肌肉特异性糖皮质激素受体基因敲除对慢性肾脏病骨骼肌消耗的影响

目的:内源性糖皮质激素水平异常升高可引起和加重慢性肾脏病(CKD)引起的骨骼肌消耗。本研究观察肌肉特异性糖皮质激素受体基因敲除(MGRKO)对CKD状态下小鼠骨骼肌消耗的影响。方法:采用5/6肾切除制作CKD小鼠模型,以假手术组(CTL组)为对照组。组织病理学检测胫骨前肌横截面积并计算肌纤维面积分布图;荧光定量PCR检测肌肉萎缩蛋白Fbox-1(Atrogin-1)和肌环指蛋白1(MuRF-1)mRNA水平;Western Blot检测其蛋白表达及Akt/FoxO1信号通路;ELISA方法检测血清皮质酮水平。结果:造模1月后,lox/lox-CKD和MGRKO-CKD小鼠的血尿素氮和皮质酮水平均明显升高;lox/lox-CKD的体重明显下降胫骨前肌湿重明显降低。而MGRKO-CTL和MGRKO-CKD两组间体重无明显差异,胫骨前肌湿重仅轻微降低。组织学观察到lox/lox-CKD胫骨前肌肌纤维明显萎缩,横截面积减少,肌纤维面积分布图明显左移。而MGRKO-CKD组的体重下降不明显,胫骨前肌仍较饱满,肌肉湿重略降低,横截面积略降低,与MGRKO-CTL组比较,肌纤维面积分布图仅轻度左移。荧光定量PCR和Western Blot结果显示lox/lox-CKD组的腓肠肌Atrogin-1和MuRF-1的mRNA和蛋白表达水平显著升高,同时其Akt磷酸化水平(pAkt/Akt)较lox/lox-CTL组显著降低3.1倍,FoxO1磷酸化水平(pFoxO1/FoxO1)较lox/lox-CTL组降低2.3倍;而MGRKO-CKD组的Atrogin-1表达仅轻微上调,MuRF-1则无明显变化;MGRKO-CKD组Atrogin-1蛋白表达水平仅比MGRKO-CTL组升高,同时pAkt/Akt、pFoxO1/FoxO1水平较对照组仅轻微降低。结论:内源性糖皮质激素水平升高在CKD肌肉消耗中发挥重要作用;MGRKO可减轻/阻止CKD状态下的体重下降和肌肉萎缩,下调肌萎缩关键基因Atrogin-1和MuRF-1表达水平;该效应与阻断糖皮质激素影响Akt/FoxO1信号途径有关,抑制糖皮质激素信号通路可能改善CKD肌肉消耗。     

肌卫星细胞功能异常在慢性肾脏病骨骼肌消耗中的机制

目的:骨骼肌消耗是慢性肾脏病(CKD)的主要特征,肌卫星细胞(muscle satellite cells,MSC)是具有自我增殖和更新能力的成肌干细胞,MSC功能异常可引起肌肉萎缩.本研究探讨MSC在CKD小鼠骨骼肌消耗中的作用机制. 方法:采用5/6肾切除制作小鼠CKD模型,以假手术组为对照组.以免疫组化标记胫骨前肌层粘连蛋白(laminin),测量肌纤维横截面积,并计算面积百分位图;RT-PCR检测腓肠肌Pax-7、MyoD、Myf-5、Myogenin及myostatin mRNA表达水平;分离并原代培养骨骼肌MSC,原位免疫组化检测Pax-7和MyoD表达,检测分化后新形成的肌纤维胚胎型肌球货白重链(eMyHC). 结果:造模一个月后,CKD小鼠体重和骨骼肌重量明显下降,形态学表现为胫骨前肌明显变细,肌纤维横截面积减少,面积百分位图明显左移;肌肉组织的Pax-7、MyoD、Myf-5和Myogenin mRNA表达水平均有不同程度下降,myostatin mRNA表达则明显上调;原代培养的MSC中,CKD组的Pax-7和MyoD阳性细胞数目明显低于对照组,分化后新形成的eMyHC阳性肌纤维数目也明显降低. 结论:CKD可引起显著的骨骼肌萎缩,使MSC增殖和分化功能下降,myostatin水平升高可能是抑制肌卫星细胞功能的重要原因.     

