Blazein of a new steroid isolated from Agaricus blazei Murrill (himematsutake) induces cell death and morphological change indicative of apoptotic chromatin condensation in human lung cancer LU99 and stomach cancer KATO III cells.

Blazein was isolated from mushroom (Agaricus blazei Murrill) and identified by Mass and 1H-NMR as blazein. The effect of blazein on the DNA of human various cancer cells was investigated. DNA fragmentations by blazein to oligonucreosomal-sized fragments, a characteristic of apoptosis, were observed in the human lung LU99 and stomach KATO III cancer cells. The DNA fragmentations by blazein were observed from day 2 (KATO III cells) or day 3 (LU99 cells) after the addition of blazein to the culture cells. These findings suggest that growth inhibition by blazein results from the induction of apoptosis by the compound.

Induction of apoptosis by rhapontin having stilbene moiety, a component of rhubarb (Rheum officinale Baillon) in human stomach cancer KATO III cells.

We have investigated the effects of rhapontin on proliferation and DNA of human stomach cancer KATO III cells. Growth inhibition and induction of apoptosis by rhapontin were observed in the KATO III cells. Morphological change showing apoptotic bodies was observed in the KATO III cells treated with rhapontin. The fragmentation of DNA by rhapontin to oligonucleosomal-sized fragments that is a characteristic of apoptosis was observed to be concentration- and time-dependent in the KATO III cells. N-acetyl-L-cysteine, an antioxidant, suppressed the DNA fragmentation caused by rhapontin. On the other hand, it was found that resveratrol having stilbene moiety as well as rhapontin induced apoptosis in the KATO III cells. So, it is considered that stilbene moiety in the molecule is essential for the induction of apoptosis. The data of the present study show that the suppression of KATO III cell-growth by rhapontin results from the induction of apoptosis by the compound, and that active oxygen is involved in the inductions of apoptosis caused by rhapontin in the KATO III cells.

Glycyrrhetic acid (a metabolic substance and aglycon of glycyrrhizin) induces apoptosis in human hepatoma, promyelotic leukemia and stomach cancer cells.

We have investigated the effect of glycyrrhetic acid (GR) which is metabolic substance of glycyrrhizin, on DNA of human hepatoma (HLE), promyelotic leukemia (HL-60) and stomach cancer (KATO III) cells. GR displayed apoptotic effects against HLE, HL-60 and KATO III cells. The fragmentation of DNA by GR to oligonucleosomal-sized fragments, a characteristic of apoptosis, was dose- and time-dependent in these cell lines. These findings suggest that growth inhibition of these cell lines by GR result from the induction of apoptosis by the compound. Inhibitors of caspases did not suppress the DNA fragmentation caused by GR. N-acetyl-L-cysteine, an antioxidant drug, weakly inhibited the DNA fragmentation caused by GR suggesting that active oxidants work partly as an apoptosis-inducing transfer substance.

Glycyrrhizin induces apoptosis in human stomach cancer KATO III and human promyelotic leukemia HL-60 cells.

We have investigated the effects of glycyrrhizin (GL) on cell proliferations of human stomach cancer KATO III and promyelotic leukemia HL-60 cells, and on DNA of those cell lines. GL displayed growth inhibitory effect against KATO III and HL-60 cells. Morphological change showing apoptotic bodies was observed in the KATO III and HL-60 cells treated with GL. The fragmentation of DNA by GL to oligonucleosomal-sized fragments that is a characteristic of apoptosis was observed to be concentration- and time-dependent in both cell lines. Caspase inhibitors such as Z-VAD-FMK and Z-Asp-CH2-DCB suppressed the DNA fragmentation induced by GL. The data of the present study show that the suppression of KATO III and HL-60 cell-growth by GL results from the induction of apoptosis by GL, and that caspase is involved in the induction of apoptosis by GL in these cells.

Isolation of five types of flavonol from seabuckthorn (Hippophae rhamnoides) and induction of apoptosis by some of the flavonols in human promyelotic leukemia HL-60 cells.

