AIM:To determine whether etomidate(ET)has a protective effect on retinal ganglion cells(RGCs)injured with hydrogen peroxide(H_(2)O_(2))and to explore the potential mechanism underlying the antioxidative stress effect ...AIM:To determine whether etomidate(ET)has a protective effect on retinal ganglion cells(RGCs)injured with hydrogen peroxide(H_(2)O_(2))and to explore the potential mechanism underlying the antioxidative stress effect of ET.METHODS:Cultured RGCs were identified by double immunofluorescent labeling of microtubule-associated protein 2 and Thy1.1.An injury model of H_(2)O_(2)-induced RGCs oxidative stress was established in vitro.Cells were pretreated with different concentrations of ET(1,5,and 10μmol/L)for 4h,followed by further exposure to H_(2)O_(2)at 1000μmol/L.Cell counting kit 8 and Annexin V/propidium iodide assays were applied to detect the viabilities and apoptosis rates of the RGCs at 12,24,and 48h after H_(2)O_(2)stimulation.The levels of nitric oxide,malondialdehyde,and glutathione in culture media were measured at these time points.Quantitative reverse transcription polymerase chain reaction(qRT-PCR)and Western blot were performed to observe the effects of ET on the messenger RNA and protein expression of inducible nitric oxide synthase(iNOS),nuclear factor erythroid 2-related factor 2(Nrf2),heme oxygenase 1(HO-1),glutathione peroxidase 1 and the level of conjugated acrolein in RGCs at 12,24,and 48h after H_(2)O_(2)stimulation and in the retina at 12h after optic nerve transection(ONT).RESULTS:The applications of 5 and 10μmol/L of ET significantly increased the viability of RGCs.Results from qRT-PCR indicated a decrease in the expression of iNOS and an increase in the expressions of Nrf2 and HO-1 in ETpretreated RGCs at 12,24 and 48h after H_(2)O_(2)stimulation,as well as in ET-treated retinas at 12h after ONT.Western blot analysis revealed a decrease in the expression of iNOS and levels of conjugated acrolein,along with an increase in the expressions of Nrf2 and HO-1 in ET-pretreated RGCs in vitro and ET-treated retinas in vivo.CONCLUSION:ET is a neuroprotective agent in primary cultured RGCs injured by H_(2)O_(2).The effect of ET is dosedependent with the greatest effect being at 10μmol/L.ET plays an antioxidant role by inhibiting iNOS,up-regulating Nrf2/HO-1,decreasing the production of acrolein,and increasing the scavenge of acrolein.展开更多
In patients with Alzheimer’s disease,gamma-glutamyl transferase 5(GGT5)expression has been observed to be downregulated in cerebrovascular endothelial cells.However,the functional role of GGT5 in the development of A...In patients with Alzheimer’s disease,gamma-glutamyl transferase 5(GGT5)expression has been observed to be downregulated in cerebrovascular endothelial cells.However,the functional role of GGT5 in the development of Alzheimer’s disease remains unclear.This study aimed to explore the effect of GGT5 on cognitive function and brain pathology in an APP/PS1 mouse model of Alzheimer’s disease,as well as the underlying mechanism.We observed a significant reduction in GGT5 expression in two in vitro models of Alzheimer’s disease(Aβ_(1-42)-treated hCMEC/D3 and bEnd.3 cells),as well as in the APP/PS1 mouse model.Additionally,injection of APP/PS1 mice with an adeno-associated virus encoding GGT5 enhanced hippocampal synaptic plasticity and mitigated cognitive deficits.Interestingly,increasing GGT5 expression in cerebrovascular endothelial cells reduced levels of both soluble and insoluble amyloid-βin the brains of APP/PS1 mice.This effect may be attributable to inhibition of the expression ofβ-site APP cleaving enzyme 1,which is mediated by nuclear factor-kappa B.Our findings demonstrate that GGT5 expression in cerebrovascular endothelial cells is inversely associated with Alzheimer’s disease pathogenesis,and that GGT5 upregulation mitigates cognitive deficits in APP/PS1 mice.These findings suggest that GGT5 expression in cerebrovascular endothelial cells is a potential therapeutic target and biomarker for Alzheimer’s disease.展开更多
