临床神经病学杂志
臨床神經病學雜誌
림상신경병학잡지
JOURNAL OF CLINICAL NEUROLOGY
2015年
2期
114-120
,共7页
张美增%逄芳芳%赵丽%常翠翠%郭云良
張美增%逄芳芳%趙麗%常翠翠%郭雲良
장미증%방방방%조려%상취취%곽운량
脑缺血%胡黄连苷Ⅱ%治疗时间窗%剂量%基质金属蛋白酶-9
腦缺血%鬍黃連苷Ⅱ%治療時間窗%劑量%基質金屬蛋白酶-9
뇌결혈%호황련감Ⅱ%치료시간창%제량%기질금속단백매-9
cerebral ischemia%picrosideⅡ%therapeutic time window%dose%matrix metalloproteinase-9
目的：探讨胡黄连苷Ⅱ对大鼠脑缺血性损伤和基质金属蛋白酶－9（ MMP－9）表达的影响，以及最佳治疗时间窗与剂量的选取。方法将174只Wistar大鼠随机分为脑缺血组、正常对照组和治疗组，按正交试验设计治疗组大鼠再分为缺血1．0 h、1．5 h、2．0 h、2．5 h，以及分别用胡黄连苷Ⅱ5 mg／kg、10 mg／kg、20 mg／kg和40 mg／kg亚组。用双侧颈总动脉结扎法制作脑缺血大鼠模型，治疗组大鼠按相应时间点和剂量经腹腔注射胡黄连苷Ⅱ。用HE染色观察脑变性细胞指数（ DCI）及形态；电镜观察脑组织超微结构。免疫组化染色和Western blot法检测脑组织MMP－9蛋白水平，逆转录（ RT）－PCR检测MMP－9 mRNA水平。结果脑缺血组大鼠脑组织损害和MMP－9表达水平明显高于正常对照组（均P＜0．05）。与脑缺血组比较，治疗组的脑组织损害及DCI和MMP－9表达水平明显减轻和降低（均P＜0．05）。治疗组各亚组间两两比较，缺血1 h亚组与2 h、2．5 h亚组间脑DCI的差异有统计学意义（均P＜0．05）；缺血2 h与2．5 h亚组间的脑MMP－9阳性细胞指数，缺血1 h与1．5 h、2 h亚组，1．5 h与2．5 h亚组，2 h与2．5 h亚组间的脑MMP－9蛋白水平的差异均有统计学意义（均P＜0．05）；以及剂量5 mg／kg与20 mg／kg亚组间的脑MMP－9 mRNA水平的差异有统计学意义（均P＜0．05）。各剂量亚组间的脑DCI、MMP－9阳性细胞指数和蛋白水平的差异均无统计学意义（均P＞0．05）。结论胡黄连苷Ⅱ能抑制大鼠脑缺血后MMP－9的表达，并有显著的脑保护作用。按治疗时间窗最大化和用药剂量最小化的原则，其最佳的治疗时间窗为脑缺血后2 h内，剂量为（10～20） mg／kg。
目的：探討鬍黃連苷Ⅱ對大鼠腦缺血性損傷和基質金屬蛋白酶－9（ MMP－9）錶達的影響，以及最佳治療時間窗與劑量的選取。方法將174隻Wistar大鼠隨機分為腦缺血組、正常對照組和治療組，按正交試驗設計治療組大鼠再分為缺血1．0 h、1．5 h、2．0 h、2．5 h，以及分彆用鬍黃連苷Ⅱ5 mg／kg、10 mg／kg、20 mg／kg和40 mg／kg亞組。用雙側頸總動脈結扎法製作腦缺血大鼠模型，治療組大鼠按相應時間點和劑量經腹腔註射鬍黃連苷Ⅱ。用HE染色觀察腦變性細胞指數（ DCI）及形態；電鏡觀察腦組織超微結構。免疫組化染色和Western blot法檢測腦組織MMP－9蛋白水平，逆轉錄（ RT）－PCR檢測MMP－9 mRNA水平。結果腦缺血組大鼠腦組織損害和MMP－9錶達水平明顯高于正常對照組（均P＜0．05）。與腦缺血組比較，治療組的腦組織損害及DCI和MMP－9錶達水平明顯減輕和降低（均P＜0．05）。治療組各亞組間兩兩比較，缺血1 h亞組與2 h、2．5 h亞組間腦DCI的差異有統計學意義（均P＜0．05）；缺血2 h與2．5 h亞組間的腦MMP－9暘性細胞指數，缺血1 h與1．5 h、2 h亞組，1．5 h與2．5 h亞組，2 h與2．5 h亞組間的腦MMP－9蛋白水平的差異均有統計學意義（均P＜0．05）；以及劑量5 mg／kg與20 mg／kg亞組間的腦MMP－9 mRNA水平的差異有統計學意義（均P＜0．05）。各劑量亞組間的腦DCI、MMP－9暘性細胞指數和蛋白水平的差異均無統計學意義（均P＞0．05）。結論鬍黃連苷Ⅱ能抑製大鼠腦缺血後MMP－9的錶達，併有顯著的腦保護作用。按治療時間窗最大化和用藥劑量最小化的原則，其最佳的治療時間窗為腦缺血後2 h內，劑量為（10～20） mg／kg。
목적：탐토호황련감Ⅱ대대서뇌결혈성손상화기질금속단백매－9（ MMP－9）표체적영향，이급최가치료시간창여제량적선취。방법장174지Wistar대서수궤분위뇌결혈조、정상대조조화치료조，안정교시험설계치료조대서재분위결혈1．0 h、1．5 h、2．0 h、2．5 h，이급분별용호황련감Ⅱ5 mg／kg、10 mg／kg、20 mg／kg화40 mg／kg아조。용쌍측경총동맥결찰법제작뇌결혈대서모형，치료조대서안상응시간점화제량경복강주사호황련감Ⅱ。용HE염색관찰뇌변성세포지수（ DCI）급형태；전경관찰뇌조직초미결구。면역조화염색화Western blot법검측뇌조직MMP－9단백수평，역전록（ RT）－PCR검측MMP－9 mRNA수평。결과뇌결혈조대서뇌조직손해화MMP－9표체수평명현고우정상대조조（균P＜0．05）。여뇌결혈조비교，치료조적뇌조직손해급DCI화MMP－9표체수평명현감경화강저（균P＜0．05）。치료조각아조간량량비교，결혈1 h아조여2 h、2．5 h아조간뇌DCI적차이유통계학의의（균P＜0．05）；결혈2 h여2．5 h아조간적뇌MMP－9양성세포지수，결혈1 h여1．5 h、2 h아조，1．5 h여2．5 h아조，2 h여2．5 h아조간적뇌MMP－9단백수평적차이균유통계학의의（균P＜0．05）；이급제량5 mg／kg여20 mg／kg아조간적뇌MMP－9 mRNA수평적차이유통계학의의（균P＜0．05）。각제량아조간적뇌DCI、MMP－9양성세포지수화단백수평적차이균무통계학의의（균P＞0．05）。결론호황련감Ⅱ능억제대서뇌결혈후MMP－9적표체，병유현저적뇌보호작용。안치료시간창최대화화용약제량최소화적원칙，기최가적치료시간창위뇌결혈후2 h내，제량위（10～20） mg／kg。
