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Biomedical subjects

Marcos Intaglietta

Publications and source records attributed to Marcos Intaglietta.

At least 55 records · Page 3Linked to original sources

Aggregate formation of erythrocytes in postcapillary venules.

The purpose of the present study was to obtain information on erythrocyte aggregate formation in vivo. The movements of erythrocytes in postcapillary venules of the rat spinotrapezius muscle at various flow rates were recorded with a high-speed video camera before and after infusion of dextran 500. To distinguish aggregates, the following criteria were used: 1) a fixed distance (4 microm) between the center points of two adjacent cells, 2) lack of visible separation between the adjacent cells, and 3) movement of the adjacent cells in the same direction. Without dextran 500 infusion, 11 and 5% of erythrocytes formed aggregates in low (33.2 +/- 28.3 s) and high pseudoshear (144.2 +/- 58.3 s) conditions, respectively, based on the above criteria. After dextran 500 infusion, 53% of erythrocytes satisfied the criteria in the low pseudoshear condition (26.5 +/- 17.0 s) and 13% of erythrocytes met the criteria in the high pseudoshear condition (240.0 +/- 85.9 s), indicating erythrocyte aggregation is strongly associated with shear rate. Approximately 90% of aggregate formation occurred in a short time period (0.15-0.30 s after entering the venule) in a region 15 to 30 microm from the entrance. The time delay may reflect rheological entrance conditions in the venule.

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Increased tissue PO2 and decreased O2 delivery and consumption after 80% exchange transfusion with polymerized hemoglobin.

The O2-carrying blood substitute based on polymerized bovine hemoglobin (PBH) was used to determine efficacy in maintaining tissue Po2 after an 80% isovolemic blood exchange leading to a hematocrit of 19% [5.4 g Hb/dl from red blood cells (RBCs) and 6.3 g Hb/dl from PBH]. Effects were studied in terms of O2 delivery, O2 extraction, and tissue Po2 at the microcirculatory level at 1, 12, and 24 h after exchange transfusion in awake hamsters prepared with a window chamber model. At 1 h after exchange, arteriolar and venular diameters were decreased compared with baseline. Arteriolar diameter did not fully recover at 12 h after exchange, but venular diameter returned to normal. At 24 h after exchange, arteriolar and venular diameters were not different from baseline. Combining diameter and flow velocity data allowed us to calculate arteriolar and venular flows. At 1 h after exchange, arteriolar and venular flow was reduced compared with baseline. Arteriolar flow was lower at 12 h after exchange and recovered after 24 h. The number of capillaries with RBC passage [functional capillary density (FCD)] at 1 h after exchange with PBH was significantly lower than baseline. FCD remained decreased at 12 h; at 24 h after exchange transfusion, FCD was fully recovered. Tissue Po2 was maximal at 1 h after exchange and decreased progressively at 12 and 24 h after exchange. O2 release to the tissue was minimal at 1 h and increased at 12 and 24 h after exchange. These results suggest the impairment of tissue O2 metabolism after introduction of PBH into the circulation, which is mitigated as PBH concentration declines.

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Microvascular PO2 during extreme hemodilution with hemoglobin site specifically PEGylated at Cys-93(beta) in hamster window chamber.

The oxygen transport capacity of nonhypertensive polyethylene glycol (PEG)-conjugated hemoglobin solutions were investigated in the hamster chamber window model. Microvascular measurements were made to determine oxygen delivery in conditions of extreme hemodilution [hematocrit (Hct) 11%]. Two isovolemic hemodilution steps were performed with a 6% Dextran 70 (70-kDa molecular mass) plasma expander until Hct was 35% of control. Isovolemic blood volume exchange was continued using two surface-modified PEGylated hemoglobins (P5K2, P(50) = 8.6, and P10K2, P(50) = 8.3; P(50) is the hemoglobin Po(2) corresponding to its 50% oxygen saturation) until Hct was 11%. P5K2 and P10K2 are PEG-conjugated hemoglobins that maintain most of the hemoglobin allosteric properties and have a cooperativity index of n = 2.2. The effects of these molecular solutions were compared with those obtained in a previous study using MP4, a PEG-modified hemoglobin whose P(50) was 5.4 and cooperativity was 1.2 (Tsai et al., Am J Physiol Heart Circ Physiol 285: H1411-H1419, 2003). Tissue oxygen levels were higher after P5K2 (7.0 +/- 2.5 mmHg) and P10K2 (6.3 +/- 2.3 mmHg) versus MP4 (1.7 +/- 0.5 mmHg) or the nonoxygen carrier Dextran 70 (1.3 +/- 1.2 mmHg). Microvascular oxygen delivery was higher after P5K2 and P10K2 (2.22 and 2.34 ml O(2)/dl blood) compared with MP4 (1.41 ml O(2)/dl blood) or Dextran 70 (0.90 ml O(2)/dl blood); however, all these values were lower than control (7.42 ml O(2)/dl blood). The total hemoglobin in blood was similar in all cases; therefore, the improvement in tissue Po(2) and oxygen delivery appears to be due to the increased cooperativity of the new molecules.