AIM:To investigate the role of nuclear factor of activated T cell 2(NFAT2),the major NFAT protein in peripheral T cells,in sustained T cell activation and intractable inflammation in human ulcerative colitis(UC). METH...AIM:To investigate the role of nuclear factor of activated T cell 2(NFAT2),the major NFAT protein in peripheral T cells,in sustained T cell activation and intractable inflammation in human ulcerative colitis(UC). METHODS:We used two-dimensional gel-electrophoresis, immunohistochemistry,double immunohistochemical staining,and confocal microscopy to inspect the expression of NFAT2 in 107,15,48 and 5 cases of UC, Crohn's disease(CD),non-specific colitis,and 5 healthy individuals,respectively. RESULTS:Up-regulation with profound nucleo- translocation/activation of NFAT2 of lamina propria mononuclear cells(LPMC)of colonic mucosa was found specifically in the affected colonic mucosa from patients with UC,as compared to CD or NC(P<0.001,Kruskal- Wallis test).Nucleo-translocation/activation of NFAT2 primarily occurred in CD8+T,but was less prominent in CD4+T cells or CD20+B cells.It was strongly associated with the disease activity,including endoscopic stage (τ=0.2145,P=0.0281)and histologic grade(τ=0.4167, P<0.001). CONCLUSION:We disclose for the first time the nucleo-translocation/activatin of NFAT2 in lamina propria mononuclear cells in ulcerative colitis.Activation of NFAT2 was specific for ulcerative colitis and highly associated with disease activity.Since activation of NFAT2is implicated in an auto-regulatory positive feedback loop of sustained T-cell activation and NFAT proteins play key roles in the calcium/calcineurin signaling pathways,our results not only provide new insights into the mechanism for sustained intractable inflammation,but also suggest the calcium-calcineurin/NFAT pathway as a new therapeutic target for ulcerative colitis.展开更多
BACKGROUND Recent studies have demonstrated that dysfunction of the intestinal barrier is a significant contributing factor to the development of severe acute pancreatitis(SAP).A stable intestinal mucosa barrier funct...BACKGROUND Recent studies have demonstrated that dysfunction of the intestinal barrier is a significant contributing factor to the development of severe acute pancreatitis(SAP).A stable intestinal mucosa barrier functions as a major anatomic and functional barrier,owing to the balance between intestinal epithelial cell(IEC)proliferation and apoptosis.There is some evidence that calcium overload may trigger IEC apoptosis and that calcineurin(CaN)/nuclear factor of activated Tcells(NFAT)signaling might play an important role in calcium-mediated apoptosis.AIM To investigate the potential mechanisms underlying the therapeutic effect of Qingyi decoction(QYD)in SAP.METHODS A rat model of SAP was created via retrograde infusion of sodium deoxycholate.Serum levels of amylase,tumor necrosis factor(TNF-α),interleukin(IL)-6,D-lactic acid,and diamine oxidase(DAO);histological changes;and apoptosis of IECs were examined in rats with or without QYD treatment.The expression of the two subunits of CaN and NFAT in intestinal tissue was measured via quantitative realtime polymerase chain reaction and western blotting.For in vitro studies,Caco-2 cells were treated with lipopolysaccharide(LPS)and QYD serum,and then cell viability and intracellular calcium levels were detected.RESULTS Retrograde infusion of sodium deoxycholate increased the severity of pancreatic and intestinal pathology and the levels of serum amylase,TNF-α,and IL-6.Both the indicators of intestinal mucosa damage(D-lactic acid and DAO)and the levels of IEC apoptosis were elevated in the SAP group.QYD treatment reduced the serum levels of amylase,TNF-α,IL-6,D-lactic acid,and DAO and attenuated the histological findings.IEC apoptosis associated with SAP was ameliorated under QYD treatment.In addition,the protein expression levels of the two subunits of CaN were remarkably elevated in the SAP group,and the NFATc3 gene was significantly upregulated at both the transcript and protein levels in the SAP group compared with the control group.QYD significantly restrained CaN and NFATc3 gene expression in the intestine,which was upregulated in the SAP group.Furthermore,QYD serum significantly decreased the LPS-induced elevation in intracellular free Ca^(2+)levels and inhibited cell death.CONCLUSION QYD can exert protective effects against intestinal mucosa damage caused by SAP and the protective effects are mediated,at least partially,by restraining IEC apoptosis via the CaN/NFATc3 pathway.展开更多
