Background Radiation-induced pulmonary fibrosis (RIPF) is a late toxicity of therapeutic radiation. mTOR signaling drives several processes implicated in RIPF, including inflammatory cytokine production, fibroblast proliferation, and epithelial senescence. We sought to determine if mTOR inhibition with rapamycin would mitigate RIPF. Methods/Materials C57BL/6NCr mice received a diet formulated with rapamycin (14 mg/kg food) or control diet two days before and continuing for 16 weeks after exposure to 5 daily fractions of 6 Gy thoracic irradiation (IR). Fibrosis was assessed with Masson-Trichrome staining and hydroxyproline assay. Cytokine expression was evaluated by quantitative real time PCR. Senescence was assessed by staining for beta-galactosidase activity. Results Administration of rapamycin extended the median survival of irradiated mice compared to control diet from 116 days to 156 days (log rank p=0.006). Treatment with rapamycin reduced hydroxyproline content compared to control diet (IR+vehicle: 45.9±11.8, IR+rapamycin: 21.4±6.0, p=0.001) and reduced visible fibrotic foci. Rapamycin treatment attenuated IL-1β and TGF-β induction in irradiated lung compared to control diet. Type II pneumocyte senescence after IR was reduced with rapamycin treatment at 16 weeks (three-fold reduction at 16 weeks, p<0.001). Conclusion Rapamycin protected against RIPF in a murine model. Rapamycin treatment reduced inflammatory cytokine expression, extra cellular matrix production, and senescence in type II pneumocytes.
Purpose/Objectives Fibrosis is a late toxicity of thoracic irradiation that can result in substantial morbidity. Plasminogen activator inhibitor-1 (PAI-1) is a critical mediator of cellular senescence and fibrin stabilization. We sought to determine if the delivery of recombinant truncated PAI-1 protein (rPAI-123) would protect from the development of radiation-induced lung injury. Methods and Materials C57Bl/6 mice received intraperitoneal injections of rPAI-123 (5.4 μg/kg/day) or vehicle for 18 weeks beginning two days prior to radiation exposure (5 daily fractions of 6 Gy). Cohorts of mice were followed for survival (n=8 per treatment) and tissue collection (n=3 per treatment and time point). Fibrosis in lung was assessed with Masson-Trichrome staining and measurement of hydroxyproline content. Senescence was assessed with staining for beta-galactosidase activity in lung and primary pneumocytes. Results Hydroxyproline content in irradiated lung was significantly reduced in mice that received rPAI-123 compared to mice that received vehicle (IR+vehicle: 84.97, IR+rPAI-123: 56.2 μg/lung, p=0.001). C57Bl/6 mice exposed to IR+vehicle had dense foci of subpleural fibrosis at 19 weeks, whereas the lungs of mice exposed to IR+rPAI-123 were largely devoid of fibrotic foci. Cellular senescence was significantly decreased by rPAI-123 treatment in primary pneumocyte cultures and in lung at multiple time points after IR. Conclusions These studies identify that rPAI-123 is capable of preventing radiation-induced fibrosis in murine lungs. These anti-fibrotic effects are associated with increased fibrin metabolism, enhanced matrix metalloproteinase-3 (MMP-3) expression and reduced senescence in type II pneumocytes. rPAI-123 is a novel therapeutic option for radiation-induced fibrosis.
Fatigue is a disabling symptom in patients with multiple sclerosis and Parkinson’s Disease, and is also common in patients with traumatic brain injury, cancer, and inflammatory disorders. Little is known about the neurobiology of fatigue, in part due to the lack of an approach to induce fatigue in laboratory animals. Fatigue is a common response to systemic challenge by pathogens, a response in part mediated through action of the pro-inflammatory cytokine interleukin-1 beta (IL-1β). We investigated the behavioral responses of mice to IL-1β. Female C57Bl/6J mice of 3 ages were administered IL-1β at various doses i.p. Interleukin-1β reduced locomotor activity, and sensitivity increased with age. Further experiments were conducted with middle-aged females. Centrally administered IL-1β dose-dependently reduced locomotor activity. Using doses of IL-1β that caused suppression of locomotor activity, we measured minimal signs of sickness, such as hyperthermia, pain or anhedonia (as measured with abdominal temperature probes, pre-treatment with the analgesic buprenorphine and through sucrose preference, respectively), all of which are responses commonly reported with higher doses. We found that middle-aged orexin-/- mice showed equivalent effects of IL-1β on locomotor activity as seen in wild-type controls, suggesting that orexins are not necessary for IL-1β -induced reductions in wheel-running. Given that the availability and success of therapeutic treatments for fatigue is currently limited, we examined the effectiveness of two potential clinical treatments, modafinil and methylphenidate. We found that these treatments were variably successful in restoring locomotor activity after IL-1β administration. This provides one step toward development of a satisfactory animal model of the multidimensional experience of fatigue, a model that could allow us to determine possible pathways through which inflammation induces fatigue, and could lead to novel treatments for reversal of fatigue.
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