多肽化合物urantide对动脉粥样硬化大鼠胸主动脉和VSMC中Ⅳ型胶原表达的影响
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摘要
目的:探讨多肽化合物urantide对动脉粥样硬化(AS)大鼠胸主动脉和血管平滑肌细胞(VSMC中Ⅳ型胶原(ColⅣ)表达的影响,阐明其防治AS的作用机制。方法:180只Wistar大鼠随机分为正常对照组(n=30)和AS模型组(n=150),采用高脂饲料喂养和腹腔注射维生素D3(VD3)的方法建立大鼠AS模型。150只AS模型鼠随机分为AS组、氟伐他汀(Flu)组和urantide组(3、7和14d组)(n=30)。免疫组织化学法检测大鼠胸主动脉壁内ColⅣ的表达水平,酶联免疫吸附试验(ELISA)法检测大鼠血清和尿液中羟脯胺酸(HYP)水平。体外培养的VSMC随机分为正常对照组、尾加压素(UⅡ)(10-8 mol·L-1)组、Flu组和urantide (10-10～10-6 mol·L-1)组,ELISA法检测各组大鼠VSMC培养上清中ColⅣ水平。结果:各组大鼠胸主动脉内膜下不规则斑块内ColⅣ表达水平比较差异有统计学意义(F=35.09,P<0.01)。与正常对照组比较,AS组大鼠主动脉中ColⅣ表达水平明显升高(P<0.01);urantide组ColⅣ阳性染色强度和范围较AS组均减少(P<0.01)。各组大鼠血清和尿液中HYP水平比较差异有统计学意义(F=24.38,P<0.01;F=26.72,P<0.01)。与正常对照组比较,AS组大鼠血清中HYP水平明显升高(P<0.01),尿液中HYP水平明显降低(P<0.01);与AS组比较,urantide组大鼠血清中HYP水平均明显降低(P<0.01),尿液中HYP水平明显升高(P<0.01),接近或优于Flu组水平。各组大鼠VSMC培养上清中ColⅣ水平比较差异有统计学意义(F=31.04,P<0.01)。与正常对照组比较,UⅡ组大鼠VSMC培养上清中ColⅣ水平明显升高(P<0.01);与UⅡ组比较,urantide组大鼠VSMC培养上清中ColⅣ水平明显降低(P<0.05或P<0.01)。结论:urantide可抑制AS大鼠胸主动脉和VSMC中ColⅣ的表达,缓解AS病变程度,为临床应用urantide治疗AS提供了实验依据。
Objective:To investigate the effects of urantide on the expressions of typeⅣ collagen(ColⅣ)in thoracic aorta and vascular smooth muscle cells(VSMC)in the rats with atherosclerosis(AS),and to clarify its mechanism of prevention and treatment of AS.Methods:A total of 180 Wistar rats were randomly divided into normal control group(n=30)and AS model group(n=150).The rat models of AS were established by feeding on high-fat diet or intraperitoneally injecting vitamin D3(VD3).The AS model rats were randomly divided into AS group,fluvestation(Flu)group and urantide group(3,7,and 14 dgroups).The expression of ColⅣin thoracic aorta wall of the rats was detected by immunohistochemistry.The levels of hydroxyproline(HYP)in serum and urine of the rats in various groups were measured by ELISA.The VSMC were randomly divided into normal control group,urotensinⅡ(UⅡ 10-8 mol·L-1)group,Flu group and urantide(10-10 to 10-6 mol·L-1)groups.The levels of ColⅣin VSMC of the rats in various groups were determined by ELISA.Results:There was a significant difference in the expression levels of ColⅣin the irregular plaques of the thoracic aorta of the rats between various groups(F=35.09,P<0.01).The expression levels of ColⅣ in thoracic aorta of the rats in AS group were significantly increased compared with normal control group(P<0.01);the intensity and extent of ColⅣ positive staining in urantide group were lower than those in AS group(P<0.01).There were significant differences in the serum and urine HYP levels between various groups(F=24.38,P<0.01;F=26.72,P<0.01).Compared with normal control group,the serum HYP level of the rats in AS group were significantly increased(P<0.01),and the urine HYP level was significantly decreased(P<0.01).Compared with AS group,the serum HYP levels in urantide groups were significantly decreased(P<0.01)and the urine HYP levels were significantly increased(P<0.01),no less than the level in Flu group.The expression levels of ColⅣin the culture supernatant of VSMC in the rats in various groups had significantly difference(F=31.04,P<0.01).The expression level of ColⅣin the culture supernatant of VSMC of the rats in UⅡ group was significantly increased compared with normal control group(P<0.01);the expression levels of ColⅣin the culture supernatant of VSMC of the rats in urantide groups were significantly decreased compared with UⅡ group(P<0.05 or P<0.01).Conclusion:Urantide can inhibit the expressions of ColⅣin the thoracic aorta and VSMC of the AS rats and alleviate the degree of AS lesions,which provides the experimental evidence for the clinical application of urantide in the treatment of AS.
