| Research Abstract |
We have suggested that annexin VI functions as a cell-surface receptor for the chondroitin sulfate chains of proteoglycan, PG-M in a Ca^<2+>-dependent manner to transduce their anti-adhesive signals into the cells. Here we have investigated the possibility and mechanisms from the following two aspects : 1) We previously succeeded in synthesizing chondroitin sulfate-phosphatidyl ethanolamine lipid (CS-PE) which can be substituted for PG-M in regard to the anti-adhesive activity and have taken advantage of dishes coated with this new molecules for anti-adhesive substrata. Their anti-adhesive properties have been investigated using both the cell attachment assay and the cell spreading assay, which are aimed preferentially to analyze the interactions between cells and substrate molecules and the subsequent cytoskeletal changes to result in the cell spreading, respectively. As far as examined, cells of any different types could attach to dishes coated with fibronectin but not spread on it w
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hen doubly coated with CS-PE.In addition, cells could also attach to dishes coated with CS-PE alone. However, A431, a cell line derived from human epithelium which we previously found to lack annexin VI could hardly attach to the CS-PE-coated dish. Any cells including A431 could attach to and subsequently spread on dishes coated with glycosaminoglycan-lipids other than CS-PE.Taken together, the results suggest the involvement of annexin VI in a specific signal transduction to inhibit the subsequent cell-spreading reaction. Various reagents such as protein kinase-inhibitors, its activators, and inophors were tested to have any effects on the anti-adhesive activity and none of them gave the significant effects. 2) Changes in cellular morphology and migration activity were analyzed when cells were transfected with the vectors containing cDNAs for various alternative spliced forms of PG-M to yield the molecules having various potential attachment sites for chondroitin sulfate chains. Transgenic mouse systems were also used to examine the effects. However, we have not observed any significant differences. The expression systems are now taken into reconsideration to yield the enough expression. Less
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