Winston W.‐Y. Kao

PURPOSE: Recent studies indicate that transforming growth factor (TGF)beta is a potent inducer of corneal myofibroblast differentiation and expression of smooth muscle-specific, alpha-actin (alpha-SMA). Although TGFbeta is known to enhance synthesis of extracellular matrix proteins and receptors, little is known about how it modulates the expression of smooth muscle proteins in nonmuscle cells. The purpose of this study was to identify the role of Arg-Gly-Asp (RGD)-dependent tyrosine phosphorylation in regulating alpha-SMA gene expression and ultimately myofibroblast development. METHODS: Because cell culture in serum-containing media mimics myofibroblast transformation, all experiments were performed on freshly isolated rabbit keratocytes plated in defined, serum-free media. Cells were exposed to TGFbeta (1 ng/ml), Gly-Arg-Gly-Asp-D-Ser-Pro (GRGDdSP, 50 microM), Gly-Arg-AL-Asp-Ser-Pro (GRADSP; 100 microM), or herbimycin A (0.1-10 nM) at 24 hours (sparse) or 7 days (confluent). Cells were evaluated by immunocytochemistry and proteins and RNA collected for western and northern blot analyses using antibodies specific for alpha-SMA, fibronectin, focal adhesion proteins, and phosphotyrosine (clones 4G10 and PY20); and probes directed against rabbit alpha-SMA. All experiments were repeated at least three times. RESULTS: Keratocytes exposed to TGFbeta showed expression of alpha-SMA that coincided with the intracellular reorganization of the actin cytoskeleton and the extracellular assembly of fibronectin fibrils. Addition of RGD containing but not control peptides blocked the organization of intracellular actin, extracellular fibronectin, and alpha-SMA protein and mRNA. Immunoprecipitation of cell proteins with 4G10 or PY20 identified the TGFbeta-associated tyrosine phosphorylation of paxillin, pp125fak, p130, PLCgamma, and tensin, which was blocked by addition of GRGDdSP. Addition of herbimycin A to keratocytes exposed to TGFbeta showed a dose-dependent loss of alpha-SMA protein and mRNA which correlated with loss of tyrosine phosphorylation, absence of actin reorganization, and fibronectin assembly. CONCLUSIONS: The data suggest that TGFbeta-mediated alpha-SMA gene expression leading to myofibroblast transformation may involve an RGD-dependent phosphotyrosine signal transduction pathway.

Collagen V is a regulatory fibril-forming collagen that forms heterotypic fibrils with collagen I. Deletion of collagen V in the mouse is associated with a lack of fibril assembly in the embryonic mesenchyme, with a resultant lethal phenotype. The current work elucidates the regulatory roles of collagen V during development and growth of tissues. A conditional mouse model with a mutation in Col5a1 was developed using a Cre-loxP approach. Col5a1 was ablated in Col5a1(flox/flox) mice using a cornea stroma-specific Kera-Cre driver mouse to produce a bitransgenic Col5a1(Δst/Δst) line that is null for collagen V. This permits analyses of the corneal stroma, a widely used model for studies of collagen V. The collagen-V-knockout stroma demonstrated severe dysfunctional regulation of fibrillogenesis. Fibril diameters were significantly increased, with an abnormal, heterogeneous distribution; fibril structure was abnormal, fibril number was decreased and lamellae were disorganized with decreased stroma thickness. The phenotype was more severe in the anterior versus posterior stroma. Opacity was demonstrated throughout the Col5a1(Δst/Δst) stroma, with significantly increased haze intensity compared with control mice. These data indicate central regulatory roles for collagen V in fibril and matrix assembly during tissue development, with dysfunctional regulation resulting in a functional loss of transparency.

