Phylogenetic relationships among JC virus strains in Japanese/Koreans and Native Americans speaking Amerind or Na-Dene

Many genetic studies using human mtDNA or the Y chromosome have been conducted to elucidate the relationships among the three Native American groups speaking Amerind, Na-Dene, and Eskimo-Aleut. Human polyomavirus JC (JCV) may also help to gain insights into this issue. JCV isolates are classified into more than 10 geographically distinct genotypes (designated subtypes here), which were generated by splits in the three superclusters, Types A, B, and C. A particular subtype of JCV (named MY) belonging to Type B is spread in both Japanese/Koreans and Native Americans speaking Amerind or Na-Dene. In this study, we evaluated the phylogenetic relationships among MY isolates worldwide, using the whole-genome approach, with which a highly reliable phylogeny of JCV isolates can be reconstructed. Thirty-six complete sequences belonging to MY (10 from Japanese/Koreans, 24 from Native Americans, and 2 from others), together with 54 belonging to other subtypes around the world, were aligned and subjected to phylogenetic analysis using the neighbor-joining and maximum-likelihood methods. In the resultant phylogenetic trees, the MY sequences diverged into two Japanese/Korean and five Native American clades with high bootstrap probabilities. Two of the Native American clades contained isolates mainly from Na-Denes and the others contained isolates mainly from Amerinds. The Na-Dene clades were not clustered together, nor were the Amerind clades. In contrast, the two Japanese/Korean clades were clustered at a high bootstrap probability. We concluded that there is no distinction between Amerinds and Na-Denes in terms of indigenous JCVs, although they are linguistically distinguished from each other.     

Assessing the Impact of Disease Vectors on Animal Populations

Many studies have attempted to assess the relative effects of different vectors of a disease on animal populations. To this end, three measures have been proposed: Vectorial efficiency, Vectorial capacity and recently Vectorial effectiveness (or Vectorial impact). In this study we relate these measures to derive some of their properties emphasising in the vectorial impact for its importance in both, population performance of parasites and the proportion of the prevalence of one parasite due to a given vector. We applied the quantitative expressions advanced in this study to a simple Chilean example with one parasite (Trypanosoma cruzi), two vectors (Triatoma infestans and Mepraia spinolai) and one animal population (humans: Chagas's disease).     

The role of lipid-protein interactions in NADH-cytochrome c reductase (rotenone-insensitive) of rat liver mitochondria

The phospholipid depletion of rat liver mitochondria, induced by acetone-extraction or by digestion with phospholipase A₂ or phospholipase C, greatly inhibited the activity of NADH-cytochrome c reductase (rotenone-insensitive). A great decrease of the reductase activity also occurred in isolated outer mitochondrial membranes after incubation with phospholipase A₂. The enzyme activity was almost completely restored by the addition of a mixture of mitochondrial phospholipids to either lipid-deficient mitochondria, or lipid-deficient outer membranes. The individual phospholipids present in the outer mitochondrial membrane induced little or no stimulation of the reductase activity. Egg phosphatidylcholine was the most active phospholipid, but dipalmitoyl phosphatidylcholine was almost ineffective. The lipid depletion of mitochondria resulted in the disappearance of the non-linear Arrhenius plot which characterized the native reductase activity. A non-linear plot almost identical to that of the native enzyme was shown by the enzyme reconstituted with mitochondrial phospholipids. Triton X-100, Tween 80 or sodium deoxycholate induced only a small activation of NADH-cytochrome c reductase (rotenone-insensitive) in lipiddeficient mitochondria. The addition of cholesterol to extracted mitochondrial phospholipids at a 1 : 1 molar ratio inhibited the reactivation of NADH-cytochrome c reductase (rotenone-insensitive) but not the binding of phospholipids to lipid-deficient mitochondria or lipid-deficient outer membranes.These results show that NADH-cytochrome c reductase (rotenone-insensitive) of the outer mitochondrial membrane requires phospholipids for its activity. A mixture of phospholipids accomplishes this requirement better than individual phospholipids or detergents. It also seems that the membrane fluidity may influence the reductase activity.     

