Role of Nuclear Transcription Factor-κB in Endotoxin-induced Shock in Rats

To investigate the role of NF-κB in endotoxic shock in rats. the model of endotoxinshock rats was induced by intravenous infusion of lipopolysaccharidc (LPS). 1 h. 2 h. 4 h and 6 h after LPS injection, the activation of NF-κB in blood mononuclear cells and the content of TNF-α and IL-6 in plasma was detected by enzyme-linked immunoadsordent assay (ELISA). The level of mean arterial pressure (MAP) and the histopathological changes of lung and liver were also observed. The activation of NF-κB in mononuclear cells increased 1 h after LPS injection and reached its peak 2 h after the injection, and its level was higher than that of normal group. The level of TNF-α was increased 1 h after the infusion and peaked 2 h after the injection, and its level was higher than that of normal group after LPS infusion. The content of IL-6 increased gradually with time. the IL-6 level was higher than that of normal group after LPS injection. MAP was decreased gradually with time and its level was lower than that of normal group after LPS injection. Pathological examination showed that endotoxic shock could cause pulmonary alveolar hemorrhage, edema and infiltration of inflammatory cell in lung tissue and congestion, edema, capillary dilation and inflammatory cell infiltration in liver tissue. It is concluded that NF-κB can up-regulate the expression of TNF-α and IL-6 in plasma and play an important role in endotoxin induced shock in rats.     

Role of Shenfu Injection (参附注射液) in rats with systemic inflammatory response syndrome

Objective： To investigate the role of Shenfu Injection （参附注射液, SFI） in rats with systemic inflammatory response syndrome （SIRS）. Methods： The SIRS rat model was induced by the intravenous injection of lipopolysaccharide （LPS）. Forty-five male Wistar rats were randomly divided into 3 groups, the sham operative control group （control group, n=5）, the SIRS model group （model group, n=20） and the SFI treatment group （SFI group, n=20）. LPS was injected through the external jugular vein （12 mg/kg, 6 mg/mL） to all rats except for those in the control group, and SFI （10 mL/kg） was given to those in the SF group only once through intraperitoneal injection, while the normal saline （10 mL/kg） was given to those in the model group. For those in the control group, normal saline was given through the external jugular vein （2 mL/kg） and intraperitoneal injection （10 mL/kg）. Then, rats in the model group and SFI group were divided into 4 subgroups according to the time points, i.e., 1 h, 2 h, 4 h and 6 h subgroups, 5 rats in each group. The activity of nuclear factor of κB （NF-κB） of in blood mononuclear cells and the plasma levels of tumor necrosis factor- α （TNF- α ） and interleukin 6-（IL-6） were determined using enzyme-linked immunoabsordent assay （ELISA） at 1 h, 2 h, 4 h and 6 h after modeling. Histopathologic changes of the lung and liver were observed under a light microscope. Results： Compared with the control group, the activity of NF-κB in mononuclear cells and the plasma level of TNF-α were obviously increased at each time points （all P〈0.01）, reaching the peaks at 2 h after modeling. The plasma level of IL-6 increased gradually as time went by in the model group （P〈0.01）. Pathological examination showed pulmonary alveoli hemorrhage, edema and inflammatory cell infiltration in the lung tissue, and angiotelectasis, congestion, and local necrosis in the liver tissue in the model group. Compared with the model group, the activity     

Changes of sulfur dioxide, nuclear factor-κB, and interleukin-8 levels in pediatric acute lymphoblastic leukemia with bacterial inflammation

