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Anabolic steroids induce injury and apoptosis of differentiated skeletal muscle
S Abu-Shakra1, M S Alhalabi, F C Nachtman
1Department of Neurology, School of Medicine, Wayne State University, Detroit, Michigan 48201, USA.
Abstract:
Apoptosis is an active form of cellular death, or suicide, which plays an important physiologic role during organ development and in cellular turnover in differentiated tissues. Apoptosis has also been demonstrated to occur in several organs in response to hypoxic/ischemic, oxidative, or drug-induced injury and is thus involved in disease pathogenesis. However, it is generally assumed that apoptosis does not occur in differentiated skeletal muscle. Apoptosis has been demonstrated in differentiated myocardial muscle, neonatal skeletal muscle, and skeletal myoblasts in response to injury. We therefore studied differentiated murine C2 skeletal muscle cells that have been injured by supraphysiologic doses (>10 microM) of an anabolic steroid, stanozolol. Stanozolol-injured muscle cells exhibited pathologic features suggestive of apoptosis: cytoplasmic shrinkage and chromatin condensation. Muscle cells also showed positive in situ nick-end labeling of nuclear chromatin, indicating DNA strand breakage. Staining with the DNA-binding dye 33342 (bisbenzimide) also showed chromatin changes characteristic of apoptotic nuclei. Total protein levels measured at 4 and 24 hr post-stanozolol injury was not significantly decreased, indicating absence of cell lysis. Cellular ATP levels (nmol ATP/mg protein) of stanozolol-injured muscle cells, measured 4 and 24 hr postinjury, also did not change significantly. In contrast, necrotic muscle cells, injured by the calcium ionophore A23187 (2 microM), showed a progressive decline in total protein and ATP levels. This study supports two other histologic studies that showed evidence of apoptosis in differentiated skeletal muscle fibers. Our data further suggest that during the early stages of apoptosis, but not necrosis, cellular energy metabolism is preserved.
Insights
Apoptosis, or programmed cell death, can occur in differentiated skeletal muscle cells. This study found that stanozolol-induced injury in muscle cells showed signs of apoptosis without significant loss of protein or ATP, preserving cellular energy metabolism.
Area of Science:
- Cell Biology
- Muscle Physiology
- Pathology
Background:
- Apoptosis is a critical physiological process involved in development and tissue homeostasis.
- While apoptosis is known in various tissues, it was traditionally thought to be absent in differentiated skeletal muscle.
- Previous studies suggested apoptosis in cardiac muscle, neonatal skeletal muscle, and myoblasts, but evidence in adult skeletal muscle remained limited.
Purpose of the Study:
- To investigate whether differentiated skeletal muscle cells undergo apoptosis in response to injury.
- To characterize the morphological and biochemical changes associated with stanozolol-induced injury in skeletal muscle cells.
- To differentiate between apoptotic and necrotic cell death pathways in skeletal muscle.
Main Methods:
- Differentiated murine C2 skeletal muscle cells were treated with supraphysiologic doses of stanozolol (>10 microM) to induce injury.
- Morphological analysis included assessing cytoplasmic shrinkage and chromatin condensation.
- In situ nick-end labeling and DNA-binding dye (33342) staining were used to detect DNA strand breaks and chromatin changes.
- Total protein and cellular ATP levels were measured at 4 and 24 hours post-injury.
- Necrotic cell death was induced using the calcium ionophore A23187 (2 microM) for comparison.
Main Results:
- Stanozolol-treated muscle cells displayed characteristic apoptotic features, including cytoplasmic shrinkage and chromatin condensation.
- Positive in situ nick-end labeling confirmed DNA strand breakage in injured nuclei.
- Chromatin changes indicative of apoptosis were observed using DNA-binding dye staining.
- Unlike necrotic cells, stanozolol-injured muscle cells showed no significant decrease in total protein or ATP levels, indicating preserved cellular energy metabolism.
- Necrotic cells induced by A23187 exhibited a progressive decline in protein and ATP levels.
Conclusions:
- Differentiated skeletal muscle cells can undergo apoptosis when exposed to specific injurious agents like stanozolol.
- The findings support histological evidence of apoptosis in differentiated skeletal muscle fibers.
- Early-stage apoptosis in skeletal muscle preserves cellular energy metabolism, distinguishing it from necrosis.