Evaluation of the use of reporter antigens in an auricular lymph node assay to assess the immunosensitizing potential of drugs.

Immune-mediated idiosyncratic drug reactions are a major problem for susceptible patients, physicians, and the pharmaceutical industry. Validated screening tools to assess the immunosensitizing capacity of orally or intravenously administered pharmaceuticals are currently not available. To date, the popliteal lymph node assay (PLNA) seems the most promising tool for this purpose. The PLNA has recently been extended with the use of reporter antigens (RA) that are coinjected together with the drug of interest. The measurement of isotypes of RA-specific antibody-secreting cells (ASC) enables the distinction of sensitizing chemicals and (nonsensitizing) irritants without radio-isotopic end points. However, the use of footpad injections raises ethical concerns. Therefore, we examined the use of RA after intradermal injection into the ear of BALB/c mice and measured RA-specific ASC in the draining auricular lymph node (ALN). We show that RA-specific IgG isotype ASC numbers are very useful and sensitive parameters to identify drug-induced hypersensitivity in both PLN and ALN. However, the type 1-associated parameters (CD8(+) cells, macrophages, IFN-gamma, TNF-alpha, and IL-1 beta) that are induced in the PLN by streptozotocin were less pronounced in the ALN. Thus, the PLNA may provide more immunologically relevant information on the mechanisms of certain chemical-induced hypersensitivity reactions. The RA-ALN assay may provide an alternative for the RA-PLNA; both assays can be used to distinguish sensitizing compounds from nonsensitizing ones.     

Presentation and interpretation of food intake data: Factors affecting comparability across studies

Non-uniform, unclear, or incomplete presentation of food intake data limits interpretation, usefulness, and comparisons across studies. In this contribution, we discuss factors affecting uniform reporting of food intake across studies. The amount of food eaten can be reported as mean portion size, number of servings or total amount of food consumed per day; the absolute intake value for the specific study depends on the denominator used because food intake data can be presented as per capita intake or for consumers only. To identify the foods mostly consumed, foods are reported and ranked according to total number of times consumed, number of consumers, total intake, or nutrient contribution by individual foods or food groups. Presentation of food intake data primarily depends on a study's aim; reported data thus often are not comparable across studies. Food intake data further depend on the dietary assessment methodology used and foods in the database consulted; and are influenced by the inherent limitations of all dietary assessments. Intake data can be presented as either single foods or as clearly defined food groups. Mixed dishes, reported as such or in terms of ingredients and items added during food preparation remain challenging. Comparable presentation of food consumption data is not always possible; presenting sufficient information will assist valid interpretation and optimal use of the presented data. A checklist was developed to strengthen the reporting of food intake data in science communication.     

A High Proliferation Rate is Critical for Reproducible and Standardized Embryoid Body Formation from Laminin-521-Based Human Pluripotent Stem Cell Cultures

When aiming for homogenous embryoid body (EB) differentiation, the use of equal-sized EBs is required to avoid a size-induced differentiation bias. In this study we developed an efficient and standardized EB formation protocol for human pluripotent stem cells (hPSC) cultured in a laminin-521-based xeno-free system. As the cell proliferation rate of the cells growing on laminin-521 strongly affected the efficiency of aggregate formation, we found that recently passaged cells, as well as the addition of ROCK inhibitor, were essential for reproducible EB formation from hPSC single-cell suspensions. EBs could be obtained in a variety of differentiation media, in 96-well round-bottom plates and in hanging drops. Gene expression studies on differentially sized EBs from three individual human embryonic stem cell lines demonstrated that the medium used for differentiation influenced the differentiation outcome to a much greater extent than the number of cells used for the initial EB formation. Our findings give a new insight into factors that influence the EB formation and differentiation process. This optimized method allows us to easily manipulate EB formation and provide an excellent starting point for downstream EB-based differentiation protocols.     

Dietary Diversity and Vegetable and Fruit Consumption of Households in a Resource-Poor Peri-Urban South Africa Community Differ by Food Security Status

Sociodemographic, living standard measure, consumption of vegetables and fruit, and dietary diversity in relation to household food security were assessed. Using a hunger score, households were categorized as food secure (n = 125) or food insecure (n = 273). Food secure respondents had a higher mean dietary diversity score (3.98; 95%CI [3.79, 4.18] versus 3.65; 95% [CI 3.53, 3.77]), were more likely to eat vitamin A–rich foods (OR 1.15; 95% CI [1.05, 1.26]), a more varied diet (DDS ≥ 4, OR 1.90; 95% CI [1.19, 3.13]), and vegetables daily (OR 3.37; 95% CI [2.00, 5.76]). Cost limited daily vegetable/fruit consumption in food insecure households. Respondents with ≥ 8 years of schooling were more likely (OR 2.07; 95% CI [1.22, 3.53]) and households receiving social grants were less likely (OR 0.37; 95% CI [0.19, 0.72]) to be food secure. Results highlight the association between dietary diversity and household food security.     

