System for inducible expression of cre-recombinase from the Foxa2 locus in endoderm, notochord, and floor plate.

We targeted the reverse tetracycline controlled transactivator (rtTA) to the Foxa2 locus (Foxa2(ITA)) to generate a system for regulating Cre-recombinase activity within Foxa2 expression domains, including the endoderm, notochord, and floor plate of early mouse embryos. The use of an internal ribosomal entry site to obtain rtTA expression preserves Foxa2 function of the targeted allele. Cre activity with this system reflects the level of endogenous Foxa2 activity and is also tightly controlled by doxycycline. The location of Cre activity within the broader Foxa2 expression domain can be restricted by altering the timing of doxycycline administration. Isolated floor plate expression can be obtained in this manner. This system will provide a useful tool for manipulating gene expression in endoderm, notochord, and floor plate, all of which are tissues with important structural and patterning functions during embryogenesis.     

树突状细胞在黏膜免疫模型小鼠中的分布及E-cadherin和ICAM-1表达的研究

目的通过研究小鼠树突状细胞在黏膜免疫模型小鼠中分布特征及上皮型钙黏附分子（E-cadherin）和细胞间黏附分子-1（ICAM-1）的表达,以探讨E-cadherin和ICAM-1在树突状细胞迁移过程中的调控作用.方法采用光镜和免疫组织化学染色方法,观察黏膜免疫模型小鼠中树突状细胞的分布特征;分析E-cadherin和ICAM-1的表达情况.结果体内的树突状细胞在黏膜免疫模型小鼠中,主要分布于肠系膜淋巴结、回肠集合淋巴小结、胃和空回肠黏膜及黏膜下层,与对照组相比有显著差异,其ICAM-1表达明显增高,E-cadherin表达下调,分别与对照组数密度和面密度比较有显著差异.结论在树突状细胞迁移运动过程中,E-cadherin和ICAM-1黏附分子可能起着关键性的调控作用.     

妊娠期间脾脏T淋巴细胞的改变与鼠胚着床

应用ＦＩＴＣ标记的Ｌ３Ｔ４及Ｌｙ２单克隆抗体，及流式细胞计细胞分选，对小鼠妊娠期间脾脏Ｔ淋巴细胞亚群进行检测。结果表明，妊娠早期小鼠脾脏ＣＤ８＋淋巴细胞比例减少，在着床时尤为明显。提示脾脏Ｔ淋巴细胞数量和功能变化，参与维持正常妊娠，尤其是参与着床。     

神经元性一氧化氮合酶活性在app/ps1双转基因AD模型小鼠皮质和海马的分布

本实验用 4～ 5月龄和 15～ 16月龄的 app/ ps1dtg小鼠各 5只 ,同龄野生型小鼠每个年龄组各 5只 ,用β-NADPH组织化学染色显示神经元一氧化氮合酶活性 ,刚果红组织学染色显示神经炎性斑 ,无偏性体视学计量皮质和海马神经炎斑的总体积 ,研究神经元一氧化氮合酶活性在 app/ ps1双转基因 (app/ ps1dtg) AD模型小鼠大脑皮质和海马的异常分布 ,探讨其在该模型小鼠及 AD患者脑内病理改变中的作用。结果显示 ,n NOS阳性神经元在各组小鼠皮质和海马内的分布没有区别 ,4～ 5月龄 app/ps1dtg小鼠皮质和海马可见神经炎斑 (NP)和营养不良性 NOS阳性神经突 (DTN) ;15～ 16月龄 app/ ps1dtg小鼠皮质和海马内NP的体积分别是 4～ 5月龄 app/ ps1dtg小鼠皮质 NP的 5倍 (P<0 .0 0 5 )和 8倍 (P<0 .0 0 1) ;15～ 16月龄 app/ ps1dtg小鼠皮质内 DTN的体积明显增大 (P<0 .0 1) ;且伴有 NOS阳性神经元形态的改变及海马 CA1 区 NOS阳性神经元数量的减少 (P<0 .0 5 ) ,DTN的形成与 NP呈正相关关系 (r=0 .85 ,P<0 .0 5 )。本实验结果表明 ,app/ ps1dtg小鼠能够模拟 AD患者脑内 NOS阳性神经元的病理改变 ,DTN的形成可能与 Aβ的毒性作用有关 ,DTN产生的 NO可能参与 app/ ps1dtg小鼠和 AD的神经病理过程。     

