EZH2-mediated regulation of NF-κB target gene expression in gastric cancer

AIM: To explore the mechanism by which over-expression of enhancer of zeste homolog 2 (EZH2) in a panel of gastric cancer cell lines is involved in tumorigenesis of gastric cancer. METHODS: Real-time PCR and Western blot were employed to examine the mRNA and protein levels of EZH2, respectively. MTS assay, cell migration and soft agar assay were performed to investigate the role of EZH2 in the regulation of stomach cancer behaviors. The effect of EZH2 on NF-κB target gene expression was determined by Luciferase reporter and real-time PCR. Co-immunoprecipitation was used to analyze the interaction of EZH2 and p65 in HEK293T cells. RESULTS: The expression levels of EZH2 were significantly increased in the gastric cancer cells compared with normal gastric epithelial cells. Pharmacological inhibition by DZNep or knockdown of EZH2 significantly compromised AGS and SNU-16 cell activity, cell migration and anchorage-independent cell growth. Moreover, siRNA knockdown of EZH2 impaired NF-κB downstream targets, such as IL-8, CXCL5 and CCL20. In addition, the interaction of EZH2 and p65 was detected. CONCLUSION: EZH2 mediates the growth of gastric cancer cells through the regulation of NF-κB downstream gene expression.     

Effect of genistein on ammonia-induced NF-κB activation in astrocytes

AIM: To explore the effect of genistein on ammonia-induced nuclear factor-κB (NF-κB) activation and the underlying mechanism.METHODS: Primary astrocyte cultures were prepared and challenged with NH₄Cl to establish a hyperammonemic model. The activation of ERK, Akt and NF-κB was examined by Western blot.RESULTS: AG1478 and genistein significantly inhibited ammonia-induced activation of ERK and Akt. Ammonia-induced NF-κB nuclear translocation was significantly inhibited by the pretreatment of LY294002, genistein and AG1478.CONCLUSION: Genistein significantly inhibited ammonia-induced ERK activation and Akt-mediated NF-κB activation, which might represent the important mechanism by which this naturally occurring substance exerts its swelling-inhibiting effect.     

Regulation of ectopic trypsin and proinflammatory cytokine expression by NF-κB and AP-1 in influenza A virus induced myocarditis

AIM: To investigate the regulatory effects of nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) on the expression of ectopic trypsin and proinflammatory cytokines in influenza A virus (IAV)-induced myocarditis. METHODS: Male BALB/c mice of 8 weeks old (n=40) were randomly divided into 4 groups: normal control group (NC), infection control group (IC), NF-κB inhibitor group (NI) and AP-1 inhibitor group (AI). The mice in NC group and IC group were instilled intranasally with 15 μL saline and 40 plaque forming units (PFU) IAV, respectively. The mice in NI group and AI group were infected intranasally with 40 PFU IAV and injected intraperitoneally with 10 mg/kg NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) or 2.5 mg/kg AP-1 inhibitor nordihydroguaiaretic acid (NDGA) once daily. The mice were euthanized at day 9 after instillation, and the hearts were removed for pathological and biochemical analysis. RESULTS: IAV infection induced significant up-regulation of ectopic trypsin, and proinflammatory cytokines interleukin 6 (IL-6), IL-1β and tumor necrosis factor-α (TNF-α) in the myocardium, and triggered acute myocarditis. PDTC significantly inhibited NF-κB activation and up-regulation of ectopic trypsin and proinflammatory cytokines, and effectively suppressed IAV replication and myocardial inflammatory response (P<0.01). NDGA effectively inhibited AP-1 activity (P<0.01) and mildly suppressed up-regulation of proinflammatory cytokines (P<0.05), but had no effects on the expression of ectopic trypsin, IAV replication and the extent of myocarditis (P>0.05). CONCLUSION: IAV infection induces up-regulation of ectopic trypsin and proinflammatory cytokines in myocardium predominantly by the activation of NF-κB. AP-1 signaling pathway might be only partially involved in the regulation of proinflammatory cytokines.     

