Crossed immunoelectrophoresis showed that factor VIII related protein (VIII R) synthesized
in the human endothelial cell (EC-VIII R) had faster electrophoretic mobility than
that secreted into the culture medium (MED-VIII R). Sodium dodecyl sulphate agarose
gel electrophoresis followed by radioimmunofixation showed that EC-VIII R consisted
of molecules varying from 0.26 × 106 to 3.76 × 106 daltons while MED-VIII R had molecules ranging from 0.93 × 106 to greater than 10 × 106 daltons, similar to that present in plasma. The smallest VIII R molecule present
in normal plasma or spent culture medium (0.93 × 106 daltons) corresponded to a tetramer of subunits of 0.22-0.24 × 106 daltons. Only molecular forms greater than 3.76 × 106 daltons possessed ristocetin cofactor activity. Sonication (15 μ. amplitued for 30
secs) effectively broke the non-covalent bonds of the VIII R multimers resulting in
smaller molecules. Thus endothelial cells in culture synthesized VIII R subunits and
assembled them into the higher multimeric forms on secretion. Different types of von
Willebrand disease could result from defects of either of the two processes.
Keywords
Human endothelial cells - Factor VIII related protein - Multimeric forms - Ristocetin
cofactor