Five types of flavonol were isolated from seabuckthorn (Hippophae rhamnoides) and identified by mass, 1H- and 13C-NMR. The proliferations of human promyelotic leukemia HL-60 cells were inhibited as the concentrations of these flavonols were increased. The order of the extent of growth inhibition by the flavonols at a concentration of 20 microM is as follows: pentamethylquercetin > syringetin > isorhamnetin > quercetin > kaempherol > myricetin. Apoptotic morphological changes of the nucleus, including chromatin condensation were induced in the HL-60 cells treated with quercetin, kaempherol and myricetin, respectively, but not in the cells treated with the other flavonols. The fragmentations of DNA by quercetin, kaempherol and myricetin, respectively, to oligonucleosomal-sized fragments, a characteristic of apoptosis, were observed to be dose-dependent in the HL-60 cells. These findings suggest that growth inhibition by quercetin, kaempherol and myricetin, respectively, results from the induction of apoptosis by these flavonols. The other flavonols (pentamethylquercetin, syringetin and isorhamnetin) having methoxy (-OCH3) group inhibited more strongly than the above 3 flavonols without induction of apoptosis in the HL-60 cells. These findings suggest that mechanisms of growth inhibition by pentamethylquercetin, syringetin and isorhamnetin are different from the apoptosis caused by quercetin, kaempherol and myricetin.

Hot-water extracts from adzuki beans (Vigna angularis) suppress not only the proliferation of KATO III cells in culture but also benzo(a)pyrene-induced tumorigenesis in mouse forestomatch.

Treatment of human stomach cancer KATO III cells with hot-water extracts from adzuki beans led to their growth inhibition as well as apoptosis induction. There are morphological changes in the cultured cells treated with the extracts, by which DNA fragmentation characteristic of apoptosis was actualized both concentration- and time-dependently. In contrast, N-acetyl-L-cysteine suppressed such DNA fragmentation, implying that the extracts from adzuki beans might exert antitumorigenicity via active oxygen-induced apoptosis. In order to verify this hypothesis in animal experiment, the 40% ethanol fraction of hot-water extracts was examined for its preventive effect against benzo(a)pyrene-induced tumorigenesis in the forestomach of A/J mice, given as drinking water containing the above fraction at 0.5-2.0% levels. Consequently, forestomach cancer has turned out to be reduced by 36-62% in tumor weight relative to the control. These results suggest that the fraction of hot-water adzuki extracts may serve as a nutrapharmaceutical or functional food available for cancer prevention.

2-O-methylisohemigossylic acid lactone, a sesquiterpene, isolated from roots of mokumen (Gossampinus malabarica) induces cell death and morphological change indicative of apoptotic chromatin condensation in human promyelotic leukemia HL-60 cells.

2-O-methylisohemigossylic acid lactone, a sesquiterpene, was purified from roots of mokumen (Gossampinus malabarica) and identified by Mass, and (1)H- and (13)-NMR. This sesquiterpene displayed strong growth inhibitory effect against human promyelotic leukemia HL-60 cells. Apoptotic morphological change of the nucleus, including chromatin condensation was induced in the HL-60 cells treated with the sesquiterpene. The fragmentation of DNA by the sesquiterpene to oligonucleosomal-sized fragments, a characteristic of apoptosis, was observed to be dose- and time-dependent in the HL-60 cells. Inhibitors of caspases suppressed the DNA fragmentation induced by the sesquiterpene. These findings suggest that growth inhibition by the sesquiterpene of HL-60 cells results from the induction of apoptosis by the sesqui-terpene, and that caspase cascade is involved in the induction of apoptosis by the compound in the HL-60 cells.

Induction of apoptosis by maackiain and trifolirhizin (maackiain glycoside) isolated from sanzukon (Sophora Subprostrate Chen et T. Chen) in human promyelotic leukemia HL-60 cells.

We have investigated the effects of maackiain and trifolirhizin (maackiain glycoside) isolated from sanzukon (Sophora Subprostrate Chen et T. Chen) on DNA of human promyelotic HL-60 leukemia cells. It was found that extent of induction of apoptosis by maackiain was larger than that by trifolirhizin in human leukemia HL-60 cells. Morphological changes showing apoptotic bodies were observed in the HL-60 cells treated with maackiain and trifolirhizin. The fragmentations of DNA by maackiain and trifolirhizin to oligonucleosomal-sized fragments that is a characteristic of apoptosis was observed to be concentration- and time-dependent in the HL-60 cells. The data of the present study show that the suppressions by maackiain and trifolrhizin of growth of the HL-60 cells result from the induction of apoptosis by these compounds, and that the extent of growth suppression and induction of apoptosis by maackiain was greater than that by the glycoside (trifolirhizin).

Induction of apoptosis by three types of procyanidin isolated from apple (Rosaceae Malus pumila) in human stomach cancer KATO III cells.

