A Plasmid vector encoding functional human keratinocyte growth factor gene in vitro—Functional human KGF gene expression in vitro — Oak Academic Publishing
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A Plasmid vector encoding functional human keratinocyte growth factor gene in vitro—Functional human KGF gene expression in vitro
Laboratory of Surgery, Children’s Hospital of Chongqing Medical University, Chongqing, China
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Laboratory of Surgery, Children’s Hospital of Chongqing Medical University, Chongqing, China;
1 Laboratory of Surgery, Children’s Hospital of Chongqing Medical University, Chongqing, China
2 Laboratory of Surgery, Children’s Hospital of Chongqing Medical University, Chongqing, China;
In this study, we cloned human KGF (hKGF) genes using RT-PCR techniques and developed a eukaryotic expression plasmid vector capable of directing the expression of functional hKGF. Monolayer culture of human embryo lung fibro-blast (HLF) was used for isolation of total RNA. Then the total RNA was purified and reverse- transcribed into cDNA using an oligo (dT) primer. A full PCR fragment for hKGF was generated and cloned. Restriction digestion and nucleo-tide sequence analysis validated the complete hKGF transcription. The hKGF cDNA fragment was inserted into pEGFP-C2 vector by means of recombinant DNA technology and verified by restriction analysis and sequencing. We have constructed pEGFP-C2-hKGF encoding the green fluorescent protein (GFP). Furthermore, hKGF had the effect on AEC II proliferation. These results suggest that the potential appli-cation of a hKGF plasmid of gene expression should be useful for sustained AEC proliferation, and its in vivo efficacy needs to be validated. Keywords:
Haddad, I. Y., Milla, C., Yang, S., Panoskaltsis-Mortari, A., Hawgood, S., Lacey, D. L. and Blazar, B. R. (2003) Surfactant protein A is a required mediator of keratinocyte growth factor after experimental marrow transplantation. American Journal of Physiology. Lung Cell Molecular Physiology, 285, L602-L610.
Yi, E. S., Williams, S. T., Lee, H., Malicki, D. M., Chin, E. M, Yin, S., Tarpley, J. and Ulich, T. R. (1996) Kerati-nocyte growth factor ameliorates radiation- and bleomycin- induced lung injury and mortality. American Journal of Pathology, 149, 1963-1970.
Keijzer, R., van Tuyl, M., Meijers, C., Post, M., Tibboel, D., Grosveld, F. and Koutsourakis, M. (2001) The tran-scription factor GATA6 is essential for branching morpho-genesis and epithelial cell differentiation during fetal pul-monary development. Development, 128, 503-511.
Ware, L. B. and Matthay, M. A. (2002) Keratinocyte and hepatocyte growth factors in the lung: Roles in lung de-velopment, inflammation, and repair. American Journal of Physiology. Lung Cell Molecular Physiology, 282, L924-L940.
Ray, P., Devaux, Y., Stolz, D. B., Yarlagadda, M., Watkins, S. C., Lu, Y., Chen, L., Yang, X. F. and Ray, A. (2003) Inducible expression of keratinocyte growth factor (KGF) in mice inhibits lung epithelial cell death induced by hyperoxia. Proceedings of the National Academy of Science, 100, 6098-6103.
Atabai, K., Ishigaki, M., Geiser, T., Ueki, I., Matthay, M. A. and Ware, L. B. (2002) Keratinocyte growth factor can enhance alveolar epithelial repair by nonmitogenic mechanisms. American Journal of Physiology. Lung Cell Molecular Physiology, 283, L163-L169.
Hohlfeld, J. M., Hoymann, H. G.., Tschernig, T., Fehren-bach, A., Krug, N. and Fehrenbach, H. (2004) Keratino-cyte growth factor transiently alters pulmonary function in rats. Journal of Applied Physiology, 96, 704-710..
Buckley, S., Barsky, L., Driscoll, B., Weinberg, K., An-derson, K. D. and Warburton, D. (1998) Apoptosis and DNA damage in type 2 alveolar epithelial cells cultured from hyperoxic rats. American Journal of Physiology. Lung Cell Molecular Physiology, 274, L714-L720.
Senaldi, G., Shaklee, C. L., Simon, B., Rowan, C. G., Lacey, D. L. and Hartung, T. (1998) Keratinocyte growth factor protects murine hepatocytes from tumor necrosis factor-induced apoptosis in vivo and in vitro. Hepatology, 27, 1584-1591.
