实验性兔主动脉粥样硬化易损斑块模型的建立与评价

目的探索建立实验性兔主动脉粥样硬化易损斑块模型的新方法。方法24只雄性日本大耳白家兔随机分为对照组8只,实验组16只,对照组给予普通饲料;实验组给予高胆固醇饲料、注射牛血清白蛋白及进行腹主动脉球囊拉伤术,分别于0周、3周、6周、10周检测血脂、ox-LDL、TNF-α、IL-1、IL-10,实验结束时取腹主动脉进行病理形态学分析及NF-κBp65亚基免疫组化染色分析。结果实验组斑块内膜面积比为53.6%,脂核面积与斑块面积比为54.9%,斑块纤维帽厚度与内膜中膜厚度（IMT）比约为8.5%;除甘油三酯变化不大外,实验组TC、LDL-C、HDL-C、LDL-C/HDL-C、TC/HDL-C、ox-LDL、TNF-α、IL-1、IL-10均有明显升高趋势,在3周、6周、10周时与对照组比较差异有统计学意义;且NF-κBp65亚基阳性染色面积较对照组高13.5倍。结论高脂喂养、免疫损伤加球囊拉伤可以成功建立家兔主动脉粥样硬化易损斑块模型。     

骨髓干细胞动员对动脉粥样硬化破裂斑块的稳定与修复作用

目的研究自体骨髓干细胞动员对兔动脉粥样硬化（AS）破裂斑块的稳定与修复作用。方法用液氮冻伤术创建兔AS破裂斑块模型,动员组注射重组人粒细胞刺激因子（rhG-CSF）动员自体骨髓干细胞,对照组注射等量生理盐水,连续5 d。动员第5天抽血分离获取单个核细胞,BrdU标记后经静脉注入动物体内;分别于动员后3d和4周末抽血,ELISA法检测兔血清MMP-9、hsC-RP及PAI-1水平;动员后4周处死兔,HE染色和Masson三色染色观察斑块病理形态,免疫组化染色观察BrdU在斑块区表达情况。结果动员5 d后,动员组兔外周血有核细胞计数及单核细胞比例明显增高;动员后4周,动员组新生内皮细胞及胶原纤维明显增多,在斑块区发现有BrdU标记的阳性细胞,动员组血清MMP-9、hsC-RP及PAI-1水平明显降低。结论应用rhG-CSF动员自体骨髓干细胞能通过促进血管内皮细胞和胶原纤维再生,降低炎症因子及凝血纤溶因子而稳定与修复AS破裂斑块。     

脾虚证IBS模型WHBE兔血浆环核苷酸水平的变化

目的观察脾虚型肠易激综合征（IBS）模型WHBE兔环核苷酸水平的变化。方法取WHBE兔和日本大耳白兔各18只,分别随机分成两组,即正常对照组6只,模型对照组12只,采用湿热应激加灌服番泻叶煎剂的方法造成脾虚证肠易激综合征（tBs）免模型,其中模型对照组造模后处理6只,剩余兔自然恢复10d,观察实验兔的血浆环核苷酸水平的变化。结果与正常对照组比较,WHBE兔模型对照组血浆cAMP和cGMP水平均显著升高（P〈0．05,P〈0．01）,且自然恢复10d后血浆cAMP和cGMP水平仍高于正常对照组（P〈O．05,P〈O．01）;日本大耳白兔模型对照组造模后及自然恢复10d后血浆cAMP和cGMP水平略有所升高,但差异均不显著（P〉O．05）;且模型对照组和自然恢复组cAMP／cGMP比值均略低于正常对照组,但差异不显著（P〉O．05）。结论本实验脾虚证IBS模型兔的血浆环核苷酸水平随阴阳盛衰而变化,具有双重特点：既有阴虚特点,又有阳虚特点,且WHBE兔更适合于脾虚证IBS模型研究理想的实验动物。     

鸡传染性支气管炎病毒E蛋白基因的表达

鸡传染性支气管炎(Infectious Bronchitis,IB)是由传染性支气管炎病毒(Infectious Bronchitis Virus,IBV)引起的鸡的一种急性、高度接触性传染病。该病是危害全球养禽业的重要传染病之一[1,2]。其感染的主要特征是气管啰音、咳嗽和打喷嚏?送猓?该病还可以引起蛋鸡产蛋量下降和蛋品质下降。雏鸡可由于呼吸道或肾脏的感染而死亡。虽然疫苗的使用对IB的流行起到了一定的预防和控制作用,但由于IBV血清型众多,不同血清型毒株之间具有较小的交叉保护性甚至无交叉保护性。因此,IB目前仍在免疫鸡群和非免疫鸡群发生和传播,给养禽业造成了严重…     

