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Heat stroke induced cerebral damage via neuroinflammation. This study aimed to approach whether heat stress would promote NOD-like receptor protein 3 (NLRP3) inflammasome via reactive oxygen species (ROS). The mice were randomly divided into the sham group, the heat stress group, and the heat stress + TEMPOL (ROS scavenger) group. And the NLRP3 -/- mice were applied and divided into the NLRP3 -/-  + sham group and the NLRP3 -/-  + heat stress group. Furthermore, the BV2 cells were divided into four groups following the intervention measures: the heat stress + TEMPOL group, the heat stress + Z-VAD-FMK (caspase-1 inhibitor) group, the heat stress group, and the control group. ROS levels were examined. The expression levels of NLRP3, caspase-1, IL-1β, and IL-18 were detected by western blotting and double immunofluorescence. We found that heat stress attack induced excessive ROS in microglia and subsequently activated NLRP3 inflammasome in both mice and BV2 cells. When ROS scavenged, the expression level of NLRP3 was downregulated. Furthermore, with NLRP3 inflammasome activation, the expression levels of caspase-1, IL-1β, and IL-18 were increased. In NLRP3 -/- mice, however, the caspase-1, IL-1β, and IL-18 were significantly declined. Further experiments showed that pretreatment of caspase-1 inhibitor decreased the expression levels of IL-1β and IL-18. These results suggest that heat stress attack caused neuroinflammation via excessive ROS activating the NLRP3 inflammasome in microglia cells., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Abstract: Objectives: This study investigated the role and potential involvement of pulmonary vascular glycocalyx degradation in acute lung injury in rats with severe heatstroke (HS). Methods: Rats in an established HS model were exposed to a heated environment for 60 min in an incubator (temperature, 40°C ± 2°C; humidity, 65% ± 5%). Following pretreatment with heparanase III (HPSE III) or heparin, pathological lung injury, arterial blood gas, alveolar barrier disruption, and hemodynamic changes were evaluated. The vascular endothelial structures of the lungs were examined using electron microscopy. The concentration of Evans blue dye in the lungs and arterial blood gas were assessed. An enzyme-linked immunosorbent assay was used to quantify the plasma concentration of heparan sulfate proteoglycan. The expression of glypican-1 and syndecan-1 in pulmonary vessels was measured using immunofluorescence. Western blots were used to detect the expression of TNF-α, IL-6, and vascular endothelial biomarkers in the rat lungs. Pulmonary apoptosis was assessed using a TUNEL (terminal dUTP nick end labeling) assay, and the concentrations of malondialdehyde were measured. Results: Glycocalyx shedding aggravated lung injuries. Severe histopathological damage was observed, and indexes of lung function deviated from abnormal ranges. In addition, pulmonary vascular endothelial cells were disrupted. Compared with the HS group, the plasma concentration of heparan sulfate proteoglycan significantly increased in the HPSE group ( P < 0.05). The expression of glypican-1 and syndecan-1 decreased, and the extravasation of Evans blue dye increased ( P < 0.01). Endothelial biomarker expression increased in the lung tissue, whereas occludin expression decreased. Moreover, TNF-α and IL-6 were overexpressed following heat stress. Furthermore, apoptosis of pulmonary tissues and the concentration of malondialdehyde in rat lungs increased in the HS and HPSE groups. Conclusions : Heatstroke induced pulmonary glycocalyx degradation, which increased vascular permeability and aggravated vascular endothelial dysfunction, contributing to apoptosis, inflammation, and oxidation in the pulmonary tissues., Competing Interests: The authors report no conflicts of interest., (Copyright © 2023 by the Shock Society.)

