皮质抑素调控钙化相关基因减轻大鼠动脉钙化

目的 探讨皮质抑素（cortistatin，CST）对大鼠主动脉钙化的影响及其可能的分子机制。方法 利用维生素D3联合尼古丁（VDN）所致的大鼠动脉钙化模型，分别采用孔雀绿直接显色法、邻甲酚酞络合酮比色法和von Kossa染色法测定大鼠血浆磷、钙水平和主动脉组织的钙含量和钙沉积，应用RT-PCR方法检测主动脉组织钙化相关基因mRNA表达。结果 与对照组相比较，VDN使大鼠主动脉的钙含量增加70.2%（P<0.05），引起弹力纤维紊乱、中断，von Kossa染色阳性的棕黑色颗粒明显增多。VDN+CST组与单独VDN组相比较，持续皮下泵入CST使主动脉的钙含量减少45.6%（P<0.05），弹力纤维紊乱中断减轻和棕黑色颗粒明显减少。而血钙、磷及钙磷乘积在各组间无显著性差异。RT-PCR结果证实VDN组主动脉组织的BMP-2 mRNA和Pit-1 mRNA表达分别增加53.2%（P<0.05）和34.0%（P<0.01），而MGP mRNA表达减少27.0%（P<0.05）。持续皮下泵入CST使主动脉组织BMP-2 mRNA和Pit-1 mRNA表达较单独VDN组分别下降38.3%（P<0.01）和17.4%（P<0.05），而MGP mRNA表达增加34.9%（P<0.01）。主动脉组织OPG mRNA表达在各组间均无显著性差异。结论 CST能够减轻VDN所致的大鼠动脉钙化，可能与其纠正促\抑钙化相关基因表达失衡有关，从而为CST防治动脉钙化提供实验依据。     

大鼠钙化血管尾加压素Ⅱ及其受体上调

目的 观察血管钙化大鼠主动脉和心肌尾加压素Ⅱ及其受体表达的变化.方法 采用维生素D3 和尼古丁诱导大鼠血管钙化模型,以Von Konsa染色检测血管钙化,以原子吸收法和磷酸苯二钠法测定血管钙含量和碱性磷酸酶活性,放射免疫法检测血浆、主动脉和心肌尾加压素Ⅱ含量,免疫组织化学法检测血管尾加压素Ⅱ的表达,RT-PCR法检测主动脉和心肌尾加压素受体mRNA水平.结果 维生素D3 和尼古丁能够诱导大鼠典型血管钙化形成.Von Kossa染色可见血管钙化大鼠主动脉有大量黑色颗粒沉淀,血管钙含量、碱性磷酸酶活性明显升高,主动脉尾加压素Ⅱ含量、主动脉和心肌尾加压素受体基因表达明显上调.精氨酸饮食能减轻血管钙化,血管钙含量、尾加压素Ⅱ水平及尾加压素受体mRNA表达与单纯钙化组相比轻度下降,但无统计学意义.蛋氨酸饮食能加重血管钙化,增加钙含量,上调尾加压素Ⅱ表达,降低碱性磷酸酶活性.各组之间血浆尾加压素Ⅱ水平差异无统计学意义.结论 大鼠钙化血管尾加压素Ⅱ表达上调,提示尾加压素Ⅱ可能参与了血管钙化的发展过程.     

罗格列酮对糖尿病大鼠胸主动脉钙化的影响

目的探讨罗格列酮对糖尿病大鼠胸主动脉钙化的影响。方法利用链脲佐菌素联合高能量饲料和维生素D3并尼古丁制备糖尿病血管钙化大鼠模型(DM+VDN组),并随机分为模型组、罗格列酮(RSG)治疗组和格列本脲(GLB)治疗组。检测大鼠一般代谢指标、血清糖基化终末产物(AGE)、丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性,胸主动脉钙含量及糖基化终末产物受体(RAGE)蛋白表达。结果 RSG治疗组和GLB治疗组血清AGE、MDA及SOD水平低于模型组,且RSG效果优于GLB(P0.05); RSG治疗组胸主动脉RAGE表达水平以及钙含量均低于模型组和GLB治疗组(P0.05)。结论罗格列酮可能通过降低AGE水平,下调RAGE表达,抑制氧化应激反应,延缓血管钙化进展。     