Objective:To investigate the effects of urantide on the expressions of typeⅣ collagen(ColⅣ)in thoracic aorta and vascular smooth muscle cells(VSMC)in the rats with atherosclerosis(AS),and to clarify its mechanism of prevention and treatment of AS.Methods:A total of 180 Wistar rats were randomly divided into normal control group(n=30)and AS model group(n=150).The rat models of AS were established by feeding on high-fat diet or intraperitoneally injecting vitamin D3(VD3).The AS model rats were randomly divided into AS group,fluvestation(Flu)group and urantide group(3,7,and 14 dgroups).The expression of ColⅣin thoracic aorta wall of the rats was detected by immunohistochemistry.The levels of hydroxyproline(HYP)in serum and urine of the rats in various groups were measured by ELISA.The VSMC were randomly divided into normal control group,urotensinⅡ(UⅡ 10-8 mol·L-1)group,Flu group and urantide(10-10 to 10-6 mol·L-1)groups.The levels of ColⅣin VSMC of the rats in various groups were determined by ELISA.Results:There was a significant difference in the expression levels of ColⅣin the irregular plaques of the thoracic aorta of the rats between various groups(F=35.09,P<0.01).The expression levels of ColⅣ in thoracic aorta of the rats in AS group were significantly increased compared with normal control group(P<0.01);the intensity and extent of ColⅣ positive staining in urantide group were lower than those in AS group(P<0.01).There were significant differences in the serum and urine HYP levels between various groups(F=24.38,P<0.01;F=26.72,P<0.01).Compared with normal control group,the serum HYP level of the rats in AS group were significantly increased(P<0.01),and the urine HYP level was significantly decreased(P<0.01).Compared with AS group,the serum HYP levels in urantide groups were significantly decreased(P<0.01)and the urine HYP levels were significantly increased(P<0.01),no less than the level in Flu group.The expression levels of ColⅣin the culture supernatant of VSMC in the rats in various groups had significantly difference(F=31.04,P<0.01).The expression level of ColⅣin the culture supernatant of VSMC of the rats in UⅡ group was significantly increased compared with normal control group(P<0.01);the expression levels of ColⅣin the culture supernatant of VSMC of the rats in urantide groups were significantly decreased compared with UⅡ group(P<0.05 or P<0.01).Conclusion:Urantide can inhibit the expressions of ColⅣin the thoracic aorta and VSMC of the AS rats and alleviate the degree of AS lesions,which provides the experimental evidence for the clinical application of urantide in the treatment of AS.
引文
[1]刘燕,刘晓辉,宋艳玲,等.急性脑梗死患者血清Ⅰ型胶原羧基末端肽改变及其与颈动脉粥样硬化斑块的关系[J].心肺血管病杂志,2018,37(1):39-45.
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[11]CHUNG E J.Targeting and therapeutic peptides in nanomedicine for atherosclerosis[J].Exp Biol Med(Maywood),2016,241(9):891-898.
[12]田祥全,万小松,杨纪才,等.血清Ⅰ型胶原羧基末端肽、基质金属蛋白-2水平与冠状动脉病变的关系[J].中国循证心血管医学杂志,2016,8(5):553-555.
[13]刘娜,资晓宏.胶原在动脉粥样硬化斑块中的作用研究进展[J].浙江临床医学,2017,19(2):377-379.
[14]赵娟.Urantide抑制动脉粥样硬化大鼠单核细胞趋化蛋白-1的表达[J].中国药理学与毒理学杂志,2011,25(5):425-429.
[15]赵娟,谢利德,王红伟,等.Urantide对实验性动脉粥样硬化大鼠白细胞介素-6表达的影响[J].中国老年学杂志,2013,33(8):1823-1824.
[16]ZHAO J,XIE L D,SONG C J,et al.Urantide improves atherosclerosis by controlling C-reactive protein,monocyte chemotactic protein-1and transforming growth factor-Urantide improves atherosclerosis by controlling C reactive protein,monocyte chemotactic protein-1 and transforming growth factor-βexpression in rats[J].Exp Therapeut Med,2014,7(10):1647-1652.
[17]ALBANESE I,DASKALOPOULOU S S,YU B,et al.The urotensinⅡsystem and carotid atherosclerosis:A role in vascular calcification[J].Front Pharmacol,2016,7:149-157.
[18]OH K S,LEE J H,YI K Y,et al.A novel urotensinⅡreceptor antagonist,KR-36996,improved cardiac function and attenuated cardiac hypertrophy in experimental heart failure[J].Eur J Pharmacol,2017,799:94-102.
