Recording lifetime behavior and movement in an invertebrate model

Characterization of lifetime behavioral changes is essential for understanding aging and aging-related diseases. However, such studies are scarce partly due to the lack of efficient tools. Here we describe and provide proof of concept for a stereo vision system that classifies and sequentially records at an extremely fine scale six different behaviors (resting, micro-movement, walking, flying, feeding and drinking) and the within-cage (3D) location of individual tephritid fruit flies by time-of-day throughout their lives. Using flies fed on two different diets, full sugar-yeast and sugar-only diets, we report for the first time their behavioral changes throughout their lives at a high resolution. We have found that the daily activity peaks at the age of 15-20 days and then gradually declines with age for flies on both diets. However, the overall daily activity is higher for flies on sugar-only diet than those on the full diet. Flies on sugar-only diet show a stronger diurnal localization pattern with higher preference to staying on the top of the cage during the period of light-off when compared to flies on the full diet. Clustering analyses of age-specific behavior patterns reveal three distinct young, middle-aged and old clusters for flies on each of the two diets. The middle-aged groups for flies on sugar-only diet consist of much younger age groups when compared to flies on full diet. This technology provides research opportunities for using a behavioral informatics approach for understanding different ways in which behavior, movement, and aging in model organisms are mutually affecting.     

14-3-3 σ expression effects G2/M response to oxygen and correlates with ovarian cancer metastasis

Background

In vitro cell culture experiments with primary cells have reported that cell proliferation is retarded in the presence of ambient compared to physiological O₂ levels. Cancer is primarily a disease of aberrant cell proliferation, therefore, studying cancer cells grown under ambient O₂ may be undesirable. To understand better the impact of O₂ on the propagation of cancer cells in vitro, we compared the growth potential of a panel of ovarian cancer cell lines under ambient (21%) or physiological (3%) O₂.

Principal Findings

Our observations demonstrate that similar to primary cells, many cancer cells maintain an inherent sensitivity to O₂, but some display insensitivity to changes in O₂ concentration. Further analysis revealed an association between defective G2/M cell cycle transition regulation and O₂ insensitivity resultant from overexpression of 14-3-3 σ. Targeting 14-3-3 σ overexpression with RNAi restored O₂ sensitivity in these cell lines. Additionally, we found that metastatic ovarian tumors frequently overexpress 14-3-3 σ, which in conjunction with phosphorylated RB, results in poor prognosis.

Conclusions

Cancer cells show differential proliferative sensitivity to changes in O₂ concentration. Although a direct link between O₂ insensitivity and metastasis was not determined, this investigation showed that an O₂ insensitive phenotype in cancer cells to correlate with metastatic tumor progression.     

Herpes simplex virus type 2, genital ulcers and HIV-1 disease progression in postpartum women

Background

Co-infection with herpes simplex virus type 2 (HSV-2) has been associated with increased HIV-1 RNA levels and immune activation, two predictors of HIV-1 progression. The impact of HSV-2 on clinical outcomes among HIV-1 infected pregnant women is unclear.

Methods

HIV-1 infected pregnant women in Nairobi were enrolled antenatally and HSV-2 serology was obtained. HIV-1 RNA and CD4 count were serially measured for 12–24 months postpartum. Survival analysis using endpoints of death, opportunistic infection (OI), and CD4<200 cells µL, and linear mixed models estimating rate of change of HIV-1 RNA and CD4, were used to determine associations between HSV-2 serostatus and HIV-1 progression.

Results

Among 296 women, 254 (86%) were HSV-2-seropositive. Only 30 (10%) women had prior or current genital ulcer disease (GUD); median baseline CD4 count was 422 cells µL. Adjusting for baseline CD4, women with GUD were significantly more likely to have incident OIs (adjusted hazard ratio (aHR) 2.79, 95% CI: 1.33–5.85), and there was a trend for association between HSV-2-seropositivity and incident OIs (aHR 3.83, 95% CI: 0.93–15.83). Rate of change in CD4 count and HIV-1 RNA did not differ by HSV-2 status or GUD, despite a trend toward higher baseline HIV-1 RNA in HSV-2-seropositive women (4.73 log10 copies/ml vs. 4.47 log10 copies/ml, P = 0.07).

Conclusions

HSV-2 was highly prevalent and pregnant HIV-1 infected women with GUD were significantly more likely to have incident OIs than women without GUD, suggesting that clinically evident HSV-2 is a more important predictor of HIV-1 disease progression than asymptomatic HSV-2.     

