A Novel Polymorphism in Glycoprotein IV (Replacement of Proline-90 by Serine) Predominates in Subjects with Platelet GPIV Deficiency

Abstract

SummaryTo clarify the molecular basis of the deficiency of glycoprotein IV (GPIV) of the platelet surface, we analyzed GPIV cDNA synthesized from platelet RNA of five unrelated Japanese subjects whose platelets did not express GPIV.We confirmed the presence of normal-sized GPIV mRNA in platelets from subjects with GPIV deficiency. The sequence of platelet GPIV cDNA from GPIV deficient subject showed three differences when compared with the published sequence; 1) a replacement of a 478CCT codon for proline-90 by TCT for serine, 2) a four-base insertion in the 3′-noncoding region, and 3) a substitution of A for 79C in the 5′-noncoding region. The replacement of Pro90 by Ser predominates in subjects with GPIV deficiency; that is, four out of five platelets with GPIV deficiency contained GPIV mRNA encoding GPIVSer-90, while all platelets from 17 GPIV positive subjects had GPIV mRNA encoding GPIVPro-90. The sequence of platelet GPIV cDNA which did not encode GPIVSer-90 from a subject with GPIV deficiency revealed no abnormality in the coding region. The four-base insertion in the 3′-noncoding region and the substitution of A for 79C in the 5′-noncoding region seems to be unrelated to the expression of GPIV.The substitution of Ser for Pro90 might alter the GPIV structure or impair GPIV biosynthesis, resulting in a lack of detectable GPIV. This hypothesis remains to be tested.

Patients with CD36 Deficiency Are Associated with Enhanced Atherosclerotic Cardiovascular Diseases

“…In patients with CD36-D, compared with healthy CD36-positive controls, metabolic phenotypes such as high TG levels, low HDL-C levels, high fasting glucose and hypertension were observed more frequent- 7,9) . Furthermore, we also found that patients with CD36-D are accompanied by insulin resistance 10) , postprandial hyperlipidemia, and high levels of remnant lipoprotein cholesterol and FFA [8][9] .…”

“…In order to confirm the diagnosis of CD36-D in these patients, immunofluorescent flow cytometric analysis was performed by using mouse monoclonal antibodies against CD36 (OKM5; Ortho Diagnostic System Inc., Raritan, NJ) at the Department of Blood Transfusion, Osaka University Hospital 7) . Briefly, 20 ml of blood was drawn, anticoagulated with heparin (10 U/ml), layered over 10 ml FicollPaque (GE Healthcare UK Ltd., Buckinghamshire, UK) and centrifuged at 1,000 g for 30 minutes.…”

“…Briefly, 20 ml of blood was drawn, anticoagulated with heparin (10 U/ml), layered over 10 ml FicollPaque (GE Healthcare UK Ltd., Buckinghamshire, UK) and centrifuged at 1,000 g for 30 minutes. A 50 l suspension of platelets (2 10 5 / l) or mononuclear cells (2 10 4 / l) was incubated with FITC-conjugat-ed anti-human CD36 monoclonal antibody OKM5 (Ortho Diagnostic Systems) (final concentration: 2.5 g/ml) or FITC-conjugated mouse IgG (final concentration: 2.5 g/ml) for 30 minutes at 40 and assayed on a FACScan ® system (Becton Dickinson Co., Mountain View, CA) as previously reported 7) . Appropriate cell fractions for the analysis of monocytes were selected by a gating method with a two-dimensional display of forward scatter and side scatter of analyzed cells 15) .…”

“…Kashiwagi et al identified several genetic mutations of human CD36 deficiency (CD36-D) 5) . We previously investigated the metabolic phenotypes of CD36-D patients [6][7] and reported that they (high fasting serum triglycerides level, low HDL-C level, fasting hyperglycemia, insulin resistance and hypertension) were frequently observed and clustered in patients with CD36-D, similar to those with MetS [8][9] . It was later reported that the accumulation of these metabolic phenotypes is not due to the deposition of abdominal visceral fat, but to insulin resistance or impaired metabolism of lipoproteins and free fatty acids (FFA) in the postprandial state in patients with CD36-D [8][9][10] .…”

Reduced uptake of oxidized low density lipoproteins in monocyte-derived macrophages from CD36-deficient subjects.

“…The physiological function of CD36 has not been well characterized although it was proposed to be a receptor for both thrombospondin and collagen or to mediate cytoadherence of plasmodium falciparum parasitized erythrocytes (17)(18)(19)(20). From the research on the subjects refractory to HLA-matched platelets transfusion (21)(22)(23)(24), Kashiwagi et al have identified the subjects whose platelets and monocytes lack CD36, and clarified three types of gene abnormalities (substitution of Pro' by Ser, deletion of two bases in exon 5, and insertion of one base in exon 10; Kashiwagi, H., Y. Tomiyama, S. Nozaki, S. Honda, H. Mizutani, S. Kosugi, M. Shiraga, N. Nagao, Y. Kanakura, Y. Kurata, and Y. Matsuzawa, manuscript in submission) (25,26). All these abnormalities result in the absence of CD36 expression on the cell surface.…”