Abstract: Heat stroke is characterized by excessive oxidative stress and inflammatory responses, both of which are implicated in vascular endothelial glycocalyx shedding and heat-stroke mortality. Although molecular hydrogen has antioxidation and anti-inflammatory potency, its effect on the vascular endothelial glycocalyx in heat stroke has not been examined. Therefore, the aim of this study was to investigate the influence of hydrogen inhalation on the survival and thickness of the vascular endothelial glycocalyx of rats subjected to heat stroke. Altogether, 98 Wistar rats were assigned to the experiments. A heat-controlled chamber set at 40°C temperature and 60% humidity was used to induce heat stroke. After preparation, the anesthetized rats that underwent the heating process were subjected to an hour of stabilization in which 0%, 2%, or 4% hydrogen gas was inhaled and maintained until the experiment ended. In addition to survival rate assessments, blood samples and left ventricles were collected to evaluate the thickness of the vascular endothelial glycocalyx and relevant biomarkers. The results showed that 2% hydrogen gas significantly improved survival in the heat-stroked rats and partially preserved the thickness of the endothelial glycocalyx. In addition, serum levels of endotoxin, syndecan-1, malondialdehyde, and tumor necrosis factor-α decreased, whereas superoxide dismutase levels increased, indicating that inhalation of 2% hydrogen attenuated the damage to the vascular endothelial glycocalyx through its antioxidative and anti-inflammatory effects., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 by the Shock Society.)

This study aims to examine microcirculation and systemic hemodynamic disturbances in severe heat stroke (HS). A total of 147 rats were divided into HS group (HS), pretreated with superoxide dismutase (SOD+HS) group, and pretreated with normal saline (NS+HS) group. Heat stress was induced by incubating the animals in certain temperatures. Blood flow and vascular reactivity were monitored dynamically with intravital microscopy. Pulmonary permeability was reflected by wet-to-dry weight ratio, the concentration of Evans Blue (EB), and histopathology of lung. The results showed that heat stress could induce blood flow rate reduced, and SOD exhibited better protective role in blood flow rate. The arteriolar reactivity threshold to norepinephrine was markedly reduced at core temperature of 41°C, but no significant decrease occurred in SOD+HS group. Water content and EB concentration in lung tissue in HS group were increased along with temperature rise. SOD treatment could attenuate those changes. The pathological lung injury caused by heat stress was also milder in SOD+HS group than that in other two groups. Mean arterial pressure decreased at early stages of heat stress, but there was no decrease in SOD+HS group. There was a significant body weight loss during heat stress in all groups. Survival time in SOD+HS group was longer than that in other two groups. These results suggest that microcirculation disturbance occurs not only at the early stage but also before systemic hemodynamic disorder, monitoring microcirculation following HS is of prognostic value, and intervention with antioxidative agents may have certain protecting effects in severe HS.

To explore the roles of mesenteric lymph on lung injury in heatstroke (HS), HS rat model was prepared in a prewarmed incubator. Vascular endothelium injury biomarkers (circulating endothelial cell [CEC] as well as von Willebrand factor [vWF] and thrombomodulin [TM]), proinflammatory factors (tumor necrosis factor-α [TNF-α], interleukin-1β [IL-1β], IL-6, and high mobility group box 1), and coagulant markers (activated partial thromboplastin time, prothrombin time, D-Dimer, and platelet count) were tested in HS and HS with mesenteric lymph duct ligation (LDL) rats. In addition, lung histopathology; arterial blood gas; Evans Blue dye (EBD) and protein lung permeability; intralung inflammatory parameters including bronchoalveolar lavage fluid (BALF) TNF-α, IL-1β, and IL-6 levels; myeloperoxidase (MPO) activity; and vWF immune staining were analyzed. LDL prolonged HS onset time but not HS survival time. LDL significantly attenuated endothelial cell injury for decreased CEC counts as well as plasma vWF and TM concentrations; downregulated systemic inflammation for decreased plasma TNF-α, IL-1β, IL-6, and high mobility group box 1 levels; and ameliorated coagulant disorders for decreased activated partial thromboplastin time, prothrombin time, and D-Dimer levels as well as increased platelet counts. LDL also significantly reduced acute lung pathological injury; improved lung function indexes including arterial blood PaO2, pH, PaCO2, and lactic acid; decreased BALF TNF-α, IL-1β, and IL-6 levels and lung MPO activity; improved EBD and protein lung permeability; and inhibited lung vascular endothelium vWF expression. However, all of these parameters were not recovered to the normal states. In summary, LDL developed protection roles systemically and alleviated lung injury in HS rats which indicated that modulating mesenteric lymph flow may have some potential benefits in HS.

