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991.
为了解水压转染法(hydrodynamics-based transfection,HDT)在小鼠肝脏不同肝叶的转染效率,将容量为小鼠体重的10%,绿色荧光蛋白基因质粒pEGFP-C1含量为35μg/只的生理盐水溶液以0.4 mL/s的速度从小鼠尾静脉注射,于注射后不同时间取小鼠各肝叶制备冰冻切片,在荧光显微镜下观察,计数各肝叶的绿色荧光蛋白表达情况。结果显示注射后12 h,绿色荧光蛋白阳性细胞比例最高,从24 h起表达量逐渐减少,至48 h时各肝叶均基本难以检测出绿色荧光蛋白阳性细胞。在12 h观察各肝叶的转染效率如下:右叶、蒂状叶的绿色荧光蛋白阳性细胞约为22%,左叶、中叶、尾状叶约为15%取材部位不同会造成数据分析的极显著差异。 相似文献
992.
993.
Ouahiba Mesbah-Benmessaoud Roza Benabdesselam H��l��ne Hardin-Pouzet Latifa Dorbani-Mamine Val��rie Grange-Messent 《The journal of histochemistry and cytochemistry》2011,59(1):88-97
Water channel aquaporin-4 (AQP4) is the most abundant water channel in the rodent brain and is mainly expressed in cerebral areas involved in central osmoreception and osmoregulation. The neurohypophysis is the release site of hypothalamic neurohormones vasopressin and oxytocin, which are involved in the regulation of the water balance. The authors investigated the cellular and subcellular distribution of AQP4 in the mouse neurohypophysis before and after chronic osmotic stimulation, using immunofluorescence microscopy and immunoperoxidase electron microscopy. They showed that AQP4 was abundant in the mouse hypophysis, mainly in the neural lobe. AQP4 was discontinuously distributed along pituicytes plasma membranes, in the dense neurosecretory granules and microvesicles of nerve endings and fibers, and along the luminal and abluminal membranes of fenestrated capillary endothelial cells. After chronic osmotic stimulation, AQP4 immunolabeling was enhanced. Taken together, these results suggest that AQP4 could be involved in the pituicyte sensor effect during osmoregulation, the modification and/or maturation mechanism of neurosecretory granules during neurohormone release, and the blood perfusion of the hypophysis. 相似文献
994.
Ghaemmaghami S Watts JC Nguyen HO Hayashi S DeArmond SJ Prusiner SB 《Journal of molecular biology》2011,413(3):527-526
Prion protein is capable of folding into multiple self-replicating prion strains that produce phenotypically distinct neurological disorders. Although prion strains often breed true upon passage, they can also transform or “mutate” despite being devoid of nucleic acids. To dissect the mechanism of prion strain transformation, we studied the physicochemical evolution of a mouse synthetic prion (MoSP) strain, MoSP1, after repeated passage in mice and cultured cells. We show that MoSP1 gradually adopted shorter incubation times and lower conformational stabilities. These changes were accompanied by structural transformation, as indicated by a shift in the molecular mass of the protease-resistant core of MoSP1 from approximately 19 kDa [MoSP1(2)] to 21 kDa [MoSP1(1)]. We show that MoSP1(1) and MoSP1(2) can breed with fidelity when cloned in cells; however, when present as a mixture, MoSP1(1) preferentially proliferated, leading to the disappearance of MoSP1(2). In culture, the rate of this transformation process can be influenced by the composition of the culture media and the presence of polyamidoamines. Our findings demonstrate that prions can exist as a conformationally diverse population of strains, each capable of replicating with high fidelity. Rare conformational conversion, followed by competitive selection among the resulting pool of conformers, provides a mechanism for the adaptation of the prion population to its host environment. 相似文献
995.
Deniz S Wersinger E Schwab Y Mura C Erdelyi F Szabó G Rendon A Sahel JA Picaud S Roux MJ 《Journal of neurochemistry》2011,116(3):350-362
Lateral interactions at the first retinal synapse have been initially proposed to involve GABA by transporter-mediated release from horizontal cells, onto GABA(A) receptors expressed on cone photoreceptor terminals and/or bipolar cell dendrites. However, in the mammalian retina, horizontal cells do not seem to contain GABA systematically or to express membrane GABA transporters. We here report that mouse retinal horizontal cells express GAD65 and/or GAD67 mRNA, and were weakly but consistently immunostained for GAD65/67. While GABA was readily detected after intracardiac perfusion, it was lost during classical preparation for histology or electrophysiology. It could not be restored by incubation in a GABA-containing medium, confirming the absence of membrane GABA transporters in these cells. However, GABA was synthesized de novo from glutamate or glutamine, upon addition of pyridoxal 5'-phosphate, a cofactor of GAD65/67. Mouse horizontal cells are thus atypical GABAergic neurons, with no functional GABA uptake, but a glutamate and/or glutamine transport system allowing GABA synthesis, probably depending physiologically from glutamate released by photoreceptors. Our results suggest that the role of GABA in lateral inhibition may have been underestimated, at least in mammals, and that tissue pre-incubation with glutamine and pyridoxal 5'-phosphate should yield a more precise estimate of outer retinal processing. 相似文献
996.
