Facilitators and barriers to adolescent participation in a TB clinical trial.

The inclusion of adolescents in TB drug trials is essential for the development of safe, child-friendly regimens for the prevention and treatment of TB. TB Trials Consortium Study 31/AIDS Clinical Trials Group A5349 (S31/A5349) enrolled adolescents as young as 12 years old. We assessed investigator and coordinator described facilitators and barriers to adolescent recruitment, enrollment, and retention.Interviews were conducted with six investigators from sites that enrolled adolescent participants and six investigators from non-enrolling sites. Additionally, two focus groups were conducted with study coordinators from enrolling sites and two focus groups with non-enrolling sites. Discussions were transcribed, analyzed, summarized, and summaries were reviewed by Community Research Advisors Group members and research group representatives for content validity.Investigators and coordinators attributed the successful enrollment of adolescents to the establishment and cultivation of external partnerships, flexibility to accommodate adolescents' schedules, staff engagement, recruitment from multiple locations, dedicated recruitment staff working onsite to access potential participants, creation of youth-friendly environments, and effective communications. Non-enrolling sites were mainly hindered by regulations. Suggestions for improvement in future trials focused on study planning and site preparations.Proactive partnerships and collaboration with institutions serving adolescents helped identify and reduce barriers to their inclusion in this trial..

Accuracy of methods for reporting inorganic element concentrations and radioactivity in oil and gas wastewaters from the Appalachian Basin, U.S. based on an inter-laboratory comparison.

Accurate and precise analyses of oil and gas (O&G) wastewaters and solids (e.g., sediments and sludge) are important for the regulatory monitoring of O&G development and tracing potential O&G contamination in the environment. In this study, 15 laboratories participated in an inter-laboratory comparison on the chemical characterization of three O&G wastewaters from the Appalachian Basin and four solids impacted by O&G development, with the goal of evaluating the quality of data and the accuracy of measurements for various analytes of concern. Using a variety of different methods, analytes in the wastewaters with high concentrations (i.e., >5 mg L-1) were easily detectable with relatively high accuracy, often within ±10% of the most probable value (MPV). In contrast, often less than 7 of the 15 labs were able to report detectable trace metal(loid) concentrations (i.e., Cr, Ni, Cu, Zn, As, and Pb) with accuracies of approximately ±40%. Despite most labs using inductively coupled plasma mass spectrometry (ICP-MS) with low instrument detection capabilities for trace metal analyses, large dilution factors during sample preparation and low trace metal concentrations in the wastewaters limited the number of quantifiable determinations and likely influenced analytical accuracy. In contrast, all the labs measuring Ra in the wastewaters were able to report detectable concentrations using a variety of methods including gamma spectroscopy and wet chemical approaches following Environmental Protection Agency (EPA) standard methods. However, the reported radium activities were often greater than ±30% different to the MPV possibly due to calibration inconsistencies among labs, radon leakage, or failing to correct for self-attenuation. Reported radium activities in solid materials had less variability (±20% from MPV) but accuracy could likely be improved by using certified radium standards and accounting for self-attenuation that results from matrix interferences or a density difference between the calibration standard and the unknown sample. This inter-laboratory comparison illustrates that numerous methods can be used to measure major cation, minor cation, and anion concentrations in O&G wastewaters with relatively high accuracy while trace metal(loid) and radioactivity analyses in liquids may often be over ±20% different from the MPV.

Defining the optimal dose of rifapentine for pulmonary tuberculosis: Exposure-response relations from two phase II clinical trials.

Rifapentine is a highly active antituberculosis antibiotic with treatment-shortening potential; however, exposure-response relations and the dose needed for maximal bactericidal activity have not been established. We used pharmacokinetic/pharmacodynamic data from 657 adults with pulmonary tuberculosis participating in treatment trials to compare rifapentine (n = 405) with rifampin (n = 252) as part of intensive-phase therapy. Population pharmacokinetic/pharmacodynamic analyses were performed with nonlinear mixed-effects modeling. Time to stable culture conversion of sputum to negative was determined in cultures obtained over 4 months of therapy. Rifapentine exposures were lower in participants who were coinfected with human immunodeficiency virus, black, male, or fasting when taking drug. Rifapentine exposure, large lung cavity size, and geographic region were independently associated with time to culture conversion in liquid media. Maximal treatment efficacy is likely achieved with rifapentine at 1,200 mg daily. Patients with large lung cavities appear less responsive to treatment, even at high rifapentine doses.

