Publications by authors named "Madalina Duta-Mare"

10 Publications

  • Page 1 of 1

Loss of function of lysosomal acid lipase (LAL) profoundly impacts osteoblastogenesis and increases fracture risk in humans.

Bone 2021 Jul 7;148:115946. Epub 2021 Apr 7.

Maine Medical Center Research Institute, 81 Research Drive, Scarborough, ME 04074, USA; Center for Bone Biology, Division of Clinical Pharmacology, Vanderbilt University Medical Center, Nashville, TN 37232, USA; Department of Medicine, Division of Clinical Pharmacology, Vanderbilt University Medical Center, Nashville, TN 37232, USA. Electronic address:

Lysosomal acid lipase (LAL) is essential for cholesteryl ester (CE) and triacylglycerol (TAG) hydrolysis in the lysosome. Clinically, an autosomal recessive LIPA mutation causes LAL deficiency (LALD), previously described as Wolman Disease or Cholesteryl Ester Storage Disease (CESD). LAL-D is associated with ectopic lipid accumulation in the liver, small intestine, spleen, adrenal glands, and blood. Considering the importance of unesterified cholesterol and fatty acids in bone metabolism, we hypothesized that LAL is essential for bone formation, and ultimately, skeletal health. To investigate the role of LAL in skeletal homeostasis, we used LAL-deficient () mice, in vitro osteoblast cultures, and novel clinical data from LAL-D patients. Both male and female LAL mice demonstarted lower trabecular and cortical bone parameters , which translated to reduced biomechanical properties. Further histological analyses revealed that LAL mice had fewer osteoblasts, with no change in osteoclast or marrow adipocyte numbers. In studying the cell-autonomous role of LAL, we observed impaired differentiation of LAL calvarial osteoblasts and in bone marrow stromal cells treated with the LAL inhibitor lalistat. Consistent with LAL's role in other tissues, lalistat resulted in profound lipid puncta accumulation and an altered intracellular lipid profile. Finally, we analyzed a large de-identified national insurance database (i.e. 2016/2017 Optum Clinformatics®) which revealed that adults (≥18 years) with CESD (n = 3076) had a higher odds ratio (OR = 1.21; 95% CI = 1.03-1.41) of all-cause fracture at any location compared to adults without CESD (n = 13.7 M) after adjusting for demographic variables and osteoporosis. These data demonstrate that alterations in LAL have significant clinical implications related to fracture risk and that LAL's modulation of lipid metabolism is a critical for osteoblast function.
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http://dx.doi.org/10.1016/j.bone.2021.115946DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8108562PMC
July 2021

Presenilin-1 Established ER-Ca Leak: a Follow Up on Its Importance for the Initial Insulin Secretion in Pancreatic Islets and β-Cells upon Elevated Glucose.

Cell Physiol Biochem 2019 ;53(3):573-586

Molecular Biology and Biochemistry, Gottfried Schatz Research Center for Cellular Signaling, Metabolism & Aging, Medical University of Graz, Graz, Austria.

Background/aims: In our recent work, the importance of GSK3β-mediated phosphorylation of presenilin-1 as crucial process to establish a Ca leak in the endoplasmic reticulum and, subsequently, the pre-activation of resting mitochondrial activity in β-cells was demonstrated. The present work is a follow-up and reveals the importance of GSK3β-phosphorylated presenilin-1 for responsiveness of pancreatic islets and β-cells to elevated glucose in terms of cytosolic Ca spiking and insulin secretion.

Methods: Freshly isolated pancreatic islets and the two pancreatic β-cell lines INS-1 and MIN-6 were used. Cytosolic Ca was fluorometrically monitored using Fura-2/AM and cellular insulin content and secretion were measured by ELISA.

