Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2006 Jul;35(1):95-102.
doi: 10.1165/rcmb.2005-0305OC. Epub 2006 Mar 2.

Modulation of MUC7 mucin expression by exogenous factors in airway cells in vitro and in vivo

Shimin Li  1 , Giuseppe Intini, Libuse A Bobek
Affiliations

Modulation of MUC7 mucin expression by exogenous factors in airway cells in vitro and in vivo

Shimin Li et al. Am J Respir Cell Mol Biol. 2006 Jul.

Abstract

The human MUC7 gene encodes a low-molecular-mass mucin that participates in the maintenance of healthy epithelium in the oral cavity, and possibly in respiratory tracts, by promoting the clearance of various bacteria. We examined whether MUC7 gene is expressed in primary normal human tracheobronchial epithelial cells and whether the expression is modulated by exogenous factors. By assessing MUC7 transcripts, we found that the MUC7 gene was induced by culturing the normal human tracheobronchial epithelial cells at the air-liquid interface, in which the cells were well differentiated. When the cells were treated with a panel of cytokines (IL-1beta, IL-4, IL-13, and TNF-alpha), epidermal growth factor, or a bacterial product (Pseudomonas aeruginosa lipopolysaccharide [LPS]), MUC7 transcripts and glycoprotein products were increased 1.7- to 3.2-fold. The effect of LPS on MUC7 gene expression was also studied in the airway tissues of MUC7 gene transgenic mice. In the in vitro cultured trachea and lung explants, the LPS-treated tissues showed over 2-fold increased levels of MUC7 mRNA compared with the untreated specimens. These results were confirmed by in vivo studies using the lungs and tracheas harvested from the transgenic mice irritated by LPS through the tracheal instillation. By immunohistochemistry, MUC7 glycoprotein was localized in tracheal submucosa within the serous cells. Upon LPS stimulation, the overexpressed MUC7 remains confined to the serous glands. In the lungs, MUC7 seems to be expressed within the respiratory epithelium at the level of the bronchioles. Upon stimulation with LPS, it seems to be overexpressed within the same cells and within the stromal tissue.

