人肥大细胞类糜蛋白酶的促血管生成作用

The Role of Human Mast Cell Chymase in Angiogenesis

目的探讨人肥大细胞类糜蛋白酶(mast cell chymase,MC-Chy)的促进血管生成作用。方法将30只BALB/c小鼠随机分为6组,每组5只。4组实验组小鼠分别予腹股沟部皮下注射基质胶与MC-Chy质粒DNA50μg(Chy50组)、100μg(Chy100组)、150μg(Chy150组)、200μg(Chy200组)的混合物;抑制剂组小鼠给予MC-Chy质粒DNA150μg与基质胶的混合物腹股沟部皮下注射,然后每天注射类糜蛋白酶抑制剂chymostatin10μg至皮下的基质胶栓;对照组小鼠行腹股沟部皮下注射生理盐水及基质胶混合物。注射后第7天,以同样剂量的质粒DNA注射至各实验组小鼠皮下的基质胶栓,于第12天处死小鼠,取出胶栓,测定MC-Chy活性及血红蛋白浓度,并进行免疫组化学染色、HE染色,测量血管密度计数。结果 Chy50组(7.5±1.4)、Chy100组(21.7±2.7)、Chy150组(28.4±2.1)、Chy200组(25.8±2.0)血红蛋白浓度与对照组(2.9±0.7)比较差异均显著(P<0.05,P<0.01,P<0.01,P<0.01)。Chy50组(9.3±2.7)、Chy100组(12.2±4.0)、Chy150组(15.4±3.1)、Chy200组(13.5±3.6)血管密度与对照组(5.9±2.7)比较,均存在显著性差异(均P<0.01)。HE染色可见到血管,免疫组化染色可见到棕黄色阳性颗粒及血管腔。结论人MC-Chy具有促进血管生成的作用。

Objective To investigate the role of the human mast cell chymase in angiogenesis. Methods Thirty BALB/c mice were randomly divided into 6 groups, 5 mice in each group. Groin subcutaneous injection of the mixtures of matrigel and MC-Chy plasmid DNA 50 μg （ chyS0 group ）, 100 μg （ chyl00 group ）, 150 μg （ chy150 group）, 200 μg （ chy200 group） were given to 4 groups of experimental mice respectively, while subcutaneous injection of the mixtures of matrigel and MC-Chy plasmid DNA 150 μg and followed daily subcutaneous injection MC-Chy inhibitor chymostatin （CHI） 10μg into the matrigel plug were given to inhibitor group. Groin subcutaneous injections of the mixtures of saline and matrigel were given to control group. The MC-Chy plasmid DNA was injected into the subcutaneous matrigel plug of mice in each groups in same doses in day 7 after the injection. Mice were sacrificed, and the matrigel plug was removed in day12. The detection of chymase activity and hemoglobin （ Hb ） concentration, HE staining, immunohistochemical staining and measurement of microvessel density （MVD） were performed in the matrigel plug. Results HB concentration of chy50 group （7.5 ± 1.4）, chyl00 group （21.7 ± 2.7 ）, chyl50 group （28.4 ±2. 1 ）, chy200 group （25.8 ±2. 0） were significant higher than control group （2. 9 ± 0.7）, the differences were statistically significant （P 〈 0.05, P 〈 0.01, P 〈 0.01, P 〈 0.01 respectively） . The MVD levels of Chy50 group （9.3 ± 2.7）, Chyl00 group （ 12.2 ± 4.0）, Chyl50 group （ 15.4 ± 3.1 ）, Chy200 group （ 13.5 ± 3.6 ） were significant higher than control group （ 5.9± 2.7） , the differences were statistically significant （ all P 〈 0. 01 ）. Microvessels were observed under HE staining, and brown positive particles and vessel lumen were shown under Immunohistochemical staining. Conclusions Angiogenesis can be elicited by human mast cell chymase.