Immunolocalization of GnRHRI,gonadotropin receptors,PGR, and PGRMCI during follicular development in the rabbit ovary

The aim of this study was to investigate the presence and localization of gonadotropin-releasing hormone receptor-I (GnRHRI), gonadotropin receptors (FSHR, LHR), progesterone receptor (PGR), and progesterone receptor membrane-binding component-I (PGRMCI) in the different developmental stages of the rabbit follicle. The ovaries were collected from four healthy New Zealand white rabbits, and the mRNA expression and protein levels of GnRHRI, FSHR, LHR, PGR, and PGRMCI were examined with real-time PCR and immunohistochemistry. The results showed that GnRHRI, FSHR, LHR, PGR, and PGRMCI mRNA was expressed in the ovary; furthermore, we show cell-type specific and follicular development stage-specific expression of these receptors at the protein level. Specifically, all of the receptors were detected in the oocytes from the primordial to the tertiary follicles and in the granulosa and theca cells from the secondary and tertiary follicles. In the mature follicles, all receptors were primarily localized in the granulosa and theca cells. In addition, LHR was also localized in the granulosa cells from the primordial and primary follicles. With follicular development, the expression level of all of the receptors, except GnRHRI, in the follicles showed a tendency to decrease because the area of the follicle increased sharply. The expression level of GnRHRI, FSHR, and PGR in the granulosa and theca cells showed an increasing trend with ongoing follicular development. Interestingly, the expression level of FSHR in the oocytes obviously decreased from the primary to the tertiary follicles, whereas LHR in the oocytes increased from the secondary to tertiary follicles. In conclusion, the expression of GnRHRI, the gonadotropin receptors, PGR, and PGRMCI decreased from the preantral follicles (primordial, primary, and secondary follicles) to the tertiary follicles. The expression of GnRHRI and LHR in the oocytes increased from the secondary to the tertiary follicles, whereas FSHR decreased from the primary to the tertiary follicles. The expression of GnRHRI and PGR in the granulosa and theca cells increased from the secondary to the mature follicles. These observations suggest that these receptors play roles in follicular development and participate in the regulation of follicular development.     

PolarCath cryoplasty enhances smooth muscle cell apoptosis in a rabbit iliac artery model: an experimental in vivo controlled study

Purpose

To in vivo investigate the histological response after single and double cryoplasty therapy in a rabbit iliac artery model.

Materials and methods

In total, 40 New Zealand White rabbits underwent percutaneous transluminal angioplasty of the iliac artery using either PolarCath balloon or a conventional angioplasty balloon of equal diameter. Arterial injury, inflammatory response and smooth muscle cells (SMC) apoptosis with the TUNEL (Terminal deoxynucleotidyl transferase dUTP Nick End Labeling) immunohistochemical assay were analyzed. Rabbits were divided between single or double balloon inflation and histological results were compared between cryoplasty and control angioplasty both at 30 min and 72 h.

Results

Arterial injury and wall inflammation scores were low and similar between cryoplasty and control groups after single and double balloon inflation. Compared to conventional balloon angioplasty, Polarcath cryoplasty demonstrated superior SMC apoptosis after single inflation at 30 min [12.0 ± 1.2 cells/(0.025 mm)2 vs 7.0 ± 1.5 cells/(0.025 mm)², p = 0.002], single inflation at 72 h [9.0 ± 1.0 cells/(0.025 mm)² vs 5.4 ± 1.4 cells/(0.025 mm)², p = 0.001], double inflation at 30 min [11.6 ± 1.5 cells/(0.025 mm)² vs 6.8 ± 1.4 cells/(0.025 mm)², p = 0.001] and double inflation at 72 h [9.2 ± 0.8 cells/(0.025 mm)² vs 5.0 ± 0.7 cells/(0.025 mm)², p = 0.001]. There were no significant differences in apoptosis between single and double cryoplasty application at 30 min and 72 h.

Conclusion

Cryoplasty demonstrated superior rates of SMC apoptosis at 30 min and 72 h and was associated to relatively low arterial injury and inflammation scores. An immediate second PolarCath inflation did not achieve superior apoptosis.     

