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Endogenous and exogenous factors affecting lipoprotein lipase activity
Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk biovetenskap, Fysiologisk kemi.
2014 (engelsk)Doktoravhandling, med artikler (Annet vitenskapelig)
Abstract [en]

Individuals with high levels of plasma triglycerides are at high risk to develop cardiovascular disease (CVD), currently one of the major causes of death worldwide. Recent epidemiological studies show that loss-of-function mutations in the APOC3 gene lower plasma triglyceride levels and reduce the incidence of coronary artery disease. The APOC3 gene encodes for apolipoprotein (APO) C3, known as an inhibitor of lipoprotein lipase (LPL) activity. Similarly, a common gain-of-function mutation in the LPL gene is associated with reduced risk for CVD.

LPL is central for the metabolism of lipids in blood. The enzyme acts at the endothelial surface of the capillary bed where it hydrolyzes triglycerides in circulating triglyceride-rich lipoproteins (TRLs) and thereby allows uptake of fatty acids in adjacent tissues. LPL activity has to be rapidly modulated to adapt to the metabolic demands of different tissues. The current view is that LPL is constitutively expressed and that the rapid modulation of the enzymatic activity occurs by some different controller proteins. Angiopoietin-like protein 4 (ANGPTL4) is one of the main candidates for control of LPL activity. ANGPTL4 causes irreversible inactivation through dissociation of the active LPL dimer to inactive monomers. Other proteins that have effects on LPL activity are the APOCs which are surface components of the substrate TRLs. APOC2 is a well-known LPL co-factor, whereas APOC1 and APOC3 independently inhibit LPL activity.

Given the important role of LPL for triglyceride homeostasis in blood, the aim of this thesis was to find small molecules that could increase LPL activity and serve as lead compounds in future drug discovery efforts. Another aim was to investigate the molecular mechanisms for how APOC1 and APOC3 inhibit LPL activity.

Using a small molecule screening library we have identified small molecules that can protect LPL from inactivation by ANGPTL4 during incubations in vitro. Following a structure-activity relationship study we have synthesized lead compounds that more efficiently protect LPL from inactivation by ANGPTL4 in vitro and also have dramatic triglyceride-lowering properties in vivo. In a separate study we show that low concentrations of fatty acids possess the ability to prevent inactivation of LPL by ANGPTL4 under in vitro conditions.

With regard to APOC1 and APOC3 we demonstrate that when bound to TRLs, these apolipoproteins prevent binding of LPL to the lipid/water interface. This results in decreased lipolysis and in an increased susceptibility of LPL to inactivation by ANGPTL4. We demonstrate that hydrophobic amino acid residues that are centrally located in the APOC3 molecule are critical for attachment of this protein to lipid emulsion particles and consequently for inhibition of LPL activity.

In summary, this work has identified a lead compound that protects LPL from inactivation by ANGPTL4 in vitro and lowers triglycerides in vivo. In addition, we propose a molecular mechanism for inhibition of LPL activity by APOC1 and APOC3.

sted, utgiver, år, opplag, sider
Umeå: Umeå University , 2014. , s. 56
Serie
Umeå University medical dissertations, ISSN 0346-6612 ; 1669
Emneord [en]
LPL, APOC1, APOC2, APOC3, ANGPTL4, enzyme inactivation, lipoprotein metabolism, triglycerides, fatty acids, hypertriglyceridemia, CVD, small molecule screening, structure-activity relationship
HSV kategori
Forskningsprogram
medicin, hjärt- och kärlforskning
Identifikatorer
URN: urn:nbn:se:umu:diva-91662ISBN: 978-91-7601-115-7 (tryckt)OAI: oai:DiVA.org:umu-91662DiVA, id: diva2:737648
Disputas
2014-08-27, NUS - Norrlands universitetssjukhus, Sal E04, Byggnad 6E, Umeå Universitet, Umeå, 14:00 (engelsk)
Opponent
Veileder
Tilgjengelig fra: 2014-08-15 Laget: 2014-08-13 Sist oppdatert: 2018-06-07bibliografisk kontrollert
Delarbeid
1. Apolipoproteins C-I and C-III Inhibit Lipoprotein Lipase Activity by Displacement of the Enzyme from Lipid Droplets
Åpne denne publikasjonen i ny fane eller vindu >>Apolipoproteins C-I and C-III Inhibit Lipoprotein Lipase Activity by Displacement of the Enzyme from Lipid Droplets
Vise andre…
2013 (engelsk)Inngår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 288, nr 47, s. 33997-34008Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

Apolipoproteins (apo) C-I and C-III are known to inhibit lipoprotein lipase (LPL) activity, but the molecular mechanisms for this remain obscure. We present evidence that either apoC-I or apoC-III, when bound to triglyceride-rich lipoproteins, prevent binding of LPL to the lipid/water interface. This results in decreased lipolytic activity of the enzyme. Site-directed mutagenesis revealed that hydrophobic amino acid residues centrally located in the apoC-III molecule are critical for attachment to lipid emulsion particles and consequently inhibition of LPL activity. Triglyceride-rich lipoproteins stabilize LPL and protect the enzyme from inactivating factors such as angiopoietin-like protein 4 (angptl4). The addition of either apoC-I or apoC-III to triglyceride-rich particles severely diminished their protective effect on LPL and rendered the enzyme more susceptible to inactivation by angptl4. These observations were seen using chylomicrons as well as the synthetic lipid emulsion Intralipid. In the presence of the LPL activator protein apoC-II, more of apoC-I or apoC-III was needed for displacement of LPL from the lipid/water interface. In conclusion, we show that apoC-I and apoC-III inhibit lipolysis by displacing LPL from lipid emulsion particles. We also propose a role for these apolipoproteins in the irreversible inactivation of LPL by factors such as angptl4.

