Poloxamer-188 improves capillary blood flow and tissue viability in a cutaneous burn wound.

After cutaneous burn injury, an area of tissue 1-2 mm thick surrounding the wound is the site of a pronounced inflammatory response where blood flow is reduced. This "zone of stasis" undergoes progressive necrosis within 24-48 h, resulting in an expansion of the burn wound. Poloxamer-188 (P-188) is a surfactant that has been shown to prevent cell death due to electrical injury in vivo and heat shock in vitro. In this study, we investigated the effect of P-188 on blood flow within and around a burn wound and on the expansion of the wound area within 24 h after administration of a full-thickness burn injury. Results show that immediately (0-2 h) after the burn, red blood cell speed decreased to zero in a zone extending up to 1 mm from the center of the burn in both P-188 (200 mg/kg)- and saline (0.9%)-treated animals. Between 1 and 3 mm from the center of the burn, red blood cell speed decreased to 50% of preburn levels in saline controls (n = 5), while no decrease occurred in P-188-treated animals (n = 5). Beyond 3 mm from the center of the burn, red blood speed was equal to the preburn levels in saline controls, while it increased by about 10% in P-188 animals. Twenty-four hours after administration of burn, the "zero red blood cell speed zone," termed as the zone of coagulation, became smaller in P-188-treated animals, with an area of 2.4 +/- 0.5 mm(2) (n = 5) compared to 3.5 +/- 0.5 mm(2) (n = 4) in saline controls (P < 0.01). These results suggest that P-188 prevented the formation of a zone of stasis within 2 h after the burn injury and reduced the area of coagulation observed 24 h after cutaneous burn injury.

In vitro and in vivo evaluation of albumin synthesis rate of porcine hepatocytes in a flat-plate bioreactor.

Several configurations of extracorporeal bioartificial liver devices have been developed for the potential treatment of fulminant hepatic failure or as a bridge to liver transplantation. Recently, we developed a microchannel flat-plate bioreactor with an internal membrane oxygenator in which porcine hepatocytes are cultured as a monolayer on the bottom glass surface. In the present study, we investigated synthetic function of porcine hepatocytes in the bioreactor in both in vitro and in vivo flow circuit models. In vitro, albumin synthesis was stable in the bioreactor for up to 4 days of perfusion. In vivo, with the extracorporeal connection of the bioreactor to rat vasculature, porcine albumin was detectable for 24 h in the rat plasma. We also developed a simple mathematical model to predict the in vivo porcine albumin concentration in rat plasma. These results indicate that this configuration of a microchannel flat-plate bioreactor has potential as a liver support device and warrants further investigation.

Effects of oxygenation and flow on the viability and function of rat hepatocytes cocultured in a microchannel flat-plate bioreactor.

The goal of this study was to investigate the viability and synthetic function of rat hepatocytes cocultured with 3T3-J2 fibroblasts in a small-scale microchannel flat-plate bioreactor with and without an internal membrane oxygenator under flow. Bioreactor channel heights ranged between 85 and 500 microm and medium flow rates ranged between 0.06 and 4.18 mL/min. The results showed that the bioreactor without the oxygenator resulted in significantly decreased viability and function of hepatocytes, whereas hepatocytes in the bioreactor with internal membrane oxygenator were able to maintain their viability and function. The shear stress calculations showed that, at lower wall shear stresses (0.01 to 0.33 dyn/cm(2)), hepatocyte functions, measured as albumin and urea synthesis rates, were as much as 2.6- and 1.9-fold greater, respectively, than those at higher wall shear stresses (5 to 21 dyn/cm(2)). Stable albumin and urea synthesis rates for 10 days of perfusion were also demonstrated in the bioreactor with internal membrane oxygenator. These results are relevant in the design of hepatocyte bioreactors and the eventual scaling-up to clinical devices.

Analysis of oxygen transport to hepatocytes in a flat-plate microchannel bioreactor.

Oxygen transfer to cultured hepatocytes in microchannel parallel-plate bioreactors with and without an internal membrane oxygenator was investigated with a mathematical model and the results were corroborated with fluorescence imaging experiments. The consumption of oxygen by hepatocytes was assumed to follow Michaelis-Menten kinetics. Our simulations indicate that under conditions of low Péclet (Pe) number (<80) and fixed Damlkohler number (= 0.14, corresponding to rat hepatocytes) the cells are hypoxic in the bioreactor without an internal membrane oxygenator. Under the same conditions, the bioreactor with an internal membrane oxygenator can avoid cell hypoxia by appropriate selection of membrane Sherwood number and/or the gas phase oxygen partial pressure, thus providing greater control of cell oxygenation. At high Pe, both bioreactors are well oxygenated. Experimentally determined oxygen concentrations within the bioreactors were in good qualitative agreement with model predictions. At low Pe, cell surface oxygen depletion was predicted from analytically derived criteria. Hepatocytes with oxygen dependent functional heterogeneity may exhibit optimal function in the bioreactor with the internal membrane oxygenator.

