Therapeutic Effects of Hydrogen Gas Inhalation on Trimethyltin-Induced Neurotoxicity and Cognitive Impairment in the C57BL/6 Mice Model

Oxidative stress (OS) is one of the causative factors in the pathogenesis of various neurodegenerative diseases, including Alzheimer’s disease (AD) and cognitive dysfunction. In the present study, we investigated the effects of hydrogen (H2) gas inhalation in trimethyltin (TMT)-induced neurotoxicity and cognitive dysfunction in the C57BL/6 mice. First, mice were divided into the following groups: mice without TMT injection (NC), TMT-only injection group (TMT only), TMT injection + lithium chloride-treated group as a positive control (PC), and TMT injection + 2% H2 inhalation-treated group (H2). The TMT injection groups were administered a single dosage of intraperitoneal TMT injection (2.6 mg/kg body weight) and the H2 group was treated with 2% H2 for 30 min once a day for four weeks. Additionally, a behavioral test was performed with Y-maze to test the cognitive abilities of the mice. Furthermore, multiple OS- and AD-related biomarkers such as reactive oxygen species (ROS), nitric oxide (NO), calcium (Ca2+), malondialdehyde (MDA), glutathione peroxidase (GPx), catalase, inflammatory cytokines, apolipoprotein E (Apo-E), amyloid β (Aβ)-40, phospho-tau (p-tau), Bcl-2, and Bcl-2- associated X (Bax) were investigated in the blood and brain. Our results demonstrated that TMT exposure alters seizure and spatial recognition memory. However, after H2 treatment, memory deficits were ameliorated. H2 treatment also decreased AD-related biomarkers, such as Apo-E, Aβ-40, p-tau, and Bax and OS markers such as ROS, NO, Ca2+, and MDA in both serum and brain. In contrast, catalase and GPx activities were significantly increased in the TMT-only group and decreased after H2 gas treatment in serum and brain. In addition, inflammatory cytokines such as granulocyte colony-stimulating factors (G-CSF), interleukin (IL)-6, and tumor necrosis factor alpha (TNF-α) were found to be significantly decreased after H2 treatment in both serum and brain lysates. In contrast, Bcl-2 and vascular endothelial growth factor (VEGF) expression levels were found to be enhanced after H2 treatment. Taken together, our results demonstrated that 2% H2 gas inhalation in TMT-treated mice exhibits memory enhancing activity and decreases the AD, OS, and inflammatory-related markers. Therefore, H2 might be a candidate for repairing neurodegenerative diseases with cognitive dysfunction. However, further mechanistic studies are needed to fully clarify the effects of H2 inhalation on TMT-induced neurotoxicity and cognitive dysfunction.

Antioxidant Properties of Hydrogen Gas Attenuates Oxidative Stress in Airway Epithelial Cells

Oxidative stress plays a crucial role in the development of airway diseases. Recently, hydrogen (H2) gas has been explored for its antioxidant properties. This study investigated the role of H2 gas in oxidative stress-induced alveolar and bronchial airway injury, where A549 and NCI-H292 cells were stimulated with hydrogen peroxide (H2O2) and lipopolysaccharide (LPS) in vitro. Results show that time-dependent administration of 2% H2 gas recovered the cells from oxidative stress. Various indicators including reactive oxygen species (ROS), nitric oxide (NO), antioxidant enzymes (catalase, glutathione peroxidase), intracellular calcium, and mitogen-activated protein kinase (MAPK) signaling pathway were examined to analyze the redox profile. The viability of A549 and NCI-H292 cells and the activity of antioxidant enzymes were reduced following induction by H2O2 and LPS but were later recovered using H2 gas. Additionally, the levels of oxidative stress markers, including ROS and NO, were elevated upon induction but were attenuated after treatment with H2 gas. Furthermore, H2 gas suppressed oxidative stress-induced MAPK activation and maintained calcium homeostasis. This study suggests that H2 gas can rescue airway epithelial cells from H2O2 and LPS-induced oxidative stress and may be a potential intervention for airway diseases.

Wound Healing Effect of Slightly Acidic Electrolyzed Water on Cutaneous Wounds in Hairless Mice via Immune-Redox Modulation

