CAJ | 학술논문

目的研究抗Ⅳ型胶原酶单链抗体scFv(3G11)对博莱霉素(BLM)诱导的小鼠肺纤维化的防治作用.方法 C57BL/6小鼠分为空白对照组、盐水对照组、BLM组和低、中、高剂量scFv(3G11)组.BLM组气管内注射BLM 400 ng/g;低、中、高剂量scFv(3G11)组注射BLM当天记为0 d,第1～7天每天腹腔注射scFv(3G11) 15、30、45 μg/g.第21天处死盐水对照组、BLM组和中剂量scFv(3G11)组小鼠各6只,取左肺行组织病理学观察,并测定肺泡隔面积比和有核细胞计数;另处死空白对照组、盐水对照组、BLM组和低、中、高剂量scFv(3G11)组小鼠各6只,取肺组织测定羟脯氨酸含量以衡量肺胶原沉积状况.取小鼠腹腔巨噬细胞进行原代培养,采用明胶酶谱法观察不同浓度scFv(3G11)(0、15、30、45、60 μmol/L)对巨噬细胞分泌基质金属蛋白酶2(MMP-2)和MMP-9的影响.结果组织病理学观察显示scFv(3G11)组小鼠肺纤维化程度明显轻于BLM组,肺泡隔面积比(45.3%± 3.2%)和有核细胞计数(451 ± 47)均明显低于BLM组(59.0% ± 3.0%,599 ± 42,均P<0.01).中、高剂量scFv(3G11)组小鼠肺组织羟脯氨酸含量[分别为(0.82±0.05)和(0.80±0.03)μg/mg]明显低于BLM组[(0.92±0.07)μg/mg,P<0.05,P<0.01].明胶酶谱分析显示,scFv(3G11)浓度为15 μmol/L时即可抑制巨噬细胞分泌的MMP-2和MMP-9的活性.结论抗Ⅳ型胶原酶单链抗体scFv(3G11)能抑制BLM诱导的小鼠肺纤维化.
목적 연구항Ⅳ형효원매단련항체scFv(3G11)대박래매소(BLM)유도적소서폐섬유화적방치작용.방법 C57BL/6소서분위공백대조조、염수대조조、BLM조화저、중、고제량scFv(3G11)조.BLM조기관내주사BLM 400 ng/g;저、중、고제량scFv(3G11)조주사BLM당천기위0 d,제1～7천매천복강주사scFv(3G11) 15、30、45 μg/g.제21천처사염수대조조、BLM조화중제량scFv(3G11)조소서각6지,취좌폐행조직병이학관찰,병측정폐포격면적비화유핵세포계수;령처사공백대조조、염수대조조、BLM조화저、중、고제량scFv(3G11)조소서각6지,취폐조직측정간포안산함량이형량폐효원침적상황.취소서복강거서세포진행원대배양,채용명효매보법관찰불동농도scFv(3G11)(0、15、30、45、60 μmol/L)대거서세포분비기질금속단백매2(MMP-2)화MMP-9적영향.결과 조직병이학관찰현시scFv(3G11)조소서폐섬유화정도명현경우BLM조,폐포격면적비(45.3%± 3.2%)화유핵세포계수(451 ± 47)균명현저우BLM조(59.0% ± 3.0%,599 ± 42,균P<0.01).중、고제량scFv(3G11)조소서폐조직간포안산함량[분별위(0.82±0.05)화(0.80±0.03)μg/mg]명현저우BLM조[(0.92±0.07)μg/mg,P<0.05,P<0.01].명효매보분석현시,scFv(3G11)농도위15 μmol/L시즉가억제거서세포분비적MMP-2화MMP-9적활성.결론 항Ⅳ형효원매단련항체scFv(3G11)능억제BLM유도적소서폐섬유화.
Objective To study the effect of anti-type Ⅳ collagenase single-chain antibody scFv(3G11) on bleomycin (BLM)-induced pulmonary fibrosis. Methods C57BL/6 mice were divided into six groups (blank, saline control, BLM, low-dose treatment, intermediate-dose treatment, and high-dose treatment), the animal model of pulmonary fibrosis was induced with 400 ng/g intratracheal BLM. The day that treatment groups were injected with BLM was marked Day 0. Then at Days 1-7, scFv(3G11) was injected once intraperitoneally each day with three dosage [low-dose treatment (15 μg/g), intermediate-dose treatment (30 μg/g) and high-dose treatment (45 μg/g)]. At Day 21, 6 mice of each group (saline control, BLM, intermediate-dose treatment) were sacrificed and pathomorphological changes of left lungs evaluated with HE stained sections. Meanwhile, 6 mice of each group (blank, saline control, BLM, low-dose treatment, intermediate-dose treatment, and high-dose treatment) were sacrificed, the content of hydroproline was detected to measure the degree of collagen deposition. Macrophages were extracted from murine abdominal cavity and primarily cultured. And the inhibition of matrix metalloproteinase (MMP)-2 and MMP-9 excretion by different concentrations of scFv(3G11)(0, 15, 30, 45, and 60 μmol/L)was determined by gelatin zymography.Results By image analysis, the degree of fibrosis in the lungs of treated group was lighter than that of the BLM group. The percentages of the area of interalveolar septum (45.3% ± 3.2%) and the number of nucleated cells (451 ± 47) of treated group were remarkably reduced as compared with the BLM group(59.0% ± 3.0%, 599 ± 42, both P<0.01). The content of hydroproline of the intermediate-dose treatment group [(0.82±0.05) μg/mg] and high-dose treatment group [(0.80±0.03) μg/mg] was lower than that of the BLM group [(0.92±0.07) μg/mg,P<0.05,P<0.01]. The result of gelatin zymography demonstrated that the excretion of MMP-2 and MMP-9 by macrophage was inhibited by scFv(3G11) at the concentration of 15 μmol/L.Conclusion Single-chain antibody directed against type Ⅳ collagenase might inhibit the development of bleomycin-induced pulmonary fibrosis.