Abstract

Introduction: Sickle cell disease (SCD) is a debilitating genetic disorder of the red blood cells, leading to acute vaso-occlusive (VOC) events and chronic disease in multiple organs. In the liver, VOC events in the hepatic sinusoids can lead to fatal sickle cell intrahepatic cholestasis (SCIC), which is estimated to occur in approximately 10% of the SCD population. Using contrast enhanced ultrasound (CEUS), we have previously shown that microvascular liver perfusion (MVLP) at basal levels in SCD mice is significantly reduced compared with non-SCD control mice and that MVLP in SCD mice can be further reduced by hypoxia-induced VOC events. Additionally, treatment with the antisickling agent, 5-Hyroxymethylfurfural (5-HMF), can prevent hypoxia-induced VOC reduction in MVLP in SCD mice. A prior study investigated a single time point (1 hour) posthypoxia; the duration and effect beyond 1 hour posthypoxia remained unknown. This investigation was performed to address the duration of the reduction in MVLP and the preventative effect of 5-HMF over a 24-hour period.

Methods: Townes sickle cell mice (SCD), homozygous for hα:βs-globulin, approximately 7−9 weeks of age (n=15) were utilized in these studies. CEUS perfusion imaging (Vevo® 2100) was performed on a central cross-section of the liver at the renal artery. Contrast agent was administered as an intravenous bolus via tail vein to anesthetized mice (isoflurane with ~21% O2) and peak perfusion intensity [PPI] measured in linear arbitrary units [l.a.u.]), and data were analyzed with VisualSonics software. Repeated CEUS measurements were obtained on SCD mice at baseline (prehypoxia), 1, 4, and 24 hours posthypoxic exposure (hypoxic exposure: 60 minutes with approximately 5.5.% O2). 5-HMF at 200 mg/kg PO or vehicle was administered following baseline PPI measurement and approximately 30 minutes before start of hypoxia.

Results:Following hypoxia-induced VOC,MVLP in vehicle-treated SCD was significantly reduced by approximately 25% from baseline levels at 1 and 4 hours posthypoxia, before returning to baseline levels at 24 hours posthypoxia (Table 1). In contrast, SCD mice administered 5-HMF at 200 mg/kg PO showed no reduction in MVLP at 1, 4, and 24 hours posthypoxia. Sickle cell counts were obtained on surrogate animals and data are still pending.

*Significant reduction from baseline, P<0.05, ANOVA with Tukey's multiple comparison test.

Summary: CEUS measured a reduction in MVLP in vehicle-treated SCD mice following hypoxia-induced VOC with a duration of at least 4 hours posthypoxia. By 24 hours posthypoxia, MVLP had returned to baseline (prehypoxia) levels. Prehypoxia treatment with the antisickling agent 5-HMF was able to prevent the hypoxia-induced VOC reduction in MVLP in SCD mice for the entire 24-hour period. These data suggest that prophylactic treatment with an antisickling agent prior to a VOC event may benefit SCD patients by maintaining organ perfusion. These results further validate CEUS as a noninvasive method to measure acute and chronic organ perfusion changes for evaluating new therapeutics for sickle cell-mediated VOC events and end-organ damage.

Table

Microvascular Liver Perfusion (PPI in l.a.u.) Over 24 Hours in SCD Mice With and Without 5-HMF (Mean ± SD)

Table

Microvascular Liver Perfusion (PPI in l.a.u.) Over 24 Hours in SCD Mice With and Without 5-HMF (Mean ± SD)

Disclosures

Abe:Bayer: Employment. Sim:Bayer: Employment. Alonso-Galicia:Bayer: Employment. Kauser:Bayer: Employment.

Author notes

*

Asterisk with author names denotes non-ASH members.