Mechanism of Action of Panaxytriol on Midazolam 1′-Hydroxylation and 4-Hydroxylation Mediated by CYP3A in Liver Microsomes and Rat Primary Hepatocytes
Mechanism of Action of Panaxytriol on Midazolam 1′-Hydroxylation and 4-Hydroxylation Mediated by CYP3A in Liver Microsomes and Rat Primary Hepatocytes
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DOI：
10.1248/bpb.b15-00143
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发表时间：
2015-10-01
影响因子：
2
通讯作者：
Liu, Mingyi
中科院分区：
文献类型：
--
作者：
He, Fang;Zhang, Wen;Liu, Mingyi
在我们以前的研究中，人参三醇（PXT）被证明能显著促进咪达唑仑（MDZ）的1 ′-羟基化，但抑制MDZ的4-羟基化。为了探讨潜在的机制，我们研究了PXT对细胞色素P450 3A（CYP 3A）介导的MDZ代谢途径的影响，使用大鼠肝微粒体（RLM），人肝微粒体（HLM），和大鼠原代肝细胞。在PXT存在下，V-max. RLM和HLM中4- 0 H MDZ的浓度分别从0.72和0.32降低到0.51和0.12 nmol/min·mg pro，K-m值分别从5.12和27.87增加到7.26和32.80 μ m。但PXT的存在降低了RLM和HLM中MDZ 1 '-羟基化的Km值，增加了Vmax值。有趣的是，在培养45分钟后，在原代大鼠肝细胞中也观察到PXT对MDZ 4-羟基化和1 '-羟基化的不同影响。PXT对大鼠肝细胞CYP 3A 1/2 mRNA表达水平无影响。PXT 4.0 μ g/mL和8.0 μ g/mL可显著抑制MDZ的4-羟基化和1 '-羟基化，使CYP 3A 1/2 mRNA表达水平分别降至对照组的87%和80%（CYP 3A 1）及89%和85%（CYP 3A 2）。这些结果表明，PXT可以激活MDZ 1 '-羟基化，但抑制MDZ 4-羟基化，通过改变CYP 3A酶的亲和力和代谢率后，短期干预。然而，长期使用PXT可通过下调CYP 3A 1/2 mRNA表达来抑制MDZ的4-羟基化和1 '-羟基化。
In our previous study, panaxytriol (PXT) was shown to enhance midazolam (MDZ) 1'-hydroxylation significantly but to inhibit MDZ 4-hydroxylation. To explore the underlying mechanism, we investigated the effects of PXT on cytochrome P450 3A (CYP3A)-mediated MDZ metabolic pathways using rat liver microsomes (RLM), human liver microsomes (HLM), and rat primary hepatocytes. In the presence of PXT, the V-max. of 4-0H MDZ decreased from 0.72 to 0.51nmol/min.mg pro in RLM and from 0.32 to 0.12nmol/min.mg pro in HLM, and the K-m value increased from 5.12 to 7.26 mu m in RLM and from 27.87 to 32.80 mu m in HLM. But the presence of PXT reduced the Km and increased the Vmax values of MDZ 1'-hydroxylation in RLM and HLM. Interestingly, the differential effect of PXT on MDZ 4-hydroxylation and 1'-hydroxylation was also observed in primary rat hepatocytes after 45-min culture. PXT did not affect the expression levels of CYP3A1/2 mRNA in rat hepatocytes. With extension of the culture time to 6h, however, PXT significantly inhibited both MDZ 4-hydroxylation and 1'-hydroxylation, and the expression level of CYP3A1/2 mRNA was decreased to 87% and 80% (CYP3A1) and to 89% and 85% (CYP3A2) of those in controls in the presence of PXT 4.0 and 8.0 mu g/mL, respectively. These results suggest that PXT could activate MDZ 1'-hydroxylation but inhibit MDZ 4-hydroxylation by changing the CYP3A enzyme affinity and metabolic rate after a short-term intervention. However, long-term treatment with PXT could inhibit both the 4-hydroxylation and 1'-hydroxylation of MDZ by downregulating CYP3A1/2 mRNA expression.