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Columbia Dissertations and Theses > Doctoral Dissertations
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| Author(s): | Tatsis-Kotsidis, Irene |
| Title: | The A1 adenosine receptor of human and mouse adipose tissues : cloning, expression, and characterization |
| Physical Description: | xi, 231 leaves, bound. |
| Issue Date: | 1997 |
| Description: | Department: Nutrition. Thesis (Ph.D.)--Columbia University, 1997. |
| Bookmark as: | http://hdl.handle.net/10022/AC:P:2980 |
| Full Text (ProQuest): | /ac/proxit.jsp?url=http://gateway.proquest.com/ope... |
| Abstract: | Aberrant functioning of the A1 adenosine receptor in fat tissue has been implicated in obesity. To better understand this relationship we have prepared a number of tools for studying A1 adenosine receptors in adipose tissue. These include cDNAs encoding the human and mouse fat A1 receptors, their transient expression in 293T cells, an ob17 preadipocyte line stably expressing the human A1 receptor, an auto-anti-idiotypic antibody that reacts with A1 receptors, and two multivalent adenosine derivatives. Omental fat tissue from an obese female subject was used to obtain the complete coding region of the human fat A1 adenosine receptor gene. Its sequence was found to be identical to the published sequence of human brain A1 adenosine receptor, lacking any mutation that would directly account for her obese state. The problem of obesity, therefore, appears not to lie, as suggested by others, in the structure of the A1 adenosine receptor, but in how its gene is regulated. The murine version of the A1 receptor coding region was isolated from mouse abdominal tissue, in a similar, but not identical, manner as the human. Its sequence was identical to the published sequence of the mouse brain A1 receptor, and highly homologous to A1 subtypes from other species. Comparison of the ligand binding properties of the recombinant human and mouse fat receptors transiently expressed in 293T cells to that of native receptors confirmed that our cDNAs encode functional A1 adenosine receptors. Interspecies differences in binding characteristics were observed with our human and mouse receptors expressed in 293T cells, even though the proteins share greater than 90% identity. We associated the binding differences with four residues, located in the proximal half of the 2nd extracellular loop of the A1 receptor, which markedly differ between the two species. We examined the possibility that changes in cellular environment of the A1 adenosine receptor could affect its binding characteristics. However, comparison of the binding properties of the human A1 receptor expressed transiently in 293T cells and stably in ob17 cells, showed no differences in the relative binding of several ligands. Binding characteristics of A1 receptors in mouse brain and fat tissues were also examined; no differences were observed from their counterparts expressed at the cellular level. In a surprising finding, stable expression of the human A1 adenosine receptor in the ob17 preadipocyte line induced rounding of the cells and triggering of intracellular triglyceride formation, as shown by Oil-red O staining of lipid droplets. This is the first demonstration of the direct participation of the A1 adenosine receptor in preadipocyte differentiation, and suggests the possibility that expression in preadipose tissue may promote obesity. |
| Collection(s): | Doctoral Dissertations |
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