Figure 1.
Changes of body weight and biochemical parameters of each rat after simvastatin treatment.
Blood samples were collected after 10 days of simvastatin treatment (80 mg/kg). Serum alanine transaminase (ALT), aspartate transaminase (AST), and plasma creatinine kinase (CK) activities were measured using commercial kits employing spectrophotometric assays. All biomarkers were analyzed at Inha University Hospital (Incheon, Korea).
Figure 2.
Representative 1H NMR spectra of urine from animals before and after simvastatin treatment.
The spectrum before the simvastatin treatment is at the top, and that after the treatment is at the bottom. Metabolite peaks were assigned using Chenomx (Spectral database; Edmonton, Alberta, Canada). The spectra were taken for urine samples containing 150 mM phosphate (pH 7.4) and 0.025% TSP as a chemical shift reference.
Figure 3.
Differentiation of pre- and post-treatment groups using multivariate analysis.
Orthogonal projections to latent structure-discriminant analysis (OPLS-DA) score plot of the pre- and post-groups. For the post-treatment group, urine samples collected after 10 days of treatment were used. Black squares: pre-treatment samples; Red circles: post-treatment samples. The model was established using one predictive and one orthogonal component.
Figure 4.
Signals contributing to the differentiation of pre- and post-treatment groups.
S-plot analysis representing the highest contributing signals for the pre- and post-groups. The pp represents modeled covariation, and p(corr)p represents modeled correlation. Potential marker signals that are significantly biased across the two groups are enclosed in dotted boxes.
Figure 5.
Levels of the marker signals in the pre- and post-treatment groups.
Student's t-test of the relative distribution of the marker signals for the pre- and post-treatment group. The resulting p-values are indicated, and all the signals showed statistical significance with p<0.01. (A) Top: 5.3916 ppm; (B) Middle: 3.0319 ppm; (C) Bottom: 3.2893 ppm.
Figure 6.
Difference between WT and HT subgroup: biochemical data.
The WT and HT subgroups among the simvastatin treated animals were categorized according to the OPLS-DA model as described in the text. Average values of AST, ALT and CK levels of each group after 10 days of treatment are plotted along with the standard deviation. Student's t-test was also performed and the resulting p values are also indicated. (A) AST; (B) ALT; (C) CK.
Figure 7.
Histopathological analysis of liver sections.
The liver tissues were collected after 10 days of simvastatin treatment. The tissues were fixed in 10% formaldehyde, and then stained using Hematoxylin and Eosin (H&E). Original magnification ×400. Control, control group; WT, weak toxicity group; HT, high toxicity group.
Figure 8.
Time course of the urine metabolite profile.
Urines collected at specified time points during the simvastatin treatment (80 mg/kg) were analyzed by NMR and OPLS-DA. The coordinates of each time point represent the average score values from the multivariate analysis. The whiskers represent one standard deviation. Open symbols represent the HT group and the filled symbols WT group. Each time point is represented by different colored symbols: Pre, black squares; 3 Day, red circles; 6 Day, blue diamonds; 10 Day, green triangles. The transitions for the 6 day to 10 day are specified by arrows. Others are not indicated to simplify the figure, but can be easily traced by following the open or filled symbols, respectively.