不同方法制备大鼠肝脏脱细胞支架及其免疫原性研究

目的:肝脏脱细胞支架(decellularized liver bioscaffold,DLB)在组织工程研究中具有良好前景,但对DLB的免疫原性尚未见探讨.本研究利用不同方案制备大鼠DLB,检测支架成分和体内重塑反应,为制备出低免疫原性的DLB提供改进依据.方法:选取文献报道的三种方案(分别以SDS,Triton X-100和NP-40为主要洗脱成分),制备F344大鼠DLB,进行HE和Masson染色,检测DLB中DNA、氨基葡聚糖(GAG)以及羟脯氨酸(HYP)含量;再将DLB埋植于C57BL/6小鼠的背部皮下,于术后第3、7和14天取出后进行形态学观察和组织学评分.结果:三种方案均成功制备出符合脱细胞标准的大鼠DLB;形态学结果提示TritonX-100和NP-40方案较SDS方案能更好地保护肝脏超微结构;成分检测结果发现NP-40方案去除DNA的能力显著强于SDS和Triton X-100方案(P＜0.05),而对GAG含量的洗脱作用最弱(P＜0.05);异种体内埋置实验提示NP-40方案制备的DLB引起的免疫反应强度明显弱于SDS和Triton X-100方案,体内重塑结局评分显著高于SDS和Triton X-100方案.结论:与SDS和TritonX-100方案相比,NP-40方案能更有效地去除肝脏DNA,较好地保留GAG,诱导移植受体对DLB的良性重塑反应,为进一步的体内研究提供更优化的支架.     

BMP9诱导人脐带间充质干细胞体内外成骨分化的作用研究

目的:探讨人骨形态发生蛋白9(bone morphogenetic proteins 9,BMP9)在体内外诱导人脐带间充质干细胞(human umbilical cord-derived mesenchymal stem cells,hUC-MSCs)成骨分化的作用研究。方法:设立Ad-BMP9处理组和Ad-GFP对照组感染hUC-MSCs,两组细胞分别于3天、5天、7天进行ALP活性检测,14天后采用免疫组织化学染色检测骨钙素(Osteocalcin,OCN)、骨桥蛋白(Osteopotin,OPN)的表达情况,21天后茜素红染色检测矿化结节的形成;然后收集不同分组hUC-MSC用于裸鼠皮下注射成骨模型的建立,4周后取出离体骨进行Micro-CT扫描和分析,并进行H&E、Masson Trichrome、Alcain Blue染色。结果:BMP9处理组的ALP活性和矿化结节形成明显高于对照组,免疫组化染色结果显示BMP9诱导组的OCN、OPG的阳性表达明显高于对照组;裸鼠皮下注射成骨模型的观察结果显示,空白对照组没有形成肉眼可见的皮下包块,仅感染Ad-BMP9的hUC-MSCs能生成异位骨,且形成的异位骨骨量明显,骨密度平均值为396.05±0.60;H&E染色结果显示BMP9诱导生成的异位骨中形成部分成熟的骨基质和骨小梁,Masson Trichrome染色结果显示BMP9明显诱导hUC-MSCs的基质矿化作用,Alcain Blue染色结果显示BMP9明显诱导hUC-MSCs的软骨内成骨作用。结论:BMP9成功诱导人脐带间充质干细胞的体内外成骨作用,为临床骨组织工程的细胞疗法提供了明确的可行性。     

利用新型聚羟丁酸酯材料构建组织工程软骨

目的:研究新型聚羟丁酸酯作为组织工程软骨支架材料的可行性.方法:取幼兔软骨组织中软骨细胞体外培养扩增.实验组接种软骨细胞于支架材料上,体外培养两周后埋植于新西兰大白兔背部皮下;对照组埋入未接种细胞的支架材料.扫描电镜观察材料表面形态及细胞生长情况.分别于第4、8、12周取出标本,大体观察后进行HE和Masson染色,观察组织工程软骨形成情况.结果:扫描电镜观察可见裸材料孔隙分布均匀,形状不规则;细胞材料复合体体外培养两周后材料表面爬满细胞且生长状态良好.埋植材料取出后可见不同时间点实验组标本大小无明显变化,对照组标本逐渐变小.HE和Masson染色显示各组支架材料至12周时已被完全吸收;实验组12周时可见较成熟软骨组织;对照组支架材料被吸收后最终被纤维结缔组织取代.结论:此新型聚羟丁酸酯材料可作为组织工程软骨支架材料.     

