Hydrogen-Rich Saline Protects Against Spinal Cord Injury in Rats

In the present study, we examined the mechanisms of hydrogen-rich saline, a reported therapeutic antioxidant, in the treatment of acute spinal cord contusion injury. Male Sprague-Dawley rats were used to produce a standardized model of contuses spinal cord injury (125 kdyn force). Hydrogen-rich saline was injected intraperitoneally (5 ml/kg) immediately, and at 24 and 48 h after injury. All rats were sacrificed at 72 h after spinal cord injury (SCI). Apoptotic cell death, oxidative stress, inflammation, level of Brain derived neurotrophic factor (BDNF) were evaluated. In addition, locomotor behavior was assessed using the Basso, Beattice and Bresnahan (BBB) scale. We observed that administration of hydrogen-rich saline decreased the number of apoptotic cells, suppressed oxidative stress, and improved locomotor functions. Hydrogen-rich saline increased the release of BDNF. In conclusion, hydrogen-rich saline reduced acute spinal cord contusion injury, possibly by reduction of oxidative stress and elevation of BDNF.     

Airway Surface Dehydration Aggravates Cigarette Smoke-Induced Hallmarks of COPD in Mice

Introduction

Airway surface dehydration, caused by an imbalance between secretion and absorption of ions and fluid across the epithelium and/or increased epithelial mucin secretion, impairs mucociliary clearance. Recent evidence suggests that this mechanism may be implicated in chronic obstructive pulmonary disease (COPD). However, the role of airway surface dehydration in the pathogenesis of cigarette smoke (CS)-induced COPD remains unknown.

Objective

We aimed to investigate in vivo the effect of airway surface dehydration on several CS-induced hallmarks of COPD in mice with airway-specific overexpression of the β-subunit of the epithelial Na⁺ channel (βENaC).

Methods

βENaC-Tg mice and wild-type (WT) littermates were exposed to air or CS for 4 or 8 weeks. Pathological hallmarks of COPD, including goblet cell metaplasia, mucin expression, pulmonary inflammation, lymphoid follicles, emphysema and airway wall remodelling were determined and lung function was measured.

Results

Airway surface dehydration in βENaC-Tg mice aggravated CS-induced airway inflammation, mucin expression and destruction of alveolar walls and accelerated the formation of pulmonary lymphoid follicles. Moreover, lung function measurements demonstrated an increased compliance and total lung capacity and a lower resistance and hysteresis in βENaC-Tg mice, compared to WT mice. CS exposure further altered lung function measurements.

Conclusions

We conclude that airway surface dehydration is a risk factor that aggravates CS-induced hallmarks of COPD.     

Beneficial Effects of Hydrogen-Rich Saline on Early Burn-Wound Progression in Rats

Introduction

Deep burn wounds undergo a dynamic process known as wound progression that results in a deepening and extension of the initial burn area. The zone of stasis is more likely to develop more severe during wound progression in the presence of hypoperfusion. Hydrogen has been reported to alleviate injury triggered by ischaemia/reperfusion and burns in various organs by selectively quenching oxygen free radicals. The aim of this study was to investigate the possible protective effects of hydrogen against early burn-wound progression.

Methods

Deep-burn models were established through contact with a boiled, rectangular, brass comb for 20 s. Fifty-six Sprague-Dawley rats were randomly divided into sham, burn plus saline, and burn plus hydrogen-rich saline (HS) groups with sacrifice and analysis at various time windows (6 h, 24 h, 48 h) post burn. Indexes of oxidative stress, apoptosis and autophagy were measured in each group. The zone of stasis was evaluated using immunofluorescence staining, ELISA, and Western blot to explore the underlying effects and mechanisms post burn.

Results

The burn-induced increase in malondialdehyde was markedly reduced with HS, while the activities of endogenous antioxidant enzymes were significantly increased. Moreover, HS treatment attenuated increases in apoptosis and autophagy postburn in wounds, according to the TUNEL staining results and the expression analysis of Bax, Bcl-2, caspase-3, Beclin-1 and Atg-5 proteins. Additionally, HS lowered the level of myeloperoxidase and expression of TNF-α, IL-1β, and IL-6 in the zone of stasis while augmenting IL-10. The elevated levels of Akt phosphorylation and NF-κB p65 expression post burn were also downregulated by HS management.

