Protective effects of carotenoids from saffron on neuronal injury in vitro and in vivo.

Crocus sativus L. (saffron) has been used as a spice for flavoring and coloring food preparations, and in Chinese traditional medicine as an anodyne or tranquilizer. Our previous study demonstrated that crocin, a carotenoid pigment of saffron, can suppress the serum deprivation-induced death of PC12 cells by increasing glutathione (GSH) synthesis and thus inhibiting neutral sphingomyelinase (nSMase) activity and ceramide formation. The carotenoid pigments of saffron consist of crocetin di-(beta-d-glucosyl)-ester [dicrocin], crocetin-(beta-d-gentiobiosyl)-(beta-d-glucosyl)-ester [tricrocin] and crocetin-di-(beta-d-gentiobiosyl)-ester [crocin]. Saffron also contains picrocrocin, the substance causing saffron's bitter taste. In this study, to confirm whether neuroprotective effects of saffron are caused solely by crocin, we examined the antioxidant and GSH-synthetic activities of these crocins in PC12 cells under serum-free and hypoxic conditions. Measurements of cell viability, peroxidized membrane lipids and caspase-3 activity showed that the rank order of the neuroprotective potency at a concentration of 10 muM was crocin>tricrocin>dicrocin and picrocrocin (the latter two crocins had a little or no potency). In addition, we show that among these saffron's constituents, crocin most effectively promotes mRNA expression of gamma-glutamylcysteinyl synthase (gamma-GCS), which contributes to GSH synthesis as the rate-limiting enzyme, and that the carotenoid can significantly reduce infarcted areas caused by occlusion of the middle cerebral artery (MCA) in mice.

Crocin prevents the death of rat pheochromyctoma (PC-12) cells by its antioxidant effects stronger than those of alpha-tocopherol.

Crocin is a pharmacologically active component of Crocus sativus L. (saffron) used in traditional Chinese medicine. We report here the effects of crocin on neuronally differentiated pheochromocytoma (PC-12) cells deprived of serum/glucose. Depriving the PC-12 cells of serum/glucose caused peroxidation of their cell membrane lipids and decreased intercellular superoxide dismutase (SOD) activity. Treating the PC-12 cells with 10 microM crocin inhibited the formation of peroxidized lipids, partly restored the SOD activity, and maintained the neuron's morphology. These antioxidant effects of crocin were more effective than those of alpha-tocopherol at the same concentration. Crocin also suppressed the activation of caspase-8 caused by serum/glucose deprivation. These results together with our previous data suggest that crocin is a unique and potent antioxidant that combats oxidative stress in neurons.

Crocin prevents the death of PC-12 cells through sphingomyelinase-ceramide signaling by increasing glutathione synthesis.

Crocin is a pharmacologically active component of Crocus sativus L. (saffron) that has been used in traditional Chinese medicine. In a previous study, we demonstrated that crocin inhibits apoptosis in PC-12 cells by affecting the function of tumor necrosis factor-alpha. In this study, we found that depriving cultured PC-12 cells of serum/glucose causes a rapid increase in cellular ceramide levels, followed by an increase in the phosphorylation of c-jun kinase (JNK). The accumulation of ceramide was found to depend on the activation of magnesium-dependent neutral sphingomyelinase (N-SMase), but not on de novo synthesis. The serum/glucose-deprived PC-12 cells also decreased the cellular levels of glutathione (GSH), which is the potent inhibitor of N-SMase. Treating the PC-12 cells with crocin prevented N-SMase activation, ceramide production, and JNK phosphorylation. We also found that the chemical can enhance the activities of GSH reductase and gamma-glutamylcysteinyl synthase (gamma-GCS), contributing to a stable GSH supply that blocks the activation of N-SMase. Thus our data suggest that crocin combats the serum/glucose deprivation-induced ceramide formation in PC-12 cells by increasing GSH levels and prevents the activation of JNK pathway, which is reported to have a role of the signaling cascade downstream ceramide for neuronal cell death.

Optimizing the dosing schedule of TNP-470 [O-(chloroacetyl-carbamoyl) fumagillol] enhances its antitumor and antiangiogenic efficacies.

Many drugs vary in potency and/or toxicity according to the time of day when they are administered. In this study, we investigated whether antitumor efficacy of angiogenesis inhibitor, TNP-470 [O-(chloroacetyl-carbamoyl) fumagillol], could be improved by optimizing the dosing schedule. Tumor-bearing mice were housed under standardized light/dark cycle conditions (lights on at 7:00 AM, off at 7:00 PM) with food and water ad libitum. The antitumor effect of TNP-470 (30 mg/kg s.c.) was more potent in mice injected with the drug at the early light phase than it was when administered at the early dark phase. The diurnal change in the antitumor effect of TNP-470 was parallel to that in its antiangiogenic activity. The variation in the effects of TNP-470 was closely related to the diurnal variations in its inhibitory action on methionine aminopeptidase activity in tumor masses. There was a significant dosing time-dependent change in the concentration of TNP-470 in plasma. The higher concentration of TNP-470 in plasma was observed when its antitumor and antiangiogenic activities were increased. These results suggest that therapeutic efficacy of TNP-470 can be enhanced by choosing the most appropriate time of day to administer the drug.