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Activation of Nrf2/ARE pathway by Anisodamine (654-2) for Inhibition of cellular aging and alleviation of Radiation-Induced lung injury
Abstract
Background:
Radiation-induced lung injury (RILI) is a common side effect of thoracic tumor radiotherapy, including early-stage radiation-induced lung injury (RP) and late-stage radiation-induced pulmonary fibrosis (RIPF). Currently, it is urgently needed to clarify the pathogenesis of RILI and find safe and effective RILI treatment methods. Irradiation causes DNA damage and oxidative stress in tissues and cells, induces cellular senescence, and promotes the occurrence and development of RILI. In recent years, Anisodamine (654-2) has shown potential therapeutic value in acute lung injury, acute kidney injury, chlamydial pneumonia, and COVID-19. However, there is currently no research on the mechanism of 654-2-mediated cellular senescence and its preventive and therapeutic effects on RILI.
Purpose:
This study aimed to investigate the protective effect and mechanism of 654-2 on X-ray-induced RILI.
Methods:
In vivo experiments involved a mouse RILI model with 18 Gy X-ray irradiation. Mice were divided into control, model, medication (control + 654-2), and treatment (model + 654-2) groups. And mice in medication and treatment groups were intraperitoneal injection of 5 mg/kg 654-2 every other day until being sacrificed at week 6. In vitro experiments used MLE-12 cells irradiated with 16 Gy and divided into control, model, and model + 654-2（2 μM and 10 μM） groups. Various assays were performed to evaluate lung tissue morphology, fibrosis, apoptosis, cytokine expression, cellular senescence, protein expression, and antioxidant capacity.
Results:
654-2 mitigated pulmonary pathological damage, inflammation, DNA damage, cellular senescence, and apoptosis in RILI mice and MLE-12 cells. It restored epithelial cell proliferation ability and enhanced antioxidant capacity. Additionally, 654-2 activated the Nrf2/ARE pathway, increased Nrf2 phosphorylation, and upregulated antioxidant gene expression. Inhibition of Nrf2 reversed the effects of 654-2 on ROS production, antioxidant capacity, and cell senescence.
Conclusion:
654-2 can activate the Nrf2/ARE pathway, enhance cellular antioxidant capacity, and inhibit cellular senescence, thereby exerting a protective effect against RILI.
... Previous studies have demonstrated that both single high-dose and multiple lowdose radiation exposures can effectively establish mouse models of radiation pneumonia [33][34][35][36]. These methods accurately reflect the pathophysiological changes observed in patients undergoing radiotherapy for lung injury. ...
Radiation pneumonitis (RP) is a prevalent and fatal complication of thoracic radiotherapy due to the lack of effective treatment options. RP primarily arises from mitochondrial injury in lung epithelial cells. The mitochondrial-derived peptide MOTS-c has demonstrated protective effects against various diseases by mitigating mitochondrial injury. C57BL/6 mice were exposed to 20 Gy of lung irradiation (IR) and received daily intraperitoneal injections of MOTS-c for 2 weeks. MOTS-c significantly ameliorated lung tissue damage, inflammation, and oxidative stress caused by radiation. Meanwhile, MOTS-c reversed the apoptosis and mitochondrial damage of alveolar epithelial cells in RP mice. Furthermore, MOTS-c significantly inhibited oxidative stress and mitochondrial damage in MLE-12 cells and primary mouse lung epithelial cells. Mechanistically, MOTS-c increased the nuclear factor erythroid 2-related factor (Nrf2) level and promoted its nuclear translocation. Notably, Nrf2 deficiency abolished the protective function of MOTS-c in mice with RP. In conclusion, MOTS-c alleviates RP by protecting mitochondrial function through an Nrf2-dependent mechanism, indicating that MOTS-c may be a novel potential protective agent against RP.
... Nrf2, a redox-sensitive transcription factor, is expressed in all cell types and regulates antioxidant defense and cell processes such as mitochondrial bioenergetics, autophagy, unfolded protein response and metabolism through the activation of cytoprotective genes 43 . Previous work has shown that the activation of Nrf2 pathway promotes antioxidant signals and inhibits cellular senescence, thereby exerting protective effects against ageassociated disease 44 . Furthermore, Nrf2 activation in diabetic ischemic muscle has been linked to improved revascularization in a mouse model of type 2 diabetes 45 . ...
Objective: Ischemia due to narrowing of the femoral artery and distal vessels is a major cause of peripheral arterial disease and morbidity affecting patients with diabetes. Diabetes-induced premature senescence of endothelial cells (EC) has been proposed as a mechanism leading to impaired ischemia-driven angiogenesis. Importantly, our previous work has shown that hyperglycemia reduced vascular endothelial growth factor (VEGF) activity in ischemic muscle of diabetic mice, which was associated with increased expression of the protein tyrosine phosphatase Src homology region 2 domain-containing phosphatase-1 (SHP-1). Here, we evaluate the impact of SHP-1 deletion on EC function and senescence.
Approach and Results: Ligation of the femoral artery was performed in nondiabetic (NDM) and 3 months diabetic (DM) mice with EC-specific deletion of SHP-1 and blood flow reperfusion was measured for 4 weeks. Blood flow reperfusion and limb function during voluntary wheel running were reduced by 43% and 82%, respectively in DM mice as compared to NDM mice. EC-specific deletion of SHP-1 in DM mice restored blood flow reperfusion by 60%, limb function by 86%, while capillary density was similar to NDM mice. Moreover, ablation of SHP-1 in EC prevented diabetes-induced expression of the senescence markers p53 and p21 and counteracted Nrf2 downregulation. In cultured EC, overexpression of dominant negative of SHP-1 prevented HG-induced inhibition of proliferation, migration, tubule formation and VEGFR2/Akt phosphorylation following VEGF stimulation. In addition, the expression of senescence markers and suppression of Nrf2 in EC exposed to HG levels were reversed by overexpression of dominant negative SHP-1.
Conclusion: SHP-1 in ECs is a central effector of diabetes-induced senescence that blocks VEGF action, and induces aberrant collateral vessel formation and blood flow reperfusion. Reduced SHP-1 expression counteracts these pathologic features suggesting the notion that it represents a promising therapeutic target.
Alkaloids are natural products that occur widely in many herbal plants. Anisodamine, widely present in the Solanaceae family, is an alkaloid extracted from the roots of the Anisodus tanguticus Maxim. It is an antagonist to M‐choline receptors and exhibits diverse pharmacological effects, such as cholinolytic effect, calcium antagonist effect, anti‐oxygenation effect. Anisodamine, a prominent constituent of the tropine alkaloid family, exhibits a range of pharmacological effects akin to those of atropine and scopolamine. owing to its low toxicity and moderate efficacy in clinical to wide applications, especially for varieties of shock treatment . However, there remains a dearth of research regarding the in vivo pharmacokinetics, mechanism of action, and toxicity of anisodamine. Consequently, this paper provides a comprehensive review of the anti‐shock effects, toxicity ,and pharmacokinetic characteristics of anisodamine to increase the understanding of its medicinal value, and provide reference and inspiration for the clinical application and further in‐depth research of anisodamine.
Intervertebral disc degeneration (IDD) is considered as a dominant contributor to low back pain (LBP), causing severe pain, limited range of lumbar motion, physical dysfunction, and restriction of social activity. However, the specific pathological mechanisms underlying IDD remain elusive, and effective strategies to delay the pathogenesis of IDD are still unclear and limited. In recent years, some studies have found that nuclear factor erythroid 2-related factor 2 (Nrf2), an important antioxidant transcription factor, may play crucial roles in the pathogenesis and progression of age-related diseases including IDD. Nrf2 can maintain redox homeostasis and protecting nucleus pulposus (NP) cells against oxidative stress, inflammatory response, extracellular matrix (ECM) catabolism, cell senescence and cell death involving in the progression of IDD. In this review, we aim to systematically describe the vital roles and pathological mechanism of Nrf2 signaling axis in the pathogenesis of IDD, which may put forward potential therapeutic strategies for the prevention and treatment of IDD by targeting Nrf2.
The limitation of cancer radiotherapy does not derive from an inability to ablate tumor, but rather to do so without excessively damaging critical tissues and organs and adversely affecting patient’s quality of life. Although cellular senescence is a normal consequence of aging, there is increasing evidence showing that the radiation-induced senescence in both tumor and adjacent normal tissues contributes to tumor recurrence, metastasis, and resistance to therapy, while chronic senescent cells in the normal tissue and organ are a source of many late damaging effects. In this review, we discuss how to identify cellular senescence using various bio-markers and the role of the so-called senescence-associated secretory phenotype characteristics on the pathogenesis of the radiation-induced late effects. We also discuss therapeutic options to eliminate cellular senescence using either senolytics and/or senostatics. Finally, a discussion of cellular reprogramming is presented, another promising avenue to improve the therapeutic gain of radiotherapy.
Radiation-induced lung injury (RILI) is one of the most common, serious, and dose-limiting toxicities of thoracic radiotherapy. A primary cause for this is the radiation-induced cell death. Ferroptosis is a recently recognized form of regulated cell death, characterized by the accumulation of lipid peroxidation products and lethal reactive oxygen species (ROS). The ROS generated by irradiation might be the original trigger of ferroptosis in RILI. In addition, activation of the P62-Kelch-like ECH-associated protein 1 (Keap1)-nuclear factor erythroid 2-related factor 2 (NRF2) pathway has been shown to blunt ferroptosis and thus acts as a protective factor. Therefore, this study aimed to explore the protective effect of the P62-Keap1-NRF2 pathway against radiation-induced ferroptosis in alveolar epithelial cells. First, we found that radiation induced ferroptosis in vitro using a RILI cell model, which could be significantly reduced by ferrostatin-1 (Fer-1), a specific ferroptosis inhibitor. Additionally, overexpression of P62 interacted with Keap1 to facilitate the translocation of NRF2 into the nucleus and promote the expression of its target proteins, including quinone oxidoreductase 1 (NQO1), heme oxygenase 1 (HO1), and ferritin heavy chain 1 (FTH1). In summary, our results demonstrated that the activation of the P62-Keap1-NRF2 pathway prevents radiation-induced ferroptosis in RILI cells, providing a theoretical basis of finding a potential therapeutic approach for RILI.