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 poss...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.展开更多
Objective： To investigate the role of peroxisome proliferator-activated receptors δ （PPARδ） in inflammatory reaction and its possible mechanism in adipocyte. Methods：Lentivirus-mediated RNA interference （RNAi）...Objective： To investigate the role of peroxisome proliferator-activated receptors δ （PPARδ） in inflammatory reaction and its possible mechanism in adipocyte. Methods：Lentivirus-mediated RNA interference （RNAi） was used to block the expression of PPARδ in 3T3-L1 cells. In order to induce inflammation in 3T3-L1, cells were stimulated with tumor necrosis factor-α（TNFα, 20 ng/ml） for 4 h. The expression of PPARδ, nuclear factor κB （NFκB） and C reactive protein （CRP） were determined by Western blot analysis. Results：The expression of PPARδ was reduced by 80% after RNAi. Blockage of PPARδ promoted the expression of CRP and NFκB in cells stimulated with TNFα but had no effect on normal cells. Conclusion： PPARδ is involved in inflammatory reaction in adipocyte. Blockage of PPARδ can promote the inflammation mediated by inflammatory factors and increase the expression of NFκB and CRP in 3T3-L1 cells stimulated with TNFα.展开更多
Obesity is a major human health problem associated with various diseases, including cardiac injury and type 2 diabetes. Trapa japonica Flerov (TJF) has been used in traditional oriental medicine to treat diabetes. In ...Obesity is a major human health problem associated with various diseases, including cardiac injury and type 2 diabetes. Trapa japonica Flerov (TJF) has been used in traditional oriental medicine to treat diabetes. In this study, we evaluated the inhibitory effect of and the mechanism underlying the effect of TJF extract on adipogenesis in 3T3-L1 cells. The effects of TJF extract on cell viability were analyzed using a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, and the anti-adipogenic effect was measured by oil red O staining. The expression of peroxisomal proliferator activated receptor (PPAR)γ, CCAAT/enhancer-binding protein-α (C/EBP)α, adenosine monophosphate-activated protein kinase (AMPK), acetyl-CoA carboxylase (ACC), adiponectin, and fatty acid binding protein (FABP)4 involved in adipogenesis was determined by western blot analysis. TJF extract effectively inhibited lipid accumulation and the expression of PPARγ and C/EBPα in 3T3-L1 cells. TJF also increased the phosphorylation of AMPK and ACC, and decreased the expression of adiponectin and FABP4. These results indicate that TJF extract exerts its anti-obesity effect through the downregulation of adipogenic transcription factors and adipogenic marker genes.展开更多
Epidermal growth factor （EGF） may increase cell motility, an event implicated in cancer cell invasion and metastasis. However, the underlying mechanisms for EGF-induced cell motility remain elusive. In this study, w...Epidermal growth factor （EGF） may increase cell motility, an event implicated in cancer cell invasion and metastasis. However, the underlying mechanisms for EGF-induced cell motility remain elusive. In this study, we found that EGF treatment could activate Ras-related C3 botulinum toxin substrate 1 （Racl）, PI3K/Akt and p21- actived kinase （PAK1） along with cell migration. Ectopic expression of PAK1 K299R, a dominant negative PAK1 mutant, could largely abolish EGF-induced cell migration. Blocking PI3K/Akt signalling with LY294002 or Akt siRNA remarkably inhibited both EGF-induced PAK1 activation and cell migration. Furthermore, expression of dominant-negative Racl （T17N） could largely block EGF-induced PI3K/Akt-PAK1 activation and cell migration. Interestingly, EGF could induce a significant production of ROS, and N-acetyl-L-cysteine, a scavenger of ROS which abolished the EGF-induced ROS generation, cell migration, as well as activation of PI3K/Akt and PAK, but not Racl. Our study demonstrated that EGF-induced cell migration involves a cascade of signalling events, including activation of Racl, generation of ROS and subsequent activation of PI3K/Akt and PAK1.展开更多