Environmental signatures and effects of an oil and gas wastewater spill in the Williston Basin, North Dakota.

Wastewaters from oil and gas development pose largely unknown risks to environmental resources. In January 2015, 11.4ML (million liters) of wastewater (300g/L TDS) from oil production in the Williston Basin was reported to have leaked from a pipeline, spilling into Blacktail Creek, North Dakota. Geochemical and biological samples were collected in February and June 2015 to identify geochemical signatures of spilled wastewaters as well as biological responses along a 44-km river reach. February water samples had elevated chloride (1030mg/L) and bromide (7.8mg/L) downstream from the spill, compared to upstream levels (11mg/L and <0.4mg/L, respectively). Lithium (0.25mg/L), boron (1.75mg/L) and strontium (7.1mg/L) were present downstream at 5-10 times upstream concentrations. Light hydrocarbon measurements indicated a persistent thermogenic source of methane in the stream. Semi-volatile hydrocarbons indicative of oil were not detected in filtered samples but low levels, including tetramethylbenzenes and di-methylnaphthalenes, were detected in unfiltered water samples downstream from the spill. Labile sediment-bound barium and strontium concentrations (June 2015) were higher downstream from the Spill Site. Radium activities in sediment downstream from the Spill Site were up to 15 times the upstream activities and, combined with Sr isotope ratios, suggest contributions from the pipeline fluid and support the conclusion that elevated concentrations in Blacktail Creek water are from the leaking pipeline. Results from June 2015 demonstrate the persistence of wastewater effects in Blacktail Creek several months after remediation efforts started. Aquatic health effects were observed in June 2015; fish bioassays showed only 2.5% survival at 7.1km downstream from the spill compared to 89% at the upstream reference site. Additional potential biological impacts were indicated by estrogenic inhibition in downstream waters. Our findings demonstrate that environmental signatures from wastewater spills are persistent and create the potential for long-term environmental health effects.

Serum biomarkers of treatment response within a randomized clinical trial for pulmonary tuberculosis.

RATIONALE: Biomarkers for monitoring response to anti-tuberculosis treatment are needed. We explored immune markers previously published as having predictive capability for 8 week culture status in 39 adults enrolled in a clinical trial in Kampala, Uganda. METHODS: We consecutively selected 20 HIV-negative pulmonary TB subjects with positive cultures, and 19 subjects with negative cultures at the end of intensive phase therapy. At baseline and after 8 weeks, serum was assayed for nine cytokines and soluble cytokine receptors using multiplexed platforms or ELISA. We evaluated their association with week 8 culture status first using single-variable logistic models, then using cross-validated estimates of the C-statistic, a measure of discrimination, of candidate models including 2 or 3 analytes in addition to age. RESULTS: All but one analyte decreased from baseline to week 8 (all p < 0.01). Individual biomarkers were not associated with 8 week culture status. Logistic models including increasing age, higher baseline soluble tumor necrosis factor receptor alpha 1 (sTNF-R1), and higher week 8 C-reactive protein (CRP) concentration classified subjects by culture status with up to 85% accuracy and acceptable discrimination (cross-validated C-statistic 0.76) and calibration (Hosmer-Lemeshow P > 0.2). CONCLUSION: Exploratory post-hoc models including sTNF-R1, CRP, and age, classified 8 week culture status with promising accuracy.

Reasons for non-participation in an international multicenter trial of a new drug for tuberculosis treatment.

SETTING: Clinical trials can provide a high standard of patient care and contribute to scientific knowledge; however, only a fraction of the patients screened participate and receive treatment as part of a trial. OBJECTIVE: To explore reasons why patients were not enrolled in an international tuberculosis (TB) treatment trial and to compare experiences among study sites. DESIGN: An analysis of reasons why patients were not enrolled was conducted among patients screened for a TB clinical trial at 26 sites in North and South America, Africa, and Europe. RESULTS: Staff at study sites screened 1119 potential candidates for the trial: 61% (n = 686) were not enrolled due to 1) failure to meet eligibility criteria (n = 405, 59%), 2) site's decision (n = 168, 24%), or 3) candidate's choice (n = 113, 16%). Study staff recorded a total of 144 reasons for why they believed patients chose not to participate, including concerns over research (28%), conflicts with work or school (21%), and lifestyle and family issues (20%). Socio-demographic and geographic factors also influenced participation. CONCLUSION: Increased evaluation of screening outcomes and of specific interventions, such as improved education and communication about trial procedures, may increase the efficiency of screening and enrollment in clinical trials.

