乙酰肝素酶在糖尿病肾病大鼠蛋白尿发生中的作用

目的 观察糖尿病肾病大鼠肾组织乙酰肝素酶（HPA）的表达，探讨其在糖尿病肾病大鼠蛋白尿发生中的作用。 方法 SD健康大鼠被随机分为健康对照组（n = 6）、糖尿病6周组(DM6w, n = 10)和糖尿病12周组(DM12w, n = 10)，采用一次性腹腔注射链脲菌素（STZ）的方法建立糖尿病大鼠模型。分别于造模后6周和12周末测定各组大鼠相对肾质量、血糖、尿素氮、血肌酐、24 h尿量及尿蛋白量(24 h)等指标，并观察肾脏病理改变。RT-PCR和免疫组织化学法检测各组大鼠肾组织HPA mRNA和蛋白表达变化，并分析其与蛋白尿发生的相关性。 结果 （1）DM6w和DM12w组大鼠的相对肾质量、血糖、尿素氮、24 h尿量及尿蛋白量(24 h)与健康对照组相比明显升高, 差异均有统计学意义(P < 0.05或P < 0.01)。（2）DM6w和DM12w组大鼠HPA mRNA和蛋白表达比健康对照组均显著增高(P < 0.01)。（3）大鼠肾组织HPA mRNA和蛋白表达与尿蛋白量(24 h)之间均呈正相关 (r = 0.783，P < 0.01；r = 0.793，P < 0.01)。 结论 HPA在糖尿病肾病中的表达升高可能参与了糖尿病肾病蛋白尿的发生。     

乙酰肝素酶在恶性肿瘤中的研究进展

乙酰肝素酶(HPSE)是目前在哺乳动物中发现的可以降解硫酸乙酰肝素蛋白聚糖(heparan sulfate proteoglycans,HSPGs)上的硫酸肝素(HS)糖链的一类β-D糖苷内切酶.其在肿瘤侵袭转移中起着关键作用.本文就HPSE分子结构、生物学特征、在恶性肿瘤中的表达、促进肿瘤转移机制及HPSE抑制剂的应...     

乙酰肝素酶与肿瘤转移

乙酰肝素酶是一种糖苷内切酶 ,能特异性的水解位于细胞表面 ,细胞外基质及基底膜的硫酸乙酰肝素蛋白聚糖为小分子片段。乙酰肝素酶普遍存在于恶性转移性肿瘤细胞中。乙酰肝素酶参与肿瘤血管生成 ,与肿瘤的侵袭、转移密切相关。本文对此进行了综述。     

乙酰肝素酶的病理生理作用及其在肾脏病的研究进展

乙酰肝素酶系一种内源性葡糖苷酸内切酶,能特异性地水解硫酸乙酰肝素侧链,调节细胞外摹质降解,并释放多种与硫酸乙酰肝素侧链结合的生物活性因子,参与多种生理和病理过程,并在肾小球疾病、特别是蛋白尿的发生发展过程中发挥重要病理作用.本文就乙酰肝素酶的病理生理作用及其在肾脏病的研究进展作一综述.     

乳腺癌浸润和转移与乙酰肝素酶表达的关系

目的 观察乙酰肝素酶(Hpa)在乳腺癌的表达,探讨它们的表达与乳腺癌浸润、转移的关系及临床意义.方法 对60例未行任何化疗、放疗和内分泌治疗的原发性乳腺癌病理标本进行分析,应用苏木素-伊红(HE)染色观察乳腺癌的组织形态和病理学变化;应用免疫组织化学法检测乳腺癌患者癌组织、癌旁2 cm乳腺组织及距癌组织5 cm以上的正常乳腺组织中Hpa的表达.结果 60例乳腺癌病理标本免疫组织化学结果显示,乳腺癌组织Hpa阳性表达率为68.33％ (41/60),癌旁组织(癌旁2 cm)Hpa阳性表达率40.00％(24/60),正常组织Hpa阳性表达率0.00％ (0/60).统计分析结果显示,与正常对照组织比较,Hpa在癌组织和癌旁组织表达升高且差异有统计学意义(P＜0.01).Hpa的表达与乳腺癌临床病理标志的相互关系分析结果显示:Hpa的表达与乳腺癌患者体内瘤块直径、组织学分级、腋下淋巴结状况及临床分期明显相关(P＜ 0.05),与患者年龄无明显相关(P＞0.05).而且具有腋下淋巴结转移的患者Hpa阳性表达率显著高于无腋下淋巴结转移组(P＜0.01).结论 Hpa在乳腺癌中的表达均较高,明显高于癌旁组织及正常乳腺组织;Hpa的表达与乳腺癌肿瘤大小、组织学分级、临床分期、淋巴结转移密切相关,与患者的年龄无明显相关.     