Results: Our data strengthened our previous findings of the existence of a presenilin-1-mediated ER-Ca leak in β-cells, since a reduction of presenilin-1 expression strongly counteracted the ER Ca leak. Furthermore, our data revealed that cytosolic Ca spiking upon administration of high D-glucose was delayed in onset time and strongly reduced in amplitude and frequency upon siRNA-mediated knock-down of presenilin-1 or the inhibition of GSK3β in the pancreatic β-cells. Moreover, glucose-triggered initial insulin secretion disappeared by depletion from presenilin-1 and inhibition of GSK3β in the pancreatic β-cells and isolated pancreatic islets, respectively.

Conclusion: These data complement our previous work and demonstrate that the sensitivity of pancreatic islets and β-cells to glucose illustrated as glucose-triggered cytosolic Ca spiking and initial but not long-lasting insulin secretion crucially depends on a strong ER Ca leak that is due to the phosphorylation of presenilin-1 by GSK3β, a phenomenon that might be involved in the development of type 2 diabetes.
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http://dx.doi.org/10.33594/000000158DOI Listing
December 2019

Glycogen Synthase Kinase 3 Beta Controls Presenilin-1-Mediated Endoplasmic Reticulum Ca²⁺ Leak Directed to Mitochondria in Pancreatic Islets and β-Cells.

Cell Physiol Biochem 2019 18;52(1):57-75. Epub 2019 Feb 18.

Molecular Biology and Biochemistry, Gottfried Schatz Research Center for Cellular Signaling, Metabolism & Aging, Medical University of Graz, Graz, Austria.

Background/aims: In pancreatic β-cells, the intracellular Ca²⁺ homeostasis is an essential regulator of the cells major functions. The endoplasmic reticulum (ER) as interactive intracellular Ca²⁺ store balances cellular Ca²⁺. In this study basal ER Ca²⁺ homeostasis was evaluated in order to reveal potential β-cell-specificity of ER Ca²⁺ handling and its consequences for mitochondrial Ca²⁺, ATP and respiration.

Methods: The two pancreatic cell lines INS-1 and MIN-6, freshly isolated pancreatic islets, and the two non-pancreatic cell lines HeLA and EA.hy926 were used. Cytosolic, ER and mitochondrial Ca²⁺ and ATP measurements were performed using single cell fluorescence microscopy and respective (genetically-encoded) sensors/dyes. Mitochondrial respiration was monitored by respirometry. GSK3β activity was measured with ELISA.

Results: An atypical ER Ca²⁺ leak was observed exclusively in pancreatic islets and β-cells. This continuous ER Ca²⁺ efflux is directed to mitochondria and increases basal respiration and organellar ATP levels, is established by GSK3β-mediated phosphorylation of presenilin-1, and is prevented by either knockdown of presenilin-1 or an inhibition/knockdown of GSK3β. Expression of a presenlin-1 mutant that mimics GSK3β-mediated phosphorylation established a β-cell-like ER Ca²⁺ leak in HeLa and EA.hy926 cells. The ER Ca²⁺ loss in β-cells was compensated at steady state by Ca²⁺ entry that is linked to the activity of TRPC3.

Conclusion: Pancreatic β-cells establish a cell-specific ER Ca²⁺ leak that is under the control of GSK3β and directed to mitochondria, thus, reflecting a cell-specific intracellular Ca²⁺ handling for basal mitochondrial activity.
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http://dx.doi.org/10.33594/000000005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6459368PMC
March 2019

Hepatocyte-specific lysosomal acid lipase deficiency protects mice from diet-induced obesity but promotes hepatic inflammation.

Biochim Biophys Acta Mol Cell Biol Lipids 2019 04 9;1864(4):500-511. Epub 2019 Jan 9.

