Browsing by Author "Talmadge, Robert J."
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- Calcineurin activation influences muscle phenotype in a muscle-specific fashionTalmadge, Robert J.; Otis, Jeffrey S.; Rittler, Matthew R.; Garcia, Nicole D.; Spencer, Shelly R.; Lees, Simon J.; Naya, Francisco J. (2004-07-28)Background The calcium activated protein phosphatase 2B, also known as calcineurin, has been implicated as a cell signaling molecule involved with transduction of physiological signals (free cytosolic Ca2+) into molecular signals that influence the expression of phenotype-specific genes in skeletal muscle. In the present study we address the role of calcineurin in mediating adaptations in myosin heavy chain (MHC) isoform expression and muscle mass using 3-month old wild-type (WT) and transgenic mice displaying high-level expression of a constitutively active form of calcineurin (MCK-CN* mice). Results Slow muscles, e.g., soleus, were significantly larger (by ~24%), whereas fast muscles, e.g., medial gastrocnemius (MG) and tibialis anterior were significantly smaller (by ~26 and ~16%, respectively) in MCK-CN* mice compared to WT. The masses of mixed phenotype muscles, such as the plantaris and the extensor digitorum longus, were not significantly changed from WT. The soleus, plantaris, MG and diaphragm displayed shifts toward slower MHC isoforms, e.g., soleus from WT mice contained ~52% MHC-I, ~39% MHC-IIa, and ~9% MHC-IIx, whereas MCK-CN* mice had ~67% MHC-I, ~26% MHC-IIa, and ~7% MHC-IIx. The specific isoforms that were either up or down-regulated were muscle-specific. For instance, the proportion of MHC-IIa was decreased in the soleus and diaphragm, but increased in the plantaris and MG of MCK-CN* mice. Also, the proportion of MHC-IIx was unchanged in the soleus, decreased in the diaphragm and increased in the plantaris and MG of MCK-CN* relative to WT mice. Fast to slow shifts in fiber type proportions were evident for the plantaris, but not the soleus. Fast, but not slow, plantaris fibers of MCK-CN* mice had higher oxidative and lower glycolytic properties than WT. Conclusion These data suggest that calcineurin activation can influence muscle phenotype and that the specific influence of calcineurin activation on the phenotypic and mass characteristics of a muscle is dependent upon the original phenotypic state of the muscle.
- The effects of congestive heart failure and functional overload on rat skeletal muscleSpangenburg, Espen E. (Virginia Tech, 2000-06-23)Numerous references have suggested that alterations in exercise capacity during congestive heart failure (CHF) are not simply due to changes in myocardial function. In fact, recent evidence has indicated that reductions in skeletal muscle strength and endurance during CHF significantly impact exercise capacity of the CHF patient. Currently, it is believed that alterations in skeletal muscle phenotype, or more specifically a slow to fast transformation in phenotypic protein isoforms contribute to the reductions in muscle function. However, currently there are few data which directly document this slow to fast transformation of the skeletal muscle. Interestingly, it is well established that exercise training can cause changes in skeletal muscle phenotype, more specifically in the fast to slow direction. This is in direct contrast to what is known to occur during CHF. However, it is unclear if similar adaptations will result from training in a CHF patient. Also, it is not clear if the adaptations are due to alterations in the myocardium or changes directly imposed upon the muscle by the exercise training. Therefore, the purpose of this study was two-fold: 1) to clarify the changes in skeletal muscle myosin heavy chain (MHC) during CHF and 2) to determine if skeletal muscle can adapt to increased activity levels, utilizing functional overload (FO) without significantly improving cardiac function. In the first study the mixed plantaris muscles from rats afflicted with severe CHF demonstrated a significant (p<0.05) increase in fast MHC (e.g. IIb expression at the expense of IIx expression) compared to the control animal (SHAM). The mixed red gastrocnemius, regardless of the severity of CHF, exhibited significant (p<0.05) changes in all of the MHC isoforms. The slow soleus and fast white gastrocnemius did not display any significant changes in MHC expression. The changes in MHC isoform significantly correlated with indicators of disease severity, suggesting there may be an existing relationship between skeletal muscle MHC expression and alterations in myocardial function. In the second study, there were no differences exhibited between CHF and SHAM absolute or specific plantaris mass. There was a significant (p<0.05) 30% increase in both absolute and specific mass of the plantaris in the CHF-FO and SHAM-FO groups compared to the CHF and SHAM groups. There was a significant (p<0.05) 3.5% increase in slow MHC I expression and a significant (p<0.05) 6.5% decrease in fast MHC IIb expression in the CHF-FO group compared to the CHF group. In the SHAM-FO group, there was a significant (p<0.05) 4% increase in MHC I expression and a subsequent 8% decrease in fast MHC IIx+IIb in the SHAM-FO compared to the SHAM groups. There were no differences detected in the rates of Ca²⁺ uptake between the CHF-FO, SHAM, and SHAM-FO. However, Ca²⁺ uptake rates were significantly (p<0.05) elevated by 44% in the CHF group when compared to the other three groups. There were very few changes in plantaris SERCA 1 or 2 protein expression between the four groups. These data suggest that during CHF there are alterations in skeletal muscle isoform expression. However, at least some of the data suggest that changes in function are not always associated with changes in phenotype. Instead, it seems that the changes in Ca²⁺ handling may be due to an alteration in a regulatory mechanism. Also, the data indicate that skeletal muscle is adaptable to increases in activity levels without significantly altering myocardial morphology.