Background and objective:
Radiation-induced lung injury (RILI) is often found in thoracic tumor patients after thoracic radiation therapy, and influences patient quality of life. However, systematic exploration of RILI, including its molecular biological mechanisms and standardized treatment, has not yet been fully elucidated. The main objective of the narrative review was to describe the available evidence concerning RILI, from the biological mechanism to the clinical management. The underlying causes of RILI are multifactorial, including gene-level changes, the influence of signaling pathways, the convergence of various cells, as well as the expression of cytokines and chemokines. Based on the various mechanisms of RILI, several novel treatment strategies have been proposed and gradually applied in clinical practice.
Methods:
PubMed was used to collect articles about RILI from 1995 to 2021. The papers included clinical trials, reviews, as well as systematic reviews and meta-analyses. Based on the mechanism, diagnosis, and treatment, we synthesized and analyzed these papers to form a clearly logical and normative suggestion to guide clinical application.
Key content and findings:
RILI is a constantly developing and changing process including radiation pneumonitis and radiation lung fibrosis. Different kinds of inflammatory and immune cells such as macrophages, fibroblasts, and T cells play key roles in the development of RILI, and transforming growth factor-β (TGF-β), interleukin-4 (IL-4), IL-13, and interferon-γ (IFN-γ) are also participants in this process. At present, glucocorticoids are mainly therapeutic drugs for the early stage of RILI, and drugs treatment should abide early period, sufficient doses, and the individual principles. Other novel drugs such as Azithromycin also have been tried in clinical application. radiation dose, combination therapy modality, the condition of the tumor, and the age and underlying conditions of patients all effect the occurrence of RILI. Importantly, RILI has a relatively higher incidence in patients who received radiotherapy combined with other treatments, especially immunotherapy.
Conclusions:
The occurrence of RILI after radiotherapy will greatly affect the prognosis and quality of life of patients. In clinical practice, early intervention, active treatment, and more effective therapeutic drugs should be found.
Radiotherapy plays an important role in the treatment of localized primary malignancies involving the chest wall or intrathoracic malignancies. Secondary effects of radiotherapy on the lung result in radiation-induced lung disease. The phases of lung injury from radiation range from acute pneumonitis to chronic pulmonary fibrosis. Radiation pneumonitis is a clinical diagnosis based on the history of radiation, imaging findings, and the presence of classic symptoms after exclusion of infection, pulmonary embolism, heart failure, drug-induced pneumonitis, and progression of the primary tumor. Computed tomography (CT) is the preferred imaging modality as it provides a better picture of parenchymal changes. Lung biopsy is rarely required for the diagnosis. Treatment is necessary only for symptomatic patients. Mild symptoms can be treated with inhaled steroids while subacute to moderate symptoms with impaired lung function require oral corticosteroids. Patients who do not tolerate or are refractory to steroids can be considered for treatment with immunosuppressive agents such as azathioprine and cyclosporine. Improvements in radiation technique, as well as early diagnosis and appropriate treatment with high-dose steroids, will lead to lower rates of pneumonitis and an overall good prognosis.
Background:
Pediatric acute lung injury (ALI) is one of the most common causes of infant mortality. Although lung-protective strategies have developed in recent years, no ALI treatment is currently available. Anisodamine (Ani) is a common drug used to treat gastrointestinal smooth muscle spasm. The protective effects of Ani against acute kidney injury and myocardial injury have been reported. However, the efficacy of Ani on bleomycin (BLM)-induced ALI has not been examined previously. In the present study, we aimed to examine the effects of Ani on bleomycin (BLM)-induced ALI on immature rats.
Methods:
The ALI rat model was established by intratracheally administration of BLM. Ani treatment was performed by an intravenous injection at different concentrations. The lung function of each rat was measured, and then lung tissue structures, apoptosis, and collagen deposition were observed by hematoxylin-eosin staining, terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling assay, and Masson's staining, respectively. Enzyme-linked immunosorbent assay was used to detect the levels of inflammatory cytokines. The expression of apoptosis-related proteins and fibrosis-related markers was determined by reverse transcription-polymerase chain reaction and/or Western blot analysis. Finally, the expression levels of Janus tyrosine kinase 2 (JAK2) and signal transducer and activator of transcription 3 (STAT3) were determined.
Results:
Our findings indicated that lung function was remarkably decreased in BLM-induced rats, which could be reversed by Ani. Ani treatment increased the levels of antioxidant enzymes, decreased the apoptotic rate and apoptosis-related proteins, and downregulated the expression of fibrosis-related markers. Additionally, Ani treatment also attenuated inflammatory response and suppressed the activation of the JAK2/STAT3 pathway.
Conclusions:
Our results demonstrated that Ani had potent activity against BLM-induced ALI in immature rats through inhibiting the JAK2/STAT3 signaling pathway. Our findings provide supporting evidence to further investigate the therapeutic effect of Ani against ALI in children.
The outbreak of coronavirus disease 2019 (COVID-19) underlined the urgent need for alleviating cytokine storm. We propose here that activating the cholinergic anti-inflammatory pathway (CAP) is a potential therapeutic strategy. However, there is currently no approved drugs targeting the regulatory pathway. It is evident that nicotine, anisodamine and some herb medicine, activate the CAP and exert anti-inflammation action in vitro and in vivo. As the vagus nerve affects both inflammation and specific immune response, we propose that vagus nerve stimulation by invasive or non-invasive devices and acupuncture at ST36, PC6, or GV20, are also feasible approaches to activate the CAP and control COVID-19. It is worth to investigate the efficacy and safety of the strategy in patients with COVID-19.
Background:
The Coronavirus Disease 2019 (COVID-19) outbreak in Wuhan, China, was caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Anisodamine hydrobromide injection (AHI), the main ingredient of which is anisodamine, is a listed drug for improving microcirculation in China. Anisodamine can improve the condition of patients with COVID-19.
Materials and methods:
Protein-protein interactions obtained from the String databases were used to construct the protein interaction network (PIN) of AHI using Cytoscape. The crucial targets of AHI PIN were screened by calculating three topological parameters. Gene ontology and pathway enrichment analyses were performed. The intersection between the AHI component proteins and angiotensin-converting enzyme 2 (ACE2) coexpression proteins was analyzed. We further investigated our predictions of crucial targets by performing molecular docking studies with anisodamine.
Results:
The PIN of AHI, including 172 nodes and 1454 interactions, was constructed. A total of 54 crucial targets were obtained based on topological feature calculations. The results of Gene Ontology showed that AHI could regulate cell death, cytokine-mediated signaling pathways, and immune system processes. KEGG disease pathways were mainly enriched in viral infections, cancer, and immune system diseases. Between AHI targets and ACE2 coexpression proteins, 26 common proteins were obtained. The results of molecular docking showed that anisodamine bound well to all the crucial targets.
Conclusion:
The network pharmacological strategy integrated molecular docking to explore the mechanism of action of AHI against COVID-19. It provides protein targets associated with COVID-19 that may be further tested as therapeutic targets of anisodamine.
Anisodamine hydrobromide (AniHBr) is a Chinese medicine used to treat septic shock. However, whether AniHBr could ameliorate septic acute kidney injury and the underlying mechanism were not investigated. In the present study, eighteen male Sprague-Dawley rats (200 g-250 g) were randomly divided into control, lipopolysaccharide (LPS) and LPS+AniHBr groups. Rats were intravenously administrated with LPS or normal saline (for control). After 4 h, the rats were intravenously administrated with AniHBr (LPS+AniHBr) or normal saline at 4 h intervals. Heamodynamic parameters including blood pressure and heart rate were measured. The histopathologic evaluation of kidney tissues was performed. Lactate, creatine kinase, inflammatory cytokines and oxidative stress indicators were determined. Using Seahorse analysis, the metabolic analysis of mitochondrial stress and glycolytic stress in human renal proximal tubular epithelial cells treated with TNF-α in the presence of AniHBr were performed. AniHBr administration significantly reduced serum creatine kinase and lactate following LPS treatment. AniHBr significantly improved heamodynamics in sepsis rats including increase of the mean atrial pressure and reduction of the heart rate. AniHBr significantly attenuated LPS-induced TNF-α, IL-6 and IL-1β in serum, and LPS-induced TNF-α and IL-1β in renal tissues. The LPS-reduced SOD activity and LPS-increased MDA content were reversed by AniHBr. In vitro, TNF-α increased mitochondrial oxygen consumption and glycolysis, but inhibited the ATP generation, which was reversed by AniHBr. Thus, AniHBr protects against the LPS-induced inflammatory cytokines, mitochondrial dysfunction and oxidative stress, and thus attenuates the LPS-induced acute kidney injury, showing AniHBr is a promising therapeutic drug for septic kidney injury.
Cellular senescence is a key process in physiological dysfunction developing upon aging or following diverse stressors including ionizing radiation. It describes the state of a permanent cell cycle arrest, in which proliferating cells become resistant to growth-stimulating factors. Senescent cells differ from quiescent cells, which can re-enter the cell cycle and from finally differentiated cells: morphological and metabolic changes, restructuring of chromatin, changes in gene expressions and the appropriation of an inflammation-promoting phenotype, called the senescence-associated secretory phenotype (SASP), characterize cellular senescence. The biological role of senescence is complex, since both protective and harmful effects have been described for senescent cells. While initially described as a mechanism to avoid malignant transformation of damaged cells, senescence can even contribute to many age-related diseases, including cancer, tissue degeneration, and inflammatory diseases, particularly when senescent cells persist in damaged tissues. Due to overwhelming evidence about the important contribution of cellular senescence to the pathogenesis of different lung diseases, specific targeting of senescent cells or of pathology-promoting SASP factors has been suggested as a potential therapeutic approach. In this review, we summarize recent advances regarding the role of cellular (fibroblastic, endothelial, and epithelial) senescence in lung pathologies, with a focus on radiation-induced senescence. Among the different cells here, a central role of epithelial senescence is suggested.
