Impact Loading and Locomotor-Respiratory Coordination Significantly Influence Breathing Dynamics in Running Humans

Locomotor-respiratory coupling (LRC), phase-locking between breathing and stepping rhythms, occurs in many vertebrates. When quadrupedal mammals gallop, 1∶1 stride per breath coupling is necessitated by pronounced mechanical interactions between locomotion and ventilation. Humans show more flexibility in breathing patterns during locomotion, using LRC ratios of 2∶1, 2.5∶1, 3∶1, or 4∶1 and sometimes no coupling. Previous studies provide conflicting evidence on the mechanical significance of LRC in running humans. Some studies suggest LRC improves breathing efficiency, but others suggest LRC is mechanically insignificant because ‘step-driven flows’ (ventilatory flows attributable to step-induced forces) contribute a negligible fraction of tidal volume. Yet, although step-driven flows are brief, they cause large fluctuations in ventilatory flow. Here we test the hypothesis that running humans use LRC to minimize antagonistic effects of step-driven flows on breathing. We measured locomotor-ventilatory dynamics in 14 subjects running at a self-selected speed (2.6±0.1 ms⁻¹) and compared breathing dynamics in their naturally ‘preferred’ and ‘avoided’ entrainment patterns. Step-driven flows occurred at 1-2X step frequency with peak magnitudes of 0.97±0.45 Ls⁻¹ (mean ±S.D). Step-driven flows varied depending on ventilatory state (high versus low lung volume), suggesting state-dependent changes in compliance and damping of thoraco-abdominal tissues. Subjects naturally preferred LRC patterns that minimized antagonistic interactions and aligned ventilatory transitions with assistive phases of the step. Ventilatory transitions initiated in ‘preferred’ phases within the step cycle occurred 2x faster than those in ‘avoided’ phases. We hypothesize that humans coordinate breathing and locomotion to minimize antagonistic loading of respiratory muscles, reduce work of breathing and minimize rate of fatigue. Future work could address the potential consequences of locomotor-ventilatory interactions for elite endurance athletes and individuals who are overweight or obese, populations in which respiratory muscle fatigue can be limiting.     

A novel method to isolate cells with conditional gene expression using fluorescence activated cell sorting.

An inducible expression system using control elements of the tetracycline resistance operon has recently shown promise for conditional gene expression of any gene of interest. However, intensive screening of multiple independent clones is often required to find cell lines with optimal induction characteristics. By coupling expression of the gene of interest with a fluorescent marker, we have developed a novel fluorescence activated cell sorting (FACS) based strategy to isolate cells with desirable expression characteristics, thus alleviating the laborious isolation and analysis of multiple independent clones.     

CHLOROPHYLL,NITROGEN, AND PHOTOSYNTHETIC PATTERNS DURING GROWTH AND SENESCENCE OF TWO BLUE-GREEN ALGAE1,2

A standardized, multiflask, batch culture system was developed to study the processes of algal senescence in Anacystis nidulans and Phormidium molle Gom, var. tenuior W. et G. West. Growth data over a 3-year period gave reproducible and comparable time-course curves. Although A. nidulans is unicellular and P. molle filamentous, the patterns of change with age were similar. Mean logarithmic doubling times and carbon yields were, respectively, 6.9 hr and 390 mg C/liter for A. nidulans and 7.2 hr and 710 mg C/liter for P. molle. Chlorophyll concentration and photo-synthetic capacity per unit carbon rose rapidly during the logarithmic phase to maximum levels in either late log phase (P. molle) or early linear phase (A. nidulans), then fell throughout the declining growth phase to low levels in the stationary phase. Nitrate was rapidly exhausted from the medium during the period of logarithmic growth and stoichiometrically converted to particulate organic form; very little subsequent fixation of molecular nitrogen occurred. The phycocyanins were rapidly destroyed during the logarithmic phase while the carotenoids remained relatively constant throughout the whole growth period and then slowly declined. Preliminary electron micrographs showed a progressive deterioration in cellular ultrastructure, especially a reduction in the number of photosynthetic thylakoids, commenting in the linear growth phase. Analysis of the results suggests that occurrence of linear growth kinetics and termination of culture growth were caused by exhaustion of nitrate. The observed decreases in chlorophylls, phycocyanins, and photosynthetic capacity during active culture growth show that senescence effects may not be, as assumed, restricted to the stationary phase of growth.     

