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Human Keratoconus Cell Contractility is Mediated by Transforming Growth Factor-Beta Isoforms.

Lyon D', McKay TB, Sarkar-Nag A, Priyadarsini S, Karamichos D - J Funct Biomater (2015)

Bottom Line: HKCs showed delayed contractility with decreased Collagen I:Collagen V ratios.We also found that HKCs have significantly decreased Collagen I:Collagen III ratios suggesting a potential link to altered collagen isoform expression in KC.Our findings show that HKCs have significant variations in collagen secretion in a 3D collagen gel and have delayed contraction of the matrix compared to HCFs.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology/Dean McGee Eye Institute, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA. desiree-lyon@ouhsc.edu.

ABSTRACT
Keratoconus (KC) is a progressive disease linked to defects in the structural components of the corneal stroma. The extracellular matrix (ECM) is secreted and assembled by corneal keratocytes and regulated by transforming growth factor-β (TGF-β). We have previously identified alterations in the TGF-β pathway in human keratoconus cells (HKCs) compared to normal corneal fibroblasts (HCFs). In our current study, we seeded HKCs and HCFs in 3D-collagen gels to identify variations in contractility, and expression of matrix metalloproteases (MMPs) by HKCs in response the TGF-β isoforms. HKCs showed delayed contractility with decreased Collagen I:Collagen V ratios. TGF-β1 significantly increased ECM contraction, Collagen I, and Collagen V expression by HKCs. We also found that HKCs have significantly decreased Collagen I:Collagen III ratios suggesting a potential link to altered collagen isoform expression in KC. Our findings show that HKCs have significant variations in collagen secretion in a 3D collagen gel and have delayed contraction of the matrix compared to HCFs. For the first time, we utilize a collagen gel model to characterize the contractility and MMP expression by HKCs that may contribute to the pathobiology of KC.

No MeSH data available.


Related in: MedlinePlus

(A) Collagen I (Col I), (B) Collagen III (Col III), and (C) Collagen V (Col V) expression and (D–E) ratios of Col I/Col V and Col I/Col III by HCFs and HKCs at week 4 measured by RT-PCR. n = 3, error bars represent standard error of the mean. (**** denotes p < 0.0001, *** denotes p < 0.001, ** denotes p < 0.01 and * denotes p < 0.05.)
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jfb-06-00422-f004: (A) Collagen I (Col I), (B) Collagen III (Col III), and (C) Collagen V (Col V) expression and (D–E) ratios of Col I/Col V and Col I/Col III by HCFs and HKCs at week 4 measured by RT-PCR. n = 3, error bars represent standard error of the mean. (**** denotes p < 0.0001, *** denotes p < 0.001, ** denotes p < 0.01 and * denotes p < 0.05.)

Mentions: In order to determine if expression of pro-collagens correlated with collagen secretion detected in the conditioned media, we quantified the expression of Col I, Col III, and Col V, using RT-PCR in HCF and HKC at day 26 following complete contraction of the matrix. We found an increase in all three collagen types by control HKCs compared to HCFs (Figure 4A–C). We also identified a significant increase of 638% and 994% in expression of Col I and Col III, respectively, by HKCs following TGF-β1 stimulation (Figure 4A,B, p < 0.05). In contrast, HCFs did not significantly increase Col I, Col III, or Col V expression following stimulation with TGF-β isoforms (Figure 4A–C). Our results show that HKCs are more responsive to TGF-β isoform treatment compared to HCFs at day 26. This data suggests that fully contracted HCFs have reduced expression of ECM components compared to HKCs.


Human Keratoconus Cell Contractility is Mediated by Transforming Growth Factor-Beta Isoforms.

Lyon D', McKay TB, Sarkar-Nag A, Priyadarsini S, Karamichos D - J Funct Biomater (2015)

(A) Collagen I (Col I), (B) Collagen III (Col III), and (C) Collagen V (Col V) expression and (D–E) ratios of Col I/Col V and Col I/Col III by HCFs and HKCs at week 4 measured by RT-PCR. n = 3, error bars represent standard error of the mean. (**** denotes p < 0.0001, *** denotes p < 0.001, ** denotes p < 0.01 and * denotes p < 0.05.)
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4493522&req=5

jfb-06-00422-f004: (A) Collagen I (Col I), (B) Collagen III (Col III), and (C) Collagen V (Col V) expression and (D–E) ratios of Col I/Col V and Col I/Col III by HCFs and HKCs at week 4 measured by RT-PCR. n = 3, error bars represent standard error of the mean. (**** denotes p < 0.0001, *** denotes p < 0.001, ** denotes p < 0.01 and * denotes p < 0.05.)
Mentions: In order to determine if expression of pro-collagens correlated with collagen secretion detected in the conditioned media, we quantified the expression of Col I, Col III, and Col V, using RT-PCR in HCF and HKC at day 26 following complete contraction of the matrix. We found an increase in all three collagen types by control HKCs compared to HCFs (Figure 4A–C). We also identified a significant increase of 638% and 994% in expression of Col I and Col III, respectively, by HKCs following TGF-β1 stimulation (Figure 4A,B, p < 0.05). In contrast, HCFs did not significantly increase Col I, Col III, or Col V expression following stimulation with TGF-β isoforms (Figure 4A–C). Our results show that HKCs are more responsive to TGF-β isoform treatment compared to HCFs at day 26. This data suggests that fully contracted HCFs have reduced expression of ECM components compared to HKCs.

Bottom Line: HKCs showed delayed contractility with decreased Collagen I:Collagen V ratios.We also found that HKCs have significantly decreased Collagen I:Collagen III ratios suggesting a potential link to altered collagen isoform expression in KC.Our findings show that HKCs have significant variations in collagen secretion in a 3D collagen gel and have delayed contraction of the matrix compared to HCFs.

View Article: PubMed Central - PubMed

Affiliation: Department of Ophthalmology/Dean McGee Eye Institute, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA. desiree-lyon@ouhsc.edu.

ABSTRACT
Keratoconus (KC) is a progressive disease linked to defects in the structural components of the corneal stroma. The extracellular matrix (ECM) is secreted and assembled by corneal keratocytes and regulated by transforming growth factor-β (TGF-β). We have previously identified alterations in the TGF-β pathway in human keratoconus cells (HKCs) compared to normal corneal fibroblasts (HCFs). In our current study, we seeded HKCs and HCFs in 3D-collagen gels to identify variations in contractility, and expression of matrix metalloproteases (MMPs) by HKCs in response the TGF-β isoforms. HKCs showed delayed contractility with decreased Collagen I:Collagen V ratios. TGF-β1 significantly increased ECM contraction, Collagen I, and Collagen V expression by HKCs. We also found that HKCs have significantly decreased Collagen I:Collagen III ratios suggesting a potential link to altered collagen isoform expression in KC. Our findings show that HKCs have significant variations in collagen secretion in a 3D collagen gel and have delayed contraction of the matrix compared to HCFs. For the first time, we utilize a collagen gel model to characterize the contractility and MMP expression by HKCs that may contribute to the pathobiology of KC.

No MeSH data available.


Related in: MedlinePlus