大鼠胚胎皮肤无瘢痕性愈合机制的实验研究
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摘要
先天性唇腭裂是我国新生儿中最常见的先天性畸形之一,唇部遗留的手术瘢痕将影响整个面部的协调美观,腭裂修复术后硬腭裸露骨面遗留的瘢痕挛缩将使患者上颌骨的生长发育受限,加重唇聘裂术后继发畸形的外观丑陋程度,加重患者的精神心理负担,不利于唇腭裂序列治疗总体目标的实现。
瘢痕的产生是由于局部皮肤、粘膜遭受手术创伤后,不能完全达到组织学的再生,为了恢复组织完整性而以结缔组织替代进行修复,并引起外观形态和/或功能改变的产物。目前,尚未找到一种理想的方法来控制瘢痕增生。
1979年,Rowlatt首次报道在人类胚胎发现无瘢痕性愈合Scarless Healing),随后,在鼠、兔、羊等其他动物胚胎中也发现了类似的现象。胚胎无瘢痕性愈合是胚胎在特定的时期受到创伤后,以周围正常组织再生完成修复,是胚胎特有的性质,其发生机理尚不清楚。通过对唇腭裂动物模型行子宫内修复术发现,手术后的胚胎局部伤口完全愈合,没有瘢痕组织形成,并且上颌骨的生长发育正常。
如果阐明胚胎无瘢痕性愈合的机理,就有可能使成体组织瘢痕性愈合向类似于胚胎样的无瘢痕性愈合转变,使瘢痕的治疗得到突破,极大
Cleft lip and palate is one of the highest incidences of congenital deformities in China. After the surgery repairing, the scar will not only influence the feature but also affect the growthing of maxillary. There is not resultful treatment to reduce the scar. In adults wound healing restores tissue integrity, but the cost is fibrosis and scar. In contrast, the early-gestation fetus has the remarkable ability to heal skin wounds without a scar. This observation was fist reported more than two decades ago and was subsequently confirmed in both animal modes and human fetuses. Since that time, an intensive research effort has focused on unraveling the mechanisms underlying scarless fetal wound repair. Fetal wounds pass from scarless repair to healing with scar formation during gestation. This transition depends on both the size of the wound and the gestational age of the fetus. The mechanism for scarless fetal repair is unknown, but it does not require systemic factors such as the fetal immune system, fetal serum, or amniotic fluid. The cellular gene regulation rather than the external environment is the critical factor in scarless repair.
Transforming growth factor-p has been implicated in the ontogenetic transition from scarless fetal repair to adult repair with scar. Fibromodulin can bind and potentially inhibit TGF-P activity. CTGF is present and frequently overexpressed in fibrotic conditions. We hypothesize that TGF-P, CTGF and fibromodulin expression be differentially regulated during the transition from early gestation wounds manifesting scarless fetal-type repair to late gestation wounds manifesting adult-type repair with scar.To investigate the mechanism for scarless fetal repair, skin from fetal Sprague-Dawley rats at time points that represented both the scarless and scar-forming periods of rat gestation was harvested. We analyzed the expression of TGF-pi,-p3, CTGF, and fibromodulin during cutaneous fetal repair by real time PCR. The result suggested that the ontogenetic transition from scarless fetal-type repair to adult-type repair weth scar correlate with the fibromodulin. Fibromodulin may have antifibrotic roles in fetal wound repair. In order to investigate the role of mechanism for fibromodulin in the fetal scarless healing, we constracted the siRNA expression carrier of fibromodulin. After transfected the fibroblast of El 6 and El 8, the fibromodulin's mRNA was effective interfered by analyzed by RT-PCR. The expression of TGF-pi and CTGF increased while the expression of TGF-P3 decreased after the RNA interference of fibromodulin. This suggested that fibromodulin may be a biologically relevant modulator of TGF-p activity and CTGF activity during scarless formation. Relative abundance of fibromodulin in early gestation wounds may partly affect scarless fetal repair through decreased TGF-P and CTGF bioavailability.Base on this foundings, increased fibromodulin induction in fetal
wounds offers a novel explanation of fetal wound collagens deposited in a more organized fashion than collagen in adult wounds. Potential strategies for the manipulation of adult wounds into being more "fetal like" may include the addition of fibromodulin to modulate both TGF-p\ CTGF activity and ECM assembly.
