ABSTRACT
Taxus suspension cell culture has the potential to provide a sustainable source of anticancer drug paclitaxel (Taxol) and other taxoids. In the cell culture of Taxus chinensis, Taxuyunnanine C (Tc) is the primary taxoid. To design a rational strategy for redirecting the precursor fluxes from other taxoids into paclitaxel production, we employed Real-time Quantitative PCR (RQ-PCR) to understand the dynamic profiling of key biosynthetic pathway genes of palcitaxel and taxoids during the culture process. Six genes (TASY, TDAT, T5alphaH, TalphaH, T10betaH and T14betaH) were quantified under the process condition of double elicitation by 2,3-dihydroxylpropanyl jasmonate (DHPJA) (100 micromol/L on day 7 and day 12), and sucrose feeding (20 g/L) on day 7. This process treatment led to a high accumulation of Tc at (554.46 +/- 21.28) mg/L 8 days after the first elicitation. Then 9 days after the second elicitation, Tc production was as high as (997.72 +/- 1.51) mg/L. The early pathway genes TASY and TDAT were significantly up-regulated by 182-fold and 98-fold, respectively for the first DHPJA elicitation and by 208-fold and 131-fold, respectively for the second elicitation. The induction occurred after each elicitation lasted for about 24 h before their abundances decreased. Things are somewhat different in the case of the other four genes T5alphaH, TalphaH, T10betaH and T14betaH. For gene TalphaH, it was highly up-regulated by 3061-fold for the first DHPJA elicitation and by 1016-fold for the second elicitation. For the other three genes T5alphaH, T10betaH, T14betaH, they were up-regulated by 13-fold, 38-fold and 20-fold, respectively for the first DHPJA elicitation and by 7-fold, 16-fold and 6-fold, respectively for the second elicitation. The RQ-PCR results showed that there is tight correlation between gene expression and Tc accumulation. Gene expression was in accordance with Tc yield. Elicitation could improve expression of six genes. While along with culture course, high expression of the genes weakened. Elicitation for the second time would promote high expression of the genes again.
Subject(s)
Cell Culture Techniques , Culture Media , Cyclopentanes , Pharmacology , Gene Expression Regulation, Plant , Oxylipins , Pharmacology , Plant Growth Regulators , Pharmacology , Sucrose , Pharmacology , Taxoids , Metabolism , Taxus , Genetics , Metabolism , TranscriptomeABSTRACT
Objective To investigate if Chondrus yendoi decoction(CYD) and polysaccharide(CYP) have the effects of scavenging superoxide anion(O?—2),antioxidation,protecting mitochondria from damage.Methods Lipid peroxidation and swelling of mouse liver and brain mitochondria were induced by Fe2+-L-Cys system in vitro.Superoxide anion was generated by reduced nicotinamide adenine dinucleotide(NADH)-N-methylphenazonium methyl sulfate(PMS)-4-nitrobluetetrazolium chloride(NBT) system.Lipid peroxide malondialdehyde(MDA),mitochondrial swelling and superoxide anion formation were determined by spectrophotometry.Results CYD and CYP could inhibit MDA production,protect mitochondria from swelling,and scavenge O?-2 obviously in a dose-dependent manner.Conclusion CYD and CYP had the effects of scavenging O?-2,antioxidation,protecting mitochondria from injury.CYP was the major bioactive component.
ABSTRACT
AIM:To investigate protective effect of ethanol extract of Forsythia suspensa flower(FFE)on mitochondrial injury induced by free radicals,antioxidation and superoxide anion(O ?-__2)scavenging activities and anti-aging mechanism.METHODS:Thiobarbituric acid(TBA)colorimetric method was used to determine malondialdehyde(MDA).Spectrometric method was used to measure the swelling of mitochondria and activities of scavenging O ?-__2 which was produced by reduced nicotinamide adenine dinucleotide(NADH)-4-Nitrobluetetrazolium chloride(NBT)-N-Methylphenazonium methyl sulfate(PMS)system.The neck back of mice were injected subcutaneously with D-galactose to induce aging model at a dose of 100 mg/(kg?d)for 7 weeks.The effects of FFE on the activities of catlase(CAT),surperoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)and anti-hydroxyl radical were assayed by ammonium molybdate colorimetry,xanthine oxidase method,dithiodinitrobenzoic acid colorimetry and Fenton reaction colorimetry in vivo respectively.RESULTS:FFE could inhibit mitochondrial injury and swelling induced by Fe 2+-L-Cys in a concentration-dependent manner and also had significant O ?-__2 scavenging effect.Moreover,the activities of CAT,SOD,GSH-Px and anti-hydroxyl radical in mice liver homogenate were increased significantly by FFE.CONCLUSION:FFE protects mitochondria by scavenging reactive oxygen species,and increasing the activities of antioxidase.So FFE has the pharmaceutical values of antioxidant and anti-aging.