[19]CHOW W Y,BIHAN D,FORMAN C J,et al.Hydroxyproline ring pucker causes frustration of helix parameters in the collagen triple helix[J].Sci Rep,2015,5:12556.
[20]钱进,邓辰亮,杨松林,等.建立SCID鼠增生性瘢痕模型的实验研究[J].组织工程与重建外科杂志,2018,14(1):31-35.
[2]马玉梅,霍清萍.基质金属蛋白酶9与动脉粥样硬化及中医药防治[J].中西医结合心脑血管病杂志,2014,12(9):1127-1128.
[3]郑辉,刘慧娟,葛焕琦,等.网膜素改善氧化应激对人动脉血管平滑肌细胞Ⅰ、Ⅳ型胶原表达的抑制作用[J].中国动脉硬化杂志,2016,24(2):141-144.
[4]DUNR P,GONALVES I,GRUFMAN H,et al.Increased aldehyde-modification of collagen typeⅣin symptomatic plaques:A possible cause of endothelial dysfunction[J].Atherosclerosis,2015,240(1):26-32.
[5]MCLEOD O,DUNR P,SAMNEGRD A,et al.Autoantibodies against basement membrane collagen typeⅣare associated with myocardial infarction[J].Int J Cardiol Heart Vasc,2015,6(1):42-47.
[6]ZHAO J,ZHANG S F,SHI Y,et al.Effects of urotensinⅡand its specific receptor antagonist urantide on rat vascular smooth muscle cells[J].Bosn J Basic Med Sci,2013,13(2):78-83.
[7]BARKAREV M A,KARPOVA A A,PICHIGIN V I,et al.Involvement of the coronary bed in patients with coronary heart disease against the background of primarily coronary and generalized atherosclerosis[J].Bull Exp Biol Med,2016,162(2):283-287.
[8]赵娟,宋成军,姜菊花,等.urantide对实验性动脉粥样硬化大鼠肝脏的影响[J].中国老年学杂志,2014,34(4):940-942.
[9]ZHAO J,YU QX,KONG W,et al.The urotensinⅡreceptor antagonist,urantide,protects against atherosclerosis in rats[J].Exp Ther Med,2013,5(6):1765-1769.
[10]KONDO T,ENDO I,AIHARA K,et al.Serum carboxyterminal telopeptide of typeⅠcollagen levels are associated with carotid atherosclerosis in patients with cardiovascular risk factors[J].Endocr J,2016,63(4):397-404.
[11]CHUNG E J.Targeting and therapeutic peptides in nanomedicine for atherosclerosis[J].Exp Biol Med(Maywood),2016,241(9):891-898.
[12]田祥全,万小松,杨纪才,等.血清Ⅰ型胶原羧基末端肽、基质金属蛋白-2水平与冠状动脉病变的关系[J].中国循证心血管医学杂志,2016,8(5):553-555.
[13]刘娜,资晓宏.胶原在动脉粥样硬化斑块中的作用研究进展[J].浙江临床医学,2017,19(2):377-379.
[14]赵娟.Urantide抑制动脉粥样硬化大鼠单核细胞趋化蛋白-1的表达[J].中国药理学与毒理学杂志,2011,25(5):425-429.
[15]赵娟,谢利德,王红伟,等.Urantide对实验性动脉粥样硬化大鼠白细胞介素-6表达的影响[J].中国老年学杂志,2013,33(8):1823-1824.
[16]ZHAO J,XIE L D,SONG C J,et al.Urantide improves atherosclerosis by controlling C-reactive protein,monocyte chemotactic protein-1and transforming growth factor-Urantide improves atherosclerosis by controlling C reactive protein,monocyte chemotactic protein-1 and transforming growth factor-βexpression in rats[J].Exp Therapeut Med,2014,7(10):1647-1652.
[17]ALBANESE I,DASKALOPOULOU S S,YU B,et al.The urotensinⅡsystem and carotid atherosclerosis:A role in vascular calcification[J].Front Pharmacol,2016,7:149-157.
[18]OH K S,LEE J H,YI K Y,et al.A novel urotensinⅡreceptor antagonist,KR-36996,improved cardiac function and attenuated cardiac hypertrophy in experimental heart failure[J].Eur J Pharmacol,2017,799:94-102.
[19]CHOW W Y,BIHAN D,FORMAN C J,et al.Hydroxyproline ring pucker causes frustration of helix parameters in the collagen triple helix[J].Sci Rep,2015,5:12556.
[20]钱进,邓辰亮,杨松林,等.建立SCID鼠增生性瘢痕模型的实验研究[J].组织工程与重建外科杂志,2018,14(1):31-35.