Conclusion

Hydrogen can attenuate early wound progression following deep burn injury. The beneficial effect of hydrogen was mediated by attenuating oxidative stress, which inhibited apoptosis and inflammation, and the Akt/NF-κB signalling pathway may be involved in regulating the release of inflammatory cytokines.     

Aging does not Enhance Experimental Cigarette Smoke-Induced COPD in the Mouse

It has been proposed that the development of COPD is driven by premature aging/premature senescence of lung parenchyma cells. There are data suggesting that old mice develop a greater inflammatory and lower anti-oxidant response after cigarette smoke compared to young mice, but whether these differences actually translate into greater levels of disease is unknown. We exposed C57Bl/6 female mice to daily cigarette smoke for 6 months starting at age 3 months (Ayoung@) or age 12 months (Aold@), with air-exposed controls. There were no differences in measures of airspace size between the two control groups and cigarette smoke induced exactly the same amount of emphysema in young and old. The severity of smoke-induced small airway remodeling using various measures was identical in both groups. Smoke increased numbers of tissue macrophages and neutrophils and levels of 8-hydroxyguanosine, a marker of oxidant damage, but there were no differences between young and old. Gene expression studies using laser capture microdissected airways and parenchyma overall showed a trend to lower levels in older animals and a somewhat lesser response to cigarette smoke in both airways and parenchyma but the differences were usually not marked. Telomere length was greatest in young control mice and was decreased by both smoking and age. The senescence marker p21^Waf1 was equally upregulated by smoke in young and old, but p16^INK4a, another senescence marker, was not upregulated at all. We conclude, in this model, animal age does not affect the development of emphysema and small airway remodeling.     

Intrathecal Infusion of Hydrogen-Rich Normal Saline Attenuates Neuropathic Pain via Inhibition of Activation of Spinal Astrocytes and Microglia in Rats

Background

Reactive oxygen and nitrogen species are key molecules that mediate neuropathic pain. Although hydrogen is an established antioxidant, its effect on chronic pain has not been characterized. This study was to investigate the efficacy and mechanisms of hydrogen-rich normal saline induced analgesia.

Methodology/Principal findings

In a rat model of neuropathic pain induced by L5 spinal nerve ligation (L5 SNL), intrathecal injection of hydrogen-rich normal saline relieved L5 SNL-induced mechanical allodynia and thermal hyperalgesia. Importantly, repeated administration of hydrogen-rich normal saline did not lead to tolerance. Preemptive treatment with hydrogen-rich normal saline prevented development of neuropathic pain behavior. Immunofluorochrome analysis revealed that hydrogen-rich normal saline treatment significantly attenuated L5 SNL-induced increase of 8-hydroxyguanosine immunoreactive cells in the ipsilateral spinal dorsal horn. Western blot analysis of SDS/PAGE-fractionated tyrosine-nitrated proteins showed that L5 SNL led to increased expression of tyrosine-nitrated Mn-containing superoxide dismutase (MnSOD) in the spinal cord, and hydrogen-rich normal saline administration reversed the tyrosine-nitrated MnSOD overexpression. We also showed that the analgesic effect of hydrogen-rich normal saline was associated with decreased activation of astrocytes and microglia, attenuated expression of interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) in the spinal cord.

Conclusion/Significance

Intrathecal injection of hydrogen-rich normal saline produced analgesic effect in neuropathic rat. Hydrogen-rich normal saline-induced analgesia in neuropathic rats is mediated by reducing the activation of spinal astrocytes and microglia, which is induced by overproduction of hydroxyl and peroxynitrite.     

