Serodiagnosis of human immunodeficiency virus (HIV) infection in the United States has traditionally relied on a sequential two-test algorithm: an initial screen with an enzyme immunoassay (EIA) and ...reflex testing of EIA-reactive specimens with a more specific supplemental test such as Western blotting or immunofluorescence. The supplemental tests are tedious, subjective, and expensive. In addition, there have been major improvements in the performance and accuracy of the EIA tests as well as the introduction of rapid serologic tests (RT) and HIV nucleic acid amplification tests (NAAT). Related to these improvements is the possibility that alternative algorithms using combinations of currently approved HIV tests may function as well as if not better than the current algorithm, with more flexibility, improved accuracy, and lower cost. To this end, we evaluated the performance of 12 currently licensed tests and 1 in-house HIV test (6 EIA, 4 RT, and 3 NAAT) on panels of plasma samples from HIV-infected (n = 621 HIV type 1 HIV-1 and 34 HIV-2) and uninfected (n = 513) people and of sequential specimens from people early in seroconversion (183 specimens from 15 patients). Test combinations were analyzed in two dual-test (sensitivity-optimized and specificity-optimized) algorithms and in a three-test (tie-breaking) algorithm, and performance was compared to the conventional algorithm. The results indicate that alternative algorithm strategies with currently licensed tests compare favorably with the conventional algorithm in detecting and confirming established HIV infection. Furthermore, there was a lower frequency of discordant or indeterminate results that require follow-up testing, and there was improved detection of early infection.
Cytochromes P450 of the CYP2C and CYP4A gene subfamilies metabolize arachidonic acid to 5,6-, 8,9-, 11,12-, and 14,15-epoxyeicosatrienoic acids (EETs) and to 19-
and 20-hydroxyeicosatetraenoic acids ...(HETEs), respectively. Abundant functional studies indicate that EETs and HETEs display
powerful and often opposing biological activities as mediators of ion channel activity and regulators of vascular tone and
systemic blood pressures. Incubation of 8,9-, 11,12-, and 14,15-EETs with microsomal and purified forms of rat CYP4A isoforms
led to rapid NADPH-dependent metabolism to the corresponding 19- and 20-hydroxylated EETs. Comparisons of reaction rates and
catalytic efficiency with those of arachidonic and lauric acids showed that EETs are one of the best endogenous substrates
so far described for rat CYP4A isoforms. CYP4A1 exhibited a preference for 8,9-EET, whereas CYP4A2, CYP4A3, and CYP4A8 preferred
11,12-EET. In general, the closer the oxido ring is to the carboxylic acid functionality, the higher the rate of EET metabolism
and the lower the regiospecificity for the EET Ï-carbon. Analysis of cis -parinaric acid displacement from the ligand-binding domain of the human peroxisome proliferator-activated receptor-α showed
that Ï-hydroxylated 14,15-EET bound to this receptor with high affinity ( K
i = 3 ± 1 n m ). Moreover, at 1 μ m , the Ï-alcohol of 14,15-EET or a 1:4 mixture of the Ï-alcohols of 8,9- and 11,12-EETs activated human and mouse peroxisome
proliferator-activated receptor-α in transient transfection assays, suggesting a role for them as endogenous ligands for these
orphan nuclear receptors.
ABSTRACT
Saturated fatty acids (SFAs) have been shown to induce endoplasmic reticulum (ER) stress and chronic inflammatory responses, as well as alter sphingolipid metabolism. Disruptions in ER ...stress and sphingolipid metabolism have also been implicated in intestinal inflammation. Therefore, to elucidate the roles of SFAs in ER stress and inflammation in intestinal epithelial cells, we examined myristate (C14:0) and palmitate (C16:0). My‐ ristate, but not palmitate, induced ER stress signaling, including activation of inositol‐requiring enzyme 1 (IRE1) and X‐box binding protein 1 (XBP1) signaling. Myristate significantly increased C14‐ceramide levels, whereas palmitate increased several long‐chain ceramides. To define the role of ceramide synthases (CerSs) in myristate‐induced ER stress, we used the pharmacologic inhibitor, fumonisin B1 (FB1), and small interfering RNA (siRNA) for CerS5 and 6, the primary isoforms that are involved in C14‐ceramide generation. FB1 and siRNA for CerS5 or 6 suppressed myristate‐induced C14‐ceramide generation and XBP1 splicing (XBP1s). Moreover, increased XBP1s induced the downstream expression of IL‐6 in a CerS5/6‐dependent manner. In addition, a myristate‐enriched milk fat‐ based diet, but not a lard‐ based diet, increased C14‐ ceramide, XBP1s, and IL‐ 6 expression in vivo. Taken together, our data suggest that myristate modulates ER stress and cytokine production in the intestinal epithelium via CerS5/6 and C14‐ ceramide generation.—Choi, S., Snider, J. M., Olakkengil, N., Lambert, J. M., Anderson, A. K., Ross‐Evans, J. S., Cowart, L. A., Snider, A. J. Myristate‐induced endoplasmic reticulum stress requires ceramide synthases 5/6 and generation of C14‐ceramide in intestinal epithelial cells. FASEB J. 32, 5724–5736 (2018). www.fasebj.org
Background
Sinonasal biofilms have been demonstrated in specimens collected from chronic rhinosinusitis (CRS) patients. Mounting evidence suggests that biofilms contribute to therapeutically ...recalcitrant CRS. Recently, the bitter taste receptor T2R38 has been implicated in the regulation of the sinonasal mucosal innate immune response. TAS2R38 gene polymorphisms affect receptor functionality and contribute to variations seen in sinonasal innate defense as well as taste perception reflected in gustatory sensitivity to the bitter compound phenylthiocarbamide (PTC). In a population of CRS patients with active infection or inflammation, we sought to determine if a correlation between T2R38 phenotype and in vitro biofilm formation existed.