The proton gradient of secretory granules and glutamate transport in blood platelets during cholesterol depletion of the plasma membrane by methyl-β-cyclodextrin

Glutamate transport in blood platelets resembles that in brain nerve terminals because platelets contain neuronal Na⁺-dependent glutamate transporters, glutamate receptors in the plasma membrane, vesicular glutamate transporters in secretory granules, which use the proton gradient as a driving force, and can release glutamate during aggregation/activation. The acidification of secretory granules and glutamate transport were assessed during acute treatment of isolated platelets with cholesterol-depleting agent methyl-β-cyclodextrin (MβCD). Confocal imaging with the cholesterol-sensitive fluorescent dye filipin showed a quick reduction of cholesterol level in platelets. Using pH-sensitive fluorescent dye acridine orange, we demonstrated that the acidification of secretory granules of human and rabbit platelets was decreased by ∼15% and 51% after the addition of 5 and 15 mM MβCD, respectively. The enrichment of platelet plasma membrane with cholesterol by the application of complex MβCD-cholesterol (1:0.2) led to the additional accumulation of acridine orange in secretory granules indicating an increase in the proton pumping activity of vesicular H⁺-ATPase. MβCD did not evoke release of glutamate from platelets that was measured with glutamate dehydrogenase assay. Flow cytometric analysis did not reveal alterations in platelet size and granularity in the presence of MβCD. These data showed that the dissipation of the proton gradient of secretory granules rather than their exocytosis caused MβCD-evoked decrease in platelet acidification. Thus, the depletion of plasma membrane cholesterol in the presence of MβCD changed the functional state of platelets affecting storage capacity of secretory granules but did not evoke glutamate release from platelets.     

Comparison studies of the structural stability of rabbit prion protein with human and mouse prion proteins

Background: Prion diseases are fatal and infectious neurodegenerative diseases affecting humans and animals. Rabbits are one of the few mammalian species reported to be resistant to infection from prion diseases isolated from other species (I. Vorberg et al., Journal of Virology 77 (3) (2003) 2003-2009). Thus the study of rabbit prion protein structure to obtain insight into the immunity of rabbits to prion diseases is very important.Findings: The paper is a straight forward molecular dynamics simulation study of wild-type rabbit prion protein (monomer cellular form) which apparently resists the formation of the scrapie form. The comparison analyses with human and mouse prion proteins done so far show that the rabbit prion protein has a stable structure. The main point is that the enhanced stability of the C-terminal ordered region especially helix 2 through the D177-R163 salt-bridge formation renders the rabbit prion protein stable. The salt bridge D201-R155 linking helixes 3 and 1 also contributes to the structural stability of rabbit prion protein. The hydrogen bond H186-R155 partially contributes to the structural stability of rabbit prion protein.Conclusions: Rabbit prion protein was found to own the structural stability, the salt bridges D177-R163, D201-R155 greatly contribute and the hydrogen bond H186-R155 partially contributes to this structural stability. The comparison of the structural stability of prion proteins from the three species rabbit, human and mouse showed that the human and mouse prion protein structures were not affected by the removing these two salt bridges. Dima et al. (Biophysical Journal 83 (2002) 1268-1280 and Proceedings of the National Academy of Sciences of the United States of America 101 (2004) 15335-15340) also confirmed this point and pointed out that “correlated mutations that reduce the frustration in the second half of helix 2 in mammalian prion proteins could inhibit the formation of PrP^Sc”.     

无线遥测和刺激技术建立兔房颤模型的探索

目的探索一种在无线遥测和刺激技术基础上的兔房颤模型的制作。方法新西兰兔皮下植入自主研发的植入式遥测刺激器,植入式遥测刺激器的制作是以TI公司（德州仪器）的MSP单片机和TI公司的RF无线收发芯片CC2250为核心开发设计。优化植入系统设计以满足新西兰兔房颤模型建立的探索实验;植入子植入新西兰兔腹部皮下,采集电极留置于左上肢和右上肢腋下皮下,两个刺激电极分别缝合于左心耳和左心房上,通过无线收发采集和刺激信号;实现利用Powerlab生理记录仪连续监测体表I导联心电信号,并通过专用计算机程序刺激软件,发放间歇（刺激2 s,暂停2 s）高频（频率20 Hz）阈上（强度2 mA,脉宽1 ms）刺激,若间歇期内出现房颤,则人为干预中止刺激,若转为窦性心律,则继续刺激。结果植入式遥测刺激器在体内可稳定工作（包括采集模拟心电信号和发放刺激）30 d,植入新西兰兔体内刺激3周后可诱导出房颤,持续时间〉48 h。结论用新西兰兔代替比格犬建立基于无线遥测和刺激基础上的房颤模型是完全可行的,同时也体现了动物福利优化和替代原则。