瘢痕的产生是由于局部皮肤、粘膜遭受手术创伤后,不能完全达到组织学的再生,为了恢复组织完整性而以结缔组织替代进行修复,并引起外观形态和/或功能改变的产物。目前,尚未找到一种理想的方法来控制瘢痕增生。
1979年,Rowlatt首次报道在人类胚胎发现无瘢痕性愈合Scarless Healing),随后,在鼠、兔、羊等其他动物胚胎中也发现了类似的现象。胚胎无瘢痕性愈合是胚胎在特定的时期受到创伤后,以周围正常组织再生完成修复,是胚胎特有的性质,其发生机理尚不清楚。通过对唇腭裂动物模型行子宫内修复术发现,手术后的胚胎局部伤口完全愈合,没有瘢痕组织形成,并且上颌骨的生长发育正常。
如果阐明胚胎无瘢痕性愈合的机理,就有可能使成体组织瘢痕性愈合向类似于胚胎样的无瘢痕性愈合转变,使瘢痕的治疗得到突破,极大
Cleft lip and palate is one of the highest incidences of congenital deformities in China. After the surgery repairing, the scar will not only influence the feature but also affect the growthing of maxillary. There is not resultful treatment to reduce the scar. In adults wound healing restores tissue integrity, but the cost is fibrosis and scar. In contrast, the early-gestation fetus has the remarkable ability to heal skin wounds without a scar. This observation was fist reported more than two decades ago and was subsequently confirmed in both animal modes and human fetuses. Since that time, an intensive research effort has focused on unraveling the mechanisms underlying scarless fetal wound repair. Fetal wounds pass from scarless repair to healing with scar formation during gestation. This transition depends on both the size of the wound and the gestational age of the fetus. The mechanism for scarless fetal repair is unknown, but it does not require systemic factors such as the fetal immune system, fetal serum, or amniotic fluid. The cellular gene regulation rather than the external environment is the critical factor in scarless repair.
Transforming growth factor-p has been implicated in the ontogenetic transition from scarless fetal repair to adult repair with scar. Fibromodulin can bind and potentially inhibit TGF-P activity. CTGF is present and frequently overexpressed in fibrotic conditions. We hypothesize that TGF-P, CTGF and fibromodulin expression be differentially regulated during the transition from early gestation wounds manifesting scarless fetal-type repair to late gestation wounds manifesting adult-type repair with scar.To investigate the mechanism for scarless fetal repair, skin from fetal Sprague-Dawley rats at time points that represented both the scarless and scar-forming periods of rat gestation was harvested. We analyzed the expression of TGF-pi,-p3, CTGF, and fibromodulin during cutaneous fetal repair by real time PCR. The result suggested that the ontogenetic transition from scarless fetal-type repair to adult-type repair weth scar correlate with the fibromodulin. Fibromodulin may have antifibrotic roles in fetal wound repair. In order to investigate the role of mechanism for fibromodulin in the fetal scarless healing, we constracted the siRNA expression carrier of fibromodulin. After transfected the fibroblast of El 6 and El 8, the fibromodulin's mRNA was effective interfered by analyzed by RT-PCR. The expression of TGF-pi and CTGF increased while the expression of TGF-P3 decreased after the RNA interference of fibromodulin. This suggested that fibromodulin may be a biologically relevant modulator of TGF-p activity and CTGF activity during scarless formation. Relative abundance of fibromodulin in early gestation wounds may partly affect scarless fetal repair through decreased TGF-P and CTGF bioavailability.Base on this foundings, increased fibromodulin induction in fetal
wounds offers a novel explanation of fetal wound collagens deposited in a more organized fashion than collagen in adult wounds. Potential strategies for the manipulation of adult wounds into being more "fetal like" may include the addition of fibromodulin to modulate both TGF-p\ CTGF activity and ECM assembly.
