Involvement of p53, p21, and Caspase-3 in Apoptosis of Coronary Artery Smooth Muscle Cells in a Kawasaki Vasculitis Mouse Model

Abstract

BACKGROUND Overexpression of p53, p21, and caspase-3 promotes apoptosis of vascular smooth muscle cells. However, the mechanisms that lead to apoptosis of coronary artery smooth muscle cells (CASMCs) is unclear in Kawasaki disease (KD). This study investigated involvement of p53, p21, and caspase-3 in the apoptosis of CASMCs from a Kawasaki vasculitis mouse model. MATERIAL AND METHODS The Kawasaki vasculitis mouse model with coronary artery lesions was generated via administration of Lactobacillus casei cell wall extract. In 2 groups of mice (healthy control and KD vasculitis mice), the levels of p53, p21, and caspase-3 protein in the root of the coronary artery were evaluated via immunohistochemistry. Receiver operating characteristic curves were plotted for determination of area under the curve, 95% confidence interval, sensitivity, specificity, and cutoff values for the ability of p53, p21, and caspase-3 expression to predict CASMC apoptosis and coronary artery lesion formation in KD vasculitis mice. RESULTS Compared with healthy mice, KD vasculitis mice had a significantly higher apoptosis index and upregulated p53, p21, and caspase-3 expression. Also, the immunoreactive score for caspase-3 was positively correlated with the immunoreactivity scores for p53 and p21. The optimal cutoff values for p53, p21, and caspase-3 expression for predicting the presence of coronary artery lesions were 4.15, 4.18, and 4.22, respectively. CONCLUSIONS Upregulated levels of p53, p21, and caspase-3 promoted apoptosis of CASMCs in KD vasculitis mice. Thus, the levels of p53, p21, and caspase-3 may serve as valuable predictors of coronary artery lesion formation in KD.

Figure 1

Pathological features of the coronary artery root in KD (A) and control (B) groups. The root of the coronary artery obtained from the KD vasculitis mice exhibited accumulation of foam cells and inflammatory cell infiltration, which were accompanied by dissolved and necrotic CASMCs, as arrows pointing to the positions. These pathological phenotypes were not present in the coronary artery of the control mice.

Figure 2

Comparison of p53 (A, C) and p21 (B, C) IRS values between the KD (n=12) and control groups (n=12). Cytoplasmic staining of p53 in CASMCs was moderate and strong in the mice of the KD group, whereas only mild or no staining was observed in mice of the control group (A). Cytoplasmic staining for p21 in CASMCs was moderate and strong in mice of the KD group, but mild or absent in mice of the control group (B). The IRS values for p53 and p21 were significantly different between the KD group and control group (C).

Figure 3

Comparison of Bcl2 (A, C) and caspase-3 (B, C) IRS values between the KD (n=12) and control groups (n=12). Comparison of Bcl2 (A) and caspase-3 (B) IRS values in the KD group and control groups. Cytoplasmic staining of Bcl2 in CASMCs was mild in mice of the KD group but was strong or moderate in mice of the control group (A). Cytoplasmic staining for caspase-3 in CASMCs was strong and moderate in mice of the KD group but mild or absent in mice of the control group (B). The IRS values for Bcl2 and caspase-3 were significantly different between the KD group and the control group (C).

Figure 4

Correlation of p53 and p21 IRS values with the caspase-3 IRS in the KD group. Pearson correlation analysis revealed that the IRS for caspase-3 was positively correlated with the IRSs for p53 and p21 in the KD group (r=4.67 and 4.83; all P<0.001).

Figure 5

Comparison of ROC curves for p53, p21, and caspase-3 in all KD vasculitis mice with CALs. The AUC values and 95% CIs for p53, p21, and caspase-3 expression. The optimal cutoff values for p53, p21, and caspase-3 expression were 4.15, 4.18, and 4.22, respectively.