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目的观察透明质酸合成酶1(HAS1)在脑出血后脑内透明质酸(HA)含量和相对分子质量变化过程中的作用。方法采用自体血注射法制作HAS1基因敲除小鼠和野生型C57BL/6J小鼠脑出血模型。分别分为出血后24 h组、出血后72 h组和对照组(不注自体血,余同),每组均n=6。提取出血侧半球脑组织的HA,经凝胶层析分离后用ELISA方法检测HA的总量和相对分子质量分布。Real time PCR方法观察HAS1、HAS2、HAS3和透明质酸分解酶(Hyal)mRNA表达水平的变化。通过对野生型C57BL/6J小鼠和HAS1基因敲除小鼠的比较,评价HAS1在此过程中的作用。结果脑出血后,①野生型C57BL/6J小鼠脑HA含量基本不变,仅在出血后72 h组呈轻微下降,但HA相对分子质量分布有显著变化,出血后24 h组呈现一过性的大分子HA的比例增加。②HAS1基因敲除小鼠脑内HA总量比野生型C57BL/6J小鼠减少1/4。出血后72 h组的HA总量明显下降,而且出血后24 h组的大分子HA比例不升反降,至出血后72 h后也未见恢复。③出血后,基因表达均随时间而增加,增幅大小顺序为HAS2>HAS3>HAS1,Hyal1>Hyal3>Hyal2。野生型C57BL/6J和HAS1基因敲除小鼠间只有HAS3的表达有差异。结论 HAS1基因敲除导致脑内HA总量减少1/4,脑出血后24 h HAS3基因表达代偿性增加,导致脑出血后脑内大分子HA含量下降。
Objective To investigate the role of hyaluronan synthase 1 (HAS1) in the changes of intracellular hyaluronic acid (HA) content and relative molecular mass after intracerebral hemorrhage. Methods The cerebral hemorrhage model of HAS1 knockout mice and wild type C57BL / 6J mice was made by autologous blood injection. Divided into 24 h after hemorrhage group, 72 h after hemorrhage group and control group (without autologous blood, Yu Tong), n = 6 in each group. The HA of hemispheric hemisphere was extracted from the hemisphere, and the total amount of HA and the relative molecular mass distribution of HA were detected by ELISA after separated by gel chromatography. Real time PCR method was used to observe the changes of mRNA expression of HAS1, HAS2, HAS3 and Hyal. The role of HAS1 in this process was evaluated by comparison of wild-type C57BL / 6J mice and HAS1 knockout mice. Results After intracerebral hemorrhage, ① The content of HA in wild-type C57BL / 6J mice remained unchanged, but slightly decreased at 72 h after hemorrhage, but the molecular mass distribution of HA changed significantly. After 24 h, The proportion of macromolecules HA increased. ② The total amount of HA in the brain of the HAS1 knockout mice was reduced by 1/4 compared with the wild type C57BL / 6J mice. The amount of HA decreased significantly at 72 h after hemorrhage, and the percentage of HA increased at 24 h after hemorrhage, but not recovered at 72 h after hemorrhage. ③ After hemorrhage, gene expression increased with time, the order of increase was HAS2> HAS3> HAS1, Hyal1> Hyal3> Hyal2. HAS3 expression differs only between wild type C57BL / 6J and HAS1 knockout mice. Conclusion HAS1 gene knockdown leads to a decrease of 1/4 of total HA in brain and HAS3 gene expression 24 h after intracerebral hemorrhage, which leads to a decrease of intracerebral macrophages HA content after intracerebral hemorrhage.