Application of induced pluripotent stem cells to hematologic disease

Induced pluripotent stem cells (iPSC) were first generated from somatic cells via the transduction of four ‘Yamanaka’ factors, Oct4, Sox2, Klf4 and c-Myc. Because iPSC are similar to embryonic stem cells (ESC) and can be differentiated into any cell type of choice, iPSC have the potential to become a platform for personalized medicine by allowing a patient's own cells to become a source of therapeutic tissue. This review describes the main challenges in iPSC technology by focusing on its application to hematologic diseases. The explosive interest in improving iPSC technology has generated numerous genetic and chemical methods for iPSC derivation, but these methods must be evaluated comparatively for their safety and efficacy because there are risks of genetic abnormalities and oncogenesis. Competent iPSC will need to be selected carefully based on physical, genetic and functional criteria, and differentiated efficiently into hematopoietic stem cells via modulation of several signaling pathways before they prove valuable in the clinic.     

A Pilot Study of Short-Duration Sputum Pretreatment Procedures for Optimizing Smear Microscopy for Tuberculosis

Background

Direct sputum smear microscopy for tuberculosis (TB) lacks sensitivity for the detection of acid fast bacilli. Sputum pretreatment procedures may enhance sensitivity. We did a pilot study to compare the diagnostic accuracy and incremental yield of two short-duration (<1 hour) sputum pretreatment procedures to optimize direct smears among patients with suspected TB at a referral hospital in India.

Methodology/Findings

Blinded laboratory comparison of bleach and universal sediment processing (USP) pretreated centrifuged auramine smears to direct Ziehl-Neelsen (ZN) and direct auramine smears and to solid (Loweinstein-Jensen (LJ)) and liquid (BACTEC 460) culture. 178 pulmonary and extrapulmonary TB suspects were prospectively recruited during a one year period. Thirty six (20.2%) were positive by either solid or liquid culture. Direct ZN smear detected 22 of 36 cases and direct auramine smears detected 26 of 36 cases. Bleach and USP centrifugation detected 24 cases each, providing no incremental yield beyond direct smears. When compared to combined culture, pretreated smears were not more sensitive than direct smears (66.6% vs 61.1 (ZN) or 72.2 (auramine)), and were not more specific (92.3% vs 93.0 (ZN) or 97.2 (auramine).

Conclusions/Significance

Short duration sputum pretreatment with bleach and USP centrifugation did not increase yield as compared to direct sputum smears. Further work is needed to confirm this in a larger study and also determine if longer duration pre-treatment might be effective in optimizing smear microscopy for TB.     

Metal bridges between the PhoQ sensor domain and the membrane regulate transmembrane signaling

Bacterial histidine kinases respond to environmental stimuli by transducing a signal from an extracytosolic sensor domain to a cytosolic catalytic domain. Among them, PhoQ promotes bacterial virulence and is tightly repressed by the divalent cations such as calcium and magnesium. We have determined the crystal structure of the PhoQ sensor domain from Salmonella typhimurium in the Ca2+-bound state, which reveals a highly negatively charged surface that is in close proximity to the inner membrane. This acidic surface binds at least three Ca2+, which mediate the PhoQ-membrane interaction. Mutagenesis analysis indicates that structural integrity at the membrane proximal region of the PhoQ sensor domain promotes metal-mediated repression. We propose that depletion or displacement of divalent cations leads to charge repulsion between PhoQ and the membrane, which initiates transmembrane signaling through a change in orientation between the PhoQ sensor domain and membrane. Therefore, both PhoQ and the membrane are required for extracytosolic sensing and transmembrane signaling.     

Risk Factors for MDR and XDR-TB in a Tertiary Referral Hospital in India

Background

India has a high burden of drug resistant TB, although there are few data on XDR-TB. Although XDR-TB has existed previously in India, the definition has not been widely applied, and surveillance using second line drug susceptibility testing has not been performed. Our objective was to analyze clinical and demographic risk factors associated with isolation of MDR and XDR TB as compared to susceptible controls, at a tertiary center.

Methodology/Findings

Retrospective chart review based on positive cultures isolated in a high volume mycobacteriology laboratory between 2002 and 2007. 47 XDR, 30 MDR and 117 susceptible controls were examined. Drug resistant cases were less likely to be extrapulmonary, and had received more previous treatment regimens. Significant risk factors for XDR-TB included residence outside the local state (OR 7.43, 3.07-18.0) and care costs subsidized (OR 0.23, 0.097-0.54) in bivariate analysis and previous use of a fluoroquinolone and injectable agent (other than streptomycin) (OR 7.00, 95% C.I. 1.14-43.03) and an initial treatment regimen which did not follow national guidelines (OR 5.68, 1.24-25.96) in multivariate analysis. Cavitation and HIV did not influence drug resistance.

Conclusions/Significance

There is significant selection bias in the sample available. Selection pressure from previous treatment and an inadequate initial regimen increases risk of drug resistance. Local patients and those requiring financial subsidies may be at lower risk of XDR-TB.