狗舌草提取物诱导L1210细胞的凋亡

以环磷酰胺、槲皮素和单猪屎豆碱为参照，观察狗舌草60％乙醇提取物体外对L1210细胞的凋亡形态变化的影响；利用流式细胞术Annexin V-FITC／PI双参数法，研究狗舌草60％乙醇提取物对L1210细胞早期凋亡的影响，探讨狗舌草60％乙醇提取物致L1210细胞凋亡的机理。结果发现，狗舌草60％乙醇提取物能使L1210细胞发生典型的细胞凋亡，形成凋亡小体；狗舌草60％乙醇提取物引起的Annexin V-FITC^＋／PI^-细胞百分比为86．8％，能诱导L1210细胞发生早期凋亡。     

狗舌草抗淋巴性白血病有效部位的急性毒性

利用体重 19～ 2 2 g的 BAL B/ c-C雌性小鼠 ,通过腹腔注射 ,从急性半数致死量 L D50 和供试鼠的临床表现等方面 ,检查了狗舌草 60 %乙醇提取物冻干粉的急性毒性。结果发现 ,狗舌草 60 %乙醇提取物对雌性 BAL B/ c-C小鼠腹腔注射的 LD50 为 ( 791.2 2± 170 .17) mg/ kg,95 %可信限为 63 9.15～ 979.49m g/ kg。急性死亡小鼠 (剂量达到 3 3 6.11mg/ kg以上 )在死前表现四肢抽搐 ,盲目运动 ,呼吸急促 ,翘尾 ;未死亡的小鼠表现食欲减退 ,反应迟钝 ,在随后的长期饲养中 ,临床表现逐步恢复正常     

狗舌草中PAs对大鼠毒性的研究

将从早花期和盛花期狗舌草中提取得到的双稠比咯啶生物碱（ＰＡｓ）,按１００ｍｇ／ｋｇ分别给两组ＳＤ大鼠腹腔注射。结果表明,两个时期狗舌草中ＰＡｓ对ＳＤ大鼠均具有一定的肝毒性、肺毒性和中枢神经毒性,但盛花期狗舌草中ＰＡｓ的毒性较弱;新发现狗舌草对心、肾也具有一定的毒性作用。     

鸡实验性淋巴细胞性白血病的病理学研究

给３５只１日龄伊莎褐蛋母鸡雏腹腔接种淋巴细胞性白血病病毒ＲＡＶ－１株，应用常规病理技术，对接毒后第１５天、１、２、３、４、５、６个月７个批次的实验鸡做了病理学研究。结果：接毒后１５ｄ和１个月，部分实验鸡发生了成髓细胞性白血病，主要表现为骨髓成髓细胞大量增生，或形成成髓细胞性肿瘤结节，肝、心、肾、法氏囊等内脏器官出现成髓细胞聚集；接毒后２～６个月，实验鸡发生了淋巴细胞性白血病，主要表现为法氏囊髓质淋巴细胞发生转化，成淋巴细胞克隆增殖形成成淋巴细胞克隆增殖灶，在肝、心、肾、脾、腺胃等器官中形成成淋巴细胞性肿瘤结节。据此，可对鸡淋巴细胞性白血病做出病理组织学诊断。     

葎草乙醇提取物对小鼠十二指肠黏膜免疫相关细胞数量的影响

为了比较不同剂量葎草乙醇提取物对小鼠十二指肠黏膜上皮内淋巴细胞(IEL)、杯状细胞(GC)及肥大细胞(MC)数量的影响,采用32只健康昆明系小鼠随机分为对照组(生理盐水)、葎草乙醇提取物试验组低(100 mg/kg体重)、中(200 mg/kg体重)、高(300 mg/kg体重)剂量组,每组8只,按体重灌胃1次/d,连续30 d,取十二指肠做石蜡切片,分别经H.E.、PAS染色和MTB染色,显微计数IEL、GC和MC的数量。结果葎草乙醇提取物中剂量组(200 mg/kg体重)的IEL、GC和MC的数量极显著高于对照组(P<0.01);中剂量组的IEL和MC与低、高剂量组比较差异不显著(P>0.05),中剂量组的GC与高剂量组比较差异极显著(P<0.01),与低剂量组比较差异不显著(P>0.05),但中剂量组细胞数量提高均优于低、高剂量组。表明葎草乙醇提取物能增加肠道黏膜免疫相关细胞的数量,保护肠道黏膜的完整性与稳定性,提高机体免疫力,中剂量(200 mg/kg体重)效果最佳。     

