【 Abstract 】 Objective Overexpressions of epidermal growth factor (EGF) and EGF receptor have been associated with progression and invasive phenotype of pancreatic cancer. However, the underlying molecular mechanism by which EGF worked in pancreatic cancer cells has not been completely understood. In this study, effect of EGF on the invasion and metastasis of pancreatic cancer cells and its regulatory mechanism were investigated. Methods The effects of EGF on the proliferation, adhesion and invasion of pancreatic cancer cells were detected by WST-1 proliferation assay, adhesion assay and invasive assay, respectively. The activity and expression of MMP-2 and MMP-9 were examined by zymography, Western blot and RT-PCR, respectively. The activity of NF- κ B was examined by EMSA. Results EGF could significantly promote the invasiveness of pancreatic cancer cells but did not affect cell proliferation or adhesion. The expressions of NF- κ B and MMP-9 were significantly increased by EGF, but EGF did not affect the activity and expression of MMP-2. Furthermore, EGF stimulated the NF- κ B binding activity. Pretreatment with NF- κ B inhibitors, pyrrolidine dithiocarbamate (PDTC), could significantly inhibit the activity of NF- κ B induced by EGF. Meanwhile, the EGF-induced expression and activity of MMP-9, as well as cell invasiveness were also inhibited by NF- κ B inhibitor. Conclusion EGF could increase the expression and promote the invasiveness of MMP-9 via the activation of NF- κ B in pancreatic cancer cells, which implies that NF- κ B inhibitant, such as PDTC, may diminish the invasiveness of pancreatic cancer cells.
ObjectiveTo investigate the possible protective effect of zerumbone on pancreatic injury in severe acute pancreatitis (SAP) rats, and to provide a theoretical basis for prevention and treatmen of severe acute pancreatitis. MethodsSeventy male Wistar rats were randomly divided into four groups:normal control group (NC group, n=10), SAP group (n=40), Zerumbone pretreatment group (ZER group, n=10), and Zerumbone drug control group (ZER-CON group, n=10). Rats of SAP group were divided into four time points of 1 h, 3 h, 6 h, and 12 h (n=10 each time point). SAP models were induced by retrograde injection of 5% sodium taurocholate (0.1 mL/100 g) in biliopancreatic duct in SAP group and ZER group. Rats were injected isotonic saline solution instead of taurocholate as a control in NC group and ZER-CON group. Zerumbone solution (10 mg/kg) was administered via femoral vein half an hour prior to establishing models in ZER group and ZER-CON group. All rats except SAP group were sacrificed at 12 h time point after the induction of SAP. The rats in SAP group were sacrificed at 1 h, 1 h, 3 h, 6 h and 12 h time point after induction of SAP. The mortality, ascites, serum amylase (AMY), phospholipase, and pathological examination of pancreas were observated or detected. The expression of nuclear factor-kappa B (NF-κB) p65 in pancreatic tissues was evaluated by immunohistochemistry assay. ResultsThere were no difference of the levels of mortality, ascites, serum AMY, phospholipase, pathological examination, and NF-κB p65 location expression of pancreas between the ZER-CON group and NC group (P>0.05). The above indexes in SAP group were significantly higher than those in NC group (P<0.05). However, those in ZER group were significantly lower than in SAP group (P<0.05), and higher than in NC group (P<0.05). ConclusionsZerumbone can reduce the mortality and ascites, effectively alleviate the enzyme, pathological injury, and NF-κB p65 location expression in pancreatic tissue following SAP. It may indicate that zerumbone can protect pancreatic injury in SAP via NF-κB pathway.