Spinal nerve ligation does not alter the expression or function of GABA(B) receptors in spinal cord and dorsal root ganglia of the rat.

Loss of GABA-mediated inhibition in the spinal cord is thought to mediate allodynia and spontaneous pain after nerve injury. Despite extensive investigation of GABA itself, relatively little is known about how nerve injury alters the receptors at which GABA acts. This study examined levels of GABA(B) receptor protein in the spinal cord dorsal horn, and in the L4 and L5 (lumbar designations) dorsal root ganglia one to 18 weeks after L5 spinal nerve ligation. Mechanical allodynia was maximal by 1 week and persisted at blunted levels for at least 18 weeks after injury. Spontaneous pain behaviors were evident for 6 weeks. Western blotting of dorsal horn detected two isoforms of the GABA(B(1)) subunit and a single GABA(B(2)) subunit. High levels of GABA(B(1a)) and low levels of GABA(B(1b)) protein were present in the dorsal root ganglia. However, GABA(B(2)) protein was not detected in the dorsal root ganglia, consistent with the proposed existence of an atypical receptor composed of GABA(B(1)) homodimers. The levels of GABA(B(1a)), GABA(B(1b)), and GABA(B(2)) protein in the ipsilateral dorsal horn were unchanged at any time after injury. Immunohistochemical staining also did not detect a change in GABA(B(1)) or GABA(B(2)) subunits in dorsal horn segments having a robust loss of isolectin B4 staining. The levels of GABA(B(1a)) protein were also unchanged in the L4 or L5 dorsal root ganglia at any time after spinal nerve ligation. Levels of GABA(B(2)) remained undetectable. Finally, baclofen-stimulated binding of guanosine-5'-(gamma-O-thio)triphosphate in dorsal horn did not differ between sham and ligated rats. Collectively, these results argue that a loss of GABA(B) receptor-mediated inhibition, particularly of central terminals of primary afferents, is unlikely to mediate the development or maintenance of allodynia or spontaneous pain behaviors after spinal nerve injury.

Surfactant-like particles mediate tissue-specific functions in epithelial cells.

Lipid-rich, unilamellar membranes appear to be relatively common structures lining the apical or 'exposed' surface of epithelial cells. They have now been described in the intestinal tract from the esophagus to the rectum and have been isolated from tissues, such as the stomach, the small bowel, the colon, and the bladder. The presence of a lining layer in the lungs has been known for some time, and its functions, structure, and metabolism have been extensively studied, as can be gleaned from the multitude of reports presented at this symposium. The 'other' surfactants, however, have attracted far less attention and have been investigated in detail in only a few reports. This paucity of information, when compared to the pulmonary system, is most likely due to the fact that a generalized function (sufficiency state) or disease (deficiency state) has not yet been recognized for either the intestinal or urinary forms of surfactant. It seems reasonable to assume that the role of the SLP will vary, at least in part, with the organ or tissue with which it is associated, although the widespread nature of the membrane assumes that some functions (e.g. protective) will be shared. Thus, pulmonary surfactant's primary function in the lung may be to reduce surface tension and prevent lung collapse; but it also plays a significant part in the lung's defenses against bacterial and/or chemical invasion. It is hoped that future studies will shed some light on the function of the various SLPs and lead to a better appreciation for their role in both maintaining a healthy environment and contributing to the proper functioning of their host tissues.

Regulation of surfactant-like particle secretion by Caco-2 cells.