Gottfried Schatz Research Center, Medical University of Graz, Graz, Austria; BioTechMed-Graz, Graz, Austria. Electronic address:

Lysosomal acid lipase (LAL) hydrolyzes cholesteryl esters (CE) and triglycerides (TG) to generate fatty acids (FA) and cholesterol. LAL deficiency (LAL-D) in both humans and mice leads to hepatomegaly, hypercholesterolemia, and shortened life span. Despite its essential role in lysosomal neutral lipid catabolism, the cell type-specific contribution of LAL to disease progression is still elusive. To investigate the role of LAL in the liver in more detail and to exclude the contribution of LAL in macrophages, we generated hepatocyte-specific LAL-deficient mice (Liv-Lipa) and fed them either chow or high fat/high cholesterol diets (HF/HCD). Comparable to systemic LAL-D, Liv-Lipa mice were resistant to diet-induced obesity independent of food intake, movement, and energy expenditure. Reduced body weight gain was mainly due to reduced white adipose tissue depots. Furthermore, Liv-Lipa mice exhibited improved glucose clearance during glucose and insulin tolerance tests compared to control mice. Analysis of hepatic lipid content revealed a massive reduction of TG, whereas CE concentrations were markedly increased, leading to CE crystal formation in the livers of Liv-Lipa mice. Elevated plasma transaminase activities, increased pro-inflammatory cytokines and chemokines as well as hepatic macrophage infiltration indicated liver inflammation. Our data provide evidence that hepatocyte-specific LAL deficiency is sufficient to alter whole-body lipid and energy homeostasis in mice. We conclude that hepatic LAL plays a pivotal role by preventing liver damage and maintaining lipid and energy homeostasis, especially during high lipid availability.
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http://dx.doi.org/10.1016/j.bbalip.2019.01.007DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6372077PMC
April 2019

N-acetylaspartate pathway is nutrient responsive and coordinates lipid and energy metabolism in brown adipocytes.

Biochim Biophys Acta Mol Cell Res 2019 03 31;1866(3):337-348. Epub 2018 Aug 31.

Institute of Biochemistry, Graz University of Technology, Graz, Austria; BioTechMed-Graz, Graz, Austria. Electronic address:

The discovery of significant amounts of metabolically active brown adipose tissue (BAT) in adult humans renders it a promising target for anti-obesity therapies by inducing weight loss through increased energy expenditure. The components of the N-acetylaspartate (NAA) pathway are highly abundant in BAT. Aspartate N-acetyltransferase (Asp-NAT, encoded by Nat8l) synthesizes NAA from acetyl-CoA and aspartate and increases energy expenditure in brown adipocytes. However, the exact mechanism how the NAA pathway contributes to accelerated mobilization and oxidation of lipids and the physiological regulation of the NAA pathway remained elusive. Here, we demonstrate that the expression of NAA pathway genes corresponds to nutrient availability and specifically responds to changes in exogenous glucose. NAA is preferentially produced from glucose-derived acetyl-CoA and aspartate and its concentration increases during adipogenesis. Overexpression of Nat8l drains glucose-derived acetyl-CoA into the NAA pool at the expense of cellular lipids and certain amino acids. Mechanistically, we elucidated that a combined activation of neutral and lysosomal (acid) lipolysis is responsible for the increased lipid degradation. Specifically, translocation of the transcription factor EB to the nucleus activates the biosynthesis of autophagosomes and lysosomes. Lipid degradation within lysosomes accompanied by adipose triglyceride lipase-mediated lipolysis delivers fatty acids for the support of elevated mitochondrial respiration. Together, our data suggest a crucial role of the NAA pathway in energy metabolism and metabolic adaptation in BAT.
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http://dx.doi.org/10.1016/j.bbamcr.2018.08.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6390944PMC
March 2019

Lysosomal Cholesterol Hydrolysis Couples Efferocytosis to Anti-Inflammatory Oxysterol Production.

Circ Res 2018 05 9;122(10):1369-1384. Epub 2018 Mar 9.

From the Institut National de la Santé et de la Recherche Médicale (Inserm) U1065, Université Côte d'Azur, Centre Méditerranéen de Médecine Moléculaire (C3M), Atip-Avenir, Fédération Hospitalo-Universitaire (FHU) Oncoage, Nice, France (M.V., S.I., L.-E.A., E.G., S.M., L.B., R.G., L.Y.-C.)

