Peroxynitrite-modified 99mTc-beta-VLDL: tissue distribution and plasma clearance rate.

Free radicals superoxide (O(2)(-)) and nitric oxide (*NO) are generated by blood vessels and can rapidly react to produce a peroxynitrite anion (ONOO(-)), a powerful oxidant that modifies lipoproteins making them more atherogenic. The aim of this study was to investigate the effect of peroxynitrite-induced modifications on beta-very-low-density lipoprotein (beta-VLDL) as to its biodistribution and plasma clearance rate, as well as the uptake of these particles by THP-1 cells. After being injected into New Zealand White rabbits, the peroxynitrite-modified beta-VLDL (99mTc-per-beta-VLDL) was cleared from circulation faster than the native beta-VLDL (99mTc-nat-beta-VLDL) in both normocholesterolemic rabbits (NC) and in hypercholesterolemic rabbits (HC). In HC rabbits, the fractional clearance of 99mTc-labeled beta-VLDL was significantly lower than in NC rabbits. The in vivo studies showed that accumulation of 99mTc-labeled beta-VLDL, expressed per gram of tissue, followed the decreasing order: kidney > liver > spleen > adrenal gland >or= lung > aortic arch > heart >or= abdominal aorta > thoracic aorta > psoas muscle. The high accumulation in the kidneys suggests the processing of 99mTc-labeled apolipoproteins by receptors present in kidney cells. The accumulation of 99mTc-nat-beta-VLDL in the whole organ was the following: liver > kidney > heart > spleen > adrenal gland > aorta in HC and NC rabbits. The uptake of 99mTc-per-beta-VLDL by the spleen was greater than the uptake by the heart in both groups. The in vitro studies showed that the uptake of 99mTc-per-beta-VLDL by THP-1 cells was higher than that of 99mTc-nat-beta-VLDL. These results show that peroxynitrite-modified beta-VLDL is rapidly removed from plasma and accumulates in several tissues, mainly in the liver and kidney. This may be particularly important in hypercholesterolemic situations that could favor the accumulation of native and peroxynitrite-modified beta-VLDL in several tissues.

Myocardial reperfusion: leukocyte accumulation in the ischemic and remote non-ischemic regions.

Neutrophil accumulation in the first hour of myocardial reperfusion was assessed in dog hearts submitted to ischemia with and without necrosis. In anesthetized dogs, the left anterior descending coronary artery was occluded for 20 min (group IS-20 n = 7) and for 90 min (group IS-90 n = 6). Immediately after reperfusion, 99m Tc-Ceretec (Exametazime-Amersham) labeled neutrophils were injected into a central vein and 60 min later the dogs were killed. The left ventricle was isolated, weighed, and sliced. Six sections, 3 from normal and 3 from reperfused regions, were divided into endocardial and epicardial layers. Myocardial and blood radiometry were used to evaluate the neutrophil accumulation during reperfusion. The differences between leukocyte accumulation in both groups were assessed comparing the ischemic/normal relations in the endocardial and epicardial layers. A second comparison considered myocardium/blood relations to allow the evaluation of differences between remote normal myocardial areas of the two experimental groups. In dogs submitted to 20 min of ischemia, leukocytes accumulated significantly more (P < 0.01) in the reperfused myocardium as compared with the non-ischemic region. The increase occurred both in the endocardial (1.49+/-0.20) and epicardial (1.48+/-0.29) regions. After 90 min ischemia, leukocyte accumulation was more intense and predominant in endocardium where there was a 4-fold (3.97+/-1.28) increase over the non-ischemic region, while in the epicardium this relation was only 2.5-fold (2.56+/-0.98). In the remote non-ischemic myocardium, leukocyte accumulation was greater in dogs submitted to 90 min of ischemia compared to the 20 min group (P < 0.01), without distinction between endocardial and epicardial layers. This accumulation in territories of non-culprit coronary arteries may be related to the blood flow abnormalities and matrix structure changes that occur in these regions during the development of an acute myocardial infarction and its natural repair.

