The effect of the diterpene 5-epi-icetexone on the cell cycle of Trypanosoma cruzi

Numerous natural compounds have been used against Trypanosoma cruzi, the causative agent of Chagas' disease. Here, we studied the effect of the diterpene 5-epi-icetexone on growth and morphology of parasites synchronized with hydroxyurea, at different periods of time after removal of the nucleotide. We observed that the diterpene does not affect the growth of the parasites when added within 10 h after removal of hydroxyurea, but the compound was effective on growth when added to the cultures after 12 h. Thymidine incorporation was somewhat inhibited when the diterpene was added at 12 h after removal of hydroxyurea, possibly on the transition S/G2. Using transmission electron microscopy we observed that the diterpene induced a delay in the progression of cell division. We conclude that the compound, at cytostatic dose, affects the cell cycle of T. cruzi, possibly in the transition S/G2 phase and cell division. Further studies will focus to identify the molecular targets for the action of 5-epi-icetexone.     

Sesquiterpene lactones and the diterpene 5-epi-icetexone affect the intracellular and extracellular stages of Trypanosoma cruzi

Chagas disease is a major health problem in Latin America and is caused by the parasitic protozoan Trypanosoma cruzi. Although many drugs have been used to alleviate the disease, these have been ineffective in the chronic phase and have also presented numerous side effects on patients. In this study we tested the effect of three sesquiterpene lactones (dehydroleucodine, helenalin and mexicanin) and a diterpene (5-epi-icetexone) on parasites (Y-strain) grown in host cells. At 48h of treatment, the number of amastigotes inside the cells was lower than in the controls. This effect was observable at concentrations of 1.5-3.8μM, which are of low cytotoxicity to host cells. In addition, the compounds caused a decrease in the percentage of infected cells. The treatments also reduced the presence of trypomastigotes in the extracellular medium. In all cases, helenalin was the most potent. The number of parasites per cell at 24h indicates the occurrence of multiple infection, which would also be affected by the compounds. However, we should not discard an effect on the proliferation and survival of parasites within the host cells. On the other hand, an additional effect on the differentiation of parasites and/or the survival of extracellular trypomastigotes might be possible. We conclude that these compounds are very effective against T. cruzi possibly by multiple mechanisms.     

Insulin-sensitizing activities of tanshinones,diterpene compounds of the root of Salvia miltiorrhiza Bunge

In this study, the effects of the extract and four tanshinone compounds from the dried root of Salvia miltiorrhiza Bunge (Labiatae) on the tyrosine phosphorylation of the insulin receptor (IR) β-subunit and the downstream signaling were examined in Chinese-hamster ovary cells expressing human insulin receptors (CHO/IR cells) as well as in 3T3-L1 adipocytes. In addition the translocation of the glucose transporter 4 was investigated in 3T3-L1 adipocytes. Total extract of Danshen (1–10 μg/ml) and the four tanshinones (10 μM) did not show any activity, but the total extract and the tanshinone I, IIA and 15, 16-dihydrotanshinone I except cryptotanshinone enhanced the activity of insulin (1 nM) on the tyrosine phosphorylation of the IR as well as the activation of the downstream kinases Akt, ERK1/2, and GSK3β. In the adipocytes the same IR-downstream signaling and the translocation of glucose transporter 4 were demonstrated by the three tanshinones in the presence of insulin. These insulin-sensitizing activities of tanshinones may be useful for developing a new class of specific IR activators as anti-diabetic agents.     

Isopimarane diterpene glycosides, apoptosis inducers, obtained from fruiting bodies of the ascomycete Xylaria polymorpha

The methanol extract of fruiting bodies of the ascomycete Xylaria polymorpha afforded three isopimarane diterpene glycosides, namely, 16-α-d-mannopyranosyloxyisopimar-7-en-19-oic acid (1), 15-hydroxy-16-α-d-mannopyranosyloxyisopimar-7-en-19-oic acid (2), and 16-α-d-glucopyranosyloxyisopimar-7-en-19-oic acid (3). Their structures were determined by spectroscopic methods and by single-crystal X-ray analysis. They showed cytotoxicity against human cancer cell lines and exhibited IC₅₀ values ranging from 71 to 607 μM. Further studies on the cytotoxicity of these compounds against HL60 cells demonstrated that they induced apoptosis along with typical DNA fragmentation. It was observed that 2 was less active than 1 and 3.     

