This study aimed to evaluate the effects of dietary yeast culture(YC)on lipopolysaccharide(LPS)-induced oxidative stress,immune and inflammatory response in P.ussuriensis.The fish were randomly assigned into three gro...This study aimed to evaluate the effects of dietary yeast culture(YC)on lipopolysaccharide(LPS)-induced oxidative stress,immune and inflammatory response in P.ussuriensis.The fish were randomly assigned into three groups as the control group,LPS group and YC+LPS group.The fish in the control were fed diet with no YC supplementation and no LPS challenge,and the fish in the LPS group or YC+LPS group were fed diet supplemented with no YC or 20 g·kg^(-1)YC,and with LPS challenge,respectively.The results showed that compared with the control group,intestinal total antioxidant capacity(T-AOC)level and superoxide dismutase(SOD)activity were significantly decreased,while intestinal malondialdehyde(MDA),plasma aspartate transaminase(AST)and alanine transaminase(ALT)levels were significantly increased in the LPS group(P<0.05).Besides,lower plasma alkaline phosphatase(ALP),alternative complement pathway(ACH50)activity and the albumin(ALB)level,as well as higher lysozyme(LZM)activity,were also found in the LPS group.However,dietary 20 g·kg^(-1)YC supplementation could relieve the above LPS-induced changes in Pseudobagrus ussuriensis.Furtherly,LPS challenge could significantly up-regulate gene expression of interleukin-8(IL-8),heat shock protein(HSP70)and NF-κBp65 except for toll-like receptors 2(TLR2),while dietary 20 g·kg^(-1)YC supplementation suppressed the increased expression of NF-κBp65 and IL-8 induced by LPS in P.ussuriensis.In summary,LPS challenge could induce immune impairment,oxidative stress and hepatic damage,and the protective effect of dietary 20 g·kg^(-1)YC supplementation on LPS-induced immune impairment and oxidative stress was observed in the present study,which was associated with the enhanced levels of antioxidant enzymes and immune parameters.Also,dietary 20 g·kg^(-1)YC supplementation could suppress LPS-induced inflammatory response by down-regulating NF-κBp65,HSP70 and IL-8 gene expression.展开更多
Endotoxins(also known as lipopolysaccharides(LPS)) are undesirable by-products of recombinant proteins,purified from Escherichia coli.LPS can be considered stable under a wide range of temperature and pH,making their ...Endotoxins(also known as lipopolysaccharides(LPS)) are undesirable by-products of recombinant proteins,purified from Escherichia coli.LPS can be considered stable under a wide range of temperature and pH,making their removal one of the most difficult tasks in downstream processes during protein purification.The inherent toxicity of LPS makes their removal an important step for the application of these proteins in several biological assays and for a safe parenteral administration.Immobilized metal affinity chromatography(IMAC) enables the affinity interactions between the metal ions(immobilized on the support through the chelating compound) and the target molecules,thus enabling high-efficiency separation of the target molecules from other components present in a mixture.Affinity chromatography is applied with Ca2+-iminodiacetic acid(IDA) to remove most of the LPS contaminants from the end product(more than90%).In this study,the adsorption of LPS on an IDA-Ca2+ was investigated.The adsorption Freundlich isotherm of LPS-IDA-Ca2+ provides a theoretical basis for LPS removal.It was found that LPS is bound mainly by interactions between the phosphate group in LPS and Ca2+ ligands on the beads.The factors such as pH(4.0 or 5.5) and ionic strength(1.0 mol/L) are essential to obtain effective removal of LPS for contaminant levels between endotoxin' concentration values less than100 EU/mL and 100 000 EU/mL.This new protocol represents a substantial advantage in time,effort,and production costs.展开更多
OBJECTIVE To establish an in vitro inflammatory model of BV2 by observing the activity,the release amount of NO and the expression of inflammatory factors of microglial cells(BV2)induced by lipopolysaccharides(LPS).ME...OBJECTIVE To establish an in vitro inflammatory model of BV2 by observing the activity,the release amount of NO and the expression of inflammatory factors of microglial cells(BV2)induced by lipopolysaccharides(LPS).METHODS BV2 was routinely cultured in vitro.Cell viability was measured by CCK-8 meth⁃od.And by drew cell growth curve to determine the logarithmic growth cycle of the cells.After 24 h of routine culture,BV2 were induced by adding different concentrations of LPS(0.1,1.0 and 10.0 mg·L-1)for 4,8,12,24 and 48 h,respectively.Meanwhile,the morphological changes of BV2 were observed under inverted microscope to compare the activation degree of microglia at dif⁃ferent time and concentration.Cell activity and nitric oxide(NO)level were determined by CCK-8 and Griess method respectively,which could help to determine the optimal concentration and time of modeling.Finally,It were determined by ELISA that the concentrations of tumor necrosis factorα(TNF-α),interleukin-6(IL-6)and IL-1βin supernatant of LPS 1 mg·L-1 culture for 24 h.RESULTS BV2 were in logarithmic growth phase for 1 to 3 d after subculture.LPS 1 mg·L-1 induced BV2 for 24 or 48 h which could increase the release amount of NO significantly(P<0.05).In order to save time,LPS induced BV2 for 24 h were selected for subsequent experiments.Microglial cells in resting state were observed to be elongated spindle shape under inverted micro⁃scope.After LPS activation,the cell body became larger and the branching processes shrank back,presenting an amoeba-like appearance.ELISA results showed that the concentrations of TNF-α,IL-6 and IL-1βin supernatant of LPS 1 mg·L-1 cultured for 24 h were significantly increased which compared with the control group(P<0.05).CONCLUSION LPS could induce the activation of BV2 and up-regulate the level of inflammatory factors.The optimal condition for establishing stable BV2 microglial inflammatory model was used LPS 1 g·L-1 induced for 24 h.展开更多