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Microvascular pressure and functional capillary density in extreme hemodilution with low- and high-viscosity dextran and a low-viscosity Hb-based O2 carrier.

Blood losses are usually corrected initially by the restitution of volume with plasma expanders and subsequently by the restoration of oxygen-carrying capacity using either a blood transfusion or possibly, in the near future, oxygen-carrying plasma expanders. The present study was carried out to test the hypothesis that high-plasma viscosity hemodilution maintains perfused functional capillary density (FCD) by preserving capillary pressure. Microvascular pressure responses to extreme hemodilution with low- (LV) and high-viscosity (HV) plasma expanders and an exchange transfusion with a polymerized bovine cell-free Hb (PBH) solution were analyzed in the awake hamster window chamber model (n = 26). Systemic hematocrit was reduced from 50% to 11%. PBH produced a greater mean arterial blood pressure than the nonoxygen carriers. FCD was higher after a HV plasma expander (70 +/- 15%) vs. PBH (47 +/- 12%). Microvascular pressure spanning the capillary network was higher after a HV plasma expander (16-19 mmHg) compared with PBH (12-16 mmHg) and a LV plasma expander (11-14 mmHg) but lower than control (22-26 mmHg). FCD was found to be directly proportional to capillary pressure. The use of a HV plasma expander in extreme hemodilution maintained the number of perfused capillaries and tissue perfusion by comparison with a LV plasma expander due to increased mean arterial blood pressure and capillary pressure. The use of PBH increased mean arterial pressure but reduced capillary pressure due to vasoconstriction and did not maintain FCD.

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Oxygen distribution and respiration by the microcirculation.

Longitudinal and radial oxygen gradients in the microcirculation due to oxygen release from arterioles show that in some tissues oxygen is primarily supplied by arterioles and secondarily by capillaries. In several tissues, the arteriolar rate of oxygen exit is too large to be explained by diffusion alone, indicating that in these tissues oxygen consumption of the arteriolar wall in vivo is much greater than that shown in in vitro studies of endothelium and vascular smooth muscle, a phenomenon that may be related to the synthesis autocoids by the endothelium in vivo. The functional significance of the high metabolic rate of the arteriolar vessels may be related to the need of providing a metabolic barrier for protecting the parenchymal tissue from high oxygen levels in arterial blood, thus reducing formation of oxygen free radicals in the perivascular tissue, a supposition supported by the finding that the radial oxygen gradient at the microvascular wall and therefore its rate of oxygen consumption are proportional to local blood oxygen partial pressure (pO(2)). Oxygen consumption by the endothelium and/or smooth muscle is also a factor in causing terminal lymphatic pO(2) to have the lowest oxygen level in the tissue, rendering this compartment most vulnerable in hypoxic conditions.

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Improving microcirculation is more effective than substitution of red blood cells to correct metabolic disorder in experimental hemorrhagic shock.

