Modulation of Radiation-Induced NADPH Oxidases in Rat's Heart Tissues by Melatonin.

BACKGROUND: Experimental studies have shown that infiltration of inflammatory cells as well as upregulation of some cytokines play a central role in the development of late effects of ionizing radiation in heart tissues. Evidences have shown that an increased level of TGF-ß has a direct correlation with late effects of exposure to ionizing radiation such as chronic oxidative stress and fibrosis. Recent studies have shown that TGF-ß, through upregulation of pro-oxidant enzymes such as NOX2 and NOX4, promotes continuous ROS production and accumulation of fibrosis. OBJECTIVE: In present study, we aimed to evaluate the expression of NOX2 and NOX4 signaling pathways as well as possible modulatory effects of melatonin on the expression of these genes. MATERIAL AND METHODS: In this experimental study, four groups of 20 rats (5 in each) were used as follows; G1: control; G2: melatonin; G3: radiation; G4: radiation + melatonin. 100 mg/kg of melatonin was administrated before irradiation of heart tissues with 15 Gy gamma rays. 10 weeks after irradiation, heart tissues were collected for real-time Polymerase chain reaction (PCR). RESULTS: Results showed a significant increase in the expression of TGF-ß, Smad2, NF-kB, NOX2 and NOX4. The upregulation of NOX2 was more obvious by 20-fold compared to other genes. Except for TGF-ß, melatonin could attenuate the expression of other genes. CONCLUSION: This study indicated that exposure of rat's heart tissues to radiation leads to upregulation of TGF-ß-NOX4 and TGF-ß-NOX2 pathways. Melatonin, through modulation of these genes, may be able to alleviate radiation-induced chronic oxidative stress and subsequent consequences.

The interactions and communications in tumor resistance to radiotherapy: Therapy perspectives.

Tumor microenvironment (TME) includes a wide range of cell types including cancer cells, cells which are involved in stromal structure and immune cells (tumor suppressor and tumor promoting cells). These cells have several interactions with each other that are mainly regulated via the release of intercellular mediators. Radiotherapy can modulate these interactions via shifting secretions into inflammatory or anti-inflammatory responses. Radiotherapy also can trigger resistance of cancer (stem) cells via activation of stromal cells. The main mechanisms of tumor resistance to radiotherapy is the exhaustion of anti-tumor immunity via suppression of CD4+ T cells and apoptosis of cytotoxic CD8+ T lymphocytes (CTLs). Cancer-associated fibroblasts (CAFs), mesenchymal-derived suppressor cells (MDSCs) and regulatory T cells (Tregs) are the main suppressor of anti-tumor immunity via the release of several chemokines, cytokines and immune suppressors. In this review, we explain the main cellular and molecular interactions and secretions in TME following radiotherapy. Furthermore, the main signaling pathways and intercellular connections that can be targeted to improve therapeutic efficiency of radiotherapy will be discussed.

Damage-associated molecular patterns in tumor radiotherapy.

Radiotherapy is one of the most common modalities for the treatment of cancer. One of the most promising effects of radiotherapy is immunologic cell death and the release of danger alarms, which are known as damage-associated molecular patterns (DAMPs). DAMPs are able to trigger cancer cells and other cells within tumor microenvironment (TME), either for suppression or promotion of tumor growth. Heat shock proteins (HSPs) including HSP70 and HSP90, high mobility group box 1 (HMGB1), and adenosine triphosphate (ATP) and its metabolites such as adenosine are the most common danger alarms that are released after radiotherapy-induced immunologic cell death. Some DAMPs including adenosine is able to interact with both cancer cells as well as other cells in TME to promote tumor growth and resistance to radiotherapy. However, others are able to trigger anti-tumor immunity or both tumor suppressive and immunosuppressive mechanisms depending on affected cells. In this review, we explain the mechanisms behind the release of radiation-induced DAMPs, and its consequences on cells within tumor. Targeting of these mechanisms may be in favor of tumor control in combination with radiotherapy and radioimmunotherapy.

Abscopal effect in radioimmunotherapy.

