MATERIALS AND METHODS: Tincture of the roots was concentrated to dryness by evaporating the ethanol in vacuo. This ethanolic extract was partitioned into 5 fractions sequentially with hexane, dichloromethane (DCM), ethyl acetate, butanol, and water. The corpus cavernosum relaxant activity of each fraction was investigated. The DCM fraction which showed the highest potency in relaxing phenylephrine-precontracted corpora cavernosa was purified by column chromatography. The effects of the most potent DCM subfraction in relaxing phenylephrine-precontracted corpora cavernosa, DCM-I, on angiotensin I- or angiotensin II-induced contractions in corpora cavernosa were investigated. The effects of DCM-I pretreatment on the responses of phenylephrine-precontracted corpora cavernosa to angiotensin II or bradykinin were also studied. An in vitro assay was conducted to evaluate the effect of DCM-I on angiotensin-converting enzyme activity.
RESULTS: Fraction DCM-I decreased the maximal contractions (100%) evoked by angiotensin I and angiotensin II to 30 ± 14% and 26 ± 16% (p < 0.001), respectively. In phenylephrine-precontracted corpora cavernosa, DCM-I pretreatment caused angiotensin II to induce 82 ± 27% relaxation of maximal contraction (p < 0.01) and enhanced (p < 0.001) bradykinin-induced relaxations from 47 ± 8% to 100 ± 5%. In vitro, DCM-I was able to reduce (p < 0.001) the maximal angiotensin-converting enzyme activity to 78 ± 0.24%.
CONCLUSION: Fraction DCM-I was able to antagonize angiotensin II-induced contraction to cause corpus cavernosum relaxation via inhibition of angiotensin II type 1 receptor and enhance bradykinin-induced relaxation through inhibition of angiotensin-converting enzyme.
OBJECTIVE: This study aimed to evaluate the anti-inflammatory and neuroprotective properties of extracts obtained from the roots of PS against beta-amyloid (Aβ)-induced microglial toxicity associated with the production of pro-inflammatory mediators.
METHOD: BV2 microglial cells were treated with hexane (RHXN), dichloromethane (RDCM), ethyl acetate (REA) and methanol (RMEOH) extracts of the roots of PS prior to activation by Aβ. The production and mRNA expression of pro-inflammatory mediators were evaluated by Griess reagent, ELISA kits and RT-qPCR respectively. The phosphorylation status of p38α MAPK was determined via western blot assay. BV2 conditioned medium was used to treat SH-SY5Y neuroblastoma cells and the neuroprotective effect was assessed using MTT assay.
RESULTS: PS root extracts, in particular RMEOH significantly attenuated the production and mRNA expression of IL-1β, IL-6 and TNF-α in Aβ-induced BV2 microglial cells. In addition, RHXN, REA and RMEOH extracts significantly reduced nitric oxide (NO) level and the inhibition of NO production was correlated with the total phenolic content of the extracts. Further mechanistic studies suggested that PS root extracts attenuated the production of cytokines by regulating the phosphorylation of p38α MAPK in microglia. Importantly, PS root extracts have protective effects against Aβ-induced indirect neurotoxicity either by inhibiting the production of NO, IL-1β, IL-6, and TNF-α in BV2 cells or by protecting SHSY5Y cells against these inflammatory mediators.
CONCLUSIONS: These findings provided evidence that PS root extracts confer neuroprotection against Aβ- induced microglial toxicity associated with the production of pro-inflammatory mediators and may be a potential therapeutic agent for inflammation-related neurological conditions including Alzheimer's disease (AD).