Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.634624
Title: A study of the P2X7 purinoceptor and vascular ATP metabolic pathways in chronic kidney disease-associated arterial calcification
Author: Fish, R. S.
ISNI:       0000 0004 5351 729X
Awarding Body: University College London (University of London)
Current Institution: University College London (University of London)
Date of Award: 2014
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Abstract:
The risk of cardiovascular-related death is several-fold higher in patients with chronic kidney disease (CKD) compared with the general population. Arterial calcification (AC) is extremely common in patients with CKD and strongly associates with cardiovascular-related mortality, however, there are currently no specific treatments to prevent its development and/or progression. Abundant evidence now suggests that AC is cell-mediated and actively-regulated, involving mechanisms linked to bone homeostasis, production of calcification inhibitors and vascular smooth muscle cell (VSMC) function. The P2X7 receptor (P2X7R) is an ATP-sensitive cation channel which has been implicated in several biological processes, in non-vascular contexts, thought to be important in the aetiology of AC. In addition, disruption to the normal function of some enzymes involved in ATP metabolism has been shown to contribute to AC, although little is known about their role in CKD-related arterial calcium deposition. The work in this thesis tested the primary hypothesis that P2X7R contributes to the pathogenesis of CKD-associated AC. Preliminary work was also conducted to examine the expression of components of the ATP-metabolising system in this clinical setting. P2X7R expression was confirmed in human and rodent vascular smooth muscle but was un-affected by calcification. In vitro, the P2X7R-specific antagonist, A438079, did not influence calcium deposition occurring in the presence of human VSMCs or segments of rat aorta exposed to ‘calcification-promoting’ medium. Calcification of cultured rat aorta was also not influenced by a second P2X7R-specific antagonist, A839977, or by BzATP (a receptor agonist). Aortic rings from mice deficient in P2X7R calcified to a similar extent to wild-type controls in vitro. A novel, adenine-based mouse model was developed to evaluate the effect of P2X7R gene deficiency on CKD-associated AC in vivo. However, the number of mice exhibiting AC in the final experiment was too low to draw any firm conclusion. Therefore, rats were fed an adenine-containing, high phosphate diet for 4 weeks (to induce CKD and AC) and administered a selective P2X7R antagonist, twice daily, throughout this period. Pharmacological blockade of P2X7R did not influence the magnitude of aortic calcification in this model. Quantification of mRNA performed on tissue obtained from the in vivo rat experiment suggested that VSMC-specific markers are down-regulated in calcified arteries, although VSMC osteogenic transformation, which is widely reported in the literature to occur in the context of AC, was not detected. Expression of the apoptosis marker, caspase-3, was increased in calcified arteries in vivo. P2X7R blockade did not influence any of these changes in mRNA expression. Expression of mRNA for ENPP-1, an ATP-metabolising enzyme responsible for the generation of the calcification inhibitor, pyrophosphate (PPi), was significantly increased in calcified arteries from CKD rats. Functional activity of ENPP-1 was also increased in these vessels. The expression of mRNA for other components of the ATP-metabolising system was also in keeping with an attempt by VSMCs to generate more PPi, possibly as an adaptive, defensive response to uraemic, calcification-promoting factors. Furthermore, an increase in ENPP-1 mRNA expression was detected in calcified inferior epigastric arteries from patients with end-stage renal disease (extracted at the time of kidney transplantation). In summary, P2X7R does not appear to contribute to the pathogenesis of CKD-associated AC, although this should be confirmed in experimental models which more closely simulate human disease. Arterial expression of enzymes involved in the metabolism of ATP does seem to change in AC. Future work should therefore focus on gauging the clinical relevance of this in order to better understand the mechanisms underlying the disease and potentially develop new therapeutic interventions.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.634624  DOI: Not available
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