河南中医药大学
河南省科技研发计划联合基金(优势学科培育类)项目(232301420020)
Henan University of Chinese Medicine
Scientific and Technological Research and development plan of Henan province (232301420020)
目的 比较不同剂量博来霉素(Bleomycin, BLM)气管内滴注诱导的大鼠肺纤维化(pulmonary disease, PF)模型的成功率及稳定性。方法 150只SD大鼠随机分为对照组、BLM 3 mg/kg低剂量组(BL-L组)和BLM 5 mg/kg高剂量组(BL-H组),造模后观察大鼠一般状态、死亡情况、体重变化,第28、42、56、84天检测大鼠深吸气量(inspiratory capacity, IC)、肺活量(vital capacity, VC)、肺准静态顺应性(chord compliance, Cchord)和肺动态顺应性(dynamic compliance, Cdyn),记录肺系数,HE、MASSON染色观察肺组织病理变化,免疫组化检测肺组织胶原蛋白III(collagen type III, COL III)表达。结果 (1) 死亡率:BL-L组与BL-H组的死亡率分别为20%和28%。(2) 体重:BL-L组体重在第0~56天低于对照组,且第14天体重下降最为明显(P < 0.01),第56天以后体重恢复。第0~56天BL-H组体重显著低于对照组和BL-L组(P < 0.01),第56天以后体重增长仍较为缓慢。(3) 肺功能:与对照组比较,第28天,BL-L组肺功能IC、VC、Cchord、Cdyn相显著降低(P < 0.01, P < 0.05),BL-H组IC和Cdyn显著下降(P < 0.05),第42天,BL-L组IC、VC、Cchord显著下降(P < 0.01, P < 0.05),BL-H组IC、VC、Cchord、Cdyn显著下降(P < 0.01, P < 0.05),且IC、VC、Cchord与BL-L组相比显著降低(P < 0.01),第56天,BL-H组Cchord显著低于对照组和BL-L组(P < 0.01),第84天,各组肺功能无明显差异。(4) 肺系数:第28天,BL-L组和BL-H组肺系数显著高于对照组(P < 0.01),第42~56天,BL-H组肺系数显著高于BL-L组和对照组(P < 0.01),第84天,各组肺系数无明显差异。(5) 肺组织病理和免疫组化:BL-L组在第28~56天可观察到炎性浸润、纤维化条索和COL III表达,病变严重程度随时间递减,第84天纤维化病变几乎消失,BL-H组在28~84天均可观察到显著纤维化病变,COL III表达程度显著高于对照组(P < 0.01)。结论 3 mg/kg与5 mg/kg的BLM均可成功制备PF大鼠模型,3 mg/kg BLM制备的PF模型28天纤维化形成,持续至42天,后逐渐恢复；5 mg/kg BLM制备的PF模型28天纤维化形成,纤维化程度较低剂量严重,纤维化病变可稳定持续到56天,至84天仍有半数存在中重度纤维化。
Objective: To compare the success rate and stability of rat pulmonary fibrosis (PF) models induced by intratracheal instillation of different doses of bleomycin (BLM). Methods: 150 SD rats were randomly divided into control group, low-dose BLM 3 mg/kg group (BL-L group), and high-dose BLM 5 mg/kg group (BL-H group). The general status, mortality, and weight changes of the rats were observed, the inspiratory capacity (IC), vital capacity (VC), chord compliance (Cchord) and dynamic compliance (Cdyn) of the lungs were detected on the 28th, 42nd, 56th and 84th days. Lung coefficients were recorded, pathological changes in lung tissue were observed by HE and MASSON staining, and collagen type III (COL III) expression in lung tissue was detected by immunohistochemistry. Results: (1) Mortality rate: The mortality rates of BL-L group and BL-H group were 20% and 28%. (2) Weight: The weight of the BL-L group was lower than that of the control group on days 0-56, and the weight loss was most significant on day 14 (P < 0.01), with weight recovery after day 56. From day 0 to day 56, the weight of the BL-H group was significantly lower than that of the control group and BL-L group (P < 0.01), and the weight gain remained relatively slow after day 56. (3) Lung function: compared with the control group, on day 28, the lung function of IC, VC, Cchord, and Cdyn were significantly lower in the BL-L group (P < 0.01, P < 0.05), and IC and Cdyn were significantly lower in the BL-H group (P < 0.01). On day 42, IC, VC, and Cchord decreased significantly in the BL-L group (P < 0.01, P < 0.05), while IC, VC, Cchord, and Cdyn significantly decreased in the BL-H group (P < 0.01, P < 0.05), and IC, VC, and Cchord were significantly lower compared with those in the BL-L group (P < 0.01). On day 56, Cchord was significantly lower in the BL-H group than in the control and BL-L groups (P < 0.01), and on day 84, there was no significant difference in lung function between the groups. (4) Lung coefficient: on day 28, the lung coefficients of the BL-L and BL-H groups were significantly higher than those of the control group (P < 0.01), and from day 42 to 56, the lung coefficients of the BL-H group were significantly higher than those of the BL-L group and the control group (P < 0.01), and on day 84, the lung coefficients of the groups did not differ significantly from those of the control group. (5) Lung histopathology and immunohistochemistry: inflammatory infiltration, fibrotic streaks and COL III expression were observed in the BL-L group from day 28 to 56, with the severity of the lesions decreasing over time, and the fibrotic lesions had almost disappeared on day 84, while in the BL-H group, significant fibrotic lesions could be observed from day 28 to 84, and the degree of COL III expression was significantly higher than that in the control group (P < 0.01). Conclusions: Both 3 mg/kg and 5 mg/kg BLM can successfully induce PF rat models. The PF model prepared with 3 mg/kg BLM developed fibrosis on day 28, which lasted until day 42 and gradually recovered. The PF model prepared with 5 mg/kg BLM showed fibrosis formation on day 28, and the degree of fibrosis was more severe than in the lower dose, with fibrotic lesions persisting stably up to day 56, and moderate-to-severe fibrosis still present in half of the cases until day 84.