引文
[1] 石冰.唇腭裂修复外科学.成都:四川大学出版社,2003.
[2] 李荟元,鲁开化,郭树忠.新编瘢痕学.西安:第四军医大学出版社,2003.
[3] Rowlatt U. Intrauterine healing in a 20-week human fetus. Virchows Arch. 1979; 381:353-361.
[4] Armstrong JR, Ferguson MWJ. Ontogeny of the skin and transition from scar free to scarring phenotype during wound healing in the pouch young of Monodelphis domestica. Dev Biol. 1995;169:242-260.
[5] Longaker MT, WhitbyDJ, Adzick NS, et al. Studies in fetal wound healing. Ⅵ. Second and third trimester fetal wounds demonstrate rapid collagen deposition without scar formation. J Pediatr Surg 1990;25: 63-9.
[6] Ihara S, Motobayashi Y, Nagao E, et al. Ontogenic transition of wound healing pattern in rat skin occurring at the fetal stage. Development 1990;110:671-680.
[7] WhitbyDJ, Ferguson MWJ. The extracellular matrix in fetal and adult wound healing. Development 1991; 112:651-668.
[8] Lorenz HP, WhitbyDJ, Longaker MT, et al. Fetal wound healing: the ontogeny of scar formation in the nonhuman primate. Ann Surg. 1993; 21:391-396.
[9] Longaker MT, Dodson TB, Kaban LB. A rabbit model for fetal cleft lip repair. J Oral Maxillofac Surg 1990;48:714-9.
[10] Longaker MT, Stern M, Lorenz P, et al. A model for fetal cleft lip repair in lambs. Plast Reconstr Surg 1992; 90: 750-6.
[11] Hedrick MH, Rice HE, Wander WKJ, et al. Delayed in utero repair of surgically created fetal cleft lip and palate. Plast Reconstr Surg 1996; 97:900-907.
[12] McCallion RL, Ferguson MWJ. Fetal wound healing and the development of antiscarring therapies for adult wound healing. In: Clark RAF, editor. The molecular and cellular biology of wound repair, vol.ⅹⅹⅲ. New York: Plenum Press; 1996. p.561-600.
[13] Cass DL, Bullard KM, Sylvester KG, et al. Wound size and gestational age modulate scar formation in fetal wound repair. J Pediatr Surg. 1997; 32:411-415.
[14] Lorenz HP, Longaker MT, Perkocha LA, et al. Scarless wound repair: a human fetal skin model. Development 1992; 114:253-259.
[15] Longaker MT, Moelleken BR, Cheng JC, et al. Fetal fracture healing in a lamb model. Plast Reconstr Surg. 1992;90:161-171.
[16] Longaker MT, WhitbyDJ, Jennings RW, et al. Fetal diaphragmatic wounds heal with scar formation. J Surg Res. 1991;50:375-85.
[17] Meuli M, Lorenz HP, Hedrick MH, et al. Scar formation in the fetal alimentary tract. J Pediatr Surg. 1995;30:392-5.
[18] Lorenz HP, Lin RY, Longaker MT, et al. The fetal fibroblast: the effect or cell of scarless wounds repair. Plast Reconst Surg. 1995;96: 1251-9.
[19] Shah M, Forman DM, Ferguson MWJ. Neutralization of TGF-β1 and TGF-β2 or exogenous addition of TGF-β3 to cutaneous rat wounds reduces scarring. J Cell Sci. 1995;108:985-1002
[20] Krummel TM, Michna BA, Thomas B. TGF-p induces fibrosis in a fetal wound model. J Pediatr Surg. 1988;23:647-52.
[21] Shah M, Foreman DM, Ferguson MWJ. Neutralizing antibody to TGF_1,2 reduces cutaneous scarring in adult rodents. J Cell Sci. 1994; 107:1137-57.