Methods
Endoscopically guided sinonasal swabs were obtained prospectively from CRS (±polyp) patients with evidence of persistent inflammation or mucopurulence. In vitro biofilm formation was assessed with a modified Calgary Biofilm Detection Assay. Patients’ phenotypic (functional) expression of the bitter taste receptor T2R38 was evaluated with a taste test including the compound PTC. Linear regression was used to determine the level of significance between mean in vitro biofilm formation levels and mean PTC taste test intensity ratings across CRS patients.
Results
Sinonasal swabs were obtained from 59 patients, with 42 of the 59 samples demonstrating in vitro biofilm formation. Analysis revealed an inverse linear association between in vitro biofilm formation and PTC taste intensity ratings (p = 0.019) for all patients. This association was exclusively driven by nonpolypoid CRS patients (p = 0.0026).
Conclusion
In vitro biofilm formation from sinonasal clinical isolates is inversely correlated with PTC taste sensitivity in nonpolypoid CRS patients.
The current study describes the development of a unique real-time PCR assay for the detection of mutations conferring drug resistance in Mycobacterium tuberculosis. The rifampicin resistance ...determinant region (RRDR) of rpoB and specific regions of katG and the inhA promoter were targeted for the detection of rifampin (RIF) and isoniazid (INH) resistance, respectively. Additionally, this assay was multiplexed to discriminate Mycobacterium tuberculosis complex (MTC) strains from nontuberculous Mycobacteria (NTM) strains by targeting the IS6110 insertion element. High-resolution melting (HRM) analysis following real-time PCR was used to identify M. tuberculosis strains containing mutations at the targeted loci, and locked nucleic acid (LNA) probes were used to enhance the detection of strains containing specific single-nucleotide polymorphism (SNP) transversion mutations. This method was used to screen 252 M. tuberculosis clinical isolates, including 154 RIF-resistant strains and 174 INH-resistant strains based on the agar proportion method of drug susceptibility testing (DST). Of the 154 RIF-resistant strains, 148 were also resistant to INH and therefore classified as multidrug resistant (MDR). The assay demonstrated sensitivity and specificity of 91% and 98%, respectively, for the detection of RIF resistance and 87% and 100% for the detection of INH resistance. Overall, this assay showed a sensitivity of 85% and a specificity of 98% for the detection of MDR strains. This method provides a rapid, robust, and inexpensive way to detect the dominant mutations known to confer MDR in M. tuberculosis strains and offers several advantages over current molecular and culture-based techniques.
Abstract A multiplex real-time PCR assay for the detection of Mycoplasma pneumoniae (MP181), Chlamydia ( Chlamydophila ) pneumoniae (CP-Arg), Legionella spp. (Pan-Leg), and the human RNase P (RNase ...P) gene was developed for rapid testing of atypical bacterial respiratory pathogens in clinical specimens. This method uses 4 distinct hydrolysis probes to detect 3 leading causes of community-acquired pneumonia. The assay was evaluated for specificity and sensitivity by testing against 35 related organisms, a dilution series of each specific target and 197 clinical specimens. Specificity testing demonstrated no cross-reactivity. A comparison to previously validated singleplex real-time PCR assays for each agent was also performed. The analytical sensitivity for specific pathogen targets in both the singleplex and multiplex was identical (50 fg), while efficiencies ranged from 82% to 97% for the singleplex assays and from 90% to 100% for the multiplex assay. The clinical sensitivity of the multiplex assay was improved for the Pan-Leg and CP-Arg targets when compared to the singleplex. The MP181 assay displayed equivalent performance. This multiplex assay provides an overall improvement in the diagnostic capability for these agents by demonstrating a sensitive, high-throughput and rapid method. This procedure may allow for a practical and efficient means to test respiratory clinical specimens for atypical pneumonia agents in health care settings and facilitate an appropriate public health response to outbreaks.