Expression and significance of TLR4/NF-κB in pulmonary vascular remodeling in smoking rats

ATM: To observe the expression of Toll-like receptor 4 (TLR4), nuclear factor-κB subunit P65 protein (NF-κB P65) and proliferating cell nuclear antigen (PCNA) in the pulmonary vascular tissues of the rats exposed to smoke, and to explore the possible mechanism of TLR4/NF-κB signaling pathway in pulmonary vascular remodeling. METHODS: SPF male healthy rats (n=48) were randomly divided into control group, smoke exposure for 4 weeks group (S4 group), smoke exposure for 8 weeks group (S8 group) and smoke exposure for 12 weeks group (S12 group), with 12 rats in each group. HE staining was used to observe the morphological changes of pulmonary vessels, and then the pulmonary vascular wall area/total vascular area (WA%) and vascular wall thickness/vascular external diameter (WT%) were measured by the medical image analysis system. The expression of TLR4, NF-κB P65 and PCNA in the pulmonary vascular tissues was detected by immunohistochemical staining. The protein content was expressed by the average integral absorbance. The mRNA expression of TLR4 in the pulmonary vessels was detected by RT-qPCR. The relationships between WA%, WT%,TLR4 protein, TLR4 mRNA, P65 protein, PCNA protein and pulmonary vascular remodeling, and another relationships between WA%, WT%, P65 protein, PCNA protein and TLR4 protein were analyzed.RESULTS: The WA% and WT% in smoke exposure groups significantly increased compared with control group, and the ratio was proportional to the time of smoke exposure. The protein expression of TLR4, p65 and PCNA, and the mRNA expression of TLR4 in smoke exposure groups also increased significantly compared with control group. CONCLUSION: The extent of pulmonary vascular remodeling in the rats increases when the protein expression of TLR4 is up-regulated. There is a positive correlation between pulmonary vascular remodeling and the protein expression of TLR4 and NF-κB P65. Pulmonary vascular remodeling may be related to the activation of TLR4/NF-κB signaling pathway.     

Effects of NF-κB inhibitor on nerve injury in rats after intracerebral hemorrhage

AIM: To investigate the regulatory effect of nuclear factor-κB (NF-κB) inhibitor, pyrrolidine dithiocarbamate (PDTC), on nerve function and neural cell apoptosis in rats after intracerebral hemorrhage (ICH). METHODS: SPF Sprague-Dawley rats were randomly divided into 4 groups with 6 rats in each group:sham group, ICH group, PDTC at low concentration (P_low) group and PDTC at high concentration (P_high) group. Autologous blood injection was used to establish ICH model. After 2 h of surgery, the rats in P_low group and P_high group were intraperitoneal injected with PDTC at 100 mg/kg and 200 mg/kg, respectively, while rats in sham group and ICH group were injected with the same volume of saline. The neurological function score was classified with modified Longa grading method. TUNEL assay was used to detected the neural cell apoptosis, and the content of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) were measured. Furthermore, the protein levels of p-P65 and cleaved caspase-3 in brain tissues were determined by Western blot. RESULTS: Compared with sham group, the rats in ICH group had higher neurological function score (P<0.05). After treatment with PDTC, the neurological function score was decreased (P<0.05), but no significant difference between P_low group and P_high group was observed. Compared with sham group, the number of apoptotic cells in ICH group was increased (P<0.05). After treatment with PDTC, the neural cell apoptosis was restrained, and the number of apoptotic cells in P_high group was lower than that in P_low group (P<0.05). Compared with sham group, the content of MDA was increased and the activity of SOD was decreased in ICH group (P<0.05). After treatment with PDTC, the content of MDA was decreased while the activity of SOD was increased, and the variation trend was more obvious in P_high group (P<0.05). Compared with sham group, the protein levels of p-P65 and cleaved caspase-3 in ICH group were increased (P<0.05). After treatment with PDTC, the protein levels of p-P65 and cleaved caspase-3 were decreased, and those in P_high group were lower than those in P_low group. CONCLUSION: NF-κB inhibitor PDTC plays a role in the se-condary brain injury after ICH, and the protective effect increases at the higher dose. The mechanism may be related to reducing MDA content and increasing SOD activity, and further inhibiting neural cell apoptosis.     

