Development and validation of models to predict personal ventilation rate for air pollution research.

Air pollution intake represents the amount of pollution inhaled into the body and may be calculated by multiplying an individual's ventilation rate with the concentration of pollutant present in their breathing zone. Ventilation rate is difficult to measure directly, and methods for estimating ventilation rate (and intake) are lacking. Therefore, the goal of this work was to examine how well linear models using heart rate and other basic physiologic data can predict personal ventilation rate. We measured personal ventilation and heart rate among a panel of subjects (n = 36) while they conducted a series of specified routine tasks of varying exertion levels. From these data, 136 candidate models were identified using a series of variable transformation and selection algorithms. A second "free­living" validation study (n = 26) served as an independent validation dataset for these candidate models. The top­performing model, which included heart rate (Hr), resting heart rate (Hrest), age, sex, and hip circumference and interactions between sex with Hr, Hrest, age, and hip predicted ventilation rate (Ve) to within 11% and 33% for moderate (Ve = 45 L/min) and low (Ve = 15 L/min) intensity activities, respectively, based on the validation study. Many of the promising candidate models performed substantially worse under independent validation. Our results indicate that while measures of air pollution exposure and intake are highly correlated within tasks for a given individual, this correlation decreases substantially across tasks (i.e., as individuals go about a series of typical daily activities). This discordance between exposure and intake may influence exposure­response estimates in epidemiological studies. New air pollution studies should consider the trade­offs between the predictive ability of intake models and the error potentially introduced by not accounting for ventilation rate.

Exposure to household air pollution from biomass-burning cookstoves and HbA1c and diabetic status among Honduran women.

Household air pollution from biomass cookstoves is estimated to be responsible for more than two and a half million premature deaths annually, primarily in low and middle-income countries where cardiometabolic disorders, such as Type II Diabetes, are increasing. Growing evidence supports a link between ambient air pollution and diabetes, but evidence for household air pollution is limited. This cross-sectional study of 142 women (72 with traditional stoves and 70 with cleaner-burning Justa stoves) in rural Honduras evaluated the association of exposure to household air pollution (stove type, 24-hour average kitchen and personal fine particulate matter [PM2.5 ] mass and black carbon) with glycated hemoglobin (HbA1c) levels and diabetic status based on HbA1c levels. The prevalence ratio (PR) per interquartile range increase in pollution concentration indicated higher prevalence of prediabetes/diabetes (vs normal HbA1c) for all pollutant measures (eg, PR per 84 µg/m3 increase in personal PM2.5 , 1.49; 95% confidence interval [CI], 1.11-2.01). Results for HbA1c as a continuous variable were generally in the hypothesized direction. These results provide some evidence linking household air pollution with the prevalence of prediabetes/diabetes, and, if confirmed, suggest that the global public health impact of household air pollution may be broader than currently estimated.

The identification of differentially expressed microRNA in osteoarthritic tissue that modulate the production of TNF-alpha and MMP13.

OBJECTIVE: To identify differentially expressed microRNAs (miRNAs) in human osteoarthritic (OA) cartilage and bone tissue and to determine their relevance to chondrocyte function. METHODS: Cartilage and bone was obtained from OA patients who underwent total knee joint replacement surgery or from post-mortem patients with no previous history of OA. MiRNA expression was quantified by real-time PCR (RT-PCR). Functional pathway analysis of miRNA was performed using Ingenuity Pathway Analysis. Primary chondrocytes were isolated by collagenase digestion and transfected with miRNA mimics and miRNA inhibitors using cationic lipid. Tumour Necrosis Factor-alpha (TNF-alpha) and Matrix metalloprotease 13 (MMP13) protein levels were measured by Enzyme-Linked ImmunoSorbent Assay (ELISA). RESULTS: In total we identified 17 miRNA that showed greater than 4-fold differential expression between OA and normal cartilage, and 30 miRNA that showed greater than 4-fold differential expression in OA bone. Functional pathway analysis of the predicted gene targets for miR-9, miR-98, which were upregulated in both OA bone and cartilage tissue, and miR-146, which was downregulated in OA cartilage, suggested that these miRNA mediate inflammatory functions and pathways. Over-expression of miR-9, miR-98 or miR-146 in isolated human chondrocytes reduced interleukin-1 beta (IL-1 beta) induced TNF-alpha production. Furthermore, inhibition and over-expression of miR-9 modulated MMP13 secretion. CONCLUSIONS: We have identified a number of differentially expressed miRNAs in late-stage human OA cartilage and bone. Functional analysis of miR-9, miR-98 and miR-146 in primary chondrocytes suggests a role in mediating the IL-1 beta induced production of TNF-alpha. MiR-9, upregulated in OA tissue, was found to inhibit secretion of the collagen type II-targeting metalloproteinase MMP13 in isolated human chondrocytes.