AKT/FOXO1信号通路在心力衰竭小鼠骨骼肌萎缩中的作用

目的:探讨AKT/FOXO1信号通路在心力衰竭(HF)小鼠骨骼肌萎缩中的作用。方法:采用主动脉弓横向结扎8周,复制小鼠HF动物模型。采用实时定量-聚合酶链式反应和Western blot技术检测HF小鼠胫骨前肌内,E3连接酶的2个肌肉萎缩特异性指标Atrogin-1和MuRF1,对胫骨前肌中转录因子FOXO1和激酶AKT的磷酸化水平和总蛋白水平进行测定,并比较磷酸化蛋白和总蛋白的比率。结果:与对照组小鼠相比,TAC 8周诱导HF小鼠的胫骨前肌肌纤维变小,质量减轻。RT-PCR分析结果显示,HF小鼠胫骨前肌内的Atrogin-1和MuRF1的mRNA表达明显上调(P0.01)。Western blot分析结果显示,HF小鼠胫骨前肌组织中Atrogin-1和MuRF1的蛋白相对表达量HF组较对照组明显增加(P0.01)。HF组小鼠胫骨前肌中p-FOXO1的表达水平较对照组增加,FOXO1的总蛋白水平显著下降;p-AKT的蛋白表达较对照组增加(P0.05),AKT的总蛋白水平差异无统计学意义。p-FOXO1/FOXO1和p-AKT/AKT比率HF组明显高于对照组(P0.01)。结论:AKT/FOXOs信号通路参与HF后骨骼肌萎缩的过程并发挥重要作用。     

肌肉萎缩蛋白Fbox-1基因沉默对糖皮质激素作用下肌管萎缩的影响

目的:研究糖皮质激素地塞米松(dexamethasone,DEX)对体外培养肌管的形态、蛋白质合成/分解代谢及肌肉萎缩蛋白Fbox-1(Atrogin-1)的表达,Atrogin-1基因沉默是否可减轻肌管萎缩.方法:体外培养小鼠肌纤维细胞株C2C12细胞并分化为肌管后,用DEX处理48h,同位素3H-酪氨酸掺入法检测蛋白合成,3H-酪氨酸释放率检测蛋白分解代谢;荧光显微镜观察肌管形态并拍照;Northern blot检测Atrogin-1 mRNA水平;应用小分子干扰RNA片段(siRNA)技术使Atrogin-1基因沉默后,观察DEX作用下肌管形态的改变.结果:DEX 5μmol/L作用后,肌管酪氨酸(tyrosine)掺入率下降17.4%,同时tyrosine释放率升高24.7%,使肌管形态变细、萎缩;剂量依赖性地升高Atroign-1蛋白表达水平,使Atroign-1 mRNA表达升高3倍;Atrogin-1基因沉默可改善DEX引起的肌管萎缩. 结论:DEX促进肌管蛋白质分解代谢,并抑制蛋白合成代谢,导致肌肉萎缩;Atrogin-1基因沉默可改善DEX引起的肌肉萎缩,Atrogin-1基因可能是逆转肌肉消耗性营养不良的有效靶点.     