Da Silva AS Fanfa VR Otto MA Gressler LT Tavares KC Lazzarotto CR Tonin AA Miletti LC Duarte MM Monteiro SG 《The Korean journal of parasitology》2011,49(4):427-430
The aim of this study was to test the susceptibility of mice to Trypanosoma evansi treated with human plasma containing different concentrations of apolipoprotein L-1 (APOL1). For this experiment, a strain of T. evansi and human plasma (plasmas 1, 2, and 3) from 3 adult males clinically healthy were used. In vivo test used 50 mice divided in 5 groups (A to E) with 10 animals in each group. Animals of groups B to E were infected, and then treated with 0.2 ml of human plasma in the following outline: negative control (A), positive control (B), treatment with plasma 1 (C), treatment with plasma 2 (D), and treatment with plasma 3 (E). Mice treated with human plasma showed an increase in longevity of 40.9 ± 0.3 (C), 20 ± 9.0 (D) and 35.6 ± 9.3 (E) days compared to the control group (B) which was 4.3 ± 0.5 days. The number of surviving mice and free of the parasite (blood smear and PCR negative) at the end of the experiment was 90%, 0%, and 60% for groups C, D, and E, respectively. The quantification of APOL1 was performed due to the large difference in the treatments that differed in the source plasma. In plasmas 1, 2, and 3 was detected the concentration of 194, 99, and 115 mg/dl of APOL1, respectively. However, we believe that this difference in the treatment efficiency is related to the level of APOL1 in plasmas. 相似文献
997.
Copper(II)-induced secondary structure changes and reduced folding stability of the prion protein 总被引:1,自引:0,他引:1
Younan ND Klewpatinond M Davies P Ruban AV Brown DR Viles JH 《Journal of molecular biology》2011,410(3):369-382
The cellular isoform of the prion protein PrPC is a Cu2+-binding cell surface glycoprotein that, when misfolded, is responsible for a range of transmissible spongiform encephalopathies. As changes in PrPC conformation are intimately linked with disease pathogenesis, the effect of Cu2+ ions on the structure and stability of the protein has been investigated. Urea unfolding studies indicate that Cu2+ ions destabilise the native fold of PrPC. The midpoint of the unfolding transition is reduced by 0.73 ± 0.07 M urea in the presence of 1 mol equiv of Cu2+. This equates to an appreciable difference in free energy of unfolding (2.02 ± 0.05 kJ mol− 1 at the midpoint of unfolding). We relate Cu2+-induced changes in secondary structure for full-length PrP(23-231) to smaller Cu2+ binding fragments. In particular, Cu2+-induced structural changes can directly be attributed to Cu2+ binding to the octarepeat region of PrPC. Furthermore, a β-sheet-like transition that is observed when Cu ions are bound to the amyloidogenic fragment of PrP (residues 90-126) is due only to local Cu2+ coordination to the individual binding sites centred at His95 and His110. Cu2+ binding does not directly generate a β-sheet conformation within PrPC; however, Cu2+ ions do destabilise the native fold of PrPC and may make the transition to a misfolded state more favourable. 相似文献
998.
999.
Li F He Z Li Y Liu P Chen F Wang M Zhu H Ding X Wangensteen KJ Hu Y Wang X 《Journal of cellular biochemistry》2011,112(4):1022-1034
Induction of definitive endoderm (DE) cells is a prerequisite for the whole process of embryonic stem (ES) cells differentiating into hepatic or pancreatic progenitor cells. We have established an efficient method to induce mouse ES cell-derived DE cells in suspension embryonic body (EB) culture. Similar to previous studies, mouse ES cell-derived DE cells, which were defined as Cxcr4(+) c-Kit(+) , Cxcr4(+) E-cadherin(+) cells or Cxcr4(+) PDGFRa(-) cells, could be induced in the serum-free EBs at Day 4 of induction. The activations of Wnt, Nodal, and FGF signaling pathways in differentiating EBs promoted DE cell differentiation, while activation of BMP4 signaling inhibited the process. In the present study, we found that chemical activation of canonical Wnt signaling pathway by LiCl could synergize with Activin A-mediated Nodal signaling pathway to promote induction of DE cells, and inhibition of Bmp4 signaling by Noggin along with Activin A/LiCl further improved the efficiency of DE cell differentiation. The derived DE cells were proved for their capacities to become hepatic progenitor cells or pancreatic progenitor cells. In conclusion, we significantly improved the efficiency of generating mouse ES cell-derived DE cells by combined Activin A/LiCl/Noggin treatment. Our work will be greatly helpful to generate ES cell-derived hepatic cells and ES cell-derived pancreatic cells for future regenerative medicine. 相似文献
1000.
Hristina Ivanova Tim Vervliet Ludwig Missiaen Jan B. ParysHumbert De Smedt Geert Bultynck 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》2014
Cell-death and -survival decisions are critically controlled by intracellular Ca2 + homeostasis and dynamics at the level of the endoplasmic reticulum (ER). Inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) play a pivotal role in these processes by mediating Ca2 + flux from the ER into the cytosol and mitochondria. Hence, it is clear that many pro-survival and pro-death signaling pathways and proteins affect Ca2 + signaling by directly targeting IP3R channels, which can happen in an IP3R-isoform-dependent manner. In this review, we will focus on how the different IP3R isoforms (IP3R1, IP3R2 and IP3R3) control cell death and survival. First, we will present an overview of the isoform-specific regulation of IP3Rs by cellular factors like IP3, Ca2 +, Ca2 +-binding proteins, adenosine triphosphate (ATP), thiol modification, phosphorylation and interacting proteins, and of IP3R-isoform specific expression patterns. Second, we will discuss the role of the ER as a Ca2 + store in cell death and survival and how IP3Rs and pro-survival/pro-death proteins can modulate the basal ER Ca2 + leak. Third, we will review the regulation of the Ca2 +-flux properties of the IP3R isoforms by the ER-resident and by the cytoplasmic proteins involved in cell death and survival as well as by redox regulation. Hence, we aim to highlight the specific roles of the various IP3R isoforms in cell-death and -survival signaling. This article is part of a Special Issue entitled: Calcium signaling in health and disease. Guest Editors: Geert Bultynck, Jacques Haiech, Claus W. Heizmann, Joachim Krebs, and Marc Moreau. 相似文献