Background Bacterial inflammation is a common complication in patients with leukemia,and sulfur dioxide （SO2） is a bioactive molecule in modulating Gram-negative bacilli infection.This study aimed to examine the changes in SO2,nuclear factor-κB （NF-κB）,and interleukin-8 （IL-8） levels in pediatric acute lymphoblastic leukemia （ALL） with Gram-negative bacterial inflammation.Methods Fifty-five ALL children were enrolled in this study,including 30 males and 25 females,aged 3-13 years,and the median age was 7.8 years.All these children who accepted chemotherapy for ALL were divided into the control group （before chemotherapy）,the infection group （after chemotherapy with infection）,and the recovery group （the infection was controlled after 1 week）.The serum level of SO2 was detected using high performance liquid chromatography with fluorescence assay,and NF-κB and IL-8 levels were measured by enzyme-linked immunosorbent assay （ELISA）.Human THP-1 cells were cultured,induced,and differentiated into macrophages,which were divided into five groups and each group was cultured with different stimulators：lipopolysaccharide （LPS） group,LPS＋L-aspartate-β-hydroxamate （HDX） group,LPS＋SO2 group,SO2,and control groups.NF-κB level and IL-8 protein contents by ELISA were examined in each group.Results In comparison with those of the control group,levels of serum SO2,NF-κB,and IL-8 of the infection group were significantly increased （P ＜0.05）,while those of the recovery group were significantly decreased （P ＜0.05）.A positive correlation was found between levels of serum SO2 and intracellular NF-κB/IL-8,and the correlation coefficients were 0.671 and 0.798 （P ＜0.05）,respectively.According to the results found in human THP-1 cells,levels of NF-κB and IL-8 in LPS group were significantly increased compared with those of the control group （P ＜0.05）; when compared with those in LPS group,levels of NF-κB in LPS＋HDX group further increased significantly （P ＜     

Role of liver X receptors alpha agonist on expressions of LPS-induced inflammatory response associated factor IRAK-4 and NF-kappaB in Kupffer cells

Objective： To explore the role of activated liver X receptor α （LXRα） on the expressions of interleukin-1 receptor associated kinase-4 （IRAK-4） and NF-kappaB （NF-κB） in the inflammatory response which induced by LPS in the Kupffer cells and to investigate the possible mechanisms of LXRα negative regulation of inflammatory response. Methods： The Kupffer cells were isolated from male Kunming mice by collagen perfusion in situ. And these cells were divided into 4 groups： normal control group, LPS treatment group, LXRct agonist T0901317 treatment group, LPS and T0901317 combined treatment group. The LPS treatment group were treated with a final concentration of 1 μg/ml LPS in RPMI 1640 and cultured for 6 h, the T0901317 treatment group were treated with a final concentration of 5 μg/ml in RPMI 1640 and cultured for 24 h, and the combined treatment group received pre-culture for 24 h with a final concentration of 1μg/ml T0901317 in RPMI 1640 and then cultured for 6 h with a final concentration of 5 μg/ml LPS in RPMI 1640. All groups were cultured for 30 h. The expression of LXRα, IRAK-4 and NF-κB at mRNA and protein levels were detected by real-time PCR and Western blotting, and the TNF-α and IL-1β levels were detected by ELISA. Results： The levels of LXRα mRNA and protein were highest in T0901317 group, and lowest in LPS group （P〈0.05）. The level of IRAK4 and NF-κB mRNAs and proteins were evidently lower in the Combined-treated group than in LPS group （P〈0.05）. And the level of TNF-α and IL-1 were observed highest in LPS group （P〈0.05）, but no difference among the Control group, T0901317 group and Combined-treated group （P〉0.05）. Conclusion： These date suggest that the LXR agonists can effectively up-regulate the expressions of LXRα mRNA and protein and inhibit the inflammatory response. This may be via down-regulating the expressions of IRAK4 and NF-κB at mRNA and protein levels.     

Role of NF-κB in protection of EPO pretreatment on neonatal rat cardiac myocytes with hypoxia/reoxygenation injury

