Geraniol and simvastatin show a synergistic effect on a human hepatocarcinoma cell line.

Simvastatin is a competitive inhibitor of 3-hydroxymethylglutaryl coenzyme A reductase activity, whereas geraniol is a monoterpene with multiple pharmacologic effects on mevalonate metabolism. Both of them inhibit growth and proliferation of many cell lines. The present study was designed to determine the action of geraniol, in combination with simvastatin, by assessing their effects in vitro on human hepatocarcinoma cell line (Hep G2). The treatment of Hep G2 cells with concentrations of simvastatin or geraniol that did not inhibit cell proliferation (5 µmol·l⁻¹ of simvastatin and 50 µmol·l⁻¹ of geraniol) resulted in a significant inhibition of cell proliferation. We also examined the effect of simvastatin, geraniol and the combination of both on the biosynthesis of lipids from [¹4C]-acetate. Our results demonstrate that the combination of simvastatin and geraniol synergistically inhibited cholesterol biosynthesis and proliferation of Hep G2 cell line, contributing to a better understanding of the action of a component of essential oils targeting a complex metabolic pathway, which would improve the use of drugs or their combination in the fight against cancer and/or cardiovascular diseases.

Effect of simvastatin on the uptake and metabolic conversion of palmitic, dihomo-gamma-linoleic and alpha-linolenic acids in A549 cells.

It is well known that simvastatin affects cholesterol synthesis. Furthermore it inhibits growth and proliferation and perturbs fatty acid metabolism in some cell lines. We have studied the effects of simvastatin on the uptake and metabolism of exogenous fatty acid in the human lung adenocarcinoma A549 cells. Simvastatin inhibited the proliferation of A549, and caused an increment in phospholipid/cholesterol ratio due to an increment in phospholipid content without affecting cholesterol content. All the fatty acids were uptaken and metabolized in both control and treated cells. The conversion of palmitic, linoleic and dihomo-gamma-linoleic acids to their metabolites and products/precursor ratios for the desaturation and elongation reactions showed that simvastatin enhanced the Delta5 desaturation step and altered some elongating steps. The machinery for unsaturated fatty acid synthesis in A549 is quite sensitive to simvastatin and its effects could have important implication taking into account that highly unsaturated fatty acids are involved in the regulation of diverse cellular functions by themselves or through their metabolites.

Simvastatin effects on a human lung carcinoma and cholesterol homeostasis of host and non-host mice.

In order to investigate the effect of a competitive inhibitor of the HMG-CoA reductase on tumor growth and cholesterol homeostasis of host and non-host mice, we maintained a human lung mucoepidermoid carcinoma (HLMC) in nude mice, treating these animals with Simvastatin for 33 days. The drug increased the total activity of hepatic HMG-CoA reductase without affecting the cholesterolemia. Non-treated host animals presented lower serum, tissue and microsomal hepatic cholesterol than non-host animals. These differences disappeared when animals were treated with Simvastatin, though the induction of the reductase activity at mid-dark was higher in non-host than in host animals. Simvastatin produced no significant effects on both final tumor volume and body weight. Synthesis and cholesterol homeostasis restoration induced by liver and tumoral reductase would account for no effect on the HLMC growth after a long treatment with Simvastatin.

Uptake and metabolic conversion of saturated and unsaturated fatty acids in Hep2 human larynx tumor cells.

Research on fatty acid metabolism in cultured human larynx tumor cells Hep2 was carried out. The cells were incubated with either a saturated (palmitic) or a polyunsaturated (linoleic, alpha-linolenic and eicosatrienoic (n-6)) radioactive fatty acid (0.66 pM, 24 h). The best incorporation capacity was observed in the linoleic acid followed by alpha-linolenic, palmitic and eicosatrienoic acids. All fatty acids tested were anabolized to higher derivatives within their own family. Palmitic acid was primarily monodesaturated rather than elongated, proving to have a very active A9 desaturase activity. With respect to polyunsaturated acid metabolism, the conversion of alpha-linolenic acid to higher homologs, although better than linoleic acid, occurred far less efficiently than that observed in other non-highly undifferentiated human tumor cells. This impairment in higher polyunsaturated fatty acid biosynthesis, reflected in the low levels of arachidonic acid in the fatty acid composition, would not reside in the A5 desaturation step since Hep2 cells can readily convert eicosatrienoic acid into arachidonic acid. Considering the potential regulatory role of specific polyunsaturated fatty acids in the cell proliferative control, the knowledge of the metabolism of fatty acids in this human tumor cell would be important for designing future experiments in order to clarify the mechanism involved in balance, proliferation and cell death.

3-Hydroxy-3-methylglutaryl coenzyme a reductase activity in liver of athymic mice with or without an implanted human carcinoma.

Hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase activities and cholesterol content in the liver of athymic mice either bearing or not an implanted human lung mucoepidormoid carcinoma (HLMC) and in the neoplasic tissue, were analyzed. The properties of the HMG-CoA reductase of HLMC grown in nude mice and those ones found in the liver of these animals, sacrificed either at mid-light or mid-dark, were similar. The hepatic reductase activity was found to be four- to five-fold greater at mid-dark than at mid-light (462 +/- 141 vs. 123 +/- 22 pmol min-1 mg protein-1). Since the Km value was not modified, the mid-dark activity could be due to an increase in the amount of enzyme. In contrast, HLMC reductase activity and cholesterol content showed similar values at mid-light and mid-dark points. HLMC reductase does not appear to have any diurnal variation and the cholesterol synthesis and content seems to be independent of food intake. HLMC-bearing nude mice undergo several alterations in the biosynthesis and homeostasis of cholesterol. Hypocholesterolemia, lower hepatic cholesterol content and higher HMG-CoA reductase activity are characteristic of host mice.

Immunohistopathological characterizatin of spontaneous metastases in a human lung mucoepidermoid adenocarcinoma (HLMC) xenograft.

The most common clinical form of lung cancer is a disseminated disease with distant metastases; several years of cancer progression precede presentation, and this ultimately limits the efficacy of curative therapy. In this immunohistochemical study, we examined a mucinous adenocarcinoma cell line, maintained by xenogeneic transplantation, and a spontaneous metastatic variant which produces distant tumors (in liver, spleen and kidney). The aim was to investigate possible parameters which characterize the metastatic process. Histopathological comparison between the two subcutaneous transplanted tumor lines showed that both lines presented a similar cellular morphology, a different pattern of cellular growth and an increased vascularization in the metastatic line with respect to its parent. All the tumor sections expressed differential immune reactivity with monoclonal antibodies against Lewis y (MAb C14), sialyl-Lewis x (MAb SNH3) and Lewis x (MAb FH2) determinants. Neither expressed MUC 1 mucins detectable with monoclonal antibodies reactive with the mucin protein core (MAbs C595 and SM3) nor was carcinoembryonic antigen (MAb C365) expressed. Neoplastic cells were reactive with an anti-pan cytokeratin monoclonal antibody confirming their epithelial histogenesis. Our findings have been evaluated with respect to defining metastatic phenotypes in lung cancer by examination of distinct histopathological and immunological parameters.

Effects of lovastatin on the fatty acid desaturation in a human lung mucoepidermoid carcinoma grown in nude mice.

The effect of lovastatin, a hypocholesterolemic drug, on tumor growth and desaturase activity was studied in a human lung mucoepidermoid carcinoma (HLMC) grown in nude mice. After administration of a diet supplemented with 25 mg% (w/w) lovastatin for 30 days the growth of HLMC was not inhibited. Liver and tumor phospholipid/cholesterol ratio was increased in lovastatin group but serum cholesterol was unaffected. Treatment with lovastatin increased delta 5 and delta 9 desaturation in tumor microsomes, whereas delta 6 desaturation did not change in tumors of treated mice. The changes were not reflected in the fatty acid composition of total tumor lipids.

[Effect of dietary supplementation with gamma-linolenic acid on the growth of a human lung carcinoma implanted in nude mice]. / Efecto de la suplementación dietética con acido gama-linolenico sobre el crecimiento de un carcinoma de pulmón humano implantado en ratones atímicos.

We have studied the effect of a gamma-linolenic acid (18:3 n-6, GLA)-supplemented diet on the growth of a human lung mucoepidermoid carcinoma (HLMC) implanted in athymic mice and on its uptake of human low density lipoproteins labeled with 99mTc (99mTc-LDL). Mice bearing the HLMC were divided into two experimental groups. One of them was administered a control diet (C diet) and the other one was given a diet supplemented with 25 mg GLA/g pellet (GLA diet) for three weeks (Table 1). A tumor growth inhibition with the GLA diet was evident at the second week of treatment, and a marked inhibition (56%) was reached at the end of the third week (Fig. 1). The GLA diet produced some changes in the total fatty acid composition of tumor, plasma and liver of host mice: GLA and arachidonic acid (20:4 n-6, AA) induced significant increases, whereas oleic (18:1 n-9, OA) and linoleic acids (18:2 n-6, LA) were decreased (Table 2). Tumors of those animals fed both diets were labeled by 99mTc-LDL, and no difference was observed in the ratio of tumor/liver and tumor/kidney uptake of host animal (Table 3). Results obtained using this experimental model suggest that the inhibitory effect of GLA on tumor growth is not related to the LDL tumor uptake.