[22] Shah M, Foreman DM, Ferguson MWJ. Control of scar in adult wounds by neutralizing antibody to transforming growth factor. Lancet 1992;339:213-4.
[23] Bradham DM, Igarashi A, Grotendorst GR. Connective tissue growth factor: a cysteine-rich mitogen secreted by human vascular endothelial cells is related to the SRC-induced immediate early gene product CEF-10. J Cell Biol 1991; 114: 1285-94.
[24] Leask A, Holmes A, Abraham DJ. Connective tissue growth factor: a new and important player in the pathogenesis of fibrosis. Curr Rheumatol Rep 2002; 4: 136-42.
[25] Hedbom, E., and D. Heinegard. 1989. Interaction of a 59-kDa connective tissue matrix protein with collagen I and collagen II. J. Biol. Chem. 264:6898-6905.
[26] Hedbom, E., and D. Heinegard. 1993. Binding of fibromodulin and decorin to separate sites on fibrillar collagens. J. Biol. Chem. 268:27307-27312.
[27] Hedlund, H., S. Mengarelli-Widholm, D. Heinegard, F.P. Fibromodulin distribution and association with collagen. Matrix Biol. 14:227-232.
[28] Hindebrand A, Romaris M, Rasmussen LM, et al. Interaction of the small interstitial proteoglycans biglycan, decorin and fibromodulin with transforming growth factor beta. Biochem J 1994; 302: 527-534.
[29] Border WA, Noble NA, et al. Natural inhibitor of transforming growth factor beta protects against scarring in experimental kidney disease. Nature 1992,360:361-4.
[30] Longaker MT, WhitbyDJ, Ferguson MWJ, et al. Adult skin wounds in the fetal environment heal with scar formation. Ann Surg. 1994;219: 65-72.
[31] Krummel TM, Michna BA, Thomas B. TGF-p induces fibrosis in a fetal wound model. J Pediatr Surg. 1988;23:647-52.
[32] Sullivan KM, Lorenz HP, Adzick NS. The role of TGF-p in human fetal wound healing. Surg Forum 1993;44:625-7.
[33] Mackool RJ, Soo C, Gittes G, et al. Endogenous expression of transforming growth factor (TGF)-p, TGF-P, receptors, and TGF-P, activity modulators as a function of gestational age in fetal rat skin. Surg Forum. 1997;48:516-9.
[34] Dang C, Beanes SR, Soo C, et al. A high ratio of TGF-p3 to TGF-β1 expression in wounds is associated with scarless repair. Wound Repair Regen. 2001 ;9:153.
[35] Hsu M, Peled ZM, Chin GS, et al. Ontogeny of expression of transforming growth factor-beta 1 (TGF-beta 1), TGF-beta 3, and TGF-beta receptors I and II in fetal rat fibroblasts and skin. Plast Reconstr Surg. 2001;107:1787-94.
[36] Kaartinen V, Voncken JW, Shuler C, et al. Abnormal lung development and cleft palate in mice lacking TGF- β 3 indicates defects of epithelial-mesenchymal interaction. Nat Genet 1995; 11:415-421.
[37] Moussad EE, Brigstock DR. Connective tissue growth factor: what's in a name? Mol Genet Metab 2000; 71: 276-92.
[38] Inn H. Pahtogenesis of fibrosis: role of TGF-beta and CTGF. Curr Opin Rheumatol 2002; 14:681-5.
[39] Wang JF, Olson ME, Reno CR, et al. Molecular and cell biology of skin wound healing in a pig model.connect Tissur Res 2000; 41: 195-211.
[40] Grotendorst GR. Connective tissue growth factor a mediator of TGF-beta action on fibroblasts. Cytokine Growth Factor Rev 1997; 8:171-9.
[41] Duncan MR, Frazier KS, Abramson S, et al. Connective tissue growth factor mediates transforming growth factor beta induced collagen synthesis: down-regulation by cAMP. FASEBJ, 1999;13( 13)1774-83.
[42]Kothapfi D , Grotendorst GR. CTGF modulates cell cycle progression in cAMP-arrested NRK fibroblasts. J Cell Physiol, 2000, 182(1): 119-24.