PURPOSE: The purpose of the present study was to examine the roles of signaling pathways potentially activated by TGFbeta (i.e., Smad and p38 mitogen-activated kinase [MAPK]) in regulation of cell migration and proliferation of healing mouse corneal epithelium. METHODS: Activation of Smads or p38MAPK was evaluated by immunohistochemistry in healing mouse corneal epithelium after debridement. The role of endogenous TGFbeta or p38MAPK in epithelial healing was determined in organ-cultured mouse corneas with an epithelial defect, in the presence or absence of a TGFbeta-neutralizing antibody or p38MAPK inhibitors, respectively. Cell proliferation was evaluated by incorporation of bromodeoxyuridine. RESULTS: Migrating mouse corneal epithelium had minimal cell proliferation. Smad3 and -4 were found in nuclei of normal corneal epithelium, whereas they were absent in nuclei of migrating cells in association with Smad7 upregulation on epithelial debridement. Administration of TGFbeta-neutralizing antibody reduced the protein expression of Smad7 in vivo after a corneal injury. In contrast, phosphorylation and nuclear translocation of p38MAPK were markedly evident in migrating epithelium during healing, but not in uninjured epithelium. In organ culture, addition of p38MAPK inhibitors blocked cell migration more markedly than neutralizing TGFbeta-antibody and enhanced cell proliferation in the injured corneal epithelium, in association with phosphorylation of Erk. CONCLUSIONS: Endogenous TGFbeta enhances migration of corneal epithelium during wound healing in mice. The p38MAPK, but not the Smad, cascade plays a major role in promoting cell migration and in suppressing cell proliferation in migrating epithelium.

Lumican is an extracellular matrix glycoprotein widely distributed inmammalian connective tissues. Corneal lumican modified with keratan sulfateconstitutes one of the major proteoglycans of the stroma. Lumican-null miceexhibit altered collagen fibril organization and loss of corneal transparency.A closely related protein, keratocan, carries the remaining keratan sulfate ofthe cornea, but keratocan-null mice exhibit a less severe corneal phenotype.In the current study, we examined the effect of lumican overexpression incorneas of wild type mice. These mice showed no alteration in collagenorganization or transparency but had increased keratocan expression at bothprotein and mRNA levels. Corneas of lumican-null mice showed decreasedkeratocan. This coupling of keratocan expression with lumican also wasobserved after intrastromal injection of a lumican expression minigene intothe corneal stroma of Lum–/– mice. Smallinterfering RNA knockdown of lumican in vitro reduced keratocanexpression, whereas co-injection of a lumican-expressing minigene with aβ-galactosidase reporter driven by the keratocan promoter demonstrated anincrease of keratocan transcriptional activity in response to lumicanexpression in Lum–/– corneas invivo. These observations demonstrate that lumican has a novel regulatoryrole in keratocan expression at the transcriptional level. Such results helpprovide an explanation for the differences in severity of cornealmanifestation found in Lum–/– andKera–/– mice. The results also suggest acritical level of small proteoglycans to be essential for collagenorganization but that overabundance is not detrimental to extracellular matrixmorphogenesis. Lumican is an extracellular matrix glycoprotein widely distributed inmammalian connective tissues. Corneal lumican modified with keratan sulfateconstitutes one of the major proteoglycans of the stroma. Lumican-null miceexhibit altered collagen fibril organization and loss of corneal transparency.A closely related protein, keratocan, carries the remaining keratan sulfate ofthe cornea, but keratocan-null mice exhibit a less severe corneal