NLRP3炎症小体在ICU获得性衰弱大鼠呼吸肌和下肢肌中的表达

目的探讨重症监护室获得性衰弱(ICU-AW)大鼠吸肌和下肢肌组织中NOD样受体热蛋白结构域相关蛋白3(NLRP3)炎症小体的表达及作用。 方法将40只健康雄性SD大鼠完全随机分为对照组(sham组)和实验组(脓毒血症组),采用盲肠结扎穿孔术在实验组大鼠中构建ICU-AW模型,对照组仅行开腹暴露盲肠手术;造模96 h后收集大鼠腓肠肌和膈肌标本,采用HE染色观察病理学变化并计算肌纤维横截面积,采用qRT-PCR和western blot的方法检测大鼠腓肠肌和膈肌中Atrogin-1、MuRF1、NLRP3、Caspase-1、IL-1β和IL-18的表达。 结果盲肠结扎穿孔术96 h后,大鼠腓肠肌和膈肌纤维排列较对照组疏松,肌纤维横截面积较对照组明显减少(P<0.05),Atrogin-1、MuRF1基因和蛋白表达量较对照组明显升高(P<0.05);ICU-AW大鼠腓肠肌和膈肌中NLRP3、Caspase-1、IL-1β和IL-18基因和蛋白表达量较对照组明显升高(P<0.05)。 结论盲肠结扎穿孔术制备的脓毒血症模型中,吸肌和下肢肌溶解参与ICU-AW的发生,NLRP3炎症小体相关信号通路参与上述过程。     

miR-21对早期糖尿病肾病db/db小鼠蛋白尿的影响

目的 了解miR-21对早期糖尿病肾病db/db小鼠24 h尿白蛋白、肾小球形态学及其分子机制的影响.方法 24只5周龄雄性小鼠分为4组:6只db/m小鼠作为对照组,18只5周龄高血糖且无蛋白尿的db/db小鼠分别随机分为pGenesil-miR-21质粒处理组、对照空质粒处理组及未处理组,每天尾静脉液压法注射质粒30 mg·kg-1·d-1,直至未处理组db/db小鼠尿白蛋白排泄(UAE)显著高于db/m小鼠组.在光学显微镜下观察肾小球形态大小,计算机计算肾小球面积,实时RT-PCR检测肾脏组织miR-21的表达水平,Western印迹、免疫组化PTEN、p-Akt(Ser473)和PI3K p85α肾脏组织的蛋白质表达水平.结果 5周龄db/db小鼠连续注射质粒4周后,未处理组db/db小鼠出现高血糖伴UAE显著增高,提示9周龄db/db小鼠进入早期糖尿病肾病阶段.实时RT-PCR结果显示,在注射pGenesil-miR-21重组质粒的db/db小鼠肾脏组织中,miR-21的表达显著增高(P<0.01).光镜观察和肾小球面积测量实验发现,pGenesil-miR-21重组质粒组肾小球显著减小.Western印迹和免疫细胞化学结果显示,miR-21处理组的db/db小鼠肾脏组织中PTEN蛋白较注射对照空质粒和未注射质粒的db/db小鼠组显著降低,而PI3K p85α和磷酸化Akt (Ser473)蛋白显著增高(均P<0.01).结论 miR-21通过特异性靶向调控PTEN/PI3K信号传导途径,以抑制肾小球肥大,可能是一个新的糖尿病肾病的保护因子.     

心脏营养素-1对失神经骨骼肌收缩功能的促进作用

目的 观察外源性心脏营养素-1(CT-1)对失神经骨骼肌收缩功能的促进作用,并探讨其机制.方法 将120只小鼠随机分为12组各10只,制作腓肠肌失神经模型,6个实验组腹腔注射重组小鼠CT-1 100μg/(kg·d),6个对照组注射等量CT-1溶媒,分别于给药后1、2、4周,检测失神经腓肠肌湿重、横截面积和收缩相关蛋白变化,并于神经修复后4、8、12周,检测神经再支配腓肠肌的复合肌肉动作电位(CMAP)和肌张力.结果 与对照组比较,实验组失神经骨骼肌湿重和纤维直径增加,α-肌动蛋白、肌球蛋白重链Ⅱ和Ca2 -ATP酶水平升高,神经再支配骨骼肌的CMAP和肌张力提高.结论 外源性CT-1可促进失神经骨骼肌重获神经支配后收缩功能的恢复;其机制为通过提高失神经骨骼肌收缩相关蛋白含量,改善失神经骨骼肌收缩的酶学基础.     