Surfactant-like particle (SLP) is a phosphatidylcholine (PC)-rich membrane produced in the small intestine, and its secretion is increased by fat feeding. In Caco-2 cells known to produce SLP, preincubation with [(3)H]palmitate labelled the SLP and was used as a marker for newly secreted membrane. SLP-associated PC and protein (d=1.07-1.08 g/ml in a linear non-equilibrium NaBr gradient) were secreted in parallel with triacylglycerols (TG) and at a rate about twice the control rate in response to feeding cells with an oleate/egg PC mixture. Cholesterol and apolipoprotein A-I identified only a small peak corresponding to high-density lipoprotein (HDL), but the largest peak corresponded with SLP (d=1.07-1.08). Palmitate incorporation into PC showed a similar small peak migrating at the density of HDL, but most labelled PC secreted from the cells was due to SLP. PC secretion, alkaline phosphatase activity, and newly synthesized immunoprecipitated SLP proteins from conditioned serum-free media migrated together at a density of >/=1.21 g/ml in a lipoprotein NaBr step gradient, and represented SLP. Glycerol incorporated into TG migrated at a peak density of 1.12 g/ml, consistent with HDL secretion from cells incubated in serum-free media. These data confirm that the secreted PC in SLP is distinct from lipoprotein particles. Incorporation of [(3)H]palmitate into the PC fraction of either whole cell homogenate or isolated brush border membranes was not affected by oleate/egg PC feeding. Both Pluronic L-81, an inhibitor of chylomicron secretion, and BMS-197636-02, a microsomal triglyceride transfer protein inhibitor, blocked the secretion of both TG and PC. Elevation of intracellular cAMP levels that stimulate surfactant secretion from type II pneumocytes caused a 50% reduction in SLP and TG secretion from Caco-2 cells. These results confirm the SLP response to fat feeding found in vivo, further supporting a role for SLP in TG secretion from the enterocyte, and show that the regulation of SLP secretion differs from that of pulmonary surfactant.

Role of intestinal surfactant-like particles as a potential reservoir of uropathogenic Escherichia coli.

The binding of uropathogenic Escherichia coli is mediated at the tips of pili by the PapG adhesin, which recognizes the Galalpha(1-4)Gal disaccharide on the uroepithelial surface. These receptors have been identified unequivocally in the human and murine urinary tracts but not in intestinal epithelium, yet uropathogenic E. coli strains are commonly found in normal colonic microflora. The gastrointestinal tract from duodenum to rectum elaborates a phospholipid-rich membrane particle with surfactant-like properties. In these studies, we report that purified murine particles contain a receptor recognized by the class I PapG adhesin because: (1) PapD-PapG complexes and class I pili bound to surfactant-like particles in a solid-phase assay, whereas binding was not detected in microvillous membranes derived from the same tissues, (2) purified PapD-PapG complex bound to a glycolipid receptor detectable in lipid extracts from the particles, and (3) soluble Galalpha(1-4)Gal inhibited the adhesin by 72% from binding to surfactant-like particles. The Galalpha(1-4)Gal receptor present in the intestinal surfactant-like particle which overlies the intestinal mucosa could provide one means to establish an intestinal habitat for uropathogenic E. coli.

Effect of tissue non-specific alkaline phosphatase in maintenance of structure of murine colon and stomach.

The gastrointestinal tract of mammals secretes a phospholipid-rich membrane that is enriched in alkaline phosphatase (AP) and surfactant proteins (surfactant-like particle, SLP). The production of this particle is stimulated in the small intestine by fat feeding and in cultured cells in vitro by transfection with intestinal alkaline phosphatase (IAP). To test whether tissue non-specific alkaline phosphatase (TNAP) was a factor in stimulating surfactant-like particle production in stomach and colon (tissues expressing TNAP), mice lacking this enzyme were studied. Mice were harvested at 8 days of life, when body weight of homozygous animals (TNAP -/-) was about half that of congenic controls (TNAP +/+) or heterozygotes (TNAP +/-), but before seizures had begun. No difference in content of the major SLP protein (65 kDa) by Western blotting or immunocytochemistry was seen in stomach or colon of TNAP -/- vs. TNAP +/+ animals, but the content was only about half in the IAP-expressing small bowel. Transmission electron microscopy of the TNAP -/- small bowel showed large dilated lysosomes and residual bodies. Colonocytes and gastric surface epithelial cells from the same animals showed mitochondria containing homogeneous dense inclusions, consistent with neutral lipid. In the underweight homozygous animals, there was a decrease in the neuronal content of submucosal ganglia in the jejunum and ileum and of myenteric ganglia in the jejunum of TNAP -/- animals. These findings suggest that (1) TNAP is not important in maintaining surfactant-like particle content of tissues that express TNAP, (2) normal fat absorption is important in maintaining SLP content in the small intestine, and (3) TNAP is important in the maintenance of some intestinal structures, and perhaps their function.

