Magnesium galvanic cells produce hydrogen and modulate the tumor microenvironment to inhibit cancer growth

Hydrogen can be used as an anti-cancer treatment. However, the continuous generation of H2 molecules within the tumor is challenging. Magnesium (Mg) and its alloys have been extensively used in the clinic as implantable metals. Here we develop, by decorating platinum on the surface of Mg rods, a Mg-based galvanic cell (MgG), which allows the continuous generation of H2 in an aqueous environment due to galvanic-cell-accelerated water etching of Mg. By implanting MgG rods into a tumor, H2 molecules can be generated within the tumor, which induces mitochondrial dysfunction and intracellular redox homeostasis destruction. Meanwhile, the Mg(OH)2 residue can neutralize the acidic tumor microenvironment (TME). Such MgG rods with the micro-galvanic cell structure enable hydrogen therapy to inhibit the growth of tumors, including murine tumor models, patient-derived xenografts (PDX), as well as VX2 tumors in rabbits. Our research suggests that the galvanic cells for hydrogen therapy based on implantable metals may be a safe and effective cancer treatment.

FDG-PET/CT Assessment of the Cerebral Protective Effects of Hydrogen in Rabbits with Cardiac Arrest

Background: Anatomical imaging methods and histological examinations have limited clinical value for early monitoring of brain function damage after cardiac arrest (CA) in vivo. Objective: We aimed to assess the cerebral protective effects of hydrogen in rabbits with CA by using fluorodeoxyglucose-positron emission tomography/computed tomography (FDG-PET/CT). Methods: Male rabbits were divided into the hydrogen-treated (n=6), control (n=6), and sham (n=3) groups. Maximum standardized uptake values (SUVmax) were measured by FDG-PET/CT at baseline and post-resuscitation. Blood Ubiquitin C-terminal hydrolase-L1 (UCH-L1) and neuron specific enolase (NSE) were measured before and after the operation. After surgical euthanasia, brain tissues were extracted for Nissl staining. Results: SUVmax values first decreased at 2 and 24 h after resuscitation before rising in the hydrogen-treated and control groups. SUVmax values in the frontal, occipital, and left temporal lobes and in the whole brain were significantly different between the hydrogen and control groups at 2 and 24 h post-resuscitation (P<0.05). The neurological deficit scores at 24 and 48 h were lower in the hydrogen-treated group (P<0.05). At 24 h, the serum UCH-L1 and NSE levels were increased in the hydrogen and control groups (P<0.05), but not in the sham group. At 48 and 72 h post-CA, the plasma UCH-L1 and NSE levels in the hydrogen and control groups gradually decreased. Neuronal damage was smaller in the hydrogen group compared with the control group at 72 h. Conclusion: FDG-PET/CT could be used to monitor early cerebral damage, indicating a novel method for evaluating the protective effects of hydrogen on the brain after CA.

Multifunctional Magnesium Anastomosis Staples for Wound Closure and Inhibition of Tumor Recurrence and Metastasis

Biodegradable magnesium (Mg) implants spontaneously releasing therapeutic agents against tumors are an intriguing therapeutic approach for both tissue repair and tumor treatment. Anastomotic staples are extensively used for wound closure after surgical resection in patients with colorectal tumors. However, the safety of Mg anastomosis implants for intestinal closure and the effect of tumor suppression remain elusive. Here, we used a high-purity Mg staple to study these issues. Based on the results, we found that it has the potential to heal wounds produced after colorectal tumor resection while inhibiting relapse of residual tumor cells in vitro and in vivo. After implantation of Mg staples for 7 weeks in rabbits, the intestinal wound gradually healed with no adverse effects such as leakage or inflammation. Furthermore, the implanted Mg staples inhibit the growth of colorectal tumor cells and block migration to normal organs because of the increased concentration of Mg ions and released hydrogen. Such an antitumor effect is further confirmed by the in vitro cell experiments. Mg significantly induces apoptosis of tumor cells as well as inhibits cell growth and migration. Our work presents a feasible therapeutic opinion to design Mg anastomotic staples to perform wound healing and simultaneously release tumor suppressor elements in vivo to decrease the risk of tumor recurrence and metastasis.

In vivo quantification of hydrogen gas concentration in bone marrow surrounding magnesium fracture fixation hardware using an electrochemical hydrogen gas sensor

Statement of significance: An electrochemical H2 sensor was used to monitor the degradation of a Mg fracture fixation system in a lapine ulna fracture model. Interestingly, the H2 concentration in the bone marrow is 82% higher than H2 saturated water solution. This suggests H2 generated in situ is trapped in the bone marrow and bone is less permeable than the surrounding tissues. The detectable H2 at the rabbit skin also demonstrates a H2 sensor’s ability to monitor the degradation process under thin layers of tissue. H2 sensing shows promise as a tool for monitoring the degradation of Mg alloy in vivo and creating in vitro test beds to more mechanistically evaluate the effects of varying H2 concentrations on cell types relevant to osteogenesis.