Objective To study the effect of baicalin on the expression of receptor activator of nuclear factor-κB ligand(RANKL)and osteoprotegerin(OPG)in cultured human periodontal ligament(HPDL)cells.Methods Small interfering ...Objective To study the effect of baicalin on the expression of receptor activator of nuclear factor-κB ligand(RANKL)and osteoprotegerin(OPG)in cultured human periodontal ligament(HPDL)cells.Methods Small interfering RNA(siRNA)eukaryotic expression vector targeted transforming growth factor βⅡ receptor(TGF-β RⅡ)was constructed and transfected into T cells.HPDL cells with T cells transfected with siRNA or not were placed in the culture medium that had been added with lipopolysaccharide(LPS)and baicalin.The obtained solution was divided into six groups according to the components(group Ⅰ:HPDL cells+LPS+T cells transfected with siRNA1+baicalin;group Ⅱ:HPDL cells+LPS+T cells transfected with siRNA1;group Ⅲ:HPDL cells+LPS+T cells+baicalin;group Ⅳ:HPDL cells+LPS+T cells;group Ⅴ:HPDL cells+baicalin;group Ⅵ:HPDL cells)and was cultured for 48 hours.RT-PCR was used to observe the effect of baicalin on the expression of OPG-RANKL in HPDL cells.Results The ratio of RANKL/OPG in group Ⅰ was lower than that in group Ⅱ(P<0.01)and higher than that in group Ⅲ(P<0.01);The ratio of RANKL/OPG in group Ⅲ was lower than that in group Ⅳ(P<0.01);the ratio of RANKL/OPG in group Ⅳ was higher than that in group Ⅵ(P<0.01);the ratio of RANKL/OPG in group Ⅴ was lower than that in group Ⅵ(P<0.05).Conclusion ① Baicalin could decrease the ratio of RANKL/OPG in HPDL cells.② The TGF-β signaling transduction plays an important role in the effect of baicalin on the RANKL/OPG ratio in HPDL cells.③ Baicalin acts not only through TGF-β to regulate RANKL/OPG in HPDL cells,but also through other pathways.展开更多
Objective: To investigate the expression relationship between nuclear transcription factor kappa B1 (NFκB1) and long non-coding RNA PACER (LncRNA-PACER) in peripheral blood mononuclear cells (PBMCs) of patients with ...Objective: To investigate the expression relationship between nuclear transcription factor kappa B1 (NFκB1) and long non-coding RNA PACER (LncRNA-PACER) in peripheral blood mononuclear cells (PBMCs) of patients with pulmonary tuberculosis. Methods: From February 2018 to March 2019, 40 patients with pulmonary tuberculosis (tuberculosis group) and 40 healthy persons (control group) were collected, the levels of TNF-α, IL-6 and IL-8 in serum were detected by enzyme-linked immunosorbent assay (ELISA);the expressions of LncRNA-PACER and NFκB1 mRNAs in PBMCs were detected by real-time fluorescence quantitative PCR;Western blot was used to detect the expressions of NFκB1 and COX 2 in PBMCs;Pearson method was used to analyze the expressions of LncRNA-PACER and NFκB1 in PBMCs of patients with pulmonary tuberculosis, and the expressions of LncRNA-PACER and NFκB1 in PBMCs of patients with pulmonary tuberculosis were analyzed. Results: Compared with the control group, the expressions of TNF-α, IL-6 and IL-8 in the serum of patients with pulmonary tuberculosis was significantly increased (P<0.05), and the expressions of LncRNA-PACER, NFκB1 mRNAs, proteins and COX-2 protein in PBMCs were significantly increased (P<0.05). The expressions of LncRNA-PACER and NFκB1 proteins in PBMCs were related to the number of pulmonary lesions and pulmonary cavity (P<0.05), and there was a positive correlation between the expression of LncRNA-PACER and the expression of NFκB1 mRNA in PBMCs of patients with pulmonary tuberculosis (r = 0.873, P<0.05). Conclusions: The expressions of NFκB1 and LncRNA-PACER in PBMCs of patients with pulmonary tuberculosis are significantly increased, they are positively correlated and both of them are related to the occurrence and development of pulmonary tuberculosis.展开更多
基金Supported by the Ministry of Science and Technology of China(No.2021ZD0203104)the Science and Technology Plan Project of Shaanxi Province of China(No.2022SF-497)Xi’an Medical University Doctoral Research Fund(No.2020DOC18).
