application) The paucity of therapeutic options for the treatment of chronic renal diseases results in the inexorable progression to end stage renal disease, characterized by interstitial fibrosis and glomerulosiderosis. In both processes, the myofibroblast has a central role as an effector cell elaborating excessive inflammatory mediators and extracellular matrix. In interstitial fibrosis, the degree of fibrosis and progression of renal disease directly correlates with the presence of myofibroblasts. Interstitial myofibroblasts arise mainly from tubular epithelial cells and fibroblasts. Factors implicated in the transformation include transforming growth factor (TGF)- beta1, endothelin- 1, and angiotensin ll- profibrogenic growth factors shown to stimulate gelatinase A production. In the glomerulus, gelatinase A transforms the mesangial cell into myofibroblasts. Similarly, gelatinase A is a key mediator of tubular epithelial cell- myofibroblast transdifferentiation in vitro. To locate the key regulatory elements for constitutive and growth factor- inducible gelatinase A transcription in the renal interstitium, a series of 5' flanking region deletion constructs of the rat gelatinase A gene will be transfected into normal rat kidney epithelial (NRK- 52e) and fibroblast (NRK- 49f) cells. The constructs extend to - 1686 bp of the immediate 5' flanking region and contain a laciferase reporter. Once the specific constitutive and growth factor- dependent enhancer sequences have been mapped, the specific interacting transcription factors will be characterized by the yeast one- hybrid system. A transgenic mouse model constitutively expressing activated gelatinase A will be constructed to directly test the relevance of gelatinase A on epithelial cell transdifferentiation to myofibroblasts and generation of interstitial fibrosis. To avoid a lethal phenotype, a tissue- specific, tetracycline- inducible transgenic system will be utilized. Using the mouse gamma- glutamyl transpeptidase II promoter, the tetracycline transactivator protein (tTA) will be restricted to the proximal tubules. The tTA mouse will be crossed with a mouse harboring the gelatinase A gene under the control of a synthetic tetracycline operator (tet0). Only those cells with tTA (i.e. the proximal tubules) will be able to express activated gelatinase A. tTA, and consequently gelatinase A expression, is inhibited by doxycycline, so temporal control over activated gelatinase A expression will be maintained by doxycycline administration. To further evaluate gelatinase A's role in interstitial fibrosis, a gelatinase A knockout strain will be crossed with a TGF- beta1 transgenic strain. Since the TGF- beta1 transgenic mouse develops fibrosis, this crossing will evaluate gelatinase A as distal effector for TGF- beta1- induced fibrosis.
| Lovett, David H; Cheng, Sunfa; Cape, Leslie et al. (2010) YB-1 alters MT1-MMP trafficking and stimulates MCF-7 breast tumor invasion and metastasis. Biochem Biophys Res Commun 398:482-8 |
| Cheng, Sunfa; Pollock, Allan S; Mahimkar, Rajeev et al. (2006) Matrix metalloproteinase 2 and basement membrane integrity: a unifying mechanism for progressive renal injury. FASEB J 20:1898-900 |
