Effects of intravenous immunoglobulin upon the overexpression and over-activation of nuclear factor-κB and matrix metalloproteinase-9 in murine model of Kawasaki disease

Abstract

To explore the changes of expression and biological activity of nuclear factor-kappa B (NF-κB) and matrix metalloproteinase-9 (MMP-9) after using intravenous immunoglobulin (IVIG) in a murine model of Kawasaki disease (KD) and elucidate the therapeutic mechanism of IVIG for the treatment of KD. A total of 72 mice were categorized randomly into IVIG, KD and control groups.Lactobacillus casei cell wall extract (LCWE) was prepared and injected intraperitoneally into C57BL/6 mice to induce KD (0.5 mg single injection).IVIG group received an intraperitoneal injection of IVIG (2 mg/g) while KD model group had an intraperitoneal injection of normal saline. At Days 14, 28 and 56, the diameter of coronary artery was by echocardiography in 8 mice of each group. At the same time, the stains of hematoxylin & eosin and elastic fiber were used to observe the pathological damage of coronary artery. Western blot was used to evaluate the expressions of NF-κB and MMP-9, electrophoretic mobility shift assay (EMSA) was used to measure the activity of NF-κB and Gelatin zymography was used to evaluate the activity of MMP-9 in heart samples of murine model of KD. The local inflammatory infiltrate, composed predominantly of mononuclear lymphocytes, of coronary artery trunk and branches was observed at Days 14 and 28 while broken elastin was observed at Day 56. And the inflammatory cell infiltrate was less severe and no apparent broken elastin was observed in IVIG and control groups. On echocardiography, the average value of diameter of left coronary artery in KD model group was higher than that in IVIG and control groups (28 d:(0.48 ± 0.07) vs (0.41 ± 0.03) and (0.35 ± 0.02) mm, all P < 0.01). Compared with the other two groups, the result of Western blot showed that the expressions of NF-κB and MMP-9 in KD model group were markedly higher than those in IVIG treatment group and that in control group at each time point (28 d: (58 ± 14) vs (25 ± 14) & (19 ± 11) µg/L, (100 ± 41) vs (39 ± 19) & (35 ± 19) µg/L, all P < 0.01). The activity of NF-κB by EMSA and the result from KD model group were much higher than those from the control and IVIG groups (28 d: (84 788 ± 2 081) vs (27 220 ± 4 990) & (50 192 ± 1 586) µg/L, all P < 0.01]. And it was in accord with the expression of NF-κB. The outcome of gelatin zymography demonstrated that the activity of MMP-9 had similar change with the expression of MMP-9(18 560 ± 7 963) vs (9 112 ± 3 398) & (11 834 ± 4 996) µg/L, all P < 0.05). NF-κB/MMP-9 is overexpressed and over-activated in the heart of KD mouse models. IVIG may inhibit the inflammatory cell infiltration and alleviate coronary artery. And such a therapeutic effect is possibly achieved by a suppression of the overexpression and over-activation of NF-κB/MMP-9 pathway.