Liposome-Encapsulated Hemoglobin Ameliorates Ischemic Stroke in Nonhuman Primates: An Acute Study

Abstract

An artificial oxygen carrier, liposome-encapsulated hemoglobin (LEH), protective in a rodent stroke model, was quantitatively evaluated in monkeys. Serial positron emission tomography studies using the steady-state 15 O-gas inhalation method were performed to quantify O 2 metabolism, which was compared based on the infarction extent and immunohistochemical evaluation in 19 monkeys undergoing middle cerebral artery occlusion (3 h), infusion of various LEH doses (n ϭ 11), empty liposome (n ϭ 4), or saline (n ϭ 4) 5 min after the onset of ischemia, and reperfusion for 5 h. There was no significant difference in O 2 metabolism until 3 h after reperfusion, when the cerebral metabolic rate of O 2 (CMRO 2 ) was significantly less suppressed in the cortex [mild suppression in CMRO 2 (71-100%) of preischemic ipsilateral control as in the ischemic penumbra: 64.7 Ϯ 14.3% in empty liposome versus 32.4 Ϯ 7.9% in LEH (2 ml/kg) treatment, P Ͻ 0.05] but not in basal ganglia. Immunohistochemical studies showed a reciprocal expression of microtubular-associated protein II expression in the cortex and LEH deposition in basal ganglia, suggesting the LEH perfusion, but not deposition, afforded the protection. Doseresponse studies revealed that as little as 0.4 ml/kg LEH (24 mg/kg hemoglobin) was effective in preserving CMRO 2 , whereas 2 and 10 ml/kg were protective in significantly reducing the area of infarction as well, by 66 and 56%, respectively, compared with animals receiving saline. CMRO 2 and histological integrity were better preserved early after 3-h occlusion and reperfusion of the middle cerebral artery of monkeys receiving LEH early after onset of ischemia.Although ischemic stroke is a major cause of morbidity and mortality in developed societies (Broderick et al., 1998), medical attention is often delayed because of low public awareness of stroke symptoms (Williams et al., 1997). Increased risk of hemorrhagic transformation (Lapchak, 2002) limits the therapeutic time window and makes it more difficult to apply reperfusion therapy (NINDS rt-PA Stroke Study Group, 1995), as with coronary circulation. Although cell-free hemoglobin has been proposed to reduce ischemic injury, the associated toxicities have limited its utility (Natanson et al., 2008). Liposome-encapsulated human hemoglobin (LEH) (Ogata Y, 2000) has been developed as an artificial O 2 carrier that is functionally and structurally similar to red blood cells (RBCs). The lipid bilayer of encapsulated hemoglobin prevents renal clearance, extravasation, and direct contact of hemoglobin with vascular smooth muscle (Ogata, 2001), thus prohibiting vascular constriction, prolonging the in vivo halflife of LEH (Kaneda et al., 2009), and reducing the dosage required for beneficial effects (Kawaguchi et al., 2007). Although liposomes are much smaller (230 nm), their O 2 -carrying capacity is equivalent to human RBCs under room air respiration (Kaneda et al., 2009). These physical characteristics prompted us to examine the therapeutic potential of LEH, not as an...

Post-ischemic intra-arterial infusion of liposome-encapsulated hemoglobin can reduce ischemia reperfusion injury

“…Since LEH can deliver oxygen more efficiently to peripheral tissues because of its smaller diameter (200-250 nm; 1/40th the size of RBC), several studies have attempted to assess the therapeutic effects of LEH on brain ischemia. Intravenous administration of LEH reduced the extent of infarction in the cortex of a rat thrombotic MCAO model (Kawaguchi et al, 2007;Fukumoto et al, 2009) and in a nonhuman primate transient MCAO model (Kawaguchi et al, 2010). Another therapeutic effect of LEH on cognitive impairment was revealed in the analysis of hippocampal dysfunction after transient two-vessel or four-vessel occlusion (Kakehata et al, 2007;Hamadate et al, 2009).…”

Getting into the brain: liposome-based strategies for effective drug delivery across the blood–brain barrier

“…Several studies reported in the literature suggest the efficacy of Hb liposomes in the treatment of stroke by enhancing the biodistribution of Hb liposomes within the ischemic area in the brain. 144,145,147,[160][161][162][163][164]167,171,191,193,194 In the same way, liposomes for the delivery of angiogenic peptides 140 and VEGF 157 to promote angiogenesis in ischemic tissue were developed, and both formulations effectively promoted vascular regeneration. 140,157 Over the years, many liposomal formulations have been developed for the treatment of stroke.…”

Liposome-Encapsulated Hemoglobin Ameliorates Impairment of Fear Memory and Hippocampal Dysfunction After Cerebral Ischemia in Rats

“…LEH reduced the size of cerebral infarction in rats subjected to photochemically induced thrombosis (PIT) of the middle cerebral artery (MCA) (6,7) and also in primates treated with MCA occlusion (MCAO) (8). Transfusion of LEH rescued lethal progressive hemodilution and improved hemodilution-induced metabolic acidosis in rats (9).…”