Dendritic cells （DCs） are bone marrow-derived professional antigen presenting cells （APCs）, they are crucial for initiation of both innate and adaptive immune responses. In this study, chicken bone marrow （chBM）...Dendritic cells （DCs） are bone marrow-derived professional antigen presenting cells （APCs）, they are crucial for initiation of both innate and adaptive immune responses. In this study, chicken bone marrow （chBM） cells were cultured in medium with recombinant chicken granulocyte-macrophage colony stimulating factor （rGM-CSF） and recombinant chicken interleukin-4 （rIL-4） for 7 days, displayed the typical morphology of DCs. These immature chicken bone marrow-derived DCs （chBM-DCs） showed signifcant up-regulation of the putative CD11c and of major histocompatibility complex class II （MHC II）, but CD40 and CD86 co-stimulatory molecules were almost no up-regulated. However, maturation with lipopolysaccharide （LPS）, surface expression of CD40, CD86 was greatly increased. The phagocytosis of chBM-DCs was assessed by neutral red, and the phagocytosis decreased after stimulation. In mixed lymphocyte responses （MLR）, stimulated chBM-DCs were more effective to T-cell stimulators than non-stimulated chBM-DCs. In addition, mRNA expression levels of IL-1β, IL-4, IL-6, IL-10, IL-12, IFN-γ, TNF-α, CXCLi1 and CXCLi2 were assessed by real-time qPCR （qRT-PCR）, and the results showed cultured chBM-DCs could be matured to a T helper cell type 1 （Th1）-promoting phenotype by LPS stimulation.展开更多
OBJECTIVE To explore the synergistic effect of baicalin and geniposide(BG)on BV2 cell activation damage caused by lipopolysaccharide(LPS).METHODS BV2 murine microglial cell line was cultured in vitro,LPS(final concent...OBJECTIVE To explore the synergistic effect of baicalin and geniposide(BG)on BV2 cell activation damage caused by lipopolysaccharide(LPS).METHODS BV2 murine microglial cell line was cultured in vitro,LPS(final concentration 500 ng·m L-1)and various concentrationof Baicalin and Geniposide(BG)(final concentration12.5,25 and 50μg·m L-1)were added tointerven,the negative control was establised.MTT method was used to value the effect of LPS on the viability of BV2 cell line.The accumulated nitrite was assayed utilizing the Griess reaction method.RESULTS(1)Morphological observation:The common marphological of quesient microglia is circle,cell bodies smaller and synaptic slender.The enlargement of microglial cell bodies and an amoeboid morphology with retraction of extensions are generally induced by LPS.BG markedly suppressed the LPS-activated BV2 microglia morphological variations,meanwhile the dose-dependent was dramaticaly performed.(2)MTT test showed that LPS-stimulated BV2 cells viability was significantly decreased compared to the control group;compared to LPS treated cells,drug group(LPS+BG)effectively improves the LPS-stimulated BV2 cells viability.(3)The Griess reaction method indicated that LPS could obviously promoted the BV2 cells′NO generation contrasted to control group;while the drug group(LPS+BG)can effectively inhibited the generation of NO which activated by LPS.CONCLUSION The treatment group could significantly enhance survival rate of LPSstimulated BV2 cells,while,the level of NO was markedly decreased in BV2 induced by LPS.These findings suggest that combination of BG could attenuate BV2 microglial cells activation and injury which induced by LPS,possessed the capacity of neuroprotective.展开更多
Aim Inducible nitric oxide synthase （iNOS） makes a great contribution to host defense and inflamma-tion. In many settings, lipopolysaccharide （LPS） induces iNOS expression through activation of the inhibitor of K...Aim Inducible nitric oxide synthase （iNOS） makes a great contribution to host defense and inflamma-tion. In many settings, lipopolysaccharide （LPS） induces iNOS expression through activation of the inhibitor of KB- α （IKB-α） -nuclear factor-KB （NF-KB） cascade, whereas interferon-γ （IFN-γ） acts through Janus kinase （ JAK）- signal transducer and activator of transcription 1 （ STAT1 ） signals. Heat shock factor 1 （ HSF1 ）, a major regulator of heat shock protein transcription, has been shown to regulate the production of pro-inflammatory cytokines such as tumor necrosis factor-α（TNF-α） and interleukin-6 （IL-6）. But it remains obscure whether and how HSF1 affects iNOS induction. Methods Western blot was used to measure the protein expression. The mRNA level was meas- ured by real time-PCR. Silence of HSF1 was achieved by small interfering RNA. Nitric oxide （NO） content and NF-KB binding activity were assayed by commercial kits. Chromatin immunoprecipitation （CHIP） was used to measure the binding activity of NF-KB and STAT1 to iNOS promoters. Results HSF1 inhibition or knockdown pre- vented the LPS- and/or IFN-γ-stimulated iNOS protein expression in cultured microglia. HSF1 inhibition blocked iNOS mRNA transcription. These inhibitory effects of HSF1 inhibition on iNOS expression were confirmed in brain tissues from endotoxemic mice. Further analysis showed that HSF1 inhibition had no effect on IKB-α degradation and NF-KB or STAT1 phosphorylation in LPS/IFN-γ-stimulated cells. The nuclear transport of active NF-KB or STAT1 was also not affected by HSF1 inhibition. But HSF1 inhibition reduced the binding of NF-KB and STAT1 to their DNA elements. In addition, HSF1 inhibition reduced NF-KB and STAT1 bindings to iNOS promoter inside the LPS/IFN-γ-stimulated cells. Conclusions This preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.展开更多
Aim Magnesium lithospermate B （MLB） is the most abundant hydrophilic active component of Salvia rniltiorrhiza Radix, a traditional Chinese herbal medicine mainly used to treat cardiovascular diseases. Studies have s...Aim Magnesium lithospermate B （MLB） is the most abundant hydrophilic active component of Salvia rniltiorrhiza Radix, a traditional Chinese herbal medicine mainly used to treat cardiovascular diseases. Studies have shown that endothelial activation contributes to the pathophysiology of cardiovascular diseases such as atherosclero- sis, diabetic vasculopathy, heart failure and hypertension. In the present study, the effects of MLB on endothelial activation were investigated. Lipopolysaccharide （LPS） 1 mg L^-1 was employed to induce endothelial activation, which was determined by relative gene expression and endothelial adhesion assay. Results showed that pretreatment with MLB attenuated LPS-induced ICAM1, VCAM1 and TNF-α upregulation in human dermal microvascular endo- thelial cells （HMEC-1） in dose-dependent manner, which contributed to the reduction of THP-1 adhesion to HMEC-1. Furthermore, it was revealed that 100 μmol · L^-1 MLB significantly decreased the nuclear translocation of NF-KB p65, a critical transcription factor in LPS-indueed inflammatory response, through the inhibition of IKBμ degradation. Besides, the transcriptional activity of NF-KB p65 was also inhibited by the pretreatment of MLB. Mo- reover, MLB pretreatment considerably inhibited LPS-induced p38 phosphorylation, which at least partly contribu- ted to the reduction of ICAM1 expression. In conclusion, these findings suggest that MLB inhibits LPS-induced nu- clear translocation and transcripitional activity of NF-KB, thus attenuates the increased expression of adhesion mole- cules and inflammatory factors, protects endothelial cells from LPS-induced activation.展开更多
