The aims of the present study are to investigate the effect of vasoconstriction and to explore the mechanism of rutae- carpine. The research findings showed that rutaecarpine could induce contractions of the rat thoracic aorta in vitro. The inhibitors of Rho-kinase and inositol 1,4,5-triphosphate receptor （IP 3 R） could suppress the effect of rutaecarpine-induced vasoconstriction. In the study of A7r5 cells （a line of smooth muscle cells）, 300 μg/L rutaecarpine promoted the concentration of intracellular Ca 2＋ and enhanced the IP 3 R expression, which connects with 1,4,5-triphosphate to evoke the release of Ca 2＋ from the intracellular stores. Rutaecarpine increased the RhoA mRNA expression when the cells were pretreated with inhibitor H-1152, and improved the levels of phosphorylation of myosin light chain phosphatase （MLCP） and myosin light chain （MLC）. These results suggest that rutaecarpine plays a role in vasoconstriction relative to the RhoA/MLCP-MLC signaling pathway, which denotes a new field of rutaecarpine in pharmacology.
Heat stress can stimulate an increase in body temperature, which is correlated with increased expression of heat shock protein 70 （HSP70） and tumor necrosis factor a （TNFa）. The exact mechanism underlying the HSP70 and TNFa induction is unclear. Berberine （BBR） can significantly inhibit the temperature rise caused by heat stress, but the mechanism responsible for the BBR effect on HSP70 and TNFa signaling has not been investigated. The aim of the present study was to explore the relationship between the expression of HSP70 and TNFa and the effects of BBR under heat conditions, using in vivo and in vitro models. The expression levels of HSP70 and YNFa were determined using RT-PCR and Western blotting analyses. The results showed that the levels of HSP70 and TNFa were ap-regulated under heat conditions （40 ~C）. HSP70 acted as a chaperone to maintain TNFa homeostasis with rising the temperature, but knockdown of HSP70 could not down-regulate the level of TNFa. Furthermore, TNFa could not influence the expression of HSP70 under aormal and heat conditions. BBR targeted both HSP70 and TNFa by suppressing their gene transcription, thereby decreasing body temperature under heat conditions. In conclusion, BBR has a potential to be developed as a therapeutic strategy for suppressing the thermal effects in hot environments.
Berberine （BBR） has a variety of pharmacological activities. Studies have reported that BBR not only reduces heat stress-induced fever but also inhibits lower body temperatures due to cold stress. Heat stress can be reduced via BBR treatment, which antagonizes HSP70-TNFa to regulate the body temperature alteration. In cold stress, however, the molecular mechanism of BBR-induced inhibition of hypothermia remains unclear. Therefore, we studied whether BBR promoted uncoupling protein 1 （UCP1, a crucial protein of thermogenesis） expression and its mechanism under cold stress. Wild type mice and Ucpl-/- mice were used for the in vivo experiments, and primary brown adipocytes and brown adipocytes HIB-1B were used for the in vitro studies. The cold stress was set at 4℃. The results showed that at 4℃, the body temperature of mice was decreased. BBR effectively inhibited this hypothermia. Simultaneously, Ucpl expression in brown adipose tissue （BAT） cells was significantly increased, and BBR promoted Ucpl expression. However, in Ucpl-knockout mice, the effect of BBR on hypothermia disappeared during cold stress, indicating that the main target for BBR regulation of body temperature was Ucpl. Further studies showed that the transcriptional response element NFE2 （nuclear factor erythroid-derived 2） in the upstream of the Ucpl promoter region contributed to the positive regulatory role on Ucpl expression at lower temperature. BBR could bind to the sequence of NFE2 response element in a temperature-dependent manner. Increased affinity of BBR binding to NFE2 response element in cold stress significantly strengthened and enhanced the expression of Ucpl. This work was important for understanding the role of BBR on thermogenesis in BAT, body temperature regulation and temperature tolerance under cold conditions.
