Neutrophil kinetics in the dog.

The production of neutrophils in dogs has been estimated from the number of postmitotic neutrophils in the marrow and the transit time of a [3H]-thymidine pulse. The number of postmitotic neutrophils was derived from the erythron iron turnover measurement of erythroid number and the neutrophil-erythroid ratio in bone marrow sections. The mean value for marrow postmitotic neutrophils in dogs was 5.61 plus or minus 0.56 times 10-9 cells/kg. The mean transit time of these neutrophils was calculated to be 82.1 h. A marrow production of 1.65 times 10-9 neutrophils/kg/day was calculated from these data. The turnover of circulating neutrophils was measured by [3H]thymidine and [32P]diisopropylphospho-fluoridate (DF32P) labeling of blood neutrophils. [3H]-Thymidine labeling gave a calculated recovery of 65 per cent, a t1/2 disappearance time of 6.7 h, and a calculated turnover of 1.66 times 10-9 cells/kg/day. Corresponding results with DF32P tagging were 51 per cent, 5.4 h, and 2.89 times 10-9 cells/kg/day. The discrepancy between these two tags persisted in doubly tagged cells and was considered to be due to elution of DF32P.     

Neutrophil kinetics in man.

A method has been developed for measuring neutrophil cellularity in normal human bone marrow, in which the neutrophil-erythroid ratio was determined from marrow sections and marrow normoblasts were estimated by the erythron iron turnover. Neutrophil maturational categories, defined by morphologic criteria, were supported by autoradiographs of marrow flashed-labeled with 3H-thymidine. Correction for multiple counting error was empirically derived by counting serial sections through cells of each maturational category. The normal neutrophil-erythroid ratio in 13 normal human subjects was 1.5 +/- 0.07. The mean number of normoblasts in the same subjects was estimated to be 5.07 +/- 0.84 X 10(9) cells/kg. Total marrow neutrophils (X 10(9) cells/kg) were 7.70 +/- 1.20, the postmitotic pool (metamyelocytes, bands, and segmented forms) was 5.59 +/- 0.90 and the mitotic pool (promyelocytes + myelocytes) was 2.11 +/- 0.36. Marrow neutrophil ("total") production has been determined from the number of neutrophils comprising the postmitotic marrow pool divided by their transit time Transit time was derived from the appearance in circulating neutrophils of injected 3H-thymidine. The postmitotic pool comprised 5.59 +/- 0.90 X 10(9) neutrophils/kg, and the transit time was 6.60 +/- 0.03 days. From these data marrow neutrophil production was calculated to be 0.85 X 10(9) cells/kg per day. Effective production, measured as the turnover of circulating neutrophils labeled with 3H-thymidine, was 0.87 +/- 0.13 X 10(9) cells/kg per day. This value correlated well with the calculation of marrow neutrophil production. A larger turnover of 1.62 +/- 0.46 X 10(9) cells/kg per day was obtained when diisopropylfluorophosphate-32P was used to label circulating neutrophils. Studies using isologous cells doubly labeled with 3H-thymidine and diisopropylfluorophosphate-32P demonstrated a lower recovery and shorter t1/2 of the 32P label.     

Neutrophil kinetics in acute infection

Neutrophil kinetics of acute experimental infection were studied with diisopropylfluorophosphate-³²P labeling in 31 dogs inoculated intrabronchially with pneumococci. In vitro neutrophil labeling indicated a rapid transit time through the blood in early infections, with an elevated marginal granulocyte pool sometimes preceding an elevation of the circulating granulocyte pool. 13 hr after infection, the circulating and total blood granulocyte pools were increased but the rate of neutrophil transit through the blood was normal. During the recovery from infection there was a marked prolongation of neutrophil blood transit time, suggesting virtually complete cessation of bone marrow release of neutrophils into the blood. Labeling of neutrophils in vivo indicated an increased rate of emptying of the bone marrow storage pool proportional to the severity of infection as measured by the fever index. The change in the blood ratio of nonsegmented to segmented neutrophils was a much more accurate index of the severity of infection than the blood granulocyte concentration, correlating significantly with the fever index.     

Liquid preservation of polymorphonuclear leukocytes: effect of various additives on chemotaxis preservation.

Adequate storage of polymorphonuclear leukocytes would allow an easier in vitro study of their structure and their functions, an easier study of polymorphonuclear leukocyte diseases (e.g. chronic granulomatous disease) and an easier use of polymorphonuclear leukocytes as a clinical tool (e.g. for localizing infections). Unfortunately, polymorphonuclear leukocytes are nearly impossible to preserve, even in short-term storage. This study proposes a model for the study of polymorphonuclear leukocyte storage in a synthetic medium: Plasmion. For storage over a period of 24 h, we found that supplementation with each of the following additives: bicarbonate buffer, glucose, adenosine triphosphate (ATP), ascorbic acid, nicotinamide adenine dinucleotide (NADH), nicotinamide adenine dinucleotide phosphate (NADPH), alpha-tocopherol acetate, amikacin and ampicillin, significantly improves (p less than 0.05) one or several functions of the polymorphonuclear leukocytes. When samples were stored for 48 h, we found that the addition of bicarbonate buffer after 24 h significantly improves the maintenance of several functions of polymorphonuclear leukocytes, in particular chemotaxis. Preservation for 96 h was achieved by making additions of supplements on each day of storage, with a chemotaxis maintenance of 83% at 24 h, 59% at 48 h, 46% at 24 h and 20% at 96 h. In conclusion, by using the Plasmion medium, and adding the above-mentioned compounds on each day of storage, chemotaxis can be satisfactorily maintained over 4 days.     