血管钙化模型大鼠salusin-α表达的实验研究

目的通过制作血管钙化模型大鼠,观察血浆和主动脉组织salusin-α的表达情况。方法将16只雄性SD大鼠随机分为正常组和钙化组,每组8只,4周后取材检测血管钙化程度,用钙离子测试盒、碱性磷酸酶(ALP)试剂盒测定大鼠主动脉钙含量和ALP活性,用放射免疫法检测大鼠血浆和主动脉组织salusin-α含量。结果与对照组比较,钙化组大鼠主动脉中膜弹性纤维间可见钙盐沉积,血浆和主动脉组织中ALP、钙含量明显升高(P0.01),salusin-α的表达明显降低(P0.01)。结论血管钙化大鼠模型salusin-α的表达下调。     

慢性间歇性低压低氧抑制内质网应激改善大鼠血管钙化

目的证实慢性间歇性低压低氧(CIHH)通过抑制内质网应激(ERS)改善大鼠血管钙化。方法使用维生素D3肌注和尼古丁灌胃(VDN)制备的大鼠在体血管钙化模型,检测主动脉组织Ca2+含量和碱性磷酸酶(ALP)活性以及血浆ALP活性,Western blot检测相关蛋白的表达水平。结果与对照组大鼠相比,VDN大鼠主动脉组织Ca2+含量和ALP活性以及血浆ALP活性均显著升高(P0.05),同时主动脉组织平滑肌细胞收缩表型标志蛋白calponin和SM22α的表达水平显著下调(P0.05),成骨样细胞表型标志蛋白骨形态发生蛋白2(BMP2)和RUNX2的表达水平显著上调(P0.05)。而CIHH能够显著改善VDN大鼠的上述改变。此外,VDN大鼠主动脉组织ERS标志蛋白葡萄糖调节蛋白78(GRP78)、CHOP(C/EBP homologous protein)和active-caspase12的表达水平较对照组大鼠显著升高(P0.05),而CIHH能够显著抑制上述蛋白表达水平的升高(P0.05)。结论 CIHH可能通过抑制ERS,发挥改善血管钙化和平滑肌细胞表型转化的作用。     

高同型半胱氨酸对高血压大鼠心肌细胞GRP78和CHOP表达及左室肥厚的影响

目的观察高同型半胱氨酸(hyperhomocysteine,Hcy)对高血压大鼠心肌内质网应激(endoplasmic reticulum stress,ERS)相关因子GRP78和CHOP表达的影响,并探讨HHcy与左室肥厚的关系。方法采用腹主动脉缩窄术制备高血压大鼠模型,术后2周用无创大鼠尾动脉测压仪测大鼠尾动脉压,择取40只高血压大鼠,随机分为对照组和高蛋氨酸组,每组20只。对照组给予普通饮食,高蛋氨酸组给于含2%蛋氨酸饮食。按分组后饲养时间不同,分别又将各组大鼠分为4W亚组及8W亚组,每组10只。无创大鼠尾动脉测压仪检测动脉收缩压(SBP),同型半胱氨酸Hcy检测仪测血清Hcy浓度、称取体重、全心质量及左心室质量计算大鼠心肌肥厚指数HWI及LVWI,通过HE染色观察心肌细胞形态学改变,免疫组化和western检测大鼠心肌组织中GRP78及CHOP表达水平。结果①对照组大鼠血清Hcy水平无明显增高,随时间延长,高蛋氨酸组大鼠血清Hcy水平增高且显著高于对照组(P0.01)。②对照组与高蛋氨酸组大鼠SBP值随时间的增长而升高。两组4W亚组间相比,高蛋氨酸组大鼠SBP略高于对照组,但两组间SBP值无统计学差异(P0.05);两组8W亚组间相比,高蛋氨酸组大鼠SBP值明显高于对照组(P0.05)。③对照组与高蛋氨酸组4W、8W亚组大鼠HWI及LVWI均有明显增高,且高蛋氨酸组相应亚组大鼠HWI及LVWI增高更为明显(P0.05)。④4W时,高蛋氨酸组大鼠心肌细胞GRP78表达明显高于对照组(P0.05);8W时,高蛋氨酸组大鼠心肌细胞CHOP表达较对照组增高更为显著(P0.05)。结论血清高Hcy浓度可促使心肌细胞ERS加重,导致ERS相关因子GRP78及CHOP表达失衡,促凋亡因子CHOP表达占优势,导致左室肥厚程度加重。ERS可能是高Hcy加重高血压左室肥厚的分子生物学机制之一。     

