大鼠脑出血灶周围小胶质细胞极化的动态变化

The dynamic changes of microgliai polarization around intracerebral hematoma in rats

目的探讨大鼠脑出血灶周围小胶质细胞极化的动态变化规律，为干预小胶质细胞的极化或研究其极化机制提供时间依据。方法健康雄性SD大鼠，随机分成sham组、脑出血4h、1d、3d、7d、14d组，每组6只。采用尾状核注射胶原酶法制备大鼠脑出血模型，分别于术后对应时间点灌注取脑；sham组注射等量生理盐水，于术后1d灌注取脑。采用免疫荧光染色法标记小胶质细胞Ⅰ型（Microglia type Ⅰ，M1；CD11b＋＋CD86＋）、小胶质细胞Ⅱ型（Microglia type Ⅱ，M2；CD11b＋ ＋Arg-1＋），评价血肿周围两种细胞数量的动态变化情况。结果（1）脑出血4h后已能检测到M1、M2，此时M1仍有部分突起存在；（2）MI在脑出血急性期（1—3d）及亚急性早期（3—7d）及慢性期（〉14d）占主要地位；M2在脑出血超急性期（24h内）及亚急性期晚期（7d）短时间内占有优势，4h时M2数量[（31．40±1．69）个]高于MI[（21．43±1．81）个]，差异有统计学意义（t=-4．085，P=0．002），7d时M2数量[（116．25±5．06）个]第二次超过M1[（85．75±7．32）个]，差异有统计学意义（t=-0．690，P=0．001）。结论M1在脑出血急性期、亚急性早期及慢性期占主要地位，M2于超急性期与亚急性晚期占有优势；于超急性期（如4h）与亚急性晚期（如7d）探讨M2的形成机制或于急性期（1～3d）干预M1的形成对于脑出血的治疗具有重要的临床意义。

Objective To investigate the dynamic changes of microglial polarization at the perihematoma area and provide timepoint evidence for interventing microglial polarization as well as studying the polarization mechanism after intracerebral hemorrhage （ICH）. Methods Healthy male Sprague Dawley （SD） rats were randomly divided into sham group, ICH-4 h, 1 d, 3 d, 7 d and 14 d groups with 6 in each group. The rats in ICH groups were injected collagenase VII-s into the caudate nucleus to establish the intracerebral hematoma model and rats in sham operated group were treated with the same amount of saline. The brains were taken at 4 h, 1 d,3 d,7 d, 14 d in the ICH group, 1 d in sham group. Microglia type I （ M1, CD11b＋ ＋CD86＋） and microglia type Ⅱ （M2, CD11b＋ ＋Arg-1＋） were examined by immunofluorescence and the number of M1 and M2 around hematoma were analyzed. Results （ 1 ） The M1 and M2 were both ob- served at 4 h after ICH and a small quantity of branches were still presented on M1. （2） M1 took the main position in acute stage （1-3 d） ,early subacute stage（3-7 d） and chronic stage （〉14 d） after ICH.The number of M2 was elevated transiently in superacute （〈24 h） and late subacute stage （7 d）.The number of M2 （31.40±1.69） was more than MI （21.43±1.81） at 4 h after ICH （ t=- 4.085, P=0.002）,and the number of M2 （116.25±5.06） significantly exceeded MI （85.75±7.32） again on day 7 （ t= -0.690, P= 0.001 ）. Conclusion M1 is in a dominant position in acute, early subacute and chronic stages after ICH; M2 is dominant in superacute and late subaeute stages. Investigating the mechanism of M2 formation at acute period （such as 4 h） or late subacute stage （such as 7 d） ,and inhibiting M1 formation in the early subacute stage （ 1 - 3 d） have important significance for clinical treatment of ICH.