Mitogen-activated protein kinase-activated protein kinase 2 (MK2) modulates key biological pathways associated with OA disease pathology.

OBJECTIVE: To examine the role of mitogen-activated protein kinase-activated protein kinase 2 (MK2) in mediating the cellular response to pro-inflammatory cytokines in human primary osteoarthritis (OA) chondrocytes. METHODS: Delivery of a dominant negative MK2 was achieved in HeLa cells by adenoviral infection. Cellular heat shock protein (HSP27) activity was determined using a Bioplex assay. Primary OA chondrocytes were isolated by collagenase digestion of human articular cartilage. Phosphorylated MK2 was detected by immunoblotting and immunohistology. Transfection of primary chondrocytes with siRNA was achieved using cationic lipid and gene expression determined by real-time polymerase chain reaction. Production of prostaglandin E2 (PGE2) and matrixmetalloproteases (MMPs) was measured by enzyme-linked immunosorbent assay. RESULTS: Over-expression of a dominant negative MK2 inhibited HSP27 phosphorylation and significantly reduced both interleukin 1 (IL-1)beta and tumour necrosis factor (TNF)-alpha mediated release of PGE2 in HeLa cells over a 24h period. Phosphorylated MK2 was detected in OA articular cartilage and in isolated primary OA chondrocytes, where it was induced by IL-1beta. Transfection of OA chondrocytes with MK2 siRNA antisense significantly reduced both basal and IL-1beta induced PGE2 release. siRNA mediated MK2 knockdown also significantly reduced both basal and IL-1beta induced MMP13 expression and MMP13 and MMP3 protein release but had no effect on MMP1. CONCLUSIONS: Our data reveal that MK2 is active in OA human articular cartilage and in isolated primary human chondrocytes and that MK2 mediates the release of PGE2, MMP3 and MMP13. These findings suggest a role for MK2 in contributing to OA algesia and OA joint structural deterioration by mediating the downstream effects of p38 activation on PGE2 release and the expression and release of catabolic proteases.

The orphan G-protein coupled receptor RDC1: evidence for a role in chondrocyte hypertrophy and articular cartilage matrix turnover.

OBJECTIVE: RDC1 is a class A orphan G-protein coupled receptor of unknown function. The purpose of this study was to identify compound RDC1 agonists and use these as tools to determine the effect of RDC1 activation in human chondrocytes and cartilage explant tissue. METHODS: Computational chemistry was employed to build a homology model of the RDC1 receptor. A virtual screen of in-house compounds was then performed and positive hits screened for their ability to invoke a Ca2+ response in a recombinant RDC1 HEK293 cell line, as measured by FLIPR. The effect of RDC1 activation on human chondrocytes and cartilage explant gene expression was determined by quantitative real-time polymerase chain reaction (PCR), and these effects validated as being mediated by RDC1 using siRNA antisense. RESULTS: Tissue expression profiling demonstrated that RDC1 expression was predominant in cartilage tissue. Treatment of human primary chondrocytes with RDC1 agonist induced a Ca2+ response, suggesting the receptor is active in this tissue type. Treatment for 24h with RDC1 agonist led to altered expression of a number of genes associated with chondrocyte hypertrophy and increased matrix degradation in human primary chondrocytes, and elevated total matrix metalloproteinase (MMP) activity in cartilage explant. Transfection with RDC1 siRNA caused a >90% reduction in human primary chondrocyte RDC1 expression and significantly reduced the impact of RDC1 agonist on the previously identified RDC1-regulated genes. CONCLUSIONS: RDC1 activation in human chondrocytes and cartilage explant leads to changes in gene expression and activity associated with chondrocyte hypertrophy, angiogenesis and increased matrix degradation, suggesting signalling via the RDC1 receptor may play an important role in the early development of osteoarthritis (OA).

Correlation between frequency of tuberculosis and compliance with control strategies.