大豆黄酮对去卵巢猪LH分泌的影响

去卵巢的德国格丁根微型小母猪、于静脉内注射或皮下埋植大豆黄酮,并以雌激素诱导后,观察其对垂体ＬＨ反馈性分泌的影响。试验分两系列。第一系列共三组,每组６头：试验组按每１０μｇ／ｋｇ体重静脉注入大豆黄酮悬浮液,对照Ⅰ组注入相等量大豆黄酮溶剂,对照Ⅱ组作空白对照。第二系列为二组,每组９头动物,试验组皮下埋植大豆黄酮硅胶囊,对照组埋植不含任何药品的硅胶管,埋植期１２天。埋植后第７天,两组同时肌注雌二醇。慢性血管导管采集血样,ＲＩＡ分析血浆ＬＨ。结果表明,系列试验一,试验组注射大豆黄酮后ＬＨ分泌呈下降趋势,ＬＨ平均水平和峰值在处理后第１小时明显低于对照组,尔后与对照组趋于相似。系列试验二,试验组在大豆黄酮埋植后第４天和第７天,ＬＨ平均水平明显低于对照组和自身埋植前。雌激素处理后４８ｈ内,试验组ＬＨ分泌几乎完全被抑制,而对照组仍有明显分泌（Ｐ＜０．００１）。ＬＨ正反馈峰,试验组较对照组晚１２ｈ出现,但峰值显著高于对照组。结果表明,大豆黄酮对去卵巢猪垂体ＬＨ分泌具有类雌激素效应,而其程度可能与大豆黄酮处理剂量和处理时间有关。     

改良大鼠子宫内膜异位症模型的建立及微血管密度观察

目的对皮下筋膜层与腹壁肌层之间移植自体子宫内膜制作的子宫内膜异位症（endometriosis,EMs）模型进行评估。方法取10只雌性未交配性成熟大鼠,术前雌激素诱导,手术开腹取右侧子宫,将自体子宫内膜种植于双侧皮下筋膜层与腹壁肌层之间,术后第29天取出异位组织,进行组织形态学观察,免疫组织化学染色观察微血管密度（Microvessel density,MVD）。结果异位内膜在腹壁内生长,呈隆起囊状小包块,内有黏液,具有正常子宫内膜基本组织结构。异位内膜中微血管密度较在位内膜和正常子宫内膜高。结论此手术方法建立的大鼠子宫内膜异位症模型异位内膜病理改变与EMs患者类似,可以作为子宫内膜异位研究的动物模型。     

PGLA编织支架的制备与初步体内评价

目的:热拉伸会改变纤维的结构和性能,进而影响由纤维编织而成的支架的性能。本文考察了PGLA纤维的拉伸倍数对编织支架在SD大鼠皮下的体内降解行为的影响。方法:制备了基于生物可降解高分子材料聚乙交酯丙交酯(PGLA,GA/LA摩尔比=90/10)的完全生物可降解编织支架,通过测试支架在大鼠体内降解过程中的失重、表面形貌、热性能、径向压缩力等变化情况,考察了纤维的不同的拉伸倍数对支架体内降解过程的影响。结果:用拉伸倍数为5的PGLA纤维编织的支架在植入SD大鼠皮下后降解最慢,重量、吸水率、结晶度、化学成分和径向压缩力的变化最慢,植入体内10天后能够保持完整的支架形态。结论:纤维的拉伸倍数会影响由纤维编织成的支架的热性能和力学性能的变化,本研究结果表明这种新的手工编织的支架具有短暂支撑管腔狭窄的潜在应用,为支架的材料选择和制备方法提供了参考,为在体内起到短暂支撑作用的支架的深入研究提供了实验基础。     

自组装短肽对角膜碱烧伤的作用

目的研究自组装短肽材料1%RADAl6-I水溶液对大鼠和兔角膜碱烧伤的作用.方法用1 mol/L NaOH溶液造SD大鼠和兔角膜碱烧伤模型,左眼每口滴加短肽水凝胶10μ1(2次/d),右眼为空白对照.在烧伤后的不同时间段(7、10、14d)处死大鼠,第7 d处死兔,取角膜行HE染色,光学显微镜观察结果.结果短肽水凝胶治疗组角膜上皮重生较快,皮下纤维排列较好,空白对照组皮下仍有较大空洞,皮下纤维排列紊乱,说明短肽材料能促进角膜恢复.     

大鼠子宫异位症模型建立方法的比较

目的将自体子宫内膜组织同时种植于大鼠体内三个不同部位,建立的子宫内膜异位症模型进行比较评估。方法取30只雌性未交配性成熟大鼠,将自体子宫内膜组织种植于左侧腹壁、子宫卵巢韧带和右侧皮下筋膜层与腹壁肌层间,术后第21天取出异位组织,进行组织形态学观察。结果异位内膜可以在大鼠不同部位同时生长,体积和重量均无显著性差异,其种植成功率差异不显著（93%-100%）,且种植物组织形态相似。结论此手术方法建立的大鼠子宫内膜异位症模型异位内膜病理改变与EMs患者类似,但皮下种植更简单,更直观,可减小损伤,便于反复实验及连续性观察研究。     

大鼠E12.5胰腺间质细胞原代培养及鉴定

目的:体外分离培养大鼠胚胎胰腺间质细胞(pancreas mesenchymal cells).方法:以孕12.5天(E12.5)的大鼠胚胎为组织来源,通过显微分离得到E12.5的胚胎胰腺,在鼠尾胶原包覆的六孔板内进行组织块培养,观察培养过程中胰芽的形态变化,间质细胞的长出,并用细胞免疫荧光的方法对间质细胞进行了鉴定.结果:显微分离得到了胚胎胰腺,为间质细胞群包绕着上皮细胞团的结构,培养第二天即发现间质细胞的长出,培养第四天时得到间质细胞群,免疫荧光鉴定为vimentin 阳性的间质细胞.结论:本方法通过大鼠E12.5胚胎胰腺显微分离.体外培养出原代胚胎胰腺间质细胞,为进一步研究大鼠胰腺间质细胞的功能提供实验平台.     

SFRP5抑制BMP9诱导人脐带间充质干细胞成骨分化的实验研究

目的:探讨脂肪因子分泌型卷曲相关蛋白(secreted frizzled-related protein 5,SFRP5)对骨形态发生蛋白9(bone morphogenetic protein 9,BMP9)诱导人脐带间充质干细胞(human umbilical cord-derived mesenchymal stem cells,hUC-MSCs)成骨分化的影响。方法:将人脐带间充质干细胞根据不同的处理因素分为4组:对照组、BMP9组、BMP9+SFRP5组和SFRP5组;分别在3天、5天和7天进行碱性磷酸酶(alkaline phosphatase,ALP)活性读数,7天进行ALP染色,21天进行茜素红染色检测钙盐沉积及油红O染色;收集不同组的细胞用于裸鼠皮下注射成骨模型的建立,4周后取出离体骨进行Micro-CT扫描和分析,获取的标本进行HE、Masson染色,Alcian blue染色及油红O染色检测。Western blot检测成骨分化相关蛋白Runx2和OPN的表达。结果:BMP9组的ALP活性读数和染色结果和茜素红染色结果均较对照组增加,而BMP9+SFRP5组则较BMP9组降低;BMP9处理后出现少量脂滴,而BMP9+SFRP5组脂滴明显增加,SFRP5组脂滴最多;裸鼠皮下注射成骨模型的观察结果显示,对照组和SFRP5组没有形成肉眼可见的皮下包块,BMP9组和BMP9+SFRP5组能生成异位骨;4周后观测大体标本以及进行MicroCT检测,发现BMP9+SFRP5组的骨密度值小于BMP9组(P0.05)。HE、Masson染色,Alcian blue染色结果显示,BMP9组的骨分化程度大于BMP9+SFRP5,油红O染色结果示BMP9+SFRP5组有较多的成脂分化;SFRP5能抑制BMP9诱导的Runx2、OPN的蛋白质表达。结论:SFRP5抑制BMP9诱导的人脐带间充质干细胞成骨分化。     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.