PubMed Disclaimer

Figures

<b>Figure 1.</b>
Figure 1.
Analysis of NHTBE cells and MUC7 expression in NHTBE cells cultured under different conditions. (A) A semiquantitative RT-PCR analysis of the MUC7 gene transcription. NHTBE cells were cultured in ALI or squamous conditions, respectively. Total RNA was isolated from those cultures every day from Day 1 to Day 10. The levels of MUC7 mRNA were assessed by semiquantitative RT-PCR, and analyzed by agarose gel electrophoresis. Lane S is a representative of the squamous culture (the cells were submerged in medium for 7 d). Lanes 1 through 10 represent ALI cultures from Day 1 to Day 10. Human GAPDH gene was used as an internal control. (B) Relative MUC7 mRNA levels expressed in densitometry units. The figure shown is a representative of three separate experiments. Error bars indicate SDs of the triplicate determinations. (C and D) Cell morphology of the cultures at Day 7 viewed by light microscopy (original magnification: ×20). (C) The cells grown in ALI conditions revealed a basal layer overlaid by columnar cells with goblet morphology. (D) The cells grown under squamous conditions revealed a stratified (three arrows), nondifferentiated pattern.
<b>Figure 2.</b>
Figure 2.
Effect of exogenous factors on MUC7 mRNA and glycoprotein production. NHTBE cells were grown in ALI for 7 d and treated with IL-1β, IL-4, IL-13, TNF-α, EGF, and P. aeruginosa LPS for 12 h. MUC7 expression in these cultures was examined at the transcriptional level and protein level. (A) Quantitative analyses of MUC7 transcription. Total RNA was isolated from the control cells and treated cells and reverse transcribed. The levels of MUC7 mRNA were assessed by real-time PCR and normalized to those of the GAPDH gene. MUC7 mRNA levels are expressed as fold of induction by comparing with untreated control cultures. Each bar represents the mean ± SE from three independent experiments analyzed in four replicates. *P < 0.05. (B) Quantitative analyses of MUC7 mucin production. The relative stimulatory effects on MUC7 mucin production by the exogenous factors were determined by ELISA as described in Materials and Methods. Cell lysates were made from the control cells and the exogenous factor–treated cells. The MUC7 mucin levels were normalized to standard curve made with purified MUC7 mucin and expressed as percentage of the control. Each bar represents the mean ± SE from three independent experiments analyzed in four replicates. *P < 0.05.
<b>Figure 3.</b>
Figure 3.
Effect of P. aeruginosa LPS on MUC7 transcription in mouse tracheal and lung explants. (A) Three human MUC7 gene transgenic mice were killed, and their tracheas and lungs were removed for in vitro culture. The tissues were cultured in bronchial epithelial growth medium for 2 h with or without LPS stimulation (10 μg/ml). RNA samples were prepared and the MUC7 (human) and muc10 (mouse) mRNA quantity determined by semiquantitative RT-PCR. Mouse GAPDH gene served as an internal control for RNA integrity and the loading amount. (B) Relative levels of MUC7 mRNA and muc10 mRNA expressed in densitometry units. Results shown are representative of triplicate experiments, and the values represent the mean ± SE. *P < 0.05. Open bars and bars with horizontal lines, MUC7; cross-hatched bars and solid bars, muc10.
<b>Figure 4.</b>
Figure 4.
Effect of P. aeruginosa LPS on MUC7 transcription in mouse airways. (A) MUC7 mRNA was examined by Northern blot analyses. Six human MUC7 gene transgenic mice were divided into two groups (control and P. aeruginosa LPS–treated, as described in Materials and Methods). Three nontransgenic mice were also treated with LPS. Tissues were collected from the animals 2 d later. The RNA samples were prepared from the tissues, and MUC7 mRNA levels were assessed. Mouse GAPDH gene served as an internal control. The RNA samples in lanes 1 to 4 were from transgenic mice; the RNA samples in lanes 5 and 6 were from nontransgenic mice. T, trachea; L lung. (B) Relative levels of MUC7 mRNA determined by densitometry. MUC7 mRNA was normalized to GAPDH mRNA bands and expressed as fold of untreated control samples. Results are representative of three independent experiments (mean ± SE). *P < 0.05.
<b>Figure 5.</b>
Figure 5.
Immunolocalization of MUC7 protein expression in mouse tracheal and lung explants. Fluorescent and phase contrast pictures are shown for each sample. (A–D) Trachea and lung of nontransgenic BCF2 mouse. (E–H) Trachea and lung of nontransgenic BCF2 mouse incubated with LPS. (I–L) Trachea and lung of transgenic TgMUC7 mouse. (M–P) Trachea and lung of transgenic TgMUC7 mouse incubated with LPS. (Q–R) Hematoxylin-eosin staining of trachea of transgenic TgMUC7 mouse incubated with LPS. (S–T) Hematoxylin-eosin staining of lung of transgenic TgMUC7 mouse incubated with LPS. Original magnification in panels A–P, Q, and S: ×200. Original magnification in panels R and T: ×400.
See this image and copyright information in PMC

References

    1. Audie JP, Janin A, Porchet N, Copin MC, Gosselin B, Aubert JP. Expression of human mucin genes in respiratory, digestive, and reproductive tracts ascertained by in situ hybridization. J Histochem Cytochem 1993;41:1479–1485. - PubMed
    1. Rose MC, Gendler SJ. Airway mucin genes and gene products. In: Lethem DRAM, editor. Airway mucus: mechanism and clinical perspectives. Basel: Birkhauser Verlag; 1997. pp. 41–66.
    1. Schenkels LCPM, Gururaja TL, Levine JM. Salivary mucins: their role in oral mucosal barrier function and drug delivery. In: Rathbone MJ, editor. Oral mucosal drug delivery. Hamilton, New Zeland: Marcel Dekker, Inc.; 1996. pp. 191–220.
    1. Widdicombe JH. Accumulation of airway mucus in cystic fibrosis. Pulm Pharmacol 1994;7:225–233. - PubMed
    1. Ho SB, Niehans GA, Lyftogt C, Yan PS, Cherwitz DL, Gum ET, Dahiya R, Kim YS. Heterogeneity of mucin gene expression in normal and neoplastic tissues. Cancer Res 1993;53:641–651. - PubMed

Publication types

MeSH terms