We have investigated the effects of three types of procyanidin isolated from apple (Rosaceae Malus pumila) on DNA of human stomach cancer KATO III cells. Induction of apoptosis by these procyanidins was observed in human stomach cancer KATO III cells. Morphological changes showing apoptotic bodies were observed in the KATO III cells treated with procyanidins. The fragmentation of DNA by procyanidins to oligonucleosomal-sized fragments, a characteristic of apoptosis, was observed to be concentration- and time-dependent in the KATO III cells. N-acetyl-L-cysteine, an antioxidant, suppressed the DNA fragmentation induced by these procyanidins. The present study shows that the suppression of KATO III cell-growth by three types of procyanidin results from the induction of apoptosis by these compounds, and that active oxygen is involved in the induction of apoptosis by these compounds in the KATO III cells.

Induction of apoptosis by lupeol isolated from mokumen (Gossampinus malabarica L. Merr) in human promyelotic leukemia HL-60 cells.

We have investigated the effect of lupeol (lup-20(29)-ene-3beta-ene-3-ol) isolated from mokumen (Gossampinus malabarica L. Merr) on DNA of human promyelotic HL-60 leukemia cells. Induction of apoptosis by lupeol was observed in human leukemia HL-60 cells. Morphological change showing apoptotic bodies was observed in the HL-60 cells treated with lupeol. The fragmentation of DNA by lupeol to oligonucleosomal-sized fragments that is a characteristic of apoptosis was observed to be concentration- and time-dependent in the HL-60 cells. Flow cytometric analysis showed that the hypodiploid nuclei of HL-60 cells were increased to 28.5, 42.0 and 70.9% after a 3-day treatment with 75, 100 and 150 micro M lupeol, respectively. The data of the present study show that the suppression by lupeol of growth of the HL-60 cells results from the induction of apoptosis by this compound.

Hot water soluble sesquiterpenes [anhydroperoxy-costunolide and 3-oxoeudesma-1,4(15),11(13)triene-12,6alpha-olide] isolated from laurel (Laurus nobilis L.) induce cell death and morphological change indicative of apoptotic chromatin condensation in leukemia cells.

Hot water soluble (HWS)-sesquiterpenes [anhydroperoxycostunolide and 3-oxo-eudesma-1,4(15),11(13)triene-12,6alpha-olide] were purified from Laurel (Laurus nobilis L.) and identified by Mass, and 1H- and 13C-NMR. These HWS-sesquiterpenes displayed strong growth inhibitory effect against human promyelotic leukemia HL-60 cells. Apoptotic morphological changes of the nucleus, including chromatin condensation were induced in the HL-60 cells treated with the sesquiterpenes. Flow cytometric analysis showed that the hypodiploid nuclei of HL-60 cells were increased to 10.5, 46.5, and 91.3% after a 3 day-treatment with 2.5, 5 and 10 micromoles anhydroperoxycostunolide, respectively. And the same analysis showed that the hypodiploid nuclei of HL-60 cells were increased to 9.8, 39.2 and 89.6% after a 3 day-treatment with 5, 10 and 20 micromoles 3-oxo-eudesma-1,4(15),11(13)triene-12,6alpha-olide, respectively. These findings suggest that growth inhibition by anhydroperoxycostunolide and 3-oxo-eudesma-1,4(15),11(13)triene-12,6alpha-olide of HL-60 cells results from the induction of chromatin condensation in the HL-60 cells. On the other hand, fragmentations by these compounds of DNA to oligonucleosomal-sized fragments were not observed in the HL-60 cells.

Sesquiterpenes (costunolide and zaluzanin D) isolated from laurel (Laurus nobilis L.) induce cell death and morphological change indicative of apoptotic chromatin condensation in leukemia HL-60 cells.

Sesquiterpenes (costunolide and zaluzanin D) were purified from laurel (Laurus nobilis L.) and identified by Mass, and 1H- and 13C-NMR. These sesquiterpenes displayed strong growth inhibitory effect against human promyelotic leukemia HL-60 cells. Apoptotic morphological changes of the nucleus, including chromatin condensation were induced in the HL-60 cells treated with the sesquiterpenes. Flow cytometric analysis showed that the hypodiploid nuclei of HL-60 cells were increased to 11.4, 47.0, and 92.5% after a 3-day-treatment with 5, 10 and 15 micro M costunolide, respectively. The same analysis showed that the hypodiploid nuclei of HL 60 cells were increased to 12.4, 28.9 and 76.7% after a 3-day-treatment with 10, 15 and 20 micro M zaluzanin D, respectively. These findings suggest that growth inhibition by costunolide and zaluzanin D of HL-60 cells results from the induction of chromatin condensation in the HL-60 cells. On the other hand, fragmentations by these compounds of DNA to oligonucleosomal-sized fragments were not observed in the HL-60 cells.