OBJECTIVE The emerging role of chronic inflammation is the major degenerative diseases of modern society such as periodontitis,atherosclerosis,rheumatoid arthritis,Parkinson′s disease and even cancer.Eight components...OBJECTIVE The emerging role of chronic inflammation is the major degenerative diseases of modern society such as periodontitis,atherosclerosis,rheumatoid arthritis,Parkinson′s disease and even cancer.Eight components were isolated from Derris laxiflora Benth.,In this study,we found these compounds from Derris laxiflora Benth suppress lipopolysaccharide-induced inflammatory response in murine macrophage(RAW 264.7)cells.METHODS RAW 264.7cells were cultured in DMEM media supplemented with 10%(V/V)heated-inactivated FBS,penicillin 100U·mL-1 and streptomycin 100μg·mL-1.The cells were incubated at 37℃in a humidified atmosphere of 5%CO2in air.RAW264.7cells were seeded in a 24-well plate at a density of 2×105 mL-1 and then incubated with or without LPS(100ng·mL-1)in the absence or presence of compounds for 24 h.Effects of these isolates on NO production were measured indirectly by analysis of nitrite levels using the Griess reaction.Quercetin was used as a positive control.RESULTS ight components were isolated from Derris laxiflora Benth.,including three new pterocarpans 7,6′-dihydroxy-3′-methoxypterocarpan(1),derrispisatin(2),derriscoumaronochromone(3)and three new flavonoids cis-3,4′-dihydroxy-5,7-dimethoxyflavan(4),derriflavanone B(5),iso-lupinenol(6)as well as two known ones,lonchocarpol A(7)and lonchocarpol D(8).The structures of these new compounds were determined by analysis of their spectroscopic data.Raw264.7 cells were treated with the compounds from Derris laxiflora Benth for 24 h.Among them,compounds 5,7 and 8 significantly suppressed the NO production in LPS-treated RAW264.7 cells with IC50 values<10μg·mL-1.CONCLUSION In this study,we found that compounds from Derris laxiflora Benth suppresses lipopolysaccharide-induced inflammatory response in murine Raw264.7 cells.展开更多
OBJECTIVE To investigate the effects of ex-tract of Ramulus Cinnamom(RC)against LPS-induced inflammation in microglia.METHODS Activated microglia releases various pro-inflammatory cytokines to induce neuroinflammation...OBJECTIVE To investigate the effects of ex-tract of Ramulus Cinnamom(RC)against LPS-induced inflammation in microglia.METHODS Activated microglia releases various pro-inflammatory cytokines to induce neuroinflammation in stroke.Lipopolysaccaride(LPS)is an endotoxin from the outer membrane of Gram-negative bacteria that activates microglia.MTT assay was used to observe the cell viability.The content of NO in supernatant was measured by Griess reagent.The levels of IL-1β,IL-6 and TNF-αin supernatant were detected by ELISA kits.The intracellular COX-2,TLR4,and My D88expression was assayed by Western blotting.RESULTS RC extract 30 and 100μg·m L-1significantly decreased the production of related inflammatory factors such as NO(P<0.05,P<0.01),IL-1β(P<0.01,P<0.01),IL-6(P<0.05,P<0.01)and TNF-α(P<0.01,P<0.01).Furthermore,RC extract significantly inhibited the COX-2,TLR4,and My D88 expression induced by LPS in BV2cells.CONCLUSION RC extract may have therapeutic potential for the improvement of neuroinflammation,and the mechanism may be involved in down-regulation of TLR4/My D88 inflammation pathway.展开更多
OBJECTIVE Xiao-xu-ming decoction(XXMD),a well-known traditional Chinese herbal prescription,has been widely used to treat stroke.It is recorded in″Bei Ji Qian Jin Yao Fang″written by Si-miao Sun of the Chinese ancie...OBJECTIVE Xiao-xu-ming decoction(XXMD),a well-known traditional Chinese herbal prescription,has been widely used to treat stroke.It is recorded in″Bei Ji Qian Jin Yao Fang″written by Si-miao Sun of the Chinese ancient Tang Dynasty.In our previous study,the active fraction of XXMD(XXM)against cerebral ischemia has been prepared by modern separation and purification techniques.This study was to investigate XXM against lipopolysaccaride(LPS)-induced neuroinflammation in mice.METHODS LPS is an endotoxin from the outer membrane of Gram-negative bacteria that activates inflammation.XXM was pre-treated in BALB/C mice followed by injected intraperitoneally with LPS(5 mg·kg-1).The effects of XXM on LPS-induced pro-inflammatory factors and proteins were measured by ELISA,Western blot,and immunofluorescence in vivo.RESULTS Mice treated with XXM showed significantly decreased proinflammatory factors level,including IL-1β(P<0.01),IL-6(P<0.01),TNF-α(P<0.05),and MCP-1(P<0.01).Furthermore,XXM also significantly inhibited the inflammatory pathway proteins expression induced by LPS,including TLR4,MyD 88,and COX-2.CONCLUSION XXM possesses anti-neuroinflammation in mice and might be a promising therapeutic agent for stroke.展开更多
OBJECTIVE To evaluate the neuroprotective effects of 4 components from Uncaria rhynchophylla(297,307,315 and 327)on long-term potentiation(LTP)deficit in neuroinflammation animal model induced by lipopolysaccha⁃ride(L...OBJECTIVE To evaluate the neuroprotective effects of 4 components from Uncaria rhynchophylla(297,307,315 and 327)on long-term potentiation(LTP)deficit in neuroinflammation animal model induced by lipopolysaccha⁃ride(LPS).METHODS Male BALB/c 18-22 g mice were divided into control group,model group and component treat⁃ment group(1μg per mouse,icv);each group contained 5 mice,model group and compound treatment group were intra⁃peritoneally injected LPS(50μg·kg^-1,ip)4 h before LTP induction.LTP of perforant path-dentate gyrus pathway in hippo⁃campus was induced by high frequency stimulation and used to evaluate the effects on synaptic plasticity.RESULTS Compared with control group,the LTP of the model group was significantly impaired.Compound 297,327,and 307 could significantly improve LPS-induced LTP impairment.315 had no significant effect on LPS-induced LTP impairment.CONCLUSION Hippocampal synaptic plasticity could be impaired by LPS.Compounds 297,327 and 307 have protec⁃tive effects against LPS induced LTP impairment,and 315 has little effect on LPS induced LTP impairment.These re⁃sults suggested that 297,327 and 307 might have potential effects on neuroinflammation induced memory deficits.展开更多
