Open Access Lazzarini et al., J Chromatogr Sep Tech 2014, 5:5 DOI: 10.4172/2157-7064.1000239 Open Access Volume 5 • Isse 5 • 1000239 J Chromatogr Sep Tech ISSN: 2157-7064 JCGST, an open access journal Analysis of Serum Sphingomyelin Species by Uflc-Ms/Ms in Patients Affected with Monoclonal Gammopathy Lazzarini A 1 , Floridi A 1 , Pugliese L 1 , Villani M 1 , Cataldi S 1 , Michela Codini 2 , Lazzarini R 1 , Beccari T 2 , Ambesi-Impiombato FS 3 , Curcio F 3 and Albi E 1 * 1 Laboratory of Nuclear Lipid BioPathology, CRABiON, Perugia, Italy 2 Department of Pharmaceutical Science, University of Perugia, Italy 3 Department of Clinical and Biological Sciences, University of Udine, Italy *Corresponding author: Albi E, Laboratory of Nuclear Lipid BioPathology, CRABiON, Perugia, Italy; Tel: 0039-075-5173544; Fax: 0039-075-5173544 e-mail: elisabetta.albi@yahoo.com Received September 11, 2014; Accepted September 11, 2014; Published October 03, 2014 Citation: Lazzarini A, Floridi A, Pugliese L, Villani M, Cataldi S, et al. (2014) Analysis of Serum Sphingomyelin Species by Ufc-Ms/Ms in Patients Affected with Monoclonal Gammopathy. J Chromatogr Sep Tech 5: 239. doi:10.4172/2157- 7064.1000239 Copyright: © 2014 Lazzarini A, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Abstract Cancer cells are hungry of cholesterol incorporated from serum with avidity and used to favour the expressions of proteins involved in cell proliferation such as RNA polymerase II, STAT3, PKCz and cyclin D1. Numerous studies have shown that exists a strong interaction between unesterifed cholesterol and saturated fatty acid sphingomyelin which arises from the Van der Waals interaction. Since sphingomyelin and cholesterol association is responsible for the formation of membrane lipid raft involved in cell signalling, we studied the possible hyposphingomyelinemia associated to hypocholesterolemia in the patients with cancer. The blood of 23 patients with monoclonal gammopathy were analyzed for cholesterol, 12:0 sphingomyelin, 16:0 sphingomyelin and 18:1sphingomyelin content. The results demonstrated that only the patients with very low level of cholesterol (65-99 mg/dl) had low amount of sphingomyelin and, in particular, of saturated sphingomyelin specie (16:0 sphingomyelin). The possibility that the hypocholesterolemia in cancer was secondary to hyposphingomyelinemia was discussed. Keywords: Cholesterol; Sphingomyelin; MS/MS-UFLC; Blood lipids; monoclonal gammopathy Introduction Lipids are structural and functional molecules localized in cell membranes and in the nucleus where interact with active chromatin and play specifc roles in cell proliferation, diferentiation, apoptosis and cancer [1]. Hypocholesterolemia is common in patients with various malignant diseases since cancer cells are hungry of cholesterol (CHO) [2]. In fact, CHO is incorporated in cancer cells from serum with avidity and used to favour the expressions of proteins involved in cell proliferation such as RNA polymerase II, STAT3, PKCz and cyclin D1 [2]. In cells, CHO is not randomly distributed over the membranes but forms lipid microdomains thank to its link with saturated fatty acid of sphingomyelin (SM) by Van der Waals interaction [3]. Te lipid microdomains, also known as lipid rafs, are platforms for specifc proteins involved in cell signaling if they are localized in cell membrane and are platforms for DNA duplication and transcription if they are associated to the inner nuclear membrane [4,5]. It has been demonstrated that lipid rafs play important mediating roles in cell migration, metastasis, cell survival and tumor progression [6]. In particular CHO has an important role in regulating the synthesis of invadopodia: protrusion in the cell membrane of some cells, displaying properties of SM and CHO-enriched membranes or lipid rafs frequently seen in invasive and metastatic cancer cells that invade surrounding tissues [7]. It remains unclear how the rafs are formed in cancer cells, i.e., if using lipids that incorporate from the blood, reducing their physiological values. Tere is evidence that low levels of serum CHO-phospholipids are associated with the antiphospholipid antibodies in monoclonal gammopathy [8]. However, while the hypocholesterolemia is widely studied in cancer nothing is known about the possible change in the content of SM in the blood of cancer patients. Tus, we aimed to study the possible association of hypocholesterolemia with hyposphingomyelinemia in the patients with monoclonal gammopathy (MG). We analysed the main species of SM by SM/SM-UFLC in order to highlight the changes of saturated fatty acid SMs that interact specifcally with CHO. Materials and Methods Patients Blood samples from patients afected by monoclonal gammopathy (MG), with diagnosis from the “Laboratorio Centralizzato di analisi chimico-cliniche Ospedale Silvestrini, Perugia”, were collected over a 24 month period. Young and without poor nutrition state or extremely dietary control patients were chosen as experimental model. From all patients (62), only those with a low level of total CHO (23 patients) were analyzed. Te population was composed of 13 males and 10 females, average age was 36 yrs. Te control group was composed of 20 healthy blood donors: 12 males and 8 females, average age 39 yrs, with normal serum CHO levels. Lipid extraction Lipid extraction was performed according to Matyash et al. [9]. 1 mL of serum was diluted with 1mL methanol. 3 mL ultra pure water and 3 mL methyl tert -butyl ether (MTBE) were added. Each Sample was vortexed for 1 min and centrifuged at 3000 g for 5 min. Te supernatant was recovered. Te extraction with MTBE was repeated on the pellet and the supernatant was added to the frst. Te organic phase was dried under nitrogen fow and resuspended in 500 µL of methanol. Research Article Journal of Chromatography Separation Techniques J o u r n a l o f C h r o m a t o g r a p h y & S e p a r a t i o n T e c h n i q u es ISSN: 2157-7064