Measuring self-efficacy in substance abuse intervention in obstetric practices.

BACKGROUND: This study presents findings on the effect of an office-based obstetrics training program (given the acronym OBIWOM) on substance use assessment, management, and referral self-efficacy among obstetricians and staff of private, community-based obstetric practices. METHODS: Participants were obstetricians and staff from 10 of 27 available community-based, private obstetric practices in the target areas, for a practice participation rate of 37%. This study used a delayed treatment design to compare self-efficacy between practice staff randomly assigned to an immediate or delayed intervention group. RESULTS: Self-efficacy increased significantly after intervention from baseline to first follow-up for the immediate group, while no change was shown for the delayed (control) group. The impact of the intervention on self-efficacy was replicated in the assessment construct only after the intervention for the delayed group. CONCLUSION: This research shows that education and training can effectively improve self-efficacy in obstetricians and their staff in the management of substance use and pregnancy.

Intestinal fatty acid binding protein may favor differential apical fatty acid binding in the intestine.

The intestinal mucosa metabolizes fatty acids differently when presented to the lumenal or basolateral membrane. Expression of both liver and intestinal fatty acid binding proteins (L- and I-FABPs) uniquely in the enterocyte offers a possible explanation of this phenomenon. An organ explant system was used to analyze the relative binding of fatty acids to each protein. More fatty acid was bound to L-FABP than to I-FABPs (28% vs. 6% of cytosolic radioactivity), no matter on which side the fatty acid was added. However, a 2-3-fold increase in fatty acid binding to the intestinal paralog was noted after apical addition of palmitic or oleic acid in mucosa from chow fed rats. When oleic acid was added apically, a 1.4-fold increase in binding to I-FABP was observed in mucosa derived from chronically fat fed rats, consistent with the previously observed 50% increase in the content of that protein. Immunocytochemical localization of both FABPs in vivo demonstrated an apical cytoplasmic localization in the fasting state, and redistribution to the entire cytoplasm after fat feeding. These data are consistent with the hypothesis that I-FABP may contribute to the metabolic compartmentalization of apically presented fatty acids in the intestine.

Binding of pili from uropathogenic Escherichia coli to membranes secreted by human colonocytes and enterocytes.

PapG adhesins mediate the binding of uropathogenic Escherichia coli. Although receptors for these adhesins have not been demonstrated in intestinal epithelia, the colonic microflora includes strains of uropathogenic E. coli. We now report that surfactant-like particles secreted by the human intestine contain receptors for PapG adhesins and may provide an intestinal habitat for uropathogenic bacteria.

Overexpression of nerve growth factor in skin selectively affects the survival and functional properties of nociceptors.

Mice that overexpress nerve growth factor (NGF-OE) in the skin have double the normal number of cutaneous sensory neurons, have increased innervation of the skin and spinal cord, and are hyperalgesic. Here, we have asked whether the increased cutaneous NGF level results in a selective survival of only certain functional types of neurons and whether it changes the properties of cutaneous neurons. Using electron microscopy, we show that the number of both myelinated and unmyelinated nociceptors increases substantially in NGF-OE mice by a factor of 3.3 and 1.5, respectively. Using extracellular recordings from single units, we demonstrate that large myelinated (Abeta) fibers are unchanged in prevalence and receptive properties. In contrast, among thin myelinated (Adelta) fibers, the percentage of nociceptors increased from a normal 65 to 97%, consistent with a selective survival of nociceptors during embryogenesis. These afferents showed a twofold increase in their mechanical responsiveness, but their heat responsiveness remained normal. Among unmyelinated (C) fibers, there was a profound increase in the percentage of heat responsive neurons from a normal 42 to 96%. This change cannot be accounted for by a selective survival of heat-sensitive neurons. Unmyelinated nociceptors increased fourfold in their thermal responsiveness but decreased in mechanical responsiveness. Therefore, target-derived NGF selectively rescues nociceptors during the period of programmed cell death with different efficacy for thin myelinated or unmyelinated fibers. NGF also affects the response to noxious heat or mechanical stimuli in each group differently, implying specific regulations of transduction processes rather than general changes of excitability.