To examine trends in HIV prevalence in the US household population, serum or urine samples from 2 National Health and Nutrition Examinations Surveys (NHANES) (1988-1994 and 1999-2002), were tested ...for HIV antibody. In the 1999 to 2002 survey, data on risk behaviors, CD4 T lymphocytes, and antiretroviral therapy (ART) were also available. In the 1988 to 1994 survey, there were 59 positive individuals of 11,203 tested. In NHANES 1999 to 2002, there were 32 positive individuals of 5926 tested. The prevalence of HIV infection among those aged 18 to 39 years in NHANES 1988 to 1994 was 0.38% (95% confidence interval CI: 0.22-0.68) as compared with 0.37% (95% CI: 0.17 to 0.80) in 1999 to 2002. Prevalence did not change significantly between surveys in any race and/or ethnic or gender group among 18- to 39-year-old participants. HIV prevalence was 3.58% (95% CI: 1.88 to 6.71) among non-Hispanic blacks in the 40- to 49-year-old age group in 1999 to 2002, but the age range available in NHANES 1988 to 1994 was 18 to 59 years and does not allow direct comparison of prevalence. Cocaine use and the presence of herpes simplex virus-2 antibody were the only significant risk factors for HIV infection for non-Hispanic blacks. Fifty-eight percent of infected individuals not reporting ART had CD4 T-lymphocyte counts < 200 cells/mm3 compared with 18.2% on therapy and 12.5% of participants newly informed of their HIV status.
Gustation assessment using the NIH Toolbox Coldwell, Susan E; Mennella, Julie A; Duffy, Valerie B ...
Neurology,
03/2013, Letnik:
80, Številka:
11 Suppl 3
Journal Article
Recenzirano
Odprti dostop
The NIH Toolbox for Assessment of Neurological and Behavioral Function (NIH Toolbox) is a set of brief measures for the assessment of cognitive function, emotional health, motor function, and sensory ...function for use in clinical trials and in epidemiologic and longitudinal studies. Gustatory perception is assessed as 1 of 6 areas of sensory function. A team of 11 scientists with expertise in taste perception selected 2 gustatory measures, 1 of which can be used in young pediatric populations. The measure selected for young pediatric populations assesses sucrose (sweet) taste preference and can also be used across the age span of 5 to 85 years. For adult populations, the selected measure is a regional test, which assesses variability in perceived intensity of quinine hydrochloride (bitter) when applied to the tongue tip as well as perceived with the whole mouth. The team also recommends the regional test for assessing other tastants, such as sodium chloride (salty). Validation studies have demonstrated that the measures modified for the NIH Toolbox correlate with more traditional assessments, and can identify known population differences in gustation.
Pulmonary fibrosis is an interstitial scarring disease of the lung characterized by poor prognosis and limited treatment options. Tissue transglutaminase 2 (TG2) is believed to promote lung fibrosis ...by crosslinking extracellular matrix components and activating latent TGFβ. This study assessed physiologic pulmonary function and metabolic alterations in the mouse bleomycin model with TG2 genetic deletion. TG2‐deficient mice demonstrated attenuated the fibrosis and preservation of lung function, with significant reduction in elastance and increases in compliance and inspiratory capacity compared to control mice treated with bleomycin. Bleomycin induced metabolic changes in the mouse lung that were consistent with increased aerobic glycolysis, including increased expression of lactate dehydrogenase A and increased production of lactate, as well as increased glutamine, glutamate, and aspartate. TG2‐deficient mice treated with bleomycin exhibited similar metabolic changes but with reduced magnitude. Our results demonstrate that TG2 is required for a typical fibrosis response to injury. In the absence of TG2, the fibrotic response is biochemically similar to wild‐type, but lesions are smaller and lung function is preserved. We also show for the first time that profibrotic pathways of tissue stiffening and metabolic reprogramming are interconnected, and that metabolic disruptions in fibrosis go beyond glycolysis.
Tissue transglutaminase 2 (TG2) knockout mice demonstrated preserved lung function and reduced lesion area following bleomycin injury. Bleomycin fibrotic injury is associated with energy and amino acid metabolic changes. These metabolic changes are similar but overall reduced in magnitude in the TG2 KO mice. We also show for the first time that profibrotic pathways of tissue stiffening and metabolic reprogramming are interconnected, and that metabolic disruptions in fibrosis go beyond glycolysis.
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