Mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin α5 in the glomerular basement membrane

In developing glomeruli, laminin α5 replaces laminin α1 in the glomerular basement membrane (GBM) at the capillary loop stage, a transition required for glomerulogenesis. To investigate domain-specific functions of laminin α5 during glomerulogenesis, we produced transgenic mice that express a chimeric laminin composed of laminin α5 domains VI through I fused to the human laminin α1 globular (G) domain, designated Mr51. Transgene-derived protein accumulated in many basement membranes, including the developing GBM. When bred onto the Lama5 −/− background, Mr51 supported GBM formation, preventing the breakdown that normally occurs in Lama5 −/− glomeruli. In addition, podocytes exhibited their typical arrangement in a single cell layer epithelium adjacent to the GBM, but convolution of glomerular capillaries did not occur. Instead, capillaries were distended and exhibited a ballooned appearance, a phenotype similar to that observed in the total absence of mesangial cells. However, here the phenotype could be attributed to the lack of mesangial cell adhesion to the GBM, suggesting that the G domain of laminin α5 is essential for this adhesion. Analysis of an additional chimeric transgene allowed us to narrow the region of the α5 G domain essential for mesangial cell adhesion to α5LG3-5. Finally, in vitro studies showed that integrin α3β1 and the Lutheran glycoprotein mediate adhesion of mesangial cells to laminin α5. Our results elucidate a mechanism whereby mesangial cells organize the glomerular capillaries by adhering to the G domain of laminin α5 in the GBM

Laminin β2 variants associated with isolated nephropathy that impact matrix regulation

Mutations in LAMB2, encoding laminin β2, cause Pierson syndrome and occasionally milder nephropathy without extrarenal abnormalities. The most deleterious missense mutations that have been identified affect primarily the N-terminus of laminin β2. On the other hand, those associated with isolated nephropathy are distributed across the entire molecule, and variants in the β2 LEa-LF-LEb domains are exclusively found in cases with isolated nephropathy. Here we report the clinical features of mild isolated nephropathy associated with 3 LAMB2 variants in the LEa-LF-LEb domains (p.R469Q, p.G699R, and p.R1078C) and their biochemical characterization. Although Pierson syndrome missense mutations often inhibit laminin β2 secretion, the 3 recombinant variants were secreted as efficiently as WT. However, the β2 variants lost pH dependency for heparin binding, resulting in aberrant binding under physiologic conditions. This suggests that the binding of laminin β2 to negatively charged molecules is involved in glomerular basement membrane (GBM) permselectivity. Moreover, the excessive binding of the β2 variants to other laminins appears to lead to their increased deposition in the GBM. Laminin β2 also serves as a potentially novel cell-adhesive ligand for integrin α4β1. Our findings define biochemical functions of laminin β2 variants influencing glomerular filtration that may underlie the pathogenesis of isolated nephropathy caused by LAMB2 abnormalities

Syndecan- and integrin-binding peptides synergistically accelerate cell adhesion

AbstractIntegrins and syndecans mediate cell adhesion to extracellular matrix and their synergistic cooperation is implicated in cell adhesion processes. We previously identified two active peptides, AG73 and EF1, from the laminin α1 chain LG4 module, that promote cell attachment through syndecan- and α2β1 integrin-binding, respectively. Here, we examined time-dependent cell attachment on the mixed peptides AG73/EF1. The AG73/EF1 promoted stronger and more rapid cell attachment, spreading, FAK phosphorylation that reached a maximum at 20min than that on AG73 (40min) or EF1 (90min) supplied singly. Thus, the syndecan- and α2β1 integrin-binding peptides synergistically affect cells and accelerate cell adhesion

A three-dimensional microfluidic tumor cell migration assay to screen the effect of anti-migratory drugs and interstitial flow