乙酰肝素酶与肿瘤转移

乙酰肝素酶是一种糖苷内切酶，能特异性的水解位于细胞表面，细胞外基质及基底膜的硫酸乙酰肝素蛋白聚糖为小分子片段。乙酰肝素酶普遍存在于恶性转移性肿瘤细胞中。乙酰肝素酶参与肿瘤血管生成，与肿瘤的侵袭、转移密切相关。本文对此进行了综述。     

乙酰肝素酶siRNA对部分肝切除大鼠术后肝再生的抑制作用

目的探讨乙酰肝素酶(HPSE)siRNA对部分肝切除大鼠术后肝再生的抑制作用。方法设计合成HPSE siRNA,以vivo-jetPEI-Gal为载体转染70%肝切除大鼠,以逆转录-聚合酶链反应(RT-PCR)检测转染后HPSE mRNA表达,以Western blot检测AFP、ALB、CK-18和CK-19蛋白表达,以原位末端标记法(Tunel)检测肝细胞凋亡,以免疫组化检测肝组织增殖细胞核抗原(PCNA)表达,并测定肝再生率。结果与各对照组相比,siRNA干扰组HPSE mRNA表达明显降低,AFP、ALB、CK-18和CK-19蛋白表达明显降低,肝细胞凋亡指数明显增高,增殖指数明显降低,肝再生率明显降低,差别显著(P0.05)。结论 siRNA沉默HPSE表达后,部分肝切除大鼠术后肝再生受到明显抑制,提示HPSE对肝再生具有调控作用。     

膀胱癌中乙酰肝素酶的表达和临床意义

目的探讨国人膀胱癌乙酰肝素酶的表达水平及其与膀胱癌发生、发展、转移和复发的关系。方法应用免疫组化EnVisionZ．步法研究58例膀胱癌患者中乙酰肝素酶蛋白的表达水平，10例正常膀胱黏膜为对照组。结果乙酰肝素酶在膀胱癌中阳性表达22例（37．9％），其表达率与肿瘤的病理分级、临床分期、复发和淋巴结转移有相关性（P〈0．05）。结论乙酰肝素酶蛋白的异常表达可作为膀胱癌患者发生和发展的预后参数。     

乙酰肝素酶与蛋白尿性肾脏疾病的关系

蛋白尿是众多肾脏疾病的主要临床表现,肾小球滤过屏障的完整性决定着对蛋白质的选择通透特性。肾小球基底膜中硫酸乙酰肝素在电荷屏障的维持中起重要作用。乙酰肝素酶作为新近克隆出来的一种内切糖苷酶,与肿瘤的转移浸润密切相关。近年来研究发现其通过选择性降解肾小球基底膜硫酸乙酰肝素侧链,也参与蛋白尿的发生。本文就其分子学特点、调控以及与蛋白尿性肾脏疾病的关系作一综述。     

阿霉素肾病大鼠外周血白细胞乙酰肝素酶基因表达与尿蛋白的相关性

研究提示，肾病综合征蛋白尿的产生主要与肾小球滤过屏障电荷筛被破坏有关，即GBM上带多阴离子的侧链硫酸乙酰肝素(heparan sulfate，HS)减少可引起蛋白尿的产生。乙酰肝素酶(heparanase，Hpa)是哺乳动物中唯一能直接分解HS的内切葡萄糖醛酸酶^【1】。肾病中HS侧链减少是否与Hpa活性增高有关?我们拟通过肾病经典动物模型一     

大鼠移植动脉硬化加快模型的建立

目的　建立一种简捷 ,有代表性且稳定的移植物动脉硬化模型。方法　将SD大鼠的腹主动脉分别冷缺血 1、2 4、48h行SD→SD及SD→Wistar的原位腹主动脉移植 ,观察术后植入段血管病理改变、TGF β1表达及手术前后过氧化脂质的变化。结果　SD→SD及SD→Wistar缺血1h组分别于术后 10周及 6周见内膜明显增厚 ,而缺血 2 4h组只需 2周 ;各组移植后 2h过氧化脂质均明显高于术前 ,术后 4、2 4h与术前比差异无显著性 (P >0 .0 5 ) ;强化缺血组TGF β1不论是SD→SD还是SD→Wistar均于术后 1周即出现高表达。结论　以SD/Wistar作为供 /受体行腹主动脉移植 ,强化冷缺血损伤 ,可加快移植物动脉硬化 ,可望成为新型慢排模型。     