Objective：To observe the protective effects of erythropoietin （EPO） pretreatment on cardiac myocyte with hypoxia/reoxygenation （H/R） injury and the role of NF-κBin this effects. Methods：After the H/R model of cardiac myocytes of neonatal rats was established, the cultured cardiac myocytes were divided into 4 groups, including EPO pretreatment group （ EPO 10 U/ml 24 h before H/R）, EPO pretreatment ＋ PDTC group（EPO 10 U/ml and PDTC 5 μg/ml 24 h before H/R）, PDTC group （PDTC 5 μg /ml 24 h before H/R） and eomrolgroup. Before and after the H/R, assay of LDH concentration in the culture medium, the survival rate of the myocytes tested by MTT chromatometry and the apoptosis by flow cytometry were undertaken. Activation of NF-κB was determined by EMSA before and after H/R. Results：EPO pretreatment markedly reduced the LDH concentration in the medium, elevated the survival rate of myocytes and inhibited the apoptosis after H/R. Addition of PDTC during the pretreatment abol- ished the protective effects of EPO pretreatment. NF-κB was markedly activated during EPO pretreatment and PDTCinhibited the activation. However, after H/R, the activity of NF-κB in myocytes with EPO pretreatment was significantly inhibited compared to the other myocytes. Conclusion：NF-κB is significantly activated during EPO pretreatment, but is inhibited after H/R, which is correlated with the protective effects of EPO pretreatment on cardiac myocytes with H/R. This phenomenon can be explained as the negative feedback mechanism of the activation of NF-κB.     

LPS-induced NF-κB activation requires Ca2+ as a mediator in isolated pancreatic acinar cells of rat

Objective To investigate the effect of Ca2+ on lipopolysaccharide (LPS)-induced NF-κB activation in pancreatic acinar cells and the role of NF-κB in LPS-induced acinar cell injury. Methods Male rat pancreatic acinar cells were isolated by collagenase digestion, then exposed to varying concentrations of LPS (from 1 to 20 mg/L) in the presence or absence of EGTA. At various time points (30 minutes, 1 hour, 2 hours, 4 hours and 10 hours) after treatment with the agents, cell viability was determined by MTT. Nuclear translocation of NF-κB’s subunit p65 was visualized by immunofluorescence staining and nuclei protein was extracted to perform EMSA which was used to assay the activity of NF-κB binding to the DNA sequence containing the recognition site of NF-κB. Results LPS induced cell damage in a time- and concentration-dependent manner while EGTA attenuated LPS-induced cell damage (P<0.05). NF-κB p65 immunofluorescence staining had increased intensity in the cytoplasm and indicated that nuclear translocation occurred within 30 minutes and its zenith was reached at 1 hour after LPS (10 mg/L) treatment. Testing of NF-κB DNA binding activity showed the same alteration phase as p65 immunofluorescence staining. NF-κB activation preceded the pathological alteration of pancreatic acinar cells. The Ca2+ chelator EGTA inhibited LPS-induced NF-κB activation. Conclusions NF-κB activation is an important early event in LPS-induced injury to pancreatic acinar cells. Ca2+ is an important mediator in the process of LPS-induced NF-κB activation.     

热毒宁注射液对脂多糖诱导急性肺损伤大鼠的抗感染作用

Objective： To evaluate the protective effects of Reduning Injection （热毒宁注射液, RDN）, a patent Chinese medicine, on lipopolysaccharide （LPS）-induced acute lung injury （ALl） in rats and its underlying mechanisms of action. Methods： Sixty male Sprague-Dawley rats were randomly divided into 6 groups, including normal control, model, dexamethasone （DEX, 5 mg/kg）, RDN-H （720 mg/kg）, RDN-M （360 mg/kg） and RDN-L （180 mg/kg） groups, with 10 rats in each group. Rats were challenged with intravenous injection of LPS 1 h after intraperitoneal treatment with RDN or DEX. At 6 h after LPS challenge, lung tissues and bronchoalveolar lavage fluid （BALF） were collected, and the number of inflammatory cells was determined. The right lungs were collected for histopathologic examination, measurement of gene and protein expressions, superoxide dismutase （SOD） and myeloperoxidase （MPO） activities. Results： In vivo pretreatment of RDN （360, 720 mg/kg） significantly reduced the weight of wet to dry （W/D） ratio of lung, protein content in BALF, and led to remarkable attenuation of LPS-induced histopathological changes in the lungs. Meanwhile, RDN enormously decreased BALF total inflammatory cells, especially neutrophil and macrophage cell numbers. Moreover, RDN increased SOD activity, inhibited MPO activity, alleviated LPS-induced tumor neurosis factor-o~ （TNF-o~） and inducible nitric oxide synthase （iNOS） expression in lung tissues. Furthermore, RDN （720 mg/kg） efficiently weakened nuclear factor- kappa B （NF- K B） gene and protein expression. Conclusion： Anti-inflammatory effects of RDN was demonstrated to be preventing pulmonary neutrophil infiltration, lowering MPO activity, TNF-oL and iNOS gene expression by inhibiting NF- K B activity in LPS-induced ALl.     