文摘AIM:To determine whether etomidate(ET)has a protective effect on retinal ganglion cells(RGCs)injured with hydrogen peroxide(H_(2)O_(2))and to explore the potential mechanism underlying the antioxidative stress effect of ET.METHODS:Cultured RGCs were identified by double immunofluorescent labeling of microtubule-associated protein 2 and Thy1.1.An injury model of H_(2)O_(2)-induced RGCs oxidative stress was established in vitro.Cells were pretreated with different concentrations of ET(1,5,and 10μmol/L)for 4h,followed by further exposure to H_(2)O_(2)at 1000μmol/L.Cell counting kit 8 and Annexin V/propidium iodide assays were applied to detect the viabilities and apoptosis rates of the RGCs at 12,24,and 48h after H_(2)O_(2)stimulation.The levels of nitric oxide,malondialdehyde,and glutathione in culture media were measured at these time points.Quantitative reverse transcription polymerase chain reaction(qRT-PCR)and Western blot were performed to observe the effects of ET on the messenger RNA and protein expression of inducible nitric oxide synthase(iNOS),nuclear factor erythroid 2-related factor 2(Nrf2),heme oxygenase 1(HO-1),glutathione peroxidase 1 and the level of conjugated acrolein in RGCs at 12,24,and 48h after H_(2)O_(2)stimulation and in the retina at 12h after optic nerve transection(ONT).RESULTS:The applications of 5 and 10μmol/L of ET significantly increased the viability of RGCs.Results from qRT-PCR indicated a decrease in the expression of iNOS and an increase in the expressions of Nrf2 and HO-1 in ETpretreated RGCs at 12,24 and 48h after H_(2)O_(2)stimulation,as well as in ET-treated retinas at 12h after ONT.Western blot analysis revealed a decrease in the expression of iNOS and levels of conjugated acrolein,along with an increase in the expressions of Nrf2 and HO-1 in ET-pretreated RGCs in vitro and ET-treated retinas in vivo.CONCLUSION:ET is a neuroprotective agent in primary cultured RGCs injured by H_(2)O_(2).The effect of ET is dosedependent with the greatest effect being at 10μmol/L.ET plays an antioxidant role by inhibiting iNOS,up-regulating Nrf2/HO-1,decreasing the production of acrolein,and increasing the scavenge of acrolein.
基金supported by STI2030-Major Projects,No.2021ZD 0201801(to JG)Shanxi Province Basic Research Program,No.20210302123429(to QS).
文摘In patients with Alzheimer’s disease,gamma-glutamyl transferase 5(GGT5)expression has been observed to be downregulated in cerebrovascular endothelial cells.However,the functional role of GGT5 in the development of Alzheimer’s disease remains unclear.This study aimed to explore the effect of GGT5 on cognitive function and brain pathology in an APP/PS1 mouse model of Alzheimer’s disease,as well as the underlying mechanism.We observed a significant reduction in GGT5 expression in two in vitro models of Alzheimer’s disease(Aβ_(1-42)-treated hCMEC/D3 and bEnd.3 cells),as well as in the APP/PS1 mouse model.Additionally,injection of APP/PS1 mice with an adeno-associated virus encoding GGT5 enhanced hippocampal synaptic plasticity and mitigated cognitive deficits.Interestingly,increasing GGT5 expression in cerebrovascular endothelial cells reduced levels of both soluble and insoluble amyloid-βin the brains of APP/PS1 mice.This effect may be attributable to inhibition of the expression ofβ-site APP cleaving enzyme 1,which is mediated by nuclear factor-kappa B.Our findings demonstrate that GGT5 expression in cerebrovascular endothelial cells is inversely associated with Alzheimer’s disease pathogenesis,and that GGT5 upregulation mitigates cognitive deficits in APP/PS1 mice.These findings suggest that GGT5 expression in cerebrovascular endothelial cells is a potential therapeutic target and biomarker for Alzheimer’s disease.