Effects of Hydrogen-Rich Saline on Hepatectomy-Induced Postoperative Cognitive Dysfunction in Old Mice

This study aims to investigate the protective effects and underlying mechanisms of hydrogen-rich saline on the cognitive functions of elder mice with partial hepatectomy-induced postoperative cognitive dysfunction (POCD). Ninety-six old male Kunming mice were randomly divided into 4 groups (n?=?24 each): control group (group C), hydrogen-rich saline group (group H), POCD group (group P), and POCD?+?hydrogen-rich saline group (group PH). Cognitive function was subsequently assessed using Morris water-maze (MWM) test. TNF-α and IL-1β levels were measured by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry, along with NF-κB activity determined by ELISA. The morphology of hippocampal tissues were further observed by HE staining. Learning and memory abilities of mice were significantly impaired at day 10 and day 14 post-surgery, as partial hepatectomy significantly prolonged the escape latency, decreased time at the original platform quadrant and frequency of crossing in group P when compared to group C (p?<?0.05). The surgery also increased the contents of TNF-α, IL-1β, and NF-κB activity at all time points after surgery (p?<?0.05). The introduction of hydrogen-rich saline (group PH) partially rescued spatial memory and learning as it shortened escape latency and increased time and crossing frequency of original platform compared to group P (p?<?0.05). Moreover, such treatment also decreased TNF-α and IL-1β levels and NF-κB activity (p?<?0.05). In addition, cell necrosis in the hippocampus induced by hepatectomy was also rescued by hydrogen-rich saline. Hydrogen-rich saline can alleviate POCD via inhibiting NF-κB activity in the hippocampus and reducing inflammatory response.     

烟雾暴露所致肺气肿小鼠模型的建立与评价

目的探讨单纯烟雾暴露所致肺气肿小鼠模型建立及病理学、气道炎症及肺功能评价,并进行支气管肺泡灌洗(bronchoalveolar lavage,BAL)技术的改进。方法 20只C57BL/6J小鼠随机分为正常对照及烟雾暴露组,烟雾暴露90d并观察30d后行小鼠肺功能检查、应用改进方法留取BALF行细胞计数及行肺组织病理切片观察,并与正常对照组进行比较。结果烟雾暴露组小鼠气道阻力(Raw)较正常对照组增高,动态肺顺应性(Cdyn)降低;BALF中细胞总数高于正常对照组,巨噬细胞数(AM)、中性粒细胞数(N)、中性粒细胞所占比例(N%)也高于对照组,差异均有统计学意义;病理学观察示烟雾暴露组肺泡腔扩大、部分肺泡间隔断裂、肺泡腔融合、肺气肿形成,气道上皮排列紊乱、部分气道上皮增生、周围炎症细胞浸润并伴有平滑肌增生;形态学计量分析示烟雾暴露组平均内衬间隔(MLI)及肺泡破坏指数(DI)较正常对照组增加。应用改进技术行BAL成功率100%,回收率高达90%。结论单纯烟雾暴露可以成功建立小鼠肺气肿模型且稳定可靠,与人类慢性阻塞性肺病相似性好,经BAL技术改进后该模型可行性高。     

Hydrogen-Rich Saline is Cerebroprotective in a Rat Model of Deep Hypothermic Circulatory Arrest

Deep hypothermic circulatory arrest (DHCA) has been widely used in the operations involving the aortic arch and brain aneurysm since 1950s; but prolonged DHCA contributes significantly to neurological deficit which remains a major cause of postoperative morbidity and mortality. It has been reported that hydrogen exerts a therapeutic antioxidant activity by selectively reducing hydroxyl radical. In this study, DHCA treated rats developed a significant oxidative stress, inflammatory reaction and apoptosis. The administration of HRS resulted in a significant decrease in the brain injury, together with lower production of IL-1β, TNF-α, 8-OHdG and MDA as well as decreased activity of NOS while increased activity of SOD. The apoptotic index as well as the expressions of caspase-3 in brain tissue was significantly decreased after treatment. HRS administration significantly attenuated the severity of DHCA induced brain injury by mechanisms involving amelioration of oxidative stress, down-regulation of inflammatory factors and reduction of apoptosis.     