紫杉醇对大鼠移植动脉内膜增生及硬化的抑制作用

目的观察紫杉醇对同种大鼠移植动脉的影响,探讨紫杉醇对移植物动脉硬化的抑制作用及机制。方法以W istar大鼠为受者,SD大鼠为供者,按供鼠、受鼠不同分为3组,每组8对:同系对照组W istar大鼠接受W istar大鼠的胸腹主动脉移植;同种对照组W istar大鼠接受SD大鼠的胸腹主动脉移植;同种实验组W istar大鼠接受SD大鼠的胸腹主动脉移植,术后1~14 d腹腔注射紫杉醇2 mg.kg-1.d-1。两个对照组每日腹腔注射相同体积的生理盐水。术后30 d,取出移植动脉,进行病理学观察,测出管腔面积和内膜截面积,计算再狭窄率,用免疫组织化学方法测定增殖细胞核抗原(PCNA)的表达。结果同种实验组移植动脉内膜厚度、内膜PCNA表达情况均较同种对照组明显减少,外膜炎症细胞浸润程度、管腔再狭窄率均较同种对照组降低,差异有统计学意义。结论紫杉醇能有效抑制同种大鼠移植动脉内膜增生,防止移植物动脉硬化,其作用机制可能与抑制血管平滑肌细胞增殖和减轻对移植物的免疫排斥反应有关。     

移植动脉慢性排斥反应的病理组织学观察

目的 了解慢性排斥反应时移植动脉硬化的病理变化。方法 采用病理图象定量分析及电镜等检测手段,观察大鼠动脉移植后3、7、14、20、30及60d的病理变化。结果 同系移植对照组除供者血管冷缺血超过1h的2只术后14d内膜轻度增生外,基余移植动脉与受者自身的正常动脉比较,差异不显著;异系移植实验组术后3d血管外膜有大量炎性细胞浸润,7d时局部内皮剥脱,14d内膜中层细胞通过弹力膜裂隙向内膜迁移,内膜增生,20d内膜全层或局部呈8半月形增生,平滑肌细胞形态由收缩型转变为合成型,30d内膜细胞数达峰值,60d内膜持续增厚,基质纤维成分增多,内膜修复。结论 移植体动脉硬化具有内皮剥脱、炎性细胞浸润、中层坏死、平滑肌细胞迁移及内膜增生4大特点。     

Endothelial nitric oxide synthase protects aortic allografts from the development of transplant arteriosclerosis

BACKGROUND: Inducible nitric oxide synthase (iNOS) is up-regulated in rejecting allografts and is protective against allograft arteriosclerosis; it suppresses neointimal smooth muscle cell accumulation and inhibits adhesion of platelets and leukocytes to the endothelium. However, the functional importance of endothelial NOS (eNOS) in the rejecting allografts remains unclear. METHODS: We examined the effects of selective eNOS deficiency in aortic allografts in a murine chronic rejection model using grafts from eNOS knockout (KO) mice (C57BL/6 background; H2b) and normal C3H (H2K) as recipients. Grafts from wild-type C57BL/6 mice served as controls. Grafts from iNOS KO mice served as a second group of controls where the contribution from iNOS was eliminated but eNOS was preserved. Aortic grafts were harvested and analyzed at days 10-14, 18-22, and 26-30 after transplantation. RESULTS: Endothelial NOS-deficient grafts showed significantly increased intima/media ratios at days 26-30 compared to controls. Immunostaining demonstrated that in eNOS KO grafts, eNOS was not detectable whereas iNOS was expressed prominently in infiltrating recipient mononuclear cells. In control grafts, eNOS expression was preserved in the endothelium even by day 30, and associated with a decrease in intimal thickening. We further demonstrated that early overexpression of iNOS by ex vivo gene transfer completely prevented the development of arteriosclerosis associated with eNOS deficiency. CONCLUSIONS: We found that eNOS plays a protective role in allografts, and that in eNOS-deficient allografts, early overexpression of iNOS is capable of preventing the development of allograft arteriosclerosis. In allografts with dysfunctional vascular endothelium and impaired eNOS activity as a result of ischemia or native arteriosclerotic disease, iNOS gene therapy may serve to improve their long-term survival and function.     

