Characterization of Interstitial Cells of Cajal in Bowel of Cattle (Bos taurus)

Interstitial cells of Cajal (ICC) have been described in the gastrointestinal tract of different mammals including humans, horses, pigs, rats, dogs, mice and guinea-pigs. In the present study, ICC were identified in the jejunum of Bos taurus using polyclonal anti-c-Kit antibodies in immunohistochemical assays. Vimentin and desmin intermediate filaments were also determined using monoclonal antibodies. ICC were found in the tunica muscularis either in a palisade distribution pattern between the outer longitudinal and the inner circular layers (ICC-MP) or freely distributed in clusters in the longitudinal layer (ICC-LM). Morphometric studies determined that ICC have a fusiform shape presenting cytoplasmic prolongations. ICC were positive to c-Kit and vimentin antigens but negative to desmin. We have observed and described for the first time the presence of ICC in a ruminant. As observed in the aforementioned mammals, bovine ICC were associated with the myenteric plexus. Nevertheless, the presence of widespread ICC in the longitudinal muscular layer of the jejunum differs from previously described studies of other mammals.     

Immunomodulatory effects of β-glucans on porcine alveolar macrophages and bone marrow haematopoietic cell-derived dendritic cells

The immunopharmacological activities of β-glucans with a backbone of β-1,3/β-1,6-linkages associated with anti-tumor, anti-viral, bacterial and fungal infections have been well documented. Dectin-1, a specific pattern recognition receptor for β-1,3/β-1,6-glucans, is expressed mainly on phagocytes, especially macrophages and dendritic cells (DCs). In this study, the encoding nucleotide for the carbohydrate-recognition domain (CRD) of porcine dectin-1 was sequenced for the first time, and the immunomodulatory functions of a synthetic particulate β-glucan (p-β-glucan) were examined. Results showed that p-β-glucan significantly enhanced cell activity and phagocytosis in porcine alveolar macrophages (AMs), immature DCs (imDCs) and mature DCs (mDCs), in a similar way to zymosan. Zymosan enhanced dectin-1/TLR2/TLR4 expression and TNF-α/IL-10 production in all of three types of cell, whereas p-β-glucan increased dectin-1/TLR4 and TNF-α/IL-12 production in AMs but inhibited IL-10 in mDCs. These results indicate that the complex collaborating interactions between dectin-1 and TLRs in the recognition of β-1,3/β-1,6-glucans with different structural features may direct different cellular responses.     

piggyBac转座子介导的家蚕细胞转基因研究初探

为探讨稳定转化家蚕细胞表达外源基因的新方法,以家蚕核型多角体病毒(BmNPV)极早期蛋白基因(ie-1)启动子元件驱动新霉素抗性基因(neor),并克隆至具有绿色荧光蛋白基因(gfp)标记的昆虫转座子载体piggyBac中,构建转基因载体pigA3-IE-Neo。以该载体转染家蚕BmN细胞,用终浓度800μg/mL的遗传霉素(geneticin,G418)筛选3个月,获得了稳定转化的细胞,呈现绿色荧光的细胞数达80%以上。通过PCR鉴定证实了细胞基因组DNA中neor基因和gfp基因的存在。     

哺乳动物成体干细胞的研究进展

干细胞具有自我更新能力,能够产生高度分化的功能细胞。干细胞按照发育阶段分为胚胎干细胞和成体干细胞。根据干细胞的发育潜能分为三类：全能干细胞、多能干细胞和单能干细胞。胚胎干细胞的发育等级较高,是全能干细胞,而成体干细胞的发育等级较低,是多能或单能干细胞。一般认为成年组织或器官内的干细胞具有组织特异性,只能分化成特定的细胞或组织,然而,研究结果表明,组织特异性干细胞同样具有分化成其他细胞或组织的潜能,这为干细胞的应用开创了更广泛的空间。作者主要综述了干细胞的研究进展及几种成体干细胞的诱导分化。     

用免疫荧光技术检测细胞培养物中兔出血症病毒

用本实验室制备的兔出血症病毒（rabbit hemorrhagic disease virus,RHDV）高免血清,按常规方法提纯出抗体（IgG）,用异硫氰酸荧光素（FITC）标记IgG。在细胞培养时,培养瓶中放入玻片,当细胞长成单层时,按常规方法接种病毒液,培养24、48、96、120h时取出玻片,用荧光抗体染色,在荧光显微镜下观察不同代次的细胞毒。经观察：兔肾上皮细胞（RK）毒培养到36—48h,羊睾丸细胞（ST）毒培养到72—96h时可观察到特异性荧光,随着培养时间的延长,荧光亮度增强,胞浆内充满特异性荧光。用肝组织强毒病料触片,呈特异性荧光,对照细胞培养48h无荧光出现,证实了两株细胞培养物中有大量的兔出血症病毒存在,从而成功的分离培养出了兔出血症病毒细胞毒。     

嗅鞘细胞对无血清培养诱导PC12细胞凋亡的作用

采用无血清培养诱导PC12细胞凋亡,研究嗅鞘细胞(Olfactory ensheathing cells,OECs)对去血清后诱导PC12细胞凋亡中的作用。通过MTT法检测细胞的活性,细胞形态学观察,以及结合流式细胞仪检测细胞的凋亡率,井对不同组细胞凋亡率进行比较。结果表明：①OECs培养上清对去血清后PC12细胞的存活有明显的促进作用;②从形态学上观察,OECs对去血清诱导PC12细胞凋亡有明显的抑制作用,使得细胞向死亡过渡受阻;②流式细胞对细胞周期分析,去血清诱导后的PC12细胞,大部分细胞滞留在G1期,细胞向S期过渡受阻,造成G1期细胞的堆积,它们所占比例分别为：73．6％,25．9％,0．5％;而OECs联合培养组所占比例：53％,42．3％,4．7％;OECs上清组：42．1％,52．2％,5．7％;正常有血清培养组：45,6％,41．4％,13％。凋亡细胞所占百分比分别为：60．3％,45,6％,37．4％,0．5％。     

利用乳汁体细胞研究共轭亚油酸钙对奶牛乳腺基因表达的作用

8头泌乳中期的奶牛被随机分为2组,分别是对照组（每头牛：基础日粮＋棕榈酸钙200g／d）和试验组（每头牛：基础日粮＋共轭亚油酸钙200g／d）,试验期14d,检测了奶产量、乳成分,分析奶牛的血液变化,并采用荧光定量PCR对乳汁体细胞中脂蛋白脂酶（LPL）、乙酰辅酶A羧化酶（ACACA）的基因表达进行检测。结果显示,与对照组相比,试验组奶牛的乳产量、乳蛋白、乳糖和乳汁体细胞含量没有显著影响,但是显著降低了乳脂肪含量（P〈0．05）,对照组和试验组奶牛乳脂肪含量分别为0．0326g／mL和0．0244g／mL。检测血液指标发现,共轭亚油酸钙显著升高了奶牛血液中高密度脂蛋白的含量（P〈0．05）。基因分析发现,试验组奶牛乳汁体细胞中LPL、ACA—CA的基因表达显著下调,表明共轭亚油酸钙抑制了奶牛乳腺细胞脂肪酸合成酶的基因表达。     

Comparing the osteogenic potential of canine mesenchymal stem cells derived from adipose tissues, bone marrow, umbilical cord blood, and Wharton's jelly for treating bone defects

Alternative sources of mesenchymal stem cells (MSCs) for replacing bone marrow (BM) have been extensively investigated in the field of bone tissue engineering. The purpose of this study was to compare the osteogenic potential of canine MSCs derived from adipose tissue (AT), BM, umbilical cord blood (UCB), and Wharton''s jelly (WJ) using in vitro culture techniques and in vivo orthotopic implantation assays. After canine MSCs were isolated from various tissues, the proliferation and osteogenic potential along with vascular endothelial growth factor (VEGF) production were measured and compared in vitro. For the in vivo assay, MSCs derived from each type of tissue were mixed with β-tricalcium phosphate and implanted into segmental bone defects in dogs. Among the different types of MSCs, AT-MSCs had a higher proliferation potential and BM-MSCs produced the most VEGF. AT-MSCs and UCB-MSCs showed greater in vitro osteogenic potential compared to the other cells. Radiographic and histological analyses showed that all tested MSCs had similar osteogenic capacities, and the level of new bone formation was much higher with implants containing MSCs than cell-free implants. These results indicate that AT-MSCs, UCB-MSCs, and WJ-MSCs can potentially be used in place of BM-MSCs for clinical bone engineering procedures.     

假蒟提取物的体外抗氧化和抗炎效果研究

本研究旨在分析假蒟提取物的活性成分并评价其体外抗氧化和抗炎活性。试验通过ABTS法检测3种假蒟提取物(石油醚、正丁醇及乙酸乙酯提取物)的抗氧化能力;采用LPS法建立猪小肠上皮细胞(IPEC-J2)体外炎症模型,用假蒟提取物预处理后,以ELISA法检测细胞上清液中肿瘤坏死因子(TNF-α)、白细胞介素-1(IL-1)和白细胞介素-6(IL-6)的含量。结果表明,3种假蒟提取物总抗氧化活性由大到小依次是假蒟正丁醇提取物、乙酸乙酯提取物、石油醚提取物(P<0.05),其中假蒟正丁醇提取物总生物碱、总酚含量最高,分别为2.81 mEq/100 g、161.82 mg/g。与空白组相比,LPS组IPEC-J2细胞的TNF-α(P<0.05)、IL-1(P>0.05)及IL-6(P<0.05)含量均升高;与LPS组相比,假蒟预处理组的这几种细胞因子含量均显著降低(P<0.05),由高到低依次为乙酸乙酯提取物>石油醚提取物>正丁醇提取物。综合试验结果,假蒟提取物具有一定的抗氧化作用,能影响IPEC-J2细胞炎性因子的分泌,抑制炎症反应。