Oxidative stress is the increase in cellular oxidant concentration in comparison to
antioxidant titer. Toxic insults and many other diseased conditions are mediated
through the formation of such condition. Once the redox equilibrium is disrupted, the
cellular antioxidant system functions to bring back the cell to redox homeostasis
state. The field players of the cytoprotective machinery are the xenobiotic‐
metabolizing enzymes that are transcriptionally controlled by upstream regulatory
pathways like the Nrf2–ARE pathway and AhR–XRE pathway. The importance of
Nrf2 lies in the fact that it is activated by a variety of compounds and has a wide
range of inducers including metals, organic toxicants and so forth. The present review
article aims to discuss the role of Nrf2 in cellular protection and also intends to
illuminate the regulatory mechanisms that control Nrf2 itself. This can add to our
knowledge of how the cell reacts and survives against such stressed conditions.
Previous studies demonstrated that Bailcalin (BAI) prevented cardiac injuries under different disease models. Whether BAI protected against type 2 diabetes mellitus- (T2DM-) associated cardiomyopathy was investigated in this study. T2DM was established by the combination of streptozotocin injection and high-fat diet in mice. BAI was administered daily for 6 months. After evaluating cardiac functions, mice hearts were removed and processed for morphological, biochemical, and molecular mechanism analyses. Neonatal rat cardiomyocytes (NRCM) were isolated and treated with high glucose and palmitate (HG/Pal) for in vitro investigation. BAI significantly ameliorated T2DM-induced cardiomyocyte hypertrophy, interstitial fibrosis, and lipid accumulation accompanied by markedly improved cardiac functions in diabetic mice. Mechanically, BAI restored decreased phosphorylation of AMPK and enhanced expression and nuclei translocation of Nrf2. In in vitro experiments, BAI also prevented NRCM from HG/Pal-induced apoptosis and oxidative stress injuries by increasing p-AMPK and Nrf2 accumulation. The means by which BAI restored p-AMPK seemed to be related to the antioxidative effects of Nrf2 after silencing AMPK or Nrf2 in NRCM. Furthermore, BAI regulated Nrf2 by inhibiting Nrf2 ubiquitination and consequent degradation mediated by Keap1. This study showed that BAI alleviated diabetes-associated cardiac dysfunction and cardiomyocyte injuries in vivo and in vitro via Keap1/Nrf2/AMPK-mediated antioxidation and lipid-lowering effects. BAI might be a potential adjuvant drug for diabetes cardiomyopathy treatment.
Background:
Anisodamine is used for the treatment of reperfusion injury in various organs. In this study, we investigated the effectiveness and mechanisms of action of anisodamine in promoting recovery from glycerol-induced acute kidney injury (AKI).
Methods:
We compared the protective effects of atropine and anisodamine in the rat model of glycerol-induced AKI. We examined signaling pathways involved in oxidative stress, inflammation and apoptosis, as well as expression of kidney injury molecule-1 (KIM-1). Renal injury was assessed by measuring serum creatinine and urea, and by histologic analysis. Rhabdomyolysis was evaluated by measuring creatine kinase levels, and oxidative stress was assessed by measuring malondialdehyde (MDA) and superoxide dismutase (SOD) levels in kidney tissues. Inflammation was assessed by quantifying interleukin 6 (IL-6) and CD45 expression. Apoptosis and necrosis were evaluated by measuring caspase-3 (including cleaved caspase 3) and RIP3 levels, respectively.
Results:
Glycerol administration resulted in a higher mean histologic damage score, as well as increases in serum creatinine, urea, creatine kinase, reactive oxygen species (ROS), MDA, IL-6, caspase-3 and KIM-1 levels. Furthermore, glycerol reduced kidney tissue SOD activity. All of these markers were significantly improved by anisodamine and atropine. However, the mean histologic damage score and levels of urea, serum creatinine, creatine kinase, ROS and IL-6 were lower in the anisodamine treatment group compared with the atropine treatment group.
Conclusion:
Pretreatment with anisodamine ameliorates renal dysfunction in the rat model of glycerol-induced rhabdomyolytic kidney injury by reducing oxidative stress, the inflammatory response and cell death.
We tested the hypothesis that 1,25‐dihydroxyvitamin D3[1α,25(OH)2D3] has antiaging effects via upregulating nuclear factor (erythroid‐derived 2)‐like 2 (Nrf2), reducing reactive oxygen species (ROS), decreasing DNA damage, reducing p16/Rb and p53/p21 signaling, increasing cell proliferation, and reducing cellular senescence and the senescence‐associated secretory phenotype (SASP). We demonstrated that 1,25(OH)2D3‐deficient [1α(OH)ase−/−] mice survived on average for only 3 months. Increased tissue oxidative stress and DNA damage, downregulated Bmi1 and upregulated p16, p53 and p21 expression levels, reduced cell proliferation, and induced cell senescence and the senescence‐associated secretory phenotype (SASP) were observed. Supplementation of 1α(OH)ase−/− mice with dietary calcium and phosphate, which normalized serum calcium and phosphorus, prolonged their average lifespan to more than 8 months with reduced oxidative stress and cellular senescence and SASP. However, supplementation with exogenous 1,25(OH)2D3 or with combined calcium/phosphate and the antioxidant N‐acetyl‐l‐cysteine prolonged their average lifespan to more than 16 months and nearly 14 months, respectively, largely rescuing the aging phenotypes. We demonstrated that 1,25(OH)2D3exerted an antioxidant role by transcriptional regulation of Nrf2 via the vitamin D receptor (VDR). Homozygous ablation of p16 or heterozygous ablation of p53 prolonged the average lifespan of 1α(OH)ase−/− mice on the normal diet from 3 to 6 months by enhancing cell proliferative ability and reducing cell senescence or apoptosis. This study suggests that 1,25(OH)2D3 plays a role in delaying aging by upregulating Nrf2, inhibiting oxidative stress and DNA damage，inactivating p53‐p21 and p16‐Rb signaling pathways, and inhibiting cell senescence and SASP.
Radiation-induced lung injury (RILI) is one of the most common and fatal complications of thoracic radiotherapy. Inflammatory cell infiltration, imbalance of inflammatory cytokines, and oxidative damage were reported to be involved during RILI pathogenesis, especially in the early phase of RILI. Nuclear factor-erythroid 2-related factor 2 (Nrf2) is a key transcriptional regulator of antioxidative cascades, and regulates life span of mice after administration of thoracic irradiation. We investigated the effects of Nrf2 on RILI and inflammation using Nrf2-knockout, Nrf2-overexpression and wild-type mice with or without 15 Gy ionizing radiation to thorax. Our results showed that Nrf2 deficiency aggravated radiation-induced histopathological changes, macrophage and neutrophil infiltration, serum levels of pro-inflammatory cytokines (IL-6, MCP-1, IFN-γ, TNF, and IL-12p70), and the levels of peroxidation products in the mouse lung. Moreover, loss of Nrf2 reduced radiation-induced serum levels of anti-inflammatory cytokine, IL-10, and antioxidative proteins. Nrf2 overexpression significantly alleviated radiation-induced histopathological changes, macrophages and neutrophils infiltration, serum levels of pro-inflammatory cytokines, and the levels of peroxidation products in lung tissues. Nrf2 overexpression also increased the serum levels of IL-10 and antioxidative proteins. These results indicated that Nrf2 had a protective role against radiation-induced acute lung injury and inflammation, and that antioxidative therapy might be a promising treatment for RILI.
Diabetes is associated with an increased risk of cardiovascular disease. A decrease in the number and functionality of endothelial progenitor cells (EPCs) leads to reduced endothelial repair and the development of cardiovascular disease. The aim of the present study was to explore the effect and underlying mechanisms of nuclear factor erythroid 2‑related factor 2 (Nrf2) on EPC dysfunction caused by diabetic mellitus. The biological functions of EPCs in streptozotocin‑induced diabetic mice were evaluated, including migration, proliferation, angiogenesis and the secretion of vascular endothelial growth factor (VEGF), stromal‑derived growth factor (SDF) and nitric oxide (NO). Oxidative stress levels in diabetic EPCs were also assessed by detecting intracellular reactive oxygen species (ROS), superoxide dismutase (SOD) and malondialdehyde (MDA). EPC senescence was evaluated by measuring p16 and b‑gal expression and observing the senescence‑associated secretory phenotype. In addition, the function of EPCs and level of oxidative stress were assessed following Nrf2 silencing or activation. Nrf2 silencing resulted in a decrease of EPC biological functions, accelerated cell senescence and increased oxidative stress, as indicated by ROS and MDA upregulation accompanied with decreased SOD activity. Furthermore, Nrf2 silencing inhibited migration, proliferation and secretion in EPCs, while it increased oxidative stress and cell senescence. Nrf2 activation protected diabetic EPCs against the effects of oxidative stress and cell senescence, ameliorating the biological dysfunction of EPCs derived from mice with diabetes. In conclusion, Nrf2 overexpression protected against oxidative stress‑induced functional damage in EPCs derived from diabetic mice by regulating cell senescence.
