Co-localization of von Willebrand factor and type VI collagen in human vascular subendothelium.

The binding of von Willebrand factor (vWF) to subendothelium constitutes an important initial step in the process of platelet adhesion to exposed subendothelium following blood vessel injury. We previously demonstrated that vWF is present in human vascular subendothelium and recently found that a 150 kd vWF-binding protein, which we extracted from subendothelium, is type VI collagen. Although we have established that vWF and type VI collagen bind in vitro, it is not known whether these two proteins are associated in the vascular subendothelium in situ. We, therefore, 1) investigated the morphological effects of our biochemical extraction procedure on human umbilical veins by scanning and transmission electron microscopy, 2) analyzed the subendothelial extract by immunofluorescence for the presence of vWF and collagens and by electron microscopy for morphological characteristics, and 3) localized vWF and type VI collagen in subendothelium by immunofluorescence and by single- and double-label immunoelectron microscopic studies with protein A-conjugated gold particles. We found that the surface exposed following de-endothelialization is composed of microfibrils and contains very little fibrillar collagen. The subendothelium is stripped after sodium dodecyl sulfate-urea extraction, and the extract itself contains immunoreactive vWF and type VI collagen but no immunoreactive type I or III fibrillar collagens. Immunofluorescence and immunoelectron microscopic studies showed that vWF and type VI collagen are both present in subendothelium, where both co-localized to microfibrils. In conclusion, vWF that binds to type VI collagen in vitro, also co-localizes with type VI collagen in subendothelium, where both are associated with microfibrils. Type VI collagen, therefore appears to serve as a biologically significant binding site for vWF in vivo and may thereby play a role in mediating platelet adhesion to exposed subendothelium following vascular injury.