诱导鸡胚精原干细胞向成骨细胞分化的研究

为了探讨体外培养的鸡胚精原干细胞(SSCs)的多向分化潜能,取孵化18~20 d的鸡胚睾丸,无菌获取SSCs,体外培养传代,通过地塞米松、β-甘油磷酸钠、维生素C诱导鸡胚SSCs向成骨细胞分化,钙结节Von Kossa s法、改良的钙钴法及免疫组化法进行成骨细胞鉴定。结果显示SSCs被诱导15~21 d后分化为成骨细胞,诱导形成率为75%~80%;诱导后的细胞Von Kossa s染色可见细胞间布满黑色颗粒,提示有矿化基质沉积;改良钙钴法碱性磷酸酶染色胞浆呈深棕色或深黑色,免疫组化法染色细胞呈阳性。因此可以得出在不同的诱导条件下,SSCs体外可被定向诱导分化为成骨细胞,证明其具有多向分化潜能。     

In vitro neuronal and osteogenic differentiation of mesenchymal stem cells from human umbilical cord blood

Mesenchymal stem cells (MSCs) have the capabilities for self-renewal and differentiation into cells with the phenotypes of bone, cartilage, neurons and fat cells. These features of MSCs have attracted the attention of investigators for using MSCs for cell-based therapies to treat several human diseases. Because bone marrow-derived cells, which are a main source of MSCs, are not always acceptable due to a significant drop in their cell number and proliferative/differentiation capacity with age, human umbilical cord blood (UCB) cells are good substitutes for BMCs due to the immaturity of newborn cells. Although the isolation of hematopoietic stem cells from UCB has been well established, the isolation and characterization of MSCs from UCB still need to be established and evaluated. In this study, we isolated and characterized MSCs. UCB-derived mononuclear cells, which gave rise to adherent cells, exhibited either an osteoclast or a mesenchymal-like phenotype. The attached cells with mesenchymal phenotypes displayed fibroblast-like morphologies, and they expressed mesenchym-related antigens (SH2 and vimentin) and periodic acid Schiff activity. Also, UCB-derived MSCs were able to transdifferentiate into bone and 2 types of neuronal cells, in vitro. Therefore, it is suggested that the MSCs from UCB might be a good alternative to bone marrow cells for transplantation or cell therapy.     

体外诱导牛BMSC向上皮样细胞分化的研究

为培育转基因肉牛提供种子细胞以及进一步丰富牛骨髓间充质干细胞(bone marrow mesenchymal stem cell,BMSC)的多向分化潜能,利用细胞免疫荧光染色和分子生物学方法,初步探讨表皮生长因子和胰岛素体外诱导牛BMSC向上皮样细胞分化的可能性。利用含细胞因子的诱导液对纯化稳定的P4代牛BMSC进行体外诱导,并对诱导后的细胞进行细胞角蛋白18的细胞免疫荧光观察和细胞角蛋白19的RT-PCR鉴定。结果表明,诱导后细胞经细胞角蛋白18免疫荧光染色后出现明显的荧光。RT-PCR结果显示诱导分化后细胞角蛋白19基因在细胞中表达。因此,在体外,表皮生长因子和胰岛素可诱导牛BMSC初步分化为上皮样细胞。     

The use of cytochemistry,immunophenotyping, flow cytometry,and in vitro differentiation to determine the ontogeny of a canine monoblastic leukemia