OBJECTIVE: To determine if compliance with annual tuberculosis skin testing correlated with the number of cases of tuberculosis seen in patients and healthcare workers. DESIGN: Survey using a written questionnaire. SETTING AND PARTICIPANTS: 159 Veterans' Administration facilities. RESULTS: Hospitals that reported that > 80% of their healthcare workers received annual skin tests saw 12.7 patient cases per 10,000 admissions and 4.0 healthcare worker cases per 10,000 personnel. Facilities in which < 20% of their healthcare workers were given annual skin tests saw 4.5 cases per 10,000 admissions and 1.6 cases in healthcare workers per 10,000 personnel (P < .001 for patients and P = .31 for healthcare workers). The ratio of the median number of patients placed in acid-fast bacilli (AFB) isolation to the median number of patients with confirmed tuberculosis was 12. There was no correlation of this ratio with the number of cases of tuberculosis in patients or healthcare workers seen in each facility. CONCLUSION: Compliance with annual tuberculosis skin testing was related directly to the rate of tuberculosis seen in patients. More standardized policies for placing patients in AFB isolation are needed to control for potentially costly variation among facilities. These measures should have highest priority in the control of tuberculosis in the healthcare setting, before implementing still more expensive interventions.

Tuberculosis control strategies: the cost of particulate respirators.

OBJECTIVE: To assess the cost of the mandatory use of high-efficiency particulate respirators to treat patients with known or suspected tuberculosis. DESIGN: A questionnaire was used to determine the number of high-efficiency particulate respirators required and the number of cases of tuberculosis in employees that could potentially be prevented. Indirect costs included the training and fitness testing of employees. The clinical efficacy of respirators is not known. To provide a best-case scenario, it was assumed that the respirators could prevent as many as 25% of tuberculosis cases in health care workers. SETTING: 159 acute care facilities administered by the Department of Veterans Affairs. PARTICIPANTS: Quality improvement, infection control, and employee health specialists. MEASUREMENTS: Cost of the respirators compared with their maximum predicted efficacy. RESULTS: The use of the respirators would cost $7 million per case of tuberculosis prevented and $100 million per life saved. CONCLUSIONS: High-efficiency particulate respirators are a costly means of trying to prevent tuberculosis. Costs could be reduced by reusing masks or by restricting the number of health care workers allowed to have contact with potentially infectious patients. As the health care budget undergoes further restrictions, specific means of accommodating the cost of new regulations must be found.

Active transport, ion movements, and pH changes : I. The chemistry of pH changes.

The transport of substances across cell membranes may be the most fundamental activity of living things. When the substance transported is any ion there can be a change in the concentration of hydrogen ions on the two sides of the membrane. These hydrogen ion concentration changes are not caused by fluxes of hydrogen ions although fluxes of hydrogen ions may sometimes be involved. The reason for the apparent contradiction is quite simple. All aqueous systems are subject to two constraints: (1) to maintain the charge balance, the sum of the cationic charges must equal the sum of the anionic charges and (2) the product of the molar concentration of H(+) and the molar concentration of OH(-), established and maintained by the association and the dissociation of water, remains always at 10(-14). As a consequence the concentrations of H(+) and OH(-) are determined uniquely by differences between the concentrations of the other cations and anions, with [H(+)] and [OH(-)] being dependent variables. Hydrogen ions and hydroxyl ions can be produced or consumed in local reactions whereas any strong ions such as Cl(-), Mg(2+), or K(+) can be neither produced nor consumed in biological reactions. Further consequences of these truisms are outlined here in terms of the chemistry of the kinds of reactions which can lead to pH changes.

Active transport, ion movements, and pH changes : II. Changes of pH and ATP synthesis.

Electron transport in chloroplasts takes place across the thylakoid membrane in such a way as to redistribute ions. This redistribution can cause transmembrane electric fields and transmembrane hydrogen ion activity differences, events that are often correlated with the ability of the membrane to phosphorylate ADP. Analysis of the chemistry responsible for the phosphorylation is difficult because there seems to be no single satisfactory description of energized state of the thylakoid vesicles responsible for the photophosphorylation, the state depending on the experimental protocol employed. Under some conditions, acidification of the lumen confers on the vesicles the ability to synthesize ATP. Thus, when electron transport or preincubation in acid causes exogenous protonated buffers to accumulate in the lumen, ATP can be made in amounts commensurate with the accumulation if the pH of the medium is raised. When permeant exogenous buffers are absent an ability to make ATP also develops during prior electron transport, presumably because of protonation of membrane components. The nature of the energized state responsible for post-illumination phosphorylation in the latter instance is unclear. The energized state probably cannot then be simply a general delocalized ionic disequilibrium because of the precisely exponential nature of its decay with time after the light is off. The nature of the energized state which drives prompt phosphorylation in single stage experiments is even more puzzling. It may not depend on the kind of ion fluxes that result in a reversible pH rise in the medium. Certainly phosphorylation can begin at high efficiency when any measurable acidification of the vesicle lumen is prevented, even under conditions where the presence of a membrane potential is unlikely.