Eeffect of HMGB1 on expression of NF-κB in BV-2 cells stimulated with Aβ25-35

AIM: To investigate the effect of high mobility group box-1 protein (HMGB1) on the expression of nuclear factor-κB (NF-κB) in BV-2 cells stimulated with amyloid β-protein (Aβ)_25-35. METHODS: Cultured BV-2 cells in logarithmic growth phase were divided into 4 groups:normal cell group (without any treatment), model group (treated with Aβ_25-35 at 40 μmol/L), RNA interference (RNAi) group (conducted with HMGB1-siRNA followed by Aβ_25-35 stimulation) and solvent control group (treated with 0.1% DMSO). After treatment with Aβ_25-35 for 24 h, the protein levels of HMGB1 and NF-κB in BV-2 cells were determined by Western blot. RESULTS: Aβ_25-35 at 40 μmol/L was used to stimulate BV-2 cells. The GFP fluorescence-tagged HMGB1-siRNA (30 nmol/L) was used to transfect BV-2 cells and its transfection efficiency was about 80%~90%. The results of Western blot showed that the protein level of HMGB1 was significantly decreased after the interference of siRNA fragment (P<0.05). The protein levels of HMGB1 and nucleic NF-κB p65 were dramatically increased in BV-2 cells stimulated with Aβ_25-35 (P<0.05). After RNA interference with HMGB1, the expression of HMGB1 and nucleic NF-κB p65 were significantly decreased in BV-2 cells stimulated with Aβ_25-35 (P<0.05). CONCLUSION: RNA interference with HMGB1 reduces the expression of nucleic NF-κB in BV-2 cells stimulated with Aβ_25-35.     

Role of TLR4-TRPC6 in LPS-induced NF-κB P65 expression and nuclear translocation in airway epithelial 16HBE cells

AIM: To investigate the role of Toll-like receptor 4 (TLR4) and transient receptor potential channel 6 (TRPC6) signaling pathway in lipopolysaccharide (LPS)-induced nuclear factor-κB (NF-κB) P65 expression and nuclear translocation in airway epithelial cells (16HBE) for supplementing the mechanism for airway inflammation. METHODS: After stimulating the 16HBE cells with LPS at 1 mg/L for 0, 0.5, 2, 6, 12 and 24 h, the expression of NF-κB P65 at mRNA and protein levels in the 16HBE cells were determined by RT-PCR and Western blot respectively, and the nuclear translocation of NF-κB P65 was detected by immunocytochemical staining method. The effects of TLR4 inhibitor CLI-095 at 5 μmol/L and TRPC6 agonist Hyp9 at 10 μmol/L on LPS (1 mg/L)-induced NF-κB P65 expression and nuclear translocation in the 16HBE cells were determined by RT-PCR, Western blot and immunocytochemical staining. RESULTS: LPS increased the mRNA and protein expression of NF-κB P65 and nuclear translocation in the 16HBE cells(P<0.05). TLR4 inhibitor CLI-095 reduced the mRNA and protein expression of NF-κB P65 and nuclear translocation induced by LPS, while Hyp9 enhanced the mRNA and protein expression of NF-κB P65 and nuclear translocation induced by LPS in the 16HBE cells(P<0.05). CONCLUSION: LPS induces the expression and nuclear translocation of NF-κB P65 in the 16HBE cells via TLR4-TRPC6 signaling pathway.     

Effect of hydrogen sulfide on myocardial fibrosis and PPARγ and NF-κB expression in rat model of diabetes

AIM: To explore the effects of hydrogen sulfide (H₂S) on the myocardial fibrosis in a rat model of diabetes and its mechanism.METHODS: Single intraperitoneal injection of streptozotocin (STZ) was utilized to establish a rat model of diabetes. Sodium hydrosulfide was used as an exogenous donor of hydrogen sulfide. Male SD rats were randomly divided into control group, STZ group, STZ+H₂S group and H₂S group. Eight weeks later, HE and VG staining methods were used to observe the collagen distribution and collagen volume fraction was measured by image analysis. The expression levels of type I collagen, PPARγ and NF-κB in the cardiac tissues were determined by Western blotting.RESULTS: Compared with control group, collagen distribution and the expression levels of type I collagen and NF-κB in the cardiac tissues were markedly increased (P<0.05), while PPARγ was significantly decreased in STZ group (P<0.05), but these indexes were reversed significantly in STZ+H₂S group (P<0.05). The expression levels of type I collagen, PPARγ and NF-κB had no significant difference between H₂S group and control group.CONCLUSION: Hydrogen sulfide attenuates cardiac fibrosis in diabetic rats, and its mechanism may be related to PPARγ-NF-κB signaling pathway.     

