Quality control for peripheral stem cell collection: the Ottawa experience.
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ADP-ribosyl Cyclase↗
Biomedical subjects
V Fuller
Publications and source records attributed to V Fuller.
Expression of stem cell factor and c-kit mRNA in cultured endothelial cells, monocytes and cloned human bone marrow stromal cells (CFU-RF).
Previously, we have shown that conditioned medium from a subpopulation of human marrow stromal cells (CFU-RF) contain an activity able to stimulate the growth of macroscopic epo-dependent erythroid colonies. The ligand for the product of the c-kit proto-oncogene (also known as stem cell factor or SCF), among other activities, has been reported to have similar effects on erythroid colony growth. We have also presented data showing that SCF together with phytohemagglutinin-stimulated leukocyte conditioned medium can stimulate erythroid colony growth in the presence of antibodies to erythropoietin. Using the human SCF cDNA probe (K. Zsebo, Amgen Inc.) we now show that cells derived from CFU-RF colonies express SCF but not c-kit. Human umbilical vein endothelial cells were also found to express SCF and this expression was increased by addition of monocyte supernatant, IL-1 beta or thrombin. Cells of the human erythroleukemia cell line HEL were found to express c-kit but not SCF. Neither c-kit nor SCF mRNA were detected in phytohemagglutinin-stimulated lymphocytes. Together, these data support the view that the behaviour of proliferating erythroid stem cells in the marrow, which may express c-kit, could be regulated by membrane-bound SCF present on surrounding stromal cells.
Base Sequence↗
Electroblotting and immunohistochemical staining for identification of platelet antibodies.
The Western blot procedure has been adapted for use with a biotinylated antiglobulin reagent and a horseradish peroxidase complex (Vectastain ABC) to determine specific sites of antigen binding and permit discrimination between various types of platelet antibodies. Monospecific anti-PlA1 antisera or sera containing mixtures of multispecific HLA and unidentified platelet specific antibodies were tested with PlA1 phenotyped platelets. Using monospecific anti-PlA1, one intense band with relative molecular weight (Mr) of 88,000 and corresponding to glycoprotein IIIa was seen with the PlA1+ platelets. With mixtures of antibodies, reactions were seen with platelet specific antibodies without interference from the HLA antibodies; with one serum a band of Mr approximately 135,000 was identified with Baka+, but not Baka- platelets, indicating the presence of an anti-Baka in the serum. The location of the Baka antigen corresponded to the area for GP IIb. With another serum, a band of Mr 88,000 was revealed with PlA1- and some PlA1+ platelets suggesting the presence of an anti-PlA2. Two additional bands of Mr 160,000 and 200,000 were present on all preparations including autologous controls, probably due to the presence of non-specific IgG. Thus, immunohistochemical staining is readily adaptable to the Western blot technique, and antibodies to platelet-specific antigens can be easily differentiated from HLA antibodies.
Antibodies↗
An in vivo assessment of factor VIII concentrates.
Replacement therapy for patients with hemophilia A requires that a specific dose of factor VIII be administered. Generally, these calculations involve the use of the manufacturer's stated dose in an equation. We have assessed factor VIII concentrates by in vitro techniques and found a considerable discrepancy (35%) between the stated and the measured contents. Confirmation of this discrepancy was obtained by in vivo assessment where posttransfusion recoveries corresponded to the measured rather than the stated contents of the vial. The half-disappearance time ranged from five to eight hours. The data suggest that constant monitoring of factor VIII preparations is necessary to ensure optimal treatment of patients with hemophilia A and help to explain the recent efforts by the Office of Biologics to achieve standardization.
Adult↗
Effect of citrate anticoagulants on factor VIII levels in plasma.
The citrate anticoagulants used during blood collection have been developed for their benefits to red cells. The concentrations in which they are used are strictly regulated in the United States: citrate-phosphate-dextrose-adenine (CPDA) is used in a 1:8 ratio for the collection of whole blood, whereas 4 percent sodium citrate (NaCit) is used in a 1:10 ratio for manual plasmapheresis. Acid-citrate-dextrose formula A (ACD-A) or formula B (ACD-B) and NaCit are commonly used in a 1:12 or 1:15 ratio during automated plasmapheresis. These anticoagulants have different initial and final pH values and citrate concentrations and different effects on the recovery of factor VIII (FVIII) in the plasma. NaCit has a higher initial pH (6.64) than ACD-A (4.98), ACD-B (5.60), or CPDA (5.12). The effects of these different anticoagulants on plasma constituents obtained from six healthy subjects were studied. In standard citrate concentrations, the FVIII level was significantly lower (p less than 0.05) in the NaCit used for manual plasmapheresis than in either of the ACD solutions used in automated plasmapheresis (104 U/dl vs. 153 and 160 U/dl). When various ratios of NaCit to blood were used, the pH increased from 7.62 at a 1:10 dilution to 7.65 at a 1:50 dilution. As expected, a progressive decrease in anticoagulant level was associated with an increase in ionized calcium and also in the level of FVIII, with the latter values rising from 104 U per dl at 1:10 to 137 at 1:20 and 148 U per dl at 1:30. Clot formation was detected only at a ratio of 1:35.(ABSTRACT TRUNCATED AT 250 WORDS)
Adenine↗
[ATP level in the whole rat liver during cold hypoxia and subsequent rewarming].
Cold storage of the whole liver at 4 degrees C in SBS and UW solution allowed to prevent from osmotic swelling of cells, which appeared at early stages of liver storage at 4 degrees C in just saline solutions. This effect of preserving solutions contributes to the preservation of quite high level of intracellular ATP content in liver at the first two stages of hypothermic storage (6 and 18 hrs), which preserves even during following normothermic reperfusion of an organ. A statistical ATP reduction in comparison with the control level (almost twice) can be explained on the one hand by the exhaustion of intracellular substrates of oxidation and on the other hand by their loss for the supporting of homeostasis under cold ischemia and following incubation of liver at 37 degrees C.
Adenosine Triphosphate↗