盐酸小檗碱对糖尿病小鼠胰岛β细胞增殖与凋亡的影响

目的:探讨盐酸小檗碱对2型糖尿病db/db小鼠胰岛β细胞增殖和凋亡的影响及其机制。方法:选取32只6周龄db/db雄性小鼠和32只db/m雄性小鼠,各分为2组,各选其中一组每日固定时间给予小檗碱(50 mg/kg)灌胃,另一组给予相同体积的生理盐水。连续给药3周后处死小鼠,实时荧光定量PCR和免疫组化法检测胰腺组织Ki-67、B细胞淋巴瘤/白血病2(Bcl-2)、Fas和Fas配体(Fas-L)mRNA和蛋白的表达,末端转移酶标记技术(TUNEL法)检测胰腺组织β细胞凋亡数。结果:实验3周后,小檗碱干预db/db组小鼠的血糖、葡萄糖曲线下面积显著降低,胰岛素分泌显著改善(均P0.05);Bcl-2阳性表达率显著升高(P<0.01);Fas及Fas-L阳性率表达水平显著降低(P<0.01,P<0.05);β细胞凋亡率下降(P     

Foxo1及泛素溶酶体系统在慢性肾脏病致骨骼肌萎缩作用中的初步观察

 目的 观察终末期肾病患者骨骼肌萎缩表现,并初步探讨腹直肌中转录因子Foxo1及泛素溶酶体系统活性变化与骨骼肌萎缩的关系。方法 对慢性肾脏病(CKD)5期的22例尿毒症患者在腹膜透析置管术中行腹直肌活检。并将8例诊断为子宫腺肌病并拟行经腹全子宫切除术患者及6例确诊为腹壁疝并拟行腹壁疝修补术患者设为对照组,于手术时留取少许腹直肌标本。观察肌纤维形态、计算肌纤维横截面积,并检测Foxo1及泛素溶酶体系统标志物Atrogin-1及MuRF1的mRNA和蛋白的表达情况。结果 CKD组患者腹直肌肌纤维的横截面面积较对照组明显缩小(P<0.01),CKD组Foxo1、Atrogin-1及MuRF1的mRNA含量及蛋白表达量均较对照组明显增加(P<0.05),而p-Foxo1蛋白表达量较对照组降低(P<0.05)。结论 CKD患者存在肌萎缩现象,推测其与Foxo1表达上调、磷酸化程度降低以及泛素溶酶体系统活性亢进有关。     

红景天、红景天苷对小鼠骨骼肌质量的调控作用及机制

目的探讨红景天、红景天苷对小鼠下肢骨骼肌质量和慢肌、快肌表达的影响及作用机制。方法 40只小鼠经过预实验筛选后,选取其中30只分为对照组、红景天组和红景天苷组。检测比目鱼肌、跖肌的骨骼肌湿重及Troponin I-SS(Tn I-SS)、Troponin I-FS(Tn I-FS)、Akt、p-Akt(Ser473)、TβR-Ⅱ的蛋白表达,Atrogin-1和MuRF-1的mRNA表达,分析调控机制。结果各组小鼠体重未见明显差异,肌湿重略有增加,差异无统计学意义。与对照组相比,红景天苷组小鼠比目鱼肌和跖肌中Tn I-SS和Tn I-FS表达增加,使骨骼肌质量得以提升。红景天苷可有效提升Akt及其磷酸化水平,抑制TβR-Ⅱ的表达,抑制Atrogin-1和MuRF-1的mRNA表达。结论红景天苷可促进小鼠骨骼肌Tn I-SS和Tn I-FS表达,从而使骨骼肌质量得以提升;促进蛋白质合成关键因子Akt及其磷酸化的表达,抑制TβR-Ⅱ表达,可能为调控小鼠骨骼肌质量的机制。     