- Effects of Reduced Muscle Glycogen on Sarcoplasmic Reticulum (SR), Muscle and Exercise PerformanceBatts, Timothy W. (Virginia Tech, 2002-04-19)Fatigue during exercise is associated with reduced muscle glycogen. However, evidence linking glycogen content to fatigue is lacking. In this study we examined whether reduced muscle glycogen content limited SR function or muscle performance. Two groups of female Sprague-Dawley rats were fasted for 24 hr and exercised for 90 min to reduce muscle glycogen; rats fasted after exercise formed the low glycogen (LG) group. Rats in the high glycogen (HG) group were allowed free access to food and a 5% sucrose solution. The LG group had 42% less muscle glycogen and 90% less glycogen associated with the sarcoplasmic reticulum (SR) than the HG group. Notably, time to exhaustion during a subsequent treadmill run (21 m/min at 10% grade) was markedly lower in the LG group (35 vs. 166.75 min). Despite less glycogen, the LG group had a higher SR Ca2+ uptake rate (45%) and Ca2+-stimulated ATPase activity (51%) possibly due to a 33% greater SERCA content. Surprisingly, in situ gastrocnemius initial twitch and tetanic forces were not different between groups although the rates of relaxation were higher in the LG group. The force responses to fatigue-inducing stimulus trains (20 Hz for 333 ms every 1 sec for 30 min) also were similar for both groups as were twitch and tetanic forces in the fatigued state. These results suggest that despite reduction in exercise performance, reduced muscle glycogen does not limit muscle performance or SR function.
- The Incidence of Stress Fractures Among All Female Division I Athletes at Virginia Polytechnic Institute and State UniversityLogsdon, Susannah M. (Virginia Tech, 1999-04-20)Stress fractures are common overuse injuries that have plagued athletes for many years. Often referred to as fatigue fractures, they are formed when the skeletal muscles fatigue and can no longer absorb the shock of repetitive pounding activities such as running. Stress fractures are most common in the weight-bearing bones of the lower extremities and are usually preceded by sudden increases or changes in training routines. Because they are most common in athletes who are least fit, it has been hypothesized that freshmen athletes who are not prepared for the increased physical demands of college athletics have the greatest risk for developing stress fractures compared to other academic classes. As of yet however, there have been very few studies that have examined the interaction of different variables such as academic class, on the formation of stress fractures. Therefore, the purpose of this study was to look at the incidence, frequency and pattern of stress fractures among the female athletes at Virginia Polytechnic Institute and State University. A retrospective analysis of 28 injury cases over four years revealed that 67% of the injuries occurred in freshmen athletes. The majority of these were in the lower leg and occurred in mid-season rather than pre-season. There were no relationships found between the variables sport, class, site and season and thus it was concluded that the incidence of injury was not affected by the interaction of these variables. However, the variables themselves did influence the formation of stress fractures. Ultimately, this study provided insight on what factors should be carefully examined in order to prevent future stress fracture injuries in collegiate athletes.