Tissue-specific tumor suppressor activity of retinoblastoma gene homologs p107 and p130

The retinoblastoma gene family consists of three genes: RB, p107, and p130. While loss of pRB causes retinoblastoma in humans and pituitary gland tumors in mice, tumorigenesis in other tissues may be suppressed by p107 and p130. To test this hypothesis, we have generated chimeric mice from embryonic stem cells carrying compound loss-of-function mutations in the Rb gene family. We found that Rb/p107- and Rb/p130-deficient mice were highly cancer prone. We conclude that in a variety of tissues tumor development by loss of pRB is suppressed by its homologs p107 and p130. The redundancy of the retinoblastoma proteins in vivo is reflected by the behavior of Rb-family-defective mouse embryonic fibroblasts in vitro.     

Immature B cells from human and mouse bone marrow can change their surface light chain expression

The capacity of bone marrow-derived surface immunoglobulin-positive (sIg⁺) human and mouse immature B cells, generated either in vitro or in vivo, to change their light (L) chain expression, has been assayed by the number of cells which change in vitro from one type of L chain to the other type, or to no sIg at all. Immature sIg⁺ B cells were generated in vitro from sIg^? precursor cells from human or mouse bone marrow. The immature sIg⁺ cells expressed RAG-1. Human sIg⁺ cells expressed xfr; and λ L chains in ratios between 1:1 and 3:1, whereas in mouse cells, this ratio ranged from 10:1 to 20:1. Upon reculture of the human and mouse xfr;⁺sIg⁺ cells, about half of them remained xfr;⁺, a quarter became λ⁺, and another quarter became sIg^?. Between 1 and 3% expressed both xfr; and λ chains. Of the human λ⁺ cells, about two-thirds remained λ⁺, only 1 to 2% became xfr;⁺, while the other third became sIg^?. Again, between 1 and 3% expressed both xfr; and λ L chains. These results indicate that expression of sIgM in the B cell membrane does not terminate L chain gene rearrangement, and that some order exists in xfr; versus λ gene rearrangements. Hence, human and mouse xfr;⁺ immature B cells can become λ⁺, but very few of the λ⁺ cells can become xfr;⁺, and both can become sIg^?. Further, human CD10⁺/sIg⁺ xfr;⁺ and λ⁺ cells and mouse B220^low/sIg^low xfr;⁺ cells enriched from bone marrow, i.e. immature B cells differentiated in vivo, changed their Ig phenotype upon in vitro culture, but in lower frequencies. By contrast, human and mouse mature B cells did not change their L chain or Ig phenotype. Hence, at least a part of the sIg⁺ immature B cells in bone marrow retain the capacity to change their L chain and Ig phenotype, and this capacity is lost when they become mature, peripheral B cells.     

Intracellular Ca2+ concentrations are not elevated in resting cultured muscle from Duchenne (DMD) patients and in MDX mouse muscle fibres