Cigarette Smoke-Induced Emphysema and Pulmonary Hypertension Can Be Prevented by Phosphodiesterase 4 and 5 Inhibition in Mice

Rationale

Chronic obstructive pulmonary disease (COPD) is a widespread disease, with no curative therapies available. Recent findings suggest a key role of NO and sGC-cGMP signaling for the pathogenesis of the disease. Previous data suggest a downregulation/inactivation of the cGMP producing soluble guanylate cyclase, and sGC stimulation prevented cigarette smoke-induced emphysema and pulmonary hypertension (PH) in mice. We thus aimed to investigate if the inhibition of the cGMP degrading phosphodiesterase (PDE)5 has similar effects. Results were compared to the effects of a PDE 4 inhibitor (cAMP elevating) and a combination of both.

Methods

C57BL6/J mice were chronically exposed to cigarette smoke and in parallel either treated with Tadalafil (PDE5 inhibitor), Piclamilast (PDE4 inhibitor) or both. Functional measurements (lung compliance, hemodynamics) and structural investigations (alveolar and vascular morphometry) as well as the heart ratio were determined after 6 months of tobacco smoke exposure. In addition, the number of alveolar macrophages in the respective lungs was counted.

Results

Preventive treatment with Tadalafil, Piclamilast or a combination of both almost completely prevented the development of emphysema, the increase in lung compliance, tidal volume, structural remodeling of the lung vasculature, right ventricular systolic pressure, and right ventricular hypertrophy induced by cigarette smoke exposure. Single, but not combination treatment prevented or reduced smoke-induced increase in alveolar macrophages.

Conclusion

Cigarette smoke-induced emphysema and PH could be prevented by inhibition of the phosphodiesterases 4 and 5 in mice.     

Treatment with Hydrogen-Rich Saline Delays Disease Progression in a Mouse Model of Amyotrophic Lateral Sclerosis

Amyotrophic lateral sclerosis (ALS) is the most frequent adult-onset motor neuron disease, and accumulating evidence indicates that oxidative mechanisms contribute to ALS pathology, but classical antioxidants have not performed well in clinical trials. The aim of this work was to investigate the effect of treatment with hydrogen molecule on the development of disease in mutant SOD1 G93A transgenic mouse model of ALS. Treatment of mutant SOD1 G93A mice with hydrogen-rich saline (HRS, i.p.) significantly delayed disease onset and prolonged survival, and attenuated loss of motor neurons and suppressed microglial and glial activation. Treatment of mutant SOD1 G93A mice with HRS inhibited the release of mitochondrial apoptogenic factors and the subsequent activation of downstream caspase-3. Furthermore, treatment of mutant SOD1 G93A mice with HRS reduced levels of protein carbonyl and 3-nitrotyrosine, and suppressed formation of reactive oxygen species (ROS), peroxynitrite, and malondialdehyde. Treatment of mutant SOD1 G93A mice with HRS preserved mitochondrial function, marked by restored activities of Complex I and IV, reduced mitochondrial ROS formation and enhanced mitochondrial adenosine triphosphate synthesis. In conclusion, hydrogen molecule may be neuroprotective against ALS, possibly through abating oxidative and nitrosative stress and preserving mitochondrial function.     

Enhanced Activation of Atrial Natriuretic Peptide (ANP) and Natriuretic Peptide Receptor-A (NPRA) in Chronic Cigarette Smoke-Induced Lung Inflammation in Experimental Rats

International Journal of Peptide Research and Therapeutics - We sought to determine the circulatory level of atrial natriuretic peptide (ANP) and its receptor (Natriuretic peptide receptor-A) on...     

富氢液对大鼠脓毒血症引起急性肺损伤的影响

目的:探讨富氢液对大鼠脓毒血症引起急性肺损伤的影响。方法:24只大鼠建立盲肠结扎穿孔法(CLP)脓毒症模型,随机均分为以下3组(n=8):假手术组(Sham组)、CLP组及CLP+富氢液组。CLP+富氢液组于CLP后即时、6及18 h腹腔分别注射富氢液5 ml/kg,CLP组及假手术组于上述相应时间点腹腔注射富氢液5 ml/kg。CLP或假手术24 h后检测肺组织TNF-α、IL-6、HMGB1、IL-10、MDA、SOD、MPO活性及干湿重比(W/D)并采用HE染色法进行肺组织学评分。结果:与Sham组比较,CLP组及CLP+富氢液组肺组织TNF-α、IL-6、HMGB1、IL-10、MDA、SOD、MPO活性、W/D及肺组织学评分明显升高而SOD活性明显降低(P<0.05);与CLP组相比,CLP+富氢液组IL-6、HMGB1、MDA、MPO明显降低及肺组织学评分明显降低而SOD明显升高(P<0.05)。结论:富氢液降低脓毒血症的炎性反应和氧化应激,从而改善ALI时的肺功能。     

