Tanaka Kazuyuki D, Kawai Yusuke K, Ikenaka Yoshinori, Harunari Tsunehito, Tanikawa Tsutomu, Fujita Shoichi, Ishizuka Mayumi
Department of Environmental Veterinary Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Kita-ku, Sapporo, Hokkaido, Japan.
J Vet Med Sci. 2013 Feb;75(2):135-9. doi: 10.1292/jvms.12-0161. Epub 2012 Sep 26.
Warfarin is a rodenticide commonly used worldwide. It inhibits coagulation of blood by inhibiting vitamin K 2,3-epoxide reductase (VKOR) activity. An inadequate supply of vitamin K blocks the production of prothrombin and causes hemorrhage. Recently, warfarin-resistant brown rats (Rattus norvegicus) were found around the Aomori area of Japan. There is no significant difference in the metabolic activity of warfarin in sensitive and resistant brown rats. To clarify the mechanism underlying warfarin resistance, we cloned the VKORC1 gene from rats and identified a novel substitution of arginine to proline at position 33 of the VKORC1 amino acid sequence. Then, we determined the differences in kinetics of VKOR activity between warfarin-resistant and sensitive rats. Hepatic microsomal VKOR-dependent activity was measured over a range of vitamin K epoxide concentrations from 6.25 to 150 µM. The Vmax values of resistant rats (0.0029 ± 0.020 nmol/min/mg) were about one tenth of those of sensitive rats (0.29 ± 0.12 nmol/min/mg). The Km values of resistant rats (47 ± 32 µM) were similar to those of sensitive rats (59 ± 18 µM). Warfarin-sensitive rats exhibited enzyme efficiencies (Vmax/Km) which were ten-fold greater than those observed in resistant rats. It may mean that VKOR activity of warfarin-resistant Aomori rats is almost lost, because their enzymatic efficiencies are very low even without warfarin. Further studies are needed to clarify how these rats can survive with a markedly reduced VKOR activity and how they simultaneously exhibit warfarin resistance.
华法林是一种在全球广泛使用的灭鼠剂。它通过抑制维生素K 2,3-环氧化物还原酶（VKOR）的活性来抑制血液凝固。维生素K供应不足会阻碍凝血酶原的产生并导致出血。最近，在日本青森地区发现了对华法林有抗性的褐家鼠（Rattus norvegicus）。敏感和抗性褐家鼠对华法林的代谢活性没有显著差异。为了阐明华法林抗性的潜在机制，我们从大鼠中克隆了VKORC1基因，并在VKORC1氨基酸序列的第33位鉴定出精氨酸到脯氨酸的新替换。然后，我们测定了抗华法林和敏感大鼠之间VKOR活性动力学的差异。在6.25至150μM的一系列维生素K环氧化物浓度范围内测量肝微粒体VKOR依赖性活性。抗性大鼠的Vmax值（0.0029±0.020 nmol/min/mg）约为敏感大鼠（0.29±0.12 nmol/min/mg）的十分之一。抗性大鼠的Km值（47±32μM）与敏感大鼠（59±18μM）相似。对华法林敏感的大鼠表现出的酶效率（Vmax/Km）比抗性大鼠高十倍。这可能意味着抗华法林的青森大鼠的VKOR活性几乎丧失，因为即使没有华法林，它们的酶效率也非常低。需要进一步研究来阐明这些大鼠如何在VKOR活性显著降低的情况下存活，以及它们如何同时表现出对华法林的抗性。