Emneord
Apolipoproteins, Dyslipidemia, Lipolysis, Lipoprotein, Lipoprotein Metabolism, Triglyceride, Apolipoprotein C-I, Apolipoprotein C-III, Lipoprotein Lipase
HSV kategori
Identifikatorer
urn:nbn:se:umu:diva-85558 (URN)10.1074/jbc.M113.495366 (DOI)000327250200041 ()2-s2.0-84888335913 (Scopus ID)
Forskningsfinansiär
Swedish Research Council, 12203Swedish Heart Lung Foundation
Tilgjengelig fra: 2014-02-07 Laget: 2014-02-06 Sist oppdatert: 2023-03-24bibliografisk kontrollert
2. Fatty acids bind tightly to the N-terminal domain of angiopoietin-like protein 4 and modulate its interaction with lipoprotein lipase
Åpne denne publikasjonen i ny fane eller vindu >>Fatty acids bind tightly to the N-terminal domain of angiopoietin-like protein 4 and modulate its interaction with lipoprotein lipase
Vise andre…
2012 (engelsk)Inngår i: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 287, nr 35, s. 29739-29752Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

Angiopoietin-like protein 4 (Angptl4), a potent regulator of plasma triglyceride metabolism, binds to lipoprotein lipase (LPL) through its N-terminal coiled-coil domain (ccd-Angptl4) inducing dissociation of the dimeric enzyme to inactive monomers. In the present study we demonstrate that fatty acids reduce the inactivation of LPL by Angptl4. This was the case both with ccd-Angptl4 and full length Angptl4 and the effect was seen in human plasma or in the presence of albumin. The effect decreased in the sequence oleic acid > palmitic acid > myristic acid >linoleic acid >linolenic acid. Surface plasmon resonance, isothermal titration calorimetry, fluorescence and chromatography measurements revealed that fatty acids bind with high affinity to ccd-Angptl4. The interactions were characterized by fast association and slow dissociation rates, indicating formation of stable complexes. The highest affinity for ccd-Angptl4 was detected for oleic acid with a sub-nanomolar equilibrium dissociation constant (Kd). The Kd values for palmitic and myristic acid were in nanomolar range. Linoleic and linolenic acid bound with much lower affinity. On binding of fatty acids, ccd-Angptl4 underwent conformational changes resulting in a decreased helical content, weakened structural stability, dissociation of oligomers, and altered fluorescence properties of the Trp38 residue which is located close to the putative LPL-binding region. Based on these results, we propose that fatty acids play an important role in modulating the effects of Angptl4.

sted, utgiver, år, opplag, sider
Bethesda: American Society for Biochemistry and Molecular Biology, 2012
HSV kategori
Identifikatorer
urn:nbn:se:umu:diva-57336 (URN)10.1074/jbc.M111.303529 (DOI)000308286900050 ()22773878 (PubMedID)2-s2.0-84865455762 (Scopus ID)
Tilgjengelig fra: 2012-07-13 Laget: 2012-07-13 Sist oppdatert: 2023-03-23bibliografisk kontrollert
3. Identification of a small molecule that stabilizes lipoprotein lipase in vitro and lowers triglycerides in vivo
Åpne denne publikasjonen i ny fane eller vindu >>Identification of a small molecule that stabilizes lipoprotein lipase in vitro and lowers triglycerides in vivo
Vise andre…
2014 (engelsk)Inngår i: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 450, nr 2, s. 1063-1069Artikkel i tidsskrift (Fagfellevurdert) Published
Abstract [en]

Patients at increased cardiovascular risk commonly display high levels of plasma triglycerides (TGs) levels, elevated LDL cholesterol, small dense LDL particles and low levels of HDL-cholesterol. Many remain at high risk even after successful statin therapy, presumably because TG levels remain high. Lipoprotein lipase (LPL) maintains TG homeostasis in blood by hydrolysis of TG-rich lipoproteins. Efficient clearance of TGs is accompanied by increased levels of HDL-cholesterol and decreased levels of small dense LDL. Given the central role of LPL in lipid metabolism we sought to find small molecules that could increase LPL activity and serve as starting points for drug development efforts against cardiovascular disease. Using a small molecule screening approach we have identified small molecules that can protect LPL from inactivation by the controller protein angiopoietin-like protein 4 during incubations in vitro. One of the selected compounds, 50F10, was directly shown to preserve the active homodimer structure of LPL, as demonstrated by heparin-Sepharose chromatography. This compound tended to reduce fasting TG levels in normal rats. On injection to hypertriglyceridemic apolipoprotein A-V deficient mice the compound ameliorated the postprandial response after an olive oil gavage. This compound is a potential lead compound for the development of drugs that could reduce the residual risk associated with elevated TGs in dyslipidemia.

sted, utgiver, år, opplag, sider
Elsevier, 2014
Emneord
Lipoprotein lipase, Angiopoietin-like protein 4, Hypertriglyceridemia, Lipoprotein metabolism, Cardiovascular disease, Small molecule screening
HSV kategori
Identifikatorer
urn:nbn:se:umu:diva-91689 (URN)10.1016/j.bbrc.2014.06.114 (DOI)000339861200023 ()24984153 (PubMedID)2-s2.0-84905109095 (Scopus ID)
Forskningsfinansiär
Swedish Research CouncilVINNOVAKnut and Alice Wallenberg FoundationCarl Tryggers foundation
Tilgjengelig fra: 2014-08-13 Laget: 2014-08-13 Sist oppdatert: 2023-03-24bibliografisk kontrollert

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