Publication Date

2013

Document Type

Honors Thesis

Department

Biological Sciences

Keywords

Proteomics, Muscle cells-Sex differences, Myoblasts, Mice, Musculuoskeletal system-Sex differences, Estrogen-Physiological effect, C2C12 cells, Sex hormones, Skeletal muscle, Cellular pathways

Abstract

Skeletal muscle tissue of males and females differs in ways beyond what can be appreciated visually. The sex hormone estrogen has been shown to impact lipid metabolism, apoptotic mechanisms, and glucose transport in muscle cells (Wend et al., 2012). Tamoxifen (Tmx) is a drug that binds estrogen receptors α and β, competitively inhibiting estrogen and its effects (Barkhem et al., 1998). Proteomic analysis provides a means of quantifying the differences in protein expression between tissues or cells that have been exposed to one or more unique conditions. Murine myoblasts (C2C12 cells) were grown in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) with or without 5 x 10-7 M Tmx. The protein extract of these cells was separated by isoelectric focusing between pH 5 and 8 on immobile pH gradient (IPG) strips, then separated further by molecular weight using sodium dodecyl sulfatepolyacrylamide gel electrophoresis (SDS-PAGE). The resulting two-dimensional gels were analyzed using PDQuest software (BioRad, ver. 8.0.1), and significant (p<0.05) protein spots that changed +/- two-fold compared to corresponding control group spots were digested with trypsin and analyzed by mass spectrometry (LC-MS/MS). Using Sequest software (BioRad, Thermo, 3.3.1 SP1), two of the analyzed spots were identified and three were predicted. Upregulation of proteins involved in the cytoskeleton and ion transport and metabolism was observed in C2C12 cells treated with tamoxifen, while proteins involved in carbohydrate transport and metabolism, cytoskeleton, and energy production and conversion were downregulated relative to expression levels observed in controls.

Language

English

Comments

vii, 54 p. : ill. (some col.) Honors project, Smith College, 2013. Includes bibliographical references (p. 45-54)

Share

COinS