血管钙化对血管组织内皮素表达的影响

通过观察血管钙化大鼠血管组织和钙化血管平滑肌细胞内皮素含量的变化 ,探讨血管钙化对血管组织内皮素表达的影响。用维生素D3加尼古丁诱导大鼠血管钙化模型 ,β 磷酸甘油制备钙化血管平滑肌细胞 ,采用VonKossa染色、钙含量测定、4 5Ca聚集及碱性磷酸酶活性测定判断钙化程度 ,放射免疫分析法测定血浆、血管和血管平滑肌细胞培养基中内皮素含量 ,用竞争性定量逆转录聚合酶链反应方法测定血管和血管平滑肌细胞中内皮素mRNA水平。结果发现 :钙化血管平滑肌细胞VonKossa染色见有大量黑色颗粒沉积 ,钙化细胞的钙含量、4 5Ca2 + 摄入及碱性磷酸酶活性分别较正常平滑肌细胞增加 1 1 8%、1 74 %和 7倍 (P均 <0 .0 1 ) ;钙化细胞培养基中内皮素含量较对照组增加 35 % (P <0 .0 5 ) ,内皮素mRNA水平较对照组高 1 2 0 % (P <0 .0 5 ) ;钙化组大鼠血管组织VonKossa染色见血管中层有大量黑色颗粒沉积 ,主动脉钙含量、4 5Ca2 + 沉积及碱性磷酸酶活性分别较对照组高 5 .0倍、1 .4倍和1 .4倍 (P均 <0 .0 1 ) ;钙化大鼠血浆和血管内皮素含量分别较对照组增加 1 0 2 %和 1 0 3% (P均 <0 .0 1 ) ;钙化血管内皮素mRNA水平较正常对照组高 2 2 % (P <0 .0 1 ) ;波生坦处理的大鼠血管钙含量、4 5Ca2 + 聚集及碱性磷酸酶活性较单     

Salusin-β在血管钙化模型的表达变化及意义

目的在大鼠血管钙化模型上观察血浆和主动脉组织salusin-β表达。方法实验动物按电脑随机数字表法随机分为正常组和钙化组,每组8只。钙化组采用维生素D3(300 000 U/kg 1次,肌肉注射)和尼古丁(25 mg/kg溶于花生油中早、晚各灌胃1次)诱导大鼠血管钙化模型。常规饲养4周后取材,用Von Kossa染色检测血管钙化程度;用钙离子测试盒、碱性磷酸酶(ALP)试剂盒测定大鼠主动脉钙含量和碱性磷酸酶活性;用放射免疫法检测大鼠血浆和主动脉组织salusin-β含量。结果维生素D3和尼古丁能够诱导典型大鼠血管钙化模型,Von Kossa染色可见血管钙化模型大鼠主动脉有大量黑色颗粒沉淀。钙化组血管钙含量、碱性磷酸酶活性明显高于正常组,差异有统计学意义(P<0.05);同时,钙化组血浆和主动脉组织中salusin-β的表达明显上调。结论大鼠血管钙化模型中salusin-β的表达上调。     

