Latiporn Udomsuk

Bimodal action of miroestrol and deoxymiroestrol, phytoestrogens from Pueraria candollei var. mirifica, on hepatic CYP2B9 and CYP1A2 expressions and antilipid peroxidation in mice.

Miroestrol and deoxymiroestrol are phytoestrogens isolated from Pueraria candollei var. mirifica. The influence of miroestrol and dexoymirosestrol on hepatic cytochrome P450 (P450) enzymes and antioxidative activity in brain was examined in C57BL/6 mice compared with that of a synthetic female sex hormone estradiol. We hypothesized that miroestrol and deoxymiroestrol would induce CYP2B9 expression, whereas CYP1A2 expression would be suppressed compared with estradiol. Miroestrol and deoxymiroestrol treatment significantly increased uterus weight and volume. In addition, both of these phytoestrogens induced the expression of CYP2B9 and suppressed the expression of CYP1A2, as expected. Hepatic P450 activities correspondingly showed that both compounds increased benzyloxyresorufin O-dealkylase activity, whereas methoxyresorufin O-dealkylase activity was reduced. These observations suggested that miroestrol and deoxymiroestrol might affect hepatic P450 enzymes, including the CYP2B9 and CYP1A2 P450 isoforms. Assessment of lipid peroxidation demonstrated that miroestrol and deoxymiroestrol markedly decreased levels of malondialdehyde formation in the mouse brain. This is the first report suggesting miroestrol and deoxymiroestrol as potential alternative medicines to estradiol because of their distinctive ability to regulate mouse hepatic P450 expression and their beneficial antioxidative activities.

Impact of Pueraria candollei var. mirifica and its potent phytoestrogen miroestrol on expression of bone-specific genes in ovariectomized mice

Miroestrol (MR) is a highly active phytoestrogen isolated from tuberous root of Pueraria candollei var. mirifica (PM). Modulatory effects of PM and MR on osteoprotegerin (OPG) and receptor activator of nuclear factor kappa B ligand (RANKL) mRNAs which are bone-specific genes were investigated in ovariectomized female ICR mice. After ovariectomy, expression of OPG mRNA was suppressed but that of RANKL was induced. Estradiol benzoate (E2) recovered OPG expression to the level comparable to the sham while that of RANKL was suppressed in ovariectomized mice. PM crude extract (PME) significantly down-regulated the expression of RANKL mRNA with no change in the OPG level whereas MR elevated the expression of OPG mRNA with lowering level of RANKL mRNA, resulting in the increased OPG/RANKL ratio, and consequently lead to lowering progression of osteoporosis at molecular level. These findings revealed potential of PME and MR on bone loss prevention via increasing the ratio of OPG to RANKL (osteoformation/osteoresorption) in liver of ovariectomized mice. Therefore, using PME and MR as alternative hormone replacement therapy of E2 might be beneficial recommended due to advantageous on regulation of osteoporosis related genes.

Down regulation of gene related sex hormone synthesis pathway in mouse testes by miroestrol and deoxymiroestrol.

Miroestrol and deoxymiroestrol are phytoestrogens isolated from tuberous root of Pueraria candollei var. mirifica. Modulatory effects of miroestrol and deoxymiroestrol on enzymes involved in sex-hormone synthesis pathway in male C57BL/6 mice were investigated using semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). Miroestrol and deoxymiroestrol suppressed the expressions of 3β-HSD, 17β-HSD1, and CYP17 while CYP19 mRNA expression was slightly decreased. In addition, the expression of 17β-HSD2 was induced in correlation with those did by estradiol. These observations supported that miroestrol and deoxymiroestrol could exhibit the same effect as estradiol regarding regulation of testicular gene related sex hormone synthesis pathway.

Isoflavonoid production in a hairy roots culture of Pueraria candollei.

A hairy roots culture of Pueraria candollei was established using Agrobacterium rhizogenes ATCC15834 and grown in half-strength Murashige and Skoog (MS) medium. The highest production of total isoflavonoids was found to be (36.48 ± 4.09) mg/g dry wt [(3.39 ± 0.20) mg/g dry wt puerarin, (29.91 ± 3.74) mg/g dry wt daidzin, (1.65 ± 0.09) mg/g dry wt genistin, (0.76 ± 0.03) mg/g dry wt daidzein, and (0.76 ± 0.03) mg/g dry wt genistein, respectively]. The total isoflavonoid content in hairy roots of P. candollei was 5.18-fold higher than that of the native tuber. Effects of sucrose content and medium type on growth and isoflavonoid production were investigated. 5% (w/v) Sucrose was an optimum content for the growth and isoflavonoid accumulation in P. candollei hairy roots. Half-strength MS medium had the highest effect for biomass production whereas woody plant medium had mostly stimulated isoflavonoid content in hairy roots

Production of isoflavonoids in callus cultures of Pueraria candollei var. mirifica.