硫辛酰胺对db/db小鼠肝损伤的保护作用研究

目的观察硫辛酰胺(ALM)对db/db小鼠肝损伤的保护作用及可能机制。方法db/db小鼠随机分为糖尿病组(DM)和ALM组,C57BL/6J小鼠为正常对照组(NC),每组各6只。ALM组于第9周时予ALM[100 mg/(kg·d)]进行灌胃干预,干预8周后处死小鼠,检测生化指标及肝组织谷丙转氨酶(ALT)、谷草转氨酶(AST)、过氧化氢酶(CAT)活性及丙二醇(MDA)表达量,油红O、HE染色观察肝脏病理学改变,Western blot法检测肝组织中核因子E2相关因子2(Nrf2)、血红素氧合酶1(HO-1)蛋白水平。结果与NC组比较,DM组体重、TG、TC、FBG升高,MDA含量升高[(0.73±0.04)vs(0.92±0.17)nmol/mg,P<0.05],CAT活性降低[(1.08±0.18)vs(0.52±0.14)U/mg,P<0.05],ALT、AST活性升高[(16.85±3.84)vs(22.42±4.56)U/g,(6.07±1.91)vs(8.19±1.51)U/g,P<0.05],Nrf2、HO-1的表达升高[(0.33±0.25)vs(1.81±0.34),(0.29±0.13)vs(1.25±0.19),P<0.05]。与DM组比较,ALM组体重、TG、TC降低,MDA降低[(0.92±0.17)vs(0.56±0.11)nmol/mg,P<0.05],CAT活性升高[(0.52±0.14)vs(0.91±0.20)U/mg,P<0.05],ALT、AST活性降低[(22.42±4.56)vs(17.08±5.08)U/g,(8.19±1.51)vs(5.10±0.46)U/g,P<0.05],Nrf2、HO-1表达降低[(1.81±0.34)vs(1.01±0.30),(1.25±0.19)vs(0.52±0.17),P<0.05]。结论ALM可抑制T2DM小鼠肝损伤的发生发展,其机制可能是通过改善肝组织脂质沉积、抑制氧化应激,调节Nrf2、HO-1蛋白表达以实现。     

STIM1/Orai1可能是防治缺血性脑血管病的新靶点

近年来,通过RNA干扰技术在果蝇细胞中鉴定出钙离子释放激活钙通道(Ca2+ releaseactivated calcium channel,CRAC)的2种重要组成蛋白,即位于内质网上的Ca2+感受器间质相互作用分子1(stromal interaction molecule 1,STIM1)和位于细胞膜上的CRAC通道蛋白Orai1.研究表明,STIM1和Orai1对血管平滑肌细胞、血小板、血管内皮细胞等多种细胞均有调控作用,在血管平滑肌细胞表型转化、止血、血栓形成、新生血管发生等方面发挥着重要作用,显示两者可能与缺血性脑血管病密切相关.文章就STIM1和Orai1蛋白在缺血性脑血管病方面的研究进展进行了综述,以探讨STIM1和Orai1作为防治缺血性脑血管病的新靶点的可能性.     

血管紧张素Ⅱ受体阻断剂治疗对db/db糖尿病小鼠胰岛的作用

目的探讨长期坎地沙坦治疗能否延缓糖尿病状态下胰岛功能的进行性衰竭和高血糖的恶化。方法将8周龄的db／db小鼠随机分为4组,分别给予坎地沙坦酯1、10mg／kg体重、马尼地平10mg／kg体重和安慰剂灌胃治疗6周,同窝出生的8周龄db／m小鼠作为非糖尿病对照亦给予安慰剂治疗。投药6周后行糖耐量试验后取胰腺,进行胰岛素、氧化应激损伤标志8-OHdG的免疫组化检测以及电镜观察胰岛β细胞的超微结构紊乱情况。结果长期坎地沙坦治疗轻微改善糖耐量,减轻了胰岛含量随糖尿病进展的丧失。坎地沙坦治疗显著减少氧化应激产物8-OHdG在胰岛中的水平,电镜观察发现坎地沙坦治疗显著增加β细胞内胰岛素颗粒的形成,明显减轻db／db糖尿病小鼠β细胞内质网和高尔基体的过度增殖以及线粒体肿胀,抗氧化作用和减轻线粒体肿胀效应均呈现剂量依赖性,而且所有保护效应均独立于血压的下降程度。结论在糖尿病起病后,坎地沙坦治疗并不能逆转糖尿病,但是能轻微改善糖耐量,并通过减轻氧化应激损伤、胰岛β细胞超微结构紊乱,从而保护胰岛β细胞功能,延缓β细胞功能衰竭,这些保护作用均独立于降压之外。     