类表皮生长因子域7在动脉硬化闭塞症和腹主动脉瘤中的表达

目的 探讨类表皮生长因子域7(epidermal growth factor-like domain 7,egfl 7)在动脉硬化闭塞症(arteriosclerosis obliterans,ASO)和腹主动脉瘤(abdominal aortic aneurysm,AAA)中的表达特点及其在动脉粥样硬化(atherosclerosis,AS)发病过程中的作用.方法收集正常动脉8例,下肢ASO标本11例,AAA标本34例,应用免疫组织化学技术和原位杂交技术检测egfl7蛋白和egfl7mRNA.应用qRT-PCR技术检测egfl 7 mRNA的表达水平.应用RNA干扰技术检测egfl 7的作用.结果 在ASO和AAA中,egfl 7蛋白主要表达于动脉中膜;egfl7 mRNA的表达部位与egfl 7蛋白一致.在ASO早期、AAA与正常动脉相比,egfl 7 mRNA的表达水平显著上调(325%±120%比100%±36%.P＜0.01,216%±133%比100%±36%,P＜0.05).干扰egfl 7后,平滑肌细胞(smooth muscle cells,SMCs)的增殖能力与对照组相比明显降低(0.85±0.05比1.34±0.04,P＜0.01),吸光度值降低(0.85±0.05比1.32±0.09,P＜0.01).结论 egfl 7可能是调控AS发病过程的一个重要基因,可能通过促进血管平滑肌细胞的增殖起作用,是早期ASO的标志.

Abstract：

Objective To investigate the expression characteristics of epidermal growth factor-like domain 7 ( egfl 7 ) in arteriosclerosis obliterans ( ASO) tissues and abdominal aortic aneurysm ( AAA ) tissues, and it's role in atherosclerosis (AS).Methods In this study, 8 normal artery, 11 lower extremity ASO and 34 AAA samples were collected.Immunohistochemistry and in situ hybridization were performed in artery sections to investigate the expression characteristics of egfl 7 at protein and mRNA levels.The relative quantitative detection of egfl 7 mRNA was detected by qRT-PCR.The effect of egfl 7 was examined by RNA interference.Results In ASO and AAA samples, the expression of egfl 7 protein was mainly in the tunica media; The expression site of egfl 7 mRNA was the same as that of egfl 7 protein.Compared with normal artery samples, the expression of egfl 7 mRNA was significantly upregulated in the early stage of ASO samples and AAA samples (325 ± 120 vs.100 ± 36, P ＜ 0.01, 216 ± 133 vs.100 ± 36, P ＜ 0.05 ).Compared with the control group, the proliferative ability of smooth muscle cells was significantly down regulated after egfl 7 interference (0.85 ± 0.05 vs.1.34 ± 0.04, P ＜ 0.01).Conclusions egfl 7 might be a gene that regulates the pathogenic process of atherosclerosis through promoting vascular smooth muscle cell proliferation.     

Chronic rejection of rat aortic allograft: II. Administration of cyclosporin induces accelerated allograft arteriosclerosis

Abstract. Rat aortic allografts immunosuppressed with cyclosporin - but not with azathioprine or steroids - develop an early inflammatory lesion in the subendothelial space. This "endothelialitis" is followed by an influx of proliferating smooth muscle cells into the intima, resulting in intimal thickening and accelerated arteriosclerosis. Administration of azathioprine and steroids largely ameliorates the development of the accelerated lesion. Similar endothelialitis and accelerated arteriosclerosis have been observed previously in the autopsy material of cardiac transplant recipients. Our results confirm the suggestion that the development of accelerated allograft arteriosclerosis is most likely linked to cyclosporin administration.     