[43] Mori T, Kawara S, Shinozaki M, et al. Role and interaction of connective tissue growth factor with transforming growth factor-beta in persistent fibrosis: A mouse fibrosis model. J Cell Physi 1999;181(l):153-8.
[44] WhitbyDJ, Ferguson MWJ. The extracellular matrix in fetal and adult wound healing. Development 1991;112:651-668.
[45] Matrisian LM. The matrix-degrading metalloproteinases. Bioessays 1992;14:455-63.
[46] Woessner JF. Matrix metalloproteinases and their inhibitors in connective tissue remodeling. Fed Am Soc Exp Biol. 1991;5:2145- 54.
[47] Bullard KM, Cass DL, Banda MJ, et al. Transforming growth factor beta-1 decreases interstitial collagenase in healing human fetal skin. J PediatrSurg. 1997;32:1023-7.
[48] Lorenz HP, Soo C, Beanes SR, et al. Differential expression of matrix metalloproteinases and their tissue-derived inhibitors in scarless fetal wound healing. Surg Forum 2001;52397-401.
[49] Fire A, Xu S, Mello CC.et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans Nature 1998 ; 391 (6669):806-11
[50] Bernstein E, Hammond SM, Harnnon GJ. Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature. 2001;409 (6818) :363-6.
[51] Hammond, S. M. , Kobayashi, R. & Hannon, G. J.et al. Argonaute2, a Link Between Genetic and Biochemical Analyses of RNAi. Science. 2001; 293 (5532): 1146-50.
[52] Sijen T, Simmer F, Fire A.et al. On the role of RNA amplification in dsRNA-triggered gene silencing. Cell .2001;107(4):465-76.
[53] Lipardi C, Wei Q, Paterson BM. RNAi as random degradative PCR: siRNA primers convert mRNA into dsRNAs that are degraded to generate new siRNAs. Cell.2001 ;107(3):297-307.
[54] Billy E, Zhang HD, Filipowicz W. Specific interference with gene expression induced by long, double-stranded RNA in mouse embryonal teratocarcinoma cell lines. Proc. Natl. Acad. Sci. USA. 2001; 98 (25): 14428-33.
[55] Elbashir SM, Lendeckel W, Tuschl T.et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature. 2001 ;411(6836): 428-9.
[56] Holen T, Wiiger M.T,Prydz H. Positional effects of short interfering RNAs targeting the human coagulation trigger Tissue Factor.Nucleic Acids Research.2002; 30(8): 1757-1766.
[57] Brummelkamp.T.R,Bernads. R,Agami.R. A system for stable expression of short interfering RNAs in Mammalian cells. Science. 2002; 296 :550-553.
[58] Brunner A, Chinn J, Neubauer M. Identification of a gene family regulated by transforming growth factor beta. DNA Cell Biol 1991; 10:293-300.
[59] Monaco JL, Lawrence WT. Acute wound healing An overview. Clin Plastic Surg 30(2003) 1-12.
[60] Longaker MT, Chiu E, Adzick NS, et al. Studies in fetal wound healing. V. A prolonged presence of hyaluronic acid characterizes fetal wound fluid. Ann Surg. 1991;213:292-6.
[61] Estes JM, Adzick NS, Harrison MR, et al. Hyaluronate metabolism undergoes an ontogenic transition during fetal development: implications for scar-free wound healing. J Pediatr Surg. 1993;28:1227—31.
[62] Longaker MT, Chiu ES, Harrison MR, et al. Studies in fetal wound healing. IV. Hyaluronic acid-stimulating activity distinguishes fetal wound fluid from adult wound fluid. Ann Surg. 1989;210:667-72.
[63] Alaish SM, Yager D, Diegelmann RF, et al. Biologyof fetal wound healing: hyaluronate receptor expression in fetal fibroblasts. J Pediatr Surg. 1994;29:1040-3.