PEGylation improves the hypoglycaemic efficacy of intranasally administered glucagon-like peptide-1 in type 2 diabetic db/db mice

Aims:  PEGylation – covalent modification of therapeutic peptides with polyethylene glycol (PEG) – is viewed as an effective way of prolonging the short lifetime of glucagon-like peptide-1 (GLP-1). In this study, we investigated the hypoglycaemic efficacies of PEGylated GLP-1s administered intranasally in type 2 diabetic db / db mice.
Methods:  Three types of site-specific (Lys³⁴) PEGylated GLP-1 analogues (PEG molecular weight: 1, 2 or 5 kDa) were synthesized. Their metabolic stabilities were evaluated in nasal mucosa enzyme pools. Oral glucose tolerance test was conducted 30, 60 and 120 min after intranasally administering these analogues in type 2 diabetic db / db mice.
Results:  PEGylated GLP-1 analogues were found to have significantly longer half-lives than native GLP-1 in nasal mucosa enzymes (2.4-fold to 11.0-fold, p < 0.005). Non-PEGylated GLP-1 at 100 nmol/kg was not found to have marked efficacy irrespective of nasal administration time [total hypoglycaemic degree (HD_total) values 2.8–17.3%]. On the contrary, PEGylated GLP-1s (100 nmol/kg) showed obvious efficacies with maximum HD_total values of >51.8 ± 5.8% (p < 0.005 vs. GLP-1).
Conclusion:  This study highlights the pharmacological potential of intranasally administered PEGylated GLP-1s in terms of stabilizing postprandial hyperglycaemia in type 2 diabetic patients.     

PEGylated glucagon-like peptide-1 displays preserved effects on insulin release in isolated pancreatic islets and improved biological activity in db/db mice

Aims/hypothesis The rapid degradation and clearance of glucagon-like peptide-1 (GLP-1) by the enzymes dipeptidyl peptidase-IV and neutral endopeptidase 24.11 are the main impediments to the development of GLP-1 as a potential glucose-lowering agent. In this study, new enzyme-resistant polyethylene glycol (PEG)-conjugated GLP-1 analogues were designed and examined for metabolic stability and biological potency.Materials and methods Two mono-PEGylated GLP-1 analogues, N-terminally modified N-PEG/GLP-1 and Lys-modified Lys-PEG/GLP-1, were prepared. Stability was tested in plasma and tissue extracts. In vitro insulin release studies were performed using isolated rat pancreatic islets, while in vivo glycaemic responses were measured in db/db mice.Results The half-life of Lys-PEG/GLP-1 was 40-, 10- and 28-fold longer than that of GLP-1 in plasma, liver and kidney homogenates, respectively. Lys-PEG/GLP-1 stimulated insulin secretion in the islets in a dose- and glucose-dependent manner, and was as potent as GLP-1. In contrast, N-PEG/GLP-1 showed extended metabolic stability but had significantly lower biological activity. The administration of Lys-PEG/GLP-1 (9 nmol/kg i.p.) to non-fasted db/db mice stabilised glycaemia (p<0.001), whereas GLP-1 (9 nmol/kg) only caused small changes in glucose level. During OGTT in fasted db/db mice, Lys-PEG/GLP-1 administered at 1, 3 and 9 nmol/kg (i.p.) reduced the glucose AUC_0–3h by 48.7±9.4, 55.0±2.9 and 63.4±2.5%, respectively, compared with placebo (p<0.01), whereas GLP-1 (9 nmol/kg) lowered the glucose level by 39.5±12.9% (p<0.01).Conclusions/interpretation This study demonstrates that site-specific PEGylated GLP-1 analogues are resistant to degradation. The enhanced biological potencies of these analogues highlight their potential as new, GLP-1-like glucose-lowering agents.     

