4-kDa FITC-dextran was added to the apical chamber and fresh medium to the basolateral chamber. After 8 h, the fluorescein level in the basolateral chamber was detected using a microplate reader .Airway organoids were placed under an inverted phase contrast microscope equipped with a high-speed digital video camera. Triplicate video recordings were made from different fields of view for every set of airway organoids and analyzed using a Sisson-Ammons Video Analysis system for their CBFs.
Multiple DEG lists were uploaded to Metascape, a web-based portal providing functional enrichment, interaction analysis, gene annotation, and membership search, leveraging over 40 independent knowledgebases [values of less than 0.05 were also submitted to Ingenuity Pathway Analysis software for pathway analysis.Apical secretion of airway organoids was collected with DPBS. Protein in the samples were extracted and digested for LC–MS/ MS analysis.
All data analysis was expressed as the mean ± SEM. When confirmed the data are normally distributed, the differences between samples were analyzed using unpaired Student’s t test for two groups and two-way ANOVA for multiple comparisons with VD treatment and H1N1/infection. For clinical data analysis, Chi-square test was used for contingency analysis of the proportion among groups and one-way ANOVA was used for multiple comparisons of every clinical indicator.
: Fig. S2 lists the clinical characteristics, categorized by different diseases. Among all the groups, the concentration of serum 25D, a relatively stable circulating form of VD, was lower than the expected cutoff level 30 ng/mL, fluctuating by 25 ng/mL. Indicators associated with VD, including parathyroid hormone , calcitonin , and serum calcium and phosphate, were all in normal ranges, as were the peripheral granulocytes, including eosinophils , basophils , and neutrophils .
To determine the degrees of VD deficiency, we set the cutoff value to be 20 ng/mL; we defined a mild low 25Dstatus of 20–30 ng/mL as “VD insufficiency” and a status lower than 20 ng/mL as “VD deficiency.” By this standard, 42.3% of the cases were of VD insufficiency, while 29.6% were of VD deficiency. In total, 72.8% of patients were in a low VD status. The proportions of VD deficiency in the CRSwNP and AR groups were, however, greater, up to 39.13% and 40.
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