Cyanidin 3-O-beta-D-glucoside isolated from skin of black Glycine max and other anthocyanins isolated from skin of red grape induce apoptosis in human lymphoid leukemia Molt 4B cells.

Cyanidin 3-O-beta-D-glucoside (CG) was purified from black bean seed coat and other anthocyanins were prepared from red grape skin. These anthocyanins were identified by Mass, and 1H- and 13C-NMR. The effects of four anthocyanins on cell viability in human leukemia Molt 4B cells were investigated. The anthocyanins displayed strong growth inhibitory effects against human leukemia Molt 4B cells. Morphological changes showing apoptotic bodies were observed in the Molt 4B cells treated with these anthocyanins. The fragmentations by these anthocyanins of DNA to oligonucleosomal-sized fragments, that is a characteristic of apoptosis, were observed to be concentration-dependent. N-acetyl-L-cysteine, an antioxidant, suppressed the fragmentation of DNA by these anthocyanins. These findings suggest that growth inhibition of Molt 4B cells by these anthocyanins result from the induction of apoptosis by these compounds and that active oxygen is involved in the induction of apoptosis in the Molt 4B cells.

PPAR-gamma, TNF-alpha messenger RNA levels and lipase activity in the pregnant and lactating rat.

Dramatic alternations in maternal metabolism occur during gestation and lactation, especially glucose and fat metabolism. For example, in rats, the amount of body fat mass increases during gestation, then decreases just prior to delivery, and remains low after parturition. To investigate the factors involved in such changes in maternal fat mass, messenger ribonucleic acid (mRNA) levels of adipocytokines, peroxisome proliferator-activated receptor-gamma (PPAR-gamma) and tumor necrosis factor-alpha (TNF-alpha), were examined in the intraabdominal adipose tissue of non-pregnant rats, pregnant rats and postpartum rats. We also examined the issue of whether apoptosis, which could be promoted by PPAR-gamma and TNF-alpha, is involved in any of the changes in maternal fat mass The activity of lipoprotein lipase (LPL) and hormone sensitive lipase (HSL) in adipose tissue was also measured. PPAR-gamma and TNF-alpha mRNA levels remained constant during the gestational and postpartum periods. Apoptosis was not detected at any time as evidenced by DNA laddering and in situ staining. LPL activity was significantly increased at day 5 and remained elevated until day 14 of gestation. HSL activity was significantly increased at day 10 of gestation and then decreased after delivery, at day 10 of lactation. In conclusion, during the gestational and postpartum period of rats, changes in maternal fat mass did not directly correlate with the levels of expression of PPAR-gamma and TNF-alpha mRNA. Apoptosis also does not appear to influence on fat mass change. The changes in LPL and HSL activities during gestation suggest that these enzymes might be regulators of maternal adipose tissue level.

Spermine induces cataract and 43-kDa protein that binds spermine possibly participates in the cataract formation.

Among polyamines (putrescine, spermidine, and spermine), spermine specifically induces cataract in an organ cultured lens. Spermine uptake nearly paralleled the cataract formation. When polyamines were added to lens soluble proteins, spermine specifically induced turbidity. When lens soluble proteins were separated by gel chromatography, heavy-molecular-weight protein (HMW, high molecular form of alpha-crystallin) and proteins between betaH- and betaL-crystallin fractions reacted with spermine and aggregated. SDS-polyacrylamide gel electrophoresis of the aggregated proteins showed that 43-kDa lens protein was commonly observed in both aggregates. Spermine-affinity chromatography of the total soluble proteins showed the binding of HMW protein to the gel and the chromatogram of the second turbidity peak in the gel chromatography showed the binding of 43-kDa protein. These results indicated that 43-kDa protein, which is present as a subunit in HMW and also in free form, binds spermine and induces turbidity of lens soluble proteins and produces cataract in a cultured lens.

Protodioscin isolated from fenugreek (Trigonella foenumgraecum L.) induces cell death and morphological change indicative of apoptosis in leukemic cell line H-60, but not in gastric cancer cell line KATO III.