Anti-Inflammatory Effects of Redlining Injection (热毒宁注射液)on Lipopolysaccharide-lnduced Acute Lung Injury of Rats

Objective:To evaluate the protective effects of Reduning Injection(热毒宁注射液.RON),a patent Chinese medicine,on lipopolysaccharide(LPS)-induced acute lung injury(ALI)in rats and its underlying mechanisms of action.Methods:Sixty male Sprague-Dawley rats were randomly divided into 6 groups,including normal control,model,dexamethasone(DEX,5 mg/kg),RDN-H(720 mg/kg),RDN-M(360 mg/kg)and RDN-L(180 mg/kg)groups,with 10 rats in each group.Rats were challenged with intravenous injection of LPS 1 h after intraperitoneal treatment with RDN or DEX.At 6 h after LPS challenge,lung tissues and bronchoaiveolar lavage fluid(BALF)were collected,and the number of inflammatory cells was determined.The right lungs were collected for histopathologic examination,measurement of gene and protein expressions,superoxide dismutase(SOD)and myeloperoxidase(MPO)activities.Results:In vivo pretreatment of RDN(360,720 mg/kg)significantly reduced the weight of wet to dry(W/D)ratio of lung,protein content in BALF,and led to remarkable attenuation of LPS-induced histopathological changes in the lungs.Meanwhile,RDN enormously decreased BALF total inflammatory cells,especially neutrophil and macrophage cell numbers.Moreover,RDN increased SOD activity,inhibited MPO activity,alleviated LPS-induced tumor neurosis factor-α(TNF-α)and inducible nitric oxide synthase(iNOS)expression in lung tissues.Furthermore,RDN(720 mg/kg)efficiently weakened nuclear factorkappa B(NF-κB)gene and protein expression.Conclusion:Anti-inflammatory effects of RDN was demonstrated to be preventing pulmonary neutrophil infiltration,lowering MPO activity,TNF-αand iNOS gene expression by inhibiting NF-κB activity in LPS-induced ALI.     

Expression of NF-κB in hRPE Cells Stimulated by PDTC and IL-1β

The constitutive expression of nuclear-factor-κB (NF-κB) in human pigment epithelial (hRPE) cells cultivated in vitro and the possible changes when incubated with PDTC and IL-I were investigated. The synchronized hRPE cells in vitro were divided into two groups. In nonPDTC group, hRPE cells were exposed respectively to IL-1β and NS (for detecting the constitutive expressions of NF-κB in hRPE cells) ; In PDTC group, PDTC-pretreated hRPE cells were exposed respectively to IL-1β?Aand NS. (for detecting the constitutive expression of NF-κB in PDTC-pretreated hRPE cells). The expression of NF-κB in hRPE cells in two groups was detected by immunofluorescence stain and flow cytometry. The results showed that the constitutive expression of NF-κB in hRPE cells in vitro was 8.05 %, and increased to 30.26 % by IL-1β. After PDTC pretreatment, the constitutive expression of NF-κB in hRPE cells was decreased to 3.74%, and 3.66 % by IL-l,respectively. It was concluded that the expressions of NF-κB in hRPE cells could be increased significantly by IL-1βand depressed effectively by PDTC. Also, PDTC could significantly inhibit the activation of NF-κB induced by IL-1β.     

