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Effect of nigranoic acid on Ca2+ influx and its downstream signal mechanism in NGF-differentiated PC12 cells
Xiao-Xi Yuana， b， Li-Ping Yanga， b， c， Zhong-Lin Yanga， b， Wei-Lie Xiaoa， ， ， Han-Dong Suna， Gui-Sheng Wua， Huai-Rong Luoa， ，
Ethnopharmacological relevance
Schisandra chinensis (Wuweizi) is a tonic in traditional Chinese herbal medicine that has been widely used for thousands of years to treat insomnia， remove fatigue， and enhance mental and intellectual functions. Many studies showed that lignans were the major active components of Schisandra chinensis， whereas the bioactivity of the abundant triterpenoids in the fruit of Schisandra chinensis， such as nigranoic acid (SBB1， 3，4-secocycloartene triterpenoid)， has not been examined yet in neuropathology.
Materials and methods
After treating with SBB1， intracellular Ca2+ concentration was analyzed by Ca2+ fluorescent indicator (Fluo-4 AM) in NGF-differentiated PC12 cells. Intracellular nitric oxide (NO) level was analyzed using NO fluorescent indicator (DAF-FM). The expression of extracellular signal regulated kinase 1 and 2 (ERK1/2) was analyzed by western blotting， and the temporal mRNA for BDNF and c-fos was analyzed using reverse transcription quantitative PCR.
Result
We found that SBB1 induced Ca2+ influx in a time- and concentration-dependent manner， which was significantly attenuated in Ca2+ free media. SBB1 promoted the intracellular NO production which depended on increasing cytoplasmic Ca2+ level Moreover， SBB1 stimulated activation of ERK1/2 through Ca2+-CaMKII pathway. In addition， we found that SBB1 increased the expression of BDNF and c-fos mRNA.
Conclusion
These results suggest that SBB1 is able to promote NO production and stimulate phosphorylation of ERK1/2 through Ca2+ influx， further impact expression of BDNF and c-fos， which provides evidence for the effects of SBB1 that may be benefit to enhance mental and intellectual functions.