Abscopal effect is an interesting phenomenon in radiobiology that causes activation of immune system against cancer cells. Traditionally, this phenomenon was known as a suppressor of non-irradiated tumors or metastasis. However, it can be used as a stimulator of the immune system against primary tumor during radiotherapy. Immunotherapy, a novel tumor therapy modality, also triggers immune system against cancer. To date, some immunotherapy types have been developed. However, immune checkpoint blockade is a more common modality and some drugs have been approved by the FDA. Studies have shown that radiotherapy or immunotherapy administered alone have low efficiency for tumor control. However, their combination has a more potent anti-tumor immunity. For this aim, it is important to induce abscopal effect in primary tumors, and also use appropriate drugs to target the mechanisms involved in the exhaustion of cytotoxic CD8+T lymphocytes (CTLs) and natural killer (NK) cells. Among the different radiotherapy techniques, stereotactic body radiation therapy (SBRT) with some few fractionations is the best choice for inducing abscopal effect. On the other hand, programmed cell death 1 (PD-1) is known as one of the best targets for triggering anti-tumor immunity. This combination is known as the best choice among various strategies for radioimmunotherapy. However, there is the need for other strategies to improve the duration of immune system's activity within tumor microenvironment (TME). In this review, we explain the cellular and molecular mechanisms behind abscopal effect by radiotherapy and evaluate the molecular targets which induce potent anti-tumor immunity.

TGF-ß in radiotherapy: Mechanisms of tumor resistance and normal tissues injury.

Emerging evidences show that changes in tumor stroma can adapt cancer cells to radiotherapy, thereby leading to a reduction in tumor response to treatment. On the other hand, radiotherapy is associated with severe reactions in normal tissues which limit the amount radiation dose received by tumor. These challenges open a window in radiobiology and radiation oncology to explore mechanisms for improving tumor response and also alleviate side effects of radiotherapy. Transforming growth factor beta (TGF-ß) is a well-known and multitasking cytokine that regulates a wide range of reactions and interactions within tumor and normal tissues. Within tumor microenvironment (TME), TGF-ß is the most potent suppressor of immune system activity against cancer cells. This effect is mediated through stimulation of CD4+ which differentiates to T regulatory cells (Tregs), infiltration of fibroblasts and differentiation into cancer associated fibroblasts (CAFs), and also polarization of macrophages to M2 cells. These changes lead to suppression of cytotoxic CD8 + T lymphocytes (CTLs) and natural killer (NK) cells to kill cancer cells. TGF-ß also plays a key role in the angiogenesis, invasion and DNA damage responses (DDR) in cancer cells. In normal tissues, TGF-ß triggers the expression of a wide range of pro-oxidant and pro-fibrosis genes, leading to fibrosis, genomic instability and some other side effects. These properties of TGF-ß make it a potential target to preserve normal tissues and sensitize tumor via its inhibition. In the current review, we aim to explain the mechanisms of upregulation of TGF-ß and its consequences in both tumor and normal tissues.

Genomic Instability and Carcinogenesis of Heavy Charged Particles Radiation: Clinical and Environmental Implications.

One of the uses of ionizing radiation is in cancer treatment. The use of heavy charged particles for treatment has been introduced in recent decades because of their priority for deposition of radiation energy in the tumor, via the Bragg peak phenomenon. In addition to medical implications, exposure to heavy charged particles is a crucial issue for environmental and space radiobiology. Ionizing radiation is one of the most powerful clastogenic and carcinogenic agents. Studies have shown that although both low and high linear energy transfer (LET) radiations are carcinogenic, their risks are different. Molecular studies have also shown that although heavy charged particles mainly induce DNA damage directly, they may be more potent inducer of endogenous generation of free radicals compared to the low LET gamma or X-rays. It seems that the severity of genotoxicity for non-irradiated bystander cells is potentiated as the quality of radiation increases. However, this is not true in all situations. Evidence suggests the involvement of some mechanisms such as upregulation of pro-oxidant enzymes and change in the methylation of DNA in the development of genomic instability and carcinogenesis. This review aimed to report important issues for genotoxicity of carcinogenic effects of heavy charged particles. Furthermore, we tried to explain some mechanisms that may be involved in cancer development following exposure to heavy charged particles.

Selenium-L-methionine modulates radiation injury and Duox1 and Duox2 upregulation in rat's heart tissues.

