Turgut Topal

Melatonin: An established antioxidant worthy of use in clinical trials

Oxidative stress plays a key role in the pathogenesis of aging and many metabolic diseases; therefore, an effective antioxidant therapy would be of great importance in these circumstances. Nutritional, environmental, and chemical factors can induce the overproduction of the superoxide anion radical in both the cytosol and mitochondria. This is the first and key event that leads to the activation of pathways involved in the development of several metabolic diseases that are related to oxidative stress. As oxidation of essential molecules continues, it turns to nitrooxidative stress because of the involvement of nitric oxide in pathogenic processes. Once peroxynitrite forms, it damages via two distinctive mechanisms. First, it has direct toxic effects leading to lipid peroxidation, protein oxidation, and DNA damage. This mechanism involves the induction of several transcription factors leading to cytokine-induced chronic inflammation. Classic antioxidants, including vitamins A, C, and E, have often failed to exhibit beneficial effects in metabolic diseases and aging. Melatonin is a multifunctional indolamine that counteracts virtually all pathophysiologic steps and displays significant beneficial actions against peroxynitrite-induced cellular toxicity. This protection is related to melatonin’s antioxidative and antiinflammatory properties. Melatonin has the capability of scavenging both oxygen- and nitrogen-based reactants, including those formed from peroxynitrite, and blocking transcriptional factors, which induce proinflammatory cytokines. Accumulating evidence suggests that this nontoxic indolamine may be useful either as a sole treatment or in conjunction with other treatments for inhibiting the biohazardous actions of nitrooxidative stress.

Pathophysiological aspects of cyclophosphamide and ifosfamide induced hemorrhagic cystitis; implication of reactive oxygen and nitrogen species as well as PARP activation.

Cyclophosphamide (CP) and ifosfamide (IF) are widely used antineoplastic agents, but their side-effect of hemorrhagic cystitis (HC) is still encountered as an important problem. Acrolein is the main molecule responsible of this side-effect and mesna (2-mercaptoethane sulfonate) is the commonly used preventive agent. Mesna binds acrolein and prevent its direct contact with uroepithelium. Current knowledge provides information about the pathophysiological mechanism of HC: several transcription factors and cytokines, free radicals and non-radical reactive molecules, as well as poly(adenosine diphosphate-ribose) polymerase (PARP) activation are now known to take part in its pathogenesis. There is no doubt that HC is an inflammatory process, including when caused by CP. Thus, many cytokines such as tumor necrosis factor (TNF) and the interleukin (IL) family and transcription factors such as nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) also play a role in its pathogenesis. When these molecular factors are taken into account, pathogenesis of CP-induced bladder toxicity can be summarized in three steps: (1) acrolein rapidly enters into the uroepithelial cells; (2) it then activates intracellular reactive oxygen species and nitric oxide production (directly or through NF-κB and AP-1) leading to peroxynitrite production; (3) finally, the increased peroxynitrite level damages lipids (lipid peroxidation), proteins (protein oxidation) and DNA (strand breaks) leading to activation of PARP, a DNA repair enzyme. DNA damage causes PARP overactivation, resulting in the depletion of oxidized nicotinamide–adenine dinucleotide and adenosine triphosphate, and consequently in necrotic cell death. For more effective prevention against HC, all pathophysiological mechanisms must be taken into consideration.

Molecular targets against mustard toxicity: implication of cell surface receptors, peroxynitrite production, and PARP activation

Despite many years of research into chemical warfare agents, cytotoxic mechanisms induced by mustards are not well understood. Reactive oxygen and nitrogen species (ROS and RNS) are likely to be involved in chemical warfare agents induced toxicity. These species lead to lipid peroxidation, protein oxidation, and DNA injury, and trigger many pathophysiological processes that harm the organism. In this article, several steps of pathophysiological mechanisms and possible ways of protection against chemical warfare agents have been discussed. In summary, pathogenesis of mustard toxicity is explained by three steps: (1) mustard binds target cell surface receptor, (2) activates intracellular ROS and RNS leading to peroxynitrite (ONOO−) production, and (3) the increased ONOO− level damages organic molecules (lipids, proteins, and DNA) leading to poly(adenosine diphosphate-ribose) polymerase (PARP) activation. Therefore, protection against mustard toxicity could also be performed in these ways: (1) blocking of cell surface receptor, (2) inhibiting the ONOO− production or scavenging the ONOO− produced, and (3) inhibiting the PARP, activated by ONOO− and hydroxyl radical (OH•) induced DNA damage. As conclusion, to be really effective, treatment against mustards must take all molecular mechanisms of cytotoxicity into account. Combination of several individual potent agents, each blocking one of the toxic mechanisms induced by mustards, would be interesting. Therefore, variations of combination of cell membrane receptor blockers, antioxidants, nitric oxide synthase inhibitors, ONOO− scavengers, and PARP inhibitors should be investigated.