phenotype.In the current study, we examined the effect of lumican overexpression incorneas of wild type mice. These mice showed no alteration in collagenorganization or transparency but had increased keratocan expression at bothprotein and mRNA levels. Corneas of lumican-null mice showed decreasedkeratocan. This coupling of keratocan expression with lumican also wasobserved after intrastromal injection of a lumican expression minigene intothe corneal stroma of Lum–/– mice. Smallinterfering RNA knockdown of lumican in vitro reduced keratocanexpression, whereas co-injection of a lumican-expressing minigene with aβ-galactosidase reporter driven by the keratocan promoter demonstrated anincrease of keratocan transcriptional activity in response to lumicanexpression in Lum–/– corneas invivo. These observations demonstrate that lumican has a novel regulatoryrole in keratocan expression at the transcriptional level. Such results helpprovide an explanation for the differences in severity of cornealmanifestation found in Lum–/– andKera–/– mice. The results also suggest acritical level of small proteoglycans to be essential for collagenorganization but that overabundance is not detrimental to extracellular matrixmorphogenesis. Lumican is a member of the small leucine-rich proteoglycan(SLRP) 1The abbreviations used are: SLRP, small leucine-rich proteoglycan; ECM,extracellular matrix; KSPG, keratan sulfate-containing proteoglycan; siRNA,small interfering RNA; RT, reverse transcription; CMTF, confocal microscopythrough focusing.1The abbreviations used are: SLRP, small leucine-rich proteoglycan; ECM,extracellular matrix; KSPG, keratan sulfate-containing proteoglycan; siRNA,small interfering RNA; RT, reverse transcription; CMTF, confocal microscopythrough focusing. family withexpression reported in cornea, sclera, aorta, cartilage, liver, skeletalmuscle, kidney, pancreas, brain, placenta, and lung(1Chakravarti S. Magnuson T. Lass J.H. Jepsen K.J. LaMantia C. Carroll H. J. CellBiol. 1998; 141: 1277-1286Crossref PubMed Scopus (570) Google Scholar, 2Funderburgh J.L. Funderburgh M.L. Mann M.M. Conrad G.W. J. Biol. Chem. 1991; 266: 24773-24777Abstract Full Text PDF PubMed Google Scholar, 3Funderburgh J.L. Funderburgh M.L. Brown S.J. Vergnes J.P. Hassell J.R. Mann M.M. Conrad G.W. J. Biol. Chem. 1993; 268: 11874-11880Abstract Full Text PDF PubMed Google Scholar, 4Grover J. Chen X.N. Korenberg J.R. Roughley P.J. J. Biol. Chem. 1995; 270: 21942-21949Abstract Full Text Full Text PDF PubMed Scopus (161) Google Scholar, 5Krull N.B. Gressner A.M. FEBS Lett. 1992; 312: 47-52Crossref PubMed Scopus (32) Google Scholar, 6Ezura Y. Chakravarti S. Oldberg A. Chervoneva I. Birk D.E. J. Cell Biol. 2000; 151: 779-788Crossref PubMed Scopus (277) Google Scholar).Lumican co-localizes with collagen fibrils in the corneal stroma and has beenhypothesized to be involved in modulation of the highly organized collagenmatrix required for corneal transparency(7Chakravarti S. Petroll W.M. Hassell J.R. Jester J.V. Lass J.H. Paul J. Birk D.E. Invest.Ophthalmol. Vis. Sci. 2000; 41: 3365-3373PubMed Google Scholar,8Birk D.E. Trelstad R.L. J.Cell Biol. 1984; 99: 2024-2033Crossref PubMed Scopus (222) Google Scholar). These predictions weresubstantiated by findings of corneal opacity, skin fragility, and abnormallylarge collagen fibril diameters and disorganized interfibrillar spacingpresent in lumican-null mice(1Chakravarti S. Magnuson T. Lass J.H. Jepsen K.J. LaMantia C. Carroll H. J. CellBiol. 