The density of long limb bones and the percentage ash weight of the skeleton of Macaca mulatta

The gravimetric density of humeri, radii, femora and tibiae from a series of 274 male and female skeletons of rhesus monkeys, Macaca mulatta, was determined for fetal, young and adult periods. The ages of 171 of the animals were known: they ranged from 57 days of gestation to 13.6 years; the ages of an additional 103 skeletons were estimated. The mean density of the fetal bones was found to increase linearly with age and was higher for males than females, and higher for the superior than for the inferior limb bones. During the young period the pattern of increase in density can be represented by a power-type curve, and the density is significantly higher in females than in males and in superior than in inferior limb bones. The densities of the long limb bones of the adult skeletons show a slight, but not significant, negative trend with increasing age. In this age group the mean densities are higher for males than females and higher for the superior than for the inferior limb bones. The percentage ash weight was determined for the total skeleton and for 21 subdivisions of 23 postnatal skeletons with estimated ages. The skull and long limb bones were found to have higher mean percentage ash weights than the vertebral segments and the sternum. Both the density and the percentage ash weight of the Macaca mulatta skeletons examined exceed those found in our earlier studies of the human skeleton.     

The effect of light-dark adaptation on the ultrastructure ofLimulus lateral eye retinular cells

Summary The fine structure of retinular cells within lateral eyes ofLimulus polyphemus which had been dark or light adapted for 12 h in vivo was studied via electron microscopy. The ommatidium to ommatidium and retinular cell to retinular cell variability observed in light microscope studies was confirmed. The rhabdomeric microvilli were longer and narrower, the area of contiguous microvillar membranes greater, the endoplasmic reticulum less abundant and the mitochondrial granules (? calcium containing) more numerous in well dark adapted than in well light adapted retinular cells (Figs. 1, 3, 4, 7, 8) and membrane whorls or vacuoles were present in the peripheral cytoplasm of very well light adapted retinular cells (Fig. 6). Phagocytotic vesicles, multivesicular bodies and lysosomes were present in the interrhabdomeral cytoplasm of partially light adapted retinular cells (Figs. 1, 2, 3, 10). The number of retinular cell microvilli in contact with the eccentric cell dendrite was smaller in very well light adapted than in well dark adapted ommatidia (Fig. 9). The possible functional significance of these light-dependent structural changes is discussed.This investigation was supported in part by Grant 2 RO1 EY 00236 National Eye Institute, National Institutes of HealthMember of the SFB 160 of the Deutsche Forschungsgemeinschaft     

Utilization of Chondroitin Sulfate by Bacteroides thetaiotaomicron Growing in Carbohydrate-Limited Continuous Culture

When Bacteroides thetaiotaomicron, an obligate anaerobe from the human colonic flora, was grown in continuous culture with the mucopolysaccharide chondroitin sulfate as the limiting source of carbohydrate, growth yields ranged from 48 g of cell dry weight per mol of equivalent monosaccharide at a growth rate of 3.5 h per generation to 32 g per mol at a growth rate of 24 h per generation. The theoretical maximum growth yield (61 g of cell dry weight per mol of equivalent monosaccharide) was comparable to that of 54 g per mol, which was obtained previously when glucuronic acid, a component of chondroitin sulfate, was the limiting carbohydrate (S. F. Kotarski and A. A. Salyers, J. Bacteriol. 146:853-860, 1981). However, the maintenance coefficient was three times higher when chondroitin sulfate was the substrate than when glucuronic acid was the substrate. The specific activity of chondroitin lyase (EC 4.2.2.4), an enzyme which cleaves chondroitin sulfate into disaccharides, declined by nearly 50% as growth rates decreased from 3.5 to 24 h per generation. By contrast, the specific activities of several glycolytic enzymes and disaccharidases remained constant over this range of growth rates. Although chondroitin sulfate was growth limiting, some carbohydrate was detectable in the extracellular fluid at all growth rates. At rapid growth rates (1 to 2 h per generation), this residual carbohydrate included fragments of chondroitin sulfate having a wide range of molecular weights. At slower growth rates (2 to 24 h per generation), the residual carbohydrate consisted mainly of a small fragment which migrated on paper chromatograms more slowly than the disaccharides produced by chondroitin lyase but faster than a tetrasaccharide. This small fragment may represent the reducing end of the chondroitin sulfate molecule.     