Brazilein is reported to have immunosuppressive effect on cardiovascular and cerebral-vascular diseases. The essential roles of innate immunity in cerebral ischemia are increasingly identified, but no studies concerning the influence of brazilein on the innate immunity receptors have been reported. The present study was designed to investigate the regulation of NOD2 （Nucleotide-binding oligomerization domain-containing protein 2） by brazilein for its protection of neuron in cerebral ischemia in vivo and oxygen-glucose deprivation in vitro. The results showed that brazilein could reverse the elevated expression of NOD2 and TNFa （tumor necrosis factor alpha） elicited by cerebral ischemia and reperfusion. This reduction could also be detected in normal mice and C 17.2 cells, indicating that this suppressive effect of brazilein was correlated with NOD2. The results from GFP reporter plasmid assay suggested brazilein inhibited NOD2 gene transcription. In conclusion, brazilein could attenuate NOD2 and TNFα expression in cerebral ischemia and NOD2 may be one possible target of brazilein for its immune suppressive effect in neuro-inflammation.
YAN Xiao-JinCHAI Yu-ShuangYUAN Zhi-YiWANG Xin-PeiJIANG Jing-FeiLEI FanXING Dong-MingDU Li-Jun
Berberine, an isoquinoline alkaloid component of Rhizoma Coptidis has been demonstrated to be the key active ingredient involved in its protective effect against cerebral ischemia-reperfusion. However, the comparison among the analogues to the protective effect against oxygen and glucose deprivation/reoxygenation （OGD-R） was mediated by inhibition of cyclooxygenase-2 （COX-2） has never been reported. The aim of this study is to investigate the protective effect of berberine and its five analogues against OGD-R in PC 12 cells, as well as to determine whether the protective effect was regulated through COX-2. An established in vitro OGD-R model of PC12 cells by oxygen glucose deprivation of 4 h and reperfusion of 24 h was used in our study. After cells were treated with berberine or its five analogues, we examined the cell viability assay by 3-（4,5-dimethylthiazol-2-yl）-2,5- diphenyltetrazolium bromide （MTT） assay. Cells were also collected to determine the levels of mRNA and protein of COX-2 by real time PCR and Western blot. We found that berberine and its analogues improved the viability of PC12 cells against OGD-R. Whereas berberine and berberrubine presented stronger activity with the most effective dose of 0.31 lag/mL and the minimum effective doses of 0.02 and 0.04 gg/mL. Palmatine possessed potentially weaker protective effect. The mRNA level of COX-2 in cells treated with berberine, coptisine and epiberberine was decreased significantly. The protein level of COX-2 was significantly down-regulated in cells treated with berberine. Studies suggested the important role of methylenedioxy groups （R2 and R3） of berberine analogues in COX-2 inhibitory effect, and methylenedioxy groups （R2, R3, R9 and R10） in berberine analogues in binding affinity with COX-2. Substituted hydroxyl group at R9 did not affect the activity of berberine. In summary, our study illustrated the protective effects of berberine and its analogues in PCI2 cells against OGD-R and to elucidate the structure-activity relat
Objective: To explore the anti-nociceptive effect of patchouli alcohol(PA), the essential oil isolated from Pogostemon cablin(Blanco) Bent, and determine the mechanism in molecular levels. Methods: The acetic acid-induced writhing test and formalin-induced plantar injection test in mice were employed to con?rm the effect in vivo. Intracellular calcium ion was imaged to verify PA on mu-opioid receptor(MOR). Cyclooxygenase 2(COX2)and MOR of mouse brain were expressed for determination of PA’s target. Cellular experiments were carried out to find out COX2 and MOR expression induced by PA. Results: PA significantly reduced latency period of visceral pain and writhing induced by acetic acid saline solution(P<0.01) and allodynia after intra-plantar formalin(P<0.01) in mice. PA also up-regulated COX2 mRNA and protein(P<0.05) with a down-regulation of MOR(P<0.05) both in in vivo and in vitro experiments, which devote to the analgesic effect of PA. A decrease in the intracellular calcium level(P<0.05) induced by PA may play an important role in its anti-nociceptive effect.PA showed the characters of enhancing the MOR expression and reducing the intracellular calcium ion similar to opioid effect. Conclusions: Both COX2 and MOR are involved in the mechanism of PA’s anti-nociceptive effect,and the up-regulation of the receptor expression and the inhibition of intracellular calcium are a new perspective to PA’s effect on MOR.