Objective: Dysfunction of the intestinal barrier plays an important role in the pathological process of heatstroke. Omega-3 (or n-3) polyunsaturated fatty acids, including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), help protect the intestinal mucosal barrier. This study assessed if pretreating rats with EPA or DHA could alleviate heat stress-induced damage to the intestinal barrier caused by experimental heatstroke., Methods: Male Wistar rats were pregavaged with either EPA, DHA, corn oil, or normal saline (all 1 g/kg) for 21 days before the heatstroke experiment (control rats were not exposed to heat). Experimental rats were exposed to an ambient temperature of 37°C and 60% humidity to induce heatstroke, and then they were allowed to recover at room temperature after rapid cooling. Survival time of rats was monitored after heatstroke. Horseradish peroxidase flux from the gut lumen and the level of plasma D-lactate were measured to analyze intestinal permeability at 6 h after heatstroke. Plasma endotoxin levels were determined using a limulus amoebocyte lysate assay. Expressions of the tight junction (TJ) proteins occludin and ZO-1 were analyzed by Western blot and localized by immunofluorescence microscopy. Tight junction protein morphology was observed by transmission electron microscopy. Fatty acids of ileal mucosa were analyzed using gas chromatography-mass selective detector., Results: Eicosapentaenoic acid significantly increased survival time after heatstroke. Eicosapentaenoic acid significantly decreased intestinal permeability and plasma endotoxin levels. Eicosapentaenoic acid effectively attenuated the heatstroke-induced disruption of the intestinal structure and improved the histology score, whereas DHA was less effective, and corn oil was ineffective. Pretreatment with EPA also increased expression of occludin and ZO-1 to effectively prevent TJ disruption. Eicosapentaenoic acid pretreatment enriched itself in the membrane of intestinal cells., Conclusions: Our results indicate that EPA pretreatment is more effective than DHA pretreatment in attenuating heat-induced intestinal dysfunction and preventing TJ damage. Enhanced expression of TJ proteins that support the epithelial barrier integrity may be important for maintaining a functional intestinal barrier during heatstroke.

Background: Heatstroke can result in significant diffuse tissue derangement, which can result in multiple organ system dysfunction. The heart can equally be affected and ischemia and infarction may occur., Objective: This study aimed to present the potential complications from heatstroke to the myocardium., Case: A case of a 15-year-old adolescent boy who collapsed after playing football in a hot summer day was found to be hyperthermic and poorly perfused. He had ischemic electrocardiographic changes and elevated cardiac enzymes but with normal coronary arteries., Conclusions: Heatstroke can lead to morbidity and mortality. Tissue damage during heatstroke is believed to result from uncoupling during oxidative phosphorylation. It is important to realize that heart damage can occur from heatstroke and that appropriate diagnostic and therapeutic measures are required for a good outcome.