Life table assay of field-caught Mediterranean fruit flies, Ceratitis capitata, reveals age bias

Though traps are used widely to sample phytophagous insects for research or management purposes, and recently in aging research, possible bias stemming from differential response of individuals of various ages to traps has never been examined. In this paper, we tested the response of Ceratitis capitata (Wiedemann) (Diptera: Tephritidae) males and females of four ages (spanning from 1 to 40 days) to McPhail-type traps baited with a synthetic food attractant in field cages and found that the probability of trapping was significantly influenced by age. The type of food on which flies were maintained before testing (sugar or protein) also had a strong effect and interacted with age. In another experiment, we collected wild C. capitata adults of unknown age using 1–3 methods and then reared them in the laboratory until death. The survival schedules of these flies were subsequently used in a life table assay to infer their age at the time of capture. Results showed that on a single sampling date, males captured in traps baited with a food attractant were younger compared with males aspirated from fruiting host trees, or males captured in traps baited with a sex attractant. Likewise, females captured in food-baited traps were younger compared with aspirated females. In addition to providing the first evidence of age-dependent sampling bias for a phytophagous insect species, this paper also provides a novel approach to estimate the differences in the age composition of samples collected with different techniques. These findings are of utmost importance for several categories of insects, medically important groups notwithstanding.     

Reproductive tactics influence cortisol levels in individual male gray-cheeked mangabeys (Lophocebus albigena)

Concentration of the hormone cortisol is often used as an indicator of stress, and chronically high cortisol levels are often associated with poor health. Among group living animals that compete for resources, agonistic social interactions can be expected to contribute to variation in cortisol levels within and among individuals over time. Reproductive tactics of males can change with individual quality, relatedness, and social structure, and affect cortisol levels. In gray-cheeked mangabey (Lophocebus albigena) groups, male rank is an important factor in social interactions, and males also move between groups while actively competing for females or sneaking copulations. During a 20-month study we observed the social behavior and collected 461 fecal samples from 24 adult male gray-cheeked mangabeys from five groups in Kibale National Park, Uganda. Aggressive interactions and the presence of females at the peak of sexual swelling were associated with elevated cortisol concentrations in all males. Independently, dominant (i.e., highest-ranking) males within groups had higher cortisol concentrations than subordinate males, and immigrant males had higher cortisol concentrations than dominant males.     

T Cell Receptor Signaling Co-regulates Multiple Golgi Genes to Enhance N-Glycan Branching

T cell receptor (TCR) signaling enhances β1,6GlcNAc-branching in N-glycans, a phenotype that promotes growth arrest and inhibits autoimmunity by increasing surface retention of cytotoxic T lymphocyte antigen-4 (CTLA-4) via interactions with galectins. N-Acetylglucosaminyltransferase V (MGAT5) mediates β1,6GlcNAc-branching by transferring N-acetylglucosamine (GlcNAc) from UDP-GlcNAc to N-glycan substrates produced by the sequential action of Golgi α1,2-mannosidase I (MIa,b,c), MGAT1, α1,2-mannosidase II (MII, IIx), and MGAT2. Here we report that TCR signaling enhances mRNA levels of MIa,b,c and MII,IIx in parallel with MGAT5, whereas limiting levels of MGAT1 and MGAT2. Blocking the increase in MI or MII enzyme activity induced by TCR signaling with deoxymannojirimycin or swainsonine, respectively, limits β1,6GlcNAc-branching, suggesting that enhanced MI and MII activity are both required for this phenotype. MGAT1 and MGAT2 have an ∼250- and ∼20-fold higher affinity for UDP-GlcNAc than MGAT5, respectively, and increasing MGAT1 expression paradoxically inhibits β1,6GlcNAc branching by limiting UDP-GlcNAc supply to MGAT5, suggesting that restricted changes in MGAT1 and MGAT2 mRNA levels in TCR-stimulated cells serves to enhance availability of UDP-GlcNAc to MGAT5. Together, these data suggest that TCR signaling differentially regulates multiple N-glycan-processing enzymes at the mRNA level to cooperatively promote β1,6GlcNAc branching, and by extension, CTLA-4 surface expression, T cell growth arrest, and self-tolerance.     