Efecto de la suplementación dietética con acido gama-linolenico sobre el crecimiento de un carcinoma de pulmón humano implantado en ratones atímicos. / [Effect of dietary supplementation with gamma-linolenic acid on the growth of a human lung carcinoma implanted in nude mice]

We have studied the effect of a gamma-linolenic acid (18:3 n-6, GLA)-supplemented diet on the growth of a human lung mucoepidermoid carcinoma (HLMC) implanted in athymic mice and on its uptake of human low density lipoproteins labeled with 99mTc (99mTc-LDL). Mice bearing the HLMC were divided into two experimental groups. One of them was administered a control diet (C diet) and the other one was given a diet supplemented with 25 mg GLA/g pellet (GLA diet) for three weeks (Table 1). A tumor growth inhibition with the GLA diet was evident at the second week of treatment, and a marked inhibition (56

) was reached at the end of the third week (Fig. 1). The GLA diet produced some changes in the total fatty acid composition of tumor, plasma and liver of host mice: GLA and arachidonic acid (20:4 n-6, AA) induced significant increases, whereas oleic (18:1 n-9, OA) and linoleic acids (18:2 n-6, LA) were decreased (Table 2). Tumors of those animals fed both diets were labeled by 99mTc-LDL, and no difference was observed in the ratio of tumor/liver and tumor/kidney uptake of host animal (Table 3). Results obtained using this experimental model suggest that the inhibitory effect of GLA on tumor growth is not related to the LDL tumor uptake.

Effects of an eicosapentaenoic and docosahexaenoic acid concentrate on a human lung carcinoma grown in nude mice.

The effects of the n-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on the growth of a human lung mucoepidermoid carcinoma (HLMC) in athymic mice were studied. The mice were divided into three groups which were given either a control chow diet (C), a chow diet supplemented with EPA/DHA (P) (25 or 50 mg of free n-3 fatty acids/g of pellet/day), or chow diet supplemented with palmitic acid (S) (isocaloric with P). Two independent experimental schedules were followed: i) host mice bearing either tumors that were allowed to reach 4000 mm3, or only 35 mm3, were fed C, P or S for 21 or 41 days; ii) animals were fed C, P and S for 9 days before tumor implant and were maintained on these diets throughout tumor growth. Food consumption, mouse weight and liver/body weight ratio showed no significant differences between supplemented diets and chow. Tumor growth was markedly inhibited (45%) in both experiments by the EPA/DHA supplemented diet. In Experiment 2, only 60% of mice fed diet P had tumors. The fatty acid composition of neutral and polar lipids of host liver and tumor reflected the dietary intake of n-3 fatty acids; the content of arachidonic acid was reduced by 50%, and EPA/DHA was increased 3- to 5-fold. Tumor prostaglandin E2 levels were reduced 7.4-fold in the P group. The reduced PGE2 content may be a factor in tumor growth inhibition.

In vitro effect of eicosapentaenoic and docosahexaenoic acids on prostaglandin E2 synthesis in a human lung carcinoma.

The capacity to synthesize both prostaglandins E1 (PGE1) and E2 (PGE2) has been determined in human lung mucoepidermoid carcinoma homogenates when [14C]-fatty acid precursors were added to the incubation medium. Only 10% of the total radioactivity recovered in PGs was found in PGF1 alpha and PGF2 alpha. The experiments were principally focused to inhibit the PGE2 synthesis either with pure eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids or with mixtures of both n-3 fatty acids obtained from fish oil. The results demonstrated that significant inhibitions were found when using 25 microM or a higher concentration of pure EPA or DHA in the incubation medium; however, 5 microM of mixtures of different EPA/DHA ratio caused the same inhibition. The results suggest that EPA and DHA, when added together, may enforce their inhibitory effect on PGE2 synthesis.

Changes in the plasma free fatty acid composition during VLDL lipolysis induced by heparin: their effect on platelet phosphatidylcholine biosynthesis.

The contribution of the acyl specificity of post heparin lipolytic enzymes to changes in the plasma free fatty acid (FFA) pattern during very low density lipoproteins (VLDL) lipolysis, was studied. The effect of these changes in the 3H-choline incorporation into platelet phosphatidylcholine (PC) during incubation, was also investigated. "In vitro" experiments with post heparin plasma and isolated VLDL showed an increase of 61.1 in the percentage of linoleic acid (C 18:2) concomitant with a decrease of 57.1% in the relative concentration of stearic acid (C 18:0) during VLDL triacylglycerol lipolysis. Similarly, a decrease of 39.2% in the plasma concentration of C 18:0 and an increase of 29.4% in the C 18:2 and 9.6% in the oleic acid (C 18:1) concentration were observed after intravenous injection of low doses of heparin. The plasma FFA compositional changes that occur during "in vitro" VLDL lipolysis accelerate not only the incorporation of labeled choline but also the incorporation of C 18:2 into platelet PC. It is suggested that the above observed changes on platelet response are due to the increase in the C 18:2 concentration during incubation.