慢性排斥反应移植动脉硬化与增殖细胞核抗原表达

目的 探讨慢性排斥反应移植动脉硬化与增殖细胞核抗原（ＰＣＮＡ）表达的关系。方法 采用免疫组化ＳＰ法研究６１只大鼠的移植动脉，１０例慢性排斥反应移植肾动脉组织中的ＰＣＮＡ表达。结果 实验组：大鼠移植动脉中层ＰＣＮＡ术后７天达最高峰，内膜层术后２０天达最高峰，且细胞数明显增多，两者呈正相关。临床组：移植肾动脉ＰＣＮＡ最表达主要集中在内膜，其次为中层，外膜极少。结论 移植动脉硬化过程中细胞增殖与ＰＣＮＡ     

Combination of clopidogrel and everolimus dramatically reduced the development of transplant arteriosclerosis in murine aortic allografts

Our group has shown that platelet inhibition with clopidogrel, an antagonist of the P2Y12 adenosine diphosphate receptor on platelets, reduced the formation of transplant arteriosclerosis. The aim of this study was to investigate whether a combination of cyclosporin or everolimus with clopidogrel has a beneficial effect on the development of transplant arteriosclerosis. Fully MHC mismatched C57Bl/6 (H2^b) donor aortas were transplanted into CBA.J (H2^k) recipients and mice received either clopidogrel alone (1 mg/kg/day) or in combination with cyclosporin (2 mg/kg/day) or everolimus (0.05 mg/kg/day). Grafts were analysed by histology and morphometry on day 30 after transplantation. In mice treated with clopidogrel alone, transplant arteriosclerosis was significantly reduced [intima proliferation 56 ± 11% vs. 81 ± 7% (control)/n = 7]. Daily application of everolimus reduced the development of transplant arteriosclerosis compared with untreated controls [intima proliferation of 29 ± 9% vs. 81 ± 7% (control)/n = 7]. Strikingly, combination of clopidogrel and everolimus almost abolished the formation of transplant arteriosclerosis [intima proliferation: 11 ± 8% vs. 81 ± 7% (control)/n = 7]. By contrast, combination of cyclosporin and clopidogrel compared with clopidogrel alone showed no additive effect. These results demonstrate that combination of platelet‐ and mammalian target of Rapamycin‐inhibition can dramatically reduce the development of transplant arteriosclerosis.     

腹主动脉移植后小鼠外周血内皮祖细胞数量的动态变化及意义

目的 观察腹主动脉移植后小鼠移植物动脉硬化的病变过程及外周血内皮祖细胞(EPC)数量的动态变化,探讨EPC与动脉内膜损伤和修复的相互关系.方法 以C57BL/6小鼠和Balb/c小鼠为供、受者,建立腹主动脉原位移植后移植性模型.术后3 d、2周、4周、6周,观察移植动脉病理改变,并采用计算机图像分析系统分析移植血管内膜增生情况.使用流式细胞仪监测术后外周血中EPC数量的变化.结果 术后3 d,移植动脉内皮细胞损伤,并伴有明显的炎症细胞浸润.术后2周,即可观察到移植动脉新生内膜形成,存在急性排斥反应;术后4周、6周内膜厚度逐渐增生,移植动脉管腔明显狭窄.术后早期外周血中EPC的数量增多,3 d时达到高峰,此后迅速减少,术后14和28 d时,显著低于术前水平(P＜0.05).结论 通过同种小鼠腹主动脉原位移植可成功复制出移植物动脉硬化的病变特点;外周血中EPC的数量与移植动脉内膜损伤的修复密切相关,可能成为移植物动脉硬化的发病指标和干预靶点.

Abstract：

Objective To investigate changes in the number of endothelial progenitor cells (EPC) from peripheral blood and pathological feature in the development of transplant arteriosclerosis in mouse abdominal aortic allografts, and discuss their correlations. Methods A segment of abdominal aorta was transplanted orthotopically from C57BL/6 to Balb/c mice. The grafts were harvested at 3rd day, 2nd week, 4th week and 6th week after the operation and studied by light and electronic microscopy. Regional changes in the lumen and intima were measured with computer imaging analysis system. EPC from peripheral blood were quantified by flow cytometry. Results Endothelium injury and inflammatory cells infiltration were seen in the aortic allografts at 3rd day after transplantation.Neointimal lesions and acute rejection were observed as early as 2nd week after surgery. The lumen of allografts was significantly narrowed due to neointima hyperplasia and had progressed at 4th and 6th week postoperatively. The number of circulation EPC was increased from 1 st day after operation and reached the peak at 3rd day. Thereafter the number of EPC was decreased rapidly and significantly less at 14th and 28th day postoperation than that pre-operation. Conclusion Abdominal aortic transplantation from C57BL/6 to Balb/c mice presents typical pathological feature of transplant arteriosclerosis. The number of EPC from peripheral blood is related to the process of injured endothelial repair and neointima formation of aortic grafts. EPC count may be considered a novel biological marker and therapeutic intervention for transplant arteriosclerosis.