Properties of a Deoxyribonucleic Acid Ligase Mutant of Escherichia coli: X-Ray Sensitivity

A deoxyribonucleic acid ligase-deficient mutant is X-ray sensitive relative to the parent strain, suggesting that deoxyribonucleic acid ligase functions in repair of X-ray-induced, single-strand scissions.     

A novel human polycomb binding site acts as a functional polycomb response element in Drosophila

Polycomb group (PcG) proteins are key chromatin regulators implicated in multiple processes including embryonic development, tissue homeostasis, genomic imprinting, X-chromosome inactivation, and germ cell differentiation. The PcG proteins recognize target genomic loci through cis DNA sequences known as Polycomb Response Elements (PREs), which are well characterized in Drosophila. However, mammalian PREs have been elusive until two groups reported putative mammalian PREs recently. Consistent with the existence of mammalian PREs, here we report the identification and characterization of a potential PRE from human T cells. The putative human PRE has enriched binding of PcG proteins, and such binding is dependent on a key PcG component SUZ12. We demonstrate that the putative human PRE carries both genetic and molecular features of Drosophila PRE in transgenic flies, implying that not only the trans PcG proteins but also certain features of the cis PREs are conserved between mammals and Drosophila.     

CSF T-Tau/Aβ42 predicts white matter microstructure in healthy adults at risk for Alzheimer's disease

Cerebrospinal fluid (CSF) biomarkers T-Tau and Aβ(42) are linked with Alzheimer's disease (AD), yet little is known about the relationship between CSF biomarkers and structural brain alteration in healthy adults. In this study we examined the extent to which AD biomarkers measured in CSF predict brain microstructure indexed by diffusion tensor imaging (DTI) and volume indexed by T1-weighted imaging. Forty-three middle-aged adults with parental family history of AD received baseline lumbar puncture and MRI approximately 3.5 years later. Voxel-wise image analysis methods were used to test whether baseline CSF Aβ(42), total tau (T-Tau), phosphorylated tau (P-Tau) and neurofilament light protein predicted brain microstructure as indexed by DTI and gray matter volume indexed by T1-weighted imaging. T-Tau and T-Tau/Aβ(42) were widely correlated with indices of brain microstructure (mean, axial, and radial diffusivity), notably in white matter regions adjacent to gray matter structures affected in the earliest stages of AD. None of the CSF biomarkers were related to gray matter volume. Elevated P-Tau and P-Tau/Aβ(42) levels were associated with lower recognition performance on the Rey Auditory Verbal Learning Test. Overall, the results suggest that CSF biomarkers are related to brain microstructure in healthy adults with elevated risk of developing AD. Furthermore, the results clearly suggest that early pathological changes in AD can be detected with DTI and occur not only in cortex, but also in white matter.     

Methods for Studying the Ecological Physiology of Feeding in Free-Ranging Howlers (Alouatta palliata) at La Pacifica, Costa Rica

We lack a general understanding of how primates perform physiologically during feeding to cope with the challenges of their natural environments. We here discuss several methods for studying the ecological physiology of feeding in mantled howlers (Alouatta palliata) at La Pacifica, Costa Rica. Our initial physiological effort focuses on recording electromyographic activity (EMG) from the jaw muscles in free-ranging howlers while they feed in their natural forest habitat. We integrate these EMG data with measurements of food material properties, dental wear rates, as well as spatial analyses of resource use and food distribution. Future work will focus on incorporating physiological measures of bone deformation, i.e., bone strain; temperatures; food nutritional data; and hormonal analyses. Collectively, these efforts will help us to better understand the challenges that howlers face in their environment and the physiological mechanisms they employ during feeding. Our initial efforts provide a proof of concept demonstrating the methodological feasibility of studying the physiology of feeding in free-ranging primates. Although howlers offer certain advantages to in vivo field research, many of the approaches described here can be applied to other primates in natural habitats. By collecting physiological data simultaneously with ecological and behavioral data, we will promote a more synthetic understanding of primate feeding and its evolutionary history.     

Prion protein expression differences in microglia and astroglia influence scrapie-induced neurodegeneration in the retina and brain of transgenic mice

Activated microglia and astroglia are known to be involved in a variety of neurodegenerative diseases, including prion diseases. In the present experiments, we studied activation of astroglia and microglia after intraocular scrapie infection in transgenic mice expressing prion protein (PrP) in multiple cell types (tg7 mice) or in neurons only (tgNSE mice). In this model, scrapie infection and protease-resistant PrP deposition occurs in the retinas of both strains of mice, but retinal degeneration is observed only in tg7 mice. Our results showed that the retinas of tg7 and tgNSE mice both had astroglial activation with increased chemokine expression during the course of infection. However, only tg7 retinas exhibited strong microglial activation compared to tgNSE retinas, which showed little microglial activation by biochemical or morphological criteria. Therefore, microglial PrP expression might be required for scrapie-induced retinal microglial activation and damage. Furthermore, microglial activation preceded retinal neurodegeneration in tg7 mice, suggesting that activated microglia might contribute to the degenerative process, rather than being a response to the damage. Surprisingly, brain differed from retina in that an altered profile of microglial activation markers was upregulated, and the profiles in the two mouse strains were indistinguishable. Microglial activation in the brain was associated with severe brain vacuolation and neurodegeneration, leading to death. Thus, retinal and brain microglia appeared to differ in their requirements for activation, suggesting that different activation pathways occur in the two tissues.     