The free intracellular calcium concentration, [Ca²⁺]_i, was studied in single myotubes using the fluorescent Ca²⁺ indicator fura-2. Myotubes cultured from satellite cells of small muscle specimens from Duchenne muscular dystrophy (DMD) patients were compared with human control myotubes and with myotubes cultured from MDX and control mouse muscle satellite cells. The resting [Ca²⁺]_i levels in DMD and control myotubes were not significantly different, i. e. 104 ±26 nM (mean ± SD, n=190 cells from eight DMD patients) compared with 97±25 nM (175/seven controls) and were not significantly lower than the corresponding murine values (154±33 nM, n=135 MDX myotubes; 159±34 nM, n=135 controls). All myotubes reacted to 10 M acetylcholine or 40 mM KCl with fast transient increases of [Ca²⁺]_i. After application of a hyposmotic (130 mOsm) solution, [Ca²⁺]_i was increased 1.5- to 3-fold within 2–3 min, the DMD myotubes tending to stronger reactions (significantly higher [Ca²⁺]_i in 2 out of 6 cases). The response was usually transient, [Ca²⁺]_i decreasing to the initial level within 10 min. Gadolinium (50 M) reduced the response by 50%–70%, indicating that the osmotic shock increased Ca²⁺ influx. During exposure to high (15 mM) [Ca²⁺]e, [Ca²⁺]_i of DMD and control cells was 1.5- to 2-fold higher. Adult muscle fibres from MDX mice and controls showed identical Ca²⁺ resting levels (n=45 fibres from three mice in each case), but did not respond to decreased external osmolarity with a change in [Ca²⁺]_i. The results indicate that lack of dystrophin in muscle fibres does not necessarily lead to increased [Ca²⁺]_i. It is suggested that increased [Ca²⁺]_i is probably a secondary consequence of fibre damage.     

Early embryonic expression patterns of the mouse Flamingo and Prickle orthologues.

The Drosophila melanogaster proteins Flamingo and Prickle act in the planar cell polarity (PCP) pathway, which is required for acquisition of epithelial polarity in the wing, eye, and epidermis. In mammals, PCP signaling has been shown to regulate cell movements and polarity in a variety of tissues. Here, we show that the murine Flamingo orthologues Celsr1-3 and the Prickle orthologues Prickle1, Prickle2, and Testin have dynamic patterns of expression during pregastrulation and gastrulation stages. Celsr1 is expressed in the anterior visceral endoderm and nascent mesoderm, Celsr2 and Celsr3 mark the prospective neuroectoderm, Prickle1 is expressed in the primitive streak and mesoderm, Prickle2 in the node, and Testin in the anterior visceral endoderm, the extraembryonic ectoderm, primitive streak, and mesoderm. Analysis of a gene-trap mutation in Testin indicates that this gene is not required for embryogenesis; therefore, other Prickle homologues may compensate for its function during development.     

实验性糖尿病对小鼠肝脏酶组织化学和超微结构的影响

实验用四氧嘧啶建造糖尿病动物模型，８周后取肝作组织学、酶组织化学和超微结构的观察，以探讨实验性糖尿病对肝组织结构和功能的影响。表明在糖尿病状态下，肝细胞的结构和功能均受到一定的损害。     

HLA-DR and H-2E transgenes differentially mediate TCR-specific positive selection

The use of HLA transgenic mice in models of immunity and diseaseassumes that human MHC molecules are able to contribute towardthe positive selection of the mouse TCR repertoire. As an initialstep towards analysis of this we have compared the relativeability of DR/Eß or E/Eß complexes to induceT cell receptor (TCR) positive selection in H-2Ea and HLA-DRAtransgenic mice lacking endogenous E. The results show that,like E/Eß, the hybrid DR/ß complexes arecapable of mediating positive selection of V_ß2⁺;,V_ß6⁺, and V_ß10⁺ cells. However, differenceswere found between the effects of the two transgenes. Thus,while V_ß6⁺ cells were efficiently selected in bothH-2Ea and DRA transgenic mice, positive selection of V_ß10⁺cells was less apparent in the DRA transgenic mice. Variationbetween Ea and DRA transgenic mice is consistent with the notionthat this process is dependent on differential binding of endogenouspeptides to the E/Eß and DR/Eß complexes.Furthermore, contrary to expectations, in neither set of micewas positive selection limited solely to the CD4⁺ subset. Thus,examples were found in which V_ß-specific positiveselection was confined to either the CD4⁺ or CD8⁺ subsets, andothers in which both subpopulations were concomltantly increased.In the case of V_ß2 positive selection, H-2Ea transgenicmice showed expansion of these cells in both the CD4⁺ and CD8⁺subpopulations whlle in DRA transgenic mice this occurred predominantlyin the CD8⁺ subpopulatlon.