Melatonin ameliorates bladder damage induced by cyclophosphamide in rats

Abstract:  Cyclophosphamide (CP), an alkylating antineoplastic agent, has potential urotoxicity including causing hemorrhagic cystitis (HC). HC is now accepted as a non‐infectious inflammation and the pathogenesis of HC includes cytokine production which leads to inducible nitric oxide synthase (iNOS) induction. Moreover, overproduction of reactive oxygen species (ROS) during inflammation leads to extensive oxidative stress, cellular injury and apoptosis/necrosis via several mechanisms. Based on these facts, the aim of this study was to evaluate the protective effects of melatonin as an antioxidant, iNOS inhibitor and peroxynitrite scavenger against CP‐induced urinary bladder damage. A total of 30 male Sprague–Dawley rats were divided into four groups. Three groups received a single dose of CP (100 mg/kg) intraperitoneally with the same times. Group 2 received CP only, group 3 received 5 mg/kg/day and group 4 received 10 mg/kg/day melatonin before and the day after CP administration. Group 1 served as the control. Increased iNOS induction, bladder malonyldialdehyde (MDA) levels and urinary nitrite–nitrate excretion were encountered in the CP‐only group leading to severe cystitis. Melatonin exhibited significant protection against CP‐induced cystitis by diminishing bladder oxidative stress and blocking iNOS and peroxynitrite production. Oxidants may have a major role in the pathogenesis of CP‐induced cystitis and iNOS is an important mediator leading to peroxynitrite production. Melatonin ameliorates bladder damage induced by CP.

The efficacy of ozone therapy in experimental caustic esophageal burn.

INTRODUCTION Ozone has been proposed as an antioxidant enzyme activator, immunomodulator and cellular metabolic activator. This study was designed to investigate the efficacy of ozone therapy in the prevention of esophageal damage and stricture formation developed after esophageal caustic injuries in the rat. MATERIALS AND METHODS Forty-five rats were allocated into three groups; sham-operated, un-treatment and treatment groups. Caustic esophageal burn was created by instilling 15% NaOH in the distal esophagus. The rats were left untreated or treated with 1 mg/kg/day ozone intraperitoneally. All rats were sacrificed at 28 days. Efficacy of the treatment was assessed by measuring the stenosis index (SI) and histopathologic damage score, and biochemically by determining tissue hydroxyproline content (HP), superoxide dismutase (SOD), glutathione peroxidase (GPx), malondialdehyde (MDA) and protein carbonyl content (PCC) in esophageal homogenates. RESULTS Whereas seven (47%) rats died in the un-treatment group, all rats in the sham-operated and the treatment group survived during the study. SI, the histopathologic damage score, was significantly lower in the ozone-therapy group than the un-treatment group. HP levels were significantly higher in the un-treatment group than the group treated with ozone. Caustic esophageal burn increased MDA and PCC levels and also decreased SOD and GPx enzyme activities. In contrast, ozone therapy decreased the elevated MDA and PCC levels and also increased the reduced SOD and GPx enzyme activities. CONCLUSION Ozone has a preventive effect in the development of fibrosis by decreasing tissue damage and increasing the antioxidant enzyme activity in an experimental model of corrosive esophageal injury.

α-Lipoic Acid and Ebselen Prevent Ischemia/Reperfusion Injury in the Rat Intestine

PurposeReactive oxygen species (ROS) and reactive nitrogen species (RNS), generated during tissue reperfusion, are characteristic of ischemia/reperfusion (I/R) injury. We conducted this study to evaluate the protective effect of α-lipoic acid (α-LA) and ebselen against intestinal I/R injury.MethodsForty Sprague-Dawley rats were divided into five groups: a sham-operated group; an I/R group, subjected to intestinal ischemia for 45 min and reperfusion for 3 days; an I/R+α-LA group; an I/R+ebselen group; and an I/R+α-LA+ebselen group. We collected ileal specimens, to measure the tissue levels of malondialdehyde (MDA), protein carbonyl content (PCC), superoxide dismutase (SOD), and glutathione peroxidase (GPx), and to evaluate the histologic changes.ResultsThere was a significant decrease in SOD and GPx levels, with an increase in MDA and PCC levels and intestinal mucosal injury in the intestinal I/R group (P < 0.05). Superoxide dismutase and GPx levels were significantly higher, MDA and PCC levels were significantly lower, and intestinal injury was significantly less severe in the I/R+α-LA+ebselen group than in the I/R group (P < 0.05). Although shortened villi and epithelial lifting were seen in the I/R group, only slight mucosal injury was seen in the treatment groups.Conclusionα-Lipoic acid and ebselen played an important role in attenuating I/R injury of the intestine by scavenging ROS and RNS.

