Androgen-sensitive Levator Ani Muscle Research Articles

Identification of potential target genes associated with the reversion of androgen-dependent skeletal muscle atrophy

Muscle wasting or atrophy is extensively associated with human systemic diseases including diabetes, cancer, and kidney failure. Accumulating evidence from transcriptional profiles has noted that a common set of genes, termed atrogenes, is modulated in atrophying muscles. However, the transcriptional changes that trigger the reversion or attenuation of muscle atrophy have not been characterized at the molecular level until now. Here, we applied cDNA microarrays to investigate the transcriptional response of androgen-sensitive Levator ani muscle (LA) during atrophy reversion. Most of the differentially expressed genes behaved as atrogenes and responded to castration-induced atrophy. However, seven genes (APLN, DUSP5, IGF1, PIK3IP1, KLHL38, PI15, and MKL1) did not respond to castration but instead responded exclusively to testosterone replacement. Considering that almost all proteins encoded by these genes are associated with the reversion of atrophy and may function as regulators of cell proliferation/growth, our results provide new perspectives on the existence of anti-atrogenes.

Enobosarm (GTx-024) Modulates Adult Skeletal Muscle Mass Independently of the Androgen Receptor in the Satellite Cell Lineage.

Androgens increase skeletal muscle mass, but their clinical use is hampered by a lack of tissue selectivity and subsequent side effects. Selective androgen receptor modulators elicit muscle-anabolic effects while only sparingly affecting reproductive tissues. The selective androgen receptor modulator, GTx-024 (enobosarm), is being investigated for cancer cachexia, sarcopenia, and muscle wasting diseases. Here we investigate the role of muscle androgen receptor (AR) in the anabolic effect of GTx-024. In mice lacking AR in the satellite cell lineage (satARKO), the weight of the androgen-sensitive levator ani muscle was lower but was decreased further upon orchidectomy. GTx-024 was as effective as DHT in restoring levator ani weights to sham levels. Expression of the muscle-specific, androgen-responsive genes S-adenosylmethionine decarboxylase and myostatin was decreased by orchidectomy and restored by GTx-024 and DHT in control mice, whereas the expression was low and unaffected by androgen status in satARKO. In contrast, insulin-like growth factor 1Ea expression was not different between satARKO and control muscle, decreased upon castration, and was restored by DHT and GTx-024 in both genotypes. These data indicate that GTx-024 does not selectively modulate AR in the satellite cell lineage and that cells outside this lineage remain androgen responsive in satARKO muscle. Indeed, residual AR-positive cells were present in satARKO muscle, coexpressing the fibroblast-lineage marker vimentin. AR positive, muscle-resident fibroblasts could therefore be involved in the indirect effects of androgens on muscle. In conclusion, both DHT and GTx-024 target AR pathways in the satellite cell lineage, but cells outside this lineage also contribute to the anabolic effects of androgens.

17β-Hydroxy-5alpha-androst-1-en-3-one (1-testosterone) is a potent androgen with anabolic properties

Since the begining of the year 2005, the use of steroid precursors (prohormones) is illegal in the United States; nevertheless, there is still an enormous abuse of such substances. One of the most frequently misused steroids, often declared to be a prohormone, is 1-testosterone (17β-hydroxy-5alpha-androst-1-en-3-one, 1-Testo). In this study, we have characterised molecular mechanisms of its action, determined its tissue specific androgenic and anabolic potency and investigated potential adverse effects. 1-Testo binds highly selective to the androgen receptor (AR) and has a high potency to stimulate AR dependent transactivation. In vivo an equimolar dose of 1-Testo has the same potency to stimulate the growth of the prostate, the seminal vesicles and the androgen sensitive levator ani muscle as the reference compound testosterone propionate (TP). Administration of 1-Testo, in contrast to TP, results in a significant increase of liver weight. Our results demonstrate that 1-Testo, even without being metabolised, is a very potent androgen. It binds selectively to the AR and transactivates AR dependent reporter genes. In vivo it has a high androgenic and anabolic potency and increases liver weight. In summary 1-Testo can be characterised as a typical anabolic steroid. It has to be assumed that consumption of this substance is associated with adverse side effects typical for this class of compounds. Therefore, a strict control of its ban is essential.

Mild dystrophic damage in the androgen-sensitive levator ani muscle of the mdx mouse

The time course of muscular dystrophy on the androgen-sensitive levator ani muscle was compared to that of the diaphragm of dystrophic (mdx) mice aged 1–20 months. Muscle growth, isometric contractile properties and caffeine-induced contractures were determined to assess the hormone myotrophic effect, muscle strength and sarcoplasmic reticulum function, respectively, of both control and dystrophic muscles. Histological analysis of mdx muscles showed variable fiber size, centronucleated cells, infiltration of connective tissue, and necrosis which was less severe in the levator ani than in the diaphragm muscle. Tetanic tension per unit weight in the mdx levator ani was reduced (29%) after aging, while the contraction time remained unchanged. The tetanic tension of the mdx diaphragm muscle decreased with age from 3 to 20 months (20–64%), and the relaxation time was prolonged after aging (22%). Gonadectomy of young adult mdx mice caused atrophy of the levator ani muscle, accelerated muscle wasting, reduced the tetanic force (31%), but it did not affect the contraction time and caffeine responses. The results showed that testosterone does not prevent the progress of muscle disease in the mdx levator ani, but androgen withdrawal accelerates muscle wasting suggesting a normonal beneficial effect.

