- Title
- Biogenesis of plasma CD36+microparticles in human diabetes and the metabolic syndrome
- Creator
- Alkhatatbeh, Mohammad Jamil
- Relation
- University of Newcastle Research Higher Degree Thesis
- Resource Type
- thesis
- Date
- 2013
- Description
- Research Doctorate - Doctor of Philosophy (PhD)
- Description
- CD36 is a widely expressed cell surface receptor that binds lipoproteins and its function has been implicated in many complications of the metabolic syndrome. A cell-free form of CD36, soluble CD36 (sCD36), has been reported in human plasma and found to be elevated in obesity and diabetes and claimed to be a marker of insulin resistance. The nature and origin of the sCD36 has not been determined previously, although it has been postulated to be either a product of proteolytic cleavage or as an intact glycoprotein in the form of circulating microparticles (MPs) which are defined as small (0.1 - 1 μm in diameter) membranous microvesicles that can be specifically and selectively released from the cell membrane and retain many features of their cell of origin. MPs are present in the peripheral blood of normal healthy individuals but their numbers can increase dramatically in various pathological states, including type 2 diabetes and various vascular diseases. Given that MPs are enriched with bioactive proteins and nucleic acids, MPs bearing CD36 may not just be a marker of insulin resistance but may in fact contribute to disease pathogenesis. Thus the overarching hypothesis for this thesis is that plasma derived CD36 (sCD36) identifies a specific subset of MPs which contributes to the pathogenesis of type 2 diabetes and/or its complications. The first objective of this thesis was to determine the nature of sCD36; in particular whether sCD36 is truly soluble or, as hypothesized, found as a component of circulating MPs. Biochemical experiments done on plasma of normal subjects revealed that the cell-free plasma CD36 was not associated with its lipoprotein ligands and was not a proteolytic fragment; rather it was associated with the plasma MP fraction suggesting that sCD36 is a product of circulating MPs. Flow cytometric and immunoblotting analyses of plasma from normal donors showed these MPs were derived mainly from platelets. Analysis of in vitro activated platelets also showed that CD36 was secreted in the form of MPs. The second objective of this thesis was to further understand the potential role of CD36 in obesity and the pathogenesis of diabetes. The aim was to determine the levels and cellular sources of the CD36+MPs in patients with type 2 diabetes compared to normal lean and obese controls. Levels of CD36+MPs were found to correspond to approximately 50% of those of platelet derived MPs, were significantly higher in obese patients with type 2 diabetes compared to the obese controls (p<0.00001), and were primarily derived from mature erythrocytes (35.8 ± 14.6%). Plasma CD36 protein concentration measured by ELISA was positively correlated with CD36+MPs measured by flow cytometry but only weakly associated with the distribution of controls and patients with diabetes. Multivariate analysis confirmed that CD36+MP levels were a much better marker of diabetes than CD36 protein concentration. The third objective of this thesis was determine if there was any pattern of elevated MP levels related to diabetic complications and medications. Analysis by self-reported diabetic complications did not show significant differences in most of the MP subsets between patient subgroups except for the significant increase in % of CD36+/PS+MPs ([PS] phosphatidylserine; activation/apoptosis marker) in patients with microangiopathy and peripheral ulcer and the significant decrease in CD14+MPs (monocyte marker) and CD36+/CD41+MPs (platelet marker) in patients with reported nephropathy. Analysis by self-reported medications showed a significant increase in absolute numbers of CD36+/CD105+MPs (endothelial marker) and CD36+/CD45+MPs (leukocyte marker) in patients with diabetes taking sulfonylurea and a significant increase in % of CD36+/CD235a+MPs (erythrocyte marker) in patients taking metformin compared to those who were not. Total CD36+MP levels were not significantly associated with any of the self-reported diabetic complications and medications except for patients who were treated with calcium blockers. The last objective of this thesis was to determine if CD36+MPs could be released from cells of organs exposed to diabetic conditions. To this end, in vitro models were developed to represent complications of type 2 diabetes including diabetic nephropathy and fatty liver disease. Treatment of cell lines using advanced glycosylation end products (AGEs) and palmitic acid (PA) induced cellular death and CD36+MP production from HK-2 cells (nephropathy model) and HepG2 cells (fatty liver model) under circumstances resembling those that occur in diabetic plasma. If similar processes occur in human liver and kidney, it will be expected that CD36+MPs could be produced from CD36 expressing tissues especially those which are involved in diabetes and its complications. Collectively, this thesis establishes that the reported diabetic marker sCD36 in human plasma is entirely associated with circulating MPs (CD36+MPs). Interestingly, CD36+MP levels were found to be a better marker of diabetes than sCD36 protein concentration. The origin of circulating MPs could be easily determined as they also express antigens of their cellular source. CD36+MPs in patients with type 2 diabetes were mainly derived from mature erythrocytes but the underlying pathophysiology behind involvement of erythrocytes in diabetes requires further investigations. In addition, further investigations are needed to determine whether CD36+MPs could contribute as mediators of diabetes or if they are purely just biomarkers.
- Subject
- microparticles; CD36; diabetes
- Identifier
- http://hdl.handle.net/1959.13/936848
- Identifier
- uon:12420
- Rights
- Copyright 2013 Mohammad Jamil Alkhatatbeh
- Language
- eng
- Full Text
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