Sepsis can cause a series of damages to various organs of the body,so it has always been regarded as a hot research topic in veterinary clinic.Aiming at the present situation of high morbidity and mortality of canine ...Sepsis can cause a series of damages to various organs of the body,so it has always been regarded as a hot research topic in veterinary clinic.Aiming at the present situation of high morbidity and mortality of canine sepsis,in order to further explore the pathogenesis of this disease,it need to establish a stable and repeatable canine sepsis model that is in line with the clinical characteristics of the disease.The study selected 12 local dogs and randomly divided into three groups:rapid bolus injection group(Group A),continuous infusion group within 30 min(Group B)and continuous infusion group(Group C).Then,the lipopolysaccharides(LPS)with 2 mg·kg^-1 were injected through the brachial vein in different modes of administration,thus the model was fully established.During the modeling period,body temperature(T),respiratory rate(RR),heart rate(HR)and mean arterial pressure(MAP)were monitored at 0,10,20,30,40,50 min and 1,2,3,4,5,6,7,8,9,12 and 24 h.Blood was collected from the canine brachial vein at 0,1,2,3,4,5,6,7,8,9,12 and 24 h,respectively,for the detection of white blood cells(WBC).The test showed that the values of T,RR,HR,MAP,WBC of the dogs in group A all changed but did not exceed the normal range,and the clinical symptoms were not significant.There were no significant changes in the values of RR,HR,T,MAP and WBC of the dogs in Group B,and the clinical symptoms were not significant.The value of T of the dogs in Group C were significantly increased at 40 min(p<0.05),which reached the fever standard and lasted for 7 h;the value of RR increased significantly at 20 min(p<0.05),and a downward trend could be observed at 12 h,then it returned to normal at 24 h;the value of HR increased significantly at 50 min(p<0.05)and recovered at 8 h;the value of HR decreased significantly at 20 min(p<0.05),which remained at 12 h(p<0.05),and returned back to normal at 24 h;the value of WBC decreased significantly from 1 h to 4 h(p<0.05),which was lower than the normal value,and increased significantly at 24 h(p<0.01);all of the four dogs in this group had clinical symptoms such as vomiting,diarrhea and depression.Based on the above results,the changes of indexes and clinical symptoms in Groups A and B did not meet the standards of sepsis.After a long-term continuous intravenous infusion of LPS,the experimental dogs in Group C showed varying degrees of clinical symptoms,such as vomiting,diarrhea and depression one after another.The indexes and clinical symptoms reached the sepsis standard about 3 h after infusion.In brief,this model not only had good stability and good regularity of repeatability,but also lasted for a long time and could be suitable for other subsequent studies.展开更多
OBJECTIVE Isofuranodiene(ISO)is a natural product isolated in Chinese herb.Our recent results showed anticancer effect in vitro.In this study,we investigated its live protective effect with a Dgalactosamine/lipopolysa...OBJECTIVE Isofuranodiene(ISO)is a natural product isolated in Chinese herb.Our recent results showed anticancer effect in vitro.In this study,we investigated its live protective effect with a Dgalactosamine/lipopolysacchride(GalN/LPS)induced rat model.METHODS SD rats were treated orally with or without ISO(20 and 50mg·kg-1,ig)for 3d and then treated with GalN/LPS for 8h.The serum were collected and the concentration of aspartate aminotransferase(AST),alanine aminotransferase(ALT),and malondialdehyde(MDA)were determined.The liver injury was examined by H&E staining.The mRNA expression of IL-1β,IL-6 and inducible nitric oxide synthase(iNOS)in liver tissues were determined by realtime PCR.RESULTS Oral administration of ISO(20 and 50mg·kg-1)dramatically inhibited GalN/LPS-induced serum elevation of AST,ALT,and MDA levels.The liver injury was also significantly ameliorated as evidenced by the histological improvement in H&E staining.Furthermore,ISO treatment significantly inhibited GalN/LPS-induced mRNA expression of IL-1β,IL-6,and inducible nitric oxide synthase(iNOS)in liver tissues.CONCLUSION This data showed that ISO has hepatoprotective effect in rats.展开更多
The model of acute lung injury(ALI)was established by intraperitoneal administration,but there was no time-point observation and comparison.ALI model was established by intraperitoneal injection of lipopolysaccharide(...The model of acute lung injury(ALI)was established by intraperitoneal administration,but there was no time-point observation and comparison.ALI model was established by intraperitoneal injection of lipopolysaccharide(LPS)at the concentration of 10 mg·kg^-1 (10 mg LPS dissolved in 1 mL normal saline to prepare 1 mL·kg^-1solution)in rats.The control group(CG)was intraperitoneally injected with saline of the same dose.In the LPS group,lung tissues were collected at 4,6,8,12 and 24 h after administration.Then,the morphology changes,the ratio of wet-to-dry weight(W/D),the expression of interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)proteins,the levels of malondialdehyde(MDA),the activities of superoxide dismutase(SOD),glutathione peroxidase(GSH)were measured.To verify the success of the model,the degrees of lung injury via Western blot,RT-PCR,ELISA and other techniques were detected at different time points,and the severe time of the ALI model established was deterimined by intraperitoneal administration,which provided a stable model basis for the study of the pathogenesis of ALI in the future.The results showed that the lung injury occurred in LPS group.W/D and lung pathological changes at 12 and 24 h of LPS group were significantly different from those in the CG.Compared with the CG,the expression of IL-1βand TNF-αproteins and the content of MDA in lung tissues of LPS group increased and most significant difference was found at 12 and 24 h(p<0.01).Compared with the CG,the activities of SOD and GSH in LPS 12 h group decreased significantly(p<0.01).In conclusion,inflammation and oxidative damage were the main causes of the ALI in rats.Lung injury was most obvious 12 h after intraperitoneal injection of 10 mg·kg^-1 LPS.展开更多