- Metabolic profile of myosin heavy chain-based fiber types in the rat soleus after spinal cord transectionOtis, Jeffrey Scott (Virginia Tech, 2000-10-27)Fully differentiated muscle fibers can undergo considerable phenotypic changes in order to adjust to changing conditions of the physiological environment. It is generally accepted that the electrical impulses a muscle receives play a role in modulating the quantities of metabolic proteins (glycolytic and oxidative enzymes) and types of contractile proteins (myosin heavy chain, MHC) that are expressed. Research has shown that decreased neuromuscular activation following spinal cord transection (ST) results in adaptations in the physiological characteristics of paralyzed muscles, including atrophy and an accompanying loss of force production, and transformations of contractile and metabolic proteins toward a more fatigable state. However, it remains unclear whether or not a strong interdependence of energy metabolism and MHC isoform composition persists. Therefore, the goal of this study was to identify and quantify relative myosin heavy chain (MHC) isoform expression and metabolic enzyme profile adaptations at multiple time points (1, 3 and 6 months) in soleus fibers of rats following spinal cord transection (ST). To accomplish this, female Sprague-Dawley rats (~150 g, n = 15) were subjected to complete transection of the spinal cord at a mid-thoracic level. Age and weight-matched, non-operated rats served as controls (n = 15). The soleus was processed for quantitative single fiber histochemical analyses for succinate dehydrogenase (SDH, oxidative marker) and a-glycerophosphate dehydrogenase (GPD, glycolytic marker) activities (~30 fibers/muscle) and immunohistochemical analysis for MHC isoform composition. The total number of soleus fibers analyzed was ~900. Oxidative capacity was increased in muscle fibers at all time points after ST. Specifically, SDH activity was significantly higher than controls by 142, 127 and 206% at 1, 3 and 6 months post-ST, respectively. ISDH, a measure of total oxidative power, also increased in muscle fibers at all time points after ST. For example, 6 months after ST ISDH activity was 93% higher than controls (91.8-3.8 vs. 47.6-0.9 OD x 10-3, respectively). Glycolytic capacity peaked one month after ST. Thereafter, glycolytic capacity of all fibers steadily declined. For example, by 6 months, GPD activity had declined by 76% compared to 1 month GPD activities (3.3-0.2 vs. 13.7-1.4 OD x 10-3, respectively). These data suggest that the increases in glycolytic capacity are transient as fibers transition toward a faster MHC phenotype and then return towards control levels as fibers of a given type become phenotypically stable. The GPD/SDH ratio, an index of metabolic substrate utilization, peaked at one month after ST (394-41) and significantly decreased at 3 months (224-10) and at 6 months (95-7) after ST. Therefore, a shift occurred such that a greater dependence on oxidative metabolism was apparent. These data suggest that the oxidative capacities of soleus muscle fibers are not compromised after ST. In fact, as the fibers transitioned toward faster MHC isoforms, the GPD/SDH ratio was maintained or decreased, suggesting a reliance on oxidative metabolism regardless of MHC isoform composition. This might imply a dissociation between the contractile and metabolic characteristics of paralyzed soleus muscle fibers. However, these data are consistent with previous data and suggest that the increased fatigability observed after chronic reductions in neuromuscular activity are not due to compromised capacities for ATP synthesis.
- Vitamin B6 Decreases Proliferation and DNA Synthesis in Human Mammary Carcinoma Cell Lines In VitroCowing, Brandy Ellen (Virginia Tech, 2000-04-04)The growth of many breast cancers is stimulated by the action of the hormone estrogen. Hormonal therapy used to treat these estrogen-dependent breast cancers acts by interfering with the action of estrogen. Current treatments, such as tamoxifen, are not consistently useful due to development of resistance to these drugs. Tamoxifen treatment can also lead to the development of other gynecological cancers, therefore the discovery of novel treatment options for breast cancer is critical. Vitamin B6 is well documented for its role as a modulator of steroid hormones. Pyridoxal phosphate (PLP), the active form of Vitamin B6, may interfere with the action of the estrogen receptor (ER) by blocking the hormone-binding and/or DNA-binding site of the ER. The objective of this study was to examine the effects of Vitamin B6 supplementation on cell proliferation and estrogen-dependent gene expression in breast cancer cells. To accomplish this, estrogen-dependent (MCF-7 and T-47D) and estrogen-independent (BT-20) breast cancer cells were grown in medium supplemented with 0,100, or 300 µM pyridoxal (PL) in the absence or presence of 0.01µM estradiol. Cell counts and [3H]-thymidine incorporation into DNA were assessed in all cell lines. The expression of pS2, an estrogen-sensitive gene, was performed using RNA extracted from MCF-7 cells. PL supplementation was found to significantly decrease total cell numbers and DNA synthesis in both the estrogen-dependent (ER+) and -independent (ER-) breast cancer cells, but did not alter the expression of pS2. These results indicate that PL significantly impairs growth of breast cancer cells and may be exerting its effects via a steroid-independent mechanism.