Connective Tissue Growth Factor Expression and Action in Human Corneal Fibroblast Cultures and Rat Corneas after Photorefractive Keratectomy

Blalock, Timothy D.; Duncan, Matthew R.; Varela, Juan Carlos; Goldstein, Michael H.; Tuli, Sonal S.; Grotendorst, Gary R.; Schultz, Gregory S.

doi:10.1167/iovs.02-0860

Cited by 89 publications

(80 citation statements)

References 36 publications

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“…9, time 0) because the homogenates of normal whole eyes includes the lens and vitreous, both of which have much higher levels of CTGF protein than found in normal cornea tissue. Similar to the findings of Blalock et al, 8 who measured total CTGF protein levels using ELISA, we found very 25, and 21 kDa) was performed from three replicates. P < 0.05 is indicated by *, 0.001 < P < 0.0099 indicated by **, and P < 0.0001 indicated by ***.…”

Section: Figuresupporting

confidence: 91%

“…Using the technique that we previously described, 8 precise, reproducible, central corneal ablations were created in rat corneas using a Nidek EC-5000 Eximer laser (Nidek, Fremont, CA). Briefly, rats were anesthetized with isofluorane/oxygen inhalation, proparacaine eye drops were applied to achieve local anesthesia of corneas, and both eyes of each rat were ablated to a depth of 80 lm with creating a 4.4-mm diameter central epithelium to stroma injury.…”

Section: Rat Corneal Ablationmentioning

confidence: 99%

“…6,7 We previously reported the expression level of CTGF was elevated in rat corneas after ablation. 8 Fragments of CTGF were first reported in biological tissues by Brigstock et al 9 in pig uterine flushings. Since this discovery, 10-, 12-, 16-, 18-, 19-, 20-, 24-, and 31-kDa fragments of CTGF have been identified in different cell types, tissues, and body fluids.…”

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confidence: 98%

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Proteolytic Processing of Connective Tissue Growth Factor in Normal Ocular Tissues and during Corneal Wound Healing

Robinson¹,

Smith²,

Patel³

et al. 2012

Invest. Ophthalmol. Vis. Sci.

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PURPOSE. Connective tissue growth factor (CTGF) is a fibrogenic cytokine that is up-regulated by TGF-b and mediates most key fibrotic actions of TGF-b, including stimulation of synthesis of extracellular matrix and differentiation of fibroblasts into myofibroblasts. This study addresses the role of proteolytic processing of CTGF in human corneal fibroblasts (HCF) stimulated with TGF-b, normal ocular tissues and wounded corneas.METHODS. Proteolytic processing of CTGF in HCF cultures, normal animal eyes, and excimer laser wounded rat corneas were examined by Western blot. The identity of a 21-kDa band was determined by tandem mass spectrometry, and possible alternative splice variants of CTGF were assessed by 5 0 Rapid Amplification of cDNA Ends (RACE).RESULTS. HCF stimulated by TGF-b contained full length 38-kDa CTGF and fragments of 25, 21, 18, and 13 kDa, while conditioned medium contained full length 38-and a 21-kDa fragment of CTGF that contained the middle ''hinge'' region of CTGF. Fragmentation of recombinant CTGF incubated in HCF extracts was blocked by the aspartate protease inhibitor, pepstatin. Normal mouse, rat, and rabbit whole eyes and rabbit ocular tissues contained abundant amounts of C-terminal 25-and 21-kDa fragments and trace amounts of 38-kDa CTGF, although no alternative transcripts were detected. All forms of CTGF (38, 25, and 21 kDa) were detected during healing of excimer ablated rat corneas, peaking on day 11.CONCLUSIONS. Proteolytic processing of 38-kDa CTGF occurs during corneal wound healing, which may have important implications in regulation of corneal scar formation. (Invest Ophthalmol Vis Sci. 2012;53:8093-8103)

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Section: Figuresupporting

confidence: 91%

Section: Rat Corneal Ablationmentioning

confidence: 99%

mentioning

confidence: 98%

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Proteolytic Processing of Connective Tissue Growth Factor in Normal Ocular Tissues and during Corneal Wound Healing

Robinson¹,

Smith²,

Patel³

et al. 2012

Invest. Ophthalmol. Vis. Sci.

