A Comparative Study on the Establishment of Animal Models of Chronic Renal Failure by Two Methods
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摘要:
目的 分别使用2种方法建立慢性肾衰竭（chronic renal failure，CRF）大鼠模型，对比2种模型的稳定性，旨在为建立一种稳定性高的CRF动物模型提供依据。 方法 分别使用单侧肾切除+腺嘌呤灌胃+高磷饲料喂养法、高磷+腺嘌呤饲料喂养法建立CRF大鼠模型。于造模结束时及造模结束后6周检测模型动物的红细胞、血红蛋白，血清尿素、肌酐，并行肾脏、骨组织病理学检查，计算肾小管间质纤维化指数（tubulointerstitial fibrosis index，TBI）、肾组织胶原纤维面积百分比。 结果 模型一组（单侧肾切除+腺嘌呤灌胃+高磷饲料喂养法）造模结束时，与正常对照组比，红细胞、血红蛋白明显下降，血清尿素、肌酐明显升高，TBI、肾组织胶原纤维面积百分比轻度升高；HE染色可见肾小管上皮细胞空泡变性、刷状缘消失，肾小管管腔扩张，肾小管呈灶状萎缩，肾间质水肿并伴有少量纤维化、可见炎症细胞浸润；可见骨小梁排列紊乱，破骨细胞数量增加；Masson染色可见肾间质内少量胶原纤维沉积。造模结束后6周，与正常对照组比，模型一组红细胞、血红蛋白轻度下降，血清尿素、肌酐、TBI、肾组织胶原纤维面积百分比轻度升高；HE染色可见部分肾小管管腔轻度扩张，肾间质轻度水肿并伴有少量纤维化；股骨组织未见明显病变 ；Masson染色可见肾间质内有少量胶原纤维沉积。模型二组（高磷+腺嘌呤饲料喂养法）造模结束时，与正常对照组比，红细胞、血红蛋白明显下降，血清尿素、肌酐、TBI、肾组织胶原纤维面积百分比明显升高；HE染色可见肾小管上皮细胞空泡变性、刷状缘消失，肾小管管腔扩张，肾小管呈灶状萎缩，肾间质轻度水肿并伴有明显纤维化，可见大量炎症细胞浸润；可见骨小梁排列紊乱、局部破骨细胞数目增多、溶骨改变明显；Masson染色可见肾间质内有大量胶原纤维沉积。造模结束后6周，与正常对照组比，模型二组红细胞、血红蛋白较正常对照组明显下降，血清尿素、肌酐、TBI、肾组织胶原纤维面积百分比较正常对照组明显升高；HE染色可见部分肾小管上皮细胞脱落、刷状缘消失，肾间质轻度水肿并伴有明显纤维化，可见大量炎症细胞浸润；可见破骨细胞数目增多、有溶骨改变；Masson染色可见肾间质内有大量胶原纤维沉积。造模结束后6周，与模型一组比，模型二组大鼠红细胞、血红蛋白明显降低，血清尿素、肌酐、TBI、肾组织胶原纤维面积百分比明显升高；HE染色可见肾间质大量炎症细胞浸润；可见骨小梁排列紊乱、局部破骨细胞数目增多、溶骨改变明显；Masson染色可见肾间质内明显胶原纤维沉积。 结论 高磷+腺嘌呤饲料喂养法可建立稳定性较高的CRF大鼠模型。 Abstract:Objective To investigate the stability of the two kinds of rat models of chronic renal failure, aiming to provide a basis for establishing a feasible and stable animal model of chronic renal failure. Methods The rat model of chronic renal failure was established by unilateral nephrectomy + adenine gavage + high-phosphorus diet and high-phosphorus + adenine diet, respectively. Erythrocyte and hemoglobin levels, serum urea and creatinine levels were detected at the end of modeling and 6 weeks after modeling, and histopathological examinations of kidney and bone were performed. The tubulointerstitial fibrosis index （TBI） and the percentage of collagen fiber area in renal tissue were calculated. Results At the end of modeling, compared with the normal group, the erythrocyte and hemoglobin in the model group 1（unilateral nephrectomy + adenine gavage + high-phosphorus） were significantly decreased, the serum urea and creatinine were significantly increased, TBI and the percentage of collagen fiber area in kidney tissue were slightly increased. HE staining revealed vacuolar degeneration of renal tubular epithelial cells, disappearance of the brush border, dilation of the renal tubular lumen, focal atrophy of the renal tubules, interstitial edema accompanied by a small amount of fibrosis, and infiltration of inflammatory cells. It was also observed that the arrangement of trabeculae was disordered and the number of osteoclasts increased. Masson staining showed a small amount of collagen fiber deposition in the renal interstitium. 