Radiation and Transforming Growth Factor-β Cooperate in Transcriptional Activation of the Profibrotic Plasminogen Activator Inhibitor-1 Gene

Abstract

Radiation-induced fibrosis is an important side effect in the treatment of cancer. Profibrotic proteins, such as plasminogen activator inhibitor-1 (PAI-1), transforming growth factor-h (TGF-h), and tissue type inhibitor of metalloproteinases-1 (Timp-1), are thought to play major roles in the development of fibrosis via the modulation of extracellular matrix integrity.We did a detailed analysis of transcriptional activation of these profibrotic genes by radiation and TGF-h. Irradiation of HepG2 cells led to a high increase in PAI-1mRNA levels and a mild increase in Timp-1mRNA levels. In contrast,TGF-h1and Smad7 were not increased. Radiation and TGF-h showed strong cooperative effects in transcription of the PAI-1 gene. The TGF-b1 gene showed a mild cooperative activation, whereas Timp-1and Smad7 were not cooperatively activated by radiation and TGF-h. Analysis using the proximal 800 bp of the human PAI-1 promoter revealed a dose-dependent increase of PAI-1levels between 2 and 32 Gy g-rays that was independent of latent TGF-h activation. Subsequent site-directed mutagenesis of the PAI-1 promoter revealed that mutation of a p53-binding element abolished radiation-induced PAI-1 transcription. In line with this, PAI-1 was not activated in p53-null Hep3B cells, indicating that p53 underlies the radiation-induced PAI-1activation and the cooperativity with theTGF-h/Smad pathway. Together, these data show that radiation and TGF-h activate PAI-1 via partially nonoverlapping signaling cascades that in concert synergize on PAI-1 transcription. This may play a role in patient-to-patient variations in susceptibility toward fibrosis after radiotherapy.

Plasminogen Activator Inhibitor-1 Controls Bone Marrow-Derived Cells Therapeutic Effect Through MMP9 Signaling: Role in Physiological and Pathological Wound Healing

“…PAI-1 expression is induced rapidly in the endothelium in irradiated skin. Moreover, ionizing radiation was shown to increase PAI-1 expression in NRK52E [30] and HEPG2 cells [31]. Additionally, PAI-1 over-expression is observed in radiation-induced nephrosclerosis [32] and in human radiation enteritis and intestinal lesion [33,34] underlining a role for PAI-1 in radiation-induced tissue damages.…”

Essential Role of Plasminogen Activator Inhibitor Type-1 in Radiation Enteropathy

“…16 Moreover, PAI-1 overexpression has been described in radiation-induced nephrosclerosis 17 and in human radiation enteritis, 18 suggesting a role of PAI-1 in radiation-induced normal tissue damage. We have clearly demonstrated that PAI-1 plays a crucial role in radiation-induced intestinal lesions following a high single dose of radiation.…”

“…PAI-1 was described as a direct target of p53 32 in tumor cells and the mutation of the p53 binding element in the PAI-1 promoter abolished the radiation-induced PAI-1 transcription in HEPG2 cells. 16 Radiation and TGF-␤ cooperate in the radiation-induced PAI-1 transcription, suggesting that a p53/Smad pathway is involved in this mechanism. Moreover, a specific cooperation and physical interaction of Smads with p53 play a key role in embryogenesis.…”

Computational analysis of the human HSPH/HSPA/DNAJ family and cloning of a human HSPH/HSPA/DNAJ expression library

“…Subsequently, the coding sequence of the different chaperones was amplified using the primers listed in supplemental Table 8. As a template source, complementary DNA (cDNA) was made from total RNA as previously described (Hageman et al 2005). As a source of total RNA, QPCR Human reference Total RNA (Stratagene) was used, which is a mixed source of RNA from the following cell line derivations: adenocarcinoma, mammary gland; hepatoblastoma, liver; adenocarcinoma, cervix; embryonal carcinoma, testis; glioblastoma, brain; melanoma, skin; liposarcoma, histiocytic lymphoma, macrophage, histocyte; lymphoblastic leukemia, T lymphoblast; plasmacytoma, myeloma, B lymphocyte.…”