In vivo high-resolution magic angle spinning magnetic and electron paramagnetic resonance spectroscopic analysis of mitochondria-targeted peptide in Drosophila melanogaster with trauma-induced thoracic injury

Abstract

Trauma is the most common cause of mortality among individuals aged between 1 and 44 years and the third leading cause of mortality overall in the US. In this study, we examined the effects of trauma on the expression of genes in Drosophila melanogaster, a useful model for investigating genetics and physiology. After trauma was induced by a non-lethal needle puncture of the thorax, we observed the differential expression of genes encoding for mitochondrial uncoupling proteins, as well as those encoding for apoptosis-related and insulin signaling-related proteins, thus indicating muscle functional dysregulation. These results prompted us to examine the link between insulin signaling and mitochondrial dysfunction using in vivo nuclear magnetic resonance (NMR) with complementary electron paramagnetic resonance (EPR) spectroscopy. Trauma significantly increased insulin resistance biomarkers, and the NMR spectral profile of the aged flies with trauma-induced thoracic injury resembled that of insulin-resistant chico mutant flies. In addition, the mitochondrial redox status, as measured by EPR, was significantly altered following trauma, indicating mitochondrial uncoupling. A mitochondria-targeted compound, Szeto-Schiller (SS)-31 that promotes adenosine triphosphate (ATP) synthesis normalized the NMR spectral profile, as well as the mitochondrial redox status of the flies with trauma-induced thoracic injury, as assessed by EPR. Based on these findings, we propose a molecular mechanism responsible for trauma-related mortality and also propose that trauma sequelae in aging are linked to insulin signaling and mitochondrial dysfunction. Our findings further suggest that SS-31 attenuates trauma-associated pathological changes.

Mitochondrial Dysfunction Underlies Cardiomyocyte Remodeling in Experimental and Clinical Atrial Fibrillation

“…Therefore, it is highly likely that SS31 is not functional at the moment of heart function measurement. However, injection of SS31 directly in the thorax of adult Drosophila was shown to confer protection against trauma-induced thoracic injury [58].…”

In-Vivo NMR Spectroscopy: A Powerful and Complimentary Tool for Understanding Environmental Toxicity

“…HR-MAS was first introduced in 1996, where researchers used a derivatized Wang resin (a linker for peptide synthesis) to demonstrate its application [ 124 ]. Since then, the approach has rapidly evolved to include in-vivo studies looking at a wide range of whole organisms, including Drosophila melanogaster , Caenorhabditis elegans , Aporrectodea caliginosa , Calanus finmarchicus , and D. magna where a range of well-resolved signals of metabolites were obtained ( Figure 3 ) [ 50 , 128 , 129 , 130 , 131 , 132 , 133 , 134 , 135 ]. One study even applied a slow magic angle spinning approach to a whole live mouse [ 136 ].…”

Lifespan and ROS levels in different Drosophila melanogaster strains after 24 h hypoxia exposure

“…Being a small animal, to quantify ROS in whole body of Drosophila m. , we used an X-Band (∼9.8 GHz) EPR spectrometer (e-Scan Bruker,BioSpin, GmbH, MA, USA). Hypoxic-treated, and untreated flies were anesthetized by cooling on ice (4°C) for 1 min ( Constantinou et al, 2016 ), rapidly homogenized with a pellet pestle (Kontes; 749521-1500) on ice, blended with solution contained: Krebs-Hepes buffer (KHB) containing 25 μM deferoxamine methane-sulfonate salt (DF) chelating agent and 5 μM sodium diethyldithio-carbamate trihydrate (DETC) at pH 7.4 with 1 mM 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine (CMH, Noxygen Science Transfer and Diagnostics, Germany) as a spin-probe which is able to trap superoxide anions, and immediately incubated in a thermostatic water (37°C). After exactly 30 min of incubations, the time necessary to rich plateau, the samples were placed in the center of 1 ml plastic syringe, according to methods previously reported ( Dikalov et al, 2018 ; Rivellini et al, 2021 ), snap frozen and stored at −80°C.…”