基金Supported by the Special Fund for Beijing Enhalor Bio-Tech Co.,Ltd.the Open Fund for National Engineering Research Laboratory of Marine Biotechnology and Engineering,Ningbo Universitythe Entrepreneurship Trainning Project of SIPT Program of Northeast Agricultural University(202310224129S)。
文摘This study aimed to evaluate the effects of dietary yeast culture(YC)on lipopolysaccharide(LPS)-induced oxidative stress,immune and inflammatory response in P.ussuriensis.The fish were randomly assigned into three groups as the control group,LPS group and YC+LPS group.The fish in the control were fed diet with no YC supplementation and no LPS challenge,and the fish in the LPS group or YC+LPS group were fed diet supplemented with no YC or 20 g·kg^(-1)YC,and with LPS challenge,respectively.The results showed that compared with the control group,intestinal total antioxidant capacity(T-AOC)level and superoxide dismutase(SOD)activity were significantly decreased,while intestinal malondialdehyde(MDA),plasma aspartate transaminase(AST)and alanine transaminase(ALT)levels were significantly increased in the LPS group(P<0.05).Besides,lower plasma alkaline phosphatase(ALP),alternative complement pathway(ACH50)activity and the albumin(ALB)level,as well as higher lysozyme(LZM)activity,were also found in the LPS group.However,dietary 20 g·kg^(-1)YC supplementation could relieve the above LPS-induced changes in Pseudobagrus ussuriensis.Furtherly,LPS challenge could significantly up-regulate gene expression of interleukin-8(IL-8),heat shock protein(HSP70)and NF-κBp65 except for toll-like receptors 2(TLR2),while dietary 20 g·kg^(-1)YC supplementation suppressed the increased expression of NF-κBp65 and IL-8 induced by LPS in P.ussuriensis.In summary,LPS challenge could induce immune impairment,oxidative stress and hepatic damage,and the protective effect of dietary 20 g·kg^(-1)YC supplementation on LPS-induced immune impairment and oxidative stress was observed in the present study,which was associated with the enhanced levels of antioxidant enzymes and immune parameters.Also,dietary 20 g·kg^(-1)YC supplementation could suppress LPS-induced inflammatory response by down-regulating NF-κBp65,HSP70 and IL-8 gene expression.
基金supported by grants from the Brazilian Agency Coordination of Graduate Level Training(CAPES,project 0366/09-9)State of So Paulo Research Support Foundation(FAPESP-Brazil,project 2005/60159-7)
文摘Endotoxins(also known as lipopolysaccharides(LPS)) are undesirable by-products of recombinant proteins,purified from Escherichia coli.LPS can be considered stable under a wide range of temperature and pH,making their removal one of the most difficult tasks in downstream processes during protein purification.The inherent toxicity of LPS makes their removal an important step for the application of these proteins in several biological assays and for a safe parenteral administration.Immobilized metal affinity chromatography(IMAC) enables the affinity interactions between the metal ions(immobilized on the support through the chelating compound) and the target molecules,thus enabling high-efficiency separation of the target molecules from other components present in a mixture.Affinity chromatography is applied with Ca2+-iminodiacetic acid(IDA) to remove most of the LPS contaminants from the end product(more than90%).In this study,the adsorption of LPS on an IDA-Ca2+ was investigated.The adsorption Freundlich isotherm of LPS-IDA-Ca2+ provides a theoretical basis for LPS removal.It was found that LPS is bound mainly by interactions between the phosphate group in LPS and Ca2+ ligands on the beads.The factors such as pH(4.0 or 5.5) and ionic strength(1.0 mol/L) are essential to obtain effective removal of LPS for contaminant levels between endotoxin' concentration values less than100 EU/mL and 100 000 EU/mL.This new protocol represents a substantial advantage in time,effort,and production costs.
基金Natural science foundation of Hebei Province(H2020405298)。
文摘OBJECTIVE To establish an in vitro inflammatory model of BV2 by observing the activity,the release amount of NO and the expression of inflammatory factors of microglial cells(BV2)induced by lipopolysaccharides(LPS).METHODS BV2 was routinely cultured in vitro.Cell viability was measured by CCK-8 meth⁃od.And by drew cell growth curve to determine the logarithmic growth cycle of the cells.After 24 h of routine culture,BV2 were induced by adding different concentrations of LPS(0.1,1.0 and 10.0 mg·L-1)for 4,8,12,24 and 48 h,respectively.Meanwhile,the morphological changes of BV2 were observed under inverted microscope to compare the activation degree of microglia at dif⁃ferent time and concentration.Cell activity and nitric oxide(NO)level were determined by CCK-8 and Griess method respectively,which could help to determine the optimal concentration and time of modeling.Finally,It were determined by ELISA that the concentrations of tumor necrosis factorα(TNF-α),interleukin-6(IL-6)and IL-1βin supernatant of LPS 1 mg·L-1 culture for 24 h.RESULTS BV2 were in logarithmic growth phase for 1 to 3 d after subculture.LPS 1 mg·L-1 induced BV2 for 24 or 48 h which could increase the release amount of NO significantly(P<0.05).In order to save time,LPS induced BV2 for 24 h were selected for subsequent experiments.Microglial cells in resting state were observed to be elongated spindle shape under inverted micro⁃scope.After LPS activation,the cell body became larger and the branching processes shrank back,presenting an amoeba-like appearance.ELISA results showed that the concentrations of TNF-α,IL-6 and IL-1βin supernatant of LPS 1 mg·L-1 cultured for 24 h were significantly increased which compared with the control group(P<0.05).CONCLUSION LPS could induce the activation of BV2 and up-regulate the level of inflammatory factors.The optimal condition for establishing stable BV2 microglial inflammatory model was used LPS 1 g·L-1 induced for 24 h.