Influence of ginsenoside Re on vascular intima hyperplasia and NF-κB p65 signaling pathway in balloon-injured rats

AIM: To investigate the inhibitory effect of ginsenoside Re on intimal hyperplasia induced by balloon-injury and to explore the role of NF-κB p65 signaling pathway in the process. METHODS: SD rats(n=40) were divided into 5 groups randomly: sham operation group, model group, low-dose ginsenoside Re group, middle-dose ginsenoside Re group and high-dose ginsenoside Re group. The carotid artery intima injury model was established by 2F balloon catheters in all groups except the sham operation group. The day after modeling, the animals in model group and sham operation group were administered intragastrically with distilled water, and the rats in low-dose, middle-dose and high-dose ginsenoside Re groups were given ginsenoside Re at doses of 12.5 mg/kg, 25mg/kg and 50 mg/kg, respectively. After 14 continuous days, the morphological changes of the injured arteries were observed by HE staining and the lumen area, intima area and media area as well as the ratio of intimal area/media area were determined. The expression of tumor necrosis factor-α(TNF-α) and interleukin-1β(IL-1β) were detected by real-time PCR. The proliferating cell nuclear antigen(PCNA) and nuclear factor-kappa B(NF-κB) p65 were examined by immunohistochemistry.RESULTS: Compared with sham operation group, the vessel cavity was narrowed(P<0.01), the mRNA levels of TNF-α and IL-1β, and the protein expression of PCNA and NF-κB p65 were increased in model group(P<0.05). Compared with model group, the vascular intimal hyperplasia was alleviated obviously(P<0.05), and the mRNA levels of TNF-α and IL-1β, and protein expression of PCNA and NF-κB p65 were decreased in medium and high-dose ginsenoside Re groups(P<0.05). CONCLUSION: Ginsenoside Re inhibits the vascular neointimal hyperplasia induced by balloon-injury in rats, and the molecular mechanism may be related to the inhibition of NF-κB p65 signaling pathway.     

Effects of metformin combined with Ge Xia Zhu Yu decoction on TLR4/NF-κB signaling pathway in rats with polycystic ovary and insulin resis-tance

AIM: To study the effect of metformin (Met) combinated with Ge Xia Zhu Yu decoction on Toll-like receptor-4 (TLR-4)/nuclear factor-κB (NF-κB) signaling pathway in the rats with polycystic ovary syndrome (PCOS) and insulin resistance induced by dehydroepiandrosterone, and to explore the mechanisms. METHODS: PCOS rats (after induced by dehydroepiandrosterone, n=110) were randomly divided into 3 groups:model group (30 rats), Met treatment group (40 rats) and Met combinated with Ge Xia Zhu Yu decoction treatment (combination) group (40 rats). The rats in model group were given the same volume of 0.9% sodium chloride daily by gavage. The rats in Met group were given Met (270 mg·kg^-1·d^-1) by gavage. The rats in combination group were given Met (270 mg·kg^-1·d^-1) and Ge Xia Zhu Yu decoction (34.5 mg·kg^-1·d^-1) by gavage. All rats were continuously intervened for 28 d. After the intervention, blood glucose[fasting plasma glucose (FPG) and 2-hour postprandial blood glucose (2hPBG)] was measured. The mRNA expression levels of follicular epithelial NF-κB, TLR-4 and oxidized low-density lipoprotein (ox-LDL) were detected by RT-PCR. The serum levels of inflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and C-reactive protein (CRP) were also detected by ELISA. RESULTS: After the intervention, FPG, 2hPBG, and serum levels of IL-6, TNF-α and CRP in Met group and combination group were lower than those in model group (P<0.05), and those in combination group were lower than those in Met group (P<0.05). Meanwhile, the mRNA expression levels of follicular epithelial NF-κB, TLR-4 and ox-LDL in Met group and combination group were lower than those in model group (P<0.05), and those in combination group were lower than those in Met group (P<0.05). CONCLUSION: Treatment with Met combined with Ge Xia Zhu Yu decoction improves insulin resistance in PCOS rats by decreasing the levels of inflammatory factors in serum and epithelial cells of ovary and inhibiting the expression of NF-κB, TLR-4 and ox-LDL in epithelial tissue of ovary.     

