Date Available

7-5-2015

Year of Publication

2015

Document Type

Doctoral Dissertation

Degree Name

Doctor of Philosophy (PhD)

College

Medicine

Department/School/Program

Physiology

Advisor

Dr. Francisco H. Andrade

Abstract

Lung tumors secrete multiple factors that contribute to cachexia, a severe wasting syndrome that includes loss of muscle mass, weakness, and fatigue. 80% of advanced lung cancer patients experience cachexia, which cannot be reversed by nutritional interventions, diminishes response to and tolerance of cancer treatments, and increases morbidity and mortality. Despite a multitude of clinical trials, there are currently no approved treatments. This deficiency suggests that not all of the factors that contribute to cachexia have been identified.

Cancer is frequently accompanied by an increase in cyclooxygenase-2 (COX-2), a hallmark of inflammation. Clinical trials for COX-2 inhibitors have resulted in restoration of muscle mass and decreased fatigue. Along with loss of myofibrillar proteins, cachexia also induces mitochondrial dysfunction, which contributes to fatigue. The amelioration of fatigue by COX-2 inhibition suggests possible alterations to mitochondrial function. We hypothesized that there were unidentified tumor-derived factors that contribute to cachectic wasting and fatigue.

Treatment of C2C12 myotubes with Lewis lung cancer-conditioned media (LCM) resulted in increased COX-2 content, myosin loss, and mitochondrial dysfunction. Mass spectrometry revealed 158 confirmed proteins in LCM. We focused on extracellular 14-3-3 proteins because they bind and regulate over 200 known partners. We found that depletion of extracellular 14-3-3 proteins diminished myosin content. CD13, an aminopeptidase, is the proposed receptor for 14-3-3 proteins. Inhibiting aminopeptidases with Bestatin also reduced myosin content.

LCM treatment decreased basal and ATP-related mitochondrial respiration, caused a transient rise in reactive oxygen species (ROS), and increased 4-Hydroxynonenal (4-HNE) in both cytosolic and mitochondrial fractions of cell lysates. COX-2 inhibition did not spare myosin content in LCM-treated myotubes, but did alter mitochondrial respiration and cytosolic oxidant levels.

Our novel findings show that extracellular 14-3-3 proteins may act as previously unidentified myokines, signaling via aminopeptidases to help maintain muscle mass. We elucidated how LCM alters mitochondrial electron flow, and increases oxidative damage by ROS and 4-HNE. Although successful in clinical trials, COX-2 inhibitors do not appear to spare muscle mass by directly working on skeletal muscle, but did alter mitochondrial function.

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