15-deoxy-Δ12, 14-prostaglandin J2 ameliorates endotoxin-induced acute lung injury in rats

Background A proinflammatory milieu emerging in the lung due to neutrophil accumulation and activation is a key in the pathogenesis of acute lung injury （ALI）.15-deoxy-△12,14-prostaglandin J2 （15d-PGJ2）,one of the terminal products of the cyclooxygenase-2 pathway,is known to be the endogenous ligand of peroxisome proliferator-activated receptor y （PPAR-y） with multiple physiological properties.Growing evidence indicates that 15d-PGJ2 has anti-inflammatory,antiproliferative,cytoprotective and pro-resolving effects.We investigated whether 15d-PGJ2 has a protective effect against endotoxin-induced acute lung injury in rats.Methods Twenty-four male Wistar rats were randomly assigned into four groups （n=6 per group）：sham＋vehicle group,sham＋15d-PGJ2 group,LPS＋vehicle group,and LPS＋15d-PGJ2 group.The rats were given either lipopolysaccharide （LPS,6 mg/kg intravenously） or saline,and pretreated with 15d-PGJ2 （0.3 mg/kg intravenously） or its vehicle （dimethyl sulphoxide） 30 minutes before LPS.Histological alterations,wet/dry weight （W/D） ratio and myeloperoxidase （MPO） activity as well as tumor necrosis factor （TNF）-α and cytokine-induced neutrophil chemoattractant-1 （CINC-1） levels were determined in lung tissues four hours after LPS injection.Immunohistochemical analysis for intercellular adhesion molecule-1 （ICAM-1） expression and Western blotting analysis for nuclear factor （NF）-κB p65 translocation and IκBα protein levels were also studied.Results 15d-PGJ2 pretreatment significantly attenuated LPS-induced lung injury,and reduced the increased W/D ratio,MPO activity,TNF-α,CINC-1 levels,and ICAM-1 expression in the lung.15d-PGJ2 also suppressed the nuclear NF-ΚB p65 translocation and increased cytosolic IKBα levels.Conclusions 15d-PGJ2 protects against endotoxin-induced acute lung injury,most likely through the reduction of proinflammatory protein levels during endotoxemia subsequent to the inhibition of NF-ΚB activation.     

Increased expression and activity of MMP-9 in C-reactive protein- induced human THP-1 mononuclear cells is related to activation of nuclear factor kappa-B

The relation between the expression and activity of MMP-9 in C-reactive protein （CRP）-induced human THP-1 mononuclear cells and the activation of nuclear factor kappa-B （NF-κB） was studied to investigate the possible role of CRP in plaque destabilization. Human THP-1 cells were incubated in the presence of CRP at 0 （control group）, 25, 50 and 100 μg/mL （CRP groups） for 24 h. In PDTC （a specific NF-κB inhibitor） group, the cells were pre-treated with PDTC at 10 μmol/L and then with 100 μg/mL CRP. The conditioned media （CM） and human THP-1 cells in different groups were harvested. MMP-9 expression in CM and human THP-1 cells was measured by ELISA and Western blotting. MMP-9 activity was assessed by fluorogenic substrates. The expression of NF-κB inhibitor α （IκB-α） and NF-κB p65 was detected by Western blotting and ELISA respectively. The results showed that CRP increased the expression and activity of MMP-9 in a dose-dependent manner in the human THP-1 cells. Western blotting revealed that IiB-α expression was decreased in the cells with the concentrations of CRP and ELISA demonstrated that NF-κB p65 expression in the CRP-induced cells was increased. After pre-treatment of the cells with PDTC at 10 μmol/L, the decrease in IκB-α expression and the increase in NF-κB p65 expression in the CRP-induced cells were inhibited, and the expression and activity of MMP-9 were lowered too. It is concluded that increased expression and activity of MMP-9 in CRP-induced human THP-1 cells may be associated with activation of NF-κB. Down-regulation of the expression and activity of MMP-9 may be a new treatment alternative for plaque stabilization by inhibiting the NF-κB activation.     