Rationale: Macrophages face a substantial amount of cholesterol after the ingestion of apoptotic cells, and the LIPA (lysosomal acid lipase) has a major role in hydrolyzing cholesteryl esters in the endocytic compartment.

Objective: Here, we directly investigated the role of LIPA-mediated clearance of apoptotic cells both in vitro and in vivo.

Methods And Results: We show that LIPA inhibition causes a defective efferocytic response because of impaired generation of 25-hydroxycholesterol and 27-hydroxycholesterol. Reduced synthesis of 25-hydroxycholesterol after LIPA inhibition contributed to defective mitochondria-associated membrane leading to mitochondrial oxidative stress-induced NLRP3 (NOD-like receptor family, pyrin domain containing) inflammasome activation and caspase-1-dependent Rac1 (Ras-related C3 botulinum toxin substrate 1) degradation. A secondary event consisting of failure to appropriately activate liver X receptor-mediated pathways led to mitigation of cholesterol efflux and apoptotic cell clearance. In mice, LIPA inhibition caused defective clearance of apoptotic lymphocytes and stressed erythrocytes by hepatic and splenic macrophages, culminating in splenomegaly and splenic iron accumulation under hypercholesterolemia.

Conclusions: Our findings position lysosomal cholesterol hydrolysis as a critical process that prevents metabolic inflammation by enabling efficient macrophage apoptotic cell clearance.
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http://dx.doi.org/10.1161/CIRCRESAHA.117.312333DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6034181PMC
May 2018

Lysosomal acid lipase regulates fatty acid channeling in brown adipose tissue to maintain thermogenesis.

Biochim Biophys Acta Mol Cell Biol Lipids 2018 Apr 31;1863(4):467-478. Epub 2018 Jan 31.

Gottfried Schatz Research Center, Medical University of Graz, Graz, Austria; BioTechMed-Graz, Graz, Austria. Electronic address:

Lysosomal acid lipase (LAL) is the only known enzyme, which hydrolyzes cholesteryl esters and triacylglycerols in lysosomes of multiple cells and tissues. Here, we explored the role of LAL in brown adipose tissue (BAT). LAL-deficient (Lal-/-) mice exhibit markedly reduced UCP1 expression in BAT, modified BAT morphology with accumulation of lysosomes, and mitochondrial dysfunction, consequently leading to regular hypothermic events in mice kept at room temperature. Cold exposure resulted in reduced lipid uptake into BAT, thereby aggravating dyslipidemia and causing life threatening hypothermia in Lal-/- mice. Linking LAL as a potential regulator of lipoprotein lipase activity, we found Angptl4 mRNA expression upregulated in BAT. Our data demonstrate that LAL is critical for shuttling fatty acids derived from circulating lipoproteins to BAT during cold exposure. We conclude that inhibited lysosomal lipid hydrolysis in BAT leads to impaired thermogenesis in Lal-/- mice.
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http://dx.doi.org/10.1016/j.bbalip.2018.01.011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839464PMC
April 2018

Impact of Endothelial Lipase on Cholesterol Efflux Capacity of Serum and High-density Lipoprotein.

Sci Rep 2017 10 2;7(1):12485. Epub 2017 Oct 2.

Institute of Molecular Biology and Biochemistry, Center of Molecular Medicine, Medical University Graz, Neue Stiftingtalstraße 6/6, 8010, Graz, Austria.