Molecular Hydrogen Attenuated N-methyl-N-Nitrosourea Induced Corneal Endothelial Injury by Upregulating Anti-Apoptotic Pathway

Purpose: Previous work by our group has demonstrated the value of N-methyl-N-nitrosourea (MNU)-induced corneal endothelial decompensation in animal models. The aim of this study was to investigate the effect of molecular hydrogen (H2) on MNU-induced corneal endothelial cell (CEC) injury and the underlying mechanism. Methods: MNU-induced animal models of CEC injury were washed with hydrogen-rich saline (HRS) for 14 days. Immunofluorescence staining, immunohistochemical staining, and corneal endothelial assessment were applied to determine architectural and cellular changes on the corneal endothelium following HRS treatment. MNU-induced cell models of CEC injury were co-cultured with H2. The effect of H2 was examined using morphological and functional assays. Results: It was shown that MNU could inhibit the proliferation and specific physiological functions of CECs by increasing apoptosis and decreasing the expression of ZO-1 and Na+/K+-ATPase, whereas H2 improved the proliferation and physiological function of CECs by anti-apoptosis. Cell experiments further confirmed that H2 could reverse MNU damage to CECs by decreasing oxidative stress injury, interfering with the NF-κB/NLRP3 pathway and the FOXO3a/p53/p21 pathway. Conclusions: This study suggests that topical application of H2 could protect CECs against corneal damage factors through anti-apoptotic effect, reduce the incidence and severity of corneal endothelial decompensation, and maintain corneal transparency.

Beneficial effects of hydrogen gas against spinal cord ischemia-reperfusion injury in rabbits

Recently, hydrogen gas (H₂) is reported to be a new therapeutic agent in organ damage induced by ischemia-reperfusion (I/R). The present study was designed to investigate the beneficial effects of H₂ against spinal cord I/R injury and its associated mechanisms. Spinal cord ischemia was induced by infrarenal aortic occlusion for 20 min in male New Zealand white rabbits. Treatment with 1%, 2% or 4% H₂ inhalation was given from 10 min before reperfusion to 60 min after reperfusion (total 70 min). Here, we found that I/R-challenged animals showed significant spinal cord damage characterized by the decreased numbers of normal motor neurons and hind-limb motor dysfunction, which was significantly improved by 2% and 4 % H₂ treatment. Furthermore, we found that the beneficial effects of H₂ treatment against spinal cord I/R injury were associated with the decreased levels of oxidative products [8-iso-prostaglandin F2α (8-iso-PGF2α) and malondialdehyde (MDA)] and pro-inflammatory cytokines [tumor necrosis factor-alpha (TNF-α) and high-mobility group box 1 (HMGB1)], as well as increased activities of antioxidant enzymes [superoxide dismutase (SOD) and catalase (CAT)] in serum and spinal cord. In addition, H₂ treatment reduced motor neuron apoptosis in the spinal cord of this model. Thus, H₂ inhalation may be an effective therapeutic strategy for spinal cord I/R damage.

Hydrogen-rich saline alleviates early brain injury via reducing oxidative stress and brain edema following experimental subarachnoid hemorrhage in rabbits

Background: Increasing experimental and clinical data indicate that early brain injury (EBI) after subarachnoid hemorrhage (SAH) largely contributes to unfavorable outcomes, and it has been proved that EBI following SAH is closely associated with oxidative stress and brain edema. The present study aimed to examine the effect of hydrogen, a mild and selective cytotoxic oxygen radical scavenger, on oxidative stress injury, brain edema and neurology outcome following experimental SAH in rabbits. Results: The level of MDA, caspase-12/3 and brain water content increased significantly at 72 hours after experimental SAH. Correspondingly, obvious brain injury was found in the SAH group by terminal deoxynucleotidyl transferase-mediated uridine 5′-triphosphate-biotin nick end-labeling (TUNEL) and Nissl staining. Similar results were found in the SAH+saline group. In contrast, the upregulated level of MDA, caspase-12/3 and brain edema was attenuated and the brain injury was substantially alleviated in the hydrogen treated rabbits, but the improvement of neurology outcome was not obvious. Conclusion: The results suggest that treatment with hydrogen in experimental SAH rabbits could alleviate brain injury via decreasing the oxidative stress injury and brain edema. Hence, we conclude that hydrogen possesses the potential to be a novel therapeutic agent for EBI after SAH.