6 weeks after the end of modeling, compared with the normal group, the erythrocyte and hemoglobin in the model group 1 were slightly decreased, serum urea, creatinine, TBI and the percentage of collagen fiber area in renal tissue were slightly increased. HE staining revealed that the lumens of some renal tubules were slightly dilated, the renal interstitium was mildly edematous and accompanied by a small amount of fibrosis. No obvious lesions were observed in the femoral tissue. Masson staining indicated that there was a small amount of collagen fiber deposition in the renal interstitium. At the end of the modeling, the erythrocyte and hemoglobin in the model group 2 （high-phosphorus + adenine diet） were significantly decreased compared with the normal group, serum urea, creatinine, TBI and the percentage of collagen fiber area in kidney tissue were significantly increased. HE staining revealed vacuolar degeneration of renal tubular epithelial cells, disappearance of the brush border, dilation of the renal tubular lumen, focal atrophy of the renal tubules, mild edema of the renal interstitium accompanied by obvious fibrosis, and a large number of inflammatory cell infiltrations. It can be observed that the arrangement of bone trabeculae was disordered, the number of local osteoclasts increased, and osteolytic changes were obvious. Masson staining indicated a large amount of collagen fiber deposition in the renal interstitium. 6 weeks after the end of modeling, compared with the normal group, the erythrocyte and hemoglobin in the model group 2 were significantly decreased, serum urea, creatinine, TBI and the percentage of collagen fiber area in kidney tissue were significantly increased. HE staining revealed the shedding of some renal tubular epithelial cells and the disappearance of the brush border, mild interstitial edema accompanied by obvious fibrosis, and a large number of inflammatory cell infiltrations. An increase in the number of osteoclasts and osteolytic changes were observed. Masson staining indicated a considerable amount of collagen fiber deposition in the renal interstitium. 6 weeks after the end of modeling, compared with the model group 1, the erythrocyte and hemoglobin in the model group 2 were significantly decreased, while the serum urea, creatinine, TBI, the collagen fiber area percentage of kidney tissue and femur lesions were significantly increased. HE staining revealed a large number of inflammatory cell infiltrations in the renal interstitium. It was observable that the arrangement of bone trabeculae was disordered, the number of local osteoclasts was increased, and the osteolytic changes were significant. Masson staining indicated obvious collagen fiber deposition in the renal interstitium. Conclusions A stable rat model of chronic renal failure can be established by high phosphorus + adenine diet. -
