Establishing a rat model for the study of vitamin K deficiency

Abstract

The main vitamin K-deficient model, minidose warfarin, is different from the pathological mechanism of vitamin K deficiency, which is a shortage of vitamin K. The objective of this study was to establish a new method of vitamin K-deficient model combining a vitamin K-deficient diet with the intragastrical administration of gentamicin in rats. The clotting was assayed by an automated coagulation analyser. The plasma PIVKA-II was assayed by ELISA. The vitamin K status was detected by an HPLC-fluorescence system. In the diet- and gentamicin-induced vitamin K-deficient 14-day group, the rats had undetected vitamin K1 and vitamin K2 in the liver and a prolonged APTT. In the 21-day group, there was also a prolonged PT and a decrease of the FIX activities. In the 28-day group, the undetected vitamin K1 and vitamin K2, the prolonged PT and APTT, and the decrease of the FII, FVII, FIX, and FX activities prompted the suggestion that there were serious deficiencies of vitamin K and vitamin K-dependent coagulation in rats. It is suggested that the diet- and gentamicin-induced vitamin K-deficient 14-day or 21-day model can be used for studies related to the status of vitamin K. The vitamin K-deficient 28-day model can be applied to research involving both the status of vitamin K and of vitamin K-dependent coagulation. In conclusion, the combination of a vitamin K-deficient diet with the administration of gentamicin results in a useful model of vitamin K-deficieny.

Keywords: animal model; coagulation factor; gentamicin; vitamin K deficiency; warfarin.

Figure 1

Representative chromatograms of vitamin K status in standard dry food (a) and vitamin K‐deficient food (b).

Figure 2

Concentrations of PIVKA‐II in plasma. Animals were administrated as follows: control (standard dry food), vitamin K‐deficient 14‐day, 21‐day and 28‐day groups (rats received a vitamin K‐deficient diet and the intragastric administration of 30 mg/kg gentamicin for 14, 21 and 28 days respectively). The values are shown as the mean ± SE, n = 10. **P < 0.01 vs. control.

Figure 3

The changes of PT in the vitamin K‐deficient model. Animals were administrated as follows: control (standard dry food), vitamin K‐deficient 14‐day, 21‐day group and 28‐day groups (rats received a vitamin K‐deficient diet and the intragastric administration of 30 mg/kg gentamicin for 14, 21 and 28 days respectively). The values are shown as the mean ± SE, n = 10. *P < 0.05, **P < 0.01 vs. control.

Figure 4

The changes of APTT in the model of vitamin K deficiency. Animals were administrated as follows: control (standard dry food), vitamin K‐deficient 14‐day, 21‐day and 28‐day groups (rats received a vitamin K‐deficient diet and the intragastric administration of 30 mg/kg gentamicin for 14, 21 and 28 days respectively). The values are shown as the mean ± SE, n = 10. **P < 0.01 vs. control.

Figure 5

Levels of coagulant factor activity in the vitamin K‐deficient model groups. a: FII activity; b: FVII activity; c: FIX activity; d: FX activity. Animals were administrated as follows: control (standard dry food), vitamin K‐deficient 14‐day, 21‐day and 28‐day groups (rats received a vitamin K‐deficient diet and the intragastric administration of 30 mg/kg gentamicin for 14, 21 and 28 days respectively). The values are shown as the mean ± SE, n = 10. *P < 0.05, **P < 0.01 vs. control.

Figure 6

Representative chromatograms of vitamin K from liver samples. a: liver sample of normal rats; b: normal liver sample with 250 ng/ml vitamin K1 standard substance; c: liver samples from rats with a vitamin K‐deficient diet and the intragastrical administration of 30 mg/kg gentamicin for 14 days; d: liver samples from rats with the diet‐ and gentamicin‐induced vitamin K deficiency for 21 days; e: liver samples from rats with a diet‐ and gentamicin‐induced vitamin K deficiency for 28 days.

Figure 7

The effect of vitamin K1 on the coagulant levels in the vitamin K‐deficient model rats. a: The changes of PT and APTT; b: The changes of the coagulant factors activities. Animals were administrated as follows: control (standard dry food), VKDM group (rats received a vitamin K‐deficient diet and the intragastric administration of 30 mg/kg gentamicin for 28 days); VKDM/VK 0.01 mg/kg and 0.1 mg/kg group (vitamin K‐deficient 28‐day rats were intravenously administered vitamin K1 injections 0.01 mg/kg and 0.1 mg/kg in the last 7 days respectively). The values are shown as the mean ± SE, n = 10. **P < 0.01 vs. control; ^# P < 0.05, ^## P < 0.01 vs. VKDM.