Estrogen stimulation of ovarian surface epithelial cell proliferation

Summary Ovarian cancer is the leading cause of gynecological cancer mortality, and 85–90% of this malignancy originates from the ovarian surface epithelium (OSE). The etiology of ovarian epithelial cancer is unknown but a role for estrogens has been suspected. However, the effect of estrogens on OSE cell proliferation remains to be determined. Using the rabbit model, our studies have demonstrated that 17β-estradiol stimulates OSE cell proliferation and the formation of a papillary ovarian surface morphology similar to that seen in human ovarian serous neoplasms of low malignant potential. Immunohistochemical staining of ovarian tissue sections with an antibody to the estrogen receptor α demonstrates its expression in both OSE cells and stromal interstitial cells. In primary ovarian cell cultures, the proliferative response of the epithelial cells to 17β-estradiol depends on the expression of the estrogen receptor α in the epithelial cells. However, when the epithelial cells are grown together with ovarian stromal cells, their proliferative response to this hormone is greatly enhanced, suggesting the involvement of stromal-epithelial interactions. These studies suggest a role for estrogens and the estrogen receptor α in OSE growth.     

Dynamics of hydrogen-deuterium exchange in Chlamydomonas centrin

Chlamydomonas reinhardtii centrin is a 169-amino acid residue calcium binding protein belonging to the EF-hand protein superfamily. Centrin is associated with the microtubule organizing center (MTOC) in all eukaryotes, and in Chlamydomonas, centrin is a component of the flagellar basal body apparatus. Recombinant full-length centrin, calmodulin, and terminal domain fragments [Ccen-N (residues 1-94) and Ccen-C (residues 99-169)] were used to examine hydrogen-deuterium (H --> D) exchange dynamics using combined attenuated total reflectance (ATR) Fourier transform-infrared (FT-IR) spectroscopy, curve fit, and two-dimensional correlation analysis. Analysis of the Ccen-N and Ccen-C fragments allowed separation of domain specific solvent exchange events and together with analysis of the full-length proteins provides novel insight into domain accessibility to the aqueous environment and the internal dynamics of the protein.     

Beta-catenin regulates differentiation of respiratory epithelial cells in vivo

An activated form of beta-catenin [Catnb(Delta(ex3))] was expressed in respiratory epithelial cells of the developing lung. Although morphogenesis was not altered at birth, air space enlargement and epithelial cell dysplasia were observed in the early postnatal period and persisted into adulthood. The Catnb(Delta(ex3)) protein caused squamous, cuboidal, and goblet cell dysplasia in intrapulmonary conducting airways. Atypical epithelial cells that stained for surfactant pro protein C (pro-SP-C) and had morphological characteristics of alveolar type II cells were observed in bronchioles of the transgenic mice. Catnb(Delta(ex3)) inhibited expression of Foxa2 and caused goblet cell hyperplasia associated with increased staining for mucins and the MUC5A/C protein. In vitro, both wild type and activated beta-catenin negatively regulated the expression of the Foxa2 promoter. Catnb(Delta(ex3)) also caused pulmonary tumors in adult mice. Activation of beta-catenin caused ectopic differentiation of alveolar type II-like cells in conducting airways, goblet cell hyperplasia, and air space enlargement, demonstrating a critical role for the Wnt/beta-catenin signal transduction pathway in the differentiation of the respiratory epithelium in the postnatal lung.     

Enhanced potency of plasmid DNA microparticle human immunodeficiency virus vaccines in rhesus macaques by using a priming-boosting regimen with recombinant proteins

DNA vaccines have been used widely in experimental primate models of human immunodeficiency virus (HIV), but their effectiveness has been limited. In this study, we evaluated three technologies for increasing the potency of DNA vaccines in rhesus macaques. These included DNA encoding Sindbis virus RNA replicons (pSINCP), cationic poly(lactide-co-glycolide) (PLG) microparticles for DNA delivery, and recombinant protein boosting. The DNA-based pSINCP replicon vaccines encoding HIV Gag and Env were approximately equal in potency to human cytomegalovirus (CMV) promoter-driven conventional DNA vaccines (pCMV). The PLG microparticle DNA delivery system was particularly effective at enhancing antibody responses induced by both pCMV and pSINCP vaccines and had less effect on T cells. Recombinant Gag and Env protein boosting elicited rapid and strong recall responses, in some cases to levels exceeding those seen after DNA or DNA/PLG priming. Of note, Env protein boosting induced serum-neutralizing antibodies and increased frequencies of gamma interferon-producing CD4 T cells severalfold. Thus, PLG microparticles are an effective means of delivering DNA vaccines in nonhuman primates, as demonstrated for two different types of DNA vaccines encoding two different antigens, and are compatible for use with DNA prime-protein boost regimens.