Opening of ATP-sensitive K+ channels protects H9c2 cardiac cells against high glucose-induced injury and inflammation by inhibiting TLR4/NF-κB pathway

AIM: To investigate whether the opening of ATP-sensitive K⁺(K_ATP) channels protects H9c2 cardiac cells against high glucose(HG)-induced injury and inflammation by inhibiting the Toll-like receptor 4(TLR4)/nuclear factor-κB(NF-κB) pathway. METHODS: The protein levels of TLR4 and NF-κB p65 were determined by Western blot. The levels of interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α) were detected by ELISA. The cell viability was measured by CCK-8 assay. Mitochondrial membrane potential(MMP) was examined by rhodamine 123(Rh 123) staining followed by photofluorography. The intracellular levels of reactive oxygen species(ROS) were detected by 2', 7'-dichlorfluorescein- diacetate(DCFH-DA) staining followed by photofluorography. The number of apoptotic cells was observed by Hoechst 33258 nuclear staining followed by photofluorography. RESULTS: After the H9c2 cardiac cells were treated with HG(35 mmol/L glucose) for 24 h, the protein levels of TLR4 and phosphorylated NF-κB p65(p-NF-κB p65) were significantly increased. Pretreatment of the cells with 100 μmol/L diazoxide(DZ, a K_ATP channel opener) for 30 min before exposure to HG considerably blocked the up-regulation of the TLR4 and p-NF-κB protein levels induced by HG. Moreover, co-treatment of the cells with 30 μmol/L TAK-242(an inhibitor of TLR4) obviously inhibited the HG-induced up-regulation of the p-NF-κB p65 protein level. On the other hand, pretreatment of the cells with 100 μmol/L DZ had a clear myocardial protection effect, which attenuated the HG-induced cytotoxicity, inflammatory response, mitochondrial damage, oxidative stress and apoptosis, evidenced by an increase in the cell viability, and decreases in the levels of IL-1β and TNF-α, MMP loss, ROS generation and the number of apoptotic cells. Similarly, co-treatment of H9c2 cardiac cells with 30 μmol/L TAK-242 or 100 μmol/L PDTC(an inhibitor of NF-κB) and HG for 24 h also obviously reduced the above injuries and inflammation induced by HG.CONCLUSION: The opening of K_ATP channels protects H9c2 cardiac cells against HG-induced injury and inflammation by inhibiting the TLR4/NF-κB pathway.     

Effect of Yiqi Huayu Huatan decoction on expression of KLF15, HMGB1, NF-κB and its downstream inflammatory factors in rats with UUO-induced renal interstitial fibrosis

AIM: To observe the effect of Yiqi Huayu Huatan decoction (YHHD) on unilaterral ureteral obstruction (UUO)-induced renal interstitial fibrosis in rats, and to investigate the possible mechanism. METHODS: Female SD rats (n=48) were randomly divided into sham group, model group, telmisartan group, and low-, middle-and high-dose YHHD groups, with 8 rats in each group. The UUO model rats was established by ligating left ureter. The rats in sham group and model group were treated with equal volume of normal saline, others were treated with the corresponding drugs daily. After 12 weeks, the rats were sacrificed. The serum samples were collected for determining the concentrations of cystatin C (Cys-C) and uric acid (UA). The morphological changes of the renal tissue were observed by PAS staining. The collagen fiber was observed by Masson staining. The mRNA expression of Krüppel-like factor 15 (KLF15), high-mo-bility group box protein 1 (HMGB1), nuclear factor-κB (NF-κB), IκB, monocyte chemotactic protein-1 (MCP-1), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), fibronectin (FN), collagen type I (Col I) and Col-Ⅳ was detected by real-time PCR. The protein expression of KLF15, HMGB1 and NF-κB was detected by Western blot. The protein expression of MCP-1 was determined by the method of immunohistochemistry. RESULTS: Compared with sham group, the deposition rate of collagen fibers and the concentration of Cys-C in model group were significantly increased (P<0.05), the mRNA and protein expression of KLF15 was significantly down-regulated (P<0.05), while the mRNA expression of HMGB1, NF-κB, IκB, MCP-1, IL-1β, TNF-α, FN, Col I and Col Ⅳ and the protein expression of HMGB1, NF-κB and MCP-1 were significantly up-regulated (P<0.05). Compared with model group, the deposition rates of collagen fibers in middle-and high-dose YHHD groups and telmisartan group were significantly decreased (P<0.05), with down-regulated protein expression of HMGB1 and NF-κB and mRNA expression of IL-1β and TNF-α (P<0.05). The protein expression of KLF15 was significantly up-regulated in high-dose YHHD group and telmisartan group (P<0.05), while the protein expression of MCP-1 and the mRNA expression of FN were significantly down-regulated (P<0.05). The mRNA expression of KLF15 was significantly up-regulated in high-dose YHHD group (P<0.05), while the mRNA expression of MCP-1, Col I and Col IV was significantly down-regulated (P<0.05). The mRNA expression of NF-κB and IκB was significantly down-regulated and the concentration of Cys-C was significantly decreased in each dose of YHHD groups and telmisartan group (P<0.05). No significant difference of UA level among the groups was observed. CONCLUSION: YHHD alleviates renal interstitial fibrosis in a dose-dependent manner, and YHHD at high dose shows the most obvious effect. The mechanism may be associated with the up-regulation of KLF15 and the down-regulation of HMGB1, NF-κB and its downstream inflammation-related factors in the renal tissue.     

