To investigate the changes and role of hydrogen sulfide (H2S) in myocardial damage in endotoxemic rats, a rat model of endotoxemia induced by injection of lipopolysaccharide (LPS) was developed. Male Wistar rats were divided into four groups: control group, LPS group, LPS + propargylglycine (PPG, a metabolic enzyme inhibitor of H2S) group and LPS + NaHS (H2S donor) group. The mean arterial pressure (MAP) of rats within 4 h was observed, TNF-alpha and H2S contents in plasma, TNF-alpha and H2S contents, lactate dehydrogenase (LDH) and myeloperoxidase (MPO) activity in cardiac muscles were determined. The morphological structure of cardiac muscle was observed. Administration of LPS caused a sustained fall in MAP within 4 h, and significant increases in TNF-alpha and H2S contents in plasma (P<0.05). Plasmic H2S content was negatively correlated with MAP (r = -0.936, -0.913 and -0.908 at 1, 2 and 4 h, respectively, P<0.05). LPS also induced increases in TNF-alpha and H2S contents, LDH and MPO activity in cardiac muscles and myocardial damage. Treatment with PPG reduced the increases in TNF-alpha and H2S contents in plasma, TNF-alpha and H2S contents, LDH and MPO activity in cardiac muscles, ameliorated the hypotensive effect and myocardial damage caused by LPS administration (P<0.05). However, treatment with NaHS increased TNF-alpha and H2S contents in plasma, TNF-alpha and H2S contents, LDH and MPO activity in cardiac muscles, and aggravated the hypotensive action and tissue injuries caused by LPS administration (P<0.05). It is suggested that hypotension and myocardial damage in endotoxemic rats are partly induced by increase in H2S content.
Animals ; Blood Pressure ; Endotoxemia ; metabolism ; pathology ; Hydrogen Sulfide ; blood ; Lipopolysaccharides ; toxicity ; Male ; Myocardium ; chemistry ; pathology ; Peroxidase ; metabolism ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVETo observe the effects and mechanisms of endotoxin pretreatment on the rat lung in endotoxemia.

METHODSEighty-four male Wistar rats were divided into seven groups (each group containing 12 rats): saline control and lipopolysaccharide (LPS)-treated 2 h, 4 h, 6 h groups and LPS-pretreated 2 h, 4 h, 6 h groups. LPS-pretreated rats were administrated with intraperitoneal injection of 0.25 mg/kg LPS. After 24 hours, they were injected with 0.5 mg/kg of LPS. Saline control and LPS-treated rats received an equivalent amount of saline. After 72 hours, LPS-treated and LPS-pretreated rats were intravenously injected with 10 mg/kg of LPS. An equivalent amount of saline was injected in the control rats. Blood was drawn from the carotid artery in LPS-treated and LPS-pretreated rats and sacrificed after intravenous injection of LPS 2, 4, 6 hours. Following saline injection of control rats, blood was drawn from the carotid artery after 6 hours. Arterial blood was drawn for blood gas analysis. The lungs were removed for detecting the mRNA levels of intercellular adhesion molecule-1 (ICAM-1) by reverse transcription polymerase chain reaction and the protein levels of inhibitor kappa B-alpha (I kappa B-alpha) by immunohistochemical staining. Bronchoalveolar lavage was performed in the right lung. Cell counts were evaluated with a light microscopy. The supernatant of bronchoalveolar lavage fluid (BALF) was assayed for the level of protein. The whole lung was weighed and the value was used to determine the lung-body index. The tissue was homogenized and centrifuged for the determination of myeloperoxidase enzyme (MPO) activity.