基金Supported by the National Technology and Research Project(2015BAD12B01-4)
文摘Dendritic cells （DCs） are bone marrow-derived professional antigen presenting cells （APCs）, they are crucial for initiation of both innate and adaptive immune responses. In this study, chicken bone marrow （chBM） cells were cultured in medium with recombinant chicken granulocyte-macrophage colony stimulating factor （rGM-CSF） and recombinant chicken interleukin-4 （rIL-4） for 7 days, displayed the typical morphology of DCs. These immature chicken bone marrow-derived DCs （chBM-DCs） showed signifcant up-regulation of the putative CD11c and of major histocompatibility complex class II （MHC II）, but CD40 and CD86 co-stimulatory molecules were almost no up-regulated. However, maturation with lipopolysaccharide （LPS）, surface expression of CD40, CD86 was greatly increased. The phagocytosis of chBM-DCs was assessed by neutral red, and the phagocytosis decreased after stimulation. In mixed lymphocyte responses （MLR）, stimulated chBM-DCs were more effective to T-cell stimulators than non-stimulated chBM-DCs. In addition, mRNA expression levels of IL-1β, IL-4, IL-6, IL-10, IL-12, IFN-γ, TNF-α, CXCLi1 and CXCLi2 were assessed by real-time qPCR （qRT-PCR）, and the results showed cultured chBM-DCs could be matured to a T helper cell type 1 （Th1）-promoting phenotype by LPS stimulation.
基金The project suppored by National Natural Science Foundation of China(81473385)Shaanxi Province Education Department Project(13JS029)by Shaanxi Province Administration of Traditional Chinese Medicine(13-ZY016)
文摘OBJECTIVE To explore the synergistic effect of baicalin and geniposide(BG)on BV2 cell activation damage caused by lipopolysaccharide(LPS).METHODS BV2 murine microglial cell line was cultured in vitro,LPS(final concentration 500 ng·m L-1)and various concentrationof Baicalin and Geniposide(BG)(final concentration12.5,25 and 50μg·m L-1)were added tointerven,the negative control was establised.MTT method was used to value the effect of LPS on the viability of BV2 cell line.The accumulated nitrite was assayed utilizing the Griess reaction method.RESULTS(1)Morphological observation:The common marphological of quesient microglia is circle,cell bodies smaller and synaptic slender.The enlargement of microglial cell bodies and an amoeboid morphology with retraction of extensions are generally induced by LPS.BG markedly suppressed the LPS-activated BV2 microglia morphological variations,meanwhile the dose-dependent was dramaticaly performed.(2)MTT test showed that LPS-stimulated BV2 cells viability was significantly decreased compared to the control group;compared to LPS treated cells,drug group(LPS+BG)effectively improves the LPS-stimulated BV2 cells viability.(3)The Griess reaction method indicated that LPS could obviously promoted the BV2 cells′NO generation contrasted to control group;while the drug group(LPS+BG)can effectively inhibited the generation of NO which activated by LPS.CONCLUSION The treatment group could significantly enhance survival rate of LPSstimulated BV2 cells,while,the level of NO was markedly decreased in BV2 induced by LPS.These findings suggest that combination of BG could attenuate BV2 microglial cells activation and injury which induced by LPS,possessed the capacity of neuroprotective.
文摘Aim Inducible nitric oxide synthase （iNOS） makes a great contribution to host defense and inflamma-tion. In many settings, lipopolysaccharide （LPS） induces iNOS expression through activation of the inhibitor of KB- α （IKB-α） -nuclear factor-KB （NF-KB） cascade, whereas interferon-γ （IFN-γ） acts through Janus kinase （ JAK）- signal transducer and activator of transcription 1 （ STAT1 ） signals. Heat shock factor 1 （ HSF1 ）, a major regulator of heat shock protein transcription, has been shown to regulate the production of pro-inflammatory cytokines such as tumor necrosis factor-α（TNF-α） and interleukin-6 （IL-6）. But it remains obscure whether and how HSF1 affects iNOS induction. Methods Western blot was used to measure the protein expression. The mRNA level was meas- ured by real time-PCR. Silence of HSF1 was achieved by small interfering RNA. Nitric oxide （NO） content and NF-KB binding activity were assayed by commercial kits. Chromatin immunoprecipitation （CHIP） was used to measure the binding activity of NF-KB and STAT1 to iNOS promoters. Results HSF1 inhibition or knockdown pre- vented the LPS- and/or IFN-γ-stimulated iNOS protein expression in cultured microglia. HSF1 inhibition blocked iNOS mRNA transcription. These inhibitory effects of HSF1 inhibition on iNOS expression were confirmed in brain tissues from endotoxemic mice. Further analysis showed that HSF1 inhibition had no effect on IKB-α degradation and NF-KB or STAT1 phosphorylation in LPS/IFN-γ-stimulated cells. The nuclear transport of active NF-KB or STAT1 was also not affected by HSF1 inhibition. But HSF1 inhibition reduced the binding of NF-KB and STAT1 to their DNA elements. In addition, HSF1 inhibition reduced NF-KB and STAT1 bindings to iNOS promoter inside the LPS/IFN-γ-stimulated cells. Conclusions This preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.
文摘Aim Magnesium lithospermate B （MLB） is the most abundant hydrophilic active component of Salvia rniltiorrhiza Radix, a traditional Chinese herbal medicine mainly used to treat cardiovascular diseases. Studies have shown that endothelial activation contributes to the pathophysiology of cardiovascular diseases such as atherosclero- sis, diabetic vasculopathy, heart failure and hypertension. In the present study, the effects of MLB on endothelial activation were investigated. Lipopolysaccharide （LPS） 1 mg L^-1 was employed to induce endothelial activation, which was determined by relative gene expression and endothelial adhesion assay. Results showed that pretreatment with MLB attenuated LPS-induced ICAM1, VCAM1 and TNF-α upregulation in human dermal microvascular endo- thelial cells （HMEC-1） in dose-dependent manner, which contributed to the reduction of THP-1 adhesion to HMEC-1. Furthermore, it was revealed that 100 μmol · L^-1 MLB significantly decreased the nuclear translocation of NF-KB p65, a critical transcription factor in LPS-indueed inflammatory response, through the inhibition of IKBμ degradation. Besides, the transcriptional activity of NF-KB p65 was also inhibited by the pretreatment of MLB. Mo- reover, MLB pretreatment considerably inhibited LPS-induced p38 phosphorylation, which at least partly contribu- ted to the reduction of ICAM1 expression. In conclusion, these findings suggest that MLB inhibits LPS-induced nu- clear translocation and transcripitional activity of NF-KB, thus attenuates the increased expression of adhesion mole- cules and inflammatory factors, protects endothelial cells from LPS-induced activation.
