title: MnTE-2-PyP Treatment, or NOX4 Inhibition, Protects against Radiation-Induced Damage in Mouse Primary Prostate Fibroblasts by Inhibiting the TGF-Beta 1 Signaling Pathway.
abstract: Prostate cancer patients who undergo radiotherapy frequently suffer from side effects caused by radiation-induced damage to normal tissues adjacent to the tumor. Exposure of these normal cells during radiation treatment can result in tissue fibrosis and cellular senescence, which ultimately leads to postirradiation-related chronic complications including urinary urgency and frequency, erectile dysfunction, urethral stricture and incontinence. Radiation-induced reactive oxygen species (ROS) have been reported as the most potent causative factor for radiation damage to normal tissue. While MnTE-2-PyP, a ROS scavenger, protects normal cells from radiation-induced damage, it does not protect cancer cells during radiation treatment. However, the mechanism by which MnTE-2-PyP provides protection from radiation-induced fibrosis has been unclear. Our current study reveals the underlying molecular mechanism of radiation protection by MnTE-2-PyP in normal mouse prostate fibroblast cells. To investigate the role of MnTE-2-PyP in normal tissue protection after irradiation, primary prostate fibroblasts from C57BL/6 mice were cultured in the presence or absence of MnTE-2-PyP and exposed to 2 Gy of X rays. We found that MnTE-2-PyP could protect primary prostate fibroblasts from radiation-induced activation, as measured by the contraction of collagen discs, and senescence, detected by beta-galactosidase staining. We observed that MnTE-2-PyP inhibited the TGF-β-mediated fibroblast activation pathway by downregulating the expression of TGF-β receptor 2, which in turn reduced the activation and/or expression of SMAD2, SMAD3 and SMAD4. As a result, SMAD2/3-mediated transcription of profibrotic markers was reduced by MnTE-2-PyP. Due to the inhibition of the TGF-β pathway, fibroblasts treated with MnTE-2-PyP could resist radiation-induced activation and senescence. NADPH oxidase 4 (NOX4) expression is upregulated after irradiation and produces ROS. As was observed with MnTE-2-PyP treatment, NOX4-/-fibroblasts were protected from radiation-induced fibroblast activation and senescence. However, NOX4-/-fibroblasts had reduced levels of active TGF-β1, which resulted in decreased TGF-β signaling. Therefore, our data suggest that reduction of ROS levels, either by MnTE-2-PyP treatment or by eliminating NOX4 activity, significantly protects normal prostate tissues from radiation-induced tissue damage, but that these approaches work on different components of the TGF-β signaling pathway. This study proposes a crucial insight into the molecular mechanism executed by MnTE-2-PyP when utilized as a radioprotector. An understanding of how this molecule works as a radioprotector will lead to a better controlled mode of treatment for post therapy complications in prostate cancer patients.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska 68198.
authors.forename: Arpita
authors.initials: A
authors.lastname: Chatterjee
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska 68198.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Kosmacek
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska 68198.
authors.forename: Rebecca E
authors.initials: RE
authors.lastname: Oberley-Deegan
mesh_terms: D000818:Animals; D048429:Cell Size / Q000187:drug effects / Q000528:radiation effects; D016922:Cellular Senescence / Q000187:drug effects / Q000528:radiation effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005110:Extracellular Space / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D005786:Gene Expression Regulation / Q000187:drug effects / Q000528:radiation effects; D008297:Male; D008665:Metalloporphyrins / Q000494:pharmacology*; D051379:Mice; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors*; D011467:Prostate / Q000166:cytology*; D011837:Radiation-Protective Agents / Q000494:pharmacology; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects* / Q000528:radiation effects*; D013481:Superoxides / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D008665:Metalloporphyrins; D011837:Radiation-Protective Agents; D018125:Receptors, Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C520506:manganese tetrakis-(N-ethyl-2 pyridyl) porphyrin; D013481:Superoxides; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C422943:Nox4 protein, mouse