Systemic inflammation promotes erythrocyte sequestration

Introduction

Changes in red blood cell (RBC) flexibility and membrane surface properties affect their circulatory clearance. Glutaraldehyde (GLUT) stiffened RBC are cleared by about 70% within 30 min after transfusion [1]. Recently, we have shown that endotoxin (ETX) increases RBC adhesion to human vascular endothelial cells [2]. In this study we investigated the effects of endotoxin and cecal ligation and perforation induced sepsis (CLP) on RBC survival and sequestration in healthy and septic rats.

Methods

Groups: Rats were randomized to six groups. Groups H/N, H/E and H/S were healthy rats (H) receiving naive (N), endotoxin treated (E) or septic (S) RBC. Groups S/N, S/E and S/S were septic rats (24 h CLP)(S) receiving naive (N), endotoxin treated (E) or septic (S) RBC. RBC solutions: Blood was harvested from healthy or septic (24 h after CLP) donor rats. ETX treated RBC were prepared by incubating whole rat blood with ETX (75 µg/ml). RBC and plasma were separated by centrifugation (15 min, 1000 g). After washing with phosphate buffered saline, RBCs were labeled with ⁵¹Chromium (⁵¹Cr) and resuspended in plasma to the experimental animal's hematocrit. Infusion of labeled RBC solution was performed by an isovolemic and simultaneous exchange transfusion. Blood samples were taken after the exchange transfusion every 5 min for 30 min and in 60 min intervals over 4 h for ⁵¹Cr measurements. Organs were harvested for tissue ⁵¹Cr measurements. Cardiac output, body temperature (TEMP), mean arterial (MAP and central venous pressure (CVP) were measured before and 4 h after exchange transfusion.

Results

Infusion of naive, ETX-treated or septic RBC did not affect cardiac index, TEMP, CVP or MAP in healthy (H/N, H/E, H/S) or septic rats (S/N, S/E, S/S). The amount of radiolabeled RBC in the circulation remained unchanged for the first 30 min in all groups. After 60 min, however, intravascular survival of septic and ETX treated RBC started to fall in healthy rats and was significantly lower than survival of naive RBC at 240 min. In septic rats, not only the E and S-RBC but also the naive RBC were cleared by 15% at 240 min. In healthy animals, sequestration of transfused S- and E-RBC in liver and spleen was higher than sequestration of N-RBC. In septic animals, no difference in sequestration was found between N-, E-, and S-RBC. In skeletal muscle, lungs, intestine, femur, diaphragm and skin, sequestration was not different between healthy or septic recipients nor between the different groups of transfused RBC.

Conclusion

Both ETX and sepsis induced alterations of the RBC membrane increase RBC clearance. However, the kinetics of ETX treated or septic RBCs clearance are completely different from the kinetics of RBC stiffened by GLUT. The majority of the injured and cleared RBC are entrapped in liver and spleen while other organ systems play only a minor role. Furthermore, RBC sequestration also appears to depend on the host's inflammatory state indicating RBC-host interactions in the microcirculation going beyond changes in RBC deformability.