Dendritic cells are less susceptible to human immunodeficiency virus type 2 (HIV-2) infection than to HIV-1 infection

Human immunodeficiency virus type 1 (HIV-1) infection of dendritic cells (DCs) has been documented in vivo and may be an important contributor to HIV-1 transmission and pathogenesis. HIV-1-specific CD4⁺ T cells respond to HIV antigens presented by HIV-1-infected DCs and in this process become infected, thereby providing a mechanism through which HIV-1-specific CD4⁺ T cells could become preferentially infected in vivo. HIV-2 disease is attenuated with respect to HIV-1 disease, and host immune responses are thought to be contributory. Here we investigated the susceptibility of primary myeloid DCs (mDCs) and plasmacytoid DCs (pDCs) to infection by HIV-2. We found that neither CCR5-tropic primary HIV-2 isolates nor a lab-adapted CXCR4-tropic HIV-2 strain could efficiently infect mDCs or pDCs, though these viruses could infect primary CD4⁺ T cells in vitro. HIV-2-exposed mDCs were also incapable of transferring virus to autologous CD4⁺ T cells. Despite this, we found that HIV-2-specific CD4⁺ T cells contained more viral DNA than memory CD4⁺ T cells of other specificities in vivo. These data suggest that either infection of DCs is not an important contributor to infection of HIV-2-specific CD4⁺ T cells in vivo or that infection of DCs by HIV-2 occurs at a level that is undetectable in vitro. The frequent carriage of HIV-2 DNA within HIV-2-specific CD4⁺ T cells, however, does not appear to be incompatible with preserved numbers and functionality of HIV-2-specific CD4⁺ T cells in vivo, suggesting that additional mechanisms contribute to maintenance of HIV-2-specific CD4⁺ T-cell help in vivo.     

Control of masculinization of the brain and behavior

Sex steroid hormones exert a profound influence on the sexual differentiation and function of the neural circuits that mediate dimorphic behaviors. Both estrogen and testosterone are essential for male typical behaviors in many species. Recent studies with genetically modified mice provide important new insights into the logic whereby these two hormones coordinate the display of sexually dimorphic behaviors: estrogen sets up the masculine repertoire of sexual and territorial behaviors and testosterone controls the extent of these male displays.     

Resistance and aerobic exercise: the influence of mode on the relationship between IL-6 and glucose tolerance in young men who are obese

Regular exercise lowers indicators of disease risk including some inflammatory cytokines; however, the relationship between different modes of acute exercise, cytokine levels, and subsequent glucose tolerance is unclear. The purpose was to determine the effects of resistance (RES) and aerobic (AER) exercises on interleukin-6 (IL-6) and its association with glucose tolerance 24 hours after exercise. After testing for 1 repetition maximum (1RM) and VO2peak, 10 obese (body mass index > 30 kg · m(-2)), untrained men aged 18-26 years completed 3 protocols: 60 minutes of RES, AER, and a resting (CON) condition. The RES was 2 sets of 8 repetitions and a third set to fatigue at 80% 1RM of 8 lifts using all major muscle groups. The AER was 60 minutes of cycling at 70% of VO2peak. On day 1, subjects completed the 60-minute exercise or resting protocol, and on day 2, they completed an oral glucose tolerance test (OGTT). Blood was collected before and after exercise, at 2 and 7 hour postexercise, and before and every 30 minutes during the OGTT and was analyzed for IL-6, glucose and insulin. Postexercise IL-6 was greater in RES (8.01 ± 2.08 pg · mL(-1)) vs. in AER (4.26 ± 0.27 pg · mL(-1)), and both were greater than in CON (1.61 ± 0.18 pg · mL(-1)). During the OGTT, there were no differences in glucose or insulin between conditions for single time points or as area under the curve. The RES caused greater IL-6 levels immediately after exercise that may be related to the greater active muscle mass compared to AER. Neither exercise produced enhanced glucose removal compared to control; thus, despite the greater elevation in IL-6 in RES, for these exercise conditions and this population, this cytokine did not influence glucose tolerance.     

Long-term continuous monitoring of dissolved oxygen in cell culture medium for perfused bioreactors using optical oxygen sensors

For long-term growth of mammalian cells in perfused bioreactors, it is essential to monitor the concentration of dissolved oxygen (DO) present in the culture medium to ascertain the health of the cells. An optical oxygen sensor based on dynamic fluorescent quenching was developed for long-term continuous measurement of DO for NASA-designed rotating perfused bioreactors. Tris(4,7-diphenyl-1,10-phenanthroline) ruthenium(II) chloride is employed as the fluorescent dye indicator. A pulsed, blue LED was chosen as the excitation light source. The sensor can be sterilized using an autoclave. The sensors were tested in a perfused rotating bioreactor supporting a BHK-21 (baby hamster kidney) cell culture over one 28-day, one 43-day, and one 180-day cell runs. The sensors were initially calibrated in sterile phosphate-buffered saline (PBS) against a blood-gas analyzer (BGA), and then used continuously during the entire cell culture without recalibration. In the 180-day cell run, two oxygen sensors were employed; one interfaced at the outlet of the bioreactor and the other at the inlet of the bioreactor. The DO concentrations determined by both sensors were compared with those sampled and measured regularly with the BGA reference. The sensor outputs were found to correlate well with the BGA data throughout the experiment using a single calibration, where the DO of the culture medium varied between 25 and 60 mm Hg at the bioreactor outlet and 80-116 mm Hg at the bioreactor inlet. During all 180 days of culture, the precision and the bias were +/-5.1 mm Hg and -3.8 mm Hg at the bioreactor outlet, and +/- 19 mm Hg and -18 mm Hg at inlet. The sensor dynamic range is between 0 and 200 mm Hg and the response time is less than 1 minute. The resolution of the sensor is 0.1 mm Hg at 50 mm Hg, and 0.25 mm Hg at 130 mm Hg.     

