12 research outputs found
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Collagenase isoforms for pancreas digestion.
The available information concerning the characteristics and composition of collagenase batches, which are effective in the digestion of human pancreas for islet transplants, is scarce and incomplete. A large inter-and intrabatched variability in activity and efficiency of blend enzymes available for isolation has been observed. The aim of this study was to characterize enzyme blend components. Liberase batches were characterized by SDS-PAGE analyses, microelectrophoresis, and then by MALDI-TOF MS analysis. Three main bands were detected by SDS-PAGE analysis and submitted to MALDI-TOF MS analysis. Two bands were found to correspond to class I (isoform Ξ² and another of 106 kDa) and one to class II (isoform Ξ΄) collagenase. These results represent an important step towards a complete characterization of enzymes, with the final aim of identifying key components for a standardized product
A Photoprotein in Mouse Embryonic Stem Cells Measures Ca2+ Mobilization in Cells and in Animals
Exogenous expression of pharmacological targets in transformed cell lines has been the traditional platform for high throughput screening of small molecules. However, exogenous expression in these cells is limited by aberrant dosage, or its toxicity, the potential lack of interaction partners, and alterations to physiology due to transformation itself. Instead, primary cells or cells differentiated from precursors are more physiological, but less amenable to exogenous expression of reporter systems. To overcome this challenge, we stably expressed c-Photina, a Ca2+-sensitive photoprotein, driven by a ubiquitous promoter in a mouse embryonic stem (mES) cell line. The same embryonic stem cell line was also used to generate a transgenic mouse that expresses c-Photina in most tissues. We show here that these cells and mice provide an efficient source of primary cells, cells differentiated from mES cells, including cardiomyocytes, neurons, astrocytes, macrophages, endothelial cells, pancreatic islet cells, stably and robustly expressing c-Photina, and may be exploited for miniaturized high throughput screening. Moreover, we provide evidence that the transgenic mice may be suitable for ex-vivo bioimaging studies in both cells and tissues
Π’Π΅Π»Π΅ΠΌΠ΅Π΄ΠΈΡΠΈΠ½Π°: ΠΏΠ΅ΡΡΠΏΠ΅ΠΊΡΠΈΠ²Ρ ΡΠ°Π·Π²ΠΈΡΠΈΡ
ΠΠ΅ΠΎΠ±Ρ
ΠΎΠ΄ΠΈΠΌΠΎΡΡΡ ΡΠ°Π·Π²ΠΈΡΠΈΡ Π² ΡΡΠ»ΠΎΠ²ΠΈΡΡ
ΡΠΈΡΡΠΎΠ²ΠΎΠΉ ΡΠΊΠΎΠ½ΠΎΠΌΠΈΠΊΠΈ ΡΠ°ΠΊΠΎΠ³ΠΎ Π½Π°ΠΏΡΠ°Π²Π»Π΅Π½ΠΈΡ ΠΊΠ°ΠΊ ΡΠ΅Π»Π΅ΠΌΠ΅Π΄ΠΈΡΠΈΠ½Π° ΠΊΠ°ΠΆΠ΅ΡΡΡ Π½Π΅ΠΎΠ±Ρ
ΠΎΠ΄ΠΈΠΌΠΎΡΡΡΡ, ΠΎΠ΄Π½Π°ΠΊΠΎ ΡΡΡΠ΅ΡΡΠ²ΡΠ΅Ρ ΠΏΡΠΎΡΠΈΠ²ΠΎΡΠ΅ΡΠΈΠ΅ ΠΌΠ΅ΠΆΠ΄Ρ ΡΡΡΠ΅ΡΡΠ²ΡΡΡΠΈΠΌΠΈ ΠΏΡΠΎΠ±Π»Π΅ΠΌΠ°ΠΌΠΈ Π² ΡΠΈΡΡΠ΅ΠΌΠ΅ ΠΎΡΠ³Π°Π½ΠΈΠ·Π°ΡΠΈΠΈ Π·Π΄ΡΠ°Π²ΠΎΠΎΡ
ΡΠ°Π½Π΅Π½ΠΈΡ Π ΠΎΡΡΠΈΠΉΡΠΊΠΎΠΉ Π€Π΅Π΄Π΅ΡΠ°ΡΠΈΠΈ ΠΈ Π²ΠΎΠ·ΠΌΠΎΠΆΠ½ΠΎΡΡΡΠΌΠΈ Π΄ΠΎΡΡΡΠΏΠ½ΠΎΡΡΠΈ ΠΊ ΡΡΠ»ΡΠ³Π°ΠΌ ΡΠ΅Π»Π΅ΠΌΠ΅Π΄ΠΈΡΠΈΠ½Ρ. ΠΠ½Π°Π»ΠΈΠ· ΠΏΠΎΠΊΠ°Π·Π°Π», ΡΡΠΎ Π΄Π°Π½Π½ΠΎΠ΅ Π½Π°ΠΏΡΠ°Π²Π»Π΅Π½ΠΈΠ΅ Π²ΠΎΡΡΡΠ΅Π±ΠΎΠ²Π°Π½ΠΎ, Π½Π΅ΠΎΠ±Ρ
ΠΎΠ΄ΠΈΠΌΠΎ, ΡΡΡΠ΅ΡΡΠ²ΡΠ΅Ρ ΡΡΠ΄ ΡΠ΅ΡΠ²ΠΈΡΠΎΠ², ΠΏΡΠ΅Π΄Π»Π°Π³Π°ΡΡΠΈΡ
ΡΡΠ»ΡΠ³ΠΈ ΠΏΠΎ ΡΠ΅Π»Π΅ΠΌΠ΅Π΄ΠΈΡΠΈΠ½Π΅, ΠΎΠ΄Π½Π°ΠΊΠΎ Π½ΠΎΡΠΌΠ°ΡΠΈΠ²Π½ΠΎ-ΠΏΡΠ°Π²ΠΎΠ²Π°Ρ Π±Π°Π·Π° Π½Π΅ ΠΏΠΎΠ·Π²ΠΎΠ»ΡΠ΅Ρ Π² ΠΏΠΎΠ»Π½ΠΎΠΉ ΠΌΠ΅ΡΠ΅ ΠΎΡΠ³Π°Π½ΠΈΠ·ΠΎΠ²Π°ΡΡ ΠΏΠΎΠ»Π½ΠΎΡΠ΅Π½Π½ΡΡ ΡΠ°Π±ΠΎΡΡ ΠΏΠΎ ΠΏΡΠ΅Π΄ΠΎΡΡΠ°Π²Π»Π΅Π½ΠΈΡ ΡΠ΅Π»Π΅ΠΌΠ΅Π΄ΠΈΡΠΈΠ½ΡΠΊΠΈΡ
ΡΡΠ»ΡΠ³, Π² ΡΠΎΠΌ ΡΠΈΡΠ»Π΅, Π±Π΅ΡΠΏΠ»Π°ΡΠ½ΡΡ
Functional characterization of the Opitz syndrome gene product (midin): evidence for homodimerization and association with microtubules throughout the cell cycle