A Novel Anti-Inflammatory and Pro-Resolving Role for Resolvin D1 in Acute Cigarette Smoke-Induced Lung Inflammation

Introduction

Cigarette smoke is a profound pro-inflammatory stimulus that contributes to acute lung injuries and to chronic lung disease including COPD (emphysema and chronic bronchitis). Until recently, it was assumed that resolution of inflammation was a passive process that occurred once the inflammatory stimulus was removed. It is now recognized that resolution of inflammation is a bioactive process, mediated by specialized lipid mediators, and that normal homeostasis is maintained by a balance between pro-inflammatory and pro-resolving pathways. These novel small lipid mediators, including the resolvins, protectins and maresins, are bioactive products mainly derived from dietary omega-3 and omega-6 polyunsaturated fatty acids (PUFA). We hypothesize that resolvin D1 (RvD1) has potent anti-inflammatory and pro-resolving effects in a model of cigarette smoke-induced lung inflammation.

Methods

Primary human lung fibroblasts, small airway epithelial cells and blood monocytes were treated with IL-1β or cigarette smoke extract in combination with RvD1 in vitro, production of pro-inflammatory mediators was measured. Mice were exposed to dilute mainstream cigarette smoke and treated with RvD1 either concurrently with smoke or after smoking cessation. The effects on lung inflammation and lung macrophage populations were assessed.

Results

RvD1 suppressed production of pro-inflammatory mediators by primary human cells in a dose-dependent manner. Treatment of mice with RvD1 concurrently with cigarette smoke exposure significantly reduced neutrophilic lung inflammation and production of pro-inflammatory cytokines, while upregulating the anti-inflammatory cytokine IL-10. RvD1 promoted differentiation of alternatively activated (M2) macrophages and neutrophil efferocytosis. RvD1 also accelerated the resolution of lung inflammation when given after the final smoke exposure.

Conclusions

RvD1 has potent anti-inflammatory and pro-resolving effects in cells and mice exposed to cigarette smoke. Resolvins have strong potential as a novel therapeutic approach to resolve lung injury caused by smoke and pulmonary toxicants.     

iNOS-Dependent Increase in Colonic Mucus Thickness in DSS-Colitic Rats

Aim

To investigate colonic mucus thickness in vivo in health and during experimental inflammatory bowel disease.

Methods

Colitis was induced with 5% DSS in drinking water for 8 days prior to experiment, when the descending colonic mucosa of anesthetized rats was studied using intravital microscopy. Mucus thickness was measured with micropipettes attached to a micromanipulator. To assess the contributions of NOS and prostaglandins in the regulation of colonic mucus thickness, the non-selective NOS-inhibitor L-NNA (10 mg/kg bolus followed by 3 mg/kg/h), the selective iNOS-inhibitor L-NIL (10 mg/kg bolus followed by 3 mg/kg/h) and the non-selective COX-inhibitor diclofenac (5 mg/kg) were administered intravenously prior to experiment. To further investigate the role of iNOS in the regulation of colonic mucus thickness, iNOS −/− mice were used.

Results

Colitic rats had a thicker firmly adherent mucus layer following 8 days of DSS treatment than untreated rats (88±2 µm vs 76±1 µm). During induction of colitis, the thickness of the colonic mucus layer initially decreased but was from day 3 significantly thicker than in untreated rats. Diclofenac reduced the mucus thickness similarly in colitic and untreated rats (−16±5 µm vs −14±2 µm). While L-NNA had no effect on colonic mucus thickness in DSS or untreated controls (+3±2 µm vs +3±1 µm), L-NIL reduced the mucus thickness significantly more in colitic rats than in controls (−33±4 µm vs −10±3 µm). The importance of iNOS in regulating the colonic mucus thickness was confirmed in iNOS−/− mice, which had thinner colonic mucus than wild-type mice (35±3 µm vs 50±2 µm, respectively). Furthermore, immunohistochemistry revealed increased levels of iNOS in the colonic surface epithelium following DSS treatment.