The trypanocidal effect of sesquiterpene lactones helenalin and mexicanin on cultured epimastigotes

Sesquiterpene lactones constitute a large group of biologically active compounds obtained from plants. The lactones, mexicanin (MXN) and helenalin (HLN), were reported recently as active against the infective form of Trypanosoma cruzi. In this work, we studied the effects of these compounds on the growth and viability of the noninfective epimastigote, to compare the sensitivity of the 2 stages and to characterize their actions. Both compounds were cytotoxic to the parasites, with HLN (inhibitory concentration 50% [IC50] 1.9 +/- 0.08 microM) more potent than MXN (IC50 3.8 +/- 0.19 microM) and the typanocidal drug, benznidazole (IC50 8.6 +/- 2.5 microM). The results showed that epimastigotes are less sensitive than trypomastigotes to the compounds. The trypanocidal effect of these lactones, irreversible after 12-hr exposure, was not reversed by the reducing agents dithiotreitol or beta-mercaptoethanol. Ultrastructurally, we observed cytoplasmic vacuolization and nuclear disorganization. Although concentrations between 0.5 and 1.5 microM of the drugs were not lethal to the parasites, epimastigotes became thinner and their nuclei became more pycnotic after exposure. We conclude that MXN and HLN are deleterious for T. cruzi epimastigotes and that their mechanism of action is different than that of the related lactone, dehydroleucodine.     

The absolute stereochemistry of a diterpene from Ballota aucheri

The semi-synthetic transformation of hispanolone, isolated from Ballota africana, into 6beta-hydroxy-15,16-epoxylabda-8,13(16),14-trien-7-one has established an ent-labdane absolute stereochemistry for a diterpene metabolite originally isolated from B. aucheri.     

Specificity of Mnt 'master residue' obtained from in vivo and in vitro selections

Mnt is a repressor from phage P22 that belongs to the ribbon–helix–helix family of DNA binding factors. Four amino acids from the N-terminus of the protein, Arg2, His6, Asn8 and Arg10, interact with the base pairs of the DNA to provide the sequence specificity. Raumann et al. (Nature Struct. Biol., 2, 1115–1122) identified position 6 as a ‘master residue’ that controls the specificity of the protein. Models for the interaction have residue 6 of Mnt interacting directly with position 5 of the operator. In vivo selections demonstrated that protein variants at residue 6 bound specifically to operator mutations at that position. Operators in which the wild-type G at position 5 was replaced by T specifically bound to several different protein variants, primarily hydrophobic residues. The obtained protein variants, plus some others, were used in in vitro selections to determine their preferred binding sites. The results showed that the residue at position 6 influenced the preference for binding site bases predominantly at position 5, but that the effects of altering it can extend over longer distances, consistent with its designation as a ‘master residue’. The similarities of binding sites for different residues do not correlate strongly with common measures of amino acid similarities.     

Salvia lavandulaefolia essential oil inhibits cholinesterase in vivo.

The essential oil of Salvia lavandulaefolia at two dosage levels was administered orally to rats for five days. Choline esterase activity was measured post mortem in three areas of the brain, both in the absence and presence of TEPP, a specific butylcholine esterase inhibitor, and was found to be significantly reduced in the striatum with both doses and also in the hippocampus at the higher dose. The activity of the enzyme in the cortex was not significantly reduced even at the higher dose. Thus it appears that S. lavandulaefolia oil, shown to inhibit choline esterase in vitro, also has an in vivo effect and this may help explain its traditional use for ailing memory.     

Possible mechanisms underlying the cholesterol-raising effect of the coffee diterpene cafestol

Cafestol, a coffee diterpene present in unfiltered coffee brews, potently raises serum lipids in humans. The mechanism through which this dietary compound influences liporotein metabolism is largely unknown. Unravelling the mechanism of action might lead to new insights into the regulation of serum cholesterol levels in humans. This review summaries ways in which cafestol may act on serum lipids.     

Protective effect of an acidic glycoprotein obtained from culture of Chlorella vulgaris against myelosuppression by 5-fluorouracil

 An acidic glycoprotein prepared from a culture of Chlorella vulgaris (CVS) was examined for its protective effect on 5-fluorouracil(5FU)-induced myelosuppression and indigenous infection in mice. Subcutaneous administration of CVS greatly reduced the mortality of non-tumor-bearing mice given a high dose of 5FU, and could increase the LD₅₀ value of 5FU for these mice. After 5FU treatment, indigenous infection developed probably as a result of the impairment of the host defense system. CVS reduced the incidence of indigenous infections and this effect was attributable to the acceleration of recovery from 5FU-induced myelosuppression. Early recovery of hematopoietic stem cells, or cells responding to interleukin-3 or granulocyte/macrophage-colony-stimulating factor, was especially observed in the bone marrow of CVS-treated mice on days 4 – 9 after the injection of 5FU. When tumor-bearing mice were given CVS during treatment with 5FU, CVS prolonged the survival of mice without affecting the antitumor activity of 5FU. In addition, CVS was itself shown to exert an antitumor effect. These results suggested that CVS may be beneficial for the alleviation of side-effects in cancer chemotherapy without affecting the antitumor activity of the chemotherapeutic agent. Received: 15 August 1995 / Accepted: 23 April 1996     