Automatic blood pressure measurement: the oscillometric waveform shape is a potential contributor to differences between oscillometric and auscultatory pressure measurements

OBJECTIVE: To explore the differences between oscillometric and auscultatory measurements. METHOD: From a simulator evaluation of a non-invasive blood pressure (NIBP) device regenerating 242 oscillometric blood pressure waveforms from 124 subjects, 10 waveforms were selected based on the differences between the NIBP (oscillometric) and auscultatory pressure measurements. Two waveforms were selected for each of five criteria: systolic over and underestimation; diastolic over and underestimation; and close agreement for both systolic and diastolic pressures. The 10 waveforms were presented to seven different devices and the oscillometric-auscultatory pressure differences were compared between devices and with the oscillometric waveform shapes. RESULTS: Consistent patterns of waveform-dependent over and underestimation of systolic and diastolic pressures were shown for all seven devices. The mean and standard deviation, for all devices, of oscillometric-auscultatory pressure differences were: for the systolic overestimated waveforms, 36 +/- 28/-6 +/- 3 and 23 +/- 2/-1 +/- 3 mmHg (systolic/diastolic differences); for systolic underestimated waveforms, -21 +/- 5/-4 +/- 3 and -11 +/- 4/-3 +/- 3 mmHg; for diastolic overestimated waveforms, 3 +/- 4/12 +/- 5 and 17 +/- 6/10 +/- 2 mmHg; for diastolic underestimated waveforms, 1 +/- 4/-22 +/- 4 and -9 +/- 6/-29 +/- 4 mmHg; and for the two waveforms with good agreement, 0 +/- 6/0 +/- 3 and -2 +/- 4/-4 +/- 3 mmHg. Waveforms for which devices showed good oscillometric and auscultatory agreement had smooth envelopes with clearly defined peaks, compared with the broader plateau and complex shapes of those waveforms for which devices over or underestimated pressures. CONCLUSION: By increasing the understanding of the characteristics and limitations of the oscillometric method and the effects of waveform shape on pressure measurements, simulator evaluation should lead to improvements in NIBP devices.     

Further delineation of the KBG syndrome caused by ANKRD11 aberrations

Loss-of-function variants in ANKRD11 were identified as the cause of KBG syndrome, an autosomal dominant syndrome with specific dental, neurobehavioural, craniofacial and skeletal anomalies. We present the largest cohort of KBG syndrome cases confirmed by ANKRD11 variants reported so far, consisting of 20 patients from 13 families. Sixteen patients were molecularly diagnosed by Sanger sequencing of ANKRD11, one familial case and three sporadic patients were diagnosed through whole-exome sequencing and one patient was identified through genomewide array analysis. All patients were evaluated by a clinical geneticist. Detailed orofacial phenotyping, including orthodontic evaluation, intra-oral photographs and orthopantomograms, was performed in 10 patients and revealed besides the hallmark feature of macrodontia of central upper incisors, several additional dental anomalies as oligodontia, talon cusps and macrodontia of other teeth. Three-dimensional (3D) stereophotogrammetry was performed in 14 patients and 3D analysis of patients compared with controls showed consistent facial dysmorphisms comprising a bulbous nasal tip, upturned nose with a broad base and a round or triangular face. Many patients exhibited neurobehavioural problems, such as autism spectrum disorder or hyperactivity. One-third of patients presented with (conductive) hearing loss. Congenital heart defects, velopharyngeal insufficiency and hip anomalies were less frequent. On the basis of our observations, we recommend cardiac assessment in children and regular hearing tests in all individuals with a molecular diagnosis of KBG syndrome. As ANKRD11 is a relatively common gene in which sequence variants have been identified in individuals with neurodevelopmental disorders, it seems an important contributor to the aetiology of both sporadic and familial cases.     

Expression of toll-like receptors 2 and 4 in rheumatoid synovial tissue and regulation by proinflammatory cytokines interleukin-12 and interleukin-18 via interferon-gamma

OBJECTIVE: To study the expression of Toll-like receptor 2 (TLR-2) and TLR-4 and its association with proinflammatory cytokines in synovial tissue from patients with rheumatoid arthritis (RA), osteoarthritis (OA), and healthy individuals. METHODS: Synovial tissue specimens from 29 RA patients were stained for TLR-2, TLR-4, and proinflammatory cytokines (interleukin-1beta [IL-1beta], IL-12, IL-17, IL-18, and tumor necrosis factor alpha [TNFalpha]). The expression of TLR-2, TLR-4, and cytokines as well as the degree of inflammation in synovial tissue were compared between patients with RA, patients with OA (n = 5), and healthy individuals (n = 3). Peripheral blood mononuclear cells (PBMCs) were incubated with IL-12 and IL-18, and TLR expression was assessed using fluorescence-activated cell sorter analysis. Production of TNFalpha and IL-6 was measured using Luminex bead array technology. RESULTS: In RA synovial tissue, the expression of TLR-2 was slightly higher than that of TLR-4. Interestingly, both TLR-2 and TLR-4 were expressed at higher levels in moderately inflamed synovium, as compared with synovial tissue with no or severe inflammation. TLR expression in both the lining and the sublining was associated with the presence of IL-12 and IL-18, but no other cytokines, in the lining. The expression of both TLRs was low in synovial tissue from OA patients and healthy donors. Stimulation of PBMCs with IL-12 and IL-18 resulted in increased expression of both TLR-2 and TLR-4; this could be blocked with anti-interferon-gamma (anti-IFNgamma) antibodies, suggesting a role for IFNgamma. Lipopolysaccharide- or lipoteichoic acid-mediated triggering of PBMCs incubated with IL-12/IL-18 or IFNgamma led to an increased production of both TNFalpha and IL-6, indicating the functionality of TLR-2 and TLR-4. CONCLUSION: TLR-2 and TLR-4 are expressed in synovial tissue of patients with clinically active disease and are associated with the levels of both IL-12 and IL-18. The synergistic effect of IL-12 and IL-18 on T cell IFNgamma production seems to regulate expression of TLR-2 and TLR-4 in the synovial tissue of RA patients.