Effects of propofol on early and late cytokines in lipopolysaccharideinduced septic shock in rats

Objective：It has been reported that the intravenous anesthetic propofol（PPF）has anti-inflammatory effects in vitro and in patients.The purpose of this study was to investigate whether PPF has anti-inflammatory effects in lipopolysaccharide（LPS）-induced septic shock by inhibiting the induction of pro-inflammatory cytokinesinterleukin-6（IL-6）and tumor necrosis factor-α（TNF-α）and high mobility group box 1（HMGB1）in rats.Methods：Thirty six male Wistar rats were randomly assigned to one of three groups（control group,PPF＋LPS group and LPS group;n=12 per group）.Control group rats received a 0.9%NaCl solution（NS）by the tail vein.The PPF＋ LPS group rats received PPF（10 mg/kg bolus,followed by infusion at 10 mg/（kg·h）through a femoral vein catheter）1 h before LPS（7.5 mg/kg）administration via the tail vein.The LPS group rats received injection of LPS（7.5 mg/kg）via the tail vein.Hemodynamic effects were recorded as well as mortality rates,and plasma cytokine concentrations（TNF-α,IL-6,HMGB1）were measured for the 24-h observation period.Results：The mean arterial pressure and heart rate of the PPF＋LPS group were more stable than those of the LPS group.The mortality at 24 h after the administration of the LPS injection was much higher in the LPS group（58.3%）compared to the PPF＋ LPS group（25.0%）.Plasma concentrations of cytokines（IL-6 and TNF-α）and HMGB1 were significantly reduced in the PPF＋LPS group compared to the LPS group（P0.05）.Conclusion：Pretreatment with PPF reduced the mortality rate of rats and attenuated the pro-inflammatory cytokine responses in an endotoxin shock model through an anti-inflammatory action inhibiting induction of HMGB1.     

The Effects of PDTC plus Leflunomide and Cyclosporine on the NF-κB Signaling Pathway in Mouse-to-rat Cardiac Xenografts

In this study, the effects of pirrolidine dithiocarbamate （PDTC） plus leflunomide （Lef） and cyclosporine （CsA） on the NF-κB signaling pathway in mouse-to-rat cardiac xeno-transplantation models were investigated. NIH mice and Wistar rats served as donors and recipients respectively. Mouse-to-rat cardiac xenotransplantation was performed. The recipients were divided into 5 groups： group A （the control group）, group B （PDTC group）, group C （PDTC plus CsA group）, group D （PDTC plus Lef group） and group E （PDTC plus Lef and CsA group）. The expressions of IKKa/[3, NF-κB-P65, IκBct, ICAM-1 and NF-κB DNA binding activity in xenograft tissues were determined by immunohistochemistry and Western blot as well as electrophoretic mobility shift assay （EMSA）. The median survival time of cardiac xenografts in the control group, PDTC group, PDTC plus CsA group, PDTC plus Lef group and PDTC plus Lef and CsA group was （2.17±0.41）, （2.33±0.52）, （4.67±1.21）, （7.00±1.79） and （9.00±1.41） days respectively. The survival time of xenografts in the PDTC plus Lef and CsA group was significantly longer than that in other four groups （P〈0.05 for all）, that in the PDTC plus Lef group longer than that in the control group, PDTC group and PDTC plus CsA group （P〈0.05 for all）, that in PDTC plus CsA group longer than the control group and PDTC group （P〈0.05 for all）. The expressions of IKKα/β, NF-κB-P65, IκBα and ICAM-1 and NF-κ3 DNA binding activity were notably increased in mouse-to-rat cardiac xenografts. The expressions were decreased in the control group, PDTC group, PDTC plus CsA group, PDTC plus Lef and PDTC plus Lef and CsA group in turn. It was concluded that PDTC plus Lef and CsA can significantly suppress the expressions of IKKα/β, NF-κB-P65, IκBα, ICAM-1 and NF-κB DNA binding activity, thereby prolonging the survival of the cardiac xenografts.     

