Natural pentacyclic triterpenes enhance antitumoral effects of doxorubicin through augmenting its intracellular concentration in human sarcoma cells

Abstracts / Chemistry and Physics of Lipids 163S (2010) S57–S65

S59

ASMase deficiency decreases fat mass but exacerbates the deleterious effects of DIO leading to hyperglycaemia and liver damage.

suggest a potential role caveolin-1 in mitochondrial cholesterol downregulation.

doi:10.1016/j.chemphyslip.2010.05.175

doi:10.1016/j.chemphyslip.2010.05.176

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Cholesterol regulates mitochondrial raft-like domains during TNF/Fas-mediated hepatocellular apoptosis

Natural pentacyclic triterpenes enhance antitumoral effects of doxorubicin through augmenting its intracellular concentration in human sarcoma cells

Nuria Matías 1,2,3 , Anna Fernández 1,2,3 , Valeria Manganelli 1,2,3 , Anna Colell 1,2,3 , Antonella Tinari 1,2,3 , Walter Malorni 1,2,3 , Carmen Garcia-Ruiz 1,2,3 , Mauricio Sorice 1,2,3 , José C. FernándezCheca 1,2,3 1 Liver Unit, Hospital Clinic, CIBEREHD, IDIBAPS, and IIBB-CSIC, Barcelona, Spain 2 Department of Experimental Medicine, “Sapienza” University of Rome, Italy 3 Department of Technology and Health, Istituto Superiore di Sanitá, Rome, Italy

Lipid rafts constitute distinctive domains of biological membranes characterized by a specific lipid composition enriched in cholesterol and sphingolipids, which determine their detergent insolubility. These domains are thought to play a critical role in signal transduction and apoptosis, particularly in the plasma membrane by providing a platform for specific interactions of membrane receptors with their corresponding ligands and adaptor molecules. However, during death receptor (Fas/TNF)-mediated apoptosis of lymphocytes mitochondrial raft-like domains have been described and characterized by a multimolecular signaling complex that recruits proapoptotic Bcl-2 family proteins. Since cholesterol is a key component of raft-like domains and its trafficking to mitochondria has been shown to regulate hepatocellular susceptibility to TNF/Fas, the purpose of this study was to examine the role of cholesterol in the regulation of mitochondrial raft-like domains. Methods: Hepatic mitochondrial fractions were prepared from models of hepatic cholesterol loading, including cholesterol or alcohol feeding, transgenic Tg-SREBP-2 mice and NPC1 knockout mice. Mitochondrial cholesterol was determined by HPLC, while ganglioside GD3 was analyzed by immunoelectron microscopy and immunoTLC. Detergent soluble and insoluble mitochondrial fractions were processed for Western blotting for caveolin-1, human Fis 1 proteins (hFis1) and VDAC-1. Results: hFis1 and VDAC-1 levels in detergent-insoluble fractions were higher in mitochondria from mice fed the hypercholesterolemic diet (MitH) compared to fractions of control mice (MitC). Furthermore, cholesterol enrichment was associated with enhanced GD3 levels as examined by immunostaining and immunoelectron microscopy in MitH fractions. Interestingly, caveolin-1 was present in detergent-insoluble mitochondrial fractions only in MitH but not in MitC samples. Similar findings regarding the association of cholesterol and GD3, and the presence of hFis1, VDAC-1 and caveolin-1 in detergent-insoluble fractions were observed in mitochondria from Tg-SREBP-2 and NPC1 KO mice. In all these models, hepatocytes were highly susceptible to TNF/Fas-mediated apoptosis that was accompanied by increased GD3 levels and recruitment of Bax. Conclusions: These findings indicate the existence of raft-like domains in hepatic mitochondria in cholesterol loading models, which may be of significance in steatohepatitis due to the hepatocellular sensitization to TNF/Fas. The findings showing the correlation of mitochondrial caveolin-1 and cholesterol levels,

Victor H. Villar 1,∗ , Manuel Gómez 1 , Oliver Vögler 1 , Jordi M. Serra 1,2 , Javier Martín 1,3 , Valentina Ruiz-Gutiérrez 4 , Francisca Barceló 1 , Regina Alemany 1 1 Clinical and Translational Research Group, University of the Balearic Island, Ctra Valldemossa, km 7.5, 07122 Palma de Mallorca, Balearic Islands, Spain 2 Department of Hematology, University Hospital Son Dureta, Doria 55, 07014 Palma de Mallorca, Balearic Islands, Spain 3 Department of Oncology, University Hospital Son Dureta, Doria 55, 07014 Palma de Mallorca, Balearic Islands, Spain 4 Instituto de la Grasa, CSIC, Av. Padre García Tejero 4, 41012 Seville, Spain

Doxorubicin (DXR), an anthracycline antibiotic, is the actual standard chemotherapy for soft tissue sarcomas (STSs), a heterogeneous group of neoplasmas. However, its severe toxicity and the development of drug resistance call for clinically important adjuvants able to improve DXR efficacy without producing additional side effects. Oleanolic acid (OLA) and maslinic acid (MLA) are non-toxic pentacyclic triterpenes present in a wide range of plants and human dietary products (e.g., orujo olive oil), which exhibit certain beneficial biological activities, such as antioxidant, anti-inflammatory and antitumoral properties. Here, we examined the anti-proliferative effects of OLA and MLA, which were isolated from pressed olive (O. europaea) fruits, as single agents or in combination with DXR in human synovial sarcoma SW982 and leiomyosarcoma SK-UT-1 cells, both prone to DXR resistance. MLA, but not OLA, inhibited cell viability with an IC50 around 50 ␮M in both sarcoma cell lines. Combination of DXR (0.1–10 ␮M) with MLA (30 ␮M) or OLA (80 ␮M) increased the anti-proliferative effect of DXR by 1.5–2.5-fold. On the molecular level, OLA and MLA increased the uptake of DXR in a time-dependent manner, leading to a faster intracellular accumulation of DXR. Our results show that natural pentacyclic triterpenes enhance the antitumoral effects of DXR, most probably by increasing its intracellular concentration. doi:10.1016/j.chemphyslip.2010.05.177 PO115 Specificities of esterified fatty acids during lymphatic absorption of plasmalogens—The differences from those of diacylglycerophospholipids Hiroshi Hara 1,∗ , Megum Nishimukai 1 , Yuya Yamazaki 2 , Toru Nezu 2 , Ryouta Maeba 3 1

Division of Applied Bioscience, Research Faculty of Agriculture, Hokkaido University, Kita-9, Nishi-9, Kitaku, Sapporo 060-8589, Japan 2 Food Development Laboratory, ADEKA Co., Higashioku, Arakawaku, Tokyo 116-8554, Japan 3 Department of Biochemistry, Teikyo University School of Medicine, Kaga Itabashiku, Tokyo 173-8605, Japan A subclass of glycerophospholipid, plasmalogen, are widely distributed in animal and marine products, and presumed to play important roles for prevention of some degenerative diseases,