1998; 141: 1277-1286Crossref PubMed Scopus (570) Google Scholar,7Chakravarti S. Petroll W.M. Hassell J.R. Jester J.V. Lass J.H. Paul J. Birk D.E. Invest.Ophthalmol. Vis. Sci. 2000; 41: 3365-3373PubMed Google Scholar,9Austin B.A. Coulon C. Liu C.Y. Kao W.W. Rada J.A. Invest Ophthalmol. Vis. Sci. 2002; 43: 1695-1701PubMed Google Scholar, 10Neame P.J. Kay C.J. McQuillan D.J. Beales M.P. Hassell J.R. Cell Mol. Life Sci. 2000; 57: 859-863Crossref PubMed Scopus (121) Google Scholar, 11Rada J.A. Cornuet P.K. Hassell J.R. Exp. Eye Res. 1993; 56: 635-648Crossref PubMed Scopus (286) Google Scholar).More recently, studies have showed lumican involvement in cell migration andproliferation during embryonic development and wound healing(12Saika S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google Scholar, 13Cornuet P.K. Blochberger T.C. Hassell J.R. Invest. Ophthalmol. Vis. Sci. 1994; 35: 870-877PubMed Google Scholar, 14Doane K.J. Ting W.H. McLaughlin J.S. Birk D.E. Exp. Eye Res. 1996; 62: 271-283Crossref PubMed Scopus (24) Google Scholar, 15Wilda M. Bachner D. Just W. Geerkens C. Kraus P. Vogel W. Hameister H. J. Bone Miner.Res. 2000; 15: 2187-2196Crossref PubMed Scopus (53) Google Scholar).The delayed epithelial wound healing phenotype inLum–/– mice is potentially due to theinvolvement of lumican in cellular migration, adhesion, and/or proliferation(12Saika S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google Scholar,16Vij N. Roberts L. Joyce S. Chakravarti S. Exp. Eye Res. 2004; 78: 957-971Crossref PubMed Scopus (90) Google Scholar). Under normal conditions,lumican is not expressed by epithelial cells, but transient expression isreported by migrating cells during wound healing and development(12Saika S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google Scholar,17Ying S. Shiraishi A. Kao C.W. Converse R.L. Funderburgh J.L. Swiergiel J. Roth M.R. Conrad G.W. Kao W.W. J. Biol. Chem. 1997; 272: 30306-30313Abstract Full Text Full Text PDF PubMed Scopus (117) Google Scholar). Impairment of cellmigration and proliferation may partially explain the delay of epithelialwound healing in lumican-null mice. Recent reports have also showed delayedepithelial-mesenchymal transition in lumican-null mice. Lumican-null mouselens epithelial cells showed decreased α-smooth muscle actin expressionand the delayed epithelial-mesenchymal transition induction by transforminggrowth factor β-2 in vitro(18Saika S. Miyamoto T. Tanaka S. Tanaka T. Ishida I. Ohnishi Y. Ooshima A. Ishiwata T. Asano G. Chikama T. Shiraishi A. Liu C.Y. Kao C.W. Kao W.W. InvestOphthalmol. Vis. Sci. 2003; 44: 2094-2102Crossref PubMed Scopus (120) Google Scholar). A role for lumican hasalso been suggested in growth and metastasis of breast, colon, and pancreaticcancer(19Leygue E. Snell L. Dotzlaw H. Troup S. Hiller-Hitchcock T. Murphy L.C. Roughley P.J. Watson P.H. J. Pathol. 2000; 192: 313-320Crossref PubMed Scopus (127) Google Scholar, 20Lu Y.P. Ishiwata T. Kawahara K. Watanabe M. Naito Z. Moriyama Y. Sugisaki Y. Asano G. Pathol. Int. 2002; 52: 519-526Crossref PubMed Scopus (58) Google Scholar, 21Ping L.Y. Ishiwata T. Asano G. J. Pathol. 2002; 196: 324-330Crossref PubMed Scopus (66) Google Scholar)and in cellular apoptosis(21Ping L.Y. Ishiwata T. Asano G. J. Pathol. 2002; 196: 324-330Crossref PubMed Scopus (66) Google Scholar). A cell surface receptorfor lumican has also been demonstrated(22Funderburgh J.L. Mitschler R.R. Funderburgh M.L. Roth M.R. Chapes S.K. Conrad G.W. InvestOphthalmol. Vis. Sci. 1997; 38: 1159-1167PubMed Google Scholar). These observationsdemonstrate lumican to have multiple functions, and it may serve as amatrikine in regulating cellular activities via interaction with integrinand/or growth factor receptors besides serving as a component of ECM, similarto what has been suggested for many other ECM components(23Swindle C.S. Tran K.T. Johnson T.D. Banerjee P. Mayes A.M. Griffith L. Wells A. J. CellBiol. 2001; 154: 459-468Crossref PubMed Scopus (219) Google Scholar, 24Duca L. Floquet N. Alix A.J. Haye B. Debelle L. Crit Rev. Oncol. Hematol. 2004; 49: 235-244Crossref PubMed Scopus (149) Google Scholar, 25Tran K.T. Griffith L. Wells A. Wound. Repair Regen. 