BackgroundIn chemotherapy, the full round of treatment must be completed as scheduled to achieve the strongest therapeutic effect. However, peripheral neuropathy, a severe side effect of the chemotherapeutic agent paclitaxel, can force the premature discontinuation of treatment. As some kampo practitioners have suggested that it may be possible to counteract such side effects, we analyzed the effects of Kamishoyosan, Shakuyakukanzoto, and Goshajinkigan in an in vitro model of paclitaxel-induced peripheral neuropathy.MethodsPaclitaxel-treated PC12 cells were assessed for neurite length and performed Western blot analysis for growth-associated protein-43 (GAP-43) and light neurofilament protein (NF-L) levels in the presence of nerve growth factor (NGF); they were re-assessed, with additional testing for acetylcholinesterase levels, after application of one of the kampo. We also compared phosphorylation of extracellular signal-regulated kinase (Erk)1/2 and Akt via Western blot analysis. About effect of kampo to anticancer efficacy, we confirmed cell cytotoxicity in A549 cells using MTT assay.ResultsAddition of Kamishoyosan or Shakuyakukanzoto, but not Goshajinkigan, significantly improved neurite length and GAP-43 and NF-L levels from paclitaxel-treated PC12 cells, relative to those of only NGF-treated PC12 cells. The promoting effect of Kamishoyosan and Shakuyakukanzoto in neurite outgrowth is confirmed when NGF promoted neurite outgrowth, and it was inhibited partially when Erk1/2 and Akt were blocked by Erk1/2 inhibitor or Akt inhibitor alone. Furthermore, neurite outgrowth induced by TJ24 and TJ68 was inhibited more strongly when Erk1/2 inhibitor and Akt inhibitor were treated at the same time. NGF with Kamishoyosan or Shakuyakukanzoto promoted the proportion of phosphorylated Erk1/2 and phosphorylated Akt compare with NGF only. On the other hand, Kamishoyosan or Shakuyakukanzoto didn’t influence cytotoxicity of paclitaxel in A549 cells.ConclusionsKamishoyosan or Shakuyakukanzoto promotes neurite outgrowth with NGF via increasing the proportion of phosphorylated Erk1/2 and phosphorylated Akt in PC12 cells. The effect applies to recovery from paclitaxel-induced axonal involvement and might promote recovery from paclitaxel-induced neuropathy without influence of anticancer effect of paclitaxel.

Aim Vinorelbine is an anticancer drug associated with vascular injury. The mechanism of vascular injury is associated with the decrease of endothelial nitric oxide synthase (eNOS), which regulates oxidative stress. Kakkonto, a traditional Japanese medicine, has anti‐inflammatory and anti‐oxidative effects. We evaluated the effect of kakkonto on vinorelbine‐induced vascular injury and its mechanism of action. Methods Human umbilical vein endothelial cells were used as the endothelial model. We compared vinorelbine‐induced cytotoxicity after pre‐incubation with the seven kakkonto components using ATP assay. Mitochondrial membrane potential was assessed using Mitochondrial Membrane Potential Assay Kit. Intracellular calcium ion concentration ([Ca2+]i) was measured with a calcium‐sensitive dye, Fluo‐4 AM. eNOS and Akt phosphorylation after ephedra extract or vinorelbine treatment were investigated on western blot. Results Ephedra, a kakkonto component, protected cells against vinorelbine‐induced cytotoxicity. Ephedra extract preserved eNOS phosphorylation by increasing [Ca2+]i, but it had no effect on Akt phosphorylation. The increase in [Ca2+]i was mediated by Ca2+ influx via l‐type calcium channels. Additionally, inhibition of Ca2+ influx or eNOS phosphorylation abolished the effects of ephedra extract. Conclusion Ephedra extract preserved eNOS phosphorylation following vinorelbine‐induced oxidative stress, thereby protecting cells from vinorelbine‐induced cytotoxicity. Moreover, the protective effect of ephedra extract is dependent on the increase of [Ca2+]i and the Ca2+/calmodulin‐dependent signal transduction pathway.