The Effects And Mechanisms Of Curcuma Compounds On LPS-Induced Neuroinflammation

Objective:Curcuma longa(turmeric)is an important medicinal and edible plant,which has drawn public attention for its pleiotropic activities.Thoughout the history of turmeric medical use,it has widely been used in chronic inflammatory diseases.It is worthy noting that,more and more studies indicate turmeric shows neuronal protective effects.Due to the strong anti-inflammatory activities and neuronal protective effects,we hypothesized that curcumin and the main component of TEO(aromatic-turmerone,Art)maybe have potential in neuroinflammation.Therefore,in this study,we established neuroinflammation model induced by lipopolysaccharide(LPS)to evaluate the anti-neuroinflammation activities of curcumin and aromatic-turmerone,and investigate the mechanisms underlying their actions.This will provide theoretical and experimental basis for further development of curcuma compounds treating neuroinflammation-related diseases.Methods:(1)The effects and mechanisms of curcumin and piperine administered individually or in combination toLPS-induced neuroinflammation models.Mice were randomly divided into seven groups,including normal control,LPS(250 ?g/kg,i.p.),LPS+Cur-L(100 mg/kg,p.o.),LPS+Cur-H(200 mg/kg,p.o.),LPS+Pip(20 mg/kg,p.o.),LPS+Cur-L+Pip(100 mg/kg+ 20 mg/kg,p.o.)and LPS+Cur-L+Pip(100 mg/kg+ 20 mg/kg,p.o.)groups.Mice were treated with curcumin and piperine consecutively for 14days followed by intraperitoneal administrationof LPS(250 ?g/kg,i.p.)for 7 days.Once the chronic neuroinflammation model established and administration of medicine done,brain tissues were harvested for histopathologic and inflammatory markers'(TNF-? and IL-1?)examination.Also,we utilized LPS to establish a microglia(BV2)cellular neuroinflammation model.Pretreated with curcumin for 1 h,andchallenged with LPS,cells were harvested then to determine the inflammatory markers' expression by Western Blotting.(2)The effects of aromatic-turmerone on LPS-induced neuroinflammation.Mice were randomly divided into six groups,including normal control,LPS(250 ?g/kg,i.p.),LPS+TTP488(5 mg/kg,i.p.),LPS+TEO(100 mg/kg,p.o.),LPS+Art-L(50 mg/kg,p.o.)and LPS+ Art-H(100 mg/kg,p.o.)groups.Mice were treated with TTP488,TEO and Art for 14 days followed by LPS(250 ?g/kg,i.p.)administration for 7 days.After 2 weeks of drugs administration,the learning and memory abilities were inspected by Morris water maze test;microPET-CT imaging was conducted to evaluate the cerebral glucose metabolism of the mice.The nissl staining experiment was conducted to detect the neurons' apoptosis in mice brains,and the neuroinflammatory biomarkers in mice brains were detected by immunohistochemistry and ELISA assay kits.(3)The mechanism study of aromatic-turmerone's alleviation effect on LPS-induced neuroinflammation in BV2 cells.Similar to above protocol,we used LPS-induced neuroinflammatory cell model to investigate the molecular mechanism of aromatic-turmerone.Results:(1)Results demonstrated that curcumin could improve LPS induced neuronal damage and related inflammatory response.Curcumin administration elevated Nissl substance in cells and increased the number of normal neurons,while decreased TNF-? and IL-1?levels.In addition,co-administration of piperine with curcumin potentiated their neuroprotective effects as compared to the treatment with curcumin alone.We further examined the molecular mechanisms of curcumin.It was found that curcumin suppressed TLR4 mediated downstream signaling,in which the protein expressions of Myd88,MAPK,and NF-?B were decreased.Further,curcumin down-regulated the release of TNF-? and IL-1? from TLR4 siRNA-transfected cells that were treated by LPS.(2)There were distinct differences in behavioral experiments,micro-PET/CT,histoligical and biochemical examination between control and LPS group.Administration of TTP488 and aromatic-turmerone could reverse these changes.In Morris water test,aromatic-turmerone attenuated LPS-induced memory and cognitive impairment;micro-PET/CT imaging showed that aromatic-turmerone up-regulated brain blood flow and cerebral glucose metabolism;pathologically,aromatic-turmerone elevated Nissl substance and increased the number of healthy neurons,all these data indicated that aromatic-turmerone showed neuroprotective effects on LPS-challenging mice.At the same time,we tested the effects of aromatic-turmerone on neuroinflammatory responses.Results showed that aromatic-turmerone inhibited the activation of microglia and generation of inflammatory cytokines,indicating aromatic-turmerone improved learning and memory function probably by decreasing LPS-elicited inflammatory responses.(3)Due to the similarity of structure and origin,we test whether TLR4 is involved in aromatic-turmerone' anti-inflammation in vitro.Cellular experiments results showed that curcumin effectively reduced cytotoxicity of LPS,and decreasedthe secretion levels of TNF-?and IL-1?,indicating the anti-inflammation activity of aromatic-turmerone as well.Further,aromatic-turmerone decreased the expressions of Myd88,MAPK and NF-?B induced by LPS,while treatment with aromatic-turmerone showed no evident effect on the expression of TLR4.To further confirm the role of TLR4 in aromatic-turmerone against LPS-induced neuroinflammation,we used siRNA to knockdown TLR4 gene.Results revealed that the levels of TNF-? and IL-1? in aromatic-turmerone-pretreated siRNA cells were down-regulated in the presence of LPS,suggesting that aromatic-turmerone targeted TLR4-mediated downstream signaling,and lowered the release of inflammatory mediators.Conclusions:Data from mice experiments suggested that curcuma compounds could relieve LPS-induced neuroinflammation,concomitant memory impairment,cerebral glucose metabolism and neuron damage.Cellular results showed that curcuma compounds inhibited TLR4-mediated downstream signaling,and lowered the release of inflammatory cytokines.The results of the current study provided scientific cluesfor the chemical modification of curcuma compounds and development of curcuma compounds in treating neuroinflammation-related diseases.