Antioxidant effects of melatonin in rats during chronic exposure to hyperbaric oxygen

Abstract:  In addition to its beneficial effects, hyperbaric oxygen (HBO) exposure causes some detrimental effects via oxidative stress. Previous experimental studies showed that melatonin is a useful agent to block single session HBO‐induced oxidative stress. In the present study, we investigated the antioxidant effect of exogenously administered as well as endogenously produced melatonin in lung and brain tissues of rats exposed to long term HBO. The HBO procedure was set as daily exposures to 2.5 ATA of oxygen for 1 hr and a total of 10 sessions. Twenty‐eight male Sprague–Dawley rats were divided into four groups as follows: control, daytime HBO, daytime HBO plus melatonin (5 mg/kg), nighttime HBO. Tissue oxidative/antioxidant status was examined by determining the protein carbonyl content as a criteria for oxidative stress and the activities of the antioxidant enzymes, superoxide dismutase (SOD) and glutathione peroxidase (GSH‐Px). HBO exposure for 10 days caused significant increases in protein carbonyl content and SOD levels of lung and brain, but GSH‐Px activities remained unaffected. The increases in protein carbonyls were blocked by exogenously administered melatonin and in part by nighttime exposure to darkness whereas the increase of SOD activity was only impeded by endogenously produced melatonin in brain tissue. Lung SOD activity was augmented by endogenous melatonin. In conclusion, melatonin blocks long‐term HBO‐induced cumulative oxidative stress as indicated changes in protein carbonyls. Both exogenously injected and physiologically secreted melatonin has this potential. The effects of HBO‐exposure and melatonin on the activities of the antioxidative enzymes are less clear.

Glucose: A vital toxin and potential utility of melatonin in protecting against the diabetic state

The molecular mechanisms including elevated oxidative and nitrosative reactants, activation of pro-inflammatory transcription factors and subsequent inflammation appear as a unified pathway leading to metabolic deterioration resulting from hyperglycemia, dyslipidemia, and insulin resistance. Consistent evidence reveals that chronically-elevated blood glucose initiates a harmful series of processes in which toxic reactive species play crucial roles. As a consequence, the resulting nitro-oxidative stress harms virtually all biomolecules including lipids, proteins and DNA leading to severely compromised metabolic activity. Melatonin is a multifunctional indoleamine which counteracts several pathophysiologic steps and displays significant beneficial effects against hyperglycemia-induced cellular toxicity. Melatonin has the capability of scavenging both oxygen and nitrogen-based reactants and blocking transcriptional factors which induce pro-inflammatory cytokines. These functions contribute to melatonins antioxidative, anti-inflammatory and possibly epigenetic regulatory properties. Additionally, melatonin restores adipocyte glucose transporter-4 loss and eases the effects of insulin resistance associated with the type 2 diabetic state and may also assist in the regulation of body weight in these patients. Current knowledge suggests the clinical use of this non-toxic indoleamine in conjunction with other treatments for inhibition of the negative consequences of hyperglycemia for reducing insulin resistance and for regulating the diabetic state.

Effect of 3-amino benzamide, a poly(adenosine diphosphate-ribose) polymerase inhibitor, in experimental caustic esophageal burn

INTRODUCTION The enzyme poly(adenosine diphosphate-ribose) polymerase affects the repair of DNA in damaged cells. However, its activation in damaged cells can lead to adenosine triphosphate depletion and death. This study was designed to investigate the efficacy of 3-amino benzamide (3-AB), a poly(adenosine diphosphate-ribose) polymerase inhibitor, on the prevention of esophageal damage and stricture-formation development after esophageal caustic injuries in rat. MATERIALS AND METHODS Forty-five rats were allocated into 3 groups: sham-operated, untreated, and treated groups. Caustic esophageal burn was created by instilling 15% NaOH to the distal esophagus. The rats were left untreated or treated with 3-AB 10 mg/kg per day intraperitoneally. All rats were killed on the 28th day. Efficacy of the treatment was assessed by measuring the stenosis index and histopathologic damage score and biochemically by determining tissue hydroxyproline content, superoxide dismutase (SOD), glutathione peroxidase (GPx), malondialdehyde (MDA), and protein carbonyl content (PCC) in esophageal homogenates. RESULTS Treatment with 3-AB decreased the stenosis index and histopathologic damage score seen in caustic esophageal burn rats. Hydroxyproline level was significantly higher in the untreated group as compared with the group treated with 3-AB. Caustic esophageal burn increased MDA and PCC levels and also decreased SOD and GPx enzyme activities. On the contrary, 3-AB treatment decreased the elevated MDA and PCC levels and also increased the reduced SOD and GPx enzyme activities. CONCLUSION 3-Amino benzamide has a preventive effect in the development of fibrosis by decreasing tissue damage and increasing the antioxidant enzyme activity in an experimental model of corrosive esophagitis in rats.

Protective effects of exogenously administered or endogenously produced melatonin on hyperbaric oxygen‐induced oxidative stress in the rat brain

1. Hyperbaric oxygen (HBO) is a widely used treatment modality in many diseases. A known side‐effect of HBO is the production of reactive oxygen species (ROS). Many anti‐oxidants, such as vitamins C and E, riboflavin and selenium, have been used successfully to scavenge the ROS produced by HBO administration.