[64] Grotendorst GR, Okochi H, Hayashi N. A novel transforming growth factor beta response element controls the expression of the connective tissur growth factor gene. Cell Growth Differ 1996;7:469-80.
[2] 李荟元,鲁开化,郭树忠.新编瘢痕学.西安:第四军医大学出版社,2003.
[3] Rowlatt U. Intrauterine healing in a 20-week human fetus. Virchows Arch. 1979; 381:353-361.
[4] Armstrong JR, Ferguson MWJ. Ontogeny of the skin and transition from scar free to scarring phenotype during wound healing in the pouch young of Monodelphis domestica. Dev Biol. 1995;169:242-260.
[5] Longaker MT, WhitbyDJ, Adzick NS, et al. Studies in fetal wound healing. Ⅵ. Second and third trimester fetal wounds demonstrate rapid collagen deposition without scar formation. J Pediatr Surg 1990;25: 63-9.
[6] Ihara S, Motobayashi Y, Nagao E, et al. Ontogenic transition of wound healing pattern in rat skin occurring at the fetal stage. Development 1990;110:671-680.
[7] WhitbyDJ, Ferguson MWJ. The extracellular matrix in fetal and adult wound healing. Development 1991; 112:651-668.
[8] Lorenz HP, WhitbyDJ, Longaker MT, et al. Fetal wound healing: the ontogeny of scar formation in the nonhuman primate. Ann Surg. 1993; 21:391-396.
[9] Longaker MT, Dodson TB, Kaban LB. A rabbit model for fetal cleft lip repair. J Oral Maxillofac Surg 1990;48:714-9.
[10] Longaker MT, Stern M, Lorenz P, et al. A model for fetal cleft lip repair in lambs. Plast Reconstr Surg 1992; 90: 750-6.
[11] Hedrick MH, Rice HE, Wander WKJ, et al. Delayed in utero repair of surgically created fetal cleft lip and palate. Plast Reconstr Surg 1996; 97:900-907.
[12] McCallion RL, Ferguson MWJ. Fetal wound healing and the development of antiscarring therapies for adult wound healing. In: Clark RAF, editor. The molecular and cellular biology of wound repair, vol.ⅹⅹⅲ. New York: Plenum Press; 1996. p.561-600.
[13] Cass DL, Bullard KM, Sylvester KG, et al. Wound size and gestational age modulate scar formation in fetal wound repair. J Pediatr Surg. 1997; 32:411-415.
[14] Lorenz HP, Longaker MT, Perkocha LA, et al. Scarless wound repair: a human fetal skin model. Development 1992; 114:253-259.
[15] Longaker MT, Moelleken BR, Cheng JC, et al. Fetal fracture healing in a lamb model. Plast Reconstr Surg. 1992;90:161-171.
[16] Longaker MT, WhitbyDJ, Jennings RW, et al. Fetal diaphragmatic wounds heal with scar formation. J Surg Res. 1991;50:375-85.
[17] Meuli M, Lorenz HP, Hedrick MH, et al. Scar formation in the fetal alimentary tract. J Pediatr Surg. 1995;30:392-5.
[18] Lorenz HP, Lin RY, Longaker MT, et al. The fetal fibroblast: the effect or cell of scarless wounds repair. Plast Reconst Surg. 1995;96: 1251-9.
[19] Shah M, Forman DM, Ferguson MWJ. Neutralization of TGF-β1 and TGF-β2 or exogenous addition of TGF-β3 to cutaneous rat wounds reduces scarring. J Cell Sci. 1995;108:985-1002
[20] Krummel TM, Michna BA, Thomas B. TGF-p induces fibrosis in a fetal wound model. J Pediatr Surg. 1988;23:647-52.
[21] Shah M, Foreman DM, Ferguson MWJ. Neutralizing antibody to TGF_1,2 reduces cutaneous scarring in adult rodents. J Cell Sci. 1994; 107:1137-57.
[22] Shah M, Foreman DM, Ferguson MWJ. Control of scar in adult wounds by neutralizing antibody to transforming growth factor. Lancet 1992;339:213-4.