Key words:
- Rat /
- Chronic renal failure /
- Adenine /
- Stability
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图 1 肾脏大体形态
A：正常对照组；B：模型一组造模结束时；C：模型一组造模结束后6周；D：模型二组造模结束时；E：模型二组造模结束后6周。
Figure 1. General morphology of kidneys
图 2 肾脏组织HE染色（100×）
A：正常对照组造模结束时；B：正常对照组造模结束后6周；C：模型一组造模结束时；D：模型一组造模结束后6周；E：模型二组造模结束时；F：模型二组造模结束后6周（实心箭头所指为肾间质炎症细胞，空心箭头所指为扩张肾小管）。
Figure 2. HE staining of renal tissue （100×）
图 3 肾脏组织Masson染色（100×）
A：正常对照组造模结束时；B：正常对照组造模结束后6周；C：模型一组造模结束时；D：模型一组造模结束后6周；E：模型二组造模结束时；F：模型二组造模结束后6周（箭头所指为肾间质纤维化）。
Figure 3. Masson staining of renal tissue （100×）
图 4 股骨HE染色（200×）
A：正常对照组造模结束时；B：正常对照组造模结束后6周；C：模型一组造模结束时；D：模型一组造模结束后6周；E：模型二组造模结束时；F：模型二组造模结束后6周（空心箭头所指为破骨细胞，空心箭头所指为排列紊乱的骨小梁）。
Figure 4. HE staining of femur （200×）
表 1 肾组织TBI评分和胶原纤维百分比（$ \bar x \pm s $）
Table 1. TBI scores and percentage of collagen fibers of renal tissue （$ \bar x \pm s $）
时间 正常对照组
（n = 10）模型一组
（n = 10）模型二组
（n = 13）F P TBI评分 造模结束时 0.00 ± 0.00 1.69 ± 0.12# 2.58 ± 0.06#* 2876.793 < 0.001 造模结束后6周 0.00 ± 0.00 0.39 ± 0.19#￥ 2.04 ± 0.14#*￥ 743.100 < 0.001 胶原纤维百分比（%） 造模结束时 0.53 ± 0.13 5.77 ± 1.03# 35.20 ± 4.49#* 492.517 < 0.001 造模结束后6周 0.85 ± 0.09 7.33 ± 0.94#￥ 37.21 ± 4.98#* 433.534 < 0.001 注：与正常对照组比较，#P < 0.05；与模型一组比较，*P< 0.05；与造模结束时比较，￥P < 0.05。 
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表 2 2种实验方法大鼠红细胞和血红蛋白水平比较（$ \bar x \pm s $）
Table 2. Comparison of erythrocyte and hemoglobin levels between the two methods
时间 正常对照组
（n = 10）模型一组
（n = 10）模型二组
（n = 13）F P 红细胞 （×1012/L） 造模结束时 7.55 ± 0.45 5.21 ± 1.27# 4.57 ± 0.71# 32.128 < 0.001 造模结束后6周 7.67 ± 0.29 6.34 ± 0.65#￥ 4.88 ± 1.11*# 34.416 < 0.001 血红蛋白 （g/L） 造模结束时 171.40 ± 9.51 109.90 ± 27.20# 83.40 ± 10.34*# 65.217 < 0.001 造模结束后6周 168.60 ± 9.25 144.60 ± 11.29#￥ 98.85 ± 16.50*#￥ 84.366 < 0.001 注：与正常对照组比较，#P < 0.05；与模型一组比较，*P < 0.05；与造模结束时比较，￥P < 0.05。
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表 3 2种实验方法大鼠血清肌酐和尿素值（$ \bar x \pm s $）
Table 3. Comparison of serum creatinine and urea values between the two methods （$ \bar x \pm s $）
时间 正常对照组
（n = 10）模型一组
（n = 10）模型二组
（n = 13）F P 肌酐（μmol/L） 造模结束时 53.95 ± 7.82 381.33 ± 112.45# 390.50 ± 40.30# 76.945 < 0.001 造模结束后6周 58.73 ± 4.05 71.55 ± 13.86￥ 213.53 ± 51.14*#￥ 78.604 < 0.001 尿素（mmol/L） 造模结束时 5.85 ± 1.52 58.33 ± 5.65# 45.23 ± 9.35*# 184.134 < 0.001 造模结束后6周 5.83 ± 1.34 8.35 ± 1.78￥ 35.25 ± 15.65*# 31.580 < 0.001 注：与正常对照组比较，#P < 0.05；与模型一组比较，*P < 0.05；与造模结束时比较，￥P < 0.05。
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