姜黄素对维生素D3和尼古丁诱导血管钙化大鼠血管中组织蛋白酶D表达的影响

目的研究组织蛋白酶D(CTSD)在大鼠动脉钙化模型中的表达以及姜黄素对其表达的影响。方法将30只雄性SD大鼠随机分成3组,每组10只,即对照组、钙化组和干预组。对照组大鼠给予等量生理盐水处理,钙化组大鼠用维生素D3和尼古丁处理,干预组大鼠在钙化组处理的基础上加用姜黄素干预。用von Kossa染色检测血管钙化,试剂盒检测血管钙含量和碱性磷酸酶活性,Western blot检测骨形态发生蛋白2(BMP-2)、Runx2和CTSD的蛋白表达,用免疫组织化学染色观察CTSD在血管中的表达及分布。结果维生素D3和尼古丁能够诱导经典大鼠钙化模型,von Kossa染色结果显示钙化组大鼠胸主动脉有大量黑色颗粒沉积,血管钙含量和碱性磷酸酶活性升高,Western blot检测结果显示BMP-2和Runx2表达显著上调;使用姜黄素干预后,与钙化组相比,大鼠胸主动脉钙化程度明显减轻,血管钙含量和碱性磷酸酶活性下降,BMP-2和Runx2表达下调。免疫组织化学染色结果显示钙化组CTSD表达上调且大量分布在细胞外基质中,姜黄素能够显著下调CTSD的表达并减少其在细胞外基质中的分布。结论姜黄素具有抑制血管钙化的作用,可能与其能够减少CTSD在细胞外基质中的表达有关。     

血管钙化大鼠模型Apelin及其受体表达的实验研究

目的观察血管钙化大鼠模型血浆和主动脉组织Apelin及其受体(APJ)的表达。方法选取16只雄性SD大鼠,随机分正常组和钙化组,每组8只。钙化组:08:00肌肉注射维生素D3 3×10^5 U/kg,同时尼古丁25 mg/kg溶于1 mL花生油灌胃,18:00时重复灌胃1次。正常组:类似钙化组操作,但肌肉注射生理盐水和灌胃单纯花生油。4周后取材检测血管钙化程度,用钙离子测试盒、碱性磷酸酶(ALP)试剂盒测定大鼠主动脉钙含量和ALP活性,用酶联免疫吸附法检测大鼠血浆和主动脉组织Apelin含量,用免疫组化法检测大鼠主动脉组织APJ的表达。结果与正常组相比,钙化组中主动脉中膜弹性纤维间可见钙盐沉积,血清和主动脉组织中ALP、钙含量明显升高(P<0.01),而Apelin的表达明显降低(P<0.01),同时,APJ表达亦下调。结论血管钙化大鼠模型Apelin及其受体的表达下调,Apelin/APJ系统可能是新的抑制血管钙化因子。     

糖尿病大鼠钙化血管中Msx2和Wnt3a的表达

目的 研究糖尿病对血管钙化的影响及Msx2和wnt3a基因在钙化血管中的表达变化.方法 将48只雄性wistar大鼠随机分为四组:维生素D3和尼古丁诱导的单纯血管钙化组(n=12)、链脲佐菌素诱导的单纯糖尿病组(n=12)、链脲佐菌素联合维生素D3和尼古丁诱导的糖尿病合并血管钙化组(n=12)和正常雄性Wistar大鼠为正常对照组(n=12).测定大鼠血糖、血清胰岛素、总胆固醇和甘油三酯水平,以血管von Kossa染色、血管钙含量和碱性磷酸酶活性作为判断血管钙化程度的指标,测定大鼠血管中Msx2和wnt3a mRNA 的表达.结果 与正常对照组相比,单纯血管钙化组大鼠血管中Msx2和wnt3a mRNA 相对表达量有所升高(P<0.05),但血管钙含量和碱性磷酸酶活性无明显变化.与正常对照组及单纯血管钙化组相比,糖尿病合并血管钙化组大鼠的血管中,可见沿中膜弹力层内广泛分布的钙盐沉积,大鼠血管中钙含量和碱性磷酸酶活性以及血管内Msx2和wnt3amRNA 相对表达量明显增高(P<0.05).结论 糖尿病可以明显加速血管钙化的发生和发展.在糖尿病大鼠钙化血管中,骨形成过程中的转录因子Msx2和Wnt3a表达增高,提示血管钙化是一个类似于骨形成的过程,Msx2和Wnt3a 参与血管钙化病变的发生.     