[23] Bradham DM, Igarashi A, Grotendorst GR. Connective tissue growth factor: a cysteine-rich mitogen secreted by human vascular endothelial cells is related to the SRC-induced immediate early gene product CEF-10. J Cell Biol 1991; 114: 1285-94.
[24] Leask A, Holmes A, Abraham DJ. Connective tissue growth factor: a new and important player in the pathogenesis of fibrosis. Curr Rheumatol Rep 2002; 4: 136-42.
[25] Hedbom, E., and D. Heinegard. 1989. Interaction of a 59-kDa connective tissue matrix protein with collagen I and collagen II. J. Biol. Chem. 264:6898-6905.
[26] Hedbom, E., and D. Heinegard. 1993. Binding of fibromodulin and decorin to separate sites on fibrillar collagens. J. Biol. Chem. 268:27307-27312.
[27] Hedlund, H., S. Mengarelli-Widholm, D. Heinegard, F.P. Fibromodulin distribution and association with collagen. Matrix Biol. 14:227-232.
[28] Hindebrand A, Romaris M, Rasmussen LM, et al. Interaction of the small interstitial proteoglycans biglycan, decorin and fibromodulin with transforming growth factor beta. Biochem J 1994; 302: 527-534.
[29] Border WA, Noble NA, et al. Natural inhibitor of transforming growth factor beta protects against scarring in experimental kidney disease. Nature 1992,360:361-4.
[30] Longaker MT, WhitbyDJ, Ferguson MWJ, et al. Adult skin wounds in the fetal environment heal with scar formation. Ann Surg. 1994;219: 65-72.
[31] Krummel TM, Michna BA, Thomas B. TGF-p induces fibrosis in a fetal wound model. J Pediatr Surg. 1988;23:647-52.
[32] Sullivan KM, Lorenz HP, Adzick NS. The role of TGF-p in human fetal wound healing. Surg Forum 1993;44:625-7.
[33] Mackool RJ, Soo C, Gittes G, et al. Endogenous expression of transforming growth factor (TGF)-p, TGF-P, receptors, and TGF-P, activity modulators as a function of gestational age in fetal rat skin. Surg Forum. 1997;48:516-9.
[34] Dang C, Beanes SR, Soo C, et al. A high ratio of TGF-p3 to TGF-β1 expression in wounds is associated with scarless repair. Wound Repair Regen. 2001 ;9:153.
[35] Hsu M, Peled ZM, Chin GS, et al. Ontogeny of expression of transforming growth factor-beta 1 (TGF-beta 1), TGF-beta 3, and TGF-beta receptors I and II in fetal rat fibroblasts and skin. Plast Reconstr Surg. 2001;107:1787-94.
[36] Kaartinen V, Voncken JW, Shuler C, et al. Abnormal lung development and cleft palate in mice lacking TGF- β 3 indicates defects of epithelial-mesenchymal interaction. Nat Genet 1995; 11:415-421.
[37] Moussad EE, Brigstock DR. Connective tissue growth factor: what's in a name? Mol Genet Metab 2000; 71: 276-92.
[38] Inn H. Pahtogenesis of fibrosis: role of TGF-beta and CTGF. Curr Opin Rheumatol 2002; 14:681-5.
[39] Wang JF, Olson ME, Reno CR, et al. Molecular and cell biology of skin wound healing in a pig model.connect Tissur Res 2000; 41: 195-211.
[40] Grotendorst GR. Connective tissue growth factor a mediator of TGF-beta action on fibroblasts. Cytokine Growth Factor Rev 1997; 8:171-9.
[41] Duncan MR, Frazier KS, Abramson S, et al. Connective tissue growth factor mediates transforming growth factor beta induced collagen synthesis: down-regulation by cAMP. FASEBJ, 1999;13( 13)1774-83.
[42]Kothapfi D , Grotendorst GR. CTGF modulates cell cycle progression in cAMP-arrested NRK fibroblasts. J Cell Physiol, 2000, 182(1): 119-24.