基金The project supported by Department of Industrial Technology,Ministry of Economic Affairs,Chinese TaipeiMedical and Pharmaceutical Industry Technology and Development Center
文摘OBJECTIVE The emerging role of chronic inflammation is the major degenerative diseases of modern society such as periodontitis,atherosclerosis,rheumatoid arthritis,Parkinson′s disease and even cancer.Eight components were isolated from Derris laxiflora Benth.,In this study,we found these compounds from Derris laxiflora Benth suppress lipopolysaccharide-induced inflammatory response in murine macrophage(RAW 264.7)cells.METHODS RAW 264.7cells were cultured in DMEM media supplemented with 10%(V/V)heated-inactivated FBS,penicillin 100U·mL-1 and streptomycin 100μg·mL-1.The cells were incubated at 37℃in a humidified atmosphere of 5%CO2in air.RAW264.7cells were seeded in a 24-well plate at a density of 2×105 mL-1 and then incubated with or without LPS(100ng·mL-1)in the absence or presence of compounds for 24 h.Effects of these isolates on NO production were measured indirectly by analysis of nitrite levels using the Griess reaction.Quercetin was used as a positive control.RESULTS ight components were isolated from Derris laxiflora Benth.,including three new pterocarpans 7,6′-dihydroxy-3′-methoxypterocarpan(1),derrispisatin(2),derriscoumaronochromone(3)and three new flavonoids cis-3,4′-dihydroxy-5,7-dimethoxyflavan(4),derriflavanone B(5),iso-lupinenol(6)as well as two known ones,lonchocarpol A(7)and lonchocarpol D(8).The structures of these new compounds were determined by analysis of their spectroscopic data.Raw264.7 cells were treated with the compounds from Derris laxiflora Benth for 24 h.Among them,compounds 5,7 and 8 significantly suppressed the NO production in LPS-treated RAW264.7 cells with IC50 values<10μg·mL-1.CONCLUSION In this study,we found that compounds from Derris laxiflora Benth suppresses lipopolysaccharide-induced inflammatory response in murine Raw264.7 cells.
基金The project supported by National Natural Science Foundation of China(81473383,81573645)
文摘OBJECTIVE To investigate the effects of ex-tract of Ramulus Cinnamom(RC)against LPS-induced inflammation in microglia.METHODS Activated microglia releases various pro-inflammatory cytokines to induce neuroinflammation in stroke.Lipopolysaccaride(LPS)is an endotoxin from the outer membrane of Gram-negative bacteria that activates microglia.MTT assay was used to observe the cell viability.The content of NO in supernatant was measured by Griess reagent.The levels of IL-1β,IL-6 and TNF-αin supernatant were detected by ELISA kits.The intracellular COX-2,TLR4,and My D88expression was assayed by Western blotting.RESULTS RC extract 30 and 100μg·m L-1significantly decreased the production of related inflammatory factors such as NO(P<0.05,P<0.01),IL-1β(P<0.01,P<0.01),IL-6(P<0.05,P<0.01)and TNF-α(P<0.01,P<0.01).Furthermore,RC extract significantly inhibited the COX-2,TLR4,and My D88 expression induced by LPS in BV2cells.CONCLUSION RC extract may have therapeutic potential for the improvement of neuroinflammation,and the mechanism may be involved in down-regulation of TLR4/My D88 inflammation pathway.
基金The project supported by National Natural Science Foundation of China(81473383,81573645)
文摘OBJECTIVE Xiao-xu-ming decoction(XXMD),a well-known traditional Chinese herbal prescription,has been widely used to treat stroke.It is recorded in″Bei Ji Qian Jin Yao Fang″written by Si-miao Sun of the Chinese ancient Tang Dynasty.In our previous study,the active fraction of XXMD(XXM)against cerebral ischemia has been prepared by modern separation and purification techniques.This study was to investigate XXM against lipopolysaccaride(LPS)-induced neuroinflammation in mice.METHODS LPS is an endotoxin from the outer membrane of Gram-negative bacteria that activates inflammation.XXM was pre-treated in BALB/C mice followed by injected intraperitoneally with LPS(5 mg·kg-1).The effects of XXM on LPS-induced pro-inflammatory factors and proteins were measured by ELISA,Western blot,and immunofluorescence in vivo.RESULTS Mice treated with XXM showed significantly decreased proinflammatory factors level,including IL-1β(P<0.01),IL-6(P<0.01),TNF-α(P<0.05),and MCP-1(P<0.01).Furthermore,XXM also significantly inhibited the inflammatory pathway proteins expression induced by LPS,including TLR4,MyD 88,and COX-2.CONCLUSION XXM possesses anti-neuroinflammation in mice and might be a promising therapeutic agent for stroke.
基金Open Fund from the State Key Laboratory of Phytochemistry and Plant Resources in West China(P2017-KF13)National Science and Technology Major Project of China(2016ZX09J16104)
文摘OBJECTIVE To evaluate the neuroprotective effects of 4 components from Uncaria rhynchophylla(297,307,315 and 327)on long-term potentiation(LTP)deficit in neuroinflammation animal model induced by lipopolysaccha⁃ride(LPS).METHODS Male BALB/c 18-22 g mice were divided into control group,model group and component treat⁃ment group(1μg per mouse,icv);each group contained 5 mice,model group and compound treatment group were intra⁃peritoneally injected LPS(50μg·kg^-1,ip)4 h before LTP induction.LTP of perforant path-dentate gyrus pathway in hippo⁃campus was induced by high frequency stimulation and used to evaluate the effects on synaptic plasticity.RESULTS Compared with control group,the LTP of the model group was significantly impaired.Compound 297,327,and 307 could significantly improve LPS-induced LTP impairment.315 had no significant effect on LPS-induced LTP impairment.CONCLUSION Hippocampal synaptic plasticity could be impaired by LPS.Compounds 297,327 and 307 have protec⁃tive effects against LPS induced LTP impairment,and 315 has little effect on LPS induced LTP impairment.These re⁃sults suggested that 297,327 and 307 might have potential effects on neuroinflammation induced memory deficits.
