The Inhibitory Effects And Mechanisms Of Bisbenzylisoquinolie Alkaloids On The Neuroinflammation

Microglial cells are a kind of macrophage cells that reside in the central nervous system(CNS),and considered to be the most important immune cell in the CNS.Microglial cells exhibit the resting state in the healthy brain,and play a role in eliminating metabolites and protecting the neurons.Microglial cells can be activated when the CNS suffers from stimuli,such as trauma or inflammation.Studies have documented that hyperactivated microglial cells can induce the damage and death of neurons by releasing of inflammatory factors,such as TNF-?,IL-6,and IL-1?,and cause or participate in the production of neuroinflammation.The present study investigated the inhibitory effects of 12 bisbenzylisoquinolie alkaloids(BBI)(Isoliensinine,Dauricine,Liensinine,Fangchinoline,D-Tetrandrine,Isotetrandrine,Neferine,Berbamine Hydrochloride,Cepharanthine,Cycleanine,D-isochonchrodendrine and(-)-Curine)on LPS-stimulated microglial activation,as well as the mechanisms of the active compounds on the NF-?B signaling pathway.In order to prevent the possible effects of reduced cell viability on NO producing,the cytotoxic influence of 12 bisbenzylisoquinolie alkaloids on LPS-activated microglial cells were evaluated by MTT assays.It was showed that 12 bisbenzylisoquinolie alkaloids(0.1–10 ?mol/L)did not affect the viability of LPS-activated microglial cells.Then,the production of NO by LPS-activated microglial cells was detected by Griess reagent.And the IC50 value was calculated.The results showed that all these12 alkaloids(0.1-10?M)suppressed the NO release to some extent.The analysis of the structure-activity relationships of these alkaloids manifested that:(1)Space structure of the attaching of head to tail enhanced torsion resistance,which improved receptor binding,benifited the elevating of the activity;(2)The molecular space configuration and conformation induced by 1 position of the chiral carbon atom had significantly impacted the activity;(3)The existing of phenolic hydroxyl group in the benzene ring may decrease activity.The levels of three inflammatory cytokines(IL-6,IL-1? and TNF-?)in cell culture supernatant fluids were measured by ELISA.The ELISA results indicated that the 12 alkaloids could greatly suppress LPS-stimulated release of TNF-?,IL-6,and IL-1? in microglial cells.In order to investigate the mechanisms of three bisbenzylisoquinolie alkaloids(Neferine,Liensinine and Isoliensinine)on LPS-induced microglial activation,Western blot was used to measure the expression of i NOS protein and the phosphorylation and degradation of I?B? in LPS-activated microglial cells after the treatment of the three bisbenzylisoquinolie alkaloids.The results showed that the three alkaloids could inhibit the producing of i NOS protein to some extent,and attenuate the phosphorylation and degradation of I?B? in LPS-stimulated microglial cells.In summary,these findings demonstrate that these 12 bisbenzylisoquinolie alkaloids exert anti-neuroinflammatory activity by suppressing the production of NO,TNF-?,IL-1?,and IL-6 in LPS-activated microglial cells.Liensinine,neferine and isoliensinine inhibit the phosphorylation and degradation of I?B? induced by LPS in microglial cells,thus prevent LPS-induced expression of i NOS protein followed by reduced NO production,and inhibit the release of cytokines(TNF-?,IL-1?,and IL-6).Bisbenzylisoquinolie alkaloids may have the potential in the treatment of neuroinflammation-related diseases caused by microglial activation.