Anti-neuroinflammatory Effects Of Lignanamides From Hemp Seed

The incidence of neurodegenerative diseases,including Alzheimer’s disease,Parkinson’s disease,multiple sclerosis,etc,is becoming higher and higher.The progress of these diseases are related to oxidative stress,immune inflammation,mitochondrial dysfunction,excitotoxin and apoptosis,etc.Studies found that microglia-mediated inflammatory response plays an important role in the development and progression of neurodegenerative diseases.The microglia could be activated by cytotoxins,external chemical,or physical stimulator,and release a large number of inflammatory factors,which aggravate inflammation and progression of neurodegenerative diseases.Microglia-induced neuroinflammation activates many related signaling pathways,such as Toll-like receptors,JAK/STAT,Nuclear factor kappa B(NF-κB),MAPK,etc.NF-κB is an important signaling pathway that mediates the occurrence of inflammation.Activation of NF-κB can induce a large number of inflammatory factors,which will stimulate microglia in return,then lead to a vicious circle of further inflammation.In the progress of the activation of microglia cells,NF-κB will coordinate with other inflammatory-related channels to trigger a variety of signal cascades,which together regulate the inflammatory response.Therefore,It’s a potential method against neurodegenerative disease to discover effective compounds which can regulate the activation of NF-κB signaling pathway.Researches showed that hemp seed extracts have potential neuroprotective activities.Previous work in our laboratory provided a series of lignanamides isolated from hemp seed.In this study,the anti-neuroinflammatory activity of these compounds were screened by establishing neuroinflammatory model using LPS-stimulated BV2 microglia cells.Grossamide and cannabisin F were selected for further mechanism study involving LPS-induced activation of NF-κB signaling pathway to prove their potential against neuroinflammation related diseases.Results showed that grossamide had potential anti-inflammatory activity.ELISA and qRT-PCR tests revealed that grossamide could inhibit the expression of LPS-induced inflammatory factors in BV2 cells.Wetsern blot and immunofluorescence analysis showed that grossamide could inhibit the phosphorylation of IKBa and NF-κKBp65 protein,and prevent the expression of NF-κBp65 into the nucleus.Meanwhile,grossamide coud prevent the expression of NF-κB upstream protein TLR4 and MyD88.Thus,grossamide can inhibit TLR4-mediated NF-κB signaling pathway in BV2 microglia cells.Cannabisin F also showed potential anti-inflammatory activity.The production of intracellular ROS in BV2 microglia cells was inhibited by cannabisin F dose-dependently.Western blot results showed that cannabisin F could promote the expression of SIRT1,Nrf-2 and the downstream protein HO-1.It could also inhibit the phosphorylation of IκBα and NF-KBp65 protein.The results of ELISA and qRT-PCR revealed that cannabisin F inhibited the production of IL-6 and TNF-a in BV2 microglia cells.Thus,cannabisin F can play a neuroprotective role through activating the SIRTl/Nrf-2 signaling pathway and inhibiting NF-κB signaling pathway.