Hagfish Vwf is expressed in endothelium and peripheral blood. (A) qPCR analysis of hemostatic factors in hagfish organs and blood. mRNA expression is represented as copy number per 106 18S copies. Data are presented as mean ± standard deviation (SD) (n = 3 fish). (B) Serial sections of the body wall and muscle of hagfish stained with hematoxylin and eosin (H&E) (left) and processed for immunofluorescence staining using polyclonal anti-human VWF antibody (right). Nuclei are stained with 4′,6-diamidino-2-phenylindole (DAPI). Positive Vwf staining is observed in the aorta (*), posterior cardinal veins (arrows), capillaries of the intestinal wall (¶), and capillaries surrounding red muscle fibers (arrowheads). All stain and magnification information for photomicrographs is provided in the supplemental Material. G, gut lumen; K, kidney; N, notochord; R, red muscle fiber; W, white muscle fiber. (C) Fluorescence microscopy images of hagfish blood processed for immunofluorescence staining of Vwf using polyclonal anti-human VWF antibody. Left image (i) shows 4 Vwf-positive cells (arrows), which are smaller than neighboring Vwf-negative erythrocytes (1 of these is outlined in white). Right image (ii) is a higher-power view showing the punctate staining pattern of a Vwf-positive cell surrounded by Vwf-negative erythrocytes and spindle cells. All stain and magnification information for photomicrographs is provided in the supplemental Material. *Representative spindle cell. See supplemental Figure 4C-D for greater detail of negatively stained cells.