In order to better understand the early pathways of the pathogenesis of, and immune response to, RSV, herein, we explored the relationship between TLR7 expression and oxidative stress induction following RSV infection in A549 cells. We studied the intervening effects of the Nrf2/ARE pathway agonist butylated hydroxyanisole (BHA) and inhibitor trigonelline (TRI) on TLR7 modulation or oxidative stress induction. For comparison purposes, we set up seven treatment groups in this study, including RSV-treated cells, BHA + RSV-treated cells, TRI + RSV-treated cells, normal cell controls, inactivated RSV controls, BHA controls and TRI controls. We measured changes in TLR7, IL-6, TNF-α mRNA using RT-PCR and IL-6, TNF-α and IL-1β protein using ELISA as well as TLR7, Nrf2 and HO-1 protein using Western blot in A549 cells from the different treatment groups. We also assessed changes in cell proliferation and measured changes in ·OH and NO in A549 cells from the different treatment groups. The results indicate that TLR7 up-regulation is related to RSV infection and the induction of oxidative stress and that TLR7 expression was mediated by the anti-inflammatory effects of Nrf2/ARE pathway inhibitors or agonists. Our experiments may help elucidate the underlying pathology of RSV infection and suggest potential therapeutic targets for drug development and the prevention of RSV-induced human diseases.
Anisodamine protects against free radical-induced cellular damage. This study aimed to investigate the protective effect of anisodamine on rhabdomyolysis-induced acute kidney injury (RIAKI). C57BL/6 J mice, TXNIP−/− and NLRP3 −/− (both were C57BL/6 J background) mice were used to construct RIAKI model. Anisodamine administration was performed on RIAKI mice only. Mice were divided into control, TXNIP-KD (knock down), LNPR3-KD, and anisodamine group (n = 15 in each group). The renal injury, renal function, renal tubular cells apoptosis and expression of Caspase-1, ASC, endoplasmic reticulum (ER) stress markers IRE-1α, CHOP, and ATF4, and interleukin (IL-1α, IL-1β, and IL-18) were detected. The knock down of TXNIP or NLRP3 expression in mice showed protective effect against RIAKI pathogenesis, as compared with the RIAKI mice. The expression of Caspase-1, ASC, and interleukins, renal injury, renal tubular cells apoptosis in TXNIP-KD and LNPR3-KD mice were significantly inhibited in comparison with the RIAKI mice. Moreover, anisodamine treatment reduced expression of ER stress markers IRE-1α, CHOP, and ATF4, TXNIP and NLRP3, as well as ACS, Caspase-1, IL-1α, IL-1β, and IL-18, showing moderate protective effect on the changes of above factors comparing with TXNIP or NLRP3 knock down. This study declared that anisodamine showed protective effect on RIAKI model may by inhibiting ER stress associated TXNIP/NLRP3 inflammasome.
The development of radiation-induced pulmonary fibrosis represents a critical clinical issue limiting delivery of therapeutic doses of radiation to non-small cell lung cancer. Identification of the cell types whose injury initiates a fibrotic response and the underlying biological factors that govern that response are needed for developing strategies that prevent or mitigate fibrosis. C57BL/6 mice (wild type, Nrf2 null, Nrf2(flox/flox), and Nrf2(Δ/Δ); SPC-Cre) were administered a thoracic dose of 12Gy and allowed to recover for 250 days. Whole slide digital and confocal microscopy imaging of H&E, Masson's trichrome and immunostaining were used to assess tissue remodeling, collagen deposition and cell renewal/mobilization during the regenerative process. Histological assessment of irradiated, fibrotic wild type lung revealed significant loss of alveolar type 2 cells 250 days after irradiation. Type 2 cell loss and the corresponding development of fibrosis were enhanced in the Nrf2 null mouse. Yet, conditional deletion of Nrf2 in alveolar type 2 cells in irradiated lung did not impair type 2 cell survival nor yield an increased fibrotic phenotype. Instead, radiation-induced ΔNp63 stem/progenitor cell mobilization was inhibited in the Nrf2 null mouse while the propensity for radiation-induced myofibroblasts derived from alveolar type 2 cells was magnified. In summary, these results indicate that Nrf2 is an important regulator of irradiated lung's capacity to maintain alveolar type 2 cells, whose injury can initiate a fibrotic phenotype. Loss of Nrf2 inhibits ΔNp63 stem/progenitor mobilization, a key event for reconstitution of injured lung, while promoting a myofibroblast phenotype that is central for fibrosis.
Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease with poor prognosis and limited therapeutic options. The incidence of IPF increases with age, and ageing-related mechanisms such as cellular senescence have been proposed as pathogenic drivers. The lung alveolar epithelium represents a major site of tissue injury in IPF and senescence of this cell population is probably detrimental to lung repair. However, the potential pathomechanisms of alveolar epithelial cell senescence and the impact of senolytic drugs on senescent lung cells and fibrosis remain unknown. Here we demonstrate that lung epithelial cells exhibit increased P16 and P21 expression as well as senescence-associated β-galactosidase activity in experimental and human lung fibrosis tissue and primary cells.
Primary fibrotic mouse alveolar epithelial type (AT)II cells secreted increased amounts of senescence-associated secretory phenotype (SASP) factors in vitro, as analysed using quantitative PCR, mass spectrometry and ELISA. Importantly, pharmacological clearance of senescent cells by induction of apoptosis in fibrotic ATII cells or ex vivo three-dimensional lung tissue cultures reduced SASP factors and extracellular matrix markers, while increasing alveolar epithelial markers.
These data indicate that alveolar epithelial cell senescence contributes to lung fibrosis development and that senolytic drugs may be a viable therapeutic option for IPF.
Senescent cells contribute to age-related pathology and loss of function, and their selective removal improves physiological function and extends longevity. Rapamycin, an inhibitor of mTOR, inhibits cell senescence in?vitro and increases longevity in several species. Nrf2 levels have been shown to decrease with aging and silencing Nrf2 gene induces premature senescence. Therefore, we explored whether Nrf2 is involved in the mechanism by which rapamycin delays cell senescence. In wild-type (WT) mouse fibroblasts, rapamycin increased the levels of Nrf2, and this correlates with the activation of autophagy and a reduction in the induction of cell senescence, as measured by SA-?-galactosidase (?-gal) staining, senescence-associated secretory phenotype (SASP), and p16 and p21 molecular markers. In Nrf2KO fibroblasts, however, rapamycin still decreased ?-gal staining and the SASP, but rapamycin did not activate the autophagy pathway or decrease p16 and p21 levels. These observations were further confirmed in?vivo using Nrf2KO mice, where rapamycin treatment led to a decrease in ?-gal staining and pro-inflammatory cytokines in serum and fat tissue; however, p16 levels were not significantly decreased in fat tissue. Consistent with literature demonstrating that the Stat3 pathway is linked to the production of SASP, we found that rapamycin decreased activation of the Stat3 pathway in cells or tissue samples from both WT and Nrf2KO mice. Our data thus suggest that cell senescence is a complex process that involves at least two arms, and rapamycin uses Nrf2 to regulate cell cycle arrest, but not the production of SASP.
Idiopathic pulmonary fibrosis (IPF) is a fatal disease characterized by interstitial remodelling, leading to compromised lung function. Cellular senescence markers are detectable within IPF lung tissue and senescent cell deletion rejuvenates pulmonary health in aged mice. Whether and how senescent cells regulate IPF or if their removal may be an efficacious intervention strategy is unknown. Here we demonstrate elevated abundance of senescence biomarkers in IPF lung, with p16 expression increasing with disease severity. We show that the secretome of senescent fibroblasts, which are selectively killed by a senolytic cocktail, dasatinib plus quercetin (DQ), is fibrogenic. Leveraging the bleomycin-injury IPF model, we demonstrate that early-intervention suicide-gene-mediated senescent cell ablation improves pulmonary function and physical health, although lung fibrosis is visibly unaltered. DQ treatment repli-cates benefits of transgenic clearance. Thus, our findings establish that fibrotic lung disease is mediated, in part, by senescent cells, which can be targeted to improve health and function.
Although it is known that the regenerative function of neural stem/progenitor cells (NSPCs) declines with age, causal mechanisms underlying this phenomenon are not understood. Here, we systematically analyze subventricular zone (SVZ) NSPCs, in various groups of rats across the aging spectrum, using in vitro and in vivo histological and behavioral techniques. These studies indicate that although NSPC function continuously declines with advancing age, there is a critical time period during middle age (13-15 months) when a striking reduction in NSPC survival and regeneration (proliferation and neuronal differentiation) occurs. The studies also indicate that this specific temporal pattern of NSPC deterioration is functionally relevant at a behavioral level and correlates with the decreasing expression of the redox-sensitive transcription factor, Nrf2, in the NSPCs. When Nrf2 expression was suppressed in 'young' NSPCs, using short interfering RNAs, the survival and regeneration of the NSPCs was significantly compromised and mirrored 'old' NSPCs. Conversely, Nrf2 overexpression in 'old' NSPCs rendered them similar to 'young' NSPCs, and they showed increased survival and regeneration. Furthermore, examination of newborn Nrf2 knockout (Nrf2 -/-) mice revealed a lower number of SVZ NSPCs in these animals, when compared to wild-type controls. In addition, the proliferative and neurogenic potential of the NSPCs was also compromised in the Nrf2-/- mice. These results identify a novel regulatory role for Nrf2 in NSPC function during aging and have important implications for developing NSPC-based strategies to support healthy aging and to treat age-related neurodegenerative disorders.
During the last few decades, improvements in the planning and application of radiotherapy in combination with surgery and chemotherapy resulted in increased survival rates of tumor patients. However, the success of radiotherapy is impaired by two reasons: firstly, the radioresistance of tumor cells and, secondly, the radiation-induced damage of normal tissue cells located in the field of ionizing radiation. These limitations demand the development of drugs for either radiosensitization of tumor cells or radioprotection of normal tissue cells. In order to identify potential targets, a detailed understanding of the cellular pathways involved in radiation response is an absolute requirement. This review describes the most important pathways of radioresponse and several key target proteins for radiosensitization.