Portnoy, J., Curran-Everett, D., Mason, R. J. (2004) Kera-tinocyte growth factor stimulates alveolar type II cell pro-liferation through the extracellular signal-regulated kinase and phosphatidylinositol 3-OH kinase pathways. American Journal of Respiratory Cell and Molecular Biology, 30, 901-907.
Fehrenbach, A., Bube, C., Hohlfeld, J. M., Stevens, P., Tschernig, T., Hoymann, H. G., Krug, N. and Fehrenbach, H. (2003) Surfactant homeostasis is maintained in vivo during keratinocyte growth factor-induced rat lung type II cell hyperplasia. American Journal of Respiratory and Critical Care Medicine, 167, 1264-1270..
Chedid, M., Rubin, J. S., Csaky, K. G. and Aaronson, S. A. (1994) Regulation of keratinocyte growth factor gene expression by interleukin 1. Journal of Biological Che-mistry, 269, 10753-10757.
Simonet, W. S., DeRose, M. L., Bucay, N., Nguyen, H. Q., Wert, S. E., Zhou, L., Ulich, T. R., Thomason, A., Danilenko, D. M. and Whitsett, J. A. (1995) Pulmonary malformation in transgenic mice expressing human kera-tinocyte growth factor in the lung. Proceedings of the National Academy of Science, 92, 12461-12465.
Spilde, T. L., Bhatia, A. M., Marosky, J. K., Preuett, B., Kobayashi, H., Hembree, M. J., Prasadan, K., Daume, E., Snyder, C. L. and Gittes, G. K. (2003) Fibroblast growth factor signaling in the developing tracheoesophageal fis-tula. Journal of Pediatric Surgery, 38, 474-477..
Fu, Y. G., Qu, Y. J., Wu, K. C., Zhai, H. H., Liu, Z. G. and Fan, D.M. (2003) Apoptosis-inducing effect of recombinant Caspase-3 expressed by constructed eukaryotic vector on gastric cancer cell line SGC7901. World Journal of Gastroenterology, 9, 1935-1939.
Morikawa, O., Walker, T. A., Nielsen, L. D., Pan, T., Cook, J. L. and Mason, R. J. (2000) Effect of adenovec-tor-mediated gene transfer of keratinocyte growth factor on the proliferation of alveolar type II cells in vitro and in vivo. American Journal of Respiratory Cell and Mole-cular Biology, 23, 626-635.
Paine, R. 3rd., Rolfe, M. W., Standiford, T. J., Burdick, M. D., Rollins, B. J. and Strieter, R. M. (1993) MCP-1 expression by rat type II alveolar epithelial cells in primary culture. Journal of Immunology, 150, 4561-4570.
Li, C., Zhu, N. L., Tan, R. C., Ballard, P. L., Derynck, R. and Minoo, P. (2002) Transforming growth factor-beta inhibits pulmonary surfactant protein B gene transcription through SMAD3 interactions with NKX2.1 and HNF-3 transcription factors. Journal of Biological Chemistry, 277, 38399-38408..
Beers, M.F., Solarin, K.O., Guttentag, S.H., Rosenbloom, J., Kormilli, A., Gonzales, L.W. and Ballard, P. L. (1998) TGF-beta1 inhibits surfactant component expression and epithelial cell maturation in cultured human fetal lung. American Journal of Physiology. Lung Cell Molecular Physiology, 275, L950-L960.
Zhao, Y., Young, S. L., McIntosh, J. C., Steele, M. P. and Silbajoris, R. (2000) Ontogeny and localization of TGF- beta type I receptor expression during lung development. American Journal of Physiology. Lung Cell Molecular Physiology, 278, L1231-L1239.
Mason, R. J., Lewis, M. C., Edeen, K. E., McCormick- Shannon, K., Nielsen, L. D. and Shannon, J. M. (2002) Maintenance of surfactant protein A and D secretion by rat alveolar type II cells in vitro. American Journal of Physiology. Lung Cell Molecular Physiology, 282, L249- L258.
Xu, X., McCormick-Shannon, K., Voelker, D. R. and-Mason, R. J. (1998) KGF increases SP-A and SP-D mRNA levels and secretion in cultured rat alveolar type II cells. American Journal of Respiratory Cell and Mole-cular Biology, 18, 168-178.