Phosphorylation of p70 s6 kinase is implicated in androgen-induced levator ani muscle anabolism in castrated rats

Androgens are known to increase muscle mass, strength and muscle protein synthesis. However, the molecular mechanisms by which androgens regulate skeletal muscle development remain poorly understood. The ribosomal protein kinase p70 s6k is a regulator of ribosome biogenesis and plays an important role in the regulation of growth-related protein synthesis. The phosphorylation of p70 s6k has been implicated in load-induced skeletal muscle hypertrophy. In the current study, we determined the effect of DHT on the phosphorylation of p70 s6k in the androgen-sensitive levator ani muscle of castrated rats. DHT induced a rapid increase in the phosphorylation of p70 s6k, which was detectable within 6 h after a single injection. Interestingly, DHT-induced phosphorylation of p70 s6k occurred only in androgen-sensitive muscles, but not prostate and seminal vesicle. Co-administration of flutamide, an AR antagonist, inhibited DHT-induced p70 s6k phosphorylation. While serum IGF-I levels were not changed by DHT treatment, IGF-I gene expression levels increased and the mRNA levels of IGFBP3 and IGFBP5 were suppressed in the LA muscle after DHT replacement in castrated rats. These results suggest that the phosphorylation of p70 s6k, likely via the IGF-I pathway, may play an important role in androgen-induced skeletal muscle hypertrophy.

Depressed translational activity in the androgen sensitive levator ani muscle of the rat

The androgen-dependent levator ani (LA) muscle of the rat provides a suitable model to explore the molecular mechanism of steroid hormone action in target tissues. The objective of the present series of experiments was to study the effect of gonadectomy (GDX) and androgen replacement therapy on the in vitro protein synthetic capacity of the LA muscle. The incorporation of labeled methionine into the contractile protein fraction of the LA muscle maintained in organ culture decreases in a time-dependent manner following GDX. Translation of total polyadenylated mRNA in the rabbit reticulocyte translation system revealed that the decrease in protein synthetic capacity was not associated with differences in the template activity of the mRNA derived from GDX tissue. However, when polyribosomes were used to direct the same in vitro synthesis system, a significant time-dependent loss of translational activity was observed following GDX. The polyribosomes of the LA muscle of control and GDX rats were shown to contain equivalent amount of rRNA and mRNA of comparable translation efficiency. Collectively the results of these experiments indicate that the decrease in protein synthetic capacity of the LA muscle in androgen deficient rats is due, in part, to a repression of the translation process associated to the functional integrity of polyribosomes.

Effect of the adrogenic status on the phenotype of the plantaris muscle of the rat

1. 1. The early demonstration of specific androgen binding sites in skeletal muscle suggested that androgen might be involved in the homeostasis of the tissue in adult mammals. 2. 2. Following five weeks of androgen withdrawal, a noticeable increase in the plantaris (PLT) myosin heavy chain cleavage occurs. No other alteration of the contractile proteins pattern was detected. 3. 3. No effect on the template activity of major mRNAs associated with the polysomal fraction was observed in GDX or GDX + TP group compared to control rats. 4. 4. As opposed to the PLT muscle, the template activity of polyribosomes isolated from the androgen sensitive levator ani muscle displays a severe alteration following GDX. 5. 5. Force-frequency analysis of PLT indicated however that the MHC modification was of no functional significance.

Effect of castration and testosterone administration on the neuromuscular junction in the levator ani muscle of the rat

The ultrastructure of the neuromuscular junction (n.m.j.) of the androgen-sensitive levator ani muscle was studied in normal adult male rats, in 8-month-old rats castrated at the age of one month and in castrated rats treated with testosterone propionate (TP). Castration does not result in significant changes of the n.m.j. The density of synaptic vesicles and the postsynaptic junctional folds remain practically normal inspite of marked atrophy of the muscle. TP administration for 7 days results in marked changes in pre- and postsynaptic structures. There is slow progressive depletion of synaptic vesicles, appearance of cisternae and coated vesicles in axon terminals, and coalescence of coated vesicles with the plasma membrane. Coated vesicles are also found inside Schwann cells and among junctional folds. Dense core vesicles appear both in the axon terminals and in the postsynaptic area. Collateral sprouting of terminal axons with the formation of new immature junctions is observed. After 35 days of TP administration depletion of synaptic vesicles continues. Glycogen beta-particles, mostly freely dispersed, occasionally seen in axon terminals 7 days after TP administration, subsequently increase in number. In the endplate zone of the muscle fibre increased protein synthesis is indicated by a rapid increase in ribosomes and irregularly located myofilaments and myofibrils. The appearance of n.m.j. after testosterone administration resembles that described after nerve stimulation; the degree of change is however less pronounced.

Effect of foreign innervation on the androgen-sensitive levator ani muscle of the rat.

Cross-union of the tibial with the pudendal nerve innervating the androgen-sensitive levator ani (LA) muscle of male rats, results in reversal of the histochemical muscle fibre pattern concerning myofibrillar ATPase, succinate dehydrogenase and phosphorylase enzyme activities. The homogeneous muscle fibre pattern of the LA muscle is changed to a mosaic pattern of muscles normally innervated by the tibial nerve. The success of the hetero-reinnervation is shown by practically full recovery of muscle weight and of isometric twitch-contraction properties of the LA muscle. Castration of 2-months duration, i. e. lack of the male sex hormone, leads to marked atrophy but no change in histochemical muscle fibre pattern. Hetero-reinnervation of the LA muscle results in change of histochemical enzyme pattern even if the cross-union of nerves is performed after long periods of castration leading to very marked decrease of muscle fibre size. However, testosterone application alone after castration increases markedly muscle fibre size but does not lead to reversal of muscle fibre pattern. The myotropic hormonal influence on the target (LA) muscle is therefore primarily of myogenic origin and specificity of hormonal action is maintained even with a foreign nerve innervating the muscle. The experiments, thus, provide evidence for the differentiation of specific neural influences affecting muscle fibre pattern and hormonal influences in respect to the myotropic action of the sex hormone on the androgen-sensitive LA muscle.