Microcirculatory perfusion deficits and impaired tissue oxygenation in nonvital organs frequently occur after hemorrhage and they contribute to potentially lethal complications. The aim of this study was to test the influence of colloid osmotic pressure, viscosity, and red blood cell (RBC) content of the resuscitative fluid on metabolic disorder, perfusion, and oxygenation in peripheral tissues. Awake hamsters were subjected to hemorrhage of 50% and were resuscitated with 25% of blood volume with solutions containing 6% pegylated bovine albumin only (PEG-BSA 0) and 6% PEG-BSA mixed with autologous RBCs to reach 4 g/dL (PEG-BSA 4) and 8 g/dL (PEG-BSA 8) of hemoglobin. PEG-BSA had a viscosity of 4.2 cP and a COP of 116 mmHg. Microhemodynamics and tissue pO2 were assessed in the hamster chamber window preparation with intravital microscopy. Arterial base excess tended to be lower than baseline for PEG-BSA 0 and PEG-BSA 4 (ns), whereas base deficit remained significantly decreased for PEG-BSA 8 (P<0.05 vs. baseline). Oxygen extraction was 91% +/- 2% of the oxygen delivery for PEG-BSA 0 compared with 85% +/- 2% for PEG-BSA 8 (P<0.05). Functional capillary density was 61%, 47%, and 45% for PEG-BSA 0 (P<0.05 vs. other groups), PEG-BSA 4 and PEG-BSA 8, respectively. We conclude that arterial base excess and oxygen extraction ratio in the tissue was better restored if a higher fraction of PEG-BSA and less RBCs were infused. This was attributed to a more homogeneous distribution of oxygen, as reflected by functional capillary density. Our results suggest that the transfusion trigger in hemorrhagic shock may be shifted toward lower hemoglobin concentrations if highly viscous and oncotic solutions are used.

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Resuscitation from hemorrhagic shock with MalPEG-albumin: comparison with MalPEG-hemoglobin.

Our aim was to determine the efficacy of polyethylene glycol-conjugated human albumin (MalPEG-Alb) in restoring circulatory volume after 1 h of hemorrhagic shock. Experiments were performed in the awake condition in the hamster skin fold preparation. Microhemodynamic parameters and tissue Po2 were assessed with intravital microscopy and the use of the phosphorescence quenching technique. One hour after shock induction by withdrawal of 50% of the blood volume, animals were resuscitated with MalPEG-Alb (n = 6). Systemic and microhemodynamic parameters following resuscitation were identical to those obtained with the same protocol using MalPEG-Hb (1). However, parameters related to microvascular oxygen distribution were significantly lower in the MalPEG-Alb group compared with the previous data from the MalPEG-Hb group in that tissue oxygen partial pressure was 5 +/- 2 mmHg (vs. 8 +/- 3 mmHg, P < 0.05), oxygen delivery was reduced to 60 +/- 27% (P < 0.05), and oxygen consumption was reduced to 69 +/- 28% (P < 0.05). Both molecules were matched in composition (4.2 g/dL) and surface chemistry. MalPEG-Alb colloid osmotic pressure was 37 mmHg (vs. 49 mmHg for MalPEG-Hb), and viscosity was 2.7 cP (vs. 2.5 cP for MalPEG-Hb). The present results show that both solutions are efficacious plasma expanders and that the hemoglobin-based solution provides improved oxygen distribution and tissue Po2 in the hamster chamber model.

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Hyperosmotic-hyperoncotic versus hyperosmotic-hyperviscous: small volume resuscitation in hemorrhagic shock.

The aim of this study was to test the effects of using a high-viscosity fluid after small-volume hyperosmotic resuscitation from hemorrhagic shock and to compare this to hyperosmotic followed by hyperoncotic resuscitation. Studies were made in the awake hamster window chamber preparation with the animals subjected to hemorrhage of 50% of blood volume and resuscitated with a small volume of a 7.5% NaCl solution, which was followed within minutes by infusion of 25% of withdrawn volume of either 0.7% or 0.8% alginate solutions (A0.7%, 7.6 cp; and A0.8%, 10.2 cp) or 5% hydroxyethyl starch (HES 5%, 2.1 cp). All modalities of resuscitation returned blood pressure to near baseline values in 5 min, which remained elevated after 90 min with A0.7% and A0.8% but returned to near shock values in 15 min with HES 5%. Microvascular flow and functional capillary density (FCD) followed the same pattern, being significantly higher for the alginate solutions than HES 5% after 90 min. Plasma viscosity 90 min after resuscitation was 2.1 and 2.6 cp for A0.7% and A0.8%, respectively, and 1.1 cP for HES 5%. There was an apparent directly proportional relationship between the concentration of alginate and blood pressure recovery, with blood pressure near normal with A0.8%, and approximately 20 mmHg lower with A0.7%. The recovery of microvascular flow and FCD, although showing a trend toward being more effective with A0.8%, was not significantly different from A0.7% but statistically different and improved relative to HES 5%. The high-viscosity fluids provide a novel small-volume method of resuscitation that maximizes microvascular perfusion for extended periods until surgical control of bleeding is possible. Results show that high-plasma-viscosity resuscitation provides a more consistent and prolonged resuscitation than hyperoncotic treatment. The increase in viscosity presents a gradual recovery in blood pressure and may be used as an alternative for small-volume hypotensive resuscitation, increasing tissue perfusion while potentially limiting hemorrhage in vascular injuries of the major blood vessels.

