Modulation of human bronchial epithelial cell IIICS fibronectin mRNA in vitro

E. Rosi, J. D. Beckmann, P. Pladsen, S. I. Rennard, Debra Romberger

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

Fibronectin (Fn) is an extracellular matrix glycoprotein which is involved in wound repair, including repair of injured airway epithelium. Bronchial epithelial cells (BECs) are known to produce Fn which has enhanced chemotactic activity compared to serum Fn. Alternative splicing of the Fn gene is an important mechanism by which cells regulate the production of Fn. Human BECs produce Fn which contains the EIIIA region, but the expression of IIICS region variants has not previously been reported. Our purpose was to better define the molecular characteristics of human BEC Fn by determining the expression of alternative splice variants of the IIICS region of Fn of human BECs in vitro. Human bronchial epithelial cells obtained from bronchoscopy were cultured. To examine the presence of IIICS messenger ribonucleic acid (mRNA) variants, we synthesized oligonucleotide primers complementary to the published human fibronectin complementary deoxyribonucleic acid (cDNA) sequence of the IIICS domain for use in polymerase chain reactions (PCR) with total ribonucleic acid (RNA) extracted from cultured human BECs. To examine the modulation of IIICS mRNA expression, the 428 base pair (bp) DNA fragment generated in the PCR was oligolabelled with 32[P]-deoxycytidine triphosphate (dCTP) for use as a probe for Northern blot analysis. Human BECs were cultured in the presence and absence of transforming growth factor-β (TGF-β) and agents which influence cyclic adenosine monophosphate (cAMP) including isoproterenol and dibutryl cAMP (db-cAMP). Total RNA from cultures was extracted, electrophoresis performed, and Northern blots obtained. Blots were hybridized with IIICS probe, total Fn cDNA, and tubulin cDNA. It was found that human BECs in culture expressed the five known human IIICS variants. TGF-β enhanced the expression of IIICS mRNA in a concentration- and time-dependent fashion. Isoproterenol and db-cAMP both reduced the expression of IIICS mRNA and attenuated the TGF-β induction. Changes in IIICS mRNA paralleled changes in total Fn mRNA, suggesting that these agents do not selectively modulate only the IIICS domain of Fn. We conclude that human airway epithelial cell Fn in vitro does contain mRNA for five IIICS variants, and that IIICS mRNA can be modulated by TGF-β and agents which influence cAMP. It is unknown whether alterations in IIICS variants contribute to the functional difference previously observed between airway epithelial cell Fn and plasma-derived Fn.

Original languageEnglish (US)
Pages (from-to)549-555
Number of pages7
JournalEuropean Respiratory Journal
Volume9
Issue number3
DOIs
StatePublished - Mar 1 1996

Fingerprint

Fibronectins
Epithelial Cells
RNA
Transforming Growth Factors
Cyclic AMP
DNA
In Vitro Techniques
Isoproterenol
Northern Blotting
Polymerase Chain Reaction
DNA Primers
Alternative Splicing
Bronchoscopy
Tubulin
Base Pairing
Extracellular Matrix
Electrophoresis
Glycoproteins
Epithelium
Cell Culture Techniques

Keywords

  • airway epithelium
  • fibronectin gene regulation
  • messenger ribonucleic acid splicing

ASJC Scopus subject areas

  • Pulmonary and Respiratory Medicine

Cite this

Modulation of human bronchial epithelial cell IIICS fibronectin mRNA in vitro. / Rosi, E.; Beckmann, J. D.; Pladsen, P.; Rennard, S. I.; Romberger, Debra.

In: European Respiratory Journal, Vol. 9, No. 3, 01.03.1996, p. 549-555.

Research output: Contribution to journalArticle

Rosi, E. ; Beckmann, J. D. ; Pladsen, P. ; Rennard, S. I. ; Romberger, Debra. / Modulation of human bronchial epithelial cell IIICS fibronectin mRNA in vitro. In: European Respiratory Journal. 1996 ; Vol. 9, No. 3. pp. 549-555.
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