Phenotypic heterogeneity in the Gray platelet syndrome extends to the expression of TREM family member, TLT-1.

The Gray platelet syndrome (GPS) is a rare inherited disorder linked to undefined molecular abnormalities that prevent the formation and maturation of alpha-granules. Here, we report studies on two patients from unrelated families that confirm phenotypic heterogeneity in the disease. First we used immunoelectron microscopy (I-EM) to confirm that TREM-like transcript-1 (TLT-1) is mostly localized to alpha-granule membranes of normal platelets.

Impaired megakaryocytopoiesis in type 2B von Willebrand disease with severe thrombocytopenia.

In type 2B von Willebrand disease, there is spontaneous binding of mutated von Willebrand factor (VWF) multimers to platelets. Here we report a family in which severe thrombocytopenia may also be linked to abnormal megakaryocytopoiesis. A heterozygous mutation in the VWF A1 domain gave a R1308P substitution in an interactive site for glycoprotein Ibalpha (GPIbalpha).

Severe deficiency of glycoprotein VI in a patient with gray platelet syndrome.

We report a novel case of gray platelet syndrome (GPS) where a severe deficiency of the platelet collagen receptor, glycoprotein (GP) VI, accompanies classical symptoms of a low platelet count and platelets lacking alpha-granules. Dense granules were normally present. Platelet aggregation with collagen was severely decreased, as was the response to convulxin (Cvx), a GPVI agonist.

Mutations in the beta3 gene giving rise to type I Glanzmann thrombasthenia in two families in Portugal.

Glazzmann thrombasthenia is an inherited bleeding syndrome in which an absence of platelet aggregation is associated with quantitative or qualitative deficiencies of the alphaIIbbeta3 integrin. We now describe biochemical and molecular studies on two Portuguese families where platelets lack both surface and intracellular pools of alphaIIbbeta3. DNA extraction was followed by PCR-SSCP analysis of all exons and intronic boundaries in the alphaIIb and beta3 genes.

A novel hemizygous Bernard-Soulier Syndrome (BSS) mutation in the amino terminal domain of glycoprotein (GP)Ibbeta--platelet characterization and transfection studies.

Glycoprotein (GP) Ib-V-IX is a unique adhesion receptor complex on platelets. Mutations in GPIbalpha, Ibbeta, and IX can lead to the rare bleeding disorder, Bernard-Soulier Syndrome (BSS). Here, we report a novel hemizygous variant of BSS in which Pro29 in one GPIbbeta allele is substituted by a Leu (GPIbbeta:P29L).

Immunolocalization of P2Y1 and TPalpha receptors in platelets showed a major pool associated with the membranes of alpha -granules and the open canalicular system.

P2Y(1) and thromboxane-prostanoid-alpha (TPalpha) receptors on platelets belong to the G-protein-coupled 7-transmembrane domain family. They transmit signals for shape change, mobilization of calcium, and platelet aggregation. Immunogold labeling with a monoclonal antibody (MoAb) to the amino-terminal domain of P2Y(1) and a polyclonal antibody to the C-terminal domain of TPalpha revealed that while present at the platelet surface, both receptors were abundantly represented inside the platelet.

Analysis of the amino acid requirement for a normal alphaIIbbeta3 maturation at alphaIIbGlu324 commonly mutated in Glanzmann thrombasthenia.

Glanzmann thrombasthenia is an inherited bleeding disorder arising from quantitative or qualitative defects of the alphaIIbbeta3 integrin of platelets. Here, we report that PCR-SSCP analysis and DNA sequencing revealed a homozygous single base pair substitution in exon 12 of the IIb gene leading to a Glu(324) (E) to Lys (K) substitution in the alphaIIb subunit in a patient with Type I disease. As this mutation is found on at least 3 continents, the codon for Glu(324) may be a mutational hotspot of the disease.

A Ser752-->Pro substitution in the cytoplasmic domain of beta3 in a Glanzmann thrombasthenia variant fails to prevent interactions between the alphaIIbbeta3 integrin and the platelet granule pool of fibrinogen.

A Glanzmann thrombasthenia variant with a beta3 Ser752-->Pro cytoplasmic domain substitution has platelets that fail to aggregate or bind soluble fibrinogen (Fg) after activation. Despite this, Fg is normally present in the alpha-granules. We have used immunoelectron microscopy to examine the reactivity of Fg with the different pools of alphaIIbbeta3 in the patient's platelets.

A novel 196Leu to Pro substitution in the beta3 subunit of the alphaIIbbeta3 integrin in a patient with a variant form of Glanzmann thrombasthenia.

Glanzmann thrombasthenia (GT) is an inherited disorder where an absence of platelet aggregation is associated with quantitative or qualitative abnormalities of the alphaIIbbeta3 integrin. In rare patients, amino acid substitutions have provided information on the functional significance of specific domains within alphaIIb or beta3. We now report an elderly male GT patient (R.

Paradoxical platelet activation was not observed on dissociation of abciximab from GpIIb-IIIa complexes.

The ability of abciximab to bind and dissociate from platelets raises the question of the conformational state of GPIIb-IIIa complexes losing abciximab and the risk of paradoxical drug-induced platelet activation. Platelets incubated with abciximab and mixed in vitro with c7E3 Fab-free platelets lost the drug to the new platelets giving a single platelet population with a unimodal abciximab distribution within 17 h. Prelabeling the receiving platelets with phycoerythrin-labeled anti-GPIb monoclonal antibody (MoAb), permitted their identification by flow cytometry.