RESULTSThe rats exposed to LPS alone demonstrated an increase in lung-body index, protein in BALF, and MPO activity in the lung tissue. In contrast, the rats exposed to LPS pretreatment exhibited a significant decrease in lung-body index, protein in BALF, and MPO activity. There was a significant decrease in the level of arterial bicarbonate in the LPS-treated rats in comparison with saline-treated and LPS-pretreated animals at 2 hours to 6 hours after LPS administration. The decrease of arterial bicarbonate was compensated by alveolar hyperventilation in LPS-treated animals, with a significant decrease in partial pressure of carbon dioxide. At the same time, partial pressure of oxygen decreased significantly compared with saline control animals and LPS-pretreated animals. LPS-treated rats showed a significant gradually increase in ICAM-1mRNA in the lung in comparison with the saline group. In contrast, ICAM-1mRNA levels in rats pretreated with LPS was lower than that in LPS-treated rats. In LPS-treated animals, LPS caused a decrease of I kappa B-alpha protein expression at 2 hours, returned to control level at 4 hours, and remained at 6 hours. There was no decrease of I kappa B-alpha protein expression in LPS-pretreated animals.

CONCLUSIONThe results in this study showed that administration of a small dose of LPS 72 hours before endotoxemia caused a attenuation effect on lung injury, which may be correlated to I kappa B-alpha expression induced by LPS pretreatment.

Animals ; Carbon Dioxide ; blood ; Endotoxemia ; metabolism ; I-kappa B Proteins ; analysis ; metabolism ; Immunohistochemistry ; Intercellular Adhesion Molecule-1 ; genetics ; Lipopolysaccharides ; pharmacology ; Lung ; metabolism ; pathology ; Male ; NF-KappaB Inhibitor alpha ; Oxygen ; blood ; Peroxidase ; metabolism ; RNA, Messenger ; analysis ; Rats ; Rats, Wistar

OBJECTIVETo study hepatic NF-κB level following endotoxemic liver injury, and its relationship with hepatic TNF-α and IL-6 levels in young rats.

METHODSForty 18-day-old rats were randomly assigned to a normal control and an endotoxemia group. Endotoxemia was induced by lipopolysaccharide injection (LPS, 5 mg/kg). The endotoxemia group was subdivided into four groups sampled at 2, 6, 12 and 24 hrs after LPS injection (n=8 each). Pathological changes in liver cells were observed under a light microscope. TNF-α and IL-6 levels in liver tissue homogenates were measured using ELISA. Reitman-Frankel was used to measure serum ALT concentrations. NF-κB activation level in liver tissues was detected by immunohistochemistry.

RESULTSLiver tissue injury was the most obvious 6 hrs after LPS injection under the light microscope, and the damage rating of liver tissues was significantly higher in the endotoxemia group than that in the normal control group at all time points (P<0.05). ALT levels in the endotoxemia group were significantly higher than those in the normal control group 6, 12 and 24 hrs after LPS injection (P<0.05). NF-κB p65 protein expression in liver cells (percentage of nuclear positive cells) in the endotoxemia groups was significantly higher than that in the normal control group (P<0.05). TNF-α and IL-6 levels in liver tissue homogenates in the endotoxemia groups were significantly higher than those in the normal control group 6 and 12 hrs after LPS injection (P<0.05).

CONCLUSIONSEndotoxemia can cause liver injury, resulting in liver cell damage and changes in liver function. NF-κB activation is involved in endotoxemic liver injury which may be mediated by inflammatory cytokines TNF-α and IL-6 synthesis.

Alanine Transaminase ; blood ; Animals ; Behavior, Animal ; Endotoxemia ; complications ; Female ; Interleukin-6 ; analysis ; Liver ; chemistry ; pathology ; Liver Diseases ; etiology ; metabolism ; Male ; NF-kappa B ; analysis ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha ; analysis