Opitz syndrome (OS) is a multiple congenital anomaly manifested by abnormal closure of midline structures. The gene responsible for the X-linked form of this disease, MID1, encodes a protein (midin) that contains a RING, two B-boxes, a coiled-coil (the so-called tripartite motif) and an RFP-like domain. The tripartite motif is characteristic of a family of proteins, named the B-box family, involved in cell proliferation and development. Since the subcellular compartmentalization and the ability to form multiprotein structures both appear to be crucial for the function of this family of proteins, we have studied these properties on the wild-type and mutated forms of midin. We found that endogenous midin is associated with microtubules throughout the cell cycle, co-localizing with cytoplasmic fibres in interphase and with the mitotic spindle and midbodies during mitosis and cytokinesis. Immunoprecipitation experiments demonstrated the ability of the tripartite motif to mediate midin homodimerization, consistent with the evidence, obtained by gel filtration analysis, that midin exists in the form of large protein complexes. Functional characterization of altered forms of midin, resulting from mutations found in OS patients, revealed that association with microtubules is compromised, while the ability to homodimerize and form multiprotein complexes is retained. We suggest that midin is involved in the formation of multiprotein structures acting as anchor points to microtubules and that impaired association with these cytoskeletal structures causes OS developmental defects
Technology-in-Practice: New Insight Into Data Warehouse Adoption
Traditionally the data warehouse (DW) has been treated in purely technical terms based on the hypothesis that the investment in technology would necessarily bring about predetermined solutions. The observation of empirical cases seems to contradict this point of view by placing the role of people and the emergent use of technology in first place. The paper examines the results of DW use in three organizations that have been using it for four years, using the same software in their recurrent practices. This project aims at identifying the emergent uses of DW and the driving forces that seem to bring forth different and unexpected results.
MID2, a homologue of the Opitz syndrome gene MID1: similarities in subcellular localization and differences in expression during development
The B-box family is an expanding new family of genes encoding proteins involved in diverse cellular functions such as developmental patterning and oncogenesis. A member of this protein family, MID1, is the gene responsible for the X-linked form of Opitz G/BBB syndrome, a developmental disorder characterized by defects of the midline structures. We now report the identification of MID2, a new transcript closely related to MID1. MID2 maps to Xq22 in human and to the syntenic region on the mouse X chromosome. The two X-linked genes share the same domains, the same exon-intron organization, a high degree of similarity at the protein level and the same subcellular localization, both being confined to the cytoplasm in association to micro-tubular structures. The expression pattern studied by RNA in situ hybridization in mouse revealed that Mid2 is expressed early in development and the highest level of expression is detected in the heart, unlike Mid1 for which no expression was detected in the developing heart. Together, these data suggest that midin and MID2 have a similar biochemical function but a different physiological role during development