Chelly, N., Mouhieddine-Gueddiche, O. B., Barlier-Mur, A. M., Chailley-Heu, B., Bourbon, J. R. (1999) Kerati-nocyte growth factor enhances maturation of fetal rat lung type II cells. American Journal of Respiratory Cell and Molecular Biology, 20, 423-432.
Xing, Z., Braciak, T., Jordana, M., Croitoru, K., Graham, F. L. and Gauldie, J. (1994) Adenovirus-mediated cyto-kine gene transfer at tissue sites. Overexpression of IL-6 induces lymphocytic hyperplasia in the lung. Journal of Immunology, 153, 4059-4069.
Yei, S., Mittereder, N., Wert, S., Whitsett, J. A., Wilmott, R. W. and Trapnell, B. C. (1994) In vivo evaluation of the safety of adenovirus-mediated transfer of the human cystic fibrosis transmembrane conductance regulator cDNA to the lung. Human Gene Therapy, 5, 731-744.
Roy, S., Gao, G., Lu, Y., Zhou, X., Lock, M., Calcedo, R. and Wilson, J. M. (2004) Characterization of a family of chimpanzee adenoviruses and development of molecular clones for gene transfer vectors. Human Gene Therapy, 15, 519-530.
Ebelt, H. and Braun, T. (2003) Optimized, highly efficient transfer of foreign genes into newborn mouse hearts in vivo. Biochemical and Biophysical Research Commu-nications, 310, 1111-1116.
Bosma, P. T., van Eert, S. J., Jaspers, N. G., Stoter, G. and Nooter, K. (2003) Functional cloning of drug resistance genes from retroviral cDNA libraries. Biochemical and Biophysical Research Communications, 309, 605-611.
Yan, J., Liang, S. H., Mao, Y. F., Li, L. W., Li, S. P. (2003) Construction of expression systems for flaA and flaB genes of Helicobacter pylori and determination of im-munoreactivity and antigenicity of recombinant proteins. World Journal of Gastroenterology, 9, 2240-2250.
Ebensen, T., Paukner, S., Link, C., Kudela, P., de Domenico, C., Lubitz, W. and Guzman, C. A. (2004) Bac-terial ghosts are an efficient delivery system for DNA vac-cines. Journal of Immunology, 172, 6858-6865.
Piao, Y. F., Tang, T. Y., Niu, J. Q. and Wang, F. (2004) Construction of eukaryotic expression plasmids of hepa-titis B surface antigen and helper T lymphocyte epitope. Hepatobiliary & Pancreatic Diseases International Jour-nal, 3, 219-222.
Schuck, S., Manninen, A., Honsho, M., Fullekrug, J. and Simons, K. (2004) Generation of single and double knockdowns in polarized epithelial cells by retrovirus-mediated RNA interference. Proceedings of the National Academy of Science, 101, 4912-4917..
Belin, D., Guzman, L.M., Bost, S., Konakova, M., Silva, F. and Beckwith, J. (2004) Functional activity of euka-ryotic signal sequences in Escherichia coli: The ovalbumin family of serine protease inhibitors. Journal of Biological Chemistry, 335, 437-453.
Baldwin, S. L., Powell, T. D., Wonderling, R. S., Keiser, K. C., Morales, T., Hunter, S., McDermott, M., Radecki, S. V. and Milhausen, M. J. (2003) Transient and stable transfection of Chinese hamster ovary cells with the re-combinant feline erythropoietin gene and expression, pu-rification, and biological activity of feline erythropoietin protein. American Journal of Veterinary Research, 64, 1465-1471.
Finch, P. W., Rubin, J. S., Miki, T., Ron, D. and Aaronson, S. A. (1989) Human KGF is FGF-related with properties of a paracrine effector of epithelial cell growth. Science, 245, 752-755.
Fujio, Y. and Walsh, K. (1999) Akt mediates cytoprotec-tion of endothelial cells by vascular endothelial growth factor in an anchorage-dependent manner. Journal of Bio-logical Chemistry, 274, 16349-16354.
Kauffmann-Zeh, A., Rodriguez-Viciana, P., Ulrich, E., Gilbert, C., Coffer, P., Downward, J. and Evan, G. (1997) Suppression of c-Myc-induced apoptosis by Ras signal-ling through PI(3)K and PKB. Nature, 385, 544-548..
Lu, Y., Parkyn, L., Otterbein, L. E., Kureishi, Y., Walsh, K., Ray, A. and Ray, P. (2001) Activated Akt protects the lung from oxidant-induced injury and delays death of mice. Journal of Experimental Medicine, 193, 545-549.