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“…The use of CCN2 antisense oligonucleotides has helped to establish the importance of CCN2 for TGF-b-induced collagen production in a variety of cell types including kidney mesangial cells, NRK cells, corneal fibroblasts, and conjunctival fibroblasts [28][29][30][31][32][33] . In in vivo animal models, CCN2 antisense oligonucleotides have proven effective in reducing matrix expansion or fibrosis in the skin or kidney [34][35][36] , and scarring around breast implants [37] .…”

Section: Discussionmentioning

confidence: 99%

Connective tissue growth factor hammerhead ribozyme attenuates human hepatic stellate cell function

Gao¹,

Brigstock²

2009

WJG

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AIM:To determine the effect of hammerhead ribozyme targeting connective tissue growth factor (CCN2) on human hepatic stellate cell (HSC) function. METHODS:CCN2 hammerhead ribozyme cDNA plus two self-cleaving sequences were inserted into pTriEx2 to produce pTriCCN2-Rz. Each vector was individually transfected into cultured LX-2 human HSCs, which were then stimulated by addition of transforming growth factor (TGF)-b1 to the culture medium. Semiquantitative RT-PCR was used to determine mRNA levels for CCN2 or collagen Ⅰ, while protein levels of each molecule in cell lysates and conditioned medium were measured by ELISA. Cell-cycle progression of the transfected cells was assessed by flow cytometry. RESULTS:In pTriEx2-transfected LX-2 cells, TGF-β1 treatment caused an increase in the mRNA level for CCN2 or collagen Ⅰ, and an increase in produced and secreted CCN2 or extracellular collagen Ⅰ protein levels. pTriCCN2-Rz-transfected LX-2 cells showed decreased basal CCN2 or collagen mRNA levels, as well as produced and secreted CCN2 or collagen Ⅰ protein. Furthermore, the TGF-b1-induced increase in mRNA or protein for CCN2 or collagen Ⅰ was inhibited partially in pTriCCN2-Rz-transfected LX-2 cells. Inhibition of CCN2 using hammerhead ribozyme cDNA resulted in fewer of the cells transitioning into S phase.CONCLUSION: Endogenous CCN2 is a mediator of basal or TGF-b1-induced collagen Ⅰ production in human HSCs and regulates entry of the cells into S phase.

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“…In the cornea, CTGF promotes transdifferentiation of the corneal fibroblast into the myofibroblast, resulting in excess production of collagen and fibronectin (FN). CTGF is also involved in the wound healing process of the corneal stroma (Folger et al 2001;Blalock et al 2003;Garrett et al 2004). One of the reasons why CTGF has so many functions may be due to its binding to other cytokines and the extracellular matrix to modulate their actions (Frazier et al 1996).…”

mentioning

confidence: 99%

Connective Tissue Growth Factor Cooperates with Fibronectin in Enhancing Attachment and Migration of Corneal Epithelial Cells

Sugioka

Yoshida

Kodama

et al. 2010

Tohoku J. Exp. Med.

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Corneal wound healing is a complex process involving the integrated actions of various growth factors, cytokines and extracellular matrix produced by corneal cells and inflammatory cells. Connective tissue growth factor (CTGF) has been linked to wound healing, and fibronectin (FN) is a major component of the extracellular matrix. However, the functions of CTGF and FN in corneal epithelial cells are not well understood. We therefore investigated the coordinated function of CTGF and FN in the attachment and migration of corneal epithelial cells. Treatment of human corneal epithelial cells (HCECs) with transforming growth factor (TGF) β 1 up-regulated the expression of CTGF, but did not noticeably affect FN expression, as judged by immunoblot analysis of cell lysates. In contrast, the amount of FN accumulated in the cultured media was increased in a time-dependent manner, but CTGF was undetectable in the cultured media. The expression level of FN was decreased by the knockdown of CTGF expression with a specific short hairpin RNA, indicating that CTGF acts as an upstream mediator of FN expression. CTGF augmented the FN-mediated increase in the attachment of HCEC by about twofold, although CTGF alone did not influence the attachment. Moreover, the migration assay with rabbit corneal blocks revealed that CTGF (390 nM) alone or in combination of FN (10 µ g/mL) promoted corneal epithelial migration; the mean migration distances of control, CTGF, and CTGF + FN were 272, 325, and 626, µ m, respectively. In conclusion, CTGF cooperates with FN in enhancing the attachment and migration of corneal epithelial cells.

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Connective Tissue Growth Factor Expression and Action in Human Corneal Fibroblast Cultures and Rat Corneas after Photorefractive Keratectomy

Cited by 89 publications

References 36 publications

Proteolytic Processing of Connective Tissue Growth Factor in Normal Ocular Tissues and during Corneal Wound Healing

Proteolytic Processing of Connective Tissue Growth Factor in Normal Ocular Tissues and during Corneal Wound Healing

Connective tissue growth factor hammerhead ribozyme attenuates human hepatic stellate cell function

Connective Tissue Growth Factor Cooperates with Fibronectin in Enhancing Attachment and Migration of Corneal Epithelial Cells

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