calcimycin and Wiskott-Aldrich-Syndrome

The Wiskott-Aldrich syndrome (WAS) is an inherited platelet/T-lymphocyte disease characterized by small platelets, thrombocytopenia and immunodeficiency. Because degradative events have a significant role, we directly examined calpain (Ca(2+)-dependent neutral protease), a prominent protease in the affected cells, by functional and antigenic quantitation. Calpain activity in platelets of seven WAS patients was decreased to 59 +/- 3.7% (P < 0.01) relative to platelets of 11 normals. Platelets of two patients with immune thrombocytopenia had normal calpain activity. By immunoblotting, mu-procalpain, the mu-calpain species in resting (unstimulated) blood cells, was decreased in platelets of nine WAS patients to 58 +/- 14.6% (P < 0.01) relative to paired normals. In contrast, mu-procalpain levels in lymphocytes of seven WAS patients did not differ from normal lymphocytes. Normal platelets and lymphocytes have different mechanisms for Ca(2+)-dependent mu-procalpain activation. On addition of ionophore and Ca2+ to stirred platelets, 80kD mu-procalpain was rapidly (0.5 min) and quantitatively converted to 76 kD active mu-calpain; this process was the same in WAS platelets. In lymphocytes, mu-procalpain activation was slow, only partially complete (40 min), and the active species was 78 kD. The marked depletion of calpain in WAS platelets demonstrated in this study may result from inappropriate stimulation of platelets and be related to the severe thrombocytopenia that characterizes this disease.

Topics: Autoimmune Diseases; Blood Platelets; Calcimycin; Calcium; Calpain; Enzyme Precursors; Humans; Immunoblotting; Lymphocytes; Male; Thrombocytopenia; Wiskott-Aldrich Syndrome

We have selected 11 patients with primary immunodeficiency disorders predominantly affecting T lymphocyte function (four with ataxia-telangiectasia (AT), four with common variable immunodeficiency (CVI) and one each with Wiskott-Aldrich syndrome, hyper-IgE syndrome and combined immunodeficiency) with defective gamma interferon (IFN-gamma) production in vitro. Induction with phytohaemagglutinin showed low interleukin 2 (IL-2) production concomitant with reduced IFN-gamma titres. However the addition of 10 U/ml of rIL-2 to cultures stimulated with staphylococcal enterotoxin B or galactose oxidase failed to restore IFN-gamma production in defective cases. IFN-gamma was titrated by both bioassay and immunoradiometric assay, ruling out the possible release of inactive or altered IFN-gamma molecules. Normal levels of IFN-gamma were found in patients of patients with AT, as well as in two AT and two CVI cases, demonstrating heterogeneity of defects within these syndromes. Soluble inhibitors or cellular suppression of IFN-gamma were not observed in mixing experiments. The possibility that defective interaction between accessory cells and T lymphocytes might account for the poor response to the inducing agents was ruled out as no IFN-gamma was produced using a calcium ionophore--which bypasses this step--in seven patients with absolute IFN-gamma deficiency.

Topics: Ataxia Telangiectasia; Calcimycin; Humans; Hypergammaglobulinemia; Immunologic Deficiency Syndromes; Interferon-gamma; Interleukin-2; Phytohemagglutinins; T-Lymphocytes; Wiskott-Aldrich Syndrome

Platelets, basophils and neutrophils from a patient with the Wiskott-Aldrich syndrome (WAS) were exposed to stimuli that activate specific membrane receptor or directly initiate biochemical events (e.g. the Ca2+ ionophore A23187 and ionomycin or arachidonic acid). Platelets from this patient did not aggregate in response to ADP, collagen, thrombin or adrenaline, which activate specific membrane receptors. Platelet aggregation, however, was normal in response to compound A23187, ionomycin or exogenous arachidonic acid. Histamine release from basophils of the WAS patient was normal in response to anti-IgE, a formylated peptide (f-met peptide), and to A23187. Similarly, the release of the lysosomal enzymes, beta-glucuronidase and lysozyme, from neutrophils of the WAS patient in response to serum treated zymosan (Zx), f-met peptide, and A23187 was not significantly different from that of his parents and 13 normal donors. These results suggest that the primary defect in WAS is selectively present in platelets and is located in a biochemical step between receptor activation and Ca2+ influx and/or initiation of arachidonate metabolism.

Topics: Arachidonic Acid; Arachidonic Acids; Basophils; Blood Platelets; Calcimycin; Child; Ethers; Histamine Release; Humans; Ionomycin; Male; Neutrophils; Platelet Aggregation; Wiskott-Aldrich Syndrome