Expression of cyclooxygenase-2 in IL-1β-stimulated mesangial cells is medicated by NF-κB/IκB signal pathway

AIM: To investigate the role of NF-κB/IκB signal pathway in the regulation of cyclooxygenase-2 (COX-2) expression in human mesangial cells (HMC). METHODS: The PGE₂ concentration in supernatants of HMC was measured by radioimmunoassay. COX-2 mRNA and protein expression were determined by RT-PCR and Western blot. Electrophoretic mobility shift assay (EMSA) and Western blot were used to detect the activity of NF-κB and degradation of IκB. RESULTS: IL-1β significantly upregulated COX-2 expression and PGE₂ production in HMC. Significant up-regulation of NF-κB activation, nuclear translocation of p65 subunit, and degradation of IκB α and IκB β were observed in IL-1β-induced HMC. CONCLUSION: Expression of COX-2 in IL-1β-induced HMC is mediated by NF-κB/IκB signal pathway.     

Expression and function of TNF-α in dorsal root ganglion of rats with chronically compressed dorsal root ganglion

AIM: To investigate the role of TNF-α and NF-κB in the mechanism of neuropathic pain due to chronically compressed dorsal root ganglion (CCD).METHODS: Based on the CCD model, von Frey filaments were used to quantify behavior test. The expression changes of TNF-α and NF-κB were determined by Western blotting, and the correlation between the expression of TNF-α and the 50% paw withdrawal threshold was also analyzed. Moreover, the location of TNF-α in dorsal root ganglion (DRG) was observed with immunofluorescence double staining.RESULTS: We found 50% paw withdrawal threshold of CCD decreased at the first day after operation. The mechanical allodynia was the most obvious at postoperative 7~14 d and lasted longer than 35 d. The expression of TNF-α and NF-κB increased significantly in DRG after operation (P<0.01), especially at 7~14 d, and then restored gradually. Moreover, there was a correlation between the protein expression of TNF-α and the changes of neuropathic behavior (P<0.05).CONCLUSION: TNF-α and NF-κB are involved in the mechanism of mechanical allodynia after chronically compressed DRG.     

Effects of baicalin on glial fibrillary acidic protein and nuclear factor-κB expression in juvenile rat hippocampus after status convulsion

AIM: To study the effects of baicalin (BC) on glial fibrillary acidic protein (GFAP) and nuclear factor-κB (NF-κB) expression and neuronal apoptosis in juvenile rat hippocampus after status convulsion (SC). METHODS: One hundred and ninety five juvenile male Sprague-Dawley rats were randomly divided into 3 groups: normal saline pretreatment group (NS group), SC group and SC with BC pretreatment group (BC group). Each of these 3 groups would be subdivided into 5 subgroups sacrificed at 4 h, 12 h, 24 h, 48 h and 72 h after SC. The rat SC model was prepared by lithium-pilocarpine chemical method. The protein expression of GFAP and NF-κB was detected by the method of immunohistochemistry. The mRNA expression of GFAP was detected by RT-PCR. The neuronal apoptosis was observed by TdT-mediated dUTP nick end labeling (TUNEL). RESULTS: Compared with NS group, the GFAP positive cells was increased in SC group (P<0.05). Compared with SC group, the expression of GFAP was significantly reduced in BC group (P<0.05). Compared with NS group, the NF-κB positive cells was increased in SC group (P<0.05). Compared with SC group, the expression of NF-κB was significantly reduced in BC group. RT-PCR showed that the expression trend of GFAP mRNA was similar to that of the protein. Compared with NS group, the TUNEL positive cells in the hippocampus CA1 area in SC group increased significantly 12 h after SC (P<0.01), and reached a peak at 48 h. After the intervention with BC, the TUNEL positive cells decreased significantly between 12~48 h after SC (P<0.05 or P<0.01), but the number of TUNEL positive cells remained significantly greater than that in NS group (P<0.05). CONCLUSION: The expression of GFAP and NF-κB in the hippocampus increased after SC in rats. Baicalin decreases the expression of GFAP and NF-κB in hippocampus of rats with pilocarpine-induced seizures, and reduces the number of neuronal apoptosis, suggesting that baicalin may protect against the brain damage caused by status convulsion.     