[43] Mori T, Kawara S, Shinozaki M, et al. Role and interaction of connective tissue growth factor with transforming growth factor-beta in persistent fibrosis: A mouse fibrosis model. J Cell Physi 1999;181(l):153-8.
[44] WhitbyDJ, Ferguson MWJ. The extracellular matrix in fetal and adult wound healing. Development 1991;112:651-668.
[45] Matrisian LM. The matrix-degrading metalloproteinases. Bioessays 1992;14:455-63.
[46] Woessner JF. Matrix metalloproteinases and their inhibitors in connective tissue remodeling. Fed Am Soc Exp Biol. 1991;5:2145- 54.
[47] Bullard KM, Cass DL, Banda MJ, et al. Transforming growth factor beta-1 decreases interstitial collagenase in healing human fetal skin. J PediatrSurg. 1997;32:1023-7.
[48] Lorenz HP, Soo C, Beanes SR, et al. Differential expression of matrix metalloproteinases and their tissue-derived inhibitors in scarless fetal wound healing. Surg Forum 2001;52397-401.
[49] Fire A, Xu S, Mello CC.et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans Nature 1998 ; 391 (6669):806-11
[50] Bernstein E, Hammond SM, Harnnon GJ. Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature. 2001;409 (6818) :363-6.
[51] Hammond, S. M. , Kobayashi, R. & Hannon, G. J.et al. Argonaute2, a Link Between Genetic and Biochemical Analyses of RNAi. Science. 2001; 293 (5532): 1146-50.
[52] Sijen T, Simmer F, Fire A.et al. On the role of RNA amplification in dsRNA-triggered gene silencing. Cell .2001;107(4):465-76.
[53] Lipardi C, Wei Q, Paterson BM. RNAi as random degradative PCR: siRNA primers convert mRNA into dsRNAs that are degraded to generate new siRNAs. Cell.2001 ;107(3):297-307.
[54] Billy E, Zhang HD, Filipowicz W. Specific interference with gene expression induced by long, double-stranded RNA in mouse embryonal teratocarcinoma cell lines. Proc. Natl. Acad. Sci. USA. 2001; 98 (25): 14428-33.
[55] Elbashir SM, Lendeckel W, Tuschl T.et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature. 2001 ;411(6836): 428-9.
[56] Holen T, Wiiger M.T,Prydz H. Positional effects of short interfering RNAs targeting the human coagulation trigger Tissue Factor.Nucleic Acids Research.2002; 30(8): 1757-1766.
[57] Brummelkamp.T.R,Bernads. R,Agami.R. A system for stable expression of short interfering RNAs in Mammalian cells. Science. 2002; 296 :550-553.
[58] Brunner A, Chinn J, Neubauer M. Identification of a gene family regulated by transforming growth factor beta. DNA Cell Biol 1991; 10:293-300.
[59] Monaco JL, Lawrence WT. Acute wound healing An overview. Clin Plastic Surg 30(2003) 1-12.
[60] Longaker MT, Chiu E, Adzick NS, et al. Studies in fetal wound healing. V. A prolonged presence of hyaluronic acid characterizes fetal wound fluid. Ann Surg. 1991;213:292-6.
[61] Estes JM, Adzick NS, Harrison MR, et al. Hyaluronate metabolism undergoes an ontogenic transition during fetal development: implications for scar-free wound healing. J Pediatr Surg. 1993;28:1227—31.
[62] Longaker MT, Chiu ES, Harrison MR, et al. Studies in fetal wound healing. IV. Hyaluronic acid-stimulating activity distinguishes fetal wound fluid from adult wound fluid. Ann Surg. 1989;210:667-72.
[63] Alaish SM, Yager D, Diegelmann RF, et al. Biologyof fetal wound healing: hyaluronate receptor expression in fetal fibroblasts. J Pediatr Surg. 1994;29:1040-3.
[64] Grotendorst GR, Okochi H, Hayashi N. A novel transforming growth factor beta response element controls the expression of the connective tissur growth factor gene. Cell Growth Differ 1996;7:469-80.