Endothelial lipase (EL) is a potent modulator of the structural and functional properties of HDL. Impact of EL on cholesterol efflux capacity (CEC) of serum and isolated HDL is not well understood and apparently contradictory data were published. Here, we systematically examined the impact of EL on composition and CEC of serum and isolated HDL, in vitro and in vivo, using EL-overexpressing cells and EL-overexpressing mice. CEC was examined in a validated assay using H-cholesterol labelled J774 macrophages. In vitro EL-modification of serum resulted in complex alterations, including enrichment of serum with lipid-free/-poor apoA-I, decreased size of human (but not mouse) HDL and altered HDL lipid composition. EL-modification of serum increased CEC, in line with increased lipid-free/-poor apoA-I formation. In contrast, CEC of isolated HDL was decreased likely through altered lipid composition. In contrast to in vitro results, EL-overexpression in mice markedly decreased HDL-cholesterol and apolipoprotein A-I serum levels associated with a decreased CEC of serum. HDL lipid composition was altered, but HDL particle size and CEC were not affected. Our study highlights the multiple and complex effects of EL on HDL composition and function and may help to clarify the seemingly contradictory data found in published articles.
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http://dx.doi.org/10.1038/s41598-017-12882-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5624901PMC
October 2017

Lysosomal lipid hydrolysis provides substrates for lipid mediator synthesis in murine macrophages.

Oncotarget 2017 Jun;8(25):40037-40051

Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.

Degradation of lysosomal lipids requires lysosomal acid lipase (LAL), the only intracellular lipase known to be active at acidic pH. We found LAL to be expressed in murine immune cells with highest mRNA expression in macrophages and neutrophils. Furthermore, we observed that loss of LAL in mice caused lipid accumulation in white blood cells in the peripheral circulation, which increased in response to an acute inflammatory stimulus. Lal-deficient (-/-) macrophages accumulate neutral lipids, mainly cholesteryl esters, within lysosomes. The cholesteryl ester fraction is particularly enriched in the PUFAs 18:2 and 20:4, important precursor molecules for lipid mediator synthesis. To investigate whether loss of LAL activity affects the generation of lipid mediators and to eliminate potential systemic effects from other cells and tissues involved in the pronounced phenotype of Lal-/- mice, we treated macrophages from Wt mice with the LAL-specific inhibitor LAListat-2. Acute inhibition of LAL resulted in reduced release of 18:2- and 20:4-derived mediators from macrophages, indicating that lipid hydrolysis by LAL is an important source for lipid mediator synthesis in macrophages. We conclude that lysosomes should be considered as organelles that provide precursor molecules for lipid mediators such as eicosanoids.
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http://dx.doi.org/10.18632/oncotarget.16673DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522325PMC
June 2017

Monoglyceride lipase deficiency affects hepatic cholesterol metabolism and lipid-dependent gut transit in ApoE-/- mice.

Oncotarget 2017 May;8(20):33122-33136

Institute of Molecular Biology and Biochemistry, Medical University of Graz, Graz, Austria.

Monoglyceride lipase (MGL) hydrolyzes monoglycerides (MGs) to glycerol and fatty acids. Among various MG species MGL also degrades 2-arachidonoylglycerol (2-AG), the most abundant endocannabinoid and potent activator of cannabinoid receptors (CBR) 1 and 2. MGL-knockout (-/-) mice exhibit pronounced 2-AG accumulation, but lack central cannabimimetic effects due to CB1R desensitization. We have previously shown that MGL affects plaque stability in apolipoprotein E (ApoE)-/- mice, an established animal model for dyslipidemia and atherosclerosis. In the current study, we investigated functional consequences of MGL deficiency on lipid and energy metabolism in ApoE/MGL double knockout (DKO) mice. MGL deficiency affected hepatic cholesterol metabolism by causing increased cholesterol elimination via the biliary pathway. Moreover, DKO mice exhibit lipid-triggered delay in gastric emptying without major effects on overall triglyceride and cholesterol absorption. The observed phenotype of DKO mice is likely not a consequence of potentiated CB1R signaling but rather dependent on the activation of alternative signaling pathways. We conclude that MGL deficiency causes complex metabolic changes including cholesterol metabolism and regulation of gut transit independent of the endocannabinoid system.
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http://dx.doi.org/10.18632/oncotarget.16529DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5464855PMC
May 2017