Isoflavones enhance pharmacokinetic exposure of active lovastatin acid via the upregulation of carboxylesterase in high-fat diet mice after oral administration of Xuezhikang capsules

Abstract

Xuezhikang capsule (XZK) is a traditional Chinese medicine that contains lovastatin (Lv) for hyperlipidemia treatment, although it has fewer side effects than Lv. However, the pharmacokinetic mechanisms contributing to its distinct efficacy and low side effects are unclear. Mice were fed a high-fat diet (HFD) for 6 weeks to induce hyperlipidemia. We first conducted the pharmacokinetic studies in HFD mice following oral administration of Lv (10 mg/kg, i.g.) and found that HFD remarkably decreased the active form of Lv (the lovastatin acid, LvA) exposure in the circulation system, especially in the targeting organ liver, with a declined conversion from Lv to LvA, whereas the Lv (responsible for myotoxicity) exposure in muscle markedly increased. Then we compared the pharmacokinetic profiles of Lv in HFD mice after the oral administration of XZK (1200 mg/kg, i.g.) or an equivalent dose of Lv (10 mg/kg, i.g.). A higher exposure of LvA and lower exposure of Lv were observed after XZK administration, suggesting a pharmacokinetic interaction of some ingredients in XZK. Further studies revealed that HFD promoted the inflammation and inhibited carboxylesterase (CES) activities in the intestine and the liver, thus contributing to the lower transformation of Lv into LvA. In contrast, XZK inhibited the inflammation and upregulated CES in the intestine and the liver. Finally, we evaluated the effects of monacolins and phytosterols, the fractional extracts of isoflavones, on inflammatory LS174T or HepG2 cells, which showed that isoflavones inhibited inflammation, upregulated CES, and markedly enhanced the conversion of Lv into LvA. For the first time, we provide evidence that isoflavones and Lv in XZK act in concert to enhance the efficacy and reduce the side effects of Lv.

Keywords: Xuezhikang capsules; carboxylesterase; hyperlipidemia; inflammation; isoflavones; lovastatin; lovastatin acid; pharmacokinetics.

Fig. 1

Pharmacokinetic profiles of Lv and LvA in the control and HFD mice. Concentration–time curves of Lv and LvA in plasma (a, e) and tissues including liver (b, f), intestine (c, g), and muscle (d, h) are presented. The experimental groups were as follows: Lv administration (10 mg/kg, i.g.) in the control mice (Control + Lv), Lv administration (10 mg/kg, i.g.) in the HFD mice (HFD + Lv), and XZK administration (1200 mg/kg, i.g., an equivalent dose to 10 mg/kg of Lv) in the HFD mice (HFD + XZK). The values are expressed as the mean ± SD of four mice for each time point in each group

Fig. 2

Expression levels and activities of CES in the intestine and the liver. Expression levels of Ces2 (a–d) and the total esterase activity (e) in the intestine; expression levels of Ces1 (f–h) and the total esterase activity (i) in the liver in each experimental group. The values are expressed as mean ± SD, n = 4. *P < 0.05, **P < 0.01 vs. Control; ^#P < 0.05, ^##P < 0.01 vs. HFD + Lv

Fig. 3

Expression levels of the inflammatory factors and PXR in the intestine and the liver. mRNA levels of TNF-α (a), IL-1β (b), and IL-6 (c), and mRNA and protein expressions of PXR (d) in the intestine; mRNA levels of TNF-α (e), IL-1β (f), and IL-6 (g), and mRNA and protein expressions of PXR (h) in the liver in each experimental group. The values are expressed as mean ± SD, n = 4. *P < 0.05, **P < 0.01 vs. Control; ^#P < 0.05, ^##P < 0.01 vs. HFD + Lv

Fig. 4

Effects of XZK and its fractions on inflammatory factors in LS174T or HepG2 cells. mRNA levels of TNF-α (a), IL-1β (b), and IL-6 (c) in LS174T cells; mRNA levels of TNF-α (d), IL-1β (e), and IL-6 (f) in HepG2 cells after the administration of Lv (10 μM), XZK (480 μg/mL, an equivalent dose to 10 μM Lv), F1 (10.9 μg/mL), F2 (4.5 μg/mL), F3 (14.2 μg/mL), or F4 (2.6 μg/mL) for 24 h under 24 h of pre-treatment of TNF-α (10 ng/mL), or LPS (1 μg/mL). All the experiments were conducted in triplicate, and data with error bars are presented as mean ± SD (n = 3). *P < 0.05, **P < 0.01 vs. Control; ^#P < 0.05, ^##P < 0.01 vs. TNF-α or LPS treatment alone

Fig. 5

Effects of XZK and its fraction 1, F1, on PXR and CES in inflammatory LS174T or HepG2 cells. mRNA and protein expressions of PXR (a) and Ces2 (b), and the total esterase activity (c) in LS174T cells; mRNA and protein expressions of PXR (d) and Ces1 (e), and the total esterase activity (f) in HepG2 cells after the administration of Lv (10 μM), XZK (480 μg/mL, an equivalent dose to 10 μM Lv), F1 (10.9 μg/mL), or F1 (10.9 μg/mL) plus Lv (10 μM) for 24 h under 24 h of pre-treatment of TNF-α (10 ng/mL) or LPS (1 μg/mL). All experiments were conducted in triplicate, and data with error bars are presented as mean ± SD (n = 3). *P < 0.05, **P < 0.01 vs. Control; ^#P < 0.05, ^##P < 0.01 vs. TNF-α or LPS treatment alone; ^$P < 0.05, ^$$P < 0.01 vs. TNF-α or LPS + Lv

Fig. 6

Effects of XZK and its fraction 1, F1, on the intracellular accumulation and efficacy or toxicity of the substrates for CES in inflammatory LS174T or HepG2 cells. Cellular accumulations of Lv (a, d) and LvA (b, e), and the conversion ratios (c, f) in LS174T cells or HepG2 cells after administration for 2 h were presented. The experimental groups were as follows: control + Lv (10 μM), TNF-α, or LPS + Lv (10 μM), TNF-α or LPS + XZK (480 μg/mL, an equivalent dose to 10 μM of Lv), TNF-α or LPS + Lv (10 μM) + F1 (isoflavones, 10.9 μg/mL). TNF-α (10 ng/mL) or LPS (1 μg/mL) pre-treated the cells for 24 h. *P < 0.05, **P < 0.01 vs. Control + Lv; ^#P < 0.05, ^##P < 0.01 vs. TNF-α or LPS + Lv; Cholesterol-lowering efficacy of Lv (g, ranging from 20 to 80 μM, 24 h), as well as cytotoxicity of irinotecan (h, at 0.3, 3.0, 30 μM, 24 h) and clopidogrel (i, at 1, 10, 100 μM, 24 h) with or without the co-administration of F1 (isoflavones, 10.9 μg/mL) under 24 h LPS pre-treatment (1 μg/mL) in HepG2 cells were presented. *P < 0.05, **P < 0.01 vs. Control; ^#P < 0.05, ^##P < 0.01 vs. LPS treatment alone. All experiments were conducted in triplicate, and data with error bars are presented as mean ± SD

Fig. 7

A schematic diagram of the pharmacokinetic mechanisms of F1 in XZK contributed to an improved efficacy and lower toxicity