The TOR (target of rapamycin) kinase limits longevity by poorly understood mechanisms. Rapamycin suppresses the mammalian TORC1 complex, which regulates translation, and extends lifespan in diverse species, including mice. We show that rapamycin selectively blunts the pro-inflammatory phenotype of senescent cells. Cellular senescence suppresses cancer by preventing cell proliferation. However, as senescent cells accumulate with age, the senescence-associated secretory phenotype (SASP) can disrupt tissues and contribute to age-related pathologies, including cancer. MTOR inhibition suppressed the secretion of inflammatory cytokines by senescent cells. Rapamycin reduced IL6 and other cytokine mRNA levels, but selectively suppressed translation of the membrane-bound cytokine IL1A. Reduced IL1A diminished NF-κB transcriptional activity, which controls much of the SASP; exogenous IL1A restored IL6 secretion to rapamycin-treated cells. Importantly, rapamycin suppressed the ability of senescent fibroblasts to stimulate prostate tumour growth in mice. Thus, rapamycin might ameliorate age-related pathologies, including late-life cancer, by suppressing senescence-associated inflammation.
Radiation is a commonly delivered therapeutic modality for cancer. The causes underlying the chronic, progressive nature of radiation injury in the lung are poorly understood.
C57Bl/6NCr mice were exposed to thoracic irradiation (n = 3 per dose and time point for tissue collection). Microarray analysis of gene expression from irradiated murine lung was performed using one-way analysis of variance with post hoc Scheffe analysis. Senescence and type II airway epithelial cell (AECII) count were assayed in irradiated murine lung tissue (n = 3 per condition). Irradiated mice were treated with diphenyleneiodonium (DPI), an inhibitor of NADPH oxidase (NOX), and fibrosis was assessed by collagen assays. All statistical tests were two-tailed.
Gene expression in lung tissue from mice irradiated to 17.5 Gy clustered with that of aged unirradiated mice. Only fibrogenic exposures led to AECII senescence (0 Gy: 0.66% +/- 0.67%; 5 Gy: 4.5% +/- 1.19%; 17.5 Gy: 18.7% +/- 3.05; P = .007) and depletion (0 Gy: 2.89 per alveolus +/- 0.26; 5 Gy: 2.41 +/- 0.19; 17.5 Gy: 1.6 +/- 0.14; P < .001) at 30 weeks. Treatment of irradiated mice with DPI for 16 weeks markedly reduced collagen accumulation (5×6 Gy: 57.26 μg/lung +/- 9.91; 5×6 Gy +/- DPI: 36.54μg/lung +/- 4.39; P = .03) and AECII senescence (5×6 Gy: 37.61% +/- 4.82%; 5×6 Gy +/- DPI: 12.38% +/- 2.78; P < .001).
These studies identify senescence as an important process in AECII in vivo and indicate that NOX is a critical mediator of radiation-induced AECII senescence and pulmonary fibrosis.
Ubiquitination has long been recognised as a key determinator of protein fate by tagging proteins for proteasomal degradation. Most recently, the ability of conjugated ubiquitin chains to confer selectivity to autophagy was demonstrated. Although autophagy was first believed to be a bulk, non-selective 'self-eating' degradative process, the molecular mechanisms of selectivity are now starting to emerge. With the discovery of autophagy receptors - which bind both ubiquitinated substrates and autophagy specific light chain 3 (LC3) modifier on the inner sheath of autophagosomes - a new pathway of selective autophagy is being unravelled. In this review, we focus on the special role of ubiquitin signals and selective autophagy receptors in sorting a variety of autophagic cargos.Cell Death and Differentiation advance online publication, 22 June 2012; doi:10.1038/cdd.2012.72.
Subsets of cancer survivors who have been subjected to thoracic irradiation face the prospect of developing pulmonary injury. Radiation-induced pulmonary fibrosis is an insidious injury that presents 6 to 24 months after irradiation and continues to progress over a period of years. TGF-β and reactive oxygen species contribute significantly to the pathogenesis of this injury. The transcription factor NRF2 controls antioxidant gene expression and therefore regulates the cellular oxidant burden. This work demonstrates an additional paradigm for NRF2: suppression of TGF-β-mediated signaling, assessed by measuring expression of a surrogate TGF-β1 target gene (PAI-1) in lung fibroblasts. Thoracic irradiation of Nfe2l2(-/-) mice resulted in rapid expression of PAI-1 and FSP-1 compared to irradiated wild-type mice. Examination of lung tissue 16 weeks after thoracic irradiation of Nfe2l2(-/-) mice revealed the presence of distended alveoli and decreased numbers of alveoli compared to wild-type mice. Suppression of NRF2 expression shortened life span in mice administered 16 Gy to the thorax. Nfe2l2(+/-) and Nfe2l2(-/-) mice exhibited a mean life span of 176 days compared to wild-type mice, which lived an average of 212 days. These novel results identify NRF2 as a susceptibility factor for the development of late tissue injury.
5-Fluorouracil (5-Fu) is the preferred drug in colorectal cancer treatment. Although 5-Fu treatment contributes to the increase in survival rates, long-term use of 5-Fu causes severe intestinal damage, eventually decreasing long-term survival. There is no standardtreatmentfor intestinal damage induced by 5-Fu. Our previous study found that 5-Fu-induced intestinal damage was connected to an increase in senescent cells, and antiaging drugs could relieve some adverse side effects caused by 5-Fu. Hence, it is essential to discover novel, potential antiaging therapeutic drugs for 5-Fu side effect treatment. According to the current study, Atorvastatincalcium (Ator) alleviated cellular senescence in human intestinal epithelial cells (HUVECs) and human umbilical vein endothelial cells (HIECs) caused by oxidative stress and 5-Fu. 5-Fu resulted in an increase in SA-β-Gal-positive cells, synchronously increased expression of aging-related proteins (p16), aging-related genes (p53, p21), and the senescence-associated secretory phenotype (SASP: IL-1β, IL-6, TNF-α), while Atorvastatincalcium (Ator) reversed the increase in these indicators. In the BALB/c mouse model, we confirmed that intestinal damage caused by 5-Fu is related to the increase in senescent cells and drug-induced inflammation, with the therapeutic effects of Ator. In addition, Ator increased the sensitivity of 5-Fu to chemotherapy in vitro and in vivo. Combination therapy significantly reduced HCT116 cell viability. Furthermore, Ator and 5-Fu present a cooperative effect on preventing the growth of tumors in CRC xenograft nude mice. In conclusion, our study demonstrates the value of Ator for treating intestinal damage. Moreover, Ator combined with 5-Fu increased the antitumor ability in CRC cells. Additionally, we provide a novel therapeutic protocol for CRC.
Anisodamine is an anticholinergic drug extracted and isolated from the Anisodus tanguticus (Maxim.) Pascher of the Solanaceae family which is also a muscarinic receptor antagonist. Owing to the lack of natural sources of anisodamine, synthetic products are now used. Using ornithine and arginine as precursor compounds, putrescine is catalyzed by different enzymes and then undergoes a series of reactions to produce anisodamine. It has been used clinically to protect cardiac function and treat septic shock, acute pancreatitis, calculous renal colic, bronchial asthma, blood circulation disturbances, jaundice, analgesia, vertigo, acute poisoning, and other conditions.This review describes the relevant pharmacokinetic parameters. Anisodamine is poorly absorbed in the gastrointestinal tract, and it is not as effective as intravenous administration. For clinical medication, intravenous infusion should be used rather than rapid intravenous injection. With the advancement of research in recent years, the application scope of anisodamine has expanded, with significant developments and application values surging.This review systematically describes the sources, pharmacokinetics, pharmacological effects and clinical application of anisodamine, in order to provide a basis for clinical use.
Background
Radiation-induced lung injury (RILI) is a common side effect in chest radiotherapy patients, and there is no good medicine to treat it. Re-Du-Ning (RDN) injection is a traditional Chinese medicine that is clinically used to treat upper respiratory tract infections and acute bronchitis. RDN has the advantage of high safety and mild side effects. The mechanism of most traditional Chinese medicine preparations is unknown.
Purpose
To illustrate the mechanisms of RDN for the treatment of RILI.
Methods
Female C57BL/6 mice were used to establish a RILI model via irradiation, and RDN injection was intraperitoneally administered at doses of 5, 10, and 20 mL/kg. The cytokines were measured by ELISA and qPCR. The data related to Absent in melanoma 2 (AIM2) inflammasome were analyzed via ELISA and a network pharmacological approach. In addition, the data related to epithelial-mesenchymal transition (EMT) were analyzed via immunofluorescence, Western blotting, and a network pharmacological approach.
Results
RDN robustly alleviated RILI. Meanwhile, RDN downregulated inflammatory cells’ infiltration and the expression of pro-inflammatory cytokines, such as IL-1β, IL-6, and TNF-α. Next, the potential molecular mechanisms of RDN were predicted through network pharmacology analysis. RDN may ameliorate radiation pneumonitis by inhibiting AIM2-mediated pyroptosis. Moreover, RDN treatment inhibited EMT and phosphoinositide-3-kinase (PI3K)/protein kinase B (Akt) pathway. The active compounds from Lonicera japonica Thunb. decreased the phosphorylation of Akt.
Conclusion
These findings demonstrate that RDN, as a traditional Chinese medicine preparation, will be a candidate drug for treating RILI.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) provoked a pandemic of acute respiratory disease, namely coronavirus disease 2019 (COVID-19). Currently, effective drugs for this disease are urgently warranted. Anisodamine is a traditional Chinese medicine that is predicted as a potential therapeutic drug for the treatment of COVID-19. Therefore, this study aimed to investigate its antiviral activity and crucial targets in SARS-CoV-2 infection. SARS-CoV-2 and anisodamine were co-cultured in Vero E6 cells, and the antiviral activity of anisodamine was assessed by immunofluorescence assay. The antiviral activity of anisodamine was further measured by pseudovirus entry assay in HEK293/hACE2 cells. Finally, the predictions of crucial targets of anisodamine on SARS-CoV-2 were analyzed by molecular docking studies. We discovered that anisodamine suppressed SARS-CoV-2 infection in Vero E6 cells, and reduced the SARS-CoV-2 pseudovirus entry to HEK293/hACE2 cells. Furthermore, molecular docking studies indicated that anisodamine may target SARS-CoV-2 main protease (Mpro) with the docking score of −6.63 kcal/mol and formed three H-bonds with Gly143, Cys145, and Cys44 amino acid residues at the predicted active site of Mpro. This study suggests that anisodamine is a potent antiviral agent for treating COVID-19.