Genome-wide association analysis in asthma subjects identifies SPATS2L as a novel bronchodilator response gene

Bronchodilator response (BDR) is an important asthma phenotype that measures reversibility of airway obstruction by comparing lung function (i.e. FEV(1)) before and after the administration of a short-acting β(2)-agonist, the most common rescue medications used for the treatment of asthma. BDR also serves as a test of β(2)-agonist efficacy. BDR is a complex trait that is partly under genetic control. A genome-wide association study (GWAS) of BDR, quantified as percent change in baseline FEV(1) after administration of a β(2)-agonist, was performed with 1,644 non-Hispanic white asthmatic subjects from six drug clinical trials: CAMP, LOCCS, LODO, a medication trial conducted by Sepracor, CARE, and ACRN. Data for 469,884 single-nucleotide polymorphisms (SNPs) were used to measure the association of SNPs with BDR using a linear regression model, while adjusting for age, sex, and height. Replication of primary P-values was attempted in 501 white subjects from SARP and 550 white subjects from DAG. Experimental evidence supporting the top gene was obtained via siRNA knockdown and Western blotting analyses. The lowest overall combined P-value was 9.7E-07 for SNP rs295137, near the SPATS2L gene. Among subjects in the primary analysis, those with rs295137 TT genotype had a median BDR of 16.0 (IQR = [6.2, 32.4]), while those with CC or TC genotypes had a median BDR of 10.9 (IQR = [5.0, 22.2]). SPATS2L mRNA knockdown resulted in increased β(2)-adrenergic receptor levels. Our results suggest that SPATS2L may be an important regulator of β(2)-adrenergic receptor down-regulation and that there is promise in gaining a better understanding of the biological mechanisms of differential response to β(2)-agonists through GWAS.     

HIV-neutralizing activity of cationic polypeptides in cervicovaginal secretions of women in HIV-serodiscordant relationships

Background

HIV exposed seronegative (HESN) women represent the population most in need of a prophylactic antiviral strategy. Mucosal cationic polypeptides can potentially be regulated for this purpose and we here aimed to determine their endogenous expression and HIV neutralizing activity in genital secretions of women at risk of HIV infection.

Methodology/Principal Findings

Cervicovaginal secretions (CVS) of Kenyan women in HIV-serodiscordant relationships (HESN, n = 164; HIV seropositive, n = 60) and low-risk controls (n = 72) were assessed for the cationic polypeptides HNP1–3, LL-37 and SLPI by ELISA and for HIV neutralizing activity by a PBMC-based assay using an HIV primary isolate. Median levels of HNP1–3 and LL-37 in CVS were similar across study groups. Neither HSV-2 serostatus, nor presence of bacterial vaginosis, correlated with levels of HNP1–3 or LL-37 in the HESN women. However, an association with their partner''s viral load was observed. High viral load (>10,000 HIV RNA copies/ml plasma) correlated with higher levels of HNP1–3 and LL-37 (p = 0.04 and 0.03, respectively). SLPI was most abundant in the low-risk group and did not correlate with male partner''s viral load in the HESN women. HIV neutralizing activity was found in CVS of all study groups. In experimental studies, selective depletion of cationic polypeptides from CVS rendered the remaining CVS fraction non-neutralizing, whereas the cationic polypeptide fraction retained the activity. Furthermore, recombinant HNP1–3 and LL-37 could induce neutralizing activity when added to CVS lacking intrinsic activity.

Conclusions/Significance

These findings show that CVS from HESN, low-risk, and HIV seropositive women contain HIV neutralizing activity. Although several innate immune proteins, including HNP1–3 and LL-37, contribute to this activity these molecules can also have inflammatory properties. This balance is influenced by hormonal and environmental factors and in the present HIV serodiscordant couple cohort study we show that a partner''s viral load is associated with levels of such molecules.     

A high-density EEG investigation into steady state binaural beat stimulation

Binaural beats are an auditory phenomenon that has been suggested to alter physiological and cognitive processes including vigilance and brainwave entrainment. Some personality traits measured by the NEO Five Factor Model have been found to alter entrainment using pulsing light stimuli, but as yet no studies have examined if this occurs using steady state presentation of binaural beats for a relatively short presentation of two minutes. This study aimed to examine if binaural beat stimulation altered vigilance or cortical frequencies and if personality traits were involved. Thirty-one participants were played binaural beat stimuli designed to elicit a response at either the Theta (7 Hz) or Beta (16 Hz) frequency bands while undertaking a zero-back vigilance task. EEG was recorded from a high-density electrode cap. No significant differences were found in vigilance or cortical frequency power during binaural beat stimulation compared to a white noise control period. Furthermore, no significant relationships were detected between the above and the Big Five personality traits. This suggests a short presentation of steady state binaural beats are not sufficient to alter vigilance or entrain cortical frequencies at the two bands examined and that certain personality traits were not more susceptible than others.