Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.616372
Title: Regulation of inflammatory signalling in adipocytes by AMPK
Author: Mancini, Sarah J.
ISNI:       0000 0004 5346 8854
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 2014
Availability of Full Text:
Access from EThOS:
Access from Institution:
Abstract:
AMP-activated protein kinase (AMPK) has been proposed to be a potential therapeutic target for patients with Type 2 diabetes and the metabolic syndrome. While the role of AMPK in muscle and liver is relatively well-characterised, less is known about the role of AMPK in the other principal metabolic tissue, adipose. Obesity is associated with the chronic, sub-clinical inflammation of adipose tissue. Characteristic hypertrophic adipocytes and the elevated infiltration and activation of macrophages stimulate production of cytokines and chemokines, including tumour necrosis factor-alpha (TNF-a), interleukin-1 beta (IL-1beta), interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1). These have autocrine, paracrine and endocrine effects which have been suggested to play a key role in the development of peripheral insulin resistance. Increasing evidence suggests that AMPK has anti-inflammatory actions, independent of its effect on carbohydrate and lipid metabolism. Previous work in our laboratory has demonstrated that AMPK inhibits TNF-alpha-stimulated MCP-1 secretion and monocyte adhesion in endothelial cells. The role of AMPK in the regulation of inflammatory signalling in adipocytes is currently poorly characterised. To address this, the effect of AMPK activation on the phosphorylation of TNF-alpha/IL-1beta and IL-6 signalling pathway intermediates was initially assessed in cultured 3T3-L1 adipocytes. Furthermore, the molecular mechanism by which AMPK elicits these effects was investigated. In addition, the effect of AMPK activation on downstream functional consequences of proinflammatory signalling in 3T3-L1 adipocytes and RAW 264.7 macrophages were examined. Finally, the effect of macrophage AMPK activation on inflammation-induced insulin resistance in 3T3-L1 adipocytes was also investigated. A769662 and infection with adenovirus expressing a constitutively active AMPK mutant suppressed IL-1beta-stimulated NFkappaB nuclear translocation in 3T3-L1 adipocytes. Conversely, this was abrogated upon adenoviral expression of a dominant negative AMPK mutant. In line with this, phosphorylation of upstream IkappaBalpha and IKK were also ameliorated upon AMPK activation. In parallel, A769662-mediated AMPK activation inhibited TNF-alpha/IL-1beta-stimulated phosphorylation of JNK, ERK1/2 and p38 MAPKs in 3T3-L1 adipocytes. Furthermore, A769662-mediated inhibition of TNF-alpha/IL-1beta proinflammatory signalling was likely to be independent of endothelial nitric oxide synthase (eNOS) activation and subsequent nitric oxide production. The target of AMPK may be downstream of TAK1, as IKK, JNK and p38 are inhibited in response to both TNF-alpha and IL-1beta; however the mechanism by which AMPK elicits these effects remains to be elucidated. A769662-mediated AMPK activation inhibited phosphorylation of IL-6-stimulated STAT3 (signal transducer and activator of transcription 3) in 3T3-L1 adipocytes independently of phosphatase action, yet A769662 was unable to inhibit constitutive Janus kinase (JAK)-mediated phosphorylation of STAT3, suggesting AMPK may inhibit JAK activity. Inhibition of mTOR was found to suppress STAT3 phosphorylation in a manner mutually exclusive with A769662 stimulation, potentially via activation of T cell protein tyrosine phosphatase (TC-PTP). Adipose tissue from AMPKalpha1-/- mice demonstrated increased basal JNK and STAT3 phosphorylation, further providing evidence for an anti-inflammatory role for AMPK in adipose tissue. In 3T3-L1 adipocytes, A769662 abrogated cytokine-stimulated MCP-1 gene expression, and secretion of chemokines IP-10 (CXCL10), KC (CXCL1) and MCP-1. Furthermore, AMPK activation reduced secretion of IL-5, MCP-1 and MIP-1alpha, but not TNF-alpha, from proinflammatory RAW 264.7 macrophages. Preliminary results indicated that chronic IL-6 and acute TNF-alpha or IL-1beta exposure suppressed insulin-stimulated glucose transport in 3T3-L1 adipocytes. Conditioned medium from activated RAW 264.7 macrophages also inhibited 3T3-L1 adipocyte insulin sensitivity; however, prior AMPK activation failed to attenuate this, potentially as a result of the presence of TNF-alpha. Overall these results suggest that activation of AMPK inhibits activation of multiple distinct proinflammatory signalling pathways in adipocytes and macrophages. AMPK activation may suppress IL-6 signalling via regulation of JAK, while the AMPK-mediated inhibition of IKK and concomitant suppression of MAPKs in response to TNF-alpha/IL-1beta suggests TAK1 as a potential AMPK target. Finally, proinflammatory stimuli induce insulin resistance in adipocytes, however whether this can be rescued by AMPK activation remains to be fully elucidated.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.616372  DOI: Not available
Keywords: Q Science (General)
Share: