Study On The Effect And Mechanism Of Cryptotanshinone On Lipopolysaccharide-induced Inflammation Of BV2 Microglia

Background:Neuroinflflammation and oxidative stress play an important role in the pathogenesis of neurodegenerative diseases.Strict regulation of proinflammatory and anti-inflammatory mediators during the inflammatory response can prevent or delay the occurrence of neurodegenerative disease.Microglial cells,activated by lipopolysaccharide(LPS),are commonly used for in vitro studies on neuroinflflammation-related diseases.Activated microglia can induce some pro-inflammatory proteases(such as inducible nitric oxide synthase and cyclooxygenase-2)by activating transcription factors such as NF-?B,thereby triggering the release of certain inflammatory mediators(such as nitric oxide Nitrogen,tumor necrosis factor-?,interleukin-1? and interleukin-6).Overproduction of these proinflflammatory mediators results in neurodegeneration or even neuronal death,accelerating the onset and progression of neurodegenerative diseases.Thus,Controlling the excessive activation of microglia may be a potential therapeutic direction for the treatment of neurodegenerative diseases related to neuroinflammation.Cryptotanshinone(CTN),a monomer compound extracted from the dried roots and rhizomes of Salvia miltiorrhiza Bge,exhibits anti-oxidative and anti-inflflammatory effects.Studies have shown that cryptotanshinone can inhibit the activity of nuclear factor-?B and cyclooxygenase-2 activated by lipopolysaccharide in macrophages;it can inhibit the expression of endothelin-1 and improve the inflammatory response.For microglia,whether cryptotanshinone can still suppress its inflammatory response and the mechanism therein is rarely reported.Objective:Observe whether cryptotanshinone can inhibit lipopolysaccharide-induced microglial inflammation and explore its potential mechanism.Methods:1.The lactate dehydrogenase method was used to detect cell viability,and the safe concentration of cryptotanshinone acting on BV-2 microglia cells was determined.;2.The NO level of the cell supernatant was detected by the nitric oxide kit;3.The levels of inflammatory factors(TNF?,IL1,IL6)were detected by RT-PCR;4.The levels of iNOS,COX 2,NF-? B,Nrf2 and HO-1 were detected by Western blot;5.The activities of NF-? B,Nrf2 and HO-1 were detected by immunofluorescence;6.Using siRNA cell transfection technology to verify whether cryptotanshinone inhibits the neuroinflammatory reaction by regulating the Nrf2 / HO-1 pathway.Results:1.1-10?mol / L cryptotanshinone is non-toxic to the cells.For the convenience of experimental operation,5 and 10?mol / L were selected as the effective concentration of cryptotanshinone;2.Cryptotanshinone can inhibit lipopolysaccharide-induced NO secretion of BV-2microglia;3.Cryptotanshinone can inhibit the secretion of inflammatory factors(TNF?,IL1? and IL6)induced by lipopolysaccharide in BV-2 microglia cells;4.Cryptotanshinone can inhibit the expression of iNOS,COX2,NF-?B in BV-2microglia,and increase the expression of Nrf2 and HO-1;5.Cryptotanshinone can inhibit the activity of NF-?B induced by lipopolysaccharide in BV-2 microglial cells,and can enhance the activity of Nrf2 and HO-1 proteins;6.Cryptotanshinone relies on the Nrf2 / HO-1 pathway to inhibit lipopolysaccharide-induced inflammation of BV-2 microglia.Conclusion:Cryptotanshinone can play a neuroprotective role by activating Nrf2 / HO-1pathway to inhibit the production of inflammatory cytokines in microglia.