Proteomic Profiling of Aging and Omega-3 Supplementation in Skeletal Muscle

Author

Francesco Maria Del Re, Smith College

Publication Date

2021-5

First Advisor

Stylianos P. Scordilis

Document Type

Masters Thesis

Degree Name

Master of Science

Department

Biological Sciences

Keywords

Skeletal Muscle, Aging, Omega-3 Diet, Sarcopenia, Proteomics, Mass Spectrometry

Abstract

Sarcopenia is one of the most relevant geriatric syndromes clinically leading to an age-associated loss of muscle. The impact of sarcopenia at a socioeconomical level has been estimated to as $18.5 billion in 2004 (Janssen et al., 2004b), and it is predicted to sharply rise by 2050 when the geriatric population (≥ 65 years old) will reach 1.6 billion people (McCormick and Vasilaki, 2018). To date, there is no treatment that can prevent, delay, or reverse the damaging effects of sarcopenia on skeletal muscles (Ubaida-Mohien et al., 2019). Although it has been proposed that exercise, protein supplementation, and dietary restriction might be beneficial for healthy aging, they are not sufficient to treat sarcopenia (Beaudart et al., 2017). Therefore, there is an urgent need to find novel biomarkers of sarcopenia to better understand the etiology of this pathology. In this complex scenario typical of a multifactorial disease such as sarcopenia, our hypothesis seeks to determine the impact omega-3 polyunsaturated fatty acids (w-3 PUFAs) have on the proteome of the soleus skeletal muscle in aging. According to the literature, w-3 PUFAs have already emerged as worthwhile in other health conditions such as heart diseases; brain development; inflammation; and cancer (reviewed by Ruxton et al., 2004). To accomplish this, our present study proposes a bottom-up large-scale shot-gun proteomic study comparing age and diet. Twenty Sprague Dawley rat soleus samples were fed different diets and divided into four different groups (n=5 for each group): young controls (8 months); aged controls (22 months); and young and aged rats fed an w-3 supplemented diet. In our proteomic analysis solei muscles were extracted, and then divided into a pellet (contractile cellular components) and supernatant (soluble proteins) fractions so as to increase the dynamic range of the protein identifications. The protein concentration of each of the 40 aliquots (supernates and pellets) was estimated by the Lowry protein procedure (Lowry et al., 1951). Next, 200 μg of the extracts were reduced with dithiothreitol, irreversibly alkylated with iodoacetamide, and finally digested with trypsin. The resulting peptide mixture was quantified in order to label exactly 10 μg of peptides from each sample with tandem mass tags (TMT). Protein identifications and quantitations were carried out using a nano liquid chromatography-coupled tandem Q Exactive HF-X Orbitrap mass spectrometer and Protein Discoverer 2.4 software. Ultimately, only the statistically significant proteins (p-value ≤ 0.05) showing a differentially expression of +/- 1.5-fold were further investigated and functionally described. From our data it emerges that w-3 PUFAs may be beneficial and protective for the proteome against aging affecting a plethora of different pathways within the cell. This result, combined with existing recommendations such as exercise and dietary restriction, could ameliorate the symptoms of sarcopenia and improve the quality of life.

Rights

©2021 Francesca Maria Del Re. Access limited to the Smith College community and other researchers while on campus. Smith College community members also may access from off-campus using a Smith College log-in. Other off-campus researchers may request a copy through Interlibrary Loan for personal use.

Language

English

Comments

xiv, 194 pages : illustrations (some color). Includes bibliographical references (pages 172-194).

Recommended Citation

Del Re, Francesco Maria, "Proteomic Profiling of Aging and Omega-3 Supplementation in Skeletal Muscle" (2021). Masters Thesis, Smith College, Northampton, MA.
https://scholarworks.smith.edu/theses/2688