane-associated cortical granules from sea urchin eggs

Cortical granules, which are specialized secretory organelles found in ova of many organisms, have been isolated from the eggs of the sea urchins Arbacia punctulata and Strongylocentrtus pupuratus by a simple, rapid procedure. Electron micropscope examination of cortical granules prepared by this procedure reveals that they are tightly attached to large segments of the plasma membrane and its associated vitelline layer. Further evidence that he cortical granules were associated with these cell surface layers was obtained by (125)I-labeling techniques. The cortical granule preparations were found to be rich in proteoesterase, which was purified 32-fold over that detected in a crude homogenate. Similarly, the specific radioactivity of a (125)I-labeled, surface glycoprotein was increased 40-fold. These facts, coupled with electron microscope observations, indicate the isolation procedure yields a preparation in which both the cortical granules and the plasma membrane-vitelline layer are purified to the same extent. Gel electrophoresis of the membrane-associated cortical granule preparation reveals the presence of at least eight polypeptides. The major polypeptide, which is a glycotprotein of apparent mol wt of 100,000, contains most of the radioactivity introduced by (125)I-labeling of the intact eggs. Lysis of the cortical granules is observed under hypotonic conditions, or under isotonic conditions if Ca(2+) ion is present. When lysis is under isotonic conditions is induced by addition of Ca(2+) ion, the electron-dense contents of the granules remain insoluble. In contrast, hypotonic lysis results in release of the contents of the granule in a soluble form. However, in both cases the (125)I-labeled glycoprotein remains insoluble, presumably because it is a component of either the plasma membrane or the vitelline layer. All these findings indicate that, using this purified preparation, it should be possible to carry out in vitro studies to better define some of the initial, surface-related events observed in vivo upon fertilization.     

Iron content correlates with peroxidase activity in cysteamine-induced astroglial organelles.

A subpopulation of astrocytes in periventricular brain regions and in cysteamine-treated neuroglial cultures contains cytoplasmic granules that exhibit an affinity for Gomori stains, orange-red autofluorescence, and non-enzymatic peroxidase activity. The autofluorescence and pseudoperoxidase activity are consistent with the presence of porphyrins and heme iron, respectively. In the present study, we employed diaminobenzidine cytochemistry, transmission electron microscopy, and energy-dispersive X-ray microanalysis (electron microprobe) in an attempt to correlate fine structure with the peroxidase activity and elemental composition of the cysteamine-induced inclusions in cultured astrocytes. In osmicated preparations, these membrane-bound inclusions varied greatly in size, were round or ovoid in shape, and exhibited an intensely electron-dense granular matrix. In non-osmicated preparations, many inclusions exhibited internal membranous partitions producing complex subcompartmentalization. Diaminobenzidine reaction product, indicative of endogenous peroxidase activity, was occasionally observed distributed diffusely throughout the granule matrix. More commonly, peroxidase activity was restricted to specific intraorganellar compartments. Elemental iron was detected in the inclusions by electron microprobe analysis. The presence and concentration of iron in these organelles correlated closely with the presence and intensity of diaminobenzidine staining, suggesting that redox-active iron mediates the pseudoperoxidase reactions in these cells. Cysteamine-induced derangements of porphyrin-heme biosynthesis may be responsible for the proliferation of iron-containing gliosomes in these astrocytes.     

Localization of phosphatases in Trypanosoma rhodesiense

Phosphatase activity in Trypanosoma rhodesiense has been examined histochemically by light and electron microscopy and by enzymatic assay in homogenate fractions. Using a method with lead as capture ion, acid phosphatase was found in lysosome-like vesicles and in the flagellar pocket. No alkaline adenosine triphosphatase (ATPase) was detectable by this method. Direct assay of p-nitrophenylphosphatase activity in homogenate fractions showed that acid phosphatase activity was strongly membrane-bound, but that activity at pH 9 was minimal in both soluble and particulate fractions. "Endogenous" ATPase activity was localized specifically and reproducibly in the mitochondrial membranes and under the plasma membrane of he flagellum. This nonenzymic reaction product could not be eradicated by glycerol extraction or glucose depletion. Unlike the membrane staining, which was manifest only after lead treatment, heat-resistant electron-dense material was found in the matrix of lysosomal vesicles in trypanosomes fixed in glutaraldehyde only and not subjected to further treatment with heavy metal reagents. X-ray emission analysis showed the presence of calcium and phosphorus, indicating that the matrix might have a phosphate storage function.     