Background
Radiotherapy of thoracic neoplasms and accidental radiation exposure often results in pneumonitis and fibrosis of lungs. Here, we investigated the potential of amifostine analogs: DRDE-07, DRDE-30, and DRDE-35, in alleviating radiation-induced lung damage.
Methods
C57BL/6 mice were exposed to 13.5 Gy thoracic irradiation, 30 min after intraperitoneal administration of the analogs, and assessed for modulation of the pathological response at 12 and 24 weeks.
Key findings
DRDE-07, DRDE-30 and DRDE-35 increased the survival of irradiated mice from 20% to 30%, 80% and 70% respectively. Reduced parenchymal opacity (X-ray CT) in the lungs of DRDE-30 pre-treated mice corroborated well with the significant decrease in Ashcroft score (p < 0.01). Two-fold increase in SOD and catalase activities (p < 0.05), coupled with a 50% increase in GSH content and a 60% decrease in MDA content (p < 0.05) suggested restoration of the antioxidant defence system. A 20% to 40% decrease in radiation-induced apoptotic and mitotic death in the lung tissue (micronuclei: p < 0.01), resulted in attenuated lung and vascular permeability (FITC-Dextran leakage) by 50% (p < 0.01), and a commensurate reduction (~50%) in leukocyte infiltration in the injured tissue (p < 0.05). DRDE-30 abrogated the activation of pro-inflammatory NF-κB and p38/MAPK signaling cascades, suppressing the release of pro-inflammatory cytokines (IL-1β: p < 0.05; TNF-α: p < 0.05; IL-6: p < 0.05) and up-regulation of CAMs on the endothelial cell surface. Reduction in hydroxyproline content (p < 0.01) and collagen suggested inhibition of lung fibrosis which was associated with attenuation of TGF-β/Smad pathway-mediated-EMT.
Conclusion
DRDE-30 could be a potential prophylactic agent against radiation-induced lung injury.
Background
Radiation exposure to lungs during nuclear catastrophes or radiotherapy poses long-term side effects and can induce pulmonary injury sufficient for causing death. The strategies for preventing or reversing radiation-induced lung injuries have not been yet developed. Quercetin-3-Rutinoside (Q-3-R), a polyphenolic bioflavonoid, has shown multifaceted pharmacological applications due to its high antioxidant and anti-inflammatory properties.
Purpose
In the current study, the potential of Q-3-R against radiation-induced lung pneumonitis/fibrosis and the possible underlying mechanism was investigated.
Study design
To evaluate the effect of Q-3-R against lung damage, C57Bl/6 mice were administered with Q-3-R (10 mg/kg b.wt.) and irradiated with a single dose of gamma radiation (12 Gy) at thoracic region.
Methods
16 weeks after irradiation lung damage was seen by histopathological studies and staining for collagen deposition. Expression of Nuclear factor kappa-B (NF-κB), transforming growth factor-β1 (TGF-β1), Smad3, intercellular adhesion molecule 1 (ICAM-1), α-smooth muscle actin protein (α-SMA), Aquaporin 5 (AQP 5), Interleukins (IL-6, IL-18, IL-1β), Tumour necrosis factor-α (TNF-α) and caspase-3 was evaluated by immunohistochemistry/western blot/Elisa. Reactive oxygen species (ROS)/ Nitric oxide (NO) scavenging potential of Q-3-R and inhibition of cell death in irradiated lungs were also assessed.
Results
Mice showed signs of pneumonitis and fibrotic changes in lungs following radiation treatment. A dramatic increase in inflammatory cells and cytokines contributing to lung disease pathogenesis was observed. Furthermore, expression of NF-κB, TGF-β1, Smad3, ICAM-1, AQP5and α-SMA was found markedly up-regulated. However, pretreatment of Q-3-R significantly attenuated radiation-induced pneumonitis and fibrosis. Histological examination revealed less structural and fibrotic changes with down-regulation of AQP 5, ICAM-1, α-SMA and caspase-3 in Q-3-R pretreated irradiated groups. The formulation significantly relieved lung injury by suppressing inflammatory and pro-fibrotic cytokines such as IL-6, IL-18, IL-1β, TNF-α and TGF-β1 via inhibition of NF-κB. Q-3-R also curtailed radiation-induced ROS/NO generation and minimized DNA damage in the irradiated lungs.
Conclusion
The findings from the current study clearly demonstrate that Q-3-R provides radioprotection to the lungs by regulating NF-κB/TGF-β1 signaling, scavenging free radicals, preventing perivascular infiltration and prolonged inflammatory cascade which could otherwise lead to chronic radiation fibrosis. Q-3-R can be proved as a potential therapeutic agent for alleviating radiation-induced lung injury in case of planned or unplanned radiation exposure scenario.
Cerebrovascular disease and its risk factors cause persistent decrease of cerebral blood flow, chronic cerebral hypoperfusion (CCH) is the major foundation of vascular cognitive impairment (VCI). The hippocampus is extremely vulnerable to cerebral ischemia and hypoxia. Oxidative stress and neuroinflammation injury are important pathophysiological mechanisms of this process, which is closely related to hippocampal neurons damage and loss. Dimethyl fumarate (DMF), an FDA-approved therapeutic for multiple sclerosis (MS), plays a protective role in multiple neurological disorders. Studies have shown that DMF exerts anti-inflammatory and antioxidant effects via the NRF2/ARE/NF-κB signaling pathway. Thus, this study aimed to evaluate the neuroprotective effect of DMF in the CCH rat model. Ferroptosis, a novel defined iron-dependent cell death form, were found to be strongly associated with the pathophysiology of CCH. Emerging evidences have shown that inhibition of ferroptosis by targeting NRF2 exerted neuroprotective effect in neurodegeneration diseases. We also investigated whether DMF can alleviate cognitive deficits through inhibition of ferroptosis by the NRF2 signaling pathway in this study. DMF was intragastric for consecutive five weeks (100 mg/kg/day). Then behavior test and histological, molecular, and biochemical analysis were performed. We found that DMF treatment significantly improved cognitive deficits and partially reversed hippocampus neuronal damage and loss caused by CCH. And DMF treatment decreased hippocampus IL-1β, TNF-α, and IL-6 pro-inflammatory cytokines concentration, and mediated the NF-κB signaling pathway. And DMF also alleviated hippocampus oxidative stress through reducing MDA, and increasing GSH and SOD levels, which are also closely associated with ferroptosis. Besides, DMF treatment reduced the expression of PTGS2, and increased the expression of FTH1 and xCT, and the iron content is also reduced, which were the important features related to ferroptosis. Furthermore, DMF activated the NRF2/ARE signaling pathway and upregulated the expression of HO-1, NQO1 and GPX4. These outcomes indicated that DMF can improve cognitive impairment in rats with CCH, possibly through alleviating neuroinflammation, oxidative stress damage and inhibiting ferroptosis of hippocampal neurons. Overall, our results provide new evidence for the neuroprotective role of DMF.
Purpose
Radiation therapy is one of the main cancer treatment modalities applied in 50–70% of cancer patients. Despite the many advantages of this treatment, such as non-invasiveness, organ-preservation, and spatiotemporal flexibility in tumor targeting, it can lead to complications in irradiated healthy cells/tissues. In this regard, the use of radio-protective agents can alleviate radiation-induced complications. This study aimed to review the potential role of alpha-lipoic acid in the prevention/reduction of radiation-induced toxicities on healthy cells/tissues.
Methods
A systematic search was performed following PRISMA guidelines to identify relevant literature on the “role of alpha-lipoic acid in the treatment of radiotherapy-induced toxicity” in the electronic databases of Web of Science, Embase, PubMed, and Scopus up to January 2021. Based on the inclusion and exclusion criteria of the present study, 278 articles were screened. Finally, 29 articles were included in this systematic review.
Results
The obtained results showed that in experimental in vivo models, the radiation-treated groups had decreased survival rate and body weight compared to the control groups. It was also found that radiation can induce mild to severe toxicities on gastrointestinal, circulatory, reproductive, central nervous, respiratory, endocrine, exocrine systems, etc. However, the use of alpha-lipoic acid could alleviate the radiation-induced toxicities in most cases. This radio-protective agent exerts its effects through mechanisms of anti-oxidant, anti-apoptosis, anti-inflammatory, and so on.
Conclusion
According to the obtained results, it can be mentioned that co-treatment of alpha-lipoic acid with radiotherapy ameliorates the radiation-induced toxicities in healthy cells/tissues.
Aims
Dysregulation of iron homeostasis in the body causes a variety of diseases. Iron deficiency leads to anemia, whereas iron overload aggravates cellular oxidative stress. Nuclear factor erythroid-2-related factor 2 (Nrf2) is a protein that is activated in the nucleus and turns on the production of antioxidant enzymes, protecting cell against oxidative damage. This study aimed to investigate whether Nrf2 gene knockout influences iron homeostasis in aging mice.
Materials and methods
Iron content and iron metabolism-related proteins were assessed in different organs and blood serum of the 18 month-old Nrf2 knockout (Nrf2−/−) mice in comparison with the wild-type (WT) mice.
Key findings
Results showed that the iron contents in spleen and liver all increased, and expression levels of iron transporters were altered in Nrf2−/− mice. In particularly, we found that the expression of iron export protein ferroportin 1 (Fpn1) in liver, spleen and small intestine all decreased in Nrf2−/− mice, which might account for the deposition of iron in different organs and the increased ROS. Surprisingly, we found that the serum iron level of Nrf2−/− mice did not decrease, but increased significantly even when the iron absorption at small intestine decreased. Our further investigation revealed that the increase of serum iron was due to the release of iron from the hemolysis of erythrocytes, which caused by the increased ROS level in red blood cells of the Nrf2−/− mice.