ObjectiveTo investigate the role and mechanism of P-selectin glycoprotein ligand-1 (PSGL-1) in hydrochloric acid-induced acute lung injury (ALI) in mice.MethodsWild-type mice (WT) and PSGL-1 knockout mice (PSGL-1 -/-) were randomly subjected to normal saline (NS) or hydrochloric acid (HCl) challenged group. The mice were intratracheally instilled with NS or HCl (1 μl/g weight) into the left lung with a catheter. After 2 hours, respiratory function index enhanced pause (Penh), PaO2 and PaO2 were analyzed. The wet to dry weight ratio (W/D) of the left lung and total protein concentration in bronchoalveolar lavage fluid (BALF) were measured. The number of leukocytes in BALF was counted too. Targeted lung tissue was processed for further HE or immunohistochemistry staining. Meanwhile, the expressions of interleukin-6 (IL-6), IL-1β, nuclear factor-κB (NF-κB), IκBa and p-IκBa in lung tissue were measured.ResultsThe Penh (4.77±1.22 vs. 5.80±0.84) and PaCO2 [(63.7±3.9) mm Hg vs. (74.4±7.4) mm Hg] in the PSGL-1 knockout mice were significantly lower than those in the WT mice after HCl stimulation (P<0.05), while the PaO2 was higher than that in the WT mice [(81.0±7.1) mm Hg vs. (62.0±8.9) mm Hg, P<0.05)]. The lung W/D ratio (4.86±0.15 vs. 5.22±0.20), protein concentration [(3.71±0.64) μg/μl vs. (4.74±0.98) μg/μl] and total leukocyte count [(13.00±2.18) ×107/L vs. (49.42±3.35) ×107/L] in BALF were significantly lower in the PSGL-1 knockout mice challenged with HCl than those in the WT mice (P<0.05). Besides, the protein expressions of IL-6, IL-1β, p65 and p-IκBa in the PSGL-1 knockout mice were lower than those in the WT mice after HCl instillation, while the IκBa expression was higher than that in the WT mice (P<0.05). More numbers of neutrophils and macrophages were found in the lung of the WT mice than the PSGL-1 knockout mice challenged with HCl. However, the differences of above values between the WT mice and the PSGL-1 knockout mice instilled with NS were not found, all of which were significantly lower than the correspongding HCl group except for IκBa (P<0.05).ConclusionPSGL-1 may play important roles in the development of HCl-induced ALI via the NF-κB signaling pathway and inflammation.
Objective To observe the effects of exogenous pulmonary surfactant (PS) on ventilation-induced lung injury (VILI) in rats, and to investigate its possible mechanisms. Methods A total of 40 Wistar rats were divided into 4 groups with randomized blocks method: control group, high tidal volume (HV) group, VILI group, and PS group, with 10 rats in each group. The control group was subjected to identical surgical procedure but was never ventilated. After 30 min of mechanical ventilation (MV) with Vt 45 ml/kg, the rats in HV group were killed immediately; rats in the VILI group were continually ventilated for up to 150 min with Vt 16 ml/kg; in the PS group, 100 mg/kg of PS administered intratracheally and with the same settings as VILI group. Mean artery pressure (MAP), blood gas analysis, lung wet to dry weight ratios (W/D), thorax-lung compliance, and cell counts in bronchoalveolar lavage fluid (BALF) were determined. Nuclear factor-κB(NF-κB) activity in lungs was measured by enzyme-linked immunosorbent assay (ELISA), interleukin-8(IL-8) in serum and BALF was determined by radioimmunoassay (RIA). Pathological examination of the lung was performed. Results Injurious ventilation significantly decreased MAP and PaO2/FiO2, but increased NF-κB activity and W/D. MAP and PaO2/FiO2 improved, but NF-κB activity, IL-8 in serum and BALF, and cell counts in BALF reduced significantly in PS group compared with those in VILI group. Histological studies showed reduced pulmonary edema and atelectasis in the PS group. Conclusion PS administered intratracheally can suppress the increased activity of NF-κB induced by VILI, exogenous PS can be used to treat VILI.
Abstract: Objective To study the preventive effect of n-3 polyunsaturated fatty acids on allograft arteriosclerosis. Methods Arterial homeotransplant model were created with 480 rats which were divided into four groups. Control group, no n-3 lyunsaturated fatty acids were taken. Group A, n-3 polyunsaturated fatty acids were taken for two weeks before operation with the dose of EPA 600mg/kg. Group B, 300 mg/kg and group C 150 mg/kg were taken respectively. The recipient’s transplanted vessel was excised after 1,7,14,21and 28 days respectively. The tissue pathological variations, ultrastructure variations and expression variations of intercellular adhesion molecule-1(ICAM-1), vascular cell adhesion molecule-1（VCAM-1）, nuclear factorkappa B（NF-κB） had been observed. Results The pathological changes occurred 7 days after operation in control group and were most prominent on the 28th day, blood vessels were obstructed and the expressions of ICAM-1, VCAM1,NF-κB were markedly intensified than those of group A, B, C (Plt;0.05). The pathological variations of transplanted vessel in group A, B, C occurred later than those in control group. The nonobstruction rates in group A, B, C were better than that in control group. The expressions of ICAM-1， VCAM-1， NF-κB in control group were ber than those in group A， B， C (Plt;0.05). The expressions of ICAM-1， VCAM-1， NF-κB after 1 day or 7 days demonstrated no statistically significant change in group A， B， C （Pgt;0.05）. The preventive effect for allograft vessel atheromatosis in group A and group B was ber than that in group C after 14, 21 and 28d (Plt;0.05). There were no significant difference between group A and group B (Pgt; 0.05). Conclusion The n-3 polyunsaturated fatty acids can prevent the allograft vessel atheromatosis, the most effective dose of n-3 polyunsaturated fatty acids is 300 mg/kg.