Relationship between Carbachol Hyperstimulation-induced Pancreatic Intracelluar Trypsinogen and NF-κB Activation in Rats in vitro

The relationship between intracelluar trypsinogen activation and NF-κB activation in rat pancreatic acinar cells induced by M3 cholinergic receptor agonist （carbachol） hyperstimulation was studied. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor （pefabloc） and NF-κB inhibitor （PDTC） in vitro. Intracelluar trypsin activity was measured by using a fluorogenic substrate. The activity of NF-κB was monitored by using electrophoretic mobility shift assay. The results showed that after pretreatment with 2 mmol/L pefabloc, the activities of trypsin and NF-κB in pancreatic acinar cells treated with high concertrations of carbachol （10^-3 mol/L） in vitro was significantly decreased as compared with control group （P〈0.01 ）. The addition of 10^-2 mol/L PDTC resulted in a significant decrease of NF-κB activities in pancreatic acinar cells after treated with high concertrations of carbachol （10^-3 mol/L） in vitro, but the intracelluar trypsinogen activity was not obviously inhibited （P〉0.05）. It was concluded that intracelluar trypsinogen activation is likely involved in the regulation of high concertrations of carbachol-induced NF-κB activation in pancreatic acinar cells in vitro. NF-κB activation is likely not necessary for high concertrations of carbachol-induced trypsinogen activation in pancreatic acinar cells in vitro.     

Effects of therapeutic hypercapnia on inflammation and apoptosis after hepatic ischemia-reperfusion injury in rats

Background Therapeutic hypercapnia （TH） has been demonstrated to protect several organs ischemia-reperfusion injury.The study aimed to investigate the effects of therapeutic hypercapnia on hepatic ischemia-reperfusion injury （HIRI）.Methods Thirty adult male Wistar rats weighing （250 ± 20） g were randomized into 3 groups （n=10 in each）, group C （control group）, group A （hypercapnia group） and group B （CO2 preconditioning group）.A segmental ischemia of the liver was induced by interrupting the blood vessels including the bile duct to the median and left lateral lobes for 60 minutes and all the animals were sacrificed after 240 minutes observation period of reperfusion.Mean arterial pressure （MAP）and the blood gases were measured before ischemia （baseline） and at 30, 60, 120, 180 and 240 minutes after reperfusion.Arterial blood samples were obtained for determination of serum levels of TNF-α, IL-10, serum aspartate aminotransferase （AST） and alanine aminotransferase （ALT）.The histopathology of liver tissues was evaluated by light microscopy.The NF-κB expression and apoptotic hepatocytes were respectively determined by immunohistochemistry and TUNEL assay.Results The serum levels of liver enzymes and TNF-α were significantly decreased while the IL-10 level was significantly increased in groups A and B than in group C （P 〈0.05）, and group B surpassed group A （P 〈0.05）.The histopathological scores, the NF-κB immunohistochemical score （IHS） and apoptotic index were significantly lower in groups A and B than in group C （P 〈0.05）, and the decrease in group B was more obvious than in group A （P〈0.05）.Conclusion Therapeutic hypercapnia attenuates ischemia-reperfusion injury to the liver.Moreover, the effects of CO2preconditioning are outstandingly notable.     

Protective effects of erythropoietin on endotoxin-related organ injury in rats

Summary： The protective effect of erythropoietin （EPO） on tissues following ischemia and reperfusion injuries remains poorly understood. We aimed to investigate the effect of EPO in preventing en- dotoxin-induced organ damage. Rat model of multiple organ failure （MOF） was established by tail vein injection of 10 mg/kg lipopolysaccharide （LPS）. Recombinant human EPO treatment （5000 U/kg） was administered by tail vein injection at 30 min after LPS challenge. Twenty-four h after EPO treatment, changes in serum enzyme levels, including aspartate aminotransferase （AST）, alanine transaminase （ALT）, blood urea nitrogen （BUN） and creatinine （Cr）, were evaluated by biochemical analysis. Serum levels of tumor necrosis factor-tx （TNF-ct） were determined by using immunoradiometric assay. Histological examination of tissue sections was carried out by hematoxylin and eosin staining, while ul- trastructure evaluation of organ tissues was assessed by transmission electron microscopy. Protein ex- pression levels were detected by using Western blotting. EPO treatment showed a modest effect in pre- venting LPS-induced elevation of AST, ALT, BUN, Cr, and TNF-ct levels, and in protecting against LPS-induced tissue degeneration and injured ultrastructure in the lung, liver, and kidney. Moreover, LPS promoted phosphorylation of alanine aminotransferase （AKT） and increased nuclear factor-r,B （NF-rB） activation in the lung, liver, and kidney （P〈0.05 vs. control）. However, EPO treatment significantly de- creased the LPS-induced pAKT up-regulation in these tissues （P〈0.05 vs. LPS treatment alone）. The present study demonstrates that EPO may play a protective role against LPS-induced MOF by reducing the inflammatory response and tissue degeneration, possibly via the phosphatidylinositol 3-kinase/AKT and NF-r,B signaling pathways.     

