Fibroblast response to hypoxia: the relationship between angiogenesis and matrix regulation.

A number of studies have demonstrated the critical role of angiogenesis for successful wound repair in the surgical patient. Vascular disruption from tissue injury due to trauma or surgery leads to a hypoxic zone in the healing wound. In this dynamic process, angiogenesis is vital for the delivery of oxygen, nutrients, and growth factors necessary to initiate the synthetic processes of wound healing. Fibroblasts, invading the wound early in the healing process, are involved in extracellular matrix (ECM) deposition as well as wound contraction. However, the exact mechanisms by which important genes are regulated remain unknown. In order to examine these processes, we studied the effects of hypoxia on fibroblasts for the expression of VEGF, type IalphaI collagen, and matrix-metalloproteinase-3, three genes essential for the regulation of angiogenesis, ECM deposition, and ECM degradation in wound healing. Primary cell cultures of normal human dermal fibroblasts (NHDFs) were placed in hypoxia for varying periods of time. Northern blot hybridization was performed with [alpha32P]dCTP-labeled cDNA probes for VEGF, type IalphaI collagen, and MMP-3. The results demonstrated a time-dependent VEGF mRNA upregulation (470% of baseline) under hypoxia. Type IalphaI collagen increased (170% of baseline) at 24 h, but was then abruptly downregulated to 3.8% of baseline at 48 h. MMP-3 was incrementally downregulated to 2.2% of baseline at 48 h. These experiments focused on the effect of hypoxia on genes thought to play a role in wound repair. VEGF upregulation in the hypoxic microenvironment of the early wound may serve to stimulate angiogenesis. Type IalphaI collagen, though upregulated early on, was abruptly downregulated at 48 h. This downregulation may reflect the in vivo requirement for angiogenesis to deliver oxygen for successful hydroxylation and collagen synthesis in the wound. MMP-3, also downregulated at 48 h, may also implicate the need for angiogenesis. These data support the theory that hypoxia-driven angiogenesis is critical for ECM formation and remodeling in successful soft tissue repair. Furthermore, they may represent the role of hypoxia as an important regulator to efficiently balance these complex processes in the healing wound.

[1]  Richard A.F. Clark,et al.  The Molecular and Cellular Biology of Wound Repair , 2012, Springer US.

[2]  J. McCarthy,et al.  Rat mandibular distraction osteogenesis: II. Molecular analysis of transforming growth factor beta-1 and osteocalcin gene expression. , 1999, Plastic and reconstructive surgery.

[3]  K. Alitalo,et al.  Comparison of VEGF, VEGF-B, VEGF-C and Ang-1 mRNA regulation by serum, growth factors, oncoproteins and hypoxia , 1997, Oncogene.

[4]  H. Dvorak,et al.  Hypoxia regulates the expression of vascular permeability factor/vascular endothelial growth factor (VPF/VEGF) and its receptors in human skin. , 1997, The Journal of investigative dermatology.

[5]  D. Wakefield,et al.  Increased expression of matrix metalloproteinases in vivo in scleritis tissue and in vitro in cultured human scleral fibroblasts. , 1997, The American journal of pathology.

[6]  A. Kurtz,et al.  Induction of VEGF and VEGF receptor gene expression by hypoxia: divergent regulation in vivo and in vitro. , 1997, Kidney international.

[7]  K. Thomas.  Vascular Endothelial Growth Factor, a Potent and Selective Angiogenic Agent (*) , 1996, The Journal of Biological Chemistry.

[8]  Atsushi Namiki,et al.  Hypoxia Induces Vascular Endothelial Growth Factor in Cultured Human Endothelial Cells (*) , 1995, The Journal of Biological Chemistry.

[9]  M. Neeman,et al.  Stabilization of vascular endothelial growth factor mRNA by hypoxia and hypoglycemia and coregulation with other ischemia-induced genes , 1995, Molecular and cellular biology.

[10]  H. Federoff,et al.  Expression of vascular endothelial growth factor from a defective herpes simplex virus type 1 amplicon vector induces angiogenesis in mice. , 1995, Circulation research.

[11]  J. Caro,et al.  Hypoxic stimulation of vascular endothelial growth factor expression in vitro and in vivo. , 1994, Laboratory investigation; a journal of technical methods and pathology.

[12]  H. Birkedal‐Hansen,et al.  Distinct populations of basal keratinocytes express stromelysin-1 and stromelysin-2 in chronic wounds. , 1994, The Journal of clinical investigation.

[13]  M. Goldberg,et al.  Similarities between the oxygen-sensing mechanisms regulating the expression of vascular endothelial growth factor and erythropoietin. , 1994, The Journal of biological chemistry.