Atelectasis can impair arterial oxygenation and decrease lung compliance. However, the effects of atelectasis on endotoxemic lungs during ventilation have not been well studied. We hypothesized that ventilation at low volumes below functional residual capacity (FRC) would accentuate lung injury in lipopolysaccharide (LPS)-pretreated rats. LPS-pretreated rats were ventilated with room air at 85 breaths/min for 2 hr at a tidal volume of 10 mL/kg with or without thoracotomy. Positive end-expiratory pressure (PEEP) was applied to restore FRC in the thoracotomy group. While LPS or thoracotomy alone did not cause significant injury, the combination of endotoxemia and thoracotomy caused significant hypoxemia and hypercapnia. The injury was observed along with a marked accumulation of inflammatory cells in the interstitium of the lungs, predominantly comprising neutrophils and mononuclear cells. Immunohistochemistry showed increased inducible nitric oxide synthase (iNOS) expression in mononuclear cells accumulated in the interstitium in the injury group. Pretreatment with PEEP or an iNOS inhibitor (1400 W) attenuated hypoxemia, hypercapnia, and the accumulation of inflammatory cells in the lung. In conclusion, the data suggest that atelectasis induced by thoracotomy causes lung injury during mechanical ventilation in endotoxemic rats through iNOS expression.
Animals ; Blood Pressure ; Carbon Dioxide/blood ; Cardiac Output ; Combined Modality Therapy ; Endotoxemia/*complications/immunology/pathology ; Functional Residual Capacity ; Immunohistochemistry ; Leukocytes, Mononuclear/pathology ; Lipopolysaccharides/pharmacology ; Lung/enzymology/pathology/physiopathology ; Lung Compliance ; Lung Volume Measurements ; Male ; Neutrophils/pathology ; Nitric Oxide Synthase Type II/metabolism ; Oxygen/blood ; Positive-Pressure Respiration/*adverse effects ; Pulmonary Atelectasis/*etiology/pathology/*therapy ; Rats ; Rats, Sprague-Dawley ; Thoracotomy/*adverse effects

In light of the anti-inflammatory properties of histone deacetylase (HDAC) inhibitors, such as suberoylanilide hydroxamic acid (SAHA) and trichostatin A (TSA), we examined a new HDAC inhibitor KBH-A42 for its anti-inflammatory activities. KBH-A42 showed noteworthy anti-inflammatory properties in vitro via suppression of the production of TNF-alpha, a proinflammatory cytokine, and nitric oxide (NO), a proinflammatory effector molecule, in LPS-stimulated RAW264.7 cells and peritoneal macrophages. It also inhibited TNF-alpha production in vivo as demonstrated in a LPS-induced mouse endotoxemia model. The levels of TNF-alpha, IL-1beta, IL-6 and iNOS mRNAs determined by RT-PCR propose that the inhibition of these pro-inflammatory mediators by KBH-A42 resulted from inhibiting expression of these genes. However, the EMSA study to see the effect of KBH-A42 on the binding of NF-kappaB, a transcription factor, to a specific DNA sequence showed that the binding of NF-kappaB to DNA was not changed regardless of increasing the concentration of KBH-A42 in the presence and absence of LPS stimulation. Interestingly, DNA binding of another transcription factor AP-1 dose-dependently increased by KBH-A42. KBH-A42 differentially regulated the phosphorylation of MAP kinases. While the phosphprylation of ERK1/2 and SAPK/JNK was not affected by KBH-A42, the phosphorylation of p38 decreased by KBH-A42. These results showed that KBH-A42 inhibits production of proinflammatory cytokines in macrophages by decreasing their mRNA levels, and p38 kinase is involved in the KBH-A42-mediated inhibition.
Animals ; Blotting, Western ; Cell Line ; Cell Survival/drug effects ; Cytokines/blood/genetics/*metabolism ; Electrophoretic Mobility Shift Assay ; Endotoxemia/blood/metabolism/pathology ; Enzyme Inhibitors/chemistry/*pharmacology ; Histone Deacetylases/*antagonists & inhibitors ; Hydroxamic Acids/chemistry/*pharmacology ; Interleukin-1beta/genetics/metabolism ; Interleukin-6/genetics/metabolism ; Macrophages/cytology/*drug effects/metabolism ; Mice ; Mitogen-Activated Protein Kinase 1/metabolism ; Mitogen-Activated Protein Kinase 3/metabolism ; Mitogen-Activated Protein Kinases/metabolism ; Molecular Structure ; NF-kappa B/metabolism ; Nitric Oxide/metabolism ; Nitric Oxide Synthase Type II/genetics/metabolism ; Phosphorylation/drug effects ; Piperidones/chemistry/*pharmacology ; Protein Binding/drug effects ; Reverse Transcriptase Polymerase Chain Reaction ; Transcription Factor AP-1/metabolism ; Tumor Necrosis Factor-alpha/blood/genetics/metabolism