Application of long-acting VLHL PAI-1 during sutureless partial nephrectomy in mice reduces bleeding

Abstract

PAI-1 prevents lysis of blood clot by inhibiting the urokinase and tPA induced conversion of plasminogen to plasmin. VLHL PAI-1 protein mutant was created to extend half-life over 700 hours. The objective of this paper was to test VLHL PAI-1 effects on bleeding during partial nephrectomy in mice. All animals had a left partial nephrectomy after intravenous infusion of saline or tPA. The animals were divided into four groups. Group 1 was infused with saline and kidney was exposed to saline too; Group 2 was infused with saline and kidney was exposed to PAI-1. Group 3 was infused with tPA and kidney was exposed to saline, while Group 4 was infused with tPA and kidney was exposed to PAI-1. Preweighed gauze containing PAI-1 or saline was then applied to the kidney for 30 minutes. The gauze was afterward weighed and blood loss was measured by subtracting the preweight of gauze from the final weight. We have observed a statistically significant (P ≤ 0.05) reduction of bleeding in PAI-1-treated group in comparison to saline and tPA-treated groups. Based on these results we propose that VLHL PAI-1 can be used therapeutically in limiting the flow of blood from renal wounds.

Figure 1

Plasma, solid line (no tPA, no VLHL PAI-1), clotted blood with mouse tPA alone (tPA 0.5 μg/mL), dash dot line, and clotted blood with mouse tPA (tPA 0.5 μg/mL) + VLHL PAI-1 (VLHL PAI-1  1.4 μg/mL), short dash line. Insert shows complex formation of VLHL PAI-1 with tPA. Lane 1, mouse tPA; lane 2, VLHL PAI-1; lane 3, mouse tPA + VLHL PAI-1.

Figure 2

((a) and (b)) Cannulation of jugular vein in preparation for infusion of tPA or saline. ((c) and (d)) Clamping of renal vascular pedicle and partial nephrectomy. ((e) and (f)) Placement of PAI-1 or saline saturated gauze onto the surface of the renal injury.

Figure 3

(a) Total blood loss; (b) normalized total blood loss. Saline treated: Group 1, VLHL PAI-1 treated: Group 2, mouse tPA treated: Group 3, and mouse tPA + VLHL treated: Group 4. Blood loss was normalized by adjusting volume of blood by ratio of weight of the resected piece of kidney to remaining kidney. Statistical significance of differences between different treatment groups are in Table 2.