Evidence That IAA Conjugates Are Slow-Release Sources of Free IAA in Plant Tissues.

Evidence that indoleacetic acid (IAA) conjugates are metabolized via enzyme-catalyzed hydrolysis to free IAA and that their biological activities are related to the rates at which they are hydrolyzed by the tissue is presented. These conclusions are based on the following observations. Slow but continuous decarboxylation of the IAA moiety of IAA-l-alanine and IAA-glycine occurs when these conjugates are applied to pea (Pisum sativum L. cv. Alaska) stem segments. Inasmuch as IAA conjugates are protected from peroxidase-catalyzed oxidative decarboxylation, the conjugates are probably hydrolyzed and the freed IAA then further metabolized. Free IAA and IAA-l-alanine are converted, by pea stem tissue, into the same metabolites. The metabolism is enzymic, since conjugates of IAA with the d-isomers of the amino acids are inactive. Ethylene production induced by IAA-l-alanine and by IAA-glycine is correlated with their hydrolysis, as indicated by their decarboxylation and with the appearance or nonappearance of IAA metabolites in the tissues.

Biological activities of indoleacetylamino acids and their use as auxins in tissue culture.

THE AUXIN ACTIVITIES OF A NUMBER OF INDOLEACETYLAMINO ACID CONJUGATES HAVE BEEN DETERMINED IN THREE TEST SYSTEMS: growth of tomato hypocotyl explants (Lycopersicon esculentum Mill. cv. Marglobe); growth of tobacco callus cultures (Nicotiana tabacum L. cv. Wisconsin 38); and ethylene production from pea stems (Pisum sativum L. cv. Alaska). The activities of the conjugates differ greatly depending on the amino acid moiety. Indoleacetyl-l-alanine supports rapid callus growth from the tomato hypocotyls while inhibiting growth of shoots and roots. Indoleacetylglycine behaves in a similar manner but is somewhat less effective in supporting callus growth and in inhibiting shoot formation. The other amino acid conjugates tested (valine, leucine, aspartic acid, threonine, methionine, phenylalanine, and proline) support shoot formation without supporting root formation or much callus growth. The tobacco callus system, which forms abundant shoots in the presence or absence of free indoleacetic acid, produces only rapid undifferentiated growth in the presence of indoleacetyl-l-alanine and indoleacetylglycine. The other conjugates inhibit shoot formation weakly if at all. Most of the conjugates induce sustained ethylene production from the pea stems but at rates well below the initial rates observed with free indoleacetic acid. Many, but not all of the effects of conjugates such as indoleacetyl-l-alanine can be mimicked by frequent renewals of the supply of free indoleacetic acid.

Photophosphorylation as a function of illumination time. II. Effects of permeant buffers.

(1) The amounts of orthophosphate, bicarbonate and tris (hydroxymethyl)-aminomethane found inside the thylakoid are almost exactly the amounts predicted by assuming that the buffers equilibrate across the membrane. Since imidazole and pyridine delay the development of post-illumination ATP formation while increasing the maximum amount of ATP formed, it follows that such relatively permeant buffers must also enter the inner aqueous space of the thylakoid. (2) Photophosphorylation begins abruptly at full steady-state efficiency and full steady-state rate as soon as the illumination time exceeds about 5 ms when permeant ions are absent or as soon as the time exceeds about 50 ms if valinomycin and KC1 are present. In either case, permeant buffers have little or no effect on the time of illumination required to initiate phosphorylation. A concentration of bicarbonate which would delay acidification of the bulk of the inner aqueous phase for at least 350 ms has no effect at all on the time of initiation of phosphorylation. In somewhat swollen chloroplasts, the combined buffering by the tris(hydroxymethyl) aminomethane and orthophosphate inside would delay acidification of the inside by 1500 ms but, even in the presence of valinomycin and KC1, the total delay in the initiation of phosphorylation is then only 65 ms. Similar discrepancies occur with all of the other buffers mentioned. (3) Since these discrepancies between internal acidification and phosphorylation are found in the presence of saturating amounts of valinomycin and KC1, it seems that photophosphorylation can occur when there are no proton concentration gradients and no electrical potential differences across the membranes which separate the medium from the greater part of the internal aqueous phase. (4) We suggest that the protons produced by electron transport may be used directly for phosphorylation without even entering the bulk of the inner aqueous phase of the lamellar system. If so, phosphorylation could proceed long before the internal pH reflected the proton activity gradients within the membrane.