肾上腺髓质素对大鼠血管钙化的影响

目的　观察肾上腺髓质素 (ADM)对血管钙化的影响。方法　维生素D3 (VitD3 )和尼古丁制备大鼠血管钙化模型 ,分别测定血管、心肌钙含量和血管碱性磷酸酶 (ALP)活性 ,血浆和血管ADM含量 ,12 5I ADM与ADM受体的结合力及血管环磷酸腺苷 (cAMP)含量。结果　与对照组相比 ,钙化组 (VDN组 )的血管钙含量和ALP活性明显升高 ;血管及血浆ADM的含量亦升高 ,但12 5I ADM与血管质膜ADM受体结合的位点减少 ,Kd值升高 ,提示亲合力降低 ,同时伴钙化血管的cAMP合成减少。提示钙化血管对ADM的反应减弱。空载脂质体对血管钙化无影响。用脂质体包裹ADM组 (VDN ADM组 )与钙化组相比 ,其血管的钙含量、ALP的活性均明显降低 ;12 5　I ADM与血管质膜ADM受体结合的位点增加 ,Kd值减少 ,cAMP的合成也增加 ,提示该组血管对ADM的反应增强。结论　血管钙化时ADM ADM受体 cAMP途径发生变化 ,外源性ADM通过改善ADM ADM受体 cAMP途径 ,发挥抑制血管钙化的作用     

Effects of adrenomedullin on vascular calcification in rats

OBJECTIVE: The aim of the present study was to investigate the effect of adrenomedullin (ADM) on vascular calcification. METHODS: The vascular calcification model was established in rats (VND group) by using vitamin D3 (300,000 IU/kg) and nicotine (25 mg/kg, two doses). The effect of liposome-encapsulated ADM was observed. Vascular calcium content, alkaline phosphatase (ALP) activity, ADM in aortic tissue and plasma, binding ability of 125I-ADM for ADM receptor on vascular plasma membrane and content of cAMP in vessels were measured. RESULTS: Compared with control rats, the aortic calcium content and vascular ALP activity in rats of the VDN group was obviously increased; in addition ADM concentrations in plasma and vessels of rats in VDN group were increased. But the maximum binding sites of 125I-ADM for ADM receptor (Bmax) on vascular plasma membrane in rats of VDN group were significantly decreased compared with control rats. The affinity of 125I-ADM for the ADM receptor was reduced, as shown by the Kd value and vascular cAMP content being reduced in rats of the VDN group compared to the control group. The in vitro response of isolated vessels to ADM incubation was weakened. Administration of empty liposome had no effect on vascular calcification. But administration of ADM significantly decreased vascular calcium content and ALP activity. The Bmax of 125I-ADM for ADM receptors on vascular plasma membrane increased by 17.7% (p < 0.01), and the value of Kd decreased by 36.2% (P < 0.01) in rats treated with ADM as compared with rats of the VDN group. In addition, the vascular cAMP content and the response to ADM in isolated aorta were markedly increased. CONCLUSION: Vascular calcification induced an alteration of the vascular ADM-ADM receptor-cAMP pathway. Treatment with exogenous ADM inhibited vascular calcification by improving the vascular ADM-ADM receptor-cAMP pathway.     