The effect of mode of agitation and type of plastic bag on storage characteristics and in vivo kinetics of platelet concentrates

We studied the characteristics of platelet concentrates stored for 5 days at 22 degrees C. Platelets were prepared in three plastic bags (PL 732, PL 1240, and CLX) and stored on one of four platelet agitators, 1- or 6-rpm elliptical and 2- or 6-rpm circular rotators. A total of 76 studies were divided among 12 groups, each group being composed of a different storage bag-rotator combination. In vivo recovery and survival were calculated using Indium-111 oxine-labeled platelets injected into autologous volunteers. Platelet recovery was assessed at 2 hours postinjection or as the y-intercept of the multiple-hit model survival curve. Survival was calculated using linear, exponential, and multiple-hit computer models. Linear T 1/2 also was calculated as an index of platelet survival. At 5 days, the pH of all concentrates was above pH 7.0 and platelet counts were above 5.5 X 10(10) per bag except for the PL 732 with the 6-rpm elliptical rotator, which was 4.6 X 10(10) per bag. This combination also showed a significantly higher poststorage lactic dehydrogenase (LDH) discharge compared to the mean of the other 11 groups (23.6 +/- 5.4% vs. 10.4 +/- 3.0%, p less than 0.05); however, the beta-thromboglobulin (beta-TG) release was not statistically different.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neutrophil collection using modified fluid gelatin. Effect on in vitro and in vivo neutrophil function

Modified fluid gelatin (MFG) is a possible alternative to hydroxyethyl starch (HES) for use as a red cell sedimenting agent in neutrophil collection procedures. Since the efficacy of neutrophil transfusion therapy depends upon the integrity of the infused cells, we examined the in vitro and in vivo function of neutrophils collecting using MFG. Neutrophils were collected from 17 normal subjects by continuous flow centrifugation using MFG in place of HES. In vitro measurement of neutrophil function included dye exclusion, phagocytosis, candidacidal activity, staphylococcal killing, chemotaxis, and random migration. For in vivo studies, neutrophils were labeled with 3H-diisopropylfluorophosphate, reinfused into the donor, and blood kinetics and skin chamber accumulation of the labeled cells were measured. In vivo results were compared with results from previous studies using neutrophils collected with HES or by phlebotomy. MFG neutrophils were normal by all in vitro functional criteria and localized normally at an in vivo inflammatory site. Intravascular survival (T 1/2 = 3.3 +/- 0.9 hours) was significantly shorter than normal (T 1/2 = 7.3 +/- 2.3 hours) but was identical to that of cells collected using HES. Thus, on the basis of neutrophil functional capabilities, MFG appears to be an acceptable alternative to HES in collection of neutrophils for transfusion.     

Neutrophil transfusion. In vivo function of neutrophils collected using cell separators

Effective transfusion therapy depends, in part, upon the ability of the transfused neutrophils to function normally in the recipient. To evaluate the in vivo function of neutrophils obtained with two widely used newer cell separators (Fenwal CS-3000 and IBM 2997), neutrophil concentrates from 12 normal subjects were labeled in vitro with 3H-diisopropylfluorophosphate, reinfused into the donor, and blood kinetics and skin chamber accumulation of the labeled cells were measured. Results were compared with previously obtained data (referred to as "normal") using neutrophils collected by phlebotomy. Blood kinetic measurements of cells obtained with the CS-3000 were normal, whereas blood recovery of cells obtained with the 2997 was decreased moderately. The percent of infused neutrophils localizing to the skin chamber was 0.101 and 0.044 for CS-3000 and 2997 cells, respectively; neither value was significantly different from normal (0.066). Thus, neutrophils collected using either of these cell separators function normally or nearly so in vivo.     

Further studies on the effect of adenine in blood preservation

Clinical and experimental studies on the effect of adenine supplementation to ACD blood are reported. The frequency and type of transfusion reactions in 5,595 transfusions of ACD-adenine blood (storage time 0–35 days) was the same as in 4,519 transfusions of ACD blood (storage time 0–21 days). There was no change in the plasma haptoglobin level in 10 recipients during and after transfusion of 29 to 35 day old ACD-adenine blood. The relation between posttransfusion viability, in vitro hemolysis, erythrocyte ATP content and duration of storage were studied. Supplementation with adenine increased the posttransfusion viability and the erythrocyte ATP content; these effects were noticeable after two weeks of storage. A negative correlation was found with respect to posttransfusion viability and the concentration of total lipids in the medium. A significant covariation between the concentrations of the three nucleotides ATP, ADP and AMP was found. This indicates that erythrocyte adenylate kinase dominates in the regulation of the relative concentrations of these nucleotides.