Inhibitory effect of the coffee diterpene kahweol on carrageenan-induced inflammation in rats

Previous studies reported that kahweol, a coffee-specific diterpene, inhibits cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression in cultured lipopolysaccharide-activated macrophages. The aim of this study was to confirm the anti-inflammatory effects of kahweol by examining its effect on the inflammatory response induced by carrageenan in a rat using an acute air pouch inflammation model. Kahweol significantly reduced the levels of the inflammatory process markers in the air pouch, such as the volume of exudates, the amount of protein and the number of leukocytes and neutrophils. The levels of nitrite, TNF-alpha and prostaglandin E2 (PGE2) were also markedly lower in the air pouch of the kahweol-treated animals than in the controls. Immunoblot analysis showed that kahweol reduced the COX-2 and iNOS expression level in the exudate cells. The histological examination showed that there was a lower inflammatory response in the pouch tissues from the kahweol-treated animals. In addition, kahweol significantly reduced the paw edema induced by carrageenan and also markedly reduced the level of PGE2 production in the inflamed paw. These results suggest that kahweol has significant anti-inflammatory effects in vivo, which might be due to the inhibition of iNOS and COX-2 expression in the inflammatory sites.     

The effect of the antiherpesvirus drug 5-methoxymethyl-2'-deoxyuridine on the humoral immune response in vivo.

5-Methoxymethyl-2'-deoxyuridine (MMUdR), a drug with potent antiviral activity in vitro against Herpes simplex virus, was investigated for its immunosuppressive effects. Doses as high as 2000mg/kg given daily for 9 days were not immunosupporessive as judged by the fact that treated animals produced normal immune responses to sheep erythrocytes, Brucella bacteria, and Herpes simplex virus.     

A diterpene quinone from the bark of Cryptomeria japonica

A diterpene, cryptoquinone, was isolated from the bark of Cryptomeria japonica, the structure, 7,11,14-trioxoabieta-8,12-diene, was established by spectral analyses and X-ray crystallography. This diterpene quinone showed moderate antifungal activities against Pyricularia orizae and Alternaria alternata, and cytotoxic activity against mouse lymphoid neoplasm (P388) cells with an IC(50) of 0.26 microg/ml.     

假地枫皮中二萜酸类化合物研究

从八角属植物假地枫皮的石油醚提取物中分离出5个二萜酸类化合物,经波谱数据分析(^1H NMR、^13C NMR、^1H-^1H COSY、NOESY、HSQC和HMBC),分别鉴定为4-epi-dehydroabietic acid(1)、4-epi-sandaraeopinaric acid(2)、4-epi-abietic acid(3)、4-epi-isopimaric acid(4)和8,11,13,15-abietatetraen-19-oic acid(4)。     

Structures of the novel diterpene glycosides from Stevia rebaudiana

From the commercial extract of the leaves of Stevia rebaudiana, two new diterpenoid glycosides were isolated besides the known steviol glycosides including stevioside, rebaudiosides A–F, rubusoside, and dulcoside A. The structures of the two new compounds were identified as 13-[(2-O-6-deoxy-β-d-glucopyranosyl-β-d-glucopyranosyl)oxy] ent-kaur-16-en-19-oic acid β-d-glucopyranosyl ester (1), and 13-[(2-O-6-deoxy-β-d-glucopyranosyl-3-O-β-d-glucopyranosyl-β-d-glucopyranosyl)oxy] ent-kaur-16-en-19-oic acid β-d-glucopyranosyl ester (2), on the basis of extensive NMR and MS spectral data as well as chemical studies.     

Nature of the trypanocidal factor in human serum

The chemical nature of the trypanocidal factor in human serum was investigated. The results show that although the trypanocidal factor is contained within the high density lipoprotein (HDL) fraction of human serum, it is apparently not one of the major apolipoproteins of the HDL complex such as apolipoprotein A-I, A-II, or apolipoprotein B. The factor would appear to be a minor component of the HDL fraction whose chemical nature is still uncertain.