eNews July 13, 2010

Transfusions of old RBCs inflame mice

Although red blood cell (RBC) transfusions benefit millions of people, they can be risky, especially in critically ill patients. The risks seem to increase the longer the RBCs are stored. A research team led by scientists at the Department of Pathology and Cell Biology, Columbia University College of Physicians and Surgeons, New York, has shown that mice transfused with old RBCs exhibit an acute rise in circulating iron levels that provokes a strong inflammatory response (Hod et al. 2010). The acute response aggravates pre-existing inflammatory states and facilitates infection by pathogens. The team's research may explain why some patients respond adversely to transfusions with long-stored RBCs.

RBC transfusions: pros and cons

Most blood donations are divided into their components: RBCs, platelets, clotting factors, plasma, antibodies and white blood cells. Transfusions of selected components reduce the risks of side effects and permit specific treatments to more individuals. RBCs are the most commonly transfused components. They restore the blood's oxygen-carrying capacity and are often given to people who are bleeding or who have severe anemia. They can be refrigerated for up to 42 days, and, sometimes (e.g., to preserve a rare type of blood), they can be frozen for up to 10 years (Merck Manuals Online Medical Library).

For reasons that are not well understood, RBCs become less viable the longer they are stored. Additionally, the 24-hour post-transfusion RBC survival rate is sometimes less than the FDA-mandated threshold of 75%. This is of particular concern because most of the RBCs that don't survive are cleared by macrophages and monocytes within an hour post-transfusion. Thus, RBC clearance of 25% or more from one blood unit acutely unloads a massive amount of hemoglobin iron to the monocyte/macrophage system. Two lines of evidence suggest that this massive iron load is responsible for the increased mortality, serious infections and multi-organ failure among some hospitalized patients (e.g., the critically ill) transfused with long-stored RBCs. First, in mice and humans, increased iron levels in macrophages are associated with increased levels of inflammatory cytokines. Second, increased circulating iron, especially nontransferrin-bound iron, facilitates the proliferation of certain pathogens.

Transfused old RBCs increase iron load in mice

JAX® Mice strain FVB/NJ. RBCs from this mouse were used to model an allogeneic transfusion in C57BL/6J recipients. The post-transfusion survival rates of fresh and two-week-stored FVB/NJ RBCs transfused into C57BL/6J mice are similar to those obtained for syngeneic transfusions.

To determine if an acute release of hemoglobin iron from long-stored, damaged RBCs causes adverse effects, the Columbia University team compared the effects of transfusing fresh and long-stored blood components from JAX® Mice strain FVB/NJ (001800) to JAX® Mice strain C57BL/6J (000664). They found that transfusions of old RBCs increase iron deposits in the liver, spleen and kidneys significantly more than transfusions of fresh RBCs.

Transfusions of old RBCs induce/aggravate inflammation

Transfusions of long-stored RBCs induce a robust inflammatory response in the liver and spleen of the recipient mice. The inflammation does not result in clinical symptoms, such as anorexia, reduced mobility, decreased alertness or lack of grooming. However, recipients first injected with lipopolysaccharide (LPS) – an endotoxin from the outer membrane of Gram-negative bacteria and that elicits a strong immune response in animals – and then transfused with stored RBCs produce higher levels of inflammatory cytokines, including IL6, MCP1, KC/CXCL1, MIP1β, IFNG, and IL10, than do LPS-injected mice transfused with fresh RBCs. Recipients treated with the most LPS become moribund, hunched and inactive 18-24 hours post transfusion.

RBC-released iron induces the inflammation

The team determined that the inflammation resulting from transfusion of old RBCs is induced by iron, not other RBC contents. When desferoxamine (DFO) and FO (an equimolar combination of DFO and ferric citrate), both iron chelators, are administered immediately before old RBCs are transfused, the inflammatory response is significantly reduced.

Transfusions of old RBCs enhance pathogen growth

Recipient plasma collected two hours after transfusions of either stored RBCs or washed stored RBCs support the growth of significantly more pathogenic E. coli than plasma from un-transfused mice, mice transfused with fresh blood, supernatant derived from stored RBCs, or ghosts prepared from stored RBCs.

The findings by the Columbia University team indicate that mice transfused with long-stored RBCs mount an inflammatory response mediated by increased levels of iron in the liver, spleen, and kidneys, and by increased circulating levels of nontransferrin-bound iron. The inflammation is exacerbated by pre-existing inflammatory states – such as that induced by LPS. The high circulating iron levels also enhance bacterial proliferation, suggesting that, in humans, they cause some of the harmful effects of transfusions with long-stored RBCs. Preventing the pro-oxidant effects of the increased iron levels may decrease these adverse effects. However, the team cautions, different storage procedures and inherent limitations of animal models do not warrant a change in human transfusion practice at this time.

Reference

Hod EA, Zhang N, Sokol SA, Wojczyk BS, Francis RO, Ansaldi D, Francis KP, Della-Latta P, Whittier S, Sheth S, Hendrickson JE, Zimring JC, Brittenham GM, Spitalnik SL. 2010. Transfusion of red blood cells after prolonged storage produces harmful effects that are mediated by iron and inflammation. Blood 115:4284-92.