Are functional and genetic components of platelets linked to coronary artery disease: A case-control study

Coronary artery disease (CAD) is a multifactorial disease process involving behavioral, inflammatory, clinical, thrombotic, and genetic components. Previous epidemiologic studies focused on identifying behavioral and demographic risk factors of CAD, but none focused on platelets. Current platelet literature lacks the known effects of platelet function and platelet receptor polymorphisms on CAD. This case-control analysis addressed these issues by analyzing data collected for a previous study. Cases were individuals who had undergone CABG and thus had been diagnosed with CAD, while the controls were volunteers presumed to be CAD free. The platelet function variables analyzed included fibrinogen Von Willebrand Factor activity (VWF), shear-induced platelet aggregation (SIPA), sCD40L, and mean platelet volume; and the platelet polymorphisms studied included PIA, α2 807, Ko, Kozak, and VNTR. Univariate analysis found fibrinogen, VWF, SIPA, and PIA to be independent risk factors of CAD. Logistic regression was used to build a predictive model for CAD using the platelet function and platelet polymorphism data adjusted for age, sex, race, and current smoking status. A model containing only platelet polymorphisms and their respective receptor densities, found polymorphisms within GPIbα to be associated with CAD, yielding an 86% (95% C.I. 0.97–3.55) increased risk with the presence of at least 1 polymorphism in Ko, Kozak, or VNTR. Another model included both platelet function and platelet polymorphism data. Fibrinogen, the receptor density of GPIbα, and the polymorphism in GPIa-IIa (α2 807) were all associated with CAD with odds ratios of 1.10, 1.04, and 2.30 for fibrinogen (10mg/dl increase), GPIbα receptors (1 MFI increase), and GPIa-IIa, respectively. In addition, risk estimates and 99% confidence intervals adjusted for race were calculated to determine if the presence of a platelet receptor polymorphism was associated with CAD. The results were as follows: PIA (1.64, 0.74–3.65); α2 807 (1.35, 0.77–2.37); Ko (1.71, 0.70–4.16); Kozak (1.17, 0.54–2.52); and VNTR (1.24, 0.52–2.91). Although not statistically significant, all platelet polymorphisms were associated with an increased risk for CAD. These exploratory findings indicate that platelets do appear to have a role in atherosclerosis and that anti-platelet drugs targeting GPI-IIa and GPIbα may be better treatment candidates for individuals with CAD. ^

Organization and function of platelet glycoprotein Ib-IX complex

Glycoprotein (GP) Ib-IX complex, the second most abundant receptor expressed on the platelet surface, plays critical roles in haemostasis and thrombosis by binding to its ligand, von Willebrand factor (vWF). Defect or malfunction of the complex leads to severe bleeding disorders, heart attack or stroke. Comprised of three type I transmembrane subunits—GPIbα, GPIbβ and GPIX, efficient expression of the GPIb-IX complex requires all three subunits, as evident from genetic mutations identified in the patients and reproduced in transfected Chinese hamster ovary (CHO) cells. However, how the subunits are assembled together and how the complex function is regulated is not fully clear. By probing the interactions among the three subunits in transfected cells, we have demonstrated that the transmembrane domains of the three subunits interact with one another, facilitating formation of the two membrane-proximal disulfide bonds between GPIbα and GPIbβ. We have also identified the interface between extracellular domains of GPIbβ and GPIX, and provided evidence suggesting a direct interaction between extracellular domains of GPIbα and GPIX. All of these interactions are not only critical for correct assembly and consequently efficient expression of the GPIb-IX complex on the cell surface, but also for its function, such as the proper ligand binding, since removing the two inter-subunit disulfide bonds significantly hampers vWF binding to the complex under both static and physiological flow conditions. The two inter-subunit disulfide bonds are also critical for regulating the ectodomain shedding of GPIbα by the GPIbβ cytoplasmic domain. Mutations in the juxtamembrane region of the GPIbβ cytoplasmic domain deregulate GPIbα shedding, and such deregulation is further enhanced when the two inter-subunit disulfide bonds are removed. In summary, we have established the overall organization of the GPIb-IX complex, and the importance of proper organization on its function. ^

Evidence for the involvement of proximal chromosome 3P loci in the progression of Von Hippel-Lindau related tumors

Von Hippel-Lindau (VHL) disease is an autosomal dominant disorder characterized by the development of retinal and central nervous system hemangioblastoma, renal cell carcinoma (RCC), pheochromocytoma and pancreatic islet cell tumors (PICT). The VHL gene maps to chromosome 3p25 and has been shown to be mutated in 57% of sporadic cases of RCC, implicating VHL in the genesis of RCC. We report a multigeneration VHL kindred in which four affected female siblings developed PICT at early ages. Analysis of the three coding exons of the VHL gene in this family revealed a single, missense mutation in codon 238. Inheritance of the 238 mutation has been reported to correlate with a 62% risk of pheochromocytoma development. In this kindred, all affected individuals carried the mutation as well as one additional sibling who showed no evidence of disease. Clinical screening of this individual indicated small ($<$1 cm) pancreatic and kidney tumors. Results suggest that inheritance of the codon 238 mutation does not correlate with early onset pheochromocytoma. Rather, the only individual in the pedigree with pheochromocytoma was the proband's mother who developed bilateral pheochromocytoma at the age of 62. Thus, the VHL codon 238 mutation may predispose to late onset pheochromocytoma in this family; however, it does not explain the preponderance of PICT in the third generation since this mutation has not been reported to increase the risk of developing pancreatic lesions. This suggests that inheritance of the codon 238 mutation and subsequent somatic inactivation of the wild type allele of the VHL gene may not be sufficient to explain the initiation and subsequent progression to malignancy in VHL-associated neoplasms. Since the two tumor types that most frequently progress to malignancy are RCC and PICT, we asked whether loss of heterozygosity (LOH) could be detected proximal to the VHL gene on chromosome 3 in distinct regions of 3p previously implicated by LOH and cytogenetic studies to contain tumor suppressor loci for RCC. LOH was performed on high molecular weight DNA isolated from peripheral blood and frozen tumor tissue of family members using microsatellite markers spanning 3p. Results indicated LOH for all informative 3p loci in tumor tissue from affected individuals with PICT. LOH was detected along the entire length of the chromosome arm and included the proximal region of 3p13-14.2 implicated in the hereditary form of renal cell carcinoma.^ If 3p LOH were a critical event in pancreatic islet cell tumorigenesis, then it should be expected that LOH in sporadic islet cell tumors would also be observed. We expanded LOH studies to include sporadic cases of PICT. Consistent LOH was observed on 3p with a highest frequency LOH in the region 3p21.2. This is the first evidence for an association between chromosome 3 loci and pancreatic islet cell tumorigenesis. (Abstract shortened by UMI.) ^