ObjectiveTo explore the protective mechanism and effect of the resveratrol for kidney injury of obstructive jaundice. MethodsThe rats were randomly divided into three groups: sham operation group receiving laparotomy without bile duct ligation (BDL), the obstructive jaundice group with BDL, and the obstructive jaundice + resveratrol group given resveratrol following BDL. The levels of total bilirubin (TBIL), direct bilirubin (DBIL), blood urea nitrogen (BUN), and creatinine (Cr) in the serum were tested. The superoxide dismutase (SOD) activity, malondialdehyde (MDA) content, glutathione (GSH) level in the renal tissues were detected. The expressions of the silent information regulator 1 (SIRT1) and nuclear factor-κB (NF-κB) proteins were tested by Western blot. The expression of SIRT1 mRNA was detected by RT-PCR and the renal cell apoptosis was examined by TUNEL staining. Results①Compared with the sham operation group, the levels of serum TBIL, DBIL, BUN and Cr were significantly higher (P < 0.05); the activity of SOD and level of GSH, and the expressions of SIRT1 mRNA and SIRT1 protein in the renal tissues were signi-ficantly lower (P < 0.05); the content of MDA, the expression of NF-κB protein, and the rate of cell apoptosis in the renal tissues were significantly higher (P < 0.05) in the obstructive jaundice group.②Compared with the obstructive jaundice group, the levels of serum TBIL, DBIL, BUN and Cr were significantly lower (P < 0.05); the activity of SOD and level of GSH, and the expressions of SIRT1 mRNA and SIRT1 protein in the renal tissues were significantly higher (P < 0.05); the content of MDA, the expression of NF-κB protein, and the rate of cell apoptosis in the renal tissues were significantly lower (P < 0.05) in the obstructive jaundice+resveratrol group. ConclusionThe resveratrol could alleviate renal damage and play a beneficial role to resist inflammation, oxidation, and apoptosis by activating the SIRT1 which probably inhibits the expression of NF-κB protein and promotes the activity of SOD in cholestatic kidney injury.
Objective To study the expression of nuclear factor-κBp65 (NF-κBp65) in gastric carcinoma and its relationship with vascular endothelial growth factor (VEGF). Methods The expression of NF-κBp65 and VEGF in 56 gastric carcinomas was detected with immunohistochemistry and compared with benign tissues. Results The positive rates of NF-κBp65 and VEGF in 56 gastric carcinomas were 62.5% and 76.8% respectively,and which were higher than those of gastric mucosal atypical hyperplasia (33.3% and 44.4%) and the normal gastric mucosa(0 and 8.3%) （P＜0.05,P＜0.01）.It was found that there was relationship between the expression of NF-κBp65 and the clinical stage, invasion depth of tumor and lymph node metastasis (P＜0.05),but there was no relation to the historica type (Pgt;0.05). There was positive correlation between NF-κBp65 and VEGF expression （r=0.36,P＜0.01）. Conclusion NF-κBp65 may play an important role in the development of gastric carcinoma by up-regulate the expression of VEGF.