Hemodynamic and metabolic effects of CO2 pneumoperitoneum in an experimental model of hemorrhagic shock due to retroperitoneal hematoma.

BACKGROUND: Diagnostic laparoscopy has been used in abdominal trauma patients, although its role is not well defined. The safety of laparoscopic evaluation in trauma patients with severe intraabdominal hemorrhage has not yet been analyzed. The purpose of this study is to evaluate the hemodynamic and metabolic effects of CO2 pneumoperitoneum (COI) in hemorrhaged animals through a retroperitoneal hematoma (RH). METHODS: Twenty-two 15-20-kg mongrel dogs were monitored for systemic and pulmonary hemodynamics, inferior vena cava pressure, and arterial blood gases. After 1 h of baseline, all animals were submitted to a RH. After 45 min the dogs were randomized into two groups. Control (CTR): dogs were submitted only to a RH; pneumoperitoneum (PN): dogs were submitted to a RH and 45 min later they were insufflated to an intraabdominal pressure of 10 mmHg with medical-grade CO2 gas for 30 min. Echocardiography was performed, only in PN animals, at baseline, 45 and 60 min after RH. RESULTS: RH induced a shock condition with low, sustained levels of arterial pressure, cardiac index, left ventricular stroke index, base excess, and oxygen delivery which were further depressed following COI. Three deaths occurred in the PN group, all of them toward the end of COI. During COI, hypercapnia was observed in one animal. COI did not impair systolic function or ejection fraction. CONCLUSIONS: COI with an IAP of 10 mmHg may be deleterious in animals with hemorrhagic shock due to an intraabdominal lesion. These findings could be clinically significant in abdominal trauma patients.

Plasma clearance and biodistribution of oxidatively modified 99mTc-beta-VLDL in rabbits.

The biodistribution and removal from plasma (measured as fractional clearance rate, FCR, per hour) of native and oxidatively modified 99mtechnetium-labeled beta-very low density lipoprotein (99mTc-beta-VLDL) were investigated in hypercholesterolemic (HC) and control (C) three-month old New Zealand rabbits. The intracellular accumulation of beta-VLDL labeled with 99mTc was studied in vitro in THP-1 cells and monocyte-derived macrophages isolated from rabbits. After intravenous injection into C rabbits, copper-oxidized beta-VLDL (99mTc-ox-beta-VLDL) was cleared from the circulation faster (0.362 +/- 0.070/h) than native beta-VLDL (99mTc-nat-beta-VLDL, 0.241 +/- 0.070/h). In contrast, the FCR of 99mTc-ox-beta-VLDL in HC rabbits was lower (0.100 +/- 0.048/h) than that of 99mTc-nat-beta-VLDL (0.163 +/- 0.043/h). The hepatic uptake of radiolabeled lipoproteins was lower in HC rabbits (0.114 +/- 0.071% injected dose/g tissue for 99mTc-nat-beta-VLDL and 0.116 +/- 0.057% injected dose/g tissue for 99mTc-ox-beta-VLDL) than in C rabbits (0.301 +/- 0.113% injected dose/g tissue for 99mTc-nat-beta-VLDL and 0.305 +/- 0.149% injected dose/g tissue for 99mTc-ox-beta-VLDL). The uptake of 99mTc-nat-beta-VLDL and 99mTc-ox-beta-VLDL by atherosclerotic aorta lesions isolated from HC rabbits (99mTc-nat-beta-VLDL: 0.033 +/- 0.012% injected dose/g tissue and 99mTc-ox-beta-VLDL: 0.039 +/- 0.017% injected dose/g tissue) was higher in comparison to that of non-atherosclerotic aortas from C rabbits (99mTc-nat-beta-VLDL: 0.023 +/- 0.010% injected dose/g tissue and 99mTc-ox-beta-VLDL: 0.019 +/- 0.010% injected dose/g tissue). However, 99mTc-nat-beta-VLDL and 99mTc-ox-beta-VLDL were taken up by atherosclerotic lesions at similar rates. In vitro studies showed that both monocyte-derived macrophages isolated from rabbits and THP-1 macrophages significantly internalized more 99mTc-ox-beta-VLDL than 99mTc-nat-beta-VLDL. These results indicate that in cholesterol-fed rabbits 99mTc-ox-beta-VLDL is slowly cleared from plasma and accumulates in atherosclerotic lesions. However, although the extent of in vitro uptake of 99mTc-ox-beta-VLDL by macrophages was high, the in vivo accumulation of this radiolabeled lipoprotein by atherosclerotic lesions did not differ from that of 99mTc-nat-beta-VLDL.