Protodioscin (PD) was purified from fenugreek (Trigonella foenumgraecum L.) and identified by Mass, and 1H- and 13C-NMR. The effects of PD on cell viability in human leukemia HL-60 and human stomach cancer KATO III cells were investigated. PD displayed strong growth inhibitory effect against HL-60 cells, but weak growth inhibitory effect on KATO III cells. Morphological change showing apoptotic bodies was observed in the HL-60 cells treated with PD, but not in KATO III cells treated with PD. Flow cytometric analysis showed that the hypodiploid nuclei of HL-60 cells were increased to 75.2, 96.3, and 100% after a 3-day treatment with 2.5, 5, and 10 microM PD, respectively. The fragmentation by PD of DNA to oligonucleosomal-sized fragments, that is a characteristic of apoptosis, was observed to be both concentration- and time-dependent in the HL-60 cells. These findings suggest that growth inhibition by PD of HL-60 cells results from the induction of apoptosis by this compound in HL-60 cells.

Diol- and triol-types of phytol induce apoptosis in lymphoid leukemia Molt 4B cells.

The exposure of human lymphoid leukemia Molt 4B cells to diol- and triol-types of phytol which were synthesized and identified by Mass, and 1H- and 13C-NMR, led to both growth inhibition and induction of programmed cell death (apoptosis). Morphological changes showing apoptotic bodies were observed in the Molt 4B cells treated with diol- and triol-types of phytol. The fragmentations of DNA by the diol- and triol-types of phytol to oligonucleosomal-sized fragments, that is a characteristic of apoptosis, were observed to be both concentration- and time-dependent. These findings suggest that growth inhibition of Molt 4B cells by the diol- and triol-types of phytol results from the induction of apoptosis in the leukemic cells.

Effect of Acanthopanax senticosus Harms on biogenic monoamine levels in the rat brain.

The extract of the stem bark of Acanthopanax senticosus Harms (ASH) is known to have healing and protective effects on stress-induced disturbance of mental status. We have analysed whether a single or chronic (2 week) administration of ASH can affect concentrations of noradrenaline (NA), dopamine (DA) and their metabolites in the normal rat brain. A single p.o. administration of ASH elevated the NA and DA levels in the whole brain of rats in a dose-dependent manner. A single or 2 week administration of ASH (500 mg/kg) showed a marked increase in the DA level only in the striatum. However, NA levels were increased by a single dose of ASH in a wide range of brain regions such as the cortex, hypothalamus, striatum, hippocampus, substantia nigra and pons. When administered for 2 weeks no increase in NA levels was seen in these brain regions, except for an increase in the frontal cortex and anterior hypothalamus. The present results suggest that ASH may act by regulating NA and DA levels in specific brain regions related to stress response and Parkinson's disease.

Specific induction of apoptosis by 1,8-cineole in two human leukemia cell lines, but not a in human stomach cancer cell line.

We have investigated the effects of 1,8-cineole [the main component of essential oil prepared from bay-leaves Laurus nobilis L.)] on DNA of human leukemia cell lines, Molt 4B, HL-60 and stomach cancer KATO III cells. Specific induction of apoptosis by 1,8-cineole was observed in human leukemia Molt 4B and HL-60 cells, but not in human stomach cancer KATO III cells. Morphological changes showing apoptotic bodies were observed in the human leukemia HL-60 cells treated with 1,8-cineole. The fragmentations of DNA by cineole to oligonucleosomal-sized fragments that is a characteristic of apoptosis were concentration- and time-dependent in Molt 4B and HL-60 cells, but not in KATO III cells. The present study shows that the suppression growth by 1,8-cineole in the leukemia cell lines results from the induction of apoptosis by this compound.

Selective induction of apoptosis by ar-turmerone isolated from turmeric (Curcuma longa L) in two human leukemia cell lines, but not in human stomach cancer cell line.

We have investigated the effects of ar-turmerone isolated from turmeric (Curcuma longa L) on DNA of human leukemia cell lines, Molt 4B, HL-60 and stomach cancer KATO III cells. It was found that selective induction of apoptosis by ar-turmerone was observed in human leukemia Molt 4B and HL-60 cells, but not in human stomach cancer KATO III cells. Morphological changes showing apoptotic bodies were observed in the human HL-60 and Molt 4B cells treated with ar-turmerone. The fragmentation of DNA by ar-turmerone to oligonucleosomal-sized fragments that is a characteristic of apoptosis was observed to be concentration- and time-dependent in Molt 4B and HL-60 cells, but not in KATO III cells. The data of the present study show that the suppression by ar-turmerone of growth of these leukemia cell lines results from the induction of apoptosis by this compound.