Hydrogen-Rich Saline Attenuates Lung Ischemia-Reperfusion Injury in Rabbits

Hydrogen gas, an antioxidant agent, was found to protect against cerebral and myocardial ischemia-reperfusion (I/R) injury. In the present study, we investigated the effect of hydrogen-rich saline (HRS) on the I/R-induced lung injury. Left lung of male New Zealand White rabbits rendered normothermic ischemia for 60 min and reperfused for up to 240 min. Treated animals received intraperitoneal injection of 5 mL/kg HRS or the same volume of normal saline 10 min before the start of reperfusion. Blood and lung tissue samples were obtained for blood gas and biochemical analyses. The tissues obtained from lower lobe of left lung were used for histologic examination. After 240 min of reperfusion, intraperitoneal administration of HRS increased PaO2/FiO2 ratio and superoxide dismutase activities, and decreased malondialdehyde contents, proinflammatory cytokines expression, and myeloperoxidase activities, along with reduced wet/dry ratio and histologic injury scores (P < 0.05 versus I/R group). These results suggest that intraperitoneal administration of HRS before reperfusion protects the lung from I/R injury. The protective effect seems to be closely related to regulating oxidative damage and antioxidant enzyme activities and neutrophil infiltration.

Nuclear factor-κB/Bcl-XL pathway is involved in the protective effect of hydrogen-rich saline on the brain following experimental subarachnoid hemorrhage in rabbits

Early brain injury (EBI), a significant contributor to poor outcome after subarachnoid hemorrhage (SAH), is intimately associated with neuronal apoptosis. Recently, the protective role of hydrogen (H2 ) in the brain has been widely studied, but the underlying mechanism remains elusive. Numerous studies have shown nuclear factor-κB (NF-κB) as a crucial survival pathway in neurons. Here we investigated the role of H2 in EBI following SAH, focusing on the NF-κB pathway. A double blood injection model was used to produce experimental SAH, and H2 -rich saline was injected intraperitoneally. NF-κB activity within the occipital cortex was measured. Immunofluorescence was performed to demonstrate the activation of NF-κB; Bcl-xL and cleaved caspase-3 were determined via Western blot. Gene expression of Bcl-xL was detected by real-time PCR, and TUNEL and Nissl staining were performed to illustrate brain injury in the occipital cortex. SAH induced a significant increase of cleaved caspase-3. Correspondingly, TUNEL staining demonstrated obvious neuronal apoptosis following SAH. In contrast, H2 treatment markedly increased NF-κB activity and the expression of Bcl-xL and decreased the level of cleaved caspase-3. Additionally, H2 treatment significantly reduced post-SAH neuronal apoptosis. The current study shows that H2 treatment alleviates EBI in the rabbits following SAH and that NF-κB/Bcl-xL pathway is involved in the protective role of H2 . © 2013 Wiley Periodicals, Inc.

Beneficial effects of hydrogen-rich saline against spinal cord ischemia-reperfusion injury in rabbits

Hydrogen-rich saline (HS) is reported to be a new therapeutic agent in ischemia-reperfusion (I/R)-induced organ damage. The present study was designed to investigate the beneficial effects of HS against spinal cord I/R injury and its associated mechanisms. Spinal cord ischemia was induced by infrarenal aortic occlusion for 20min in male New Zealand white rabbits. Different doses of HS were intravenously (i.v.) administered at 5min before or after the beginning of reperfusion. Moreover, the roles of mitochondrial ATP-sensitive potassium channels (mitoKATP), oxidative stress, inflammatory cytokines and apoptosis was assessed. Here, we found that I/R-challenged rabbits exhibited significant spinal cord injury characterized by the decreased numbers of normal motor neurons and hind-limb motor dysfunction, which was significantly ameliorated by 5mL/kg and 10mL/kg HS treatment before reperfusion or 10mL/kg HS treatment after reperfusion. However, the protective effects of HS treatment in spinal cord I/R injury were partially abolished by the selective mitoKATP channel blocker 5-hydroxydecanoate (5-HD). Moreover, we showed that the beneficial effects of 10mL/kg HS treatment against spinal cord I/R damage were associated with the decreased levels of oxidative products [8-iso-prostaglandin F2α (8-iso-PGF2α) and malondialdehyde (MDA)] and pro-inflammatory cytokines [tumor necrosis factor-alpha (TNF-α) and high-mobility group box 1 (HMGB1)], as well as the increased activities of antioxidant enzymes [superoxide dismutase (SOD) and catalase (CAT)] in serum at 6h, 12h, 24h, 48h and 72h after reperfusion and in spinal cord at 72h after reperfusion. Furthermore, HS treatment (10mL/kg) reduced caspase-3 activity in the spinal cord of this model. Thus, HS may be an effective therapeutic agent for spinal cord I/R injury via activation of mitoKATP channels as well as reduction of oxidative stress, inflammatory cytokines and apoptosis.