Rong Xing, Xiang Ru-lian, Chu Mao-ping, Wu Rong-zhou, Chen Qi, Xu Qiang, Zhang Yuan-hai
Department of Cardiovascular Diseases, Yuying Children's Hospital Affiliated to Wenzhou Medical College, Wenzhou 325000, China.
Zhonghua Er Ke Za Zhi. 2007 Dec;45(12):893-7.
To investigate the role of heme oxygenase-1 (HO-1) and its catalyst carbon monoxide (CO) in the development of myocardial damage and the effects of zinc protoporphyrin IX (ZnPPIX), an inhibitor of HO-1 on myocardium of mice with acute viral myocarditis.
A total of 112 inbred male Balb/C mice 4 - 6 weeks of age were divided randomly into 3 groups: the control group (C group, n = 32), the viral myocarditis group (V group, n = 40) and ZnPPIX group (Z group, n = 40). The Z and V groups were inoculated intraperitoneally (i.p.) with 0.1 ml of 10(-4.36) tissue culture infectious dose 50% (TCID(50))/ml Coxsackie virus B3 (CVB(3)) to produce viral myocarditis model on day 0, C group was injected i.p. with virus-free 1640 culture culture medium 0.1 ml at the same time, then operation was done as follows: the mice of group C and group V were injected i.p. with 0.1 ml NS each day. The mice of group Z were injected i.p. with 40 micromol per kilogram of body weight ZnPPIX (HO-1 inhibitor) qod. Eight mice of each group were sacrificed on days 4, 8, 15 and 21, respectively. The blood specimens were collected by taking out the eyeballs to test for the content of carboxyhemoglobin (COHb) using spectrophotometry and cardiac troponin I (cTnI) using chemiluminescent immunoassay. The hearts tissue slides were also stained by immunohistochemistry (IHC) for HO-1 and in situ hybridization (ISH) for HO-1 mRNA. The histological and ultrastructural changes were observed under light and electron microscopes.
(1) The histopathological changes of myocardial cells: in the V and Z groups myocardial inflammatory cells infiltration reached the peak on day 8, the Z group histopathological scores were significantly lower than those in V group on day 8 (2.40 +/- 0.31 vs. 1.73 +/- 0.24, P < 0.01) and on day 15 (1.78 +/- 0.29 vs. 1.43 +/- 0.23, P < 0.05). No inflammation was present in group C. (2) The changes of serum cTnI level in both V and Z groups were significantly higher than those in C group on day 4, 8 and 15 (P < 0.01). The level in Z group was significantly lower than that in V group on day 4 [(6.074 +/- 1.475) ng/ml vs (7.911 +/- 1.225) ng/ml, P < 0.05] and day 8 [(0.821 +/- 0.294) ng/ml vs (1.480 +/- 0.454) ng/ml, P < 0.05]. (3) The changes of blood COHb level: compared with V group, in Z group the COHb level was lower on day 4 (P < 0.05) and day 15 (P < 0.01) after CVB(3) inoculation. Surprisingly, in Z group COHb level elevated suddenly on day 8 and showed conspicuously higher than that of V group (P < 0.01). (4) The result of HO-1 IHC staining: in both V and Z group myocardial cells had positive expression, while C group did not. (5) The results of HO-1 ISH were similar to those of HO-1 IHC, the A values of group Z was significantly lower than that of group V on day 4, 15 and 21(P < 0.01), but on day 8 it was higher than that of group C (P < 0.05).
HO-1 inhibitor, ZnPP not only could inhibit HO-1 overexpression but also could induce HO-1 expression temporarily and protect against myocardial injury at the early stage of acute viral myocarditis.
探讨血红素加氧酶-1（HO-1）及其催化产物一氧化碳（CO）在心肌损伤发生发展中的作用，以及HO-1抑制剂锌原卟啉IX（ZnPPIX）对急性病毒性心肌炎小鼠心肌的影响。
将112只4-6周龄的近交系雄性Balb/C小鼠随机分为3组：对照组（C组，n = 32）、病毒性心肌炎组（V组，n = 40）和ZnPPIX组（Z组，n = 40）。Z组和V组于第0天腹腔注射0.1 ml含10(-4.36)组织培养感染剂量50%（TCID(50)）/ml柯萨奇病毒B3（CVB(3)）的病毒液制备病毒性心肌炎模型，C组同时腹腔注射0.1 ml无病毒的1640培养液，然后按以下方法处理：C组和V组小鼠每天腹腔注射0.1 ml生理盐水；Z组小鼠每隔一天腹腔注射40 μmol/kg体重的ZnPPIX（HO-1抑制剂）。分别于第4、8、15和21天每组处死8只小鼠，摘眼球取血标本，采用分光光度法检测碳氧血红蛋白（COHb）含量，采用化学发光免疫分析法检测心肌肌钙蛋白I（cTnI）。心脏组织切片行HO-1免疫组织化学（IHC）染色及HO-1 mRNA原位杂交（ISH）。在光镜和电镜下观察组织学和超微结构变化。
（1）心肌细胞组织病理学变化：V组和Z组心肌炎性细胞浸润在第8天达高峰，Z组在第8天（2.40±0.31 vs. 1.73±0.24，P < 0.01）和第15天（1.78±0.29 vs. 1.43±0.23，P < 0.05）的组织病理学评分显著低于V组。C组无炎症。（2）V组和Z组血清cTnI水平在第4、8和15天均显著高于C组（P < 0.01）。Z组在第4天[（6.074±1.475）ng/ml vs（7.911±1.225）ng/ml，P < 0.05]和第8天[（0.821±0.294）ng/ml vs（1.480±0.454）ng/ml，P < 0.05]的水平显著低于V组。（3）血COHb水平变化：与V组相比，Z组在CVB(3)接种后第4天（P < 0.05）和第15天（P < 0.01）的COHb水平较低。令人惊讶的是，Z组COHb水平在第8天突然升高，且显著高于V组（P < 0.01）。（4）HO-1 IHC染色结果：V组和Z组心肌细胞均有阳性表达，C组无。（5）HO-1 ISH结果与HO-1 IHC结果相似，Z组在第4、15和21天的A值显著低于V组（P < 0.01），但在第8天高于C组（P < 0.05）。
HO-1抑制剂ZnPP不仅能抑制HO-1的过度表达，还能在急性病毒性心肌炎早期暂时诱导HO-1表达并保护心肌免受损伤。
