Screening and identification of antagonistic actinomycete LA-5 against Botrytis cinerea.

In this study, actinomycetes were isolated from rhizosphere soil of tomato by spread plate method, with the pathogen Botrytis cinerea as the target fungus. An actinomycete isolate LA-5 with strong antifungal activity was obtained by confrontation culture and Oxford cup double screening method. According to cultural characteristics, physiological and biochemical properties and 16S rDNA analysis, the strain LA-5 was primarily identified as Streptomyces chungwhensis. The second screening results showed that the fermentation filtrate of LA-5 could substantially inhibit the spore germination and mycelium growth of B. cinerea. Both conidium germination and mycelia growth could be inhibited by 100 times fermentation broth of LA-5, with the inhibiting rate being higher than 50%. Furthermore, the inhibited colony showed white mycelia, with sparse and thin aerial hypha, and the obviously decreased branches. Results from in vitro biocontrol experiment showed that control efficiency of filtrate from LA-5 fermentation liquid on B. cinerea could be up to 83.4%, indicating that the strain was a potential antagonist against tomato gray mold.

Investigation of the active components in Tripterygium wilfordii leading to its acute hepatotoxicty and nephrotoxicity.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional herbal medicine Tripterygium wilfordii Hook. f. (TW) has been widely used for the treatment of rheumatoid arthritis and autoimmune disease in the clinic. However, adverse reactions of TW including hepatotoxicity and nephrotoxicity have been frequently reported. Terpenes and alkaloids are among the most important active components in TW. Triptolide (TP), a major terpene in TW, has been found to induce toxicity, and metabolic pathways could lead to detoxification of TP. In this study, whether other major terpenes or alkaloids in TW contribute to its toxicity was investigated. The role of metabolic eliminations in their potential detoxification process was also evaluated. MATERIALS AND METHODS: The toxicity of TW and its five major active components (one terpene and four alkaloids) in mice was evaluated in terms of mortality and blood biochemical levels (ALT, AST, BUN and CREA). TP was used as a positive control. Metabolic pathways leading to potential detoxification of TW or its two representative components (triptonide and wilforgine) were evaluated in glutathione (GSH)-depleted (treated with L-buthionine-S,R-sulfoxinine, BSO) and aminobenzotriazole (ABT; a nonspecific inhibitor for P450s)-treated mice. RESULTS: In normal mice, the major metabolic pathways for the terpene compounds TP and triptonide (TN) were hydroxylation and cysteine conjugation, and the alkaloid wilforgine (WG) mainly underwent oxidative metabolism and hydrolysis. In ABT/BSO-treated mice, the hydroxylated metabolites of TP, TN and WG were found at a lower level than normal mice, and the level of cysteine conjugates of TN increased probably due to the stress response. Compared with normal mice, mortality and levels of ALT (but not BUN) were significantly higher (P<0.01) in TW (or TP)-treated mice (1.2 mg kg(-1)), indicating the acute toxicity (may not nephrotoxicity) of TW and its active component TP. Pretreatment with ABT and/or BSO increased the acute toxicity (including hepatotoxicity and nephrotoxicity) caused by TW or TP. No significant toxicity was found for TN or four alkaloids in normal mice or ABT/BSO-treated mice. CONCLUSIONS: TP was probably the main contributor to the toxicity of TW, and the terpene TN and alkaloids in TW may be of no toxicological concern at dosage levels up to 20-fold of the therapeutic dose. Metabolic eliminations to less reactive metabolites implied a high potential for detoxification of TW, and caution should be taken for TW clinical use during co-administration with other CYP inhibitors or GSH-depleting agents.

Effect of interrupted endogenous BMP/Smad signaling on growth and steroidogenesis of porcine granulosa cells.

Bone morphogenetic proteins (BMPs) play a critical role in the growth and steroidogenesis of granulosa cells (GCs). BMP signals act through membrane-bound heteromeric serine/threonine kinase receptors. Upon ligand binding, BMPs activate intracellular Smad proteins and regulate growth and apoptosis in various cell types. The objective of this study was to demonstrate the effects of BMP/Smad signal on growth and steroidogenesis of porcine GCs. A strategy of RNA interference (RNAi)-mediated 'gene silencing' of Smad4, a core molecule mediating the intracellular BMP/Smad signal transduction pathways, was used to interrupt endogenous BMP/Smad signaling. Results indicate that Smad4-small interfering RNA (siRNA) caused specific inhibition of Smad4 mRNA and protein expression after transfection. Interrupted endogenous BMP/Smad signaling significantly inhibited growth, and induced apoptosis of porcine GCs, while decreasing estradiol production. In addition, interrupted BMP/Smad signaling significantly (P<0.05) changed the expression of Cyclin D2, CDK4, Bcl-2, and Cyp19a1. These findings provide new insights into how BMP/Smad signaling regulates the growth and steroidogenesis of porcine GCs.