Synthesis of high sensitivity C-reactive protein in Beas2B cells and nasal epithelial cells of patients with cystic fibrosis

Introduction High sensitivity C-reactive protein (hsCRP) has an upcoming role in predicting pulmonary inflammation and clinical disease progression in lung diseases. HsCRP may predict clinical disease activity and risk of pulmonary exacerbations in adult cystic fibrosis (CF) patients. Furthermore, hsCRP can be synthesized by airway epithelial cells in patients with chronic obstructive pulmonary disease (COPD). Preliminary analysis of 32 CF patients showed a strong correlation of hsCRP with proinflammatory cytokines and lung clearance index (LCI). Our aim was to investigate the role of airway epithelium as a source of systemic hsCRP levels in CF patients.

Methods Beas2Bs (human immortalized bronchial epithelial cells) and human nasal epithelial cells (hNEC, on air liquid interface) from CF patients (n=4, mean age (SD): 14.85 (1.92) years, pseudomonas negative, 3 male, F508del/F508del (2x), F508del/K68X, Q525X/Q525X) and healthy controls (n=4, mean age (SD): 42.73 (1.46) years, 2 male) were stimulated for 24h (interleukin 6 (IL-6), interleukin 1β (IL-1β), phosphate buffered saline (PBS), 10 ng/ml and 100 ng/ml). The media was collected and concentrated for determining the secreted CRP via Enzyme-linked Immunosorbent Assay (ELISA, Human C-Reactive Protein ELISA Kit, ab99995, abcam, UK). Furthermore, we performed a quantitative reverse transcription Polymerase Chain Reaction (RT-qPCR, PrimeTime Std qPCR Assay, CRP: Hs.PT.58.3341389.g, GAPDH: Hs.PT.39a.22214836, integrated DNA technologies, US) with isolated RNA to investigate the expression levels of CRP.

Results IL-6- and IL-1β-stimulated Beas2Bs showed significant higher CRP expression levels (Mann-Whitney U test; IL-6 10 ng/ml: Δct=-15.06, p<0.05; IL-6 100 ng/ml: Δct=-14.36, p<0.001; IL-1β 10 ng/ml: Δct=-14.93, p<0.05, IL-1β 100 ng/ml: Δct=-14.14, p<0.001; PBS: Δct=-15.72; N=8) in RT-qPCR and higher CRP concentrations in the supernatant measured via ELISA in comparison to the control (PBS) after 24h. There was no expression of CRP in differentiated hNEC from CF patients or healthy controls. In line with this, CRP was undetectable via ELISA in the supernatant of hNEC.

Conclusion Our results showed that IL-6 and IL-1β stimulated Beas2Bs produce CRP. These findings could not be replicated in hNEC of CF patients and healthy controls. It remains unclear, which cell type contributes to elevated hsCRP serum levels in CF patients and worsens LCI. Thus, further investigation is needed.

Publication History

Article published online:
09 March 2023