Most anti-cancer drug screening assays are currently performed in two dimensions, on flat, rigid surfaces. However, there are increasing indications that three-dimensional (3D) platforms provide a more realistic setting to investigate accurate morphology, growth, and sensitivity of tumor cells to chemical factors. Moreover, interstitial flow plays a pivotal role in tumor growth. Here, we present a microfluidic 3D platform to investigate behaviors of tumor cells in flow conditions with anti-migratory compounds. Our results show that interstitial flow and its direction have significant impact on migration and growth of hepatocellular carcinoma cell lines such as HepG2 and HLE. In particular, HepG2/HLE cells tend to migrate against interstitial flow, and their growth increases in interstitial flow conditions regardless of the flow direction. Furthermore, this migratory activity of HepG2 cells is enhanced when they are co-cultured with human umbilical vein endothelial cells. We also found that migration activity of HepG2 cells attenuates under hypoxic conditions. In addition, the effect of Artemisinin, an anti-migratory compound, on HepG2 cells was quantitatively analyzed. The microfluidic 3D platform described here is useful to investigate more accurately the effect of anti-migratory drugs on tumor cells and the critical influence of interstitial flow than 2D culture models.Japan Society for the Promotion of Science (22680037)Japan Society for the Promotion of Science (G2212)National Cancer Institute (U.S.) (R21CA140096)Japan. Ministry of Education, Culture, Sports, Science and Technology (2009-00631)Japan. Ministry of Education, Culture, Sports, Science and Technology (2012-0009565)Korea (South). Ministry of Education & Human Resources Development (MOEHRD)Korea (South). Ministry of Education & Human Resources Development (MOEHRD) (20124010203250

Laminin isoforms in human embryonic stem cells : synthesis, receptor usage and growth support

Sanna Vuoristo, Ismo Virtanen, Minna Takkunen, Jaan Palgi, Yamato Kikkawa, Patricia Rousselle, Kiyotoshi Sekiguchi, Timo Tuuri, and Timo Otonkoski, "Laminin isoforms in human embryonic stem cells: synthesis, receptor usage and growth support", Journal of Cellular and Molecular Medicine, 13, 8b, pp. 2622-2633, Wiley, 200

Purification and Characterization of Human Laminin-8 : LAMININ-8 STIMULATES CELL ADHESION AND MIGRATION THROUGH α3β1 AND α6β1INTEGRINS

This research was originally published in the Journal of Biological Chemistry. Hironobu Fujiwara, Yamato Kikkawa, Noriko Sanzen and Kiyotoshi Sekiguchi. Purification and Characterization of Human Laminin-8 : LAMININ-8 STIMULATES CELL ADHESION AND MIGRATION THROUGH α3β1 AND α6β1INTEGRINS. J. Biol. Chem. 2001; 276: 17550-17558 © the American Society for Biochemistry and Molecular Biolog

Die noodsaaklikheid van ’n veelsydige persoon- likheidsontwikkeling en in waiter mate die lees van keurleesstof daartoe kan hydra

This research was originally published in the Journal of Biological Chemistry. Yamato Kikkawa, Noriko Sanzen and Kiyotoshi Sekiguchi. Isolation and Characterization of Laminin-10/11 Secreted by Human Lung Carcinoma Cells : LAMININ-10/11 MEDIATES CELL ADHESION THROUGH INTEGRIN α3β1. J. Biol. Chem. 1998; 273: 15854-15859 © the American Society for Biochemistry and Molecular Biolog

Critical Examination of a Revision of Bloom's Taxonomy for the Development of Art Education Curriculum

This paper is aimed at developing Art Education Taxonomy Table through critically examining a revision of Bloom's Taxonomy of Educational Objectives contrived by Anderson and Krathwohl. First, the overview of the history of Bloom's Taxonomy is provided in relation to art education. Second, the influence of Bloom's theory of educational evaluation and Taxonomy on the classroom practice of art education in Japan is discussed. Third, aiming at using for the development of art education curriculum at Mihara Elementary School and Junior High School attached to Hiroshima University, Art Education Taxonomy Table is constructed and its characteristics are specified in relation to the research theme of Mihara Educational Institution