Heat stroke (HS) is defined clinically as a condition when core body temperature rises above 40°C and is accompanied by central nervous system abnormalities. In this study, we established a rat model of HS by exposing anesthetized rats to elevated ambient temperature (40°C) until core temperature reaching 40.5°C (HS onset). The rat was immediately removed from heating chamber, allowed recovery for various time periods, and killed for histological and biochemical studies. Our results indicated neuronal shrinkage and pyknosis of the nucleus and sustained up to 12 h recovery time in cerebral cortex. Elevated expression of autophagy-related proteins, including microtubule associated protein light chain 3 and beclin 1 in cortical tissue at various times (3, 6, 12 h) of recovery was observed. In addition, the number of autophagosomes stained by monodansylcadaverine, a specific autophagosome marker, increased after heat exposure but was reduced by pretreatment with 3-methyladenine, an autophagy inhibitor. Furthermore, heat exposure increased neuronal degeneration in cortical tissue, as evidenced by staining with the fluorescent dye Fluoro-Jade B for degenerating neuron. Pretreatment with 3-methyladenine in HS rats aggravated neurodegeneration. Taken together, these results suggest that HS induces autophagy as a protection mechanism against neurodegeneration. Modulation of autophagy may provide a potential therapeutic approach for HS and await further research.

Objective: We tested the hypothesis that the antithrombotic and cytoprotective effects of recombinant human activated protein C (rhAPC) protect baboons against the lethal effects of heatstroke., Methods and Results: Fourteen anesthetized baboons assigned randomly to rhAPC (n=7) or control group (n=7) were heat-stressed in a prewarmed incubator at 44 to 47 degrees C until systolic blood pressure fell below 90 mm Hg, which signaled severe heatstroke. rhAPC was administered intravenously (24 microg/kg/h) for 12 hours at onset of heatstroke. Heat stress induced coagulation and fibrinolysis activation as evidenced by a significant increase from baseline levels in plasma levels of thrombin-antithrombin (TAT) complexes, tissue plasminogen activator, and D-dimer. Heat stress elicited cell activation/injury as assessed by the release of interleukin (IL)-6, soluble thrombomodulin, and procoagulant microparticles (MPs). rhAPC did not significantly reduce heatstroke-induced thrombin generation, and D-dimer and had no effect on fibrinolytic activity. In contrast, rhAPC infusion attenuated significantly the plasma rise of IL-6 and inhibited the release of soluble thrombomodulin and MPs as compared with control group. No difference in survival was observed between rhAPC-treated and control group., Conclusions: rhAPC given to heatstroke baboons provided cytoprotection, but had no effect on heatstroke-induced coagulation activation and fibrin formation. Inhibition of MPs by rhAPC suggested a novel mechanism of action for this protein.

Objective: Severe heatstroke is a leading cause of morbidity and mortality during heat waves. The pathogenesis of tissue injury, organ failure, and death in heatstroke is not well understood., Methods and Result: We investigated the pathways of heatstroke-induced tissue injury and cell death in anesthetized baboons (Papio hamadyras) subjected to environmental heat stress until core temperature attained 42.5 degrees C (moderate heatstroke; n = 3) or onset of severe heatstroke (n = 4) signaled by a fall in systolic blood pressure to < 90 mm Hg and rise in core temperature to 43.1+/-0.1 degrees C. Three sham-heated animals served as controls. Light and electron microscopy revealed widespread hemorrhage and thrombosis, transmural migration of leukocytes, and microvascular endothelium injury in severe heatstroke. Immunohistology and ultrastructural analysis demonstrated increased staining of endothelial von Willebrand factor (vWF), tissue factor (TF), and endothelial leukocyte-platelet interaction. Extensive apoptosis was noted in spleen, gut, and lung, and in hematopoeitic cells populating these organs. Double-labeling studies colocalized active caspase-3 and TF with apoptotic cells. Findings in sham-heated animals were unremarkable., Conclusions: These data suggested that microvascular injury, thrombosis, inflammation, and apoptosis may play an important role in the pathogenesis of heatstroke injury.