Plasma clearance and biodistribution of oxidatively modified 99mTc-Beta-VLDL in rabbits

The biodistribution and removal from plasma (measured as fractional clerance rate, FCR, per hour) of native and oxidatively modified (99m)technetium-labeled Beta-very low density lipoprotein ((99m)Tc-Beta-VLDL)) were investigated in hypercholesterolemic (HC) and control (C) three-month old New Zealand rabbits. The intracellular accumulation of Beta-VLDL labeled with (99m)Tc was studied in vitro in THP-1 cells and monocyte-derived macrophages isolated from rabbits. After intravenous injection into C rabbits, copper-oxidized Beta-VLDL ((99m)Tc-ox-Beta-VLDL)) was cleared from the circulation faster (0.362 + 0.070/h) than native Beta-VLDL ((99m)Tc-nat-Beta-VLDL, 0.241 + 0.070/h)). In contrast, the FCR of (99m)Tc-ox-Beta-VLDL in HC rabbits was lower (0.100 + 0.048/h) than that of (99m)Tc-nat-Beta-VLDL (0.163 + 0.043/h). The hepatic uptake of radiolabeled lipoproteins was lower in HC rabbits (0.114 + 0.071 percent injected dose/g tissue for (99m)Tc-nat-Beta-VLDL and 0.116 + 0.057 percent injected dose/g tissue for (99m) Tc-ox-Beta-VLDL) than in C rabbits (0.301 + 0.113 percent injected dose/g tissue for (99m)Tc-nat-Beta-VLDL and 0.305 + 0.149 percent injected dose/g tissue for ((99m)Tc-ox-Beta-VLDL). The uptake of (99m)Tc-nat-Beta-VLDL and (99m)Tc-ox-Beta-VLDL by atherosclerotic aorta lesions isolated from HC rabbits ((99m)Tc-nat-Beta-VLDL:0.033 + 0.012 percent injected dose/g tissue and (99m)Tc-ox-Beta-VLDL: 0.039 + 0.017 percent injected dose/g tissue) was higher in comparison to that of non-atherosclerotic aortas from C rabbits (99m)Tc-nat-Beta-VLDL: 0.023 + 0.010 percent injected dose/g tissue and (99m)Tc-ox-Beta VLDL: 0.019 + 0.010 percent injected dose/g tissue). However, (99m) Tc-nat-Beta-VLDL and (99m)Tc-ox-Beta-VLDL were taken up by atherosclerotic lesions at similar rates. In vitro studies showed that both monocyte-derived macrophages isolated from rabbits and THP-1 macrophages significantly internalized more (99m)Tc-ox-Beta-VLDL than (99m)Tc-nat-Beta-VLDL. These results indicate that in cholesterol-fed rabbits (99m)Tc-ox-Beta-VLDL is slowly cleared from plasma and accumulates in atherosclerotic lesions. However, although the extent of in vitro uptake of (99m)Tc-ox-Beta-VLDL by macrophages was high, the in vivo accumulation of this radiolabeled lipoprotein by atherosclerotic lesions did not differ from that of (99m)Tc-nat-Beta-VLDL.

Distribution of pleural injectate. Effect of volume of injectate and animal rotation.