Effects of adrenomedullin on vascular calcification in rats

Objective The aim of the present study was to investigate the effect of adrenomedullin (ADM) on vascular calcification. Methods The vascular calcification model was established in rats (VND group) by using vitamin D₃ (300000IU/kg) and nicotine (25mg/kg, two doses). The effect of liposome-encapsulated ADM was observed. Vascular calcium content, alkaline phosphatase (ALP) activity, ADM in aortic tissue and plasma, binding ability of ¹²⁵I-ADM for ADM receptor on vascular plasma membrane and content of cAMP in vessels were measured. Results Compared with control rats, the aortic calcium content and vascular ALP activity in rats of the VDN group was obviously increased; in addition ADM concentrations in plasma and vessels of rats in VDN group were increased. But the maximum binding sites of ¹²⁵I-ADM for ADM receptor (Bmax) on vascular plasma membrane in rats of VDN group were significantly decreased compared with control rats. The affinity of ¹²⁵I-ADM for the ADM receptor was reduced, as shown by the Kd value and vascular cAMP content being reduced in rats of the VDN group compared to the control group. The in vitro response of isolated vessels to ADM incubation was weakened. Administration of empty liposome had no effect on vascular calcification. But administration of ADM significantly decreased vascular calcium content and ALP activity. The Bmax of ¹²⁵I-ADM for ADM receptors on vascular plasma membrane increased by 17.7% (p<0.01), and the value of Kd decreased by 36.2% (P<0.01) in rats treated with ADM as compared with rats of the VDN group. In addition, the vascular cAMP content and the response to ADM in isolated aorta were markedly increased. Conclusion Vascular calcification induced an alteration of the vascular ADM-ADM receptor-cAMP pathway. Treatment with exogenous ADM inhibited vascular calcification by improving the vascular ADM-ADM receptor-cAMP pathway.     

The consequences of aortic calcium overload following vitamin D3 plus nicotine treatment in young rats.

Treatment of young rats with vitamin D3 plus nicotine has been proposed as a model of cardiovascular calcium overload. This treatment produced a 20-35-fold increase in the calcium content of the aorta, a compliance vessel, and this increase was accompanied by a 1.6-fold elevation of pulse pressure. In aortic rings, the maximal inhibition by the endothelium-dependent vasodilator, carbachol, of vasoconstriction induced by noradrenaline decreased from 90% in controls to 61% in treated animals. There were significant correlations between aortic calcium content and pulse pressure and aortic calcium content and carbachol-induced relaxation. In conclusion, the vitamin D3 plus nicotine model may be useful for the study of the role of calcium overload in decreased arterial compliance coupled with endothelial injury.     

Osteoprotegerin inhibits artery calcification induced by warfarin and by vitamin D

The present experiments were carried out to test the hypothesis that arterial calcification is linked to bone resorption by determining whether the selective inhibition of bone resorption with osteoprotegerin will inhibit arterial calcification. In the first test, arterial calcification was induced by treating 22-day-old male rats with warfarin, a procedure that inhibits the gamma-carboxylation of matrix Gla protein and causes extensive calcification of the arterial media. Compared with rats treated for 1 week with warfarin alone, rats treated with warfarin plus osteoprotegerin at a dose of 1 mg/kg per day had dramatically reduced alizarin red staining for calcification in the aorta and in the carotid, hepatic, mesenteric, renal, and femoral arteries, and they had 90% lower levels of calcium and phosphate in the abdominal aorta (P<0.001) and in tracheal ring cartilage (P<0.01). More rapid arterial calcification was induced by treating 49-day-old male rats with toxic doses of vitamin D. Treatment for 96 hours with vitamin D caused widespread alizarin red staining for calcification in the aorta and the femoral, mesenteric, hepatic, renal, and carotid arteries, and osteoprotegerin completely prevented calcification in each of these arteries and reduced the levels of calcium and phosphate in the abdominal aorta to control levels (P<0.001). Treatment with vitamin D also caused extensive calcification in the lungs, trachea, kidneys, stomach, and small intestine, and treatment with osteoprotegerin reduced or prevented calcification in each of these sites. Measurement of serum levels of cross-linked N-teleopeptides showed that osteoprotegerin dramatically reduced bone resorption activity in each of these experiments (P<0.001). Therefore, we conclude that doses of osteoprotegerin that inhibit bone resorption are able to potently inhibit the calcification of arteries that is induced by warfarin treatment and by vitamin D treatment. These results support the hypothesis that arterial calcification is linked to bone resorption.     

吡格列酮对糖尿病大鼠血管钙化的影响及机制

目的研究吡格列酮对糖尿病大鼠血管钙化的影响及其可能机制。方法将36只SD雄性大鼠随机平均分为6组:对照组、糖尿病组、钙化组、糖尿病+钙化组、钙化+吡格列酮组、糖尿病+钙化+吡格列酮组;建立大鼠血管钙化模型(维生素D3+华法林)和糖尿病模型(链尿佐菌素);并对血管组织进行Von Kossa染色、钙含量和碱性磷酸酶活性检测,qRT-PCR检测mRNA表达,免疫组织化学法检测骨保护素蛋白表达。结果钙化组血管平滑肌细胞及其间质内有大量黑色颗粒沉积;糖尿病+钙化组较糖尿病组和钙化组血管组织钙含量、碱性磷酸酶活性分别升高3.63倍、1.35倍和3.69倍、1.30倍(P<0.05),骨保护素mRNA含量及其蛋白表达降低(P<0.05);糖尿病+钙化+吡格列酮组较糖尿病+钙化组钙含量、碱性磷酸酶活性分别下调13.70%、18.04%(P<0.05),骨保护素mRNA含量及其蛋白表达升高(P<0.05)。结论吡格列酮可以减轻血管钙化程度并上调骨保护素mRNA含量及蛋白表达,骨保护素可能是抑制血管钙化主要因素之一。     

睾酮对维生素D3和尼古丁诱导的主动脉血管钙化的影响

目的建立SD大鼠血管钙化模型,并观察睾酮在主动脉血管钙化中的作用。方法将10周龄SD雄性大鼠分为:对照组、钙化组、钙化+低剂量睾酮组和钙化+高剂量睾酮组,每组8只。除对照组外,其余三组采用维生素D3(300 k U/kg一次肌肉注射)和尼古丁(25 mg/kg溶于花生油中早、晚各灌胃1次)诱导大鼠血管钙化模型;低剂量睾酮组注射1 mg/kg外源性睾酮(隔日注射1次),高剂量睾酮组注射2 mg/kg睾酮(隔日注射1次),持续8周后处死。采用ELISA法测定大鼠血清睾酮和骨形态发生蛋白4(BMP-4)含量,采用试剂盒检测血管组织钙离子及碱性磷酸酶(ALP)含量,蛋白免疫印迹分析(Western blot)检测主动脉血管组织BMP-4、骨桥蛋白(OPN)的蛋白表达水平,Von Kossa染色法观察血管钙化情况。结果 (1)成功制备了大鼠血管钙化模型:Von Kossa染色可见钙化组大鼠血管中膜大量黑色颗粒样钙盐沉积,而对照组血管结构完好,未见黑色钙盐沉积物。(2)睾酮对血管钙化的影响:睾酮组钙含量、ALP、BMP-4、OPN水平显著低于钙化组(P0.01),且高剂量睾酮组低于低剂量睾酮组,对照组水平最低; Von Kossa染色可见钙化组血管中膜出现大量黑色颗粒样钙盐沉积,而低剂量睾酮组和高剂量睾酮组均见少量钙盐沉积,对照组无钙盐沉积。结论外源性睾酮能一定程度上减轻维生素D3和尼古丁诱导的大鼠血管钙化。