Pueraria candollei Wall. ex Benth. var. mirifica (Airy Shaw & Suvat.) Niyomdham was investigated for callus induction using Murashige and Skoog (MS) medium containing different plant growth regulators. After 8 weeks of culture, 66 - 100% of leaf or stem explants formed calli. Calli from stem explants cultured on MS medium supplemented with 0.5 mg/l thidiazuron (TDZ) gave the maximum of shoot induction (16%) and the highest level of total isoflavonoids [(50.39 ± 7.06) mg/g dry wt], which was 7-fold higher than that of the native tuber [(7.04 ± 0.29) mg/g dry wt]. These results suggest that addition of TDZ to the culture medium markedly enhances the production of isoflavonoids in calli induced from stem explants of P. candollei var. mirifica.

Different profiles of hepatic alkoxyresorufin O‐dealkylase activities in small rodents

The aims of the present study were to determine cytochrome P450 enzyme activity in six strains of experimental rodents (n = 5/sex/species): ICR, C57BL/6 and DBA/2 mice; Sprague Dawley and Wistar rats; and Dunkin Hartley guinea pigs. After animals were treated with the typical inducers β‐naphthoflavone (BNF), dexamethasone (DEX) and phenobarbital (PB), the levels of O‐dealkylation of ethoxyresorufin (EROD), methoxyresorufin (MROD), pentoxyresorufin (PROD) and benzyloxyresorufin (BROD) activity were determined using responsive catalytic reactions to study CYP1A1, CYP1A2 and CYP2B, respectively. A maximal induction of EROD and MROD was found in BNF‐treated animals from all strains (2.4‐ to 15.1‐fold) except DBA/2 (0.9‐ to 1.8‐fold). C57BL/6 mice had the strongest BNF‐induced EROD (15.1‐fold) and MROD (8.3‐fold) activities. No differences in BNF‐induced EROD and MROD activities were observed between males and females. However, the EROD activity of Wistar rats and the MROD activity of Sprague Dawley rats were higher in males than females. DEX induced PROD activity only in mice (1.3‐ to 7.1‐fold), but not in rats and guinea pigs (0.2‐ to 1.1‐fold). However, induction of BROD activity was found in DEX‐treated mice and rats (1.5 to 12.5‐fold), but not in guinea pigs (0.3 to 0.4‐fold). PB caused a significant elevation of PROD (1.7‐ to 10.4‐fold) and BROD (31‐ to 13.2‐fold) activities in all the animals. PB‐induced BROD activity was higher in females than males in Sprague Dawley rats. These observations strongly suggest that the choice of experimental animal strain, species and inducer is of critical importance for studies of drug metabolism and interaction. Copyright

Suppression of BSEP and MRP2 in mouse liver by miroestrol and deoxymiroestrol isolated from Pueraria candollei

Miroestrol and deoxymiroestrol are highly active phytoestrogens isolated from the tuberous root of Pueraria candollei var. mirifica (Leguminosae). Modulatory effects of miroestrol and deoxymiroestrol on the mRNAs of BSEP and MRP2 genes involved in bile salt transportation, in C57BL/6 mice were investigated. In contrast to estradiol, miroestrol and deoxymiroestrol suppressed the expression of BSEP and MRP2 mRNA in both male and female mice. The results suggest for the first time that the use of miroestrol and deoxymiroestrol-containing products as alternative medicines or health supplements should be concerned according to their effects on key genes that regulate the bile salt export pump, which could result in the risk of hepatotoxicity and intrahepatic cholestasis.

Miroestrol Improves the Expression of Genes Involved in Osteoporosis and Bile Salt Transportation in the ï¢-Naphthoflavone-Treated Mouse Liver

Background : Miroestrol, a potent phytoestrogen isolated from the tuberous roots of Pueraria candollei var. mirifica , exhibits estrogenic activity and has long been used as a Thai traditional medicine for rejuvenation. Objective : To investigate the influence of miroestrol on the regulation of osteoporosis, a worldwide public health problem, and bile salt transportation-related genes was examined as an additional factor in the risk/benefit assessment of the use of miroestrol as an alternative estrogen replacement therapy. Results : Miroestrol at a dose of 0.5 mg/kg/day elevated the expression of OPG mRNA while suppressing the RANKL levels, increasing the OPG/RANKL ratio and consequently inhibiting the progression of osteoporosis at the molecular level. Miroestrol additionally improved the ratio of OPG/RANKL in the livers of I²-naphthoflavone-induced mice. In addition to ameliorating osteoclastogenesis, the 0.5 mg/kg/day dose of miroestrol decreased the extent of I²-naphthoflavone-induced MRP2 expression without affecting the level of BSEP mRNA in the mouse livers. Conclusion : This is the first report suggesting that miroestrol may be a promising candidate for alternative estrogen replacement therapy due to the beneficial effects on the osteoporosis and bile salt transportation regulatory pathways.