Objective To explore the effect of picrosideⅡon cerebral ischemic injury and expression of matrix metalloproteinase-9 ( MMP-9) in rats, and the selection of the best time window and dose of treatment.Methods A total of 174 Wistar rats were randomly divided into cerebral ischemia group, normal control group and treatment group.The rats in treatment group were randomly divided into four subgroups with therapeutic time window designed:1.0 h, 1.5 h, 2.0 h, 2.5 h after ischemia, and four subgroups with therapeutic drug picroside Ⅱ dose which has four levels as follows:5 mg/kg, 10 mg/kg, 20 mg/kg, 40 mg/kg body weight according to the principle of orthogonal experimental design.The forebrain ischemia models were established by bilateral common carotid artery occlusion ( BCCAO) methods.The rats in treatment group were injected intraperitoneally according to the corresponding doses designed in the orthogonal layout with picrosideⅡ.The morphology of nerve cell and the denatured cell index( DCI) were observed by HE staining.The ultrastructure of brain tissue was observed by transmission electron microscopy (TEM).The expression of MMP-9 protein was determined by immunohistochemical staining and Western blot method.The expression of MMP-9 mRNA was detected by reverse transcription-polymerase chain reaction ( RT-PCR).Results The cerebral ischemic injury and the expression of MMP-9 in cerebral ischemia group rats increased significantly compared with those in normal control group ( all P<0.05 ) , and the cerebral ischemic injury and the expression of MMP-9 in treatment group were alleviated and reduced obviously compared with cerebral ischemia group ( all P<0.05 ) . All data from treatment group was analyzed by the least significant difference for pairwise comparisons and the results showed that there was significant deviation of DCI between ischemia 1.0 h and 2.0 h, and ischemia 1.0 h and 2.5 h(all P<0.05).Also there was significant deviation of the positive cell index of MMP-9 between ischemia 2.0 h and 2.5 h,MMP-9 protein expression between ischemia 1.0 h and 1.5 h, ischemia 1.0 h and 2.0 h, ischemia 1.5 h and 2.5 h,and ischemia 2.0 h and 2.5 h(all P<0.05).There was significant deviation of the expression of MMP-9 mRNA between therapeutic dose 5 mg/kg and 20 mg/kg(all P<0.05).And there were no significant deviation of DCI, positive cell index of MMP-9 and MMP-9 protein expression between different therapeutic doses( all P>0.05 ) .Conclusions Picroside Ⅱ can inhibit the expression of MMP-9 after cerebral ischemic injury in rats, having a significant cerebral protection effect.Considering the minimization of medication dose and maximization of therapeutic time, the best time window of picrosideⅡis within 2 h after cerebral ischemia, and the therapeutic dose is(10-20) mg/kg.