We evaluated the utility of cytochemistry, immunophenotyping, flow cytometry, and in vitro culture with forced differentiation of leukemic cells as diagnostic aids to identify the malignant cell ontogeny in a dog with leukemia. A tentative diagnosis of monoblastic leukemia was established by microscopic examination of Romanowsky-stained blood smears and bone marrow aspirate smears. This diagnosis also was supported by the light scatter signature that identified the blast cells as large, non-granular monocytic cells using a CellDyn 3500 automated hematology analyzer; as well as by the detection of N-butyrate esterase and the lack of choloroacetate esterase or leukocyte peroxidase by cytochemical staining. Subsequently, leukemic cells were isolated from the dog's peripheral blood and placed into tissue culture or cryopreserved. The leukemic cells grew in suspension cultures and proliferated spontaneously for up to 4 days. By day 7, proliferation was negligible. Upon culture with conditioned supernatant using mitogen-stimulated human T cells as a source of cytokines, an increased proportion of cells entered S phase by day 2 of culture; however, proliferation declined markedly by day 4, at which time the cells had apparently differentiated to adherent, vacuolated macrophages. The cytokine-stimulated leukemic cells were positive for the monocyte/macrophage specific markers alpha-1-antitrypsin, alpha-1-antichymotrypsin, lysozyme, CD14, MHC class II, and calprotectin, an antigen found in differentiated macrophages and granulocytes. Despite the strong tendency of the leukemic cells towards monocytic differentiation, our results suggested that they retained some features of a myelomonocytic precursor. These data show that cytochemistry, immunophenotyping, flow cytometry, and in vitro differentiation of canine leukemia cells are useful tools for confirming the lineage of malignant hematopoietic cells.     

UBE2L3对牛骨骼肌卫星细胞增殖分化的影响

将分离的牛骨骼肌卫星细胞(BSMSCs)进行体外培养,首先检测泛素结合酶UBE2L3在BSMSCs增殖分化过程中mRNA以及蛋白表达水平的变化.设计UBE2L3的3个干扰RNA(si-UBE2 L3-1、si-UBE2L3-2、si-UBE2L3-3),对干扰效果进行筛选.构建UBE2L3过表达质粒载体pcDNA3.1...     

妊娠中期猪羊水来源干细胞EGFP基因转化及体外分化

本研究旨在获得妊娠中期猪羊水来源千细胞,并通过用EGFP对干细胞进行标记,为以EGFP作为示踪标记对干细胞进行体内移植研究奠定基础.利用羊水来源干细胞培养技术体系,从胎龄60 d猪胎儿羊水中分离获得干细胞,通过脂质体介导转染将EGFP基因导入干细胞,诱导转基因干细胞向肌细胞和神经细胞分化,观察其分化特点.采用RT-PCR技术检测干细胞和分化细胞表面标志或相关基因.结果成功分离培养出妊娠中期猪羊水来源干细胞,并获得转EGFP基因干细胞.干细胞在表达EGFP的同时仍具有分化潜能.干细胞中Oct4、CD-90和Sox2表达阳性;体外诱导的干细胞能分化为肌细胞(表达myf-6和myoD)、星形胶质细胞(表达GFAP)、少突胶质细胞(表达GalC)和神经元细胞(表达NF、NSE和MAP2).研究表明,从妊娠中期猪胎儿羊水中分离干细胞具有可行性和有效性,转EGFP基因干细胞具有自我更新、增殖和分化潜能,可以用EGFP对羊水来源干细胞进行标记、追踪,为EGFP作为示踪标记对干细胞用于体内移植研究奠定了基础.     

口蹄疫病毒诱导BHK-21细胞产生凋亡的研究

试验利用口蹄疫病毒感染BHK-21细胞,通过MTT法、Hoechst 33258染色、原位末端标记技术(TUNEL)、流式细胞术和链特异性荧光定量RT-PCR,分别就口蹄疫病毒对BHK-21细胞生长的抑制作用、凋亡细胞的形态学和分子生物学特征、凋亡峰的出现和细胞周期的变化以及口蹄疫病毒基因组在BHK-21细胞内的复制情况进行了检测。结果表明:口蹄疫病毒可抑制BHK-21细胞的生长并诱导其产生凋亡,呈现典型的凋亡细胞特征,出现细胞凋亡峰并且细胞周期明显被阻滞在G1/G0期,同时对凋亡率和口蹄疫病毒基因组复制的关系做了初步研究。     

补骨脂种子提取物对苹果腐烂病菌的抑菌活性研究

旨在探究补骨脂种子提取物对苹果腐烂病菌的杀菌活性及田间防效。采用菌丝生长和孢子萌发抑制率法对补骨脂种子提取物的抑菌活性进行测定,得到的EC50值分别为6.766mg/L、0.172 mg/L,说明补骨脂种子提取物对苹果腐烂病菌菌丝生长和孢子萌发均具有明显的抑制作用。田间试验表明,补骨脂种子提取物微乳剂稀释600倍液～200倍液后,对苹果腐烂病菌引起的病斑进行刮疤涂抹处理,防治效果可达71%～81%左右。经田间系统观察,供试药剂在试验剂量范围内对苹果树生长发育无不良影响,使用安全。