ABSTRACT
Objective To evaluate the effect of ulinastatin on the expression of metabotropic glutamate receptor Ⅱ ( mGluRⅡ) during cerebral ischemia-reperfusion ( I∕R) in rats. Methods Forty-eight male Sprague-Dawley rats, aged 6-8 weeks, weighing 230-280 g, were divided into 3 groups ( n=16 each) by a random number table method: sham operation group (S group), cerebral I∕R group (I∕R group) and ulinastatin group ( U group) . The model of cerebral I∕R injury was established by occluding the right middle cerebral artery using a nylon thread with a rounded tip inserted into internal carotid artery and advanced cranially until resistance was met. Middle cerebral artery occlusion was maintained for 2 h followed by 24-h reperfusion. Ulinastatin 100000 U∕kg was injected via the tail vein immediately after onset of reperfusion in group U. The neurological deficit score ( NDS) was assessed after 24 h of reperfusion. The rats were then sacrificed, and brain tissues were obtained for determination of brain infarction ( by TTC staining) , expression of IκB-α in cerebral ischemic penumbra ( by Western blot) and expression of mGluRⅡ in cerebral ischemic penumbra ( by immunofluorescent staining) . The percentage of cerebral infarct vol-ume was calculated. Results Compared with S group, the NDS and percentage of cerebral infarct volume were significantly increased, the expression of mGluRⅡ in cerebral ischemic penumbra was up-regulated, and the expression of IκB-α in cerebral ischemic penumbra was down-regulated in I∕R and U groups ( P<0. 05). Compared with I∕R group, the NDS and percentage of cerebral infarct volume were significantly de-creased, the expression of mGluRⅡ in cerebral ischemic penumbra was down-regulated, and the expres-sion of IκB-α in cerebral ischemic penumbra was up-regulated in U group ( P<0. 05) . Conclusion The mechanism by which ulinastatin mitigates cerebral I∕R injury may be related to inhibiting the expression of mGluR Ⅱ in cerebral ischemic penumbra of rats.
ABSTRACT
Objective To evaluate the effect of methylene blue (MB) on hydrogen peroxide (H2O2)-induced apoptosis in macrophages through mitochondria-dependent pathway in mice.Methods Mouse peritoneal macrophage line RAW264.7 cells were cultured in DMEM culture medium containing 10％ fetal bovine serum.Cells were divided into 6 groups (n =24 each) using a random number table method:control group (group C),H2O2 group,prophylactic different concentrations of MB groups (MB1,2 groups) and therapeutic different concentrations of MB groups (MB3.4 groups).H2O2 50 μmol/L was added to the culture medium in group H2O2.MB was added to the culture medium with the final concentrations of 0.1 μmol/L (in MB1 and MB3 groups) and 1.0 μmol/L (in MB2 and MB4 groups) at 30 min before adding H2O2 in MB1.2 groups and 30 min after adding H2O2 in MB3.4 groups.At 24 h of culture or incubation in each group,the cell survival rate was measured by methyl thiazolyl tetrazolium assay,the activity of reactive oxygen species (ROS) in cells was determined with the fluorescent probe,the lactate dehydrogenase (LDH) activity in supernatant was detected by spectrophotometry,the activity of superoxide dismutase (SOD) in cells was detected by colorimetric method,mitochondrial membrane potential (MMP) was measured using rhodamine 123 staining,the content of ATP was determined by an ATP bioluminescent method,the expression of pro-caspase-3 and spliceosomes P20 protein and P 18 protein was detected by Western blot,and cell apoptosis was detected using flow cytometry.Results Compared with group C,the cell survival rate,SOD activity and contents of MMP and ATP were significantly decreased,the ROS activity and activity of LDH in supernatant were increased,the expression of pro-caspase-3 and spliceosomes P20 protein and P18 protein was up-regulated,and early and late apoptosis rates were increased in the other five groups (P＜0.05).Compared with group H2O2,the cell survival rate,SOD activity and contents of MMP and ATP were significantly increased,the ROS activity and activity of LDH in supernatant were decreased,the expression of pro-caspase-3 and spliceosomes P20 protein and P18 protein was down-regulated,and early and late apoptosis rates were decreased in MB1-4 groups (P＜0.05).Compared with group MB1,the cell survival rate was significantly decreased,and the expression of caspase-3 spliceosome P 18 was down-regulated in group MB2,and the cell survival rate and SOD activity were significantly decreased,and the activity of ROS was increased in group MB3 (P＜0.05).Compared with group MB4,the expression of caspase-3 spliceosome P 18 was significantly down-regulated,early and late apoptosis rates were decreased,and the activity of ROS was increased in group MB2,and the activity of ROS was significantly increased in group MB3 (P＜0.05).Conclusion The mechanism by which MB attenuates H2O2-induced oxidative damage to macrophages is related to inhibiting cell apoptosis in macrophages through mitochondria-dependent pathway in mice.
ABSTRACT
Objective To evaluate the effect of dexmedetomidine on high-mobility group box 1 protein (HMGB1)/Toll-like receptors (TLRs) signaling pathway during lung injury in septic rats.Methods Twenty-four SPF healthy adult male Wistar rats,aged 15-18 weeks,weighing 200-250 g,were divided into 3 groups (n =8 each) using a random number table:sham operation group (group S),sepsis group (group Sep) and dexmedetomidine group (group D).Dexmedetomidine 25 μg/kg was intraperitoneally injected in D group,while the equal volume of normal saline was given instead in S and Sep groups.Sepsis was produced by cecal ligation and puncture in Sep and D groups.The rats were sacrificed at 24 h after operation,and the right lung was removed for examination of the pathological changes which were scored and for determination of myeloperoxidase (MPO) activity,content of interleukin-1β (IL-1β),IL-6 and tumor necrosis factor-α (TNF-α) in lung tissues (by enzyme-linked immunosorbent assay),wet to dry weight ratio (W/D ratio) and expression of HMGB1,TLR2 and TLR4 in lung tissues (by Western blot).Results Compared with group S,the MPO activity,lung injury score,W/D ratio,content of IL-1β,IL-6,TNF-α and expression of HMGB1,TLR2 and TLR4 were significantly increased in Sep and D groups (P＜0.05).Compared with group Sep,the MPO activity,lung injury score,W/D ratio,content of IL-1β,IL-6,TNF-α and expression of HMGB1,TLR2 and TLR4 were significantly decreased in group D (P＜0.05).Conclusion Dexmedetomidine reduces lung injury through inhibiting HMGB1/TLRs signaling pathway in septic rats.