Introduction: Redox interactions play a key role in radiation injury including heart diseases. In present study, we aimed to detect the possible protective role of selenium-L-methionine on infiltration of immune cells and Duox1&2 upregulation in rat's heart tissues. Methods: In this study, 20 rats were divided into 4 groups (5 rats in each) namely: irradiation; irradiation plus Selenium-L-methionine; control; and Selenium-L-methionine treatment. Irradiation (15 Gy to chest) was performed using a cobalt-60 gamma ray source while 4 mg/kg of selenium-L-methionine was administered intraperitoneally. Ten weeks after irradiation, rats were sacrificed for detection of IL-4 and IL-13 cytokines, infiltration of macrophages and lymphocytes as well as the expressions of IL4Ra1, Duox1, IL13Ra2 and Duox2. Results: Results showed an increase in the level of IL-4 as well as the expressions of IL4Ra1, Duox1 and Duox2. Similarly, there was an increase in the infiltration of lymphocytes and macrophages. There was significant attenuation of all these changes following treatment with selenium-L-methionine. Conclusion: Selenium-L-methionine has the potential to protect heart tissues against radiation injury. Downregulation of pro-oxidant genes and modulation of some cytokines such as IL-4 are involved in the radioprotective effect of selenium-L-methionine on heart tissues.

Prevalence of high blood pressure in Iranian adults based on the 2017 ACC/AHA guideline.

Background: : In 2017, American College of Cardiology (ACC) and the American Heart Association (AHA) presented a new guideline for assessing blood pressure in adults. This study aimed to assess the prevalence of hypertension in Iranian adults based on ACC/AHA 2017 guideline. Methods: Data from 9801 Iranian adults (59.2% women) aged between 20-69 years were obtained from the sixth round of National Surveillance of Risk Factors of Non-Communicable Diseases (SuRFNCD) performed in 2011. Blood pressure was classified as normal, elevated blood pressure, and stage 1 and 2 hypertension using a weighted analysis and 2017 ACC/AHA guidelines. Data were presented as prevalence and 95% confidence interval (95% CI). All analyses were performed in Stata/SE 14.0. Results: Overall prevalence of hypertension in Iranian men was 52.0%. Also, 32.9% (95% CI: 29.9-36.0) and 19.1% (95% CI: 16.9-21.6) of men had stage 1 and 2 hypertension, respectively. In addition, 44.3% of women had hypertension, of whom 26.3% (95% CI: 24.5 - 28.2) had stage 1 and 18.0% (95% CI: 16.1-20.1) stage 2 hypertension. Furthermore, 16.5% (95% CI: 14.4-18.9) and 9.6% (95% CI: 7.86-11.7) of men and women had elevated blood pressure, respectively. Conclusion: The findings of this study indicated that adopting the 2017 ACC/AHA guidelines showed a higher prevalence of adult hypertension (48.2%) in Iran. In this study, the prevalence of hypertension in men was higher than in women, which was steadily increased by age in older adults in both sexes.

Melatonin Attenuates Upregulation of Duox1 and Duox2 and Protects against Lung Injury following Chest Irradiation in Rats.

OBJECTIVE: The Lung is one of the most radiosensitive organs of the body. The infiltration of macrophages and lymphocytes into the lung is mediated via the stimulation of T-helper 2 cytokines such as IL-4 and IL-13, which play a key role in the development of fibrosis. It is likely that these cytokines induce chronic oxidative damage and inflammation through the upregulation of Duox1 and Duox2, which can increase the risk of late effects of ionizing radiation (IR) such as fibrosis and carcinogenesis. In the present study, we aimed to evaluate the possible increase of IL-4 and IL-13 levels, as well as their downstream genes such as IL4ra1, IL13ra2, Duox1, and Duox2. MATERIALS AND METHODS: In this experimental animal study, male rats were divided into 4 groups: i. Control, ii. Melatonintreated, iii. Radiation, and iv. Melatonin (100 mg/kg) plus radiation. Rats were irradiated with 15 Gy 60Co gamma rays and then sacrificed after 67 days. The expressions of IL4ra1, IL13ra2, Duox1, and Duox2, as well as the levels of IL-4 and IL-13, were evaluated. The histopathological changes such as the infiltration of inflammatory cells, edema, and fibrosis were also examined. Moreover, the protective effect of melatonin on these parameters was also determined. RESULTS: Results showed a 1.5-fold increase in the level of IL-4, a 5-fold increase in the expression of IL4ra1, and a 3-fold increase in the expressions of Duox1 and Duox2. However, results showed no change for IL-13 and no detectable expression of IL13ra2. This was associated with increased infiltration of macrophages, lymphocytes, and mast cells. Melatonin treatment before irradiation completely reversed these changes. CONCLUSION: This study has shown the upregulation of IL-4-IL4ra1-Duox2 signaling pathway following lung irradiation. It is possible that melatonin protects against IR-induced lung injury via the downregulation of this pathway and attenuation of inflammatory cells infiltration.