Objective: Heat stroke is a life-threatening illness characterized by an increased core body temperature as a result of exposure to high ambient temperature. Despite advances in supportive care, heat stroke is often fatal, and no specific and effective therapies exist. The pathophysiological responses to heat stroke involve a systemic inflammatory response and a disseminated intravascular coagulation in the host, which lead to a multiorgan dysfunction syndrome. Poly(ADP-ribose) polymerase-1 (PARP-1) is a nuclear DNA-binding protein that has been shown to play a relevant role in cell necrosis and organ failure in various diseases associated with inflammation. Therefore, we set out to investigate whether inhibition of PARP activity might affect the heat stroke-induced injury., Design: Controlled animal study., Setting: Research laboratory of an academic institution., Subjects: PARP-1-deficient mice (Parp-1(-/-)) and wild-type mice (C57BL/6J)., Interventions: Wild-type mice untreated or treated with either PJ34 or 3-AB, two generic PARP inhibitors, and Parp-1(-/-) mice were subjected to heat exposure as a model to study heat stroke., Measurements and Main Results: We measured rectal temperature, serum interleukin-1beta and interleukin-6, liver histology, and heat shock proteins expression. We found that the heat stroke-induced injury was attenuated in mice lacking PARP-1 and was markedly reduced in wild-type mice treated with PARP inhibitors. Interestingly, heat-induced expression of heat shock proteins 27 and 70 was boosted after PARP inhibition. Indeed, PARP inhibition increased expression of heat shock proteins 27 and 70 even in the absence of heat exposure. Accordingly, PARP inhibition increased thermal tolerance that may contribute to attenuate the clinical effects of heat stroke, resulting in increased survival., Conclusions: Our results find a new protective function of PARP inhibitors and support their potential therapeutic application in the treatment of heat stroke.

Objective: Heatstroke is characterized by hyperthermia, vasoplegic shock, and cerebral ischemia and hypoxia. Hyperbaric oxygen (HBO) has been shown to reduce brain ischemia and behavioral dysfunction during cerebral artery occlusion. The efficacy of HBO therapy for resuscitation from heatstroke remains to be determined in the laboratory., Design: Anesthetized rats were randomized to several groups and administered: 1) no resuscitation (normobaric air) after onset of heatstroke, 2) HBO for 1 hr (100% oxygen at 253 kPa for 1 hr), 3) cyclic HBO intermitted by a 5-min air break for 1 hr of treatment (100% oxygen at 253 kPa), 4) hyperbaric air (air at 253 kPa for 1 hr), 5) normobaric hyperoxia (100% oxygen at 101 kPa for 1 hr), or 6) 8% HBO (hyperbaric 8% oxygen at 253 kPa for 1 hr)., Setting: Laboratory investigation., Subjects: Sprague-Dawley rats (300- to 400-g males)., Interventions: Rats were exposed to an ambient temperature of 43 degrees C to induce heatstroke. Their colonic temperature; mean arterial pressure; heart rate; arterial blood levels of pH, Paco2, Pao2, So2%, and tumor necrosis factor-alpha; the cortical levels of ischemic and damage markers, and cortical neuronal damage scores were determined. The moment at which mean arterial pressure began to decrease from peak levels was arbitrarily taken as the onset of heatstroke., Main Results: Survival time (interval between onset of heatstroke and animal death) was 19 +/- 1 (n = 10), 131 +/- 18 (n = 14), 159 +/- 28 (n = 13), 72 +/- 14 (n = 10), 68 +/- 12 (n = 10), and 45 +/- 11 (n = 10) mins, respectively, for normobaric air, HBO for 1 hr, cyclic HBO, hyperbaric air, normobaric hyperoxia, and 8% HBO groups. The heatstroke induced arterial hypotension and bradycardia, decreased arterial levels of pH, Pao2, and So2%, increased arterial levels of tumor necrosis factor-alpha, and increased values of cellular ischemia and damage markers. In addition, neuronal damage scores in the cortex were significantly reduced by HBO for 1 hr and cyclic HBO resuscitation., Conclusion: We successfully demonstrated that HBO and, to some extent, hyperbaric air, normobaric hyperoxia, or HBO 8% was found beneficial in resuscitating rats with experimental heatstroke. HBO effectively reduced heatstroke-induced arterial hypotension, hypoxia, plasma tumor necrosis factor-alpha overproduction, and cerebral ischemia and damage and improved survival.