Albumin therapy augments the effect of thrombolysis on local vascular dynamics in a rat model of arteriolar thrombosis: a two-photon laser-scanning microscopy study

Abstract

Background and purpose: Results of our recent pilot clinical trial suggest that the efficacy of thrombolytic therapy in acute ischemic stroke may be enhanced by the coadministration of high-dose albumin. Here, we explored the microvascular hemodynamic effects of this combined therapy in a laboratory model of cortical arteriolar thrombosis.

Methods: We studied the cortical microcirculation of physiologically monitored rats in vivo by two-photon laser-scanning microscopy after plasma-labeling with fluorescein-dextran. We induced focal thrombosis in 30- to 50-microm cortical arterioles by laser irradiation and measured arteriolar flow velocity by repeated line-scanning. At 30 minutes post-thrombosis, we treated animals with the thrombolytic agent, reteplase, which was coadministered with either human albumin, 2 g/kg, or with saline control.

Results: Baseline arteriolar flow velocity averaged 3.8+/-0.7 mm/s, was immediately reduced by thrombosis to 22% to 25% of control values, and remained unchanged before treatment. Subthrombolytic doses of reteplase combined with saline led to a median increase in flow velocity to 37% of control distal to the thrombus (P=nonsignificant versus pretreatment). By contrast, reteplase combined with albumin therapy resulted in a prompt, highly significant increase of median flow velocity to 58% of control levels (P=0.013 versus reteplase+saline), which remained significantly higher than the reteplase+saline group at multiple time-points over the subsequent hour.

Conclusions: The beneficial effect of subthrombolytic doses of reteplase on microvascular hemodynamics distal to a cortical arteriolar thrombosis is markedly enhanced by the coadministration of high-dose albumin therapy; these results have important clinical implications for the management of patients with acute ischemic stroke.

Figure 1

Representative experiments in which 30- to 50-µm cortical arterioles were visualized by two-photon laser-scanning microscopy after IV administration of FITC-dextran, showing the effects of laser-induced thrombosis and treatment paradigms. Thrombosis is evident as an interruption of the FITC dye column. The thrombosed vascular segments are variably dilated, and there is focal plasma extravasation into the vessel wall, suggesting vascular injury (20× objective). Panel A: Reteplase, 1.84 g/kg, plus ALB. Panel B: Reteplase, 1.84 g/kg, plus saline; the thrombus is bifid. Panel C: Reteplase, 1.84 g/kg, plus ALB. At 30 minutes post-treatment, the thrombus is divided into two segments separated by plasma column; later, there is partial lysis of the thrombus. Panel D: Reteplase, 1.84 g/kg, plus ALB. At 60 minutes post-treatment, incipient separation of the thrombus from the vessel wall is apparent (arrow). Abbreviations: Baseline indicates before thrombosis; Isch, after thrombus induction, before treatment; Post-Rx, after administration of reteplase along with either ALB or saline; T, thrombus. *site of flow-velocity measurement, downstream from thrombus site.

Figure 2

Arteriolar flow velocity, normalized to each animal’s baseline velocity value, measured at baseline, during 30-minute thrombosis before treatment (Isch-5 through -30), and up to 90 minutes after treatment with either 2.5 g/kg of 25% human albumin (Rx-5 through -90) (upper panel) or isotonic saline control (lower panel). All animals received reteplase. Bars span the 25th to 75th percentiles; whiskers denote the 10% and 90% percentiles; black dots represent outliers; lines connect median values. Median flow velocity declined to 22% to 25% of control values within 5 minutes of thrombus induction. Reteplase+ALB administration was followed promptly by statistically significant median flow-velocity increments of 52% to 64% of control during the subsequent 60 minutes. By contrast, reteplase+saline led to median increments of 37% to 47%, which did not attain statistical significance. *different from corresponding reteplase+saline value, repeated-measures ANOVA followed by Student-Newman-Keuls comparisons.

Figure 3

Median normalized flow velocities in the reteplase+ALB and reteplase+saline groups, illustrating the increment in flow-velocity conferred by ALB therapy during the first post-treatment hour.

Figure 4

Arteriolar flow velocities in rats receiving laser-induced arteriolar thrombosis and subsequent treatment paradigms. The groups receiving ALB or saline (without reteplase) are replotted from our previous study. The reteplase+ALB and reteplase+saline groups are from the present study. Bars represent median values; whisker represents third quartile. The black horizontal line denotes the median flow velocity in the saline-alone group. Relative to this group, all other treatments are associated with incremental flow-velocity improvements.