Protective Effect of Emodin against Lipopolysaccharidesinduced Corneal Injury in Rats

Objective To investigate the effect of emodin on lipopolysaccharides （LPS）-induced corneal injury in rats. Methods Three parallel incisions on the central surface of corneal epithelium were made and LPS was applied on them to induce corneal injury in Wistar rats. All rats were randomly divided into emodin group （n=40） and keratitis group （n=40）. Rats in the emodin group received subconjunctival injection of emodin and rats in the keratitis group received its vehicle 30 minutes before LPS exposure. At different time points-1 3, 6, 12, and 24 hours after LPS exposure, the symptoms of all rats were observed and the severity of their ocular inflammation was examined with a slit lamp microscope, then 8 rats in each group were killed through cervical dislocation and their eyes were enucleated and prepared to observe pathological changes of corneal tissue under a light microscope. The activation of nuclear factor-loB （NF-κB） under different condi- tions was determined by Western blot. Immunocytochemistry staining with an antibody against intercellular adhesion molecule-1 （ICAM-1） was performed to identify positive cells in corneal tissues. Results The model of acute keratitis was successfully established in Wistar rats. LPS could induce a typical corneal inflammatory response, such as hyperemia, corneal edema and opacity, which were observed in model rats. Compared with keratitis group, both ocular behaviors and damages of the corneal structure were improved in emodin group. Furthermore, the activation of NF-κB and the expression of ICAM-1 induced by LPS were markedly inhibited in emodin group. Conclusion Emodin can inhibit the activation of NF-κB and the expression of ICAM-I induced by LPS in corneas, protect against acute corneal injury, and improve symptoms in rats.     

Effects of interleukin-18 on asthmatic airway inflammation and nuclear fa ctor kappa-B in murine models

Objective To investigate the effects of Interleukin-18 (IL-18) on asthmatic airway infla mmation and nuclear factor kappa-B (NF-κB) in a murine asthmatic model. Methods BALB/C mice were randomly divided into three groups: group A(control group,n=10) ; group B (asthmatic model group, n=10); group C (IL-18 injection group, n=10) . The asthmatic model was established in group B and C by respiratory syncytial virus (RSV) killed by ultraviolet light. Saline solution (0.1 ml) and IL-18 (0.1 ml, 1 μg) was injected in groups B and C at seven time points (1, 2, 7, 8 , 9, 21, 22 d). The symptoms and the numbers of eosinophils and plasmacytes in the airways were observed and the expression of NF-κB activation in the lung w as analyzed by Immohistochemistry (IHC) and Western blot. Results The symtoms of group C were more severe than in groups A and B. Group A did no t have EOS and plasmacytes in the airway submucosal while the numbers of eosinop hils ［15±3 (average cell counts per microscopic visual field, the same below) ］ and plasmacytes (10±2) in group B were found to have increased significantly . But the numbers of eosinophils and plasmacytes in group C were decreased sig nificantly when compared with group B (6±2 and 2±1, respectively, both P<0 .05). ISH showed that the expression of NF-κB activation in group B was stro nger than that in groups A and C. The amount of NF-κB inhibitor (IκB) in gro up A and group C were 3.5 times and 2.5 times more than that of group B respec tively via Western blot. Conclusion IL-18 can inhibit asthmatic airway inflammation in mice and its mechamism may b e due to the fact that IL-18 can inhibit the activation of NF-κB in the murin e asthmatic model.