2004; 12: 262-268Crossref PubMed Scopus (136) Google Scholar).The full extent of cellular functions mediated by lumican, however, remains tobe determined. Keratan sulfate-containing proteoglycans (KSPGs) are uniquely abundant inthe cornea and have long been thought to be essential for cornealtransparency. Lumican constitutes only about half of corneal KSPG. Most of theremaining corneal keratan sulfate modifies keratocan, a protein with highsequence similarity to lumican. In adult tissues, keratocan is limited tocorneal stroma, and keratocan expression is considered a phenotypic marker forkeratocytes. Lumican-null (Lum–/–) micemanifest corneal opacity, skin fragility, and impaired collagenfibrillogenesis (1Chakravarti S. Magnuson T. Lass J.H. Jepsen K.J. LaMantia C. Carroll H. J. CellBiol. 1998; 141: 1277-1286Crossref PubMed Scopus (570) Google S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google however, of only results in a of cornea in of the collagen matrix C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google A.J. C. Liu C.Y. Kao W.W. Biol. 2003; PubMed Scopus Google Scholar). of the of similarity in and has been no explanation for the the and In the current study, we have the as to the ofthe in the cornea by lumican in the corneal stroma of This the of a corneal collagen KSPG. The results that the lumican not have corneal or corneal a lumican expression found to effect the expression of These findings a for the differences of in corneas andKera–/– mice and a novel of lumican. of a a by the of used to lumican of as a in K. Liu C.Y. R.L. Birk D.E. Funderburgh J.L. Kao W.W. 2003; PubMed Google Scholar). The by and and to promoter in the C. H. Kao C. Kao W.W. 2000; PubMed Scopus Google Scholar). The of minigene by in A of is minigene by and and injection of by the mice used for with a and a the for for for for and for cornea used in of of of reverse and of corneal RNA to the to The for for and for and A of the to of a of of of of and The for at for for and for for by a at at the of and corneas and and in of and to Corneas a for at and the at in a by for in of in The and with a at about of protein be with at as J.L. Funderburgh M.L. Liu C.Y. Kao W.W. Conrad G.W. Invest. Ophthalmol. Vis. Sci. 1995; Google and development to of to lumican keratocan used as S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). In confocal microscopythrough used to corneal epithelial and corneal in the mice for by injection of and cornea a confocal with a surface J.V. L.Y. J. Chakravarti S. Paul J. Petroll W.M. 2001; 41: PubMed Scopus Google Scholar, J.V. Petroll W.M. PubMed Scopus Google Scholar, J. Jester J.V. T.D. Petroll W.M. Invest Ophthalmol. Vis. Sci. 2000; 41: Google to and a of the ofthe to serve as an a the cornea at that be to a of corneal and be in the to the of and for the stroma J.V. Petroll W.M. PubMed Scopus Google J. Jester J.V. T.D. Petroll W.M. Invest Ophthalmol. Vis. Sci. 2000; 41: Google Scholar). the the to the corneal stroma of J. Jester J.V. T.D. Petroll W.M. Invest Ophthalmol. Vis. Sci. 2000; 41: Google Petroll W.M. T. Jester J.V. 1997; PubMed Scopus Google Scholar). The and corneal the and the and mice by the in in with for D.E. Trelstad R.L. J.Cell Biol. 1984; 99: 2024-2033Crossref PubMed Scopus (222) Google Scholar).The corneas with and with in an by the corneas and a of and a and a and with cornea and the and stroma examined at a for the and fibril diameters in of the and stroma the at in a the and and diameters RNA corneas of RNA to keratocan as C.Y. Shiraishi A. Kao C.W. Converse R.L. Funderburgh J.L. Conrad G.W. Kao W.W. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). with a mice after for or S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google Scholar). and to the for in and the of in by the and of the by and of to to as The mice to intrastromal Liu C.Y. M. B. J. Invest Sci. 2004; PubMed Scopus Google and during in a A.M. Y. J. C. Invest Ophthalmol. Vis. Sci. 2001; Google Scholar). A small in the corneal a to a the the stroma. The the stroma, and of K. Liu C.Y. R.L. Birk D.E. Funderburgh J.L. Kao W.W. 2003; PubMed Google corneal stroma injection and to and for lumican and and lumican in the and for lumican and keratocan as used as as S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). with The a and by Lumican of at for as C.J. Roth M.R. Funderburgh M. Conrad G.W. Funderburgh J.L. Chem. 2000; 275: Full Text Full Text PDF PubMed Scopus Google Scholar). These or a of RNA lumican at a of in The the lumican a The and of the as after and cells in and proteoglycans after as the by and J.L. Mann M.M. Funderburgh M.L. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). in at with Lumican and keratocan in the after a a to lumican by RNA at for of keratocan, and J.L. Mann M.M. Funderburgh M.L. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). injection of in lumican-null mice as Liu C.Y. M. B. J. Invest Sci. 2004; PubMed Scopus Google Scholar). with of in a K. Liu C.Y. R.L. Birk D.E. Funderburgh J.L. Kao W.W. 2003; PubMed Google Scholar). corneas and activity reporter aβ-galactosidase to In corneal with a in of reporter in the a protein to the of used to the activity and to used to and The of to the role of lumican inthe corneal stroma by mice with a lumican minigene a protein the of the keratocan promoter C. H. Kao C. Kao W.W. 2000; PubMed Scopus Google of the minigene used to mice by with a and and the used for for the reporter The expression by and for a of mRNA the and used for and other also for the not by for the of mRNA of the by not not and mice the of protein in with S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google Scholar).Lumican in corneas of the that of corneas of of expression by the in the of the of protein, and mice showed a protein by in of lumican by the in the The in demonstrate that no differences and mice for epithelial and The epithelial and in mice and in the mice the level of in the mice with the not mice. mice of corneal with the of expression of the of of expression corneal epithelial and with the is for and and as by in a that the and the and and in normal differences and stroma. in the to be limited with less not in diameters the stroma and whereas stroma and for and diameters in the for lumican expression and corneal of keratocan expression to lumican overexpression corneal expression C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google keratocan expression in cornea of that of an by lumican. lumican keratocan proteoglycans corneas and mice. the results of a in corneal by expression decreased in the with wild type but a keratocan expression in the the lumican keratocan mRNA with corneal RNA and mice mRNA of that the keratocan mRNA mice decreased with and mice. the showed an in keratocan mRNA as with the These results the that lumican expression by the The results of to novel role of lumican in the to expression of a reporter in the A.M. Y. J. C. Invest Ophthalmol. Vis. Sci. 2001; Google Scholar). role of lumican in keratocan the corneal stroma of lumican-null mice to in lumican an in keratocan used in and with and for lumican. corneas exhibit the lumican Lumican in the increased expression by to keratocan in intrastromal of the Lum–/– of to the corneal stroma results in lumicanexpression by no lumican be the at a in keratocan expression by of as with the the This the keratocan for lumican and of corneal mice injection of or as intrastromal injection also after injection as with the In to the in keratocan examined in in vitro in been reduced by that lumican the during a a with in the by mRNA showed a of after lumican the other not altered in the of