Enhanced upregulation of smooth muscle-related transcripts by TGFβ₂in asthmatic (myo)fibroblasts
Wicks, J., Haitchi, H.M., Holgate, S.T., Davies, D.E. and Powell, R.M. (2006) Enhanced upregulation of smooth muscle-related transcripts by TGFβ₂in asthmatic (myo)fibroblasts. Thorax, 61, (4), 313-319. (doi:10.1136/thx.2005.050005).
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Description/Abstract
Background: Transforming growth factor beta (TGFß) upregulates a number of smooth muscle specific genes in (myo)fibroblasts. As asthma is characterised by an increase in airway smooth muscle, we postulated that TGFß₂ favours differentiation of asthmatic (myo)fibroblasts towards a smooth muscle phenotype.
Methods: Primary fibroblasts were grown from bronchial biopsy specimens from normal (n = 6) and asthmatic (n = 7) donors and treated with TGFß₂ to induce myofibroblast differentiation. The most stable genes for normalisation were identified using RT-qPCR and the geNorm software applied to a panel of 12 "housekeeping" genes. Expression of α-smooth muscle actin (αSMA), heavy chain myosin (HCM), calponin 1 (CPN 1), desmin, and γ-actin were measured by RT-qPCR. Protein expression was assessed by immunocytochemistry and western blotting.
Results: Phospholipase A2 and ubiquitin C were identified as the most stably expressed and practically useful genes for normalisation of gene expression during myofibroblast differentiation. TGFß₂ induced mRNA expression for all five smooth muscle related transcripts; αSMA, HCM and CPN 1 protein were also increased but desmin protein was not detectable. Although there was no difference in basal expression, HCM, CPN 1 and desmin were induced to a significantly greater extent in asthmatic fibroblasts than in those from normal controls (p = 0.041 and 0.011, respectively).
Conclusions: Although TGFß₂ induced the transcription of several smooth muscle related genes, not all were translated into protein. Thus, while TGFß₂ is unable to induce a bona fide smooth muscle cell phenotype, it may "prime" (myo)fibroblasts for further differentiation, especially if the cells are derived from asthmatic airways.
Abbreviations: CPN 1, calponin 1; DMEM, Dulbecco’s modified Eagle’s medium; FBS, fetal bovine serum; HCM, heavy chain myosin; RT-qPCR, reverse transcription quantitative polymerase chain reaction; SFM, serum free medium; SMA, smooth muscle actin; TGFß, transforming growth factor ß
| Item Type: | Article |
|---|---|
| Additional Information: | Asthma |
| ISSNs: | 0040-6376 (print) |
| Related URLs: | |
| Keywords: | asthma, smooth muscle markers, transforming growth factor ß |
| Subjects: | R Medicine Q Science > QH Natural history > QH426 Genetics |
| Divisions: | University Structure - Pre August 2011 > School of Medicine > Infection, Inflammation and Repair |
| Item ID: | 27485 |
| Date Deposited: | 27 Apr 2006 |
| Last Modified: | 02 Mar 2012 11:46 |
| Contributors: | Wicks, J. (Author) Haitchi, H.M. (Author) Holgate, S.T. (Author) Davies, D.E. (Author) Powell, R.M. (Author) |
| Date: | 2006 |
| Additional Information: | Asthma |
| Status: | Published |
| Contact Email Address: | rmp2@soton.ac.uk |
| URI: | http://eprints.soton.ac.uk/id/eprint/27485 |
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