Jejunal and ileal d-glucose transport in isolated brush border membranes

d-Glucose transport was investigated in isolated brush border membranes from small intestine. The transport properties of membranes from upper jejunum were compared with those from terminal ileum. The jejunal membranes accumulated d-glucose to a greater extent than the ileal membranes when supplied with energy in the form of a NASCN gradient. This difference is behavior is similar to that of the more intact epithelial preparations and suggests that the isolated membranes actually reflect the state present in intact cells. Ileal membranes transported d-glucose about two to three times slower than the jejunal ones, which can partially explain the lower sugar accumulation.     

Viral evolution and interferon resistance of hepatitis C virus RNA replication in a cell culture model

Hepatitis C virus (HCV) replicates through an error-prone process that may support the evolution of genetic variants resistant to the host cell antiviral response and interferon (IFN)-based therapy. We evaluated HCV-IFN interactions within a long-term culture system of Huh7 cell lines harboring different variants of an HCV type 1b subgenomic RNA replicon that differed at only two sites within the NS5A-encoding region. A replicon with a K insertion at HCV codon 2040 replicated efficiently and exhibited sequence stability in the absence of host antiviral pressure. In contrast, a replicon with an L2198S point mutation replicated poorly and triggered a cellular response characterized by IFN-beta production and low-level IFN-stimulated gene (ISG) expression. When maintained in long term-culture, the L2198S RNA evolved into a stable high-passage (HP) variant with six additional point mutations throughout the HCV protein-encoding region that enhanced viral replication. The HP RNA transduced Huh7 cells with more than 1,000-fold greater efficiency than its L2198S progenitor or the K2040 sequence. Replication of the HP RNA resisted suppression by IFN-alpha treatment and was associated with virus-directed reduction in host cell expression of ISG56, an antagonist of HCV RNA translation. Accordingly, the HP RNA was retained within polyribosome complexes in vivo that were refractory to IFN-induced disassembly. These results identify ISG56 as a translational control effector of the host response to HCV and provide direct evidence to link this response to viral sequence evolution, ISG regulation, and selection of the IFN-resistant viral phenotype.     

Regulating intracellular antiviral defense and permissiveness to hepatitis C virus RNA replication through a cellular RNA helicase, RIG-I

Virus-responsive signaling pathways that induce alpha/beta interferon production and engage intracellular immune defenses influence the outcome of many viral infections. The processes that trigger these defenses and their effect upon host permissiveness for specific viral pathogens are not well understood. We show that structured hepatitis C virus (HCV) genomic RNA activates interferon regulatory factor 3 (IRF3), thereby inducing interferon in cultured cells. This response is absent in cells selected for permissiveness for HCV RNA replication. Studies including genetic complementation revealed that permissiveness is due to mutational inactivation of RIG-I, an interferon-inducible cellular DExD/H box RNA helicase. Its helicase domain binds HCV RNA and transduces the activation signal for IRF3 by its caspase recruiting domain homolog. RIG-I is thus a pathogen receptor that regulates cellular permissiveness to HCV replication and, as an interferon-responsive gene, may play a key role in interferon-based therapies for the treatment of HCV infection.     

Increases in summer temperatures decrease the survival of an invasive forest insect

Higher temperatures projected under current climate change models are generally predicted to exert an overall positive effect on the success of invasive insects through increased survivability, developmental rates and fecundity, and by facilitating geographic range expansion. However, these effects have primarily focused on the shifts in winter temperatures with limited attention to the role that summer heat may play in shaping species ranges or fitness. We examined the thermal ecology of an ecologically important invasive forest insect, the hemlock woolly adelgid (Adelges tsugae), by determining survival during its summer dormancy phase under increasing temperature regimens. From laboratory and field experiments, we documented a positive association between increased temperatures and duration of exposure, and A. tsugae mortality. Adelges tsugae mortality was minimal (<20%) when exposed to summer temperatures characteristic to its native range (<25 °C), but markedly increased (up to 100%) when exposed to temperatures that occur occasionally or rarely in natural settings (>30 °C). At the warmest, southernmost edge of their range, field mortality of A. tsugae ranged from 8.5 to 81.9% and was strongly correlated with site temperature regimens. Further, we found no significant differences in A. tsugae survival between populations collected from Maine and Georgia, and over a 3-year period within Georgia, indicating that A. tsugae may not be acclimating to heat. These results highlight the importance of including summer temperatures in studies regarding increased temperatures on insect dynamics, and may alter historical predictions of climate change impacts on invasive insects and the conservation of forest ecosystems.     

