Kak-Ming Ling1, Erika N Sutanto1,2, Thomas Iosifidis3,4, Elizabeth Kicic-Starcevich1,2, Kevin Looi3, Luke W Garratt1,3, Kelly M Martinovich1, Francis J Lannigan5, Darryl A Knight6,7, Stephen M Stick1,2,3,4, Anthony Kicic8,9,10,11. 1. Telethon Kids Institute, Centre for Child Health Research, The University of Western Australia, Nedlands, Western Australia, Australia. 2. Department of Respiratory Medicine, Princess Margaret Hospital for Children, Perth, Western Australia, Australia. 3. School of Paediatrics and Child Health, The University of Western Australia, Nedlands, Western Australia, Australia. 4. Centre for Cell Therapy and Regenerative Medicine, The University of Western Australia and Harry Perkins Institute, Nedlands, Western Australia, Australia. 5. School of Medicine, Notre Dame University, Fremantle, Western Australia, Australia. 6. School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, New South Wales, Australia. 7. Department of Anesthesiology, Pharmacology and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada. 8. Telethon Kids Institute, Centre for Child Health Research, The University of Western Australia, Nedlands, Western Australia, Australia. Anthony.Kicic@telethonkids.org.au. 9. Department of Respiratory Medicine, Princess Margaret Hospital for Children, Perth, Western Australia, Australia. Anthony.Kicic@telethonkids.org.au. 10. School of Paediatrics and Child Health, The University of Western Australia, Nedlands, Western Australia, Australia. Anthony.Kicic@telethonkids.org.au. 11. Centre for Cell Therapy and Regenerative Medicine, The University of Western Australia and Harry Perkins Institute, Nedlands, Western Australia, Australia. Anthony.Kicic@telethonkids.org.au.
Abstract
BACKGROUND AND OBJECTIVE: Evidence into the role of TGF-β1 in airway epithelial repair in asthma is still controversial. This study tested the hypothesis that the reduced TGF-β1 levels previously observed in paediatric asthmatic airway epithelial cells directly contribute to the dysregulated repair seen in these cells. METHODS: Primary airway epithelial cells (pAEC) from children with asthma (n = 16) and non-asthmatic subjects (n = 20) were isolated, and subcultured for investigation of TGF-β1 gene and protein via quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Expression of other associated genes such as integrins αvβ6, αvβ8 and MT1-MMP were also tested. Small interfering RNA (siRNA) was employed to assess the role of TGF-β1 during wound repair. RESULTS: TGF-β1 gene and protein expression were significantly downregulated in asthmatic pAEC over the course of repair, compared with cells from non-asthmatic children. Messenger RNA (mRNA) expression of TGF-β1 was also directly implicated in non-asthmatic and asthmatic pAEC proliferation over their quiescent counterparts. Small interfering RNA-mediated knockdown of TGF-β1 compromised repair in non-asthmatic pAEC and exacerbated the dysregulated repair seen in asthmatic pAEC. Expression of major TGF-β1 activators of epithelial cells, integrin αvβ6 and αvβ8 was also measured and there was no difference in αvβ6 gene expression between the two cohorts. Although integrin αvβ8 gene expression was significantly higher in asthmatic pAEC, the expression of MT1-MMP (MMP14) which facilitates the αvβ8 mediated TGF-β1 activation was significantly downregulated. CONCLUSION: Our data has highlighted the importance of TGF-β1 in pAEC wound repair in vitro. The significantly lower levels seen in asthmatic pAEC subsequently contributes to the dysregulated repair observed in these cells.
BACKGROUND AND OBJECTIVE: Evidence into the role of TGF-β1 in airway epithelial repair in asthma is still controversial. This study tested the hypothesis that the reduced TGF-β1 levels previously observed in paediatric asthmatic airway epithelial cells directly contribute to the dysregulated repair seen in these cells. METHODS: Primary airway epithelial cells (pAEC) from children with asthma (n = 16) and non-asthmatic subjects (n = 20) were isolated, and subcultured for investigation of TGF-β1 gene and protein via quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Expression of other associated genes such as integrins αvβ6, αvβ8 and MT1-MMP were also tested. Small interfering RNA (siRNA) was employed to assess the role of TGF-β1 during wound repair. RESULTS: TGF-β1 gene and protein expression were significantly downregulated in asthmatic pAEC over the course of repair, compared with cells from non-asthmatic children. Messenger RNA (mRNA) expression of TGF-β1 was also directly implicated in non-asthmatic and asthmatic pAEC proliferation over their quiescent counterparts. Small interfering RNA-mediated knockdown of TGF-β1 compromised repair in non-asthmatic pAEC and exacerbated the dysregulated repair seen in asthmatic pAEC. Expression of major TGF-β1 activators of epithelial cells, integrin αvβ6 and αvβ8 was also measured and there was no difference in αvβ6 gene expression between the two cohorts. Although integrin αvβ8 gene expression was significantly higher in asthmatic pAEC, the expression of MT1-MMP (MMP14) which facilitates the αvβ8 mediated TGF-β1 activation was significantly downregulated. CONCLUSION: Our data has highlighted the importance of TGF-β1 in pAEC wound repair in vitro. The significantly lower levels seen in asthmatic pAEC subsequently contributes to the dysregulated repair observed in these cells.
Authors: Thomas Iosifidis; Erika N Sutanto; Alysia G Buckley; Laura Coleman; Erin E Gill; Amy H Lee; Kak-Ming Ling; Jessica Hillas; Kevin Looi; Luke W Garratt; Kelly M Martinovich; Nicole C Shaw; Samuel T Montgomery; Elizabeth Kicic-Starcevich; Yuliya V Karpievitch; Peter Le Souëf; Ingrid A Laing; Shyan Vijayasekaran; Francis J Lannigan; Paul J Rigby; Robert Ew Hancock; Darryl A Knight; Stephen M Stick; Anthony Kicic Journal: JCI Insight Date: 2020-04-09
Authors: Marta Michalik; Katarzyna Wójcik-Pszczoła; Milena Paw; Dawid Wnuk; Paulina Koczurkiewicz; Marek Sanak; Elżbieta Pękala; Zbigniew Madeja Journal: Cell Mol Life Sci Date: 2018-08-12 Impact factor: 9.261
Authors: Thomas Iosifidis; Erika N Sutanto; Samuel T Montgomery; Patricia Agudelo-Romero; Kevin Looi; Kak-Ming Ling; Nicole C Shaw; Luke W Garratt; Jessica Hillas; Kelly M Martinovich; Elizabeth Kicic-Starcevich; Shyan Vijayasekaran; Francis J Lannigan; Paul J Rigby; Darryl A Knight; Stephen M Stick; Anthony Kicic Journal: J Pers Med Date: 2021-12-07