Dynamics of tissue neutrophil sequestration after cutaneous burns in rats.

BACKGROUND: Neutrophil recruitment in organs after burns may cause local vascular damage, which can be reduced by agents blocking neutrophil adhesion to the vascular wall. Because these agents may increase susceptibility to infection, it is important to characterize the dynamics of neutrophil sequestration in order to optimize an eventual anti-adhesion therapy. MATERIALS AND METHODS: Rats were scald burned over 20 or 40% of their total body surface area (TBSA) and saline resuscitated. Sham controls were used. Myeloperoxidase (MPO) activity was measured in lungs, liver, kidney, gut, and burned skin up to 1 week postburn. Extravascular accumulation of (125)I-labeled bovine serum albumin ((125)I-BSA) was measured at 12 h postburn. RESULTS: MPO activity in lungs, liver, and kidney was increased within 3 h postburn and returned to normal within 24-48 h. Peak MPO levels occurred at 6-12 h postburn and were similar for both burn sizes. No MPO increase was observed in gut. MPO levels in burned skin did not increase before 6 h, peaked at 24 h, decreased at 48 h, but remained elevated for up to 7 days. Neutrophil recruitment in lungs and liver was confirmed histochemically. No neutrophils were found in kidneys. Extravascular (125)I-BSA was increased in lungs, liver, kidneys, and gut, in the 40% TBSA group only. CONCLUSIONS: Neutrophil sequestration in remote organs is a transient phenomenon while neutrophil homing into the wound site is sustained. Neutrophil accumulation dynamics are independent of burn size, although a minimum size is required to trigger vascular damage. Temporary early anti-adhesion therapy to reduce lung and liver neutrophil sequestration with little impact on neutrophil homing into the burn wound may be possible.

Enhancement of O2 and CO2 transfer through microporous hollow fibers by pressure cycling.

An intravascular gas exchange device for the treatment of respiratory failure consisted of a multitude of blind-ended hollow fibers glued in a pine-needle arrangement to a central gas supply catheter. It has previously been shown that gas desorption rates can be significantly enhanced by cycling gas pressure between a hypobaric level of 130 and an ambient level of 775 Torr. In this study, influences of the cycling frequency (f) and the cycle fraction during which hypobaric pressure is applied (theta) were investigated. Rates of O2 desorption from O2-saturated water and CO2 desorption from CO2-saturated water into a manifold containing 198 fibers, 380 microm in diameter, were measured over a range of f from 0.2 to 1.0 Hz. theta from 0.1 to 0.8, and fiber lengths from 4 to 16 cm. Relative to operation at ambient pressure, pressure cycling increased O2 transfer 3-4 times and CO2 transfer 4-6 times when the water flowed over the fiber manifold at 2.3 l/min. Transfer rates were relatively insensitive tof and theta with 80%-90% of maximum enhancement obtained when theta was as low as 0.2. Transfer rates increased continuously with fiber length, implying that pressure cycling reduced the intra-fiber resistance to gas diffusion. A mathematical diffusion model which utilized only two adjustable parameters, a mass transfer coefficient for O2 and for CO2, simulated the trends exhibited by desorption data.

Small intrapulmonary artery lung prototypes. Mathematical modeling of gas transfer.

Two diffusion models have been developed to analyze gas transfer data previously measured in an intravascular artificial lung consisting of a central gas supply catheter from which are tethered a large number of blind-ended microporous fibers of equal length. A convective-diffusion model (CD) describes the countercurrent transfer of a binary gas pair when gas is supplied at constant pressure conditions, and a well mixed (WM) cycled pressure model predicts transfer when the gas supply pressure is time cycled between compression and vacuum conditions. Regression of gas to gas and liquid to gas excretion data with the CD model resulted in estimates of the liquid phase mass transfer coefficient kAI. Because these values were intermediate between the kAI expected for flow parallel to a cylinder and for flow normal to a cylinder, gas transfer was influenced by both the tethered region of the fiber that was nearly perpendicular to the axis of the test section and the free end of the fiber that rested along the wall of the test section. With a time cycled gas supply pressure, the enhanced carbon dioxide and oxygen excretion predicted by the WM model was similar to the data, but a loss in transfer efficiency with fiber length was not accounted for by the theory.