Human papillomavirus DNA and liquid-based cervical cytology cotesting in screening and follow-up patient groups

OBJECTIVE: To compare the use of human papillomavirus (HPV) DNA and cervical cytology cotesting in screening and follow-up of patients with previous cervical abnormalities and to assess the significance of a positive HPV DNA test result in re-screening of cytologically normal cases. STUDY DESIGN: Cellular samples collected in liquid-based fixative were used for both cervical cytology and HPV DNA testing. The cervical cytology slides were manually screened by cytotechnologists followed by rapid re-screening by pathologists. The HPV DNA tests were performed using hybrid capture test kits. Statistical analyses of cervical cytology results and HPV DNA tests for high- and low-risk HPV from both patient groups were carried out. RESULTS: The prevalence of HPV DNA-positive cases was higher in younger patients. There was a poor correlation between cervical cytology results and HPV DNA tests for the screening group (kappa = 0.23), but a fair to good correlation was obtained for the follow-up group (kappa = 0.51). The false negative fraction of cytology negative/HPV DNA positive cases (0.1317), as compared with cytology negative/HPV DNA negative cases (0.0056), was statistically significant (p = 0.000001). CONCLUSION: The prevalence of HPV DNA decreased with increasing age in both the screening and follow-up patient groups. Virus clearance was delayed in the follow-up group as compared with the screening group. There was a poor correlation between cervical cytology and HPV DNA tests in the screening group but a fair to good correlation in the follow-up patient group. Cotesting of HPV DNA and cervical cytology increases the sensitivity and decreases the false negative fraction, suggesting that cotesting could be used to increase the interval of screening.     

Chronic hypoxic decreases in soluble guanylate cyclase protein and enzyme activity are age dependent in fetal and adult ovine carotid arteries.

The present study tests the hypothesis that chronic hypoxia enhances reactivity to nitric oxide (NO) through age-dependent increases in soluble guanylate cyclase (sGC) and protein kinase G (PKG) activity. In term fetal and adult ovine carotids, chronic hypoxia had no significant effect on mRNA levels for the beta1-subunit of sGC, but depressed sGC abundance by 16% in fetal and 50% in adult arteries, through possible depression of rates of mRNA translation (15% in fetal and 50% in adult) and/or increased protein turnover. Chronic hypoxia also depressed the catalytic activity of sGC, but only in fetal arteries (63%). Total sGC activity was reduced by chronic hypoxia in both fetal (69%) and adult (37%) carotid homogenates, but this effect was not observed in intact arteries when sGC activity was measured by timed accumulation of cGMP. In intact arteries treated with 300 microM 3-isobutyl-1-methylxanthine (IBMX), chronic hypoxia dramatically enhanced sGC activity in fetal (186%) but not adult (89%) arteries. This latter observation suggests that homogenization either removed an sGC activator, released an sGC inhibitor, or altered the phosphorylation state of the enzyme, resulting in reduced activity. In the absence of IBMX, chronic hypoxia had no significant effect on rates of cGMP accumulation. Chronic hypoxia also depressed the ability of the cGMP analog, 8-(p-chlorophenylthio)-cGMP, to promote vasorelaxation in both fetal (8%) and adult (12%) arteries. Together, these results emphasize the fact that intact and homogenized artery studies of sGC activity do not always yield equivalent results. The results further suggest that enhancement of reactivity to NO by chronic hypoxia must occur upstream of PKG and can only be possible if changes in cGMP occurred in functional compartments that afforded either temporal or chemical protection to the actions of phosphodiesterase. The range and age dependence of hypoxic effects observed also suggest that some responses to hypoxia must be compensatory and homeostatic, with reactivity to NO as the primary regulated variable.     

Physiological 3D tissue model of the airway wall and mucosa

This protocol describes the setup, maintenance and characteristics of a tissue-engineered model of the human bronchial mucosa that can be used for basic physiology and pathophysiology studies. The model includes a well-differentiated epithelium with functional cilia, mucus secretion and subepithelial fibroblasts within type I collagen. The tissue is created within porous polymeric wells to prevent gel contraction and allow culture at the air-liquid interface. It requires at least 2 wk to be established and can be maintained thereafter for over 4 wk, with tissue differentiation moving towards a more physiologically relevant phenotype with increasing time in culture. Over time, the extracellular matrix also remodels, depositing proteins such as types III and IV collagen and fibronectin. Because it recapitulates many key anatomical and functional features of the airway wall, this model is well suited for a wide range of studies, including those on airway remodeling, transepithelial transport and inflammatory cell interactions with the mucosa. The entire protocol takes 4-6 wk, including cell expansion, depending on the extent of ciliogenesis desired.