Supplementary Material for: Real-Time Sexing of Chicken Embryos and Compatibility with in ovo Protocols

<p>The chicken embryo is an established model system for studying early vertebrate development. One of the major advantages of this model is the facility to perform manipulations in ovo and then continue incubation and observe the effects on embryonic development. However, in common with other vertebrate models, there is a tendency to disregard the sex of the experimental chicken embryos, and this can lead to erroneous conclusions, a lack of reproducibility, and wasted efforts. That this neglect is untenable is emphasised by the recent demonstration that avian cells and tissues have an inherent sex identity and that male and female tissues respond differently to the same stimulus. These sexually dimorphic characteristics dictate that analyses and manipulations involving chicken embryos should always be performed using tissues/embryos of known sex. Current sexing protocols are unsuitable in many instances because of the time constraints imposed by most in ovo procedures. To address this lack, we have developed a real-time chicken sexing assay that is compatible with in ovo manipulations, reduces the number of embryos required, and conserves resources.</p

Erratum: The Functional IgE-Blocking Factor Induced by Allergen-Specific Immunotherapy Correlates with IgG4 Antibodies and a Decrease of Symptoms in House Dust Mite-Allergic Children

<b><i>Background:</i></b> At present, there are no validated biomarkers reflecting or predicting the clinical efficacy of allergen-specific immunotherapy (AIT) . We aimed to investigate the correlations between clinical and immunological responses of patients undergoing house dust mite (HDM) AIT. <b><i>Methods:</i></b> Sixty-nine children diagnosed with HDM allergic rhinitis and/or asthma received standardized <i>Dermatophagoides</i><i>pteronyssinus</i> (Dp) subcutaneous AIT for 12 months. Twenty HDM-allergic children served as an open control group. Clinical symptom and medication scores were recorded and Dp-specific IgE, IgG4 and IgE-blocking factor were measured before AIT and after 4 and 12 months of AIT. <b><i>Results:</i></b> Symptom scores decreased after 4 months and continued to decrease during 12 months of AIT. No differences in medication scores were observed between AIT and the control group during the study period. Levels of Dp IgG4 increased after 4 months and correlated to symptom scores at 12 months (r = -0.296, p = 0.013) of AIT. The Dp IgE-blocking factor increased after 4 months of AIT, and correlated with symptom scores at 4 months (r = -0.307, p = 0.010) and 12 months (r = -0.288, p = 0.016) of AIT. A strong correlation between Dp IgE-blocking factor and Dp IgG4 during AIT (4 months: r = 0.680; 12 months: r = 0.636, both p < 0.0001) was observed. Patients with IgE-blocking factor ≥0.2 after 4 months of AIT showed lower symptom scores at 12 months of AIT (p = 0.0093). <b><i>Conclusions:</i></b> Subcutaneous HDM AIT results in a decrease of allergic symptoms among HDM-allergic children. IgE-blocking activity increased after 4 months of AIT and correlated with clinical symptoms. A high IgE-blocking factor at an early stage of AIT is associated with fewer symptoms at a later stage of AIT

Supplementary Material for: Serum Vitamin D Concentrations and Cognitive Change Over 20 Years: The Atherosclerosis Risk in Communities Neurocognitive Study

<b><i>Background/Aims:</i></b> 25-hydroxyvitamin D (25[OH]D) concentrations have been associated with cognitive decline and incident dementia in elderly populations; however, these relationships are susceptible to reverse causation. Less is known about the association of midlife 25(OH)D with long-term cognitive decline. <b><i>Methods:</i></b> This was a prospective cohort study of 13,044 participants (mean age 57 years at baseline) in the Atherosclerosis Risk in Communities Study. 25(OH)D was measured from serum collected at baseline (1990–1992) using liquid chromatography tandem high-sensitivity mass spectrometry. Cognition was assessed using 3 neuropsychological tests at 3 time points, which were combined into a composite cognitive <i>Z</i>-score. Multivariable-adjusted linear mixed-effects models with random intercepts and slopes were used to estimate associations between 25(OH)D and cognitive change over 20 years. <b><i>Results:</i></b> Compared to persons with sufficient 25(OH)D (≥30 ng/mL), those with deficient (< 20 ng/mL) and intermediate (20–< 30 ng/mL) 25(OH)D concentrations had similar cognitive decline in composite cognitive <i>Z</i>-scores (deficient versus sufficient: –0.035 [95% CI –0.104 to 0.033] and intermediate versus sufficient: –0.029 [95% CI –0.080 to 0.023]). <b><i>Conclusions:</i></b> Lower concentrations of 25(OH)D measured in midlife were not significantly associated with more rapid cognitive decline over a 20-year follow-up period. The results of this prospective study are less susceptible to reverse causation than prior studies