Selenium as an adjuvant for modification of radiation response.

Ionizing radiation plays a central role in several medical and industrial purposes. In spite of the beneficial effects of ionizing radiation, there are some concerns related to accidental exposure that could pose a threat to the lives of exposed people. This issue is also very critical for triage of injured people in a possible terror event or nuclear disaster. The most common side effects of ionizing radiation are experienced in cancer patients who had undergone radiotherapy. For complete eradication of tumors, there is a need for high doses of ionizing radiation. However, these high doses lead to severe toxicities in adjacent organs. Management of normal tissue toxicity may be achieved via modulation of radiation responses in both normal and malignant cells. It has been suggested that treatment of patients with some adjuvant agents may be useful for amelioration of radiation toxicity or sensitization of tumor cells. However, there are always some concerns for possible severe toxicities and protection of tumor cells, which in turn affect radiotherapy outcomes. Selenium is a trace element in the body that has shown potent antioxidant and radioprotective effects for many years. Selenium can potently stimulate antioxidant defense of cells, especially via upregulation of glutathione (GSH) level and glutathione peroxidase activity. Some studies in recent years have shown that selenium is able to mitigate radiation toxicity when administered after exposure. These studies suggest that selenium may be a useful radiomitigator for an accidental radiation event. Molecular and cellular studies have revealed that selenium protects different normal cells against radiation, while it may sensitize tumor cells. These differential effects of selenium have also been revealed in some clinical studies. In the present study, we aimed to review the radiomitigative and radioprotective effects of selenium on normal cells/tissues, as well as its radiosensitive effect on cancer cells.

Evaluating the protective effect of resveratrol, Q10, and alpha-lipoic acid on radiation-induced mice spermatogenesis injury: A histopathological study.

BACKGROUND: Testis is one of the most sensitive organs against the toxic effect of ionizing radiation. Exposure to even a low dose of radiation during radiotherapy, diagnostic radiology, or a radiological event could pose a threat to spermatogenesis. This may lead to temporary or permanent infertility or even transfer of genomic instability to the next generations. OBJECTIVE: In this study, we evaluated the protective effect of treatment with three natural antioxidants; resveratrol, alpha lipoic acid, and coenzyme Q10 on radiation-induced spermatogenesis injury. MATERIALS AND METHODS: 30 NMRI mice (6-8 wk, 30 ± 5 gr) were randomly divided into six groups (n = 5/each) as 1) control; 2) radiation; 3) radiation + resveratrol; 4) radiation + alpha lipoic acid; 5) radiation + resveratrol + alpha lipoic acid; and 6) radiation+ Q10. Mice were treated with 100 mg/kg resveratrol or 200 mg/kg alpha lipoic acid or a combination of these drugs. Also, Q10 was administered at 200 mg/kg. All treatments were performed daily from two days before to 30 min before irradiation. Afterward, mice were exposed to 2 Gy 60 Co gamma rays; 37 days after irradiation, the testicular samples were collected and evaluated for histopathological parameters. RESULTS: Results showed that these agents are able to alleviate some toxicological parameters such as basal lamina and epididymis decreased sperm density. Also, all agents were able to increase Johnsen score. However, they could not protect against radiation-induced edema, atrophy of seminiferous tubules, and hyperplasia in Leydig cells. CONCLUSION: This study indicates that resveratrol, alpha-lipoic acid, and Q10 have the potential to reduce some of the side effects of radiation on mice spermatogenesis. However, they cannot protect Leydig cells as a source of testosterone and seminiferous tubules as the location of sperm maturation.

Metformin Protects Against Radiation-Induced Pneumonitis and Fibrosis and Attenuates Upregulation of Dual Oxidase Genes Expression.