[14]  R. Kirsner,et al.  Low oxygen tension increases mRNA levels of alpha 1 (I) procollagen in human dermal fibroblasts , 1993, Journal of cellular physiology.

[15]  V. Falanga,et al.  Growth factors and wound healing. , 1993, Dermatologic clinics.

[16]  M. Matsubara,et al.  Stromal fibroblasts synthesize collagenase and stromelysin during long-term tissue remodeling. , 1993, Journal of cell science.

[17]  G. Semenza,et al.  A nuclear factor induced by hypoxia via de novo protein synthesis binds to the human erythropoietin gene enhancer at a site required for transcriptional activation , 1992, Molecular and cellular biology.

[18]  K. Sueishi,et al.  Media conditioned by smooth muscle cells cultured in a variety of hypoxic environments stimulates in vitro angiogenesis. A relationship to transforming growth factor-beta 1. , 1992, The American journal of pathology.

[19]  H. Dvorak,et al.  Expression of vascular permeability factor (vascular endothelial growth factor) by epidermal keratinocytes during wound healing , 1992, The Journal of experimental medicine.

[20]  J. Uitto,et al.  Leukoregulin, A T‐cell derived cytokine, upregulates stromelysin‐1 gene expression in human dermal fibroblasts: Evidence for the role of AP‐1 in transcriptional activiation , 1992, Journal of cellular biochemistry.

[21]  M. Diaz-Meco,et al.  Protein kinase C-independent expression of stromelysin by platelet-derived growth factor, ras oncogene, and phosphatidylcholine-hydrolyzing phospholipase C. , 1991, The Journal of biological chemistry.

[22]  T. K. Hunt,et al.  Tissue oxygenation, anemia, and perfusion in relation to wound healing in surgical patients. , 1991, Annals of surgery.

[23]  D. Knighton,et al.  Regulation of cutaneous wound healing by growth factors and the microenvironment. , 1991, Investigative radiology.

[24]  J. Montani,et al.  Growth regulation of the vascular system: evidence for a metabolic hypothesis. , 1990, The American journal of physiology.

[25]  D. Connolly,et al.  Vascular permeability factor, an endothelial cell mitogen related to PDGF. , 1989, Science.

[26]  D. Goeddel,et al.  Vascular endothelial growth factor is a secreted angiogenic mitogen. , 1989, Science.

[27]  D. Connolly,et al.  Tumor vascular permeability factor stimulates endothelial cell growth and angiogenesis. , 1989, The Journal of clinical investigation.

[28]  D. Rappolee,et al.  Genes for extracellular-matrix-degrading metalloproteinases and their inhibitor, TIMP, are expressed during early mammalian development. , 1989, Genes & development.

[29]  Y. Otani,et al.  Transcriptional regulation of human stromelysin. , 1989, The Journal of biological chemistry.

[30]  W. Eaglstein,et al.  Dermal pericapillary fibrin in venous disease and venous ulceration. , 1987, Archives of dermatology.

[31]  W. Swartz,et al.  The causes of skin ulcerations associated with venous insufficiency: a unifying hypothesis. , 1987, Plastic and reconstructive surgery.

[32]  T. K. Hunt,et al.  Regulation of wound-healing angiogenesis-effect of oxygen gradients and inspired oxygen concentration. , 1981, Surgery.

[33]  K. Kivirikko,et al.  [Biosynthesis of collagen and its disorders]. , 1979, Duodecim; laaketieteellinen aikakauskirja.

[34]  T. K. Hunt,et al.  The effect of varying ambient oxygen tensions on wound metabolism and collagen synthesis. , 1972, Surgery, gynecology & obstetrics.

[35]  S. Udenfriend.  Formation of Hydroxyproline in Collagen , 1966, Science.

[36]  C. Brinckerhoff,et al.  Post-transcriptional regulation of collagenase and stromelysin gene expression by epidermal growth factor and dexamethasone in cultured human fibroblasts. , 1992, Journal of cellular biochemistry.

[37]  L. Matrisian,et al.  Metalloproteinases and their inhibitors in matrix remodeling. , 1990, Trends in genetics : TIG.

[38]  H. Tk,et al.  Prospective: a retrospective perspective on the nature of wounds. , 1988, Progress in clinical and biological research.

[39]  D. Rifkin,et al.  Proteinases and Tissue Remodeling , 1988 .

[40]  M. Chvapil,et al.  The influence of various oxygen tensions upon proline hydroxylation and the metabolism of collagenous and non-collagenous proteins in skin slices. , 1968, Hoppe-Seyler's Zeitschrift fur physiologische Chemie.