Role of AMP-protein kinase (AMPK) in regulation of perivascular adipose tissue (PVAT) function

Almabrouk, Tarek Ali Mohamed (2017) Role of AMP-protein kinase (AMPK) in regulation of perivascular adipose tissue (PVAT) function. PhD thesis, University of Glasgow.

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This thesis, entitled: ‘Role of AMP-protein kinase (AMPK) in regulation of perivascular
adipose tissue (PVAT) function’, has been submitted by author Tarek Ali Mohamed
Almabrouk for a degree of Doctor of Philosophy (PhD) in the College of Medical,
Veterinary and Life Sciences at the University of Glasgow, October 2016.

Apart from the cerebral circulation, all vasculature is surrounded by layers of adipose
tissue known as perivascular adipose tissue (PVAT). In health, PVAT can function as an
endocrine organ to produce a wide range of adipocytokines which can attenuate vascular
contraction. The exact mechanism of this anti-contractile effect is still ill-defined, although
much evidence suggests that PVAT-released adipocytokines may activate K+ channels on
VSMCs or eNOS on endothelial layer possibly via AMP-activated protein kinase (AMPK).
However, obesity results in oxidative stress and inflammation of the PVAT leading to
abnormal adipocytokine release and PVAT dysfunction. AMPK is a serine/threonine
kinase with many potential physiological functions, including regulation of energy
heamostasis. AMPK is expressed in the three layers of the blood vessel: smooth muscle
(VSM), the endothelium and PVAT and it is known that activation of AMPK leads to
vascular dilatation via both endothelium- and non-endothelium-dependent mechanisms.
Although it is known that AMPK can modulate VSM and endothelial function, it is
unknown whether AMPK can influence the anti-contractile activity of PVAT. Therefore,
this project aimed to investigate the mechanism of the anticontractile effect of PVAT by
determining the functions of AMPK within adipocytes, as well as to assess the importance
of vascular AMPK to the PVAT anti-contractile function.

Experiments were conducted using wild type (WT) and global AMPKα1 knockout (KO)
mice aortae. The phenotypic features of the PVAT were assessed by both histological,
immunohistochemical and immunofluorescent methods. Secretory function of the PVAT
was tested using an immunoblotting array and ELISA, whereas the anti-contractile effect
of PVAT was studied using wire myography. Immunoblotting methods were used to test
AMPK activity in the PVAT and VSMCs.
Aortic rings from WT and KO mice were denuded of endothelium and mounted on a wire
myograph in the presence and absence of PVAT. The responses to an AMPK activator
(AICAR) and the AMPK-independent vasodilator cromakalim were subsequently assessed.
Relaxation responses to AICAR or cromakalim in the Sv129 (wild type) mouse were

significantly enhanced in the presence of endogenous attached or unattached PVAT, an
effect that was absent in vessels from KO mice. Furthermore, enhanced relaxation was
observed in vessels from KO mice incubated with PVAT from Sv129 mice, whereas
PVAT from KO mice had no effect on relaxation of vessels from Sv129 mice.

Furthermore, conditioned medium (CM) transfer experiments demonstrated the presence of
an anticontractile factor released from PVAT that was absent in KO mice. Adiponectin
secretion was reduced in PVAT from KO mice and PVAT-enhanced relaxation was
attenuated in the presence of adiponectin blocking peptide. Adipokine array and ELISA
demonstrated that adiponectin release is significantly reduced in the KO conditioned media
in comparison with wild type CM. Globular adiponectin restores the relaxation response in
both wild type aortae without PVAT and in KO aortae with and without PVAT.
High fat diet (HFD) fed mice showed a reduction in the relaxation response to cromakalim
in wild type vessels with intact PVAT in comparison with animals fed a normal chow diet
(ND). HFD animals had increased inflammatory infiltrates in the PVAT which were
associated with reduced AMPK activity and adiponectin release in comparison with ND
fed WT mice. In KO mice, AMPK activity was also reduced and increased inflammatory
infiltration was observed in both ND and HFD mice.

In conclusion, the current project demonstrates that AMPKα1 has a critical role in
maintaining PVAT’s anti-contractile effect; likely mediated through altered adiponectin
secretion or sensitivity, and through protection of PVAT against inflammation. Marked
reduction in AMPK activity in WT PVAT, accompanied with the reduction in the release
of adiponectin in HFD and KO animal may explain the impaired vascular function
observed in obesity.

Item Type: Thesis (PhD)
Qualification Level: Doctoral
Keywords: Perivascular adipose tissue (PVAT), AMPK, adiponectin, anticontractile effect, High fat diet (HFD), vascular function.
Subjects: R Medicine > RM Therapeutics. Pharmacology
Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health
Supervisor's Name: Kennedy, Dr. Simon and Salt, Dr. Ian
Date of Award: 2017
Depositing User: Tarek Ali Mohamed Almabrouk
Unique ID: glathesis:2017-8178
Copyright: Copyright of this thesis is held by the author.
Date Deposited: 16 Jun 2017 07:30
Last Modified: 14 Jul 2017 14:53
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