lumican to keratocan in and in vitro the by that lumican keratocan a keratocan promoter driven reporter mice with a or an corneas and determined. a of the of the or lumican minigene Lumican expression the the keratocan the of lumican to expression at the promoter level. and lumican are of collagen matrix organization in the corneal W.W. Liu C.Y. J. 2002; PubMed Scopus Google Scholar). lumican a role in as wound epithelial-mesenchymal and S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google S. Miyamoto T. Tanaka S. Tanaka T. Ishida I. Ohnishi Y. Ooshima A. Ishiwata T. Asano G. Chikama T. Shiraishi A. Liu C.Y. Kao C.W. Kao W.W. InvestOphthalmol. Vis. Sci. 2003; 44: 2094-2102Crossref PubMed Scopus (120) Google E. Snell L. Dotzlaw H. Troup S. Hiller-Hitchcock T. Murphy L.C. Roughley P.J. Watson P.H. J. Pathol. 2000; 192: 313-320Crossref PubMed Scopus (127) Google L.Y. Ishiwata T. Asano G. J. Pathol. 2002; 196: 324-330Crossref PubMed Scopus (66) Google L.Y. Ishiwata T. Asano G. J. Pathol. 2002; 196: 324-330Crossref PubMed Scopus (66) Google E. Snell L. Dotzlaw H. K. Hiller-Hitchcock T. Roughley P.J. Watson P.H. Murphy L.C. Res. 1998; Google Scholar). The results reported a novel role of lumican as a of keratocan by The of the lumican and keratocan-null reported (1Chakravarti S. Magnuson T. Lass J.H. Jepsen K.J. LaMantia C. Carroll H. J. CellBiol. 1998; 141: 1277-1286Crossref PubMed Scopus (570) Google S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). the has a corneal and skin to altered collagen delayed corneal (1Chakravarti S. Magnuson T. Lass J.H. Jepsen K.J. LaMantia C. Carroll H. J. CellBiol. 1998; 141: 1277-1286Crossref PubMed Scopus (570) Google S. Shiraishi A. Saika S. Liu C-Y. Funderburgh J.L. Kao C.W.C. Converse R.L. Kao W.W.Y. J. Biol. Chem. 2000; 275: 2607-2612Abstract Full Text Full Text PDF PubMed Scopus (206) Google and delayedepithelial-mesenchymal transition of S. Miyamoto T. Tanaka S. Tanaka T. Ishida I. Ohnishi Y. Ooshima A. Ishiwata T. Asano G. Chikama T. Shiraishi A. Liu C.Y. Kao C.W. Kao W.W. InvestOphthalmol. Vis. Sci. 2003; 44: 2094-2102Crossref PubMed Scopus (120) Google the keratocan-null of but cornea and corneal as examined to C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google A.J. C. Liu C.Y. Kao W.W. Biol. 2003; PubMed Scopus Google Scholar). ofthe keratocan-null showed no alteration of other in the C.Y. Birk D.E. Hassell J.R. B. Kao W.W. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). The the expression in the and a in the Lum–/– mice as type This that and in the lumican are not the of of lumican expression but also the of a expression by corneal stroma mice. is of to a of lumican expression in of mice suggested that of lumican in the phenotype K.J. J.H. Paul J. Roberts L. Y. Oldberg A. Birk D.E. Chakravarti S. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). The that the expression of of the family may be other in the The results of studies to a The of to the role of lumican inthe cornea lumican a wild collagen and collagen in an overexpression be corneal fibril and transparency of The of corneal of is not detrimental to The keratocan and mice have normal and only the Lum–/– mice current that the lumican lumican but also have reduced that corneal a and that a is not be This may be in by the of a of collagen L. I. Oldberg A. FEBS Lett. 2000; PubMed Scopus Google Scholar). a in lumican expression in intrastromal injection of lumican in lumican but anincrease in keratocan the lumican expression in the keratocan of regulating keratocan expression in the adult The lumican may serve as a for keratocan expression to the by lumican an in promoter the to keratocan promoter activity inLum–/– mice as observations are with the that of may have functions besides serving as of J. 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