Differential Lung Uptake of ^99mTc-Hexamethylpropyleneamine Oxime and ^99mTc-Duramycin in the Chronic Hyperoxia Rat Model

Abstract

Noninvasive radionuclide imaging has the potential to identify and assess mechanisms involved in particular stages of lung injury which occur with acute respiratory distress syndrome, for example. Lung uptake of 99mTc-hexamethylpropyleneamine oxime (HMPAO) is reported to be partially dependent on the redox status of the lung tissue while 99mTc-duramycin, a new marker of cell injury, senses cell death via apoptosis and/or necrosis. Thus, we investigated changes in lung uptake of these agents in rat exposed to hyperoxia for prolonged periods, a common model of acute lung injury. Methods Male Sprague-Dawley rats were pre-exposed to either normoxia (21% O2) or hyperoxia (85% O2) for up to 21 days. For imaging, the rats were anesthetized, injected i.v. with either 99mTc-HMPAO or 99mTc-duramycin (37-74 MBq) and planar images were acquired using a high sensitivity modular gamma camera. Subsequently, 99mTc-macroagreggated albumin (37 MBq, diam=10-40 μm) was injected i.v., imaged, and used to define a lung region-of-interest. The lung to background ratio was used as a measure of lung uptake. Results Hyperoxia exposure resulted in a 74% increase in 99mTc-HMPAO lung uptake, which peaked at 7 days and persisted for the 21 days of exposure. 99mTc-duramycin lung uptake was also maximal at 7 days of exposure but decreased to near control levels by 21 days. The sustained elevation of 99mTc-HMPAO uptake suggests ongoing changes in lung redox status whereas cell death appears to have subsided by 21 days. Conclusion These results suggest the potential use of 99mTc-HMPAO and 99mTc-duramycin as redox and cell-death imaging biomarkers, respectively, for in vivo identification and assessment of different stages of lung injury.

In vivo detection of hyperoxia-induced pulmonary endothelial cell death using 99mTc-Duramycin

“…Duramycin (3,035 g/mole MW) kits were prepared as previously described [12], and technetium-labeled macroaggregated albumin ( 99m Tc-MAA, particle sizes 20 – 40 μm) was purchased from Cardinal Health (Wauwatosa, WI). Antibodies to cleaved caspase 3 (Biocare, #CP229B), CD31 (Acris, #AP15436PU-N), keratin 7 (Abcam, #9021), and myeloperoxidase (Abcam, #45977) were used with appropriate secondary antibodies (Jackson Immuno #711-066-152 for CD31, Jackson Immuno #715-066-151 for keratin 7, and Abcam #6829 HRP for MPO) to identify endpoints of interest.…”

“…These sample sizes achieve a power > 85% for detecting changes in lung uptake of DU greater than 15% based on previously published results with DU [12] (ANOVA power; SigmaPlot version 12.0).…”

“…There is strong evidence that the pulmonary capillary endothelium is a primary and early site of oxygen toxicity injury [7,9,10]. This and the large pulmonary capillary surface area in direct apposition with blood-borne compounds suggest the utility of biomarker imaging for detecting early lung endothelial injury due to hyperoxic ALI [11,12]. …”

“…99m Tc-duramycin, DU, is a new SPECT biomarker sensing cell death via apoptosis and/or necrosis [12-14]. DU serves as a molecular probe that binds to phosphatidylethanolamine (PE), which has little presence on the surface of normal viable cells, but becomes exposed onto the cell surface and/or accessible to the extracellular milieu with apoptosis and necrosis, respectively [13,14].…”

“…DU serves as a molecular probe that binds to phosphatidylethanolamine (PE), which has little presence on the surface of normal viable cells, but becomes exposed onto the cell surface and/or accessible to the extracellular milieu with apoptosis and necrosis, respectively [13,14]. Recently we reported an increase in the lung uptake of DU in rats exposed to sublethal 85% O 2 [12]. The objective of the present study is to determine the extent of DU lung uptake in rats injured with >98% O 2 exposure for up to 60 hours, and to correlate that uptake with capillary endothelial cell apoptosis detected histologically.…”