Acidic electrolyzed water is an innovative sanitizer having a wide-spectrum of applications in food industry, and healthcare industry but little is known on its effect and mechanism in wound healing. The study was conducted to identify the effect and mechanism of slightly acidic electrolyzed water (SAEW) on cutaneous wounds in hairless mice. SAEW (pH: 5–6.5, oxidation reduction potential: 800 mV, chlorine concentration: 25 ppm) was prepared through electrolysis of water and was applied to the wounds of hairless mice three times a day for seven days. Wound size, immune response and oxidative stress were explored and compared to conventional agents such as Betadine and alcohol. We found that SAEW-treated group showed the highest wound reduction percentage (p<0.01). Antioxidant activities such as glutathione peroxidase, catalase and myeloperoxidase activities of SAEW group surpassed the total reactive oxygen species in skin. Nuclear factor erythroid-2-related-factor-2 and aryl hydrocarbon receptor were upregulated in SAEW group. Further, SAEW recruited the production of intracellular calcium and promoted its utilization for faster healing. In line, SAEW treatment decreased pro-inflammatory cytokines [interleukin (IL)-1β, IL-6, keratinocyte chemoattractant, and tumor necrosis factor-α] in serum. Other hallmarks of wound healing, matrixmetalloproteinases (MMP)1 and MMP9 were also upregulated. Collectively, our study indicates that SAEW is effective in wound healing of hairless mice via immune-redox modulation, and heals better/faster than conventional agents.

Potential Therapeutic and Preventive Effect of Drinking Hydrogen-rich Water on Acute Fatigue

Acute fatigue is a condition defined as a sudden onset of physical and mental exhaustion particularly after a short but strenuous period of physical exercise due to effect of waste product of muscle contraction within muscle fiber and accumulation of metabolites within the muscle fiber. Until recently, it’s believed that lactic acid build-up and reactive oxygen species (ROS) resulting oxidative stress are the most common causes of muscle fatigue. With prevalence of this condition and due to the lack of effective therapeutics, fatigue-related disorders turn to alternative medicine and other non-traditional practices. Previous studies on molecular hydrogen have reported that hydrogen exerts antioxidant, anti-inflammatory and metabolic modulation properties that are beneficial to the cell. Herein we hypothesized that hydrogen-rich water (HW) might ameliorate various impairment of acute fatigue through ROS scavenging activity and elevation of metabolic profile which were related to fatigue such as blood lactate and increasing storage of muscle glycogen, thereby improving physical endurance.

Hydrogen Water Drinking Exerts Antifatigue Effects in Chronic Forced Swimming Mice via Antioxidative and Anti-Inflammatory Activities

Purpose: This study was performed to evaluate antifatigue effect of hydrogen water (HW) drinking in chronic forced exercise mice model. Materials and methods: Twelve-week-old C57BL6 female mice were divided into nonstressed normal control (NC) group and stressed group: (purified water/PW-treated group and HW-treated group). Stressed groups were supplied with PW and HW, respectively, ad libitum and forced to swim for the stress induction every day for 4 consecutive weeks. Gross antifatigue effects of HW were assessed by swimming endurance capacity (once weekly for 4 wk), metabolic activities, and immune-redox activities. Metabolic activities such as blood glucose, lactate, glycogen, blood urea nitrogen (BUN), and lactate dehydrogenase (LDH) as well as immune-redox activities such as reactive oxygen species (ROS), nitric oxide (NO), glutathione peroxidase (GPx), catalase, and the related cytokines were evaluated to elucidate underlying mechanism. Blood glucose and lactate were measured at 0 wk (before swimming) and 4 wk (after swimming). Results: HW group showed a higher swimming endurance capacity (p < 0.001) than NC and PW groups. Positive metabolic effects in HW group were revealed by the significant reduction of blood glucose, lactate, and BUN in serum after 4 wk (p < 0.01, resp.), as well as the significant increase of liver glycogen (p < 0.001) and serum LDH (p < 0.05) than PW group. In parallel, redox balance was represented by lower NO in serum (p < 0.01) and increased level of GPx in both serum and liver (p < 0.05) than PW group. In line, the decreased levels of serum TNF-α (p < 0.01), IL-6, IL-17, and liver IL-1β (p < 0.05) in HW group revealed positive cytokine profile compared to PW and NC group. Conclusion: This study shows antifatigue effects of HW drinking in chronic forced swimming mice via metabolic coordination and immune-redox balance. In that context, drinking HW could be applied to the alternative and safety fluid remedy for chronic fatigue control.

Effect of Hydrogen Water Drinking on Dementia Patient through Mechanism of Oxidative Stress and Immune Redox Balance