Significance
These findings provide a more comprehensive understanding of the important role of Nrf2 in the regulation of systemic iron metabolism.
Aims
Radiation-induced lung injury (RILI) is a serious complication of radiation therapy. Development of an effective drug that selectively protects normal lung tissues and sensitizes tumor cells to radiotherapy is an unmet need. 2-Methoxyestradiol (2ME2) possesses polypharmacological properties, which qualifies it as an effective radioprotector. Our aim is to explore the potential protective effects of 2ME2 against early and late stages of RILI and the underlying mechanisms.
Main methods
BALB/c mice were either treated with 2ME2 (50 mg/kg/day i.p., for 4 weeks); or received a single dose of 10 Gy ionizing radiation (IR) delivered to the lungs; or 10 Gy IR and 2ME2. Animal survival and pulmonary functions were evaluated. Immune-phenotyping of alveolar macrophages (AM) in the broncho-alveolar lavage fluids (BALF) was determined by flow cytometry. ELISA was used to evaluate the expression levels of TNF-α, TGF-β; and IL-10 in BALF. Lung tissues were used for histopathological examination or immunofluorescence staining for CD68 (pan-macrophage marker), Arginase-1 (Arg1, M2-specific marker), inducible nitric oxide synthase (iNOS, M1-specific marker) and HIF-1α. VEGF and γH2AX expression in lung tissues were detected by western blot.
Key findings
The results demonstrated that 2ME2 improved the survival, lung functions and histopathological parameters of irradiated mice. Additionally, it attenuated the radiation-induced AM polarization and reduced the pneumonitis and fibrosis markers in lung tissues. Significant reduction of TNF-α and TGF-β with concomitant increase in IL-10 concentrations were observed. Moreover, the expression of HIF-1α, VEGF and γH2AX declined.
Significance
2ME2 is a promising radioprotectant with fewer anticipated side effects.
Purpose: Anisodamine (An) has anti-inflammatory effects, but its role in acute pancreatitis is still unknown. This study aimed to explore the action mechanism of An pretreatment in lipopolysaccharide (LPS)-induced pancreatic acinar cells, hoping to provide a research basis for the disease treatment.
Materials and methods: Pancreatic acinar cells were pretreated with An at different concentrations and then induced by LPS. The viability and apoptosis of the treated cells were measured by Cell Counting Kit-8 and flow cytometry. The releases of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and IL-18 were measured by enzyme-linked immunosorbent assay. The expressions of thioredoxin-interacting protein (TXNIP), apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), NOD-like receptor protein 3 (NLRP3), Caspase-1, p65, and inhibitor of kappa B alpha (IκBα) in the treated cells were detected by Western blot and quantitative real-time polymerase chain reaction assay.
Results: LPS promoted apoptosis of pancreatic acinar cells, suppressed cell viability, increased TNF-α, IL-1β, and IL-18 releases and the expression levels of TXNIP, ASC, NLRP3, Caspase-1, p-p65, and p-IκBα, however, such effects of LPS could be alleviated by An pretreatment with the strongest effect when the concentration of An was set at 100 μg/mL. Moreover, overexpressed NLRP3 aggravated the effects of LPS in pancreatic acinar cells, which could be reversed by pretreatment of 100 μg/mL An.
Conclusion: An pretreatment attenuated LPS-induced apoptosis and inflammatory response of pancreatic acinar cells through suppressing NLRP3 and inactivating NF-κB signaling pathway, thus, it could be explored as a potential therapy for treating acute pancreatitis.
Oxidative stress-induced cellular senescence and inflammation are important biological events in diabetic nephropathy (DN). Our recent studies have found that pyrroloquinoline quinine (PQQ) has protective effects against HG-induced oxidative stress damage and apoptosis in HK-2 cells. Nevertheless, whether PPQ has the effect of anti-inflammation and anti-senescence in HK-2 cells remains unclear. Here, we showed that low-dose PPQ treatment (100 nM) downregulates the expression of P16, P21, IL-1β, TNF-α and NF-κB in HG cultured HK-2 cells. A low dose of PPQ also upregulated the protein expression of SOD2, CAT and inhibited the generation of ROS. We also indicated that PPQ affected the activity of Keap1/Nrf2 pathway, increased the nuclear accumulation of Nrf2 and the downstream pathway protein expression of Keap1/Nrf2 signaling pathway (HO-1, NQO-1, GST and GPx-3). When ML385 was added to inhibit the activity of Keap1/Nrf2 signaling pathway, the effects of PPQ on anti-oxidative stress, anti-inflammation and anti-senescence in HK-2 cells under HG condition were weakened. In conclusion, our results suggest that PPQ could modulate HG-induced inflammation and senescence in HK-2 cells via the inhibition of ROS generation and achieves the protective effects through Keap1/Nrf2 pathway and upregulating the expression of its target protein.
Aims:
Endoplasmic reticulum stress (ERS) is an evolutionarily conserved cell stress response. Recently, it was found that ERS induces not only apoptosis but also endoplasmic reticulophagy (ER-phagy). A previous study demonstrated that inhibition of ER-phagy alleviates cell injury. The purpose of this study was to investigate the involvement of the protein kinase R-like ER kinase (PERK)/nuclear factor erythroid 2-related factor 2 (Nrf2) pathway in ERS-induced ER-phagy in H9c2 cardiomyoblasts. To address this aim, cells were treated with ERS inhibitors and a Nrf2 inhibitor before establishment of thapsigargin (TG)- or tunicamycin (TM)-induced ERS models in H9c2 cardiomyoblasts.
Main methods:
Transmission electron microscopy and immunofluorescence staining were used to detect ER-phagy. Western blotting was employed to detect the levels of calreticulin (CRT), total and phosphorylated PERK, nuclear Nrf2, activated transcription factor 4 (ATF4), light chain 3B (LC3B)-II and Beclin 1. Immunofluorescence staining was used to assess subcellular location of Nrf2.
Key finding:
TG or TM induced H9c2 cell injury and ER-phagy and upregulated CRT expression, PERK phosphorylation, Nrf2 nuclear translocation, and expression of ATF4, Beclin 1, and LC3B-II compared with control cells. Treatment with ERS inhibitors decreased TG- or TM-induced ER-phagy, downregulated CRT expression, PERK phosphorylation, Nrf2 nuclear translocation and the expression of ATF4, Beclin 1 and LC3B-II. Moreover, a Nrf2 inhibitor downregulated the expression of ATF4, Beclin 1 and LC3B-II and alleviated TG- or TM-induced ER-phagy and H9c2 cell injury.
Significance:
These findings suggest that the PERK/Nrf2 pathway mediates upregulation of ER-phagy, thereby inducing cell injury in H9c2 cardiomyoblasts.
Intervertebral disc degeneration (IVDD) is a prevalent disease characterized by the progressive loss of the extracellular matrix in the local nucleus pulposus region. Metalloproteinases and pro-inflammatory cytokines play an important role in this process. Thus, anti-inflammatory strategies are an important component of IVDD treatment. Wogonin, a naturally existing monoflavonoid, has been reported to have potential anti-inflammatory effects in some inflammatory diseases. Hence, in our present study we investigated the protective effects and potential mechanisms of wogonin in rat nucleus pulposus cells that had been treated with interleukin-1beta (IL-1β) to induce severe IVDD. An in vivo rat caudal vertebrae needle-stab model was also designed and its validity was evaluated as an IVDD model. The results demonstrated that wogonin suppressed IL-1β-induced inflammatory mediators (iNOS, IL-6 and COX2) and matrix-degrading proteinases (MMP1, MMP3, MMP13 and ADAMTS4). Wogonin also upregulated some of the key components of the extracellular matrix, such as collagen II. Furthermore, we discovered that wogonin exerted anti-inflammatory effects by activating the Nrf2/HO-1-SOD2-NQO1-GCLC signaling axis. Moreover, the IL-1β-induced stimulation of the MAPK signaling pathway was reversed by wogonin treatment. The in vivo MRI and histological results also revealed that wogonin protected the nucleus pulposus from the progression of IVDD. Therefore, wogonin may be a potential agent for the treatment of IVDD.
Models of thoracic irradiation have been developed as clinicians and scientists have attempted to decipher the events that led up to the pulmonary toxicity seen in human subjects following radiation treatment. The most common model is that of whole thorax irradiation (WTI), applied in a single dose. Mice, particularly the C57BL/6J strain, has been frequently used in these investigations, and has greatly informed our current understanding of the initiation and progression of radiation-induced lung injury (RILI). In this review, we highlight the sequential progression and dynamic nature of RILI, focusing primarily on the vast array of information that has been gleaned from the murine model. Ample evidence indicates a wide array of biological responses that can be seen following irradiation, including DNA damage, oxidative stress, cellular senescence and inflammation, all triggered by the initial exposure to ionizing radiation (IR) and heterogeneously maintained throughout the temporal progression of injury, which manifests as acute pneumonitis and later fibrosis. It appears that the early responses of specific cell types may promote further injury, disrupting the microenvironment and preventing a return to homeostasis, although the exact mechanisms driving these responses remains somewhat unclear. Attempts to either prevent or treat RILI in preclinical models have shown some success by targeting these disparate radiobiological processes. As our understanding of the dynamic cellular responses to radiation improves through the use of such models, so does the likelihood of preventing or treating RILI.
Aim:
The constitutive expression of the Nrf2 genes during development and aging in both sexes would help our understanding of the Nrf2/Keap1 pathway in toxicological studies.
