Methods for extracting platelets and compositions obtained therefrom

#The present invention relates to methods for extracting platelets, compositions obtained therefrom, and methods for using the same.

#New blood vessel formation, or angiogenesis, is requirement for the repair and regeneration of damaged body tissues. This process requires the coordinated activity of multiple cell types including endothelial cells, smooth muscle and other mesenchymal cells, as well as cells of the monocyte/macrophage lineage. A great amount of work has been put into identifying growth factors that can stimulate angiogenesis for therapeutic purposes. Much attention has been focused on the molecule VEGF (vascular endothelial growth factor) because of its central role in the process.

#VEGF is believed to be a very specific stimulator of endothelial cells that line the interior of blood vessels. Augmentation of motility, mitogenesis, and differentiation of endothelial cells can be demonstrated in response to VEGF readily in cell culture. In addition, it has been shown that animals lacking the gene for VEGF die early in development from a failure of blood vessel formation. Thus, it is believed that VEGF could be used for therapeutic purposes to stimulate angiogenesis, for example, in treatment of heart disease and peripheral vascular insufficiency. However, it has been shown that in some instances delivery of VEGF to ischemic tissue leads to the formation of very weak, leaky blood vessels lacking the support needed from the other cell types. In these instances, VEGF alone was not able to stimulate the formation of entire mature, therapeutically valuable, blood vessels. Without being bound by any theory, it is believed that a single growth factor alone is not sufficient to stimulate the coordinated activity of all of the necessary cell types to bring about angiogenesis for therapeutic purposes, e.g., in wound healing.

#FIGS. 1A and 1B are magnified views of the cross-section of a Matrigel plug without and with the composition of the invention, respectively.

#FIG. 2 is a graph showing counts of tubes/field within Matrigel plugs placed subcutaneously in mice for two units of platelets obtained from different patients processed by sonication in physiologic media followed by dialysis against 10 mM HCl followed by lyophilization and resolubilization in concentrated urea and then dialysis again against 10 mM HCl.

#FIG. 3 is a bar graph showing the corresponding counts of capillaries/field of FIG. 2.

#The term “pelletized platelets” refers to platelets that have been concentrated (e.g., by centrifugation or filtration) such that the bulk (typically 90%, often 95%, and more often 98%) of the natural media (e.g., serum, plasma, anticoagulant solution) in which the platelets are naturally found in have been removed.

#When referring to platelets, the term “disrupted” means membranes of platelet cells and granules have been destroyed such that the contents are released.

#“Low molecular weight proteins” refers to proteins having a molecular weight of 10 kD or less, typically 8 kD or less, and often 2 kD or less.