PPARs signaling pathway is involved in diabetic hepatopathy in mice

AIM: To investigate the role of peroxisome proliferator-activated receptors (PPARs)-inflammation signaling pathways in diabetic hepatopathy. METHODS: Diabetic mouse model was established by feeding the mice with a high-energy diet for 4 weeks combined with intraperitoneal injection of streptozotocin (STZ; 40 mg·kg^-1·d^-1 for 5 d). The hepatopathy model was confirmed by histopathological observation and the indexes of liver function, such as alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP), after another 4 weeks. Moreover, fasting blood glucose (FBG), and serum levels of total cholesterol (TC), triglyceride (TG) and insulin were measured, and the HOMA insulin resistance index (HOMA-IR) was calculated. The mRNA and protein expression levels of PPARs and inflammation-related factors were measured by qPCR and Western blot, respectively. RESULTS: After treatment with STZ for 7 d, the FBG of mice exceeded 11.1 mmol/L, suggesting that the diabetic model was established. After 4 weeks, the structural deformation of the hepatocytes (including hepatocytes containing abundant fat vacuoles, and inflammatory cell infiltration), and the increases in the serum levels of insulin, HOMA-IR, TC, TG, ALT, AST and ALP were observed (P<0.01), indicating the occurrence and progression of hepatopathy in diabetic mice. Meanwhile, compared with the control group, the mRNA and protein expression of PPARα, PPARβ and PPARγ decreased, but the expression of nuclear factor-κB (NF-κB), cyclooxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS) significantly increased in the diabetic hepatopathy mice (P<0.01). CONCLUSION: Down-regulation of PPARα, PPARβ and PPARγ and activation of NF-κB-COX-2/iNOS signaling pathways may be involved in the diabetic hepatopathy in mice induced by long-term high-energy diet feeding combined with intraperitoneal injection of STZ.     

Signal transduction pathway and blocking strategies of NF-κB

NF-κB is thought of as a genetic switch to control expressions of many target genes and directly participates in pathogenesis of infection, inflammation, stress, immunoresponse, cellular apoptosis, toxic shock and tumor as well as cell-cycle regulation and cell differentiation.The overactivation of NF-κB is intimately involved in many human diseases.Various therapeutic strategies against NF-κB, to date, in-clude anti-inflammatory drugs, antioxidants, immunosuppressive agents, inhibitors of protease and protea-some, prostaglandings, nitric oxide, IL-10, microbial products, synthetic inhibitors, antisense oligon cleotides and decoy deoxyoligonucleotides.Studies are underway to develop NF-κB member-specific and cell type-specific drugs that can inhibit the activation of NF-κB only in target cells and that may become a novel way to treat the human diseases.     

Role of NF-κB in inducing HEL cell proliferation and apoptosis during human cytomegalovirus infection

AIM:To investigate whether human cytomegalovirus(HCMV) regulate human embryonic lung fibroblast(HEL) cell proliferation and apoptosis by activating NF-κB.METHODS:Immunohistochemistry and Western blot analysis were used to detect the NF-κB translocation and/or Bcl-2 and the levels of I-κBα during HCMV infection. Apoptotic cell were examined by flow cytometry, and the HEL cell proliferation was determined by MTT.RESULTS:The levels of NF-κB in the nucleus reached highly 48 h postinfection, and the levels of I-κBα were low 24 h postinfection. The activity of NF-κB was inhibited 120 h postinfection. The levels ofbcl-2was accorded with the activity of NF-κB. HCMV promoted HEL cells to proliferate before 72 h postinfection and induced apoptosis 120 h postinfection.CONCLUSION:NF-κB plays a role in HEL cell proliferation and apoptosis during HCMV infection, and it involves in the pathological mechanisms of diseases associated with HCMV infection.