Main methods:
Sprague Dawley rat livers were collected at 11 age points from prenatal (-2d), neonatal (1, 7, 14 and 21d), at puberty (28 and 35d), at adulthood (60 and 180d), to aging (540 and 800d) from both sexes. Total RNA and proteins were extracted for real-time RT-PCR and Western-blot analysis.
Key findings:
The abundant mRNA expression was in the order of Nrf2, Gclm, Nqo1, Gclc, Ho-1, and Keap1. The expression of these genes except Gclc was high in fetal livers, decreased at birth, reached the first peak at 7 days of age, and gradually decreased to adult levels till 180 days of age. All these genes remained high at 540 days of age, but declined at 800 days of age, with more increases with Nqo1 and Ho-1. Females had lower fetal, neonatal, and aged levels than males. Protein expressions of Nrf2, Nqo1, Ho-1, GCLC and GCLM agree with mRNA analysis.
Significance:
This study characterized the age- and sex-related changes of Nrf2-related gene/proteins in livers of rats, and higher expressions in newborns and aged rats could cope with increased oxidative stress in infants and elderly.
The mechanisms of aging that are involved in the development of idiopathic pulmonary fibrosis (IPF) are still unclear. Although it has been hypothesized that the proliferation and activation of human lung fibroblasts (hLF) are essential in IPF, no studies have assessed how this process works in an aging lung. Our goal was to elucidate if there were age-related changes on primary hLF isolated from IPF lungs compared to age-matched controls. We investigated several hallmarks of aging in hLF from IPF patients and age-matched controls. IPF hLF have increased cellular senescence with higher expression of β-galactosidase, p21, p16, p53 and cytokines related to the senescence-associated secretory phenotype (SASP) as well as decreased proliferation/apoptosis compared to age-match controls. Additionally, we observed shorter telomeres, mitochondrial dysfunction, and upon TGF-β stimulation, increased markers of ER stress. Our data suggests that IPF hLF develop senescence resulting in a decreased apoptosis and that the development of SASP may be an important contributor to the fibrotic process observed in IPF. These results might change the existing paradigm, which describes fibroblasts as aberrantly activated cells, to a cell with a senescence phenotype.
Purpose:
Ionizing radiation (IR)-induced pulmonary fibrosis (PF) is an irreversible and severe late effect of thoracic radiation therapy. The goal of this study was to determine whether clearance of senescent cells with ABT-263, a senolytic drug that can selectively kill senescent cells, can reverse PF.
Methods and materials:
C57BL/6J mice were exposed to a single dose of 17 Gy on the right side of the thorax. Sixteen weeks after IR, they were treated with 2 cycles of vehicle or ABT-263 (50 mg/kg per day for 5 days per cycle) by gavage. The effects of ABT-263 on IR-induced increases in senescent cells; elevation in the expression of selective inflammatory cytokines, matrix metalloproteinases, and tissue inhibitors of matrix metalloproteinases; and the severity of the tissue injury and fibrosis in the irradiated lungs were evaluated 3 weeks after the last treatment, in comparison with the changes observed in the irradiated lungs before treatment or after vehicle treatment.
Results:
At 16 weeks after exposure of C57BL/6 mice to a single dose of 17 Gy, thoracic irradiation resulted in persistent PF associated with a significant increase in senescent cells. Treatment of the irradiated mice with ABT-263 after persistent PF had developed reduced senescent cells and reversed the disease.
Conclusions:
To our knowledge, this is the first study to demonstrate that PF can be reversed by a senolytic drug such as ABT-263 after it becomes a progressive disease. Therefore, ABT-263 has the potential to be developed as a new treatment for PF.
Purpose:
Radiation-induced pulmonary fibrosis (RIPF) is a late toxicity of therapeutic radiation. Signaling of the mammalian target of rapamycin drives several processes implicated in RIPF, including inflammatory cytokine production, fibroblast proliferation, and epithelial senescence. We sought to determine if mammalian target of rapamycin inhibition with rapamycin would mitigate RIPF.
Methods and materials:
C57BL/6NCr mice received a diet formulated with rapamycin (14 mg/kg food) or a control diet 2 days before and continuing for 16 weeks after exposure to 5 daily fractions of 6 Gy of thoracic irradiation. Fibrosis was assessed with Masson trichrome staining and hydroxyproline assay. Cytokine expression was evaluated by quantitative real-time polymerase chain reaction. Senescence was assessed by staining for β-galactosidase activity.
Results:
Administration of rapamycin extended the median survival of irradiated mice compared with the control diet from 116 days to 156 days (P=.006, log-rank test). Treatment with rapamycin reduced hydroxyproline content compared with the control diet (irradiation plus vehicle, 45.9 ± 11.8 μg per lung; irradiation plus rapamycin, 21.4 ± 6.0 μg per lung; P=.001) and reduced visible fibrotic foci. Rapamycin treatment attenuated interleukin 1β and transforming growth factor β induction in irradiated lungs compared with the control diet. Type II pneumocyte senescence after irradiation was reduced with rapamycin treatment at 16 weeks (3-fold reduction at 16 weeks, P<.001).
Conclusions:
Rapamycin protected against RIPF in a murine model. Rapamycin treatment reduced inflammatory cytokine expression, extracellular matrix production, and senescence in type II pneumocytes.
Introduction:
To investigate the impact of pre-existing radiological interstitial lung disease (ILD) findings on the incidence of radiation pneumonitis (RP) and clinical outcomes after stereotactic body radiation therapy (SBRT) for stage I non-small-cell lung cancer.
Methods:
We included 157 consecutive patients who underwent SBRT alone for stage I non-small-cell lung cancer and whose pretreatment lung computed tomography images were available for retrospective review. The pretreatment computed tomography images were evaluated retrospectively for the presence of ILD. The incidence of RP, overall survival (OS) rate, and the incidence of disease progression and local progression were evaluated between patients with ILD (ILD[+]) and without ILD (ILD[-]).
Results:
Pre-existing ILD was identified in 20 patients. The median follow-up period was 39.5 months. The incidences of RP worse than grade 2 (≥ Gr2 RP) and worse than grade 3 (≥ Gr3 RP) were significantly higher in ILD(+) than ILD(-) (1 year ≥ Gr2 RP rate, 55.0% versus 13.3%; p < 0.001 and 1year ≥ Gr3 RP rate 10.0% versus 1.5%; p = 0.020). Multivariate analysis also indicated that ILD(+) was a risk factor for ≥ Gr2 and ≥ Gr3 RP, and the volume of the irradiated lung. The OS rate tended to be worse in ILD(+) than ILD(-) (3-year OS, 53.8% versus 70.8%; p = 0.28). No difference was observed in the disease progression or local progression rates.
Conclusions:
Pre-existing ILD was a significant risk factor for symptomatic and severe RP. Prescreening for ILD findings is important for determining the radiation pneumonitis risk when planning SBRT.
There are various types of autophagy, which can be categorized as nonselective or selective. Macroautophagy is an evolutionarily conserved process through which cells degrade and recycle cytoplasm. Nonselective macroautophagy randomly engulfs a portion of the cytoplasm into autophagosomes and then delivers them to the vacuole (in fungi or plants) or the lysosome (in other higher eukaryotes) for degradation. Selective macroautophagy, however, specifically recognizes and degrades a particular cargo, either a protein complex, an organelle, or an invading microbe. The morphological hallmark of macroautophagy is the formation of an initial sequestering compartment, the phagophore, which expands into the double-membrane autophagosome; the initial sequestration occurs in a compartment that is separate from the degradative organelle. Selective microautophagy utilizes the same cellular machinery, but in this case, the sequestration event takes place directly at the limiting membrane of the lysosome/vacuole. In higher eukaryotes, selective types of autophagy also include chaperone-mediated autophagy (CMA), and two similar processes, endosomal microautophagy (e-MI) and chaperone-assisted selective autophagy (CASA), each of which involves uptake at the limiting membrane of either the lysosome or endosome. In all cases, how a substrate is targeted for sequestration and segregated from other parts of the cell is one of the major questions in this research field. Nonselective autophagy is primarily a starvation response, whereas cells use selective autophagy for a variety of purposes, including remodeling to adapt to changing environmental/nutritional conditions and to eliminate damaged organelles. Accordingly, defects in selective autophagy are associated with a range of pathophysiologies in humans, including certain types of neurodegenerative diseases.
Although anisodamine, a muscarinic acetylcholine receptor antagonist, has been used in China for treating various shocks for many years, the mechanisms are not well understood. Our previous studies have demonstrated anisodamine exerts its cholinergic anti-inflammatory action through indirectly activating α7 nicotinic acetylcholine receptors (α7 nAChR). Because IL-10 is a critical anti-inflammatory factor, we investigated its potential role in the antishock action of anisodamine.
C57BL/6 and IL-10 -/- mice were intraperitoneally administered LPS and/or anisodamine, and the 24h survival rate, cytokine production and α7 nAChR expression were examined. In addition, RAW264.7 cells were stimulated with LPS, anisodamine and/or IL-10, and cytokine production and α7 nAChR expression were investigated.
Anisodamine dose-dependently increased the 24h survival rate of C57BL/6 mice treated with LPS. The antishock role of anisodamine was significantly attenuated in IL-10 -/- mice. Anisodamine significantly decreased TNF-α and IL-1β production in LPS-treated RAW264.7 cells and C57BL/6 mice. However, it did not increase the level of IL-10 in the same experiments. In RAW264.7 cells, IL-10 treatment increased α7 nAChR expression, which was further augmented in the presence of anisodamine. Spleens from IL-10 -/- mice expressed significantly lower α7 nAChRs levels compared to IL-10 +/+ mice. Although anisodamine markedly increased the expression of α7 nAChRs in spleens from LPS-treated IL-10 +/+ mice, it only induced a marginal increase of the receptor in spleens from LPS-treated IL-10 -/- mice.
These findings demonstrate that IL-10 plays an important role in the antishock action of anisodamine. It acts through upregulating α7 nAChR synergistically with anisodamine.