The regulation of hemopoiesis in long-term bone marrow cultures. II. Stimulation and inhibition of stem cell proliferation

The isolation of a DNA synthesis inhibitor (NBME fraction IV) and stimulator (RBME fraction III) specific for the hemopoietic stem cell (CFU-s) from freshly isolated normal adult and regenerating murine bone marrow, respectively, has been well documented. We have utilized long- term liquid bone marrow cultures in a further analysis of the role of these factors in the regulation of CFU-s proliferation. Our results show that shortly after feeding, at a time when the cultured CFU-s are actively proliferating, high levels of the hemopoietic stem cell proliferation stimulator fraction III can be isolated from the culture medium. In contrast, the presence of essentially noncycling CFU-s found in cultures fed 8-10 days previously correlates with high levels of the hemopoietic stem cell inhibitor fraction IV. These results suggest that a certain balance between these factors determines CFU-s proliferation in the long-term cultures. In support of this, DNA synthesis in actively cycling CFU-s in the long-term cultures is inhibited for at least 3 days by the addition of excess NBME fraction IV (inhibitor). Furthermore, DNA synthesis in noncycling cultured CFU-s is stimulated for at least 5 days by the addition of RBME fraction III (stimulator).     

Preparation of factor IX deficient human plasma by immunoaffinity chromatography using a monoclonal antibody

A murine hybridoma clone is described that grows continuously in culture and produces a monoclonal antibody we have called Royal Free Monoclonal Antibody to factor IX No. 1 (RFF-IX/1). This has high affinity for a coagulation site on factor IX. RFF-IX/1 immobilised on sepharose can be used to deplete factor IX from normal human plasma. This immunoaffinity depleted plasma is indistinguishable from severe Christmas disease plasma and can be used as the substrate in a one stage coagulation assay for factor IX. The affinity column has high capacity and can be regenerated so that large scale production from normal plasma of factor IX deficient plasma as a diagnostic reagent is now feasible.     

Severe perinatal thrombosis in double and triple heterozygous offspring of a family segregating two independent protein S mutations and a protein C mutation

Molecular genetic and phenotypic analyses were performed in a highly unusual case of combined protein S and protein C deficiency manifesting in a family in which a child had died perinatally from renal vein thrombosis. Antenatal diagnosis in a second pregnancy was initially performed by indirect restriction fragment length polymorphism (RFLP) tracking using a neutral dimorphism within the PROS gene and served to exclude severe protein S deficiency. Am umbilical vein blood sample at 22 weeks gestation showed isolated protein C deficiency. This pregnancy proceeded to a full-term delivery without thrombotic complications. Molecular genetic analysis of the PROC and PROS gene segregating in the family then yielded one PROC gene lesion in the father and two PROS gene lesions, one in each parent. These lesions were shown to segregate with the respective deficiency states through the family pedigree. Analysis of DNA from paraffin-embedded liver tissue taken from the deceased child showed the presence of both PROS mutations, as well as the PROC mutation. Genotypic analysis of the second child showed a PROC mutation, but neither PROS mutation consistent with its possession of normal protein S levels and a low/borderline protein C level. Antenatal diagnosis was then performed in a third pregnancy by direct mutation detection. However, although the fetus carried only the paternal PROS and PROC gene lesions, the child developed renal thrombosis in utero. It may be that a further genetic lesion at a third locus still remains to be defined. Alternatively, the intrauterine development of thrombosis in this infant could have been caused, at least in part by a transplacental thrombotic stimulus arising in the protein S-deficient maternal circulation. This analysis may, therefore, serve as a warning against extrapolating too readily from genotype to phenotype in families with a complex thrombotic disorder.     

Scientific Session III: Impact of Neurosonology on Therapeutic Decisions

Background and Purpose: A study of carotid endarterectomies performed without angiography was conducted in order to evaluate the safety of this approach in a selected group of patients.     

Postthaw stability of fibrinogen in cryoprecipitate stored between 1 and 6 degrees C

The fibrinogen activity in thawed cryoprecipitate stored between 1 and 6 degrees C is maintained essentially unchanged in most bags for a month. Occasionally, a bag will have a reduction in fibrinogen. If pooling has not occurred, thawed cryoprecipitate should be useful as a source of fibrinogen for a period of time considerably in excess of the 6 hours allowed for its use as a source of factor VIII or von Willebrand factor.