基金a grant from Chang Gung Memorial Hospital,No.CMRPG33074a grant from National Science Council,Taiwan
文摘AIM:To investigate the role of nuclear factor of activated T cell 2(NFAT2),the major NFAT protein in peripheral T cells,in sustained T cell activation and intractable inflammation in human ulcerative colitis(UC). METHODS:We used two-dimensional gel-electrophoresis, immunohistochemistry,double immunohistochemical staining,and confocal microscopy to inspect the expression of NFAT2 in 107,15,48 and 5 cases of UC, Crohn's disease(CD),non-specific colitis,and 5 healthy individuals,respectively. RESULTS:Up-regulation with profound nucleo- translocation/activation of NFAT2 of lamina propria mononuclear cells(LPMC)of colonic mucosa was found specifically in the affected colonic mucosa from patients with UC,as compared to CD or NC(P<0.001,Kruskal- Wallis test).Nucleo-translocation/activation of NFAT2 primarily occurred in CD8+T,but was less prominent in CD4+T cells or CD20+B cells.It was strongly associated with the disease activity,including endoscopic stage (τ=0.2145,P=0.0281)and histologic grade(τ=0.4167, P<0.001). CONCLUSION:We disclose for the first time the nucleo-translocation/activatin of NFAT2 in lamina propria mononuclear cells in ulcerative colitis.Activation of NFAT2 was specific for ulcerative colitis and highly associated with disease activity.Since activation of NFAT2is implicated in an auto-regulatory positive feedback loop of sustained T-cell activation and NFAT proteins play key roles in the calcium/calcineurin signaling pathways,our results not only provide new insights into the mechanism for sustained intractable inflammation,but also suggest the calcium-calcineurin/NFAT pathway as a new therapeutic target for ulcerative colitis.
基金Supported by the National Key R and D Program of China,No.2019YFE0119300National Natural Science Foundation of China,No.82074158+2 种基金Project funded by China Postdoctoral Science Foundation,No.2018M631793Natural Science Foundation of Liaoning Province,No.2019-ZD-0624Dalian Traditional Chinese Medicine-Related Scientific Research Project,No.18Z2002.
文摘BACKGROUND Recent studies have demonstrated that dysfunction of the intestinal barrier is a significant contributing factor to the development of severe acute pancreatitis(SAP).A stable intestinal mucosa barrier functions as a major anatomic and functional barrier,owing to the balance between intestinal epithelial cell(IEC)proliferation and apoptosis.There is some evidence that calcium overload may trigger IEC apoptosis and that calcineurin(CaN)/nuclear factor of activated Tcells(NFAT)signaling might play an important role in calcium-mediated apoptosis.AIM To investigate the potential mechanisms underlying the therapeutic effect of Qingyi decoction(QYD)in SAP.METHODS A rat model of SAP was created via retrograde infusion of sodium deoxycholate.Serum levels of amylase,tumor necrosis factor(TNF-α),interleukin(IL)-6,D-lactic acid,and diamine oxidase(DAO);histological changes;and apoptosis of IECs were examined in rats with or without QYD treatment.The expression of the two subunits of CaN and NFAT in intestinal tissue was measured via quantitative realtime polymerase chain reaction and western blotting.For in vitro studies,Caco-2 cells were treated with lipopolysaccharide(LPS)and QYD serum,and then cell viability and intracellular calcium levels were detected.RESULTS Retrograde infusion of sodium deoxycholate increased the severity of pancreatic and intestinal pathology and the levels of serum amylase,TNF-α,and IL-6.Both the indicators of intestinal mucosa damage(D-lactic acid and DAO)and the levels of IEC apoptosis were elevated in the SAP group.QYD treatment reduced the serum levels of amylase,TNF-α,IL-6,D-lactic acid,and DAO and attenuated the histological findings.IEC apoptosis associated with SAP was ameliorated under QYD treatment.In addition,the protein expression levels of the two subunits of CaN were remarkably elevated in the SAP group,and the NFATc3 gene was significantly upregulated at both the transcript and protein levels in the SAP group compared with the control group.QYD significantly restrained CaN and NFATc3 gene expression in the intestine,which was upregulated in the SAP group.Furthermore,QYD serum significantly decreased the LPS-induced elevation in intracellular free Ca^(2+)levels and inhibited cell death.CONCLUSION QYD can exert protective effects against intestinal mucosa damage caused by SAP and the protective effects are mediated,at least partially,by restraining IEC apoptosis via the CaN/NFATc3 pathway.