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Microvascular perfusion upon exchange transfusion with stored red blood cells in normovolemic anemic conditions.

BACKGROUND: Transfusions are intended to augment oxygen-carrying capacity. The ability of fresh and stored red blood cells (RBCs) to maintain microvascular perfusion and oxygen delivery to the tissue has not been directly measured. STUDY DESIGN AND METHODS: Microvascular responses to exchange transfusion with fresh and stored RBCs after acute isovolemic hemodilution with a plasma expander were investigated with the hamster window chamber model. In-vivo functional capillary density (FCD), blood flow, and high-resolution oxygen distribution in microvascular networks were measured by noninvasive methods. RESULTS: Exchange transfusion with an RBC suspension after a 60 percent isovolemic hemodilution with dextran 70 (6% MW = 70 kDa) resulted in a hematocrit of 18 percent (5.6 +/- 0.2 g/dL hemoglobin [Hb]). All other systemic variables were unchanged. Stored RBCs (28 days in citrate-phosphate-dextrose-adenine-1) resuspended in fresh frozen plasma matched to the Hct and Hb concentration were exchange transfused until 25 percent of the circulating RBCs were stored RBCs. Stored RBCs reduced microvascular flow and FCD by 63 and 54 percent, respectively, of the level achieved when fresh RBCs were exchange transfused. Microvascular oxygen extraction by the stored RBC was 54 percent lower than that of the fresh RBCs. The tissue oxygen levels were 3.5 and 14.4 mmHg for the stored and fresh RBCs, respectively. CONCLUSION: Circulation of stored RBCs in a hemodiluted animal resulted in significantly malperfused and underoxygenated microvasculature that was not detectable at the systemic level.

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Oxygen delivery and consumption in the microcirculation after extreme hemodilution with perfluorocarbons.

The oxygen transport capacity of fluorocarbons was investigated in the hamster chamber window model microcirculation to determine the rate at which oxygen is delivered to the tissue in conditions of extreme hemodilution [hematocrit (Hct) 11%]. Hydroxyethlyl starch (HES 200; 200 kDa molecular mass) was used as a plasma expander for two isovolemic hemodilutions performed with 10% HES 200 until a Hct of 65%. A third step reduced the Hct to 75% of baseline and was performed with either HES 200 or a 60% perfluorocarbon (PFC) emulsion. Comparisons of HES 200-only-hemodiluted animals versus 4.2 g/kg PFC emulsion-hemodiluted animals were made at 21% and 100% normobaric oxygen ventilation. It was found that systemic and microvascular oxygen delivery was 25% and 400% higher in the PFC animals compared with HES 200 animals, respectively, showing that PFCs deliver oxygen to the tissue when combined with hyperoxic ventilation in the present experiments, with no evidence of vasoconstriction or impaired microvascular function. Oxygen ventilation (100%) led to a positive base excess for the PFC group (5.5 +/- 2.5 mmol/l) versus a negative balance (-0.8 +/- 1.4 mmol/l) for the HES 200 group, suggesting that microvascular findings corresponded to systemic events.

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Oxygen-carrying blood substitutes: a microvascular perspective.

