The present study aimed to evaluate the ability of a novel serum-free medium (SFM) to culture human airway epithelium cells (hAECs). hAECs were cultured in the novel SFM as the experimental group in the PneumaCult-Ex medium and Dulbecco's modified eagle medium (DMEM) and fetal bovine serum (FBS) as the control groups. Cell morphology, proliferative capacity, differentiation capacity and expression levels of basal cell markers were assessed accordingly in both culture systems. Optical microscope photos of hAECs were collected for cell morphology assessment. Cell Counting Kit-8 assay was conducted to assess the proliferation ability, and an air-liquid interface (ALI) assay was conducted to assess the differentiation capacity. Markers for proliferating basal and differentiated cells were relatively identified by immunohistochemical and immunofluorescent analysis. The results show that whether grown in the novel SFM or Ex medium, hAECs exhibited similar morphology at every passage, whereas cells could hardly form colonies in the DMEM + FBS group. Cells typically exhibited cobblestone shape, while a proportion of them in the novel SFM at late passage exhibited a larger shape. White vesicles appeared in the cytoplasm of some control cells at the later stage of culture. Basal cell markers (P63 + KRT5 + KI67 + CC10 -) for proliferating ability were found in the hAECs cultured by the novel SFM and Ex medium. hAECs at passage 3 cultured in the novel SFM and Ex medium both had the capacity to differentiate into ciliated cells (acetylated tubulin + ), goblet cells (MUC5AC + ) and club cells (CC10 + ) in the ALI culture assay. In conclusion, the novel SFM was capable of culturing hAECs. The hAECs cultured by the novel SFM could proliferate and differentiate in vitro. The novel SFM does not change the morphological characteristics or biomarkers of hAECs. The novel SFM has the potential for the amplification of hAECs for scientific research and clinical application.

Background: Recently, the central position of small airways in the pathogenesis of idiopathic pulmonary fibrosis(IPF) has been gradually recognized and accepted with evidence of small airway anatomical and genetic abnormalities. However, the small airway physiology of IPF patients remained unclear. This study aimed to assess the small airway physiology of IPF patients using pulmonary function test (PFT). Methods: We retrospectively analyzed 138 IPF patients' and 186 control patients' small airway lung function data and medical records. A 1:1 propensity score match(PSM) for age, sex, smoking status and BMI was completed priors to the group comparison between IPF patients and controls. In addition, patients were divided into three groups according to FVC decline degree with cut-off values of 75% and 55%, and the small airway function was compared between the three groups. The impulse oscillometry system (IOS) diagnosis value was evaluated in 60 IPF patients. Results: The total cohort comprised 138 subjects, with 53(38.4%) diagnosed with SAD. After PSM, IPF patients were predisposed to SAD compared with controls (38.0% vs.15.2%, p=0.001). Compared small airway function between patients with different disease severities, we found even for patients in the mild disease group whose FVC is almost normal, 30.6% (n=22) were diagnosed with spirometry-SAD. And we found MEF50(98.2% vs. 87.4% vs. 52.4%, p=0.001), MEF25(74.2% vs.71.9% vs. 61.3%, p=0.062) and MMEF(77.2% vs. 74.6% vs. 44.6%, p=0.002) were deteriorated with the severity of disease. The incidence of spirometry-SAD in more severe patients was higher than those with less severe disease(p=0.021). Applying IOS to 60 IPF patients, 41(68.3%) were diagnosed with IOS-SAD. There was poor concordance between spirometry and IOS for the diagnosis of SAD (Kappa value=-0.068, p=0.542). IOS showed higher sensitivity than spirometry. For patients with abnormal lung function (FVC<80%),this advantage could be even more obvious. Conclusions: This study revealed that SAD was a common lesion of IPF patients. It should be emphasized that SAD could occur at the early stages of disease when the lung capacity has not decreased, and the incidence and degree of dysfunction were progressively aggravated with disease progression. Additionally, IOS was a powerful complementary tool for diagnosing SAD, especially for IPF patients with decreased lung function.