Hydrogen water (HW) which is produced by electrolysis of water has characteristics of extremely low oxidation-reduction potential (ORP) value and high dissolved hydrogen (DH). It has been proved to have various beneficial effects including antioxidant and anti-inflammatory effects; however, HW effect on dementia that is associated with cognitive impairment and memory loss, is poorly documented. In the present study, we investigated the drinking effects of hydrogen water on dementia patient for prevention through oxidative stress and immuno redox mechanism. Dementia patients were administered with HW and purified water (PW) for 6 months. Further, we evaluated hydrogen water effect on the cognitive impairment by checking the serum concentration of Alzheimer’s disease (AD) biomarkers [Amyloid β (Aβ) 40, Aβ 42 and tau], inflammatory cytokines such as IL-1β, IL-6, IL-13, TNF-α, IL-10 and IL-4, Th1/Th2 cytokines such as IL-2, IFN-γ, IL-12p70 and IL-5, reactive oxygen species (ROS), and nitric oxide (NO) levels. We found that HW treatment significantly decreased the serum level of Aβ 40 after 6 mon of treatment. In parallel, inflammatory cytokines were significantly inhibited in experimental group compared to control group. In line, Th1/Th2 cytokines were significantly decreased in HW than PW group. Furthermore, oxidative stress markers such as ROS and nitric oxide level were decreased in HW treated group than PW control group. Overall our clinical results strongly recommended that drinking hydrogen water might be a promising preventative approach for age-related neurodegenerative disease like dementia and have an enormous impact on future healthcare for the elderly patients. This might be the first note on the clinical application of drinking HW on the serum of dementia patients.

Effects of Strong Acidic Electrolyzed Water in Wound Healing via Inflammatory and Oxidative Stress Response

Strong acidic electrolyzed water (StAEW) is known to inactivate microorganisms but is not fully explored in the medical field. This study is aimed at exploring StAEW as a potential wound care agent and its mechanism. StAEW (pH: 2.65, ORP: 1159 mV, ACC: 32.1 ppm) was sprayed three times a day to the cutaneous wounds of hairless mice for seven days. Wound morphological and histological features and immune-redox markers were compared with saline- (Sal-) and alcohol- (Alc-) treated groups. Results showed that the StAEW group showed a significantly higher wound healing percentage than the Sal group on days 2, 4, 5, and 6 and the Alc group on day 4. The StAEW group also showed earlier mediation on proinflammatory cytokines such as tumor necrosis factor-α, interleukin- (IL-) 6, IL-1β, and keratinocyte chemoattractant. In addition, basic fibroblast growth factor and platelet-derived growth factor were found to be significantly changed in favor of the fibroblast synthesis and angiogenesis. In line, the StAEW group showed a controlled amount of ROS and significantly decreased compared to the Alc group. The StAEW group also favored oxidative stress balance through antioxidant responses. Additionally, matrix metalloproteinases (MMP) 9 and MMP1 were also modulated for keratinocyte and cell migration. Taken together, this study has proven the wound healing effect of StAEW and its earlier mediation through oxidative and inflammatory responses.

Molecular hydrogen protects against oxidative stress-induced RAW 264.7 macrophage cells through the activation of Nrf2 and inhibition of MAPK signaling pathway

Background Oxidative stress is involved in the development of many inflammatory, metabolic and aging diseases.Objective In this study we investigated, the protective effects of H2 on RAW 264.7 macrophage cell against LPS-and H2O2-induced oxidative stress by the inhibition of MAPK pathway and also activate the Nrf2 pathway.ResultsOur results showed H2 increased the macrophage cell proliferation and generated ROS and NO against LPS stimulation to exert an active immune response. Similarly, H2 protected the macrophage cell from H2O2-induced oxidative stress. H2 reduced the LPS-and H2O2-induced inflammatory cytokine production and intracellular calcium influxes. H2 inhibited the LPS-and H2O2-induced phosphorylation of MAPK pathway and its downstream signaling molecules. Furthermore, H2 protected the macrophage cell from mitochondrial apoptosis. H2 increased Nrf2 protein expression indicating its strong anti-oxidative effects against oxidative stress.Conclusion Collectively, our results indicate the strong antioxidant role of H2 against LPS-and H2O2-induced oxidative stress on macrophage cells by activating the Nrf2 pathway and inhibiting the MAPK-signaling pathway.Graphic AbstractOur results clearly showed that LPS increased the cellular ROS by recognizing the TLR4 and H2O2 rapidly increased the cellular (1) and mitochondrial (2) oxidative stress. Excessive ROS/NO molecules cause intracellular calcium influxes (3) As a results imbalance the cellular membrane homeostasis and activate the stress response MAPK signaling pathway with its downstream signaling protein and mitochondrial caspase protein (4) that collapse the anti-oxidant mechanisms and induced the inflammatory cytokine secretion; leads to cell apoptosis (5) Whether H2 reduced the cellular and mitochondrial oxidative stress, intracellular calcium influxes and inhibits the stress response MAPK, caspase cell signaling pathway through the activation of Nrf2/ARE signaling pathway (6) Consequently, increased the antioxidant enzymes and reduced the inflammatory cytokine that influences the macrophage cell proliferation (7) to protect the cell from apoptosis. The different effects of ROS and H2 used in this study are indicated in red and green, red arrow depicts ROS effects in the cell, green arrow depicts H2 effects.