Physiology

MARIANA NIKOLOVA KARAKASHIAN, Ph.D. Professor Ph.D. Bulgarian Academy of Science, 1992 Office: MS-579 Medical Center 0298 Tel: (859) 323-8210 Lab: MS-531 Tel: (859) 323-1115 E-mail: mnikolo@uky.edu Curriculum Vita (pdf)

Sphingolipids are diverse class of lipid molecules that participate in regulation of various cell functions. Sphingomyelin is one of the main sphingolipids in mammalian cells and can be hydrolyzed to ceramide by a family of enzymes termed sphingomyelinases. Pro-inflammatory cytokines, oxidative stress, and aging control sphingomyelinase activity, thus generating quantities of ceramide, which in turn acts to regulate distinct down-stream targets resulting in a variety of cellular responses.  Our lab uses state-of the art molecular and biochemical approach to understand the mechanisms that determines the activity of sphingomyelinases, specifically that of neutral sphingomyelinase-2 (smpd3), and to define new ceramide-dependent down-stream targets in the context of the IL-1b signaling pathway.
In addition to mediating signaling pathways, sphingolipids play a role in maintaining lipid homeostasis of the cells.  Our ongoing study suggests that disruption of sphingolipid balance in the liver has strong influence on the partitioning of the fat into neutral lipids, the development of steatotic liver and respectively insulin resistance and hyperglycemia. Our goal is by combining classical in vitro/in vivo approach to decipher the coordinated regulation of sphingolipid, glycerophospholipid, and TAG metabolism and define new mechanisms that maintain sphingolipid homeostasis in cells.
Our long-term objective is to apply the results of our research towards better understanding the role of lipids in the onset of key liver pathologies associated with aging, including fatty liver and chronic inflammation

Techniques often used in the lab:

Biochemistry: Western blot,  protein purification, co-immunoprecipitation;  Real time and classical RT-PCR, gel-shift analyses, enzyme activity assays, lipid extractions, and lipid analysis by  TLC, HPLC, and MasSpectrometry. Sub-cellular fractionation.

Molecular biology:  Cloning and expresison of various genes of interest in bacterial and mammalian systems using plasmids or adenoviruses, site-directed mutagenesis.

Cell biology:  Maintaining of different cell lines. Isolation of primary hepatocytes from rats and mice. Creation and work with stably transfected cell lines. Transient overexpression or silencing of proteins in mammalian cells by plasmid- or adenovirus-mediated gene transfer.  Fluorescent microscopy of subcellular organelles. Indirect immunofluorescence.Receptor internalization and ligand-receptor interactions.

Work with small animals:  Breeding of transgenic animals, genotyping, In vivo gene overexpression/silencing through adenoviral-mediated gene trasnfer, Mouse and rat modes of inflammation (LPS administration), diet-induced diabetes.  Organ and tissue collection.

Key publications:

Interleukin 1β Regulation of FoxO1 Protein Content and Localization: Evidence for a Novel Ceramide-dependent Mechanism.  Dobierzewska A, Shi L, Karakashian AA, Nikolova-Karakashian MN. J Biol Chem. 2012 Oct 26. [Epub ahead of print]. http://www.ncbi.nlm.nih.gov/pubmed/23105097

Characterization of secretory sphingomyelinase activity, lipoprotein sphingolipid content and LDL aggregation in ldlr-/- mice fed on a high-fat diet. Deevska GM, Sunkara M, Morris AJ, Nikolova-Karakashian MN. Biosci Rep. 2012 Oct;32(5):479-90. http://www.ncbi.nlm.nih.gov/pubmed/22712892

Protein phosphatase 2A and neutral sphingomyelinase 2 regulate IRAK-1 Ubiquitination and degradation in response to IL-1{beta} Dobierzewska A, Giltiay NV, Sabapathi S, Karakashian AA, Nikolova-Karakashian MN. J Biol Chem. 2011 Jun 27. [Epub ahead of print]
http://www.ncbi.nlm.nih.gov/pubmed/21708940

Sphingolipid Metabolism, Oxidant Signaling, and Contractile Function of Skeletal Muscle. Nikolova-Karakashian MN, Reid MB. Antioxid Redox Signal. 2011 Jun 8. [Epub ahead of print]
http://www.ncbi.nlm.nih.gov/pubmed/21453197

Ceramide in stress response. Nikolova-Karakashian MN, Rozenova KA. Adv Exp Med Biol. 2010;688:86-108. Review.
http://www.ncbi.nlm.nih.gov/pubmed/20919648

The twists and turns of sphingolipid pathway in glucose regulation. Deevska GM, Nikolova-Karakashian MN. Biochimie. 2011 Jan;93(1):32-8. Epub 2010 May 31. Review.
http://www.ncbi.nlm.nih.gov/pubmed/20561942

Studies on the role of acid sphingomyelinase and ceramide in the regulation of tumor necrosis factor alpha (TNFalpha)-converting enzyme activity and TNFalpha secretion in macrophages. Rozenova KA, Deevska GM, Karakashian AA, Nikolova-Karakashian MN. J Biol Chem. 2010 Jul 2;285(27):21103-13. Epub 2010 Mar 17.
http://www.ncbi.nlm.nih.gov/pubmed/20236926

Acid Sphingomyelinase Deficiency Prevents Diet-induced Hepatic Triacylglycerol Accumulation and Hyperglycemia in Mice. Deevska GM, Rozenova KA, Giltiay NV, Chambers MA, White J, Boyanovsky BB, Wei J, Daugherty A, Smart EJ, Reid MB, Merrill AH Jr, Nikolova-Karakashian M. J Biol Chem. 2009 Mar 27;284(13):8359-68. Epub 2008 Dec 11.
http://www.ncbi.nlm.nih.gov/pubmed/19074137

Rutkute K, Asmis RH, Nikolova-Karakashian MN. Regulation of neutral sphingomyelinase-2 by GSH: a new insight to the role of oxidative stress in aging-associated inflammation. J Lipid Res. 48(11):2443-52, 2007.

Rutkute K, Karakashian AA, Giltiay NV, Dobierzewska A, Nikolova-Karakashian MN. Aging in rat causes hepatic hyperresposiveness to interleukin-1beta which is mediated by neutral sphingomyelinase-2. Hepatology. 46(4):1166-1176, 2007.

Rutkute K, Nikolova-Karakashian MN. Regulation of insulin-like growth factor binding protein-1 expression during aging. Biochem Biophys Res Commun. 361(2): 263-9, 2007.

Other Publications