小鼠去细胞拟胚体对小鼠Lewis肺癌细胞体内生长的影响

目的:阐明小鼠去细胞拟胚体对小鼠Lewis肺癌细胞在体内生长的影响。方法:先制备来源于小鼠胚胎干细胞的拟胚体,然后用SDS去细胞处理。实验分成3组:小鼠Lewis肺癌细胞与去细胞拟胚体培养组,癌细胞与Matrigel培养组和单纯癌细胞组(每组n=12)。培养3天后注射入裸鼠体内,观察肿瘤生长情况。28天取出瘤体,Ki67和CD31免疫组化染色(n=12)检测细胞增殖和肿瘤微血管密度(MVD),Western blot检测组织Paxillin,E-cadherin和β-actin水平(n=6)。结果:去细胞拟胚体组肿瘤生长明显较单纯细胞组和Matrigel组慢。去细胞拟胚体组Ki67指数((17.1±2.6)%)明显小于单纯细胞组((34.5±4.7)%)和Matrigel组((48.4±8.6)%)(P0.05);去细胞拟胚体组的MVD(18.7±3.6个/mm2)明显小于单纯细胞组(32.1±6.4个/mm2)和Matrigel组(42.6±7.1个/mm2)(P0.05)。Western blot结果提示去细胞拟胚体组的Paxillin表达小于单纯细胞组和Matrigel组(P0.05),而E-cadherin表达大于单纯细胞组和Matrigel组(P0.05)。结论:小鼠去细胞拟胚体对小鼠Lewis肺癌细胞在体内有明显的促分化作用。     

Differentiation of embryonic stem cells into inner ear vestibular hair cells using vestibular cell derived-conditioned medium

Vestibular hair cells (V–HCs) in the inner ear have important roles and various functions. When V–HCs are damaged, crippling symptoms, such as vertigo, visual field oscillation, and imbalance, are often seen. Recently, several studies have reported differentiation of embryonic stem (ES) cells, as pluripotent stem cells, to HCs, though a method for producing V–HCs has yet to be established. In the present study, we used vestibular cell conditioned medium (V-CM) and effectively induced ES cells to differentiate into V–HCs. Expressions of V-HC-related markers (Math1, Myosin6, Brn3c, Dnah5) were significantly increased in ES cells cultured in V-CM for 2 weeks, while those were not observed in ES cells cultured without V-CM. On the other hand, the cochlear HC-related marker Lmod3 was either not detected or detected only faintly in those cells when cultured in V-CM. Our results demonstrate that V-CM has an ability to specifically induce differentiation of ES cells into V–HCs.     

Impact of transient stress and stress enzymes on development

Stress enzymes triggered by transient stress mediate reprioritization of developmental and homeostatic events to flexibly accomplish the next essential developmental event. This review analyzes recent studies on stress and stress enzyme function during early mammalian development and describes the diverse consequences that result from measurement, analysis of function, and management of stress and stress enzymes during development.     

Apoptosis regulates notochord development in Xenopus

The notochord is the defining characteristic of the chordate embryo and plays critical roles as a signaling center and as the primitive skeleton. In this study we show that early notochord development in Xenopus embryos is regulated by apoptosis. We find apoptotic cells in the notochord beginning at the neural groove stage and increasing in number as the embryo develops. These dying cells are distributed in an anterior to posterior pattern that is correlated with notochord extension through vacuolization. In axial mesoderm explants, inhibition of this apoptosis causes the length of the notochord to approximately double compared to controls. In embryos, however, inhibition of apoptosis decreases the length of the notochord and it is severely kinked. This kinking also spreads from the anterior with developmental stage such that, by the tadpole stage, the notochord lacks any recognizable structure, although notochord markers are expressed in a normal temporal pattern. Extension of the somites and neural plate mirrors that of the notochord in these embryos, and the somites are severely disorganized. These data indicate that apoptosis is required for normal notochord development during the formation of the anterior-posterior axis, and its role in this process is discussed.     

A mutation of keratin 18 within the coil 1A consensus motif causes widespread keratin aggregation but cell type-restricted lethality in mice

Mutations in genes encoding epidermal keratins cause skin disorders, while those in internal epithelial keratins, such as K8 and K18, are risk factors for liver diseases. The effect of dominant mutations in K8 or K18 during embryonic development and tissue homeostasis has not been examined so far. Here we demonstrate that the dominant mutation hK18 R89C, that is highly similar to hK14 R125C, causing EBS in humans, leads to cell type-specific lethality in mice, depending on the ratio of mutant to endogenous keratins. Mice expressing hK18 R89C in the absence of endogenous K19 and K18 died at mid-gestation from defects in trophoblast giant cells, accompanied by haematomas. A single, endogenous K18 allele rescued embryonic lethality but caused aggregation of keratins in all adult internal epithelia, surprisingly without spontaneous cell fragility. Closer analysis revealed that both filaments and aggregates coexisted in the same cell, depending on the ratio of mutant to endogenous keratins. Our results demonstrate that balanced overexpression of a wild-type keratin rescued the lethal consequences of a dominant-negative mutation. This has important implications for therapy approaches of keratinopathies, suggesting that suppressing the mutant allele is not necessary in vivo.     

The context of embryonic development and its ethical relevance

Research on human stem cells and embryos creates ethical issues. Here I discuss ten frequently used arguments against research and point out their weaknesses. These arguments include the possessed potentiality of the embryo per se and, in contrast to other cell systems, the "slippery slope" argument, the right of disposal of parents, totipotency versus pluripotency, the burden of proof for research, natural versus artificial, and three arguments based on the precaution principle (the open biological questions, uncertainty regarding clinically applicable therapies, and the problem solving rule). I finally suggest a different answer to the ethical questions concerning research on human embryos and embryonic stem cells, which takes into consideration their biological context.     

根癌农杆菌介导的高效大豆遗传转化体系的建立

利用根癌农杆菌对来自大豆成熟种子的胚尖进行遗传转化,研究了影响农杆菌介导大豆转化的各种因素,建立了一套优化的大豆遗传转化体系。研究结果表明:菌株KYRT1比EHA105和LBA4404具有更强的侵染能力;较酸的共培养基(pH5.4)、较低的培养温度(22℃)均有利于提高转化效率;恢复培养和分步抗性筛选方式有利于提高抗性组织的存活率和分化率。同时应用这种优化的遗传转化体系,获得了7个大豆品系的转基因植株,转化频率为4.29%-18%。经过PCR和Southern分析证明外源的双价抗虫基因cryIA(c)和pta已经整合到大豆的基因组中。