文摘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.
文摘Objective： To investigate the role of peroxisome proliferator-activated receptors δ （PPARδ） in inflammatory reaction and its possible mechanism in adipocyte. Methods：Lentivirus-mediated RNA interference （RNAi） was used to block the expression of PPARδ in 3T3-L1 cells. In order to induce inflammation in 3T3-L1, cells were stimulated with tumor necrosis factor-α（TNFα, 20 ng/ml） for 4 h. The expression of PPARδ, nuclear factor κB （NFκB） and C reactive protein （CRP） were determined by Western blot analysis. Results：The expression of PPARδ was reduced by 80% after RNAi. Blockage of PPARδ promoted the expression of CRP and NFκB in cells stimulated with TNFα but had no effect on normal cells. Conclusion： PPARδ is involved in inflammatory reaction in adipocyte. Blockage of PPARδ can promote the inflammation mediated by inflammatory factors and increase the expression of NFκB and CRP in 3T3-L1 cells stimulated with TNFα.
文摘Obesity is a major human health problem associated with various diseases, including cardiac injury and type 2 diabetes. Trapa japonica Flerov (TJF) has been used in traditional oriental medicine to treat diabetes. In this study, we evaluated the inhibitory effect of and the mechanism underlying the effect of TJF extract on adipogenesis in 3T3-L1 cells. The effects of TJF extract on cell viability were analyzed using a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, and the anti-adipogenic effect was measured by oil red O staining. The expression of peroxisomal proliferator activated receptor (PPAR)γ, CCAAT/enhancer-binding protein-α (C/EBP)α, adenosine monophosphate-activated protein kinase (AMPK), acetyl-CoA carboxylase (ACC), adiponectin, and fatty acid binding protein (FABP)4 involved in adipogenesis was determined by western blot analysis. TJF extract effectively inhibited lipid accumulation and the expression of PPARγ and C/EBPα in 3T3-L1 cells. TJF also increased the phosphorylation of AMPK and ACC, and decreased the expression of adiponectin and FABP4. These results indicate that TJF extract exerts its anti-obesity effect through the downregulation of adipogenic transcription factors and adipogenic marker genes.
基金supported by grants from the National Natural Science Foundation of China (No. 30872926)the Program for AdvancedTalents within Six Industries of Jiangsu Province (08-D) to Dr. Luo Gu+1 种基金the Science Development Foundation of Nanjing Medical University (No. 2010NJMUZ35)the Research Program funded by Schoolof Basic Medical Science, Nanjing Medical University to Dr. Jun Du
文摘Epidermal growth factor （EGF） may increase cell motility, an event implicated in cancer cell invasion and metastasis. However, the underlying mechanisms for EGF-induced cell motility remain elusive. In this study, we found that EGF treatment could activate Ras-related C3 botulinum toxin substrate 1 （Racl）, PI3K/Akt and p21- actived kinase （PAK1） along with cell migration. Ectopic expression of PAK1 K299R, a dominant negative PAK1 mutant, could largely abolish EGF-induced cell migration. Blocking PI3K/Akt signalling with LY294002 or Akt siRNA remarkably inhibited both EGF-induced PAK1 activation and cell migration. Furthermore, expression of dominant-negative Racl （T17N） could largely block EGF-induced PI3K/Akt-PAK1 activation and cell migration. Interestingly, EGF could induce a significant production of ROS, and N-acetyl-L-cysteine, a scavenger of ROS which abolished the EGF-induced ROS generation, cell migration, as well as activation of PI3K/Akt and PAK, but not Racl. Our study demonstrated that EGF-induced cell migration involves a cascade of signalling events, including activation of Racl, generation of ROS and subsequent activation of PI3K/Akt and PAK1.