目的探讨髓样细胞分化蛋白88（MyD88）在重症急性胰腺炎（SAP）发病机理的作用。 方法将48只小鼠按随机数字表法随机分为SAP组（32只）与正常对照组（16只）；再将2组小鼠随机（随机数字表法）分为6、12、24及48 h组，SAP组各亚组每组8只，正常对照组每亚组4只。SAP组小鼠腹腔注射20% L-精氨酸以诱导SAP模型，正常对照组小鼠仅腹腔注射生理盐水。分别于建模术后6、12、24及48 h处死小鼠，取其动脉血，采用ELISA方法检测血清中白细胞介素-1β（IL-1β）、白细胞介素-10（IL-10）及肿瘤坏死因子-α（TNF-α）浓度；同时取其胰腺组织（正常对照组仅术后6 h取材），用逆转录-聚合酶链反应（RT-PCR）方法检测胰腺组织中MyD88 mRNA和核因子-κB（NF-κB）mRNA的表达水平，并进行HE染色。 结果镜下见SAP组小鼠的胰腺组织随时间进展其炎症逐渐加重。各时点SAP组小鼠的IL-1β、IL-10及TNF-α浓度均高于正常对照组（P＜0.05）；各时点SAP组与正常对照组（6 h组）相比较，其胰腺组织中MyD88 mRNA及NF-κB mRNA的表达水平均较高（P＜0.05）。各时点SAP组小鼠MyD88 mRNA的表达水平与血清IL-1β、IL-10及TNF-α的浓度和NF-κB mRNA的表达水平均呈正相关（P＜0.01）。 结论MyD88的表达对SAP的发生和发展可能均具有重要的作用。
Objective To investigate the changes in osteoprotegerin (OPG) / receptor activator of nuclear factor-κB ligand (RANKL) ratio in sepsis-associated acute lung injury (SA-ALI) and the role of regulation of this ratio on the inflammatory response in SA-ALI. Methods Eighteen C57BL/6 male mice were randomly divided into sham operation group, cecal ligation and perforation (CLP) group and RANKL group, with 6 mice in each group. Before the experiment, the RANKL group was intraperitoneally injected with 5 μg (0.2 mL) of recombinant RANKL antibody, whereas both the sham operation group and the CLP group were intraperitoneally injected with a volume-matched normal saline. One hour later, the sham operation group underwent only abdominal exploration and repositioning, while the other groups underwent the CLP surgery to induce the SA-ALI model. After 24 h of modelling, all mice were sacrificed and samples were collected. Pathological evaluation of lung tissues was performed by haematoxylin-eosin staining; enzyme-linked immunosorbent assay was used to detect serum concentrations of interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-1β; while the mRNA and protein expression of OPG and RANKL, along with their ratio values, were detected by real-time polymerase chain reaction for quantitative analysis and protein immunoblotting. Results The SA-ALI mouse model was successfully established. Compared with the sham operation group, mice in the CLP group showed disturbed alveolar structure, obvious alveolar and interstitial haemorrhage and inflammatory cell infiltration, elevated serum levels of IL-6, TNF-α and IL-1β (P<0.05), significantly increased mRNA and protein expression of OPG and elevated OPG/RANKL ratio in lung tissue (P<0.05), whereas RANKL mRNA and protein expression was significantly decreased (P<0.05). Compared with the CLP group, the pathological damage of lung tissue in the RANKL group was reduced, the infiltration of alveolar and interstitial inflammatory cells was significantly improved, and the alveolar structure and morphology were more regular, with lower serum levels of IL-6, TNF-α and IL-1β (P<0.05), significantly lower mRNA and protein expression of OPG and OPG/RANKL ratio in lung tissue (P<0.05), and significantly higher mRNA and protein expression of RANKL in lung tissue (P<0.05). Conclusion The alteration of OPG/RANKL ratio may be related to the pathophysiological process of SA-ALI, and the decrease in its level may reflect the attenuation of the inflammatory response in SA-ALI.
ObjectiveTo investigate the effect of Metformin (MET) on the anxiety behavior of mice with Pentylenetetrazol (PTZ)-induced epilepsy and the mechanisms. MethodsSixty male 8-week-old C57BL/6 mice were randomly divided into normal control group (Normal), Temporal lobe epilepsy (TLE) model control group (TLE-con), TLE + MET treatment group (TLE-MET), and normal mice + MET intervention group (MET-con) (n=15/group). In the TLE-con group and the TLE-MET group, mice were injected intraperitoneally with PTZ every other day to establish the TLE model, while mice in the Normal group and the MET group were given the same dose of normal saline. During PTZ administration, mice in the TLE-MET treatment group and the MET-con group were intraperitoneally injected with MET at 200 mg/(kg·d) every other day, for 14 times in a total of 28 days. The mice in the Normal group and the TLE-con group were intraperitoneally injected with the same amount of normal saline. Open field test (OFT) and elevated cross maze (EPM) were used to evaluate the anxiety behavior of mice in each group, and the Western blotting analysis was performed to detect expression of Toll like receptor 4 (TLR4), Nuclear factor-kappa B (NF-κB) p65 in brain tissues. ResultsCompared with the Normal group, the TLE-con group showed decreased times in the open arm in the EPM test (P<0.01) and in the center of open field in the OFT test (P<0.01), while MET intervention could increase the times of epileptic mice in the central area and the open arm (P<0.05). Compared with the Normal group, the expression of TLR4 and NF-κB in the cerebral cortex in the TLE-con group was increased significantly (P<0.05), while MET intervention could partially decrease the expression of TLR4 and NF-κB in the cerebral cortex of epileptic mice (P<0.05). ConclusionMET may improve the anxiety behavior of epileptic mice by reducing the inflammatory TLR4–NF-κB pathway.