It is controversial whether rotation is necessary for patients undergoing pleurodesis. In addition, the optimal volume of the injectate remains to be determined. The purpose of this study was to determine the importance of rotation and the volume of the agent on the intrapleural dispersion of agents injected into the pleural space of rabbits. Technetium 99m pertechnetate (99mTc) in 0.5, 1.0, or 2.0 ml of saline solution was injected into ten lightly anesthetized rabbits, half of whom were rotated for 1 min after the injection. Static images were obtained in the anterior projection 1 and 5 min after the injection. After the second scan, the limits of the lung were defined by obtaining a perfusion scan immediately after the intravenous injection of macroaggregates of 99mTc-labeled serum albumin. The degree of dispersion was significantly greater in the nonrotated groups both at 1 min (F = 8.11, p = 0.0085) and at 5 min (F = 5.89, p = 0.0274). In addition, the homogeneity of the distribution of the injectate was not improved with rotation. From this study, we conclude that rotation of the animal for 1 min after the intrapleural injection does not improve the distribution of the injectate throughout the pleural space. Furthermore, a volume of 0.5 ml is sufficient for all pleural surfaces to be exposed.

Plasma kinetics and biodistribution of a lipid emulsion resembling low-density lipoprotein in patients with acute leukemia.

Low-density lipoprotein (LDL) could be used as a carrier of chemotherapeutic agents to neoplastic cells that overexpress LDL receptors (rLDL), but LDL is difficult to obtain and handle. Recently, it was observed that a protein-free emulsion resembling the lipid portion of LDL (LDE) behave like native LDL when injected into the bloodstream. In this study, the evidence that LDE is taken up by rLDL was expanded by comparing LDL and LDE plasma decay curves in rabbits and by competition experiments with lymphocytes. To verify whether LDE could be removed from the plasma by neoplastic cells with increased rLDL, LDE labeled with 14Ccholesteryl ester was injected into 14 patients with acute myeloid leukemia (AML) and into 7 with acute lymphocytic leukemia (ALL). In AML rLDL expression is increased but in ALL it is normal. LDE plasma fractional clearance rate (FCR, in h-1) was calculated from the remaining radioactivity measured in plasma samples collected during 24 h following injection. LDE FCR was 3-fold greater in AML than in ALL patients 0.192 +/- 0.210 (SD) and 0.066 +/- 0.033 h-1, respectively, P < 0.035. When LDE injection was repeated in 9 AML patients in hematological remission, LDE FCR diminished 66% compared to the pretreatment values (from 0.192 +/- 0.210 to 0.065 +/- 0.038 h-1, P < 0.02), so that it could be estimated that nearly 66% of the emulsion was taken up by AML cells and only 34% by the normal tissues. As expected, LDE FCR was unchanged in 4 patients with ALL in hematological remission (0.069 +/- 0.044 h-1). Gamma camera images obtained 6 h after the injection of 99mTc-label LDE into one patient with ALL showed biodistribution similar to that of LDL. In one AML patient LDE was comparatively more concentrated over the areas corresponding to the bone marrow infiltrated by AML cells. Our results indicate that LDE FCR is increased in a disease known to contain malignant cells that overexpress rLDL, suggesting that LDE is taken up by malignant cells with increased rLDL.

A quantitative analysis of transcapillary refill in severe hemorrhagic hypotension in dogs.

In pressure-driven hemorrhage (PDH), where the rate of bleeding is a function of prevailing arterial pressure, survival time, arterial pressure, cardiac output, oxygen consumption, and base excess are functions of initial bleeding rate. The quantitative rate of transcapillary refill (TR) throughout PDH leading to death was determined in splenectomized dogs, through serial analysis of Cr51-tagged red cell dilution. Mild, moderate, and severe levels of PDH were produced by varying initial bleeding rate (10, 25, and 50 ml/min, respectively). The rate of TR is a function of the severity of PDH, but does not correlate with arterial pressure, cardiac output, or systemic resistance. The volume of transferred fluid represents an ever increasing fraction of total plasma volume, and accounts for more than 75% of plasma volume in preterminal stages of shock. TR sustains a relatively fixed level of plasma volume, equivalent to two-third of the initial plasma volume, irrespective of the rate of bleeding. Hypertonic NaCl (7.5%) enhances TR, while isotonic NaCl reverses it.