Conclusion

Both prostaglandins and nitric oxide regulate basal colonic mucus thickness. During onset of colitis, the thickness of the mucus layer is initially reduced followed by an iNOS mediated increase.     

Proliferative Activity of Liver Growth Factor is Associated with an Improvement of Cigarette Smoke-Induced Emphysema in Mice

Cigarette smoke (CS)-induced emphysema is a major component of chronic obstructive pulmonary disease (COPD). COPD treatment is based on the administration of bronchodilators and corticosteroids to control symptoms and exacerbations, however, to date, there are no effective therapies to reverse disease progression. Liver growth factor (LGF) is an albumin-bilirubin complex with mitogenic properties, whose therapeutic effects have previously been reported in a model of emphysema and several rodent models of human disease. To approach the therapeutic effect of LGF in a model of previously established emphysema, morphometric and lung function parameters, matrix metalloproteinase (MMP) activity and the expression of several markers, such as VEGF, PCNA, 3NT and Nrf2, were assessed in air-exposed and CS-exposed C57BL/6J male mice with and without intraperitoneal (i.p.) injection of LGF. CS-exposed mice presented a significant enlargement of alveolar spaces, higher alveolar internal area and loss of lung function that correlated with higher MMP activity, higher expression of 3NT and lower expression of VEGF. CS-exposed mice injected with LGF, showed an amelioration of emphysema and improved lung function, which correlated with lower MMP activity and 3NT expression and higher levels of VEGF, PCNA and Nrf2. Taken together, this study suggests that LGF administration ameliorates CS-induced emphysema, highlights the ability of LGF to promote alveolar cell proliferation and may be a promising strategy to revert COPD progression.     

The Saline Lakes of Saskatchewan IV. Primary Production by Phytoplankton in Selected Saline Ecosystems

Primary production by phytoplankton, efficiency of photosynthesis, and chlorophyll-a concentrations were determined for seven saline lakes that varied widely in ionic concentration and composition. The investigations were done during the summer months of 1972 and 1973. Productivity ranged from 0.001 to 11.135 g C m⁻³ day⁻¹ and 0.053 to 7.968 g C m⁻² day⁻¹. Highest productivities were measured in two lakes that supported blooms of Aphanizomenon flos-aquae and Nodularia spumigena, respectively. Species of Cyanophyceae, Bacillariophyceae and Chlorophyceae dominated the phytoplankton of the study lakes. Active chlorophyll-a ranged from 0.01 to 116 mg m⁻³. Integral photosynthetic efficiency estimates were <1% except during phytoplankton blooms when they were considerably higher. The overall range of 0.03 to 3.8% is concordant with estimates for other lacustrine ecosystems. The extinction of light caused by photo-synthetic processes, or in situ efficiency, was <1% in the trophogenic zone for most lakes but, it was considerably higher during blooms. In situ efficiencies invariably increased with depth in ail lakes.     

哮喘气道重塑大鼠气道上皮细胞及线粒体超微结构变化

目的探讨哮喘气道重塑大鼠气道上皮细线粒体超微结构变化,为哮喘的治疗寻求新的理论基础。方法雌性Wistar大鼠6只、随机分为2组,采用卵蛋白吸人法制作哮喘气道重塑大鼠模型;取两组大鼠气管制作电镜切片,观察两组大鼠气道纤毛上皮,气道上皮细胞问胶原沉积及细胞内线粒体超微结构改变。结果哮喘组大鼠气道多层纤毛上皮层,纤毛脱落,杯状细胞增多,排列不整齐,上皮细胞中线粒体基膜密度下降,线粒体峡减少,上皮细胞细胞核多切迹,形状不规则;气道上皮细胞间胶原沉积;板层小体和线粒体空泡化。结论哮喘大鼠气道上皮细胞出现早期凋亡改变,线粒体失去正常形态。