Modulation of erythroid adhesion receptor expression by hydroxyurea in children with sickle cell disease.

Background: We investigated adhesion receptor levels on red blood cells, reticulocytes and erythroid progenitors from children with sickle cell disease treated or not with hydroxyurea.

Design And Methods: Four groups of patients were investigated: (i) children receiving hydroxyurea for severe vaso-occlusive events (n=26); (ii) untreated children with a history of vaso-occlusive events (n=20); (iii) children with no history of vaso-occlusive events (n=28); and (iv) healthy African controls (n=27). Expression of adhesion receptors was analyzed by flow cytometry with specific mono-clonal antibodies.

Generation of mice with inactivated Rh or Rhag genes.

Mice carrying inactivated Rh and Rhag genes were generated by insertional targeting. KO animals exhibited normal growth, development and fertility and both types were indistinguishable at a gross phenotypic level from their wild type littermates. Preliminary analysis revealed that red cells from Rh-/- mice lack Rh protein and have a moderate decrease of Rhag protein, whereas those from Rhag-/- mice have a total absence of Rhag and Rh proteins.

Flow cytometric analysis of the association between blood group-related proteins and the detergent-insoluble material of K562 cells and erythroid precursors.

The linkage between blood group-related cell surface proteins and the detergent-insoluble material (DIM) was estimated by flow cytometry using a panel of specific monoclonal antibodies (mAbs) as a comparison of the antibody-binding capacity of intact and Triton-X100-treated cells. Studies were performed with K562 cells expressing endogenous or recombinant proteins and with human erythroid progenitors during their proliferation and differentiation in vitro. Glycophorin C (GPC) was found to be Triton-insoluble in both cellular models.

Time-course expression of polypeptides carrying blood group antigens during human erythroid differentiation.

The time course expression of blood group antigens was examined by flow cytometry using a two-phase liquid culture system that supports the proliferation and maturation of human erythroid progenitors from adult peripheral blood. The progression towards erythroid differentiation was followed by the expression changes of the transferrin receptor (CD71++) and glycophorin A (GPA+). Four main categories of blood group markers were identified: (i) those characterized by an early expression like ABO (A), Kell (K:2) and Rh50 which were detected in the Epo-independent phase 1, (ii) those including GPC (Gerbich, Ge antigens) and Fy6 which were expressed in the late phase 1, (iii) GPA (MN antigens), Wrb (Band 3/GPA interaction), Rh(D, Cc/Ee) and LW which appeared during the Epo-dependent phase 2 and (iv) those like Jk3 and Lub which were expressed in late phase 2.

Shift from Rh-positive to Rh-negative phenotype caused by a somatic mutation within the RHD gene in a patient with chronic myelocytic leukaemia.

We report a female patient whose Rh phenotype shifted from RhD-positive to RhD-negative over a 3-year period (1991-94), during which time she was treated with mastectomy (1992) and local irradiation for a low-grade recurrent breast cancer. She was diagnosed with chronic myeloid leukaemia in 1994, and has since then received chemotherapy. The patient was repeatedly typed as O, RhD-positive between 1965 and 1991 and was repeatedly found RhD-negative after 1994.

The Lutheran blood group glycoproteins, the erythroid receptors for laminin, are adhesion molecules.

The Lutheran antigens are recently characterized glycoproteins in which the extracellular region contains five immunoglobulin like domains, suggesting some recognition function. A recent abstract suggests that the Lutheran glycoproteins (Lu gps) act as erythrocyte receptors for soluble laminin (Udani, M., Jefferson, S.

Tumoricidal potential of human macrophages grown in vitro from blood monocytes.

Adoptive transfer of activated autologous human macrophages obtained by in vitro differentiation of monocytes in culture has successfully undergone phase I clinical trials in patients with metastatic cancer. The efficacy of these autologous macrophages in the in vivo killing of human tumors has now to be demonstrated. GM-CSF was shown to increase the number of monocytes differentiating in culture into macrophages.