Purpose: Lung tissue is one of the most sensitive organs to ionizing radiation (IR). Early and late side effects of exposure to IR can limit the radiation doses delivered to tumors that are within or adjacent to this organ. Pneumonitis and fibrosis are the main side effects of radiotherapy for this organ. IL-4 and IL-13 have a key role in the development of pneumonitis and fibrosis. Metformin is a potent anti-fibrosis and redox modulatory agent that has shown radioprotective effects. In this study, we aimed to evaluate possible upregulation of these cytokines and subsequent cascades such as IL4-R1, IL-13R1, Dual oxidase 1 (DUOX1) and DUOX2. In addition, we examined the potential protective effect of metformin in these cytokines and genes, as well as histopathological changes in rat's lung tissues. Methods: 20 rats were divided into 4 groups: control; metformin treated; radiation + metformin; and radiation. Irradiation was performed with a 60Co source delivering 15 Gray (Gy) to the chest area. After 10 weeks, rats were sacrificed and their lung tissues were removed for histopathological, real-time PCR and ELISA assays. Results: Irradiation of lung was associated with an increase in IL-4 cytokine level, as well as the expression of IL-4 receptor-a1 (IL4ra1) and DUOX2 genes. However, there was no change in the level of IL-13 and its downstream gene including IL-13 receptor-a2 (IL13ra2). Moreover, histopathological evaluations showed significant infiltration of lymphocytes and macrophages, fibrosis, as well as vascular and alveolar damages. Treatment with metformin caused suppression of upregulated genes and IL-4 cytokine level, associated with amelioration of pathological changes. Conclusion: Results of this study showed remarkable pathological damages, an increase in the levels of IL-4, IL4Ra1 and Duox2, while that of IL-13 decreased. Treatment with metformin showed ability to attenuate upregulation of IL-4-DUOX2 pathway and other pathological damages to the lung after exposure to a high dose of IR.

Evaluating the Expression of NOX2 and NOX4 Signaling Pathways in Rats' Lung Tissues Following Local Chest Irradiation; Modulatory Effect of Melatonin.

Lung injury is one of the major concerns for chest cancer patients that undergo radiotherapy as well as persons exposed to an accidental radiological event. Reduction/oxidation (redox) system plays a key role in lung injury via chronic upregulation of pro-oxidant enzymes. NOX2 and NOX4 are two important reactive oxygen species generating enzymes that are involved in radiation toxicity in some organs such as the bone marrow. In this study, we aimed to evaluate the expression of NOX2 and NOX4 signaling in rat's lung tissues. Upregulation of these genes may be involved in radiation-induced lung injury. Moreover, we evaluated the role of pre-treatment with melatonin on the expression of these genes. Twenty male rats were divided into 4 groups as control; melatonin treated; irradiation; and irradiation with melatonin pre-treatment. Rats were exposed to 15 Gy 60Co gamma rays and sacrificed after 10 weeks for evaluation of NF-κB, TGFßR1, SMAD2, NOX2, and NOX4 gene expression by real-time PCR. Results showed the upregulation of all five genes. The expression of NOX2 was more obvious compared to other genes. Administration of melatonin before irradiation could attenuate the expression of all mentioned genes. Results indicate that upregulation of NADPH oxidase genes such as NOX2 and NOX4 may be involved in the late effects of lung exposure to ionizing radiation. Melatonin via downregulation of these pro-oxidant genes is able to attenuate radiation toxicity in the lung.

Metformin Protects Against Radiation-Induced Heart Injury and Attenuates the Upregulation of Dual Oxidase Genes Following Rat's Chest Irradiation.

Radiation-induced heart toxicity is one of the serious side effects after a radiation disaster or radiotherapy for patients with chest cancers, leading to a reduction in the quality of life of the patients. Evidence has shown that infiltration of inflammatory cells plays a key role in the development of functional damages to the heart via chronic upregulation of some pro-fibrotic and pro-inflammatory cytokines. These changes are associated with continuous free radical production and increased stiffness of heart muscle. IL-4 and IL-13 are two important pro-fibrotic cytokines which contribute to the side effects of ionizing radiation exposure. Recent studies have proposed that IL-4 through upregulation of DUOX2, and IL-13 via stimulation of DUOX1 gene expression, are involved in the development of radiation late effects. In the present study, we aimed to detect changes in the expression of these pathways following irradiation of rat's heart. Furthermore, we evaluated the possible protective effect of metformin on the development of these abnormal changes. 20 male rats were divided into 4 groups (control, radiation, metformin treated, metformin + radiation). These rats were irradiated with 15 Gy 60Co gamma rays, and sacrificed after 10 weeks for evaluation of the changes in the expression of IL4R1, IL-13R2a, DUOX1 and DUOX2. In addition, the levels of IL-4 and IL-13 cytokines, as well as infiltration of macrophages and lymphocytes were detected. Results showed an upregulation of both DUOX1 and DUOX2 pathways in the presence of metformin, while the level of IL-13 did not show any significant change. This was associated with infiltration of macrophages and lymphocytes. Also, treatment with metformin could significantly attenuate accumulation of inflammatory cells, and upregulate these pathways. Therefore, suppression of dual oxidase genes by metformin may be a contributory factor to its protective effect.