Fibrin deposition in tissues from endotoxin-treated mice correlates with decreases in the expression of urokinase-type but not tissue-type plasminogen activator.

Abstract

The primary hypothesis of this report is that the formation and subsequent removal of fibrin in specific tissues during pathologic processes reflects temporal changes in the local expression of key procoagulant and fibrinolytic genes. To begin to test this hypothesis, we have used quantitative PCR assays and in situ hybridization analysis to examine the effects of endotoxin on the expression of specific genes in murine tissues, and to relate these changes to fibrin deposition/ dissolution using immunohistochemical approaches. Endotoxin caused large increases in plasminogen activator inhibitor-1 mRNA and modest increases in tissue factor mRNA in most tissues examined. However, fibrin was only detected in the kidneys and adrenals of endotoxin-treated mice, and it was transient. Unexpectedly, changes in urokinase-type plasminogen activator mRNA but not tissue-type plasminogen activator mRNA correlated with fibrin deposition/dissolution in these tissues. Pretreatment of mice with the fibrinolytic inhibitor epsilon-aminocaproic acid before endotoxin increased both the number of fibrin-positive tissues and the duration of fibrin deposition in the kidneys and adrenals. These results suggest that the absence of fibrin in some tissues reflects ongoing local fibrinolysis, and that increases in plasminogen activator inhibitor-1 and tissue factor gene expression and decreases in urokinase-type plasminogen activator expression are necessary for tissue-specific fibrin deposition. Changes in tissue-type plasminogen activator gene expression do not appear to be essential for fibrin deposition/dissolution in this murine model of sepsis. ( J. Clin. Invest. 1996. 97:2440-2451.)

60 The kidneys of mice with autoimmune disease acquire a hypofibrinolytic/procoagulant state that correlates with the development of glomerulonephritis and tissue microthrombosis

“…13 Endotoxin also caused a significant reduction in u-PA gene expression in the kidneys and adrenals but did not alter u-PA in other tissues and did not decrease t-PA gene expression in most tissues examined. These initial studies led to the hypothesis that, in sepsis, increases in PAI-1 and TF are necessary but not sufficient for tissue-specific fibrin deposition and that u-PA must also decrease.…”

“…These initial studies led to the hypothesis that, in sepsis, increases in PAI-1 and TF are necessary but not sufficient for tissue-specific fibrin deposition and that u-PA must also decrease. 13 In this report, we begin to investigate the generality of the above hypothesis using mice that develop autoimmune disease. SLE is an autoimmune disorder also associated with renal thrombosis and nephropathy.1 8 Several recent reports demonstrate an increase in plasma PAI-1 levels in SLE patients17 19 and an induction in renal PAI-1 mRNA in experimental glomerulonephritis in rats33 and mice.…”

“…Thus, the changes in the plasminogen activators in kidneys of lupus-prone mice also resemble the changes observed in kidneys of a septic model. 13 The resulting changes in the balance between the plasminogen activators and PAI-1, their primary inhibitor, should result in "fibrinolytic shut-down", a process that will favor the accumulation of fibrin and extracellular matrix in nephritic glomeruli. u-PA is synthesized by renal tubular epithelial cells,12 and its pattern of expression in Figure 3 suggests that its synthesis by these cells is decreased.…”

Expression of Transforming Growth Factor-β and Tumor Necrosis Factor-α in the Plasma and Tissues of Mice with Lupus Nephritis

“…A quantitative RT-PCR approach was employed to determine the concentration of specific mRNAs in murine tissues (Platzer et al, 1992;Yamamoto and Loskutoff, 1996). Competitive RT-PCR was performed using 10 5 molecules of competitor RNA (cRNA) for TGF-␤, TNF-␣, and PAI-1, and 10 7 molecules of cRNA for ␤-actin, all in the presence of 1 g total tissue RNA and 5 ϫ 10 5 cpm of 32 P-labeled oligonucleotide as described (Samad et al, 1996;Yamamoto and Loskutoff, 1996).…”

“…Competitive RT-PCR was performed using 10 5 molecules of competitor RNA (cRNA) for TGF-␤, TNF-␣, and PAI-1, and 10 7 molecules of cRNA for ␤-actin, all in the presence of 1 g total tissue RNA and 5 ϫ 10 5 cpm of 32 P-labeled oligonucleotide as described (Samad et al, 1996;Yamamoto and Loskutoff, 1996). Aliquots (20 l) of the PCR products were electrophoresed through 2% to 2.5% agarose gels containing ethidium bromide.…”