Altered bone metabolism in db/db mice]

Bone and calcium metabolism was investigated in genetically obese, diabetic db/db mice and compared with that in a new hypoglycemic agent (AS-6) treated db/db mice and in their lean litter mates as controls. The 5-week-old db/db mice (serum Ca 9.88 +/- 0.22 mg/dl, glucose 258.6 +/- 13.3 mg/dl) were randomly divided into two groups. One group, together with their lean litter mates, was fed a commercial diet (CE-2). The other db/db group was fed CE-2 diet containing 0.1% of AS-6. Both groups were fed for 20 weeks. The serum glucose and calcium levels in db/db control groups (serum Ca 12.3 +/- 0.1 mg/dl, glucose 650.2 +/- 23.9 mg/dl) were higher than those in lean control groups (Ca 9.8 +/- 0.2 mg/dl, glucose 180.7 +/- 10.1 mg/dl). The wet, dry and ashed weights of the femur in db/db control were significantly lower and the length of femur in db/db control was significantly shorter than those of lean controls. These data suggest that retarded bone growth in db/db mice is related to progression of diabetes. Although, there was no change in Ca/P, Ca/ash and total perimeter in femurs, the cortical area in the femurs of db/db control mice (0.65 +/- 0.02 mm2) was significantly smaller than that of the femurs of lean control mice (0.74 +/- 0.02 mm2). The cortical bone thinning observed in the db/db control could have been caused by increased bone resorption. Treatment with AS-6 for 20 weeks resulted in a 48.6% decrease of serum glucose and 5.2% decrease of calcium as compared with db/db controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Oxidative damage in cerebral vessels of diabetic db/db mice

BACKGROUND: Oxidative stress in diabetes mellitus has recently received increasing attention as it has been proven to be associated with the development of diabetic vascular complications. Our aim was to examine whether microvascular changes, including oxidative damage, were induced in the brains of diabetic animals. METHODS: The expression of 8-hydroxy-2'-deoxyguanosine (8-OHdG), a marker of oxidative DNA damage, the binding of cationized ferritin, a marker for evaluating endothelial glycocalyx, to the endothelial cells of capillaries and vascular permeability of intravenously injected horseradish peroxidase were examined in the cortices of 12- and 20-week-old db/db and db/+m mice. RESULTS: Immunostaining for 8-OHdG was clearly seen in the vessels of the cortex of 20-week-old db/db mice, but was hardly seen in those of mice in the other groups. The immunopositive area of 8-OHdG was significantly increased in the cortex of 20-week-old db/db mice compared with that of 20-week-old db/+m mice. No extravasated leakage of horseradish peroxidase was seen in any groups of mice, while the numbers of cationized ferritin particles binding to the endothelial cells was significantly decreased in 12- and 20-week-old db/db mice compared with that of db/+m mice at the same age, respectively. CONCLUSION: These findings suggest that changes in endothelial glycocalyx are induced in db/db mice and, in addition, the long-term diabetic condition of these mice induces oxidative DNA damage to the cerebral vessels.     

替米沙坦对db/db小鼠胰岛内微血管结构的影响

目的探讨替米沙坦阻断肾素血管紧张素系统对db/db小鼠胰岛内微血管结构的影响。方法将22只8周龄db/db小鼠随机分为2组:替米沙坦组11只和安慰剂组11只;另选11只同周龄db/m小鼠为非糖尿病组。每周监测小鼠体重、血糖,6周后行腹腔葡萄糖耐量试验(IPGTT)并取胰腺行免疫组织化学检测,分析胰岛内微血管中CD31表达情况。结果治疗6周后,与安慰剂组比较,替米沙坦组小鼠空腹血糖明显降低,胰岛素敏感指数明显升高,差异有统计学意义(P0.05);非糖尿病组小鼠的体重和空腹血糖明显降低,差异有统计学意义(P0.05)。替米沙坦组小鼠葡萄糖曲线下面积明显下降,0～30 min胰岛素曲线下面积明显上升,差异有统计学意义(P0.05);替米沙坦组小鼠胰岛中血管内皮细胞CD31染色强度明显增强,血管数量明显增加。结论替米沙坦阻断血管紧张素Ⅱ1型受体能够改善胰岛微环境,增加胰岛内微血管数量,促进早期相胰岛素分泌,从而保护胰岛β细胞。