Development of a viable blood substitute began by focusing on recreating the oxygen-carrying capacity of blood, leading to the recognition that haemoglobin (Hb) is presently unequalled for this function. However, as human Hb is the only realistic source of this protein, the production of a blood substitute that solves transfusional blood availability problems and shortages must introduce a multiplying factor between supply of natural blood and blood substitute, while maintaining equivalency of function/efficacy. In other words, a unit of blood should produce several units of equivalent blood substitute. This expansion is now possible because of new understanding of how blood delivers oxygen in the microcirculation and the consequences of reducing oxygen-carrying capacity in haemorrhage. This information is used to provide improved resuscitation capacity and maintenance of tissue metabolism by tailoring the properties of a blood substitute to the task of maintaining microvascular function, rather than oxygen delivery capacity. Resuscitation in an organism that is haemorrhaging requires maintenance perfusion, a process directly linked to the maintenance of adequate levels of shear stress on the endothelium, induced by either increased blood/plasma viscosity or increased blood flow velocity in the microcirculation. This process must also be intimately coupled with the requirement that no portion of the tissue is anoxic. This disparate set of requirements can be satisfied with high viscosity Hb solutions that have high affinity for oxygen, a combination of properties that causes the microcirculation to remain functional, and a requirement that supersedes restoration of oxygen-carrying capacity in the treatment of haemorrhage.

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Activated clotting time and heparin administration in Sprague-Dawley rats and Syrian golden hamsters.

To determine the activated clotting time (ACT) in rats and hamsters from our colony and to evaluate the response of this parameter to different heparin doses in these species, ACTs were measured using a Medtronic HemoTec ACT measurement system in samples obtained by intracardiac puncture from normal, nonanticoagulated, anesthetized rats and hamsters. Another groups of animals received different intravenous boluses of heparin to determine the dose needed to maintain ACT values > 480 sec for at least 30 min. The ACT (mean +/- SEM) was 48.0 +/- 2.17 sec for the 50 rats sampled and 42.5 +/- 2.35 sec for the 48 hamsters. Rats required a bolus of 1200 IU/kg intravenous heparin to maintain an ACT > 480 sec for 30 min; hamsters required 1000 IU/kg heparin for the same effect. We concluded that compared with humans, rats and hamsters from our colony have short ACTs and low sensitivity to heparin, in terms of the dose needed to reach a target ACT as well as the time required to sustain it. Further the ACT values in these animals showed great variability.

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Microlymphatic and tissue oxygen tension in the rat mesentery.

Oxygen phosphorescence quenching was used to measure tissue Po(2) of lymphatic vessels of 43.6 +/- 23.1 microm (mean +/- SD) diameter in tissue locations of the rat mesentery classified according to anatomic location. Lymph and adipose tissue Po(2) were 20.6 +/- 9.1 and 34.1 +/- 7.8 mmHg, respectively, with the difference being statistically significant. Rare microlymphatic vessels in connective tissue not surrounded by microvessels had a Po(2) of 0.8 +/- 0.2 mmHg, whereas the surrounding tissue Po(2) was 3.0 +/- 3.2 mmHg, with both values being significantly lower than those of adipose tissue. Lower of lymph fluid Po(2) relative to the surrounding tissue was also evident in paired measurements of Po(2) in the lymphatic vessels and perilymphatic adipose tissue, which was significantly lower than the Po(2) in paired adipose tissue. The Po(2) of the lymphatic fluid of the mesenteric microlymphatics is consistently lower than that of the surrounding adipose tissue by approximately 11 mmHg; therefore, lymph fluid has the lowest Po(2) of this tissue. The disparity between lymph and tissue Po(2) is attributed to the microlymphatic vessel wall and lymphocyte oxygen consumption.

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Radial displacement of red blood cells during hemodilution and the effect on arteriolar oxygen profile.

In this study, we assessed the magnitude of the erratic deviations in the radial position of red blood cells (RBCs) in the laminar flow regime of arterioles in a hamster window preparation and the intraluminal Po(2) profile to determine whether this variability affects the intraluminal distribution of oxygen in conditions of normal hematocrit and hemodilution. A gated image intensifier was used to visualize fluorescently labeled RBCs in tracer quantities and obtain multiple measurements of RBC radial and longitudinal positions at time intervals on the order of 5 ms within single arterioles (diameter range 40-95 microm). RBCs in the velocity range of 0.3-14 mm/s exhibit a mean coefficient of variation of velocity of 16.9 +/- 10.5% and a SD of the radial position of 1.98 +/- 0.98 microm. Both quantities were inversely related to hematocrit, and the former was significantly lowered by hemodilution. Our experimental results presented very similar values and shape compared with the intraluminal oxygen profile derived theoretically for normal hematocrit, suggesting that shear-augmented diffusion due to the measured radial displacement of RBCs did not significantly affect oxygen diffusion from blood into the arteriolar vessel wall. Po(2) profiles in the arterioles assumed an increasingly parabolic configuration with increasing levels of hemodilution.

