Homing of long-term and short-term engrafting cells in vivo.

Long-term repopulating hematopoietic stem cells can be separated from cells which provided radioprotection (short-term repopulating cells) on the basis of size. This might be a result of the quiescent nature of long-term repopulating cells. To define the activity of these populations we utilized a dye, PKH26, which incorporates into the membrane of cells and is equally distributed to daughter cells when they divide. We were able to retrieve PKH26(+)-labeled cells posttransplant in the hematopoietic tissues of the recipients. We could also assess their cell cycle status and their ability, short- and long-term, to reconstitute secondary lethally irradiated hosts in limiting dilution. The results suggest that long-term repopulating cells remain quiescent in the bone marrow shortly after engraftment, whereas cells which radioprotect are more rapidly dividing. We could not detect labeled cells in the peripheral blood posttransplant, and even though cells homed to both the spleen and bone marrow the cells in the bone marrow were significantly more competent at reconstituting lethally irradiated secondary hosts.

Hematopoietic stem cell tracking in vivo: a comparison of short-term and long-term repopulating cells.

We have previously demonstrated that we could separate long-term repopulating stem cells from cells that provided radioprotection (short-term repopulating cells) on the basis of size and suggested that this might be due to the quiescent nature of long-term repopulating cells. To further define the activity of these populations, we used a dye (PKH26), which incorporates into the membrane of cells and is equally distributed to daughter cells when they divide. We developed an assay, which allowed us to retrieve PKH26(+) long-term and short-term repopulating cells in the hematopoietic tissues of the recipients posttransplant. We were able to recover the labeled cells and determine their cell cycle activity, as well as their ability to reconstitute secondary lethally irradiated hosts in limiting dilution. The results of our assay suggest that long-term repopulating cells are quiescent in the bone marrow (BM) 48 hours after transplant. We were able to detect only a few labeled cells in the peripheral blood posttransplant and even though cells homed to both the spleen and BM, more long-term repopulating cells homed to the marrow and only these cells, which homed to the marrow, were capable of reconstituting lethally irradiated secondary hosts long-term.

Cytokine induction of platelet activation.

Recent studies have focused on the accumulation of cytokines in stored platelet concentrates and the role that these cytokines play in mediating transfusion reactions. To elucidate any additional adverse effects that may be associated with cytokine accumulation, the authors examined whether cytokines, which normally accumulate during routine platelet storage, can cause platelet activation in vitro. Concentrations of IL-6 and IL-8 were first determined for random donor platelet concentrates on days 1 through 4 of storage. Fresh platelets were then incubated with these levels of exogenous cytokines, and activation measured by flow cytometry using a monoclonal antibody directed against p-Selectin. Significant platelet activation was observed with concentrations of cytokines which are normally present in days 3 and 4 of shelf life. The study data demonstrate that levels of cytokines that routinely accumulate in stored platelet products can affect platelet biology. Strategies to reduce cytokine generation during platelet storage may be a method to improve the function and viability of stored platelets used for transfusion.