基金Supported by the National Natural Science Foundation of China(31772806)Undergraduate Innovative Entrepreneurship Program in Heilongjiang Province(201810224054)the National Key Research and Development Program of China(2016YED0501008)
文摘Sepsis can cause a series of damages to various organs of the body,so it has always been regarded as a hot research topic in veterinary clinic.Aiming at the present situation of high morbidity and mortality of canine sepsis,in order to further explore the pathogenesis of this disease,it need to establish a stable and repeatable canine sepsis model that is in line with the clinical characteristics of the disease.The study selected 12 local dogs and randomly divided into three groups:rapid bolus injection group(Group A),continuous infusion group within 30 min(Group B)and continuous infusion group(Group C).Then,the lipopolysaccharides(LPS)with 2 mg·kg^-1 were injected through the brachial vein in different modes of administration,thus the model was fully established.During the modeling period,body temperature(T),respiratory rate(RR),heart rate(HR)and mean arterial pressure(MAP)were monitored at 0,10,20,30,40,50 min and 1,2,3,4,5,6,7,8,9,12 and 24 h.Blood was collected from the canine brachial vein at 0,1,2,3,4,5,6,7,8,9,12 and 24 h,respectively,for the detection of white blood cells(WBC).The test showed that the values of T,RR,HR,MAP,WBC of the dogs in group A all changed but did not exceed the normal range,and the clinical symptoms were not significant.There were no significant changes in the values of RR,HR,T,MAP and WBC of the dogs in Group B,and the clinical symptoms were not significant.The value of T of the dogs in Group C were significantly increased at 40 min(p<0.05),which reached the fever standard and lasted for 7 h;the value of RR increased significantly at 20 min(p<0.05),and a downward trend could be observed at 12 h,then it returned to normal at 24 h;the value of HR increased significantly at 50 min(p<0.05)and recovered at 8 h;the value of HR decreased significantly at 20 min(p<0.05),which remained at 12 h(p<0.05),and returned back to normal at 24 h;the value of WBC decreased significantly from 1 h to 4 h(p<0.05),which was lower than the normal value,and increased significantly at 24 h(p<0.01);all of the four dogs in this group had clinical symptoms such as vomiting,diarrhea and depression.Based on the above results,the changes of indexes and clinical symptoms in Groups A and B did not meet the standards of sepsis.After a long-term continuous intravenous infusion of LPS,the experimental dogs in Group C showed varying degrees of clinical symptoms,such as vomiting,diarrhea and depression one after another.The indexes and clinical symptoms reached the sepsis standard about 3 h after infusion.In brief,this model not only had good stability and good regularity of repeatability,but also lasted for a long time and could be suitable for other subsequent studies.
基金The project supported by the National Natural Science Foundation of China(81160048)the Science and Technology Development Fund,Macao S.A.R(FDCT)(021/2012/A1)the Research Fund of University of Macao(MRG007/CXP/2013/ICMS)
文摘OBJECTIVE Isofuranodiene(ISO)is a natural product isolated in Chinese herb.Our recent results showed anticancer effect in vitro.In this study,we investigated its live protective effect with a Dgalactosamine/lipopolysacchride(GalN/LPS)induced rat model.METHODS SD rats were treated orally with or without ISO(20 and 50mg·kg-1,ig)for 3d and then treated with GalN/LPS for 8h.The serum were collected and the concentration of aspartate aminotransferase(AST),alanine aminotransferase(ALT),and malondialdehyde(MDA)were determined.The liver injury was examined by H&E staining.The mRNA expression of IL-1β,IL-6 and inducible nitric oxide synthase(iNOS)in liver tissues were determined by realtime PCR.RESULTS Oral administration of ISO(20 and 50mg·kg-1)dramatically inhibited GalN/LPS-induced serum elevation of AST,ALT,and MDA levels.The liver injury was also significantly ameliorated as evidenced by the histological improvement in H&E staining.Furthermore,ISO treatment significantly inhibited GalN/LPS-induced mRNA expression of IL-1β,IL-6,and inducible nitric oxide synthase(iNOS)in liver tissues.CONCLUSION This data showed that ISO has hepatoprotective effect in rats.
基金Supported by the National Key Research and Development Program of China(2016YED0501008)the National Natural Science Foundation of China(31772806)the Natural Science Foundation of Heilongjiang Province(C2017022)。
文摘The model of acute lung injury(ALI)was established by intraperitoneal administration,but there was no time-point observation and comparison.ALI model was established by intraperitoneal injection of lipopolysaccharide(LPS)at the concentration of 10 mg·kg^-1 (10 mg LPS dissolved in 1 mL normal saline to prepare 1 mL·kg^-1solution)in rats.The control group(CG)was intraperitoneally injected with saline of the same dose.In the LPS group,lung tissues were collected at 4,6,8,12 and 24 h after administration.Then,the morphology changes,the ratio of wet-to-dry weight(W/D),the expression of interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)proteins,the levels of malondialdehyde(MDA),the activities of superoxide dismutase(SOD),glutathione peroxidase(GSH)were measured.To verify the success of the model,the degrees of lung injury via Western blot,RT-PCR,ELISA and other techniques were detected at different time points,and the severe time of the ALI model established was deterimined by intraperitoneal administration,which provided a stable model basis for the study of the pathogenesis of ALI in the future.The results showed that the lung injury occurred in LPS group.W/D and lung pathological changes at 12 and 24 h of LPS group were significantly different from those in the CG.Compared with the CG,the expression of IL-1βand TNF-αproteins and the content of MDA in lung tissues of LPS group increased and most significant difference was found at 12 and 24 h(p<0.01).Compared with the CG,the activities of SOD and GSH in LPS 12 h group decreased significantly(p<0.01).In conclusion,inflammation and oxidative damage were the main causes of the ALI in rats.Lung injury was most obvious 12 h after intraperitoneal injection of 10 mg·kg^-1 LPS.