Effect of lipids on avian satellite cell proliferation, differentiation and heparan sulfate proteoglycan expression

The objective of this study was to determine the effects of fatty acids on the proliferation, differentiation, and expression of syndecan-4 and glypican-1 in avian myogenic satellite cells (SC). SC derived from the pectoralis major (PM) and biceps femoris (BF) muscles of the turkey and chicken were individually administered 8 different fatty acids in defined medium during proliferation. A parallel set of turkey SC was induced to differentiate. Highest levels of proliferation of turkey PM and BF SC occurred in cultures containing oleate. Linoleate and oleate were equipotent in supporting proliferation of chicken SC. Microscopic examination revealed that inclusion of docosahexaenoate or eicosapentaenoate was toxic towards both PM and BF SC from both species. Linolenate and arachidonate diminished levels of differentiation. Expression of glypican-1 varied between treatments to a greater extent with turkey BF than with PM SC. Expression in chicken PM and BF SC demonstrated a similar pattern in response to treatments. Turkey PM syndecan-4 expression varied between treatments, whereas expression in turkey BF SC was similar between treatments. Expression in chicken SC varied little between treatments. The results demonstrate species and muscle-specific differences in the parameters examined. It is proposed that changes in lipid raft receptor interactions may contribute to these observed differences.     

Biologic properties of mesenchymal stem cells derived from bone marrow and adipose tissue

The biologic characteristics of mesenchymal stem cells (MSCs) isolated from two distinct tissues, bone marrow and adipose tissue were evaluated in these studies. MSCs derived from human and non-human primate (rhesus monkey) tissue sources were compared. The data indicate that MSCs isolated from rhesus bone marrow (rBMSCs) and human adipose tissue (hASCs) had more similar biologic properties than MSCs of rhesus adipose tissue (rASCs) and human bone marrow MSCs (hBMSCs). Analyses of in vitro growth kinetics revealed shorter doubling time for rBMSCs and hASCs. rBMSCs and hASCs underwent significantly more population doublings than the other MSCs. MSCs from all sources showed a marked decrease in telomerase activity over extended culture; however, they maintained their mean telomere length. All of the MSCs expressed embryonic stem cell markers, Oct-4, Rex-1, and Sox-2 for at least 10 passages. Early populations of MSCs types showed similar multilineage differentiation capability. However, only the rBMSCs and hASCs retain greater differentiation efficiency at higher passages. Overall in vitro characterization of MSCs from these two species and tissue sources revealed a high level of common biologic properties. However, the results demonstrate clear biologic distinctions, as well.     

oca2 regulation of chromatophore differentiation and number is cell type specific in zebrafish

We characterized a zebrafish mutant that displays defects in melanin synthesis and in the differentiation of melanophores and iridophores of the skin and retinal pigment epithelium. Positional cloning and candidate gene sequencing link this mutation to a 410‐kb region on chromosome 6, containing the oculocutaneous albinism 2 (oca2) gene. Quantification of oca2 mutant melanophores shows a reduction in the number of differentiated melanophores compared with wildtype siblings. Consistent with the analysis of mouse Oca2‐deficient melanocytes, zebrafish mutant melanophores have immature melanosomes which are partially rescued following treatment with vacuolar‐type ATPase inhibitor/cytoplasmic pH modifier, bafilomycin A1. Melanophore‐specific gene expression is detected at the correct time and in anticipated locations. While oca2 zebrafish display unpigmented gaps on the head region of mutants 3 days post‐fertilization, melanoblast quantification indicates that oca2 mutants have the correct number of melanoblasts, suggesting a differentiation defect explains the reduced melanophore number. Unlike melanophores, which are reduced in number in oca2 mutants, differentiated iridophores are present at significantly higher numbers. These data suggest distinct mechanisms for oca2 in establishing differentiated chromatophore number in developing zebrafish.     

Anti-idiotype responses abrogate anti-CD4–induced tolerance to a tumor-specific antigen and promote systemic tumor immunity

Purpose: Immunologic-based cancer treatment modalities represent an active area of investigation. Included in these strategies are passive administration of monoclonal antibodies which recognize tumor-associated antigens and active vaccination with identified tumor antigens. However, several problems associated with these types of treatment strategies have been identified. Methods: In this report, we address certain issues by employing a murine model for experimental pulmonary metastasis and a tumor antigen vaccination strategy that induces complete tumor immunity in this system. Utilizing this model, we attempt to address issues related to unresponsiveness to tumor antigen immunization induced by passive administration of a rat monoclonal anti-CD4 and the induction of anti-idiotype responses to a passively administered monoclonal antibody and the effects on the induction of tumor immunity. Results: The results presented indicate that passive administration of rat monoclonal anti-CD4 exhibits immunosuppressive effects that inhibit the production of antibodies to the tumor antigen immunization and abolishes tumor immunity. Repeated administration of the rat monoclonal anti-CD4 results in an anti-idiotype response that can abrogate unresponsiveness to tumor antigen immunization and promote systemic tumor immunity. Conclusions: The data examine a number of potential problems associated with immunologic-based treatments for cancer. These problems include the potential for tolerance to the tumor antigen and establishing an immunocompromised state where immunization with a tumor antigen failed to generate tumor immunity. Approaches to eliminate tolerant T cells by targeting anti-CD4 via anti-idiotype responses that could be generated in vivo without CD4⁺ T cells allowed for recovery of nontolerant T cells, and an antibody response to the tumor antigen that results in tumor immunity.Abbreviations CTL Cytotoxic T lymphocyte - FITC Fluorescein isothiocyanate - OD Optical density - PBS Phosphate-buffered saline - SV40 Simian virus 40