Physical Mapping of Human 7q and 14q Subtelomeric DNA Sequences in the Great Apes

Phylogenetic divergence of the members of the Pongidae familyhas been based on genetic evidence. The terminal repeat array(T₂AG₃) has lately been considered as an additional basis toanalyze genomes of highly related species. The recent isolationof subtelomeric DNA probes specific for human (HSA) chromosomes7q and 14q has prompted us to cross-hybridize them to the chromosomesof the chimpanzee (PTR), gorilla (GGO) and orangutan (PPY) tosearch for its equivalent locations in the great ape species.Both probes hybridized to the equivalent telomeric sites ofthe long (q) arms of all three great ape species. Hybridizationsignals to the 7q subtelomeric DNA sequence probe were observedat the telomeres of HSA 7q, PTR 6q, GGO 6q and PPY 10q, whilehybridization signals to the 14q subtelomeric DNA sequence probewere observed at the telomeres of HSA 14q, PTR 15q, GGO 18qand PPY 15q. No hybridization signals to the chromosome 7-specificalpha satellite DNA probe on the centromeric regions of theape chromosomes were observed. Our observations demonstratesequence homology of the subtelomeric repeat families D7S427and D14S308 in the ape chromosomes. An analogous number of subtelomericrepeat units exists in these chromosomes and has been preservedthrough the course of differentiation of the hominoid species.Our investigation also suggests a difference in the number ofalpha satellite DNA repeat units in the equivalent ape chromosomes,possibly derived from interchromosomal transfers and subsequentamplification of ancestral alpha satellite sequences.     

Effectiveness Of Sch1ff Variants In The Periodic-Schiff and Feulgen Nucleal Technics

The presence of additional S02 in Schiff reagents above that required for bleaching of the basic fuchsin has no significant effect on the behavior of such reagents in performing the periodic-Schiff or Feulgen nucleal reactions. Schiff reagents made from aqueous solutions containing less than 0.125% basic fuchsin will not demonstrate in these reactions all the tissue elements stained by stronger solutions. Schiff reagents made from aqueous solutions containing more than 0.5% basic fuchsin will in time precipitate considerable amounts of leucofuchsin. Fresh activated charcoal discriminates much more sharply between the yellow contaminant which occurs in basic fuchsin when the fuchsin has been converted to the leuco state than it does when the fuchsin is in the colored condition. Precipitation of leucofuchsin occurs at pH 3 or slightly above in Schiff reagents of generally useful concentration, but at lower levels down to the pH of 10 N HC1, hydrogen ion concentration does not influence their histochemical effectiveness.     

Natural killer cells in the blood and bone marrow of the rhesus monkey

Natural killer (NK) cells in peripheral blood lymphocytes (PBL) and bone marrow (BM) cells of the rhesus monkey were detected by their functional activity against K562 cells. Animals could be grouped into "high" or "low" NK responders, a trait found to be consistent over a period of 2 years. NK active cells in PBL were in the nonadherent population, with the majority bearing Fc receptors and a further subdivision of these into CR+ (complement receptor) and CR- NK cells. Of 10 monoclonal antibodies directed against different epitopes of human lymphocytes, OKT11, OKT10, and Leu 11 showed reactivity with rhesus NK cells. Only OKT10 was reactive with the effector site of the cell, as shown by its capability to block NK function. Of the Leu 11 monoclonal antibodies (a, b, c), Leu 11c was nonreactive while Leu 11a and Leu 11b were shown by immunofluorescence to bind to 7 to 21% of PBL; Leu 11b was also cytotoxic to the NK cells. Leu 11b did not prevent binding of Leu 11a to PBL, suggesting reactivity of these antibodies with different epitopes. Percoll fractionation of PBL and BM revealed a greater enrichment of NK activity with BM; also, with PBL peak NK activity occurred in fractions 4 and 5 while this occurred in fraction 5 with BM. Although Percoll PBL fractions contained a higher percentage of Leu 11b cells, the NK activity of the BM fractions was proportionately greater. The majority of PBL cells with NK activity were FcR+ while significant activity could be attributed to FcR- cells of BM, in both the unseparated and Percoll fractions of each tissue. The data suggest NK active cells of BM may be distinct from those found in PBL.     

Staining properties of aldehyde fuchsin analogs.

This investigation was designed to clarify the role of the aldehyde component of aldehyde fuchsin in its staining reactions. Several aldehyde fuchsin analogs were prepared by using different aldehydes. The staining quality of these analogs and pararosaniline-HCl was compared with that of aldehyde fuchsin prepared with paraldehyde in the usual way. The major findings of this investigation include: 1) Aldehyde fuchsin staining of nonoxidized pancreatic B cells requires a stain prepared with either paraldehyde or acetaldehyde. 2) An aldehyde moiety is required for aldehyde fuchsin staining of strong tissue anions. 3) Staining of elastic tissue with aldehyde fuchsin analogs resembles staining of strong tissue anions more than staining of nonoxidized pancreatic B cells. Possible reaction mechanisms of aldehyde fuchsin with tissue substrates are discussed.