Aggregation-dependent turnover of flagellar adhesion molecules in chlamydomonas gametes

Previous studies on flagellar adhesion in chlamydomonas (Snell, W. and S. Roseman. 1979. J. Biol. Chem. 254:10820-10829.) have shown that as gametes adhere to flagella isolated from gametes of the opposite mating type, the adhsiveness of the added flagella but not of the gametes is lost. The studies reported here show that the addition of protein synthesis inhibitors (cycloheximide [CH] or anisomycin) to the medium of such cell- flagella mixtures causes the cells to lose their adhesiveness. This loss, however, occurs only after the cells have interacted with 4-8 flagella/cell and does not occur if the cells are kept in CH (7 h) without aggregating. The availability of an impotent (imp) mating type plus (MT(+)) mutant (provided by U.W. Goodenough), which adheres but is unable to undergo the fusion that normally follows adhesion, made it possible to determine whether a similar loss of adhesiveness occurs in mixtures of matting type minus (mt(-)) and imp mt(+) gametes. In the absence of inhibitor, mt(-) and imp mt(+) gametes adhered to each other (without fusing) for several hours; however, in the presence of CH or anisomycin, the gametes began to de-adhere 35 min after mixing, and, by 90 min, 100 percent of the cells were single again. This effect was reversible, and the rapid turnover of cells were single again. This effect was reversible, and the rapid turnover of molecules involved in adhesion occurred only during adhesion inasmuch as gametes pretreated for 4 h with CH were able to aggregate in CH for the same length of time as nonpretreated cells aggregated in CH. By the addition of CH at various times after the mt(-) and imp mt(+) gametes were mixed, measurements were made of the “pool size” of the molecules involved in adhesion. The pool reached a minimum after 25 min of aggregation, rapidly increased for the next 25 min, and then leveled off at the premixing level. These results suggest that flagellar adhesion in chlamydomonas causes modification of surface molecules (receptors, ligands), which brings about their inactivation and stimulates their replacement.     

Tracking fatty acid kinetics in distinct lipoprotein fractions in vivo: a novel high-throughput approach for studying dyslipidemia in rodent models

Isotopic tracers have been used to examine lipid trafficking for many years, and data from those studies have typically yielded novel insight regarding the pathophysiology of dyslipidemia. Previous experimental designs were suitable for studies in humans because relatively large volumes of plasma could be regularly sampled. We have expanded on the earlier logic by applying high-throughput analytical methods that require reduced sample volumes. Specifically, we have examined the possibility of coupling gel-based separations of lipoproteins (e.g., lipoprint) with LC-MS/MS analyses of complex lipid mixtures as a way to routinely measure the labeling profiles of distinct lipids in discrete lipoprotein subfractions. We demonstrate the ability to measure the incorporation of [U-¹³C]oleate into triglycerides (TG), PLs (PL), and cholesterol esters (CE) in VLDL, LDL, and HDL particles in mice. Although rodent models of dyslipidemia are inherently different from humans because of alterations in enzyme activities and underlying metabolism, rodent models can be used to screen novel compounds for efficacy in altering a given biochemical pathway and therein enable studies of target engagement in vivo. We expect that it is possible to translate our approach for application in other systems, including studies in humans.     

Modeling Hurricane Exposure in a Caribbean Lower Montane Tropical Wet Forest: The Effects of Frequent,Intermediate Disturbances and Topography on Forest Structural Dynamics and Composition

Frequent intermediate disturbances can produce qualitatively different spatial heterogeneity and environmental variability than large infrequent disturbances, which facilitates the coexistence of disparate species types. We hypothesized that species coexistence will be maximized at sites exposed to recurring hurricanes with intermediate frequencies and effects. Consequently, we sought to determine if exposure vulnerability (EV) from three hurricanes with intermediate effects and frequencies (Ivan-2004, Dennis-2005 and Dean-2007), or the interaction between exposure and topography, could be used to explain forest structural dynamics and composition. We used data obtained in 2006 and 2012 from within 45, 25 × 25 m (2.8125 ha) permanent sample plots, established according to a randomized block design, and stratified according to elevation and aspect/sites (NE and SW facing ridges) in a tropical montane wet forest, John Crow Mountains, Jamaica. There was a significant reduction in basal area (BA) (14%), tree volume (10%), and density (26%) and there was a negative shift in the height profile of trees. Understory light (2008), stem density, and mortality increased with EV but decreased with aspect. The NE aspect/sites had higher EV after Dean and the other hurricanes. Consequently, BA, volume and density increased significantly for light-demanding species at the NE sites, but declined significantly overall with minimal changes at SW sites. Moreover, diversity was significantly higher at sites with higher EV for the three hurricanes. The frequent hurricanes with intermediate effects may have therefore maintained/increased spatial heterogeneity, which promoted the coexistence of species with disparate life histories at more exposed sites.