Infusion of large quantities of autologous blood monocyte-derived macrophages in two cancer patients did not induce increased concentration of IL-6, TNF-alpha, soluble CD14 and nitrate in blood plasma.

In an attempt to increase the number of macrophages available for reinfusion in immunotherapy trials, GM-CSF was injected in vivo to mobilize circulating blood monocytes in 2 cancer patients. Subsequently mononuclear cells were collected by apheresis, cultured in the presence of GM-CSF and activated with IFN-gamma. This procedure resulted in the harvesting of 1.

[Positive selection of hematopoietic CD 34 stem cells for autograft].

The CD 34 antigen is a glycoprotein found on the surface of hematopoietic stem cells and early committed progenitors. The CE 34 stem cells from 14 samples of bone marrow, cord blood or leucapheresis were isolated using a positive selection procedure involving an anti CD 34 biotinylated monoclonal antibody and an avidin immunoabsorption device. Results showed that in 60 percent of samples, the positively-selected fractions contained more than 70 percent CD 34 cells.

New developments in stem cell detection and stem cell selection in the context of autologous transplantations.

The concept of bone marrow transplantation is now enlarged to hemopoietic stem cell transplantation. Basic research on hemopoiesis, its regulation and markers, allowed concrete progress in the field of autologous transplantation. We report two applications: 1) the improvement of methods for detection of stem cells, which ensure engraftment, using recombinant growth factors, allowed a more sensitive determination of chemotherapeutic in vitro treatments toxicity upon normal progenitor cells; 2) large scale selection of hemopoietic progenitor/stem cell using the CD 34 immunological marker: its application to autologous transplantation.

Lack of differential sensitivity of normal hematopoietic stem cells and murine lymphoblastic leukemic cells to a lysosomotropic agent, N-dodecyl morpholine.

The effect of N-dodecyl morpholine (NDM), a lysosomotropic compound, on the clonogenic capacity of GK15, Sp2.0, Hb131, and L1210 lymphoblastic tumor cells and CFU-GM and CFU-S progenitor cells from DBA/2 mice was measured in order to evaluate the potential use of this compound for the purging of tumor-contaminated bone marrow (BM) in autologous BM transplantation. The growth of clonogenic tumor cells from all of the tested cell lines was inhibited with doses of NDM that also killed 100% CFU-GM and CFU-S, and no optimal dose could be found in this animal model to purge marrow while sparing sufficient stem cells to ensure engraftment in syngeneic BM transplantation.

Cytotoxicity of anti-PP1Pk antibodies and possible relationship with early abortions of p mothers.

IgG and IgM anti-PP1Pk antibodies of human sera from p individuals were examined for their ability to agglutinate red blood cells (RBC) of various P phenotypes and to mediate ADCC of these RBC by human effector cells. Agglutinating antibodies were found in either the IgG or the IgM fraction, while ADCC active antibodies were found mainly in the IgG fraction and were essentially cytotoxic with Pk RBC. The possible relationship of these antibodies with early abortions of p mothers is discussed.

Activity of IgG and IgM ABO antibodies against some weak A (A3, Ax, Aend) and weak B (B3, Bx) red cells.

The IgG and IgM anti-A and anti-B activities from several immune and non-immune O, A and B sera were tested against a panel of weak (A (A3, AX, AND Aend) and weak B (B3 and Bx) red cells. In all cases it is the IgM which agglutinated optimally Ax (or Bx) cells, while IgG and IgM anti-A (or anti-B) reacted similarly with A3 and Aend (or B3) cells. The agglutinating activity of all these ABO antibodies was found straightly related to their association constant for the A (or the B) receptor.

Cad antigen: comparative study of 50 samples.

50 Cad (+) blood samples were tested using various anti-Cad reagents. Results demonstrated a wide Cad antigenic expression, even among the members of one family. In particular experimental procedures, almost all Cad (+) cells were polyagglutinable.