基金supported by the Foundation of Stomatology Hospital,Xi'an Jiaotong University
文摘Objective To study the effect of baicalin on the expression of receptor activator of nuclear factor-κB ligand(RANKL)and osteoprotegerin(OPG)in cultured human periodontal ligament(HPDL)cells.Methods Small interfering RNA(siRNA)eukaryotic expression vector targeted transforming growth factor βⅡ receptor(TGF-β RⅡ)was constructed and transfected into T cells.HPDL cells with T cells transfected with siRNA or not were placed in the culture medium that had been added with lipopolysaccharide(LPS)and baicalin.The obtained solution was divided into six groups according to the components(group Ⅰ:HPDL cells+LPS+T cells transfected with siRNA1+baicalin;group Ⅱ:HPDL cells+LPS+T cells transfected with siRNA1;group Ⅲ:HPDL cells+LPS+T cells+baicalin;group Ⅳ:HPDL cells+LPS+T cells;group Ⅴ:HPDL cells+baicalin;group Ⅵ:HPDL cells)and was cultured for 48 hours.RT-PCR was used to observe the effect of baicalin on the expression of OPG-RANKL in HPDL cells.Results The ratio of RANKL/OPG in group Ⅰ was lower than that in group Ⅱ(P<0.01)and higher than that in group Ⅲ(P<0.01);The ratio of RANKL/OPG in group Ⅲ was lower than that in group Ⅳ(P<0.01);the ratio of RANKL/OPG in group Ⅳ was higher than that in group Ⅵ(P<0.01);the ratio of RANKL/OPG in group Ⅴ was lower than that in group Ⅵ(P<0.05).Conclusion ① Baicalin could decrease the ratio of RANKL/OPG in HPDL cells.② The TGF-β signaling transduction plays an important role in the effect of baicalin on the RANKL/OPG ratio in HPDL cells.③ Baicalin acts not only through TGF-β to regulate RANKL/OPG in HPDL cells,but also through other pathways.
基金Shenzhen Science and Technology Plan(No.JCYJ20180306172419505).
文摘Objective: To investigate the expression relationship between nuclear transcription factor kappa B1 (NFκB1) and long non-coding RNA PACER (LncRNA-PACER) in peripheral blood mononuclear cells (PBMCs) of patients with pulmonary tuberculosis. Methods: From February 2018 to March 2019, 40 patients with pulmonary tuberculosis (tuberculosis group) and 40 healthy persons (control group) were collected, the levels of TNF-α, IL-6 and IL-8 in serum were detected by enzyme-linked immunosorbent assay (ELISA);the expressions of LncRNA-PACER and NFκB1 mRNAs in PBMCs were detected by real-time fluorescence quantitative PCR;Western blot was used to detect the expressions of NFκB1 and COX 2 in PBMCs;Pearson method was used to analyze the expressions of LncRNA-PACER and NFκB1 in PBMCs of patients with pulmonary tuberculosis, and the expressions of LncRNA-PACER and NFκB1 in PBMCs of patients with pulmonary tuberculosis were analyzed. Results: Compared with the control group, the expressions of TNF-α, IL-6 and IL-8 in the serum of patients with pulmonary tuberculosis was significantly increased (P<0.05), and the expressions of LncRNA-PACER, NFκB1 mRNAs, proteins and COX-2 protein in PBMCs were significantly increased (P<0.05). The expressions of LncRNA-PACER and NFκB1 proteins in PBMCs were related to the number of pulmonary lesions and pulmonary cavity (P<0.05), and there was a positive correlation between the expression of LncRNA-PACER and the expression of NFκB1 mRNA in PBMCs of patients with pulmonary tuberculosis (r = 0.873, P<0.05). Conclusions: The expressions of NFκB1 and LncRNA-PACER in PBMCs of patients with pulmonary tuberculosis are significantly increased, they are positively correlated and both of them are related to the occurrence and development of pulmonary tuberculosis.
