Abstract. As a traditional Chinese medicine, dragon's blood (DB) is widely used in treating various pains for thousands of years due to its potent anti-inflammatory and analgesic effects. In the present study, we observed that intragastric administration of DB at dosages of 0.14, 0.56, and 1.12 g/kg potently inhibited paw edema, hyperalgesia, cyclooxygenase-2 (COX-2) protein expression, or preprotachykinin-A mRNA expression in carrageenan-inflamed or sciatic nerve-injured (chronic constriction injury) rats, respectively. A short-term (15 s or 10 min) pre-exposure of cultured rat dorsal root ganglion (DRG) neurons to DB (0.3, 3, and 30 μg/ml) or its component cochinchinenin B (CB; 0.1, 1, and 10 μM) blocked capsaicin-evoked increases in both the intracellular calcium ion concentration and the substance P release. Moreover, a long-term (180 min) exposure of cultured rat DRG neurons to DB or CB significantly attenuated bradykinin-induced substance P release. These findings indicate that DB exerts anti-inflammatory and analgesic effects by blocking the synthesis and release of substance P through inhibition of COX-2 protein induction and intracellular calcium ion concentration. Therefore, DB may serve as a promising potent therapeutic agent for treatment of chronic pain, and its effective component CB might partly contribute to anti-inflammatory and analgesic effects.

To examine regulatory effects of β-catenin on the biosynthesis and release of substance P, a rat chronic constriction injury (CCI) model and a rat dorsal root ganglion (DRG) cell culture model were used in the present study. The CCI treatment significantly induced the overall expression of β-catenin (158 ± 6% of sham) in the ipsilateral L5 DRGs in comparison with the sham group (109 ± 4% of sham). The CCI-induced aberrant expression of β-catenin was significantly attenuated by oral administration of diclofenac (119 ± 6% of the sham value; 10 mg/kg). Importantly, aberrant nuclear accumulation of β-catenin in cultured DRG cells resulted in up-regulation of the PPT-A mRNA expression and the substance P release. The up-regulation of both the PPT-A mRNA expression and the substance P release by either a GSK-3β inhibitor TWS119 (10 μM) or a Wnt signaling agonist Wnt-3a (100 ng/ml) were significantly abolished by an inhibitor of cyclooxygenase-2 (COX-2; NS-398, 1 μM). Collectively, these data suggest that nociceptive input-activated β-catenin signaling plays an important role in regulating the biosynthesis and release of substance P, which may contribute to the inflammation responses related to chronic pain.

Liu-Shen-Wan (LSW), an ancient preparation used to treat localized infection with pain, was recently reported to possess anticancer activity. The mechanism responsible for LSW's analgesic and anticancer activity is unclear. In the present study, we obtained a LSW supernatant (LSWS) fraction from ultrasoundassisted ethanol extraction (yield 15.9%) which proved to be safer than LSW in terms of hepatotoxicity. The LSWS (1 and 10 µg/mL) exhibited a potent inhibitory effect on the bradykinin-evoked rapid release of substance P from dorsal root ganglion (DRG) cells. At concentrations of 0.1 µg/mL and higher, the LSWS resulted in a concentration-related growth inhibitory effect on HepG2, a representative cancer cell lines. The LSWS significantly down-regulated the neurokinin-1 (NK-1) receptor expression in both HepG2 and bradykinin-treated DRG cells. In addition to the NK-1 receptor-dependent growth inhibition in HepG2 cells (0.1-100 µg/mL), the LSWS induced mitochondria-mediated apoptosis at a higher concentration (1-100 µg/mL). In conclusion, we recently isolated a safer LSW fraction which maintained its analgesic and anticancer activity, and found that the substance P/NK-1 receptor system was partly responsible for these effects. Our findings will be useful for developing more effective and less toxic LSW preparations.Key words Liu-Shen-Wan; anticancer; analgesic; substance P; neurokinin-1 receptor Liu-Shen-Wan (LSW), a well-known heat clearing and detoxifying herbal preparation, is mainly composed of Calculus Bovis Cowherb Seed, Margarita Pecazine, Venenum Bufonis Toborinone, Broneolum Syntheticum Boro-Scopol, Moschus Muskone, and Realgar Reapam. This ancient prescription is effective for treating localized infections and inflammationassociated pain.1) LSW's analgesic effect has been known and used for more than 200 years.2) During the past 200 years, many other applications have been uncovered, including its use for the treatment of diphtheria, scarlet fever, pharyngotonsillitis, acute tonsillitis, purulent parotitis, encephalitis B and viral pneumonia.3) It was also recently reported that LSW had a potential anticancer activity. 2,3)Despite its approval by the China Food and Drug Administration and its use for nearly 200 years, traditional LSW is associated with some toxicity because it contains realgar and Venenum Bufonis, 4) both of which were found to be toxic if administered alone.5) This toxicity made LSW unacceptable to the international community as a therapeutic agent.6) In the present study, we isolated an ethanol fraction of LSW which maintained its analgesic and anticancer activities, but with a decreased level of toxicity. However, to the best of our knowledge, the mechanism(s) underlying LSW's analgesic and anticancer activity are still unclear.Because LSW has both analgesic and anticancer activities, we suppose that there might be some common mechanisms underlying these two activities. Substance P and its endogenous receptor, the neurokinin-1 (NK-1) receptor, 7) which play important roles in both pa...