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Relationship between erythrocyte aggregate size and flow rate in skeletal muscle venules.

In previous studies we showed that intravenous infusion of Dextran 500 in the rat causes blunting of the velocity profile of red blood cells in venules at low shear rates. To determine whether this blunting is associated with the formation of red blood cell aggregates, we measured the length and width of particles in the venular flow stream at systemic hematocrits up to 20% with a high-speed video camera and a new image analysis technique. Data were obtained at various shear rates under normal (nonaggregating) conditions as well as after infusion of Dextran 500. Under normal conditions, particle length (parallel to the vessel axis) was 6.5 +/- 2.7 microm and width (perpendicular to the axis) was 6.1 +/- 1.7 microm, in agreement with published dimensions of individual red blood cells for this species. After Dextran 500 infusion, particle length and width increased significantly to 8.7 +/- 5.1 and 10.4 +/- 4.4 microm, respectively. Particle dimensions were greater in the central region of the flow stream for both normal and dextran-treated blood and increased at low flow rates with dextran-treated blood. This study provides direct confirmation of aggregate formation at low shear in venules with high-molecular-weight dextran as well as an estimate of aggregate size and range.

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Oxygen tension distribution in postcapillary venules in resting skeletal muscle.

We tested the hypothesis that blood flow is distributed among capillary networks in resting skeletal muscle in such a manner as to maintain uniform end-capillary PO2. Oxygen tension in venules draining two to five capillaries was obtained by using the phosphorescence decay methodology in rat spinotrapezius muscle. For 64 postcapillary venules among 18 networks in 10 animals, the mean PO2 was 30.1 Torr (range, 9.7-43.5 Torr) with a coefficient of variation (CV; standard deviation/mean) of 0.26. Oxygen levels of postcapillary venules within a single network or single animal, however, displayed a much smaller CV (0.064 and 0.094, respectively). By comparison, the CV of blood flow in 57 postcapillary venules of 17 networks in 9 animals was 1.27 with a mean flow of 0.011 +/- 0.014 nl/s and a range of 3.7 x 10(-4) to 6.5 x 10(-2) nl/s. Blood flow of postcapillary venules within single networks displayed a lower CV (mean, 0.51), whereas that in individual animals was 0.78. Results indicate that among venular networks, heterogeneity of oxygen tension is less than that of blood flow and within venular networks the heterogeneity of oxygen tension is much less than that of blood flow. In addition, postcapillary PO2 was independent of flow among venules in which both were measured. Results of this study may be attributable to three factors: 1) O2 diffusion between adjacent capillaries and venules, 2) structural remodeling in regions of lower PO2, and 3) O2-dependent local control mechanisms.

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Microvascular oxygen distribution in awake hamster window chamber model during hyperoxia.

The microvascular effects and hemodynamic events following exposure to normobaric hyperoxia (because of inspiration of 100% O2) were studied in the awake hamster window chamber model and compared with normoxia. Hyperoxia increased arterial blood Po2 to 477.9 +/- 19.9 from 60.0 +/- 1.2 mmHg (P < 0.05). Heart rate and blood pressure were unaltered, whereas cardiac index was reduced from 196 +/- 13 to 144 +/- 31 ml.min-1.kg-1 (P < 0.05) in hyperoxia. Direct measurements in the microcirculation showed there was arteriolar vasoconstriction, reduction of microvascular flow (83% of control, P < 0.05), and functional capillary density (FCD, 74 +/- 16% of control), the latter change being significant (P < 0.05). Calculations of oxygen delivery and oxygen consumption based on the measured changes in microvascular blood flow velocity and diameter and estimates of oxygen saturation corrected for the Bohr effect due to the lowered pH and increased Pco2 showed that oxygen transport in the microvascular network did not change between normal and hyperoxic condition. The congruence of systemic and microvascular hemodynamics events found with hyperoxia suggests that the microvascular findings are common to most tissues in the organism, and that hyperoxia, due to vasoconstriction and the decrease of FCD, causes a maldistribution of perfusion in the microcirculation.

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