20-Hydroxyecdysone increases fiber size in a muscle-specific fashion in rat.Abstract 20-Hydroxyecdysone (20E) is an ecdysteroid hormone that regulates moulting in insects. Interestingly, 20E is also found most abundantly in plant species and has anabolic anabolic pertaining to or arising from anabolism. anabolic steroid steroids with a tissue-building effect. Testosterone is an example of a natural anabolic steroid with the, sometimes undesirable, effect of causing masculinization. effects in vertebrates, i.e. increasing muscle size without androgen androgen (ăn`drəjən): see testosterone. androgen Any of a group of hormones that mainly influence the development of the male reproductive system. influence. The effect of 20E on slow and fast fiber types of skeletal muscle has not been reported yet. Here we present that 20E affects the size (cross-sectional area, CSA (1) (Canadian Standards Association, Toronto, Ontario, www.csa.ca) A standards-defining organization founded in 1919. It is involved in many industries, including electronics, communications and information technology. ) of the different fiber types in a muscle-specific manner. The effect on fiber size was modified by the distance from the site of the treatment and the presence of a regenerating soleus muscle Noun 1. soleus muscle - a broad flat muscle in the calf of the leg under the gastrocnemius muscle soleus skeletal muscle, striated muscle - a muscle that is connected at either or both ends to a bone and so move parts of the skeleton; a muscle that is in the animal. Besides the fiber size, 20E also increased the myonuclear number in the fibers of normal and regenerating muscles, suggesting the activation of satellite cells. According to our results 20E may provide an alternative for substitution of anabolic-androgenic steroids in therapeutic treatments against muscle atrophy. [c] 2008 Elsevier GmbH. All rights reserved. Keywords: 20-Hydroxyecdysone; Muscle fiber size; Regeneration Introduction Ecdysteroids represent a large family of polyhydroxy-lated steroid invertebrate invertebrate (ĭn'vûr`təbrət, –brāt'), any animal lacking a backbone. The invertebrates include the tunicates and lancelets of phylum Chordata, as well as all animal phyla other than Chordata. hormones that regulate moulting, metamorphosis and reproduction of arthropods (Schnal, 1989). These substances are also synthesized by 5-6% of the plant species (phytoecdysteroids) may be as a defence against phytophagus insects (Dinan, 1995, 2001). Generally ecdysteroids are present in a higher concentration in plants than in arthropods and are easier to extract from plants in order to investigate their pharmacological effect in mammals. The acute toxicity acute toxicity Pharmacology Illness caused by a single exposure to a toxic substance of ecdysteroids is very low in mice: the [LD.sub.50] of 20E is >9g/kg BW when given orally and 6.4g/kg BW if injected intraperitoneally (Ogawa et al., 1974; Matsuda et al., 1970). Ecdysteroids have no side effects Side effects Effects of a proposed project on other parts of the firm. in mammals, including humans (Slama and Lafont, 1995). Moreover, some of the ecdystero id-containing plants are included in the human diet and used in the traditional medicine as well (Spinacia oleracea Spinacia oleracea plant in the family Chenopodiaceae; can cause soluble oxalate poisoning comprising nephrosis, urolithiasis. Called also spinach. (Bathori et al., 1982) or Rhaponticum carthamoides (Syrov and Kurmukov, 1976)). Although ecdysteroids do not bind to the vertebrate steroid receptors and their mechanism of action is still unknown, many beneficial pharmacological properties are attributed to them (reviewed by Slama and Lafont, 1995; Dinan and Lafont, 2006; Bathori et al. 2008). Increased body, organ and muscle weight and protein synthesis have been reported in case of oral or intraperitoneal administration of ecdysteroids in several animal species: Japanese quails (Slama et al., 1996), mice (Stopka et al., 1999), rats (Syrov, 2000) and pigs (Kratky et al., 1997). There is an important difference between the effect of ecdysteroids and anabolic steroids: castration castration, removal of the sex glands of an animal, i.e., testes in the male, or ovaries and often the uterus in the female. Castration of the female animal is commonly referred to as spaying. in rats sharply decreases the protein synthesis stimulating property of ecdysteroids, while steranabols (androgen steroids) still remain effective under this condition (Syrov, 2000). The non-androgenic effect was suggested because the studied phytoecdysteroids did not produce any significant changes in the weight of ventral prostate and seminal vesicles in the tested animals (normal or castrated mature or immature rats, Syrov, 2000). Phytoecdysteroids also showed an anabolic effect in female rats (Syrov, 2000). While in the previous reports the muscle and body weight or the rate of protein synthesis have been measured, a more sophisticated study has not been carried out yet. It is not known whether 20E acts preferentially on fast or slow fiber types like the anabolic steroids (Ustunel et al., 2003). Therefore, we examined the effect of 20E on the size of muscle fiber types and the number of myonuclei reflecting the accretion of myoblasts into the fibers. We chose the soleus so·le·us n. A muscle with origin from the head and shaft of the fibula, the medial margin of the tibia, and the tendinous arch passing between the tibia and fibula, with insertion into the tuberosity of the calcaneus, with nerve supply from the tibial , a predomi nantly slow and the extensor extensor /ex·ten·sor/ (-ser) [L.] 1. causing extension. 2. a muscle that extends a joint. ex·ten·sor n. A muscle that extends or straightens a limb or body part. digitorum longus (EDL See nonlinear video editing. (language) EDL - 1. Experiment Description Language. 2. Event Description Language. ), a predominantly fast type muscle. We also investigated the effect of 20E on the regenerating fibers of the soleus. We found that 20E changes the size and the myonuclear number of the myosin myosin (mī`əsĭn), one of the two major protein constituents responsible for contraction of muscle. In muscle cells myosin is arranged in long filaments called thick filaments that lie parallel to the microfilaments of actin. fiber types in a muscle-specific manner in the normal and regenerating soleus and the normal EDL. This effect is influenced by the applied dose, the distance from the site of 20E injection and, in case of the normal muscles, by the presence of a regenerating soleus in the other hindlimb hindlimb the pelvic limb; back leg. . Materials and methods 20-Hydroxyecdysone 20E was isolated from Silene viridiflora by Toth and Bathori (2008) in the University of Szeged, Faculty of Pharmacy, Department of Pharmacognosy pharmacognosy /phar·ma·cog·no·sy/ (fahr?mah-kog´nah-se) the branch of pharmacology dealing with natural drugs and their constituents. phar·ma·cog·no·sy n. , and provided for the examination in a purity of 99%. The physical and spectroscopic spec·tro·scope n. An instrument for producing and observing spectra. spec  tro·scop data of the isolated com pound are in
accordance with those of the literature (Lafont et al., 2002). The
chemical structure of 20E is shown in Fig. 1.
[FIGURE 1 OMITTED] Animals and treatment Animals were treated according to the regulations of the Ethical Committee of the University of Szeged. Twenty male Wistar rats (304[+ or -]19 g) were divided into five groups, with four animals in each, and treated according to the scheme shown in Fig. 2. The group N20E received 5mg/kg BW of 20E daily as an sc. injection in the left thigh. On the 8th day the soleus and EDL muscles were dissected from the left and right hindlimb and the animals were killed with an overdose of chloralhydrate. Group C, the control of N20E received 0.9% NaCl instead of 20E. Three groups (RC, R20E-1 and R20E-2) received snake venom (notexin) injection into the left soleus muscle. The rats were narcotised and treated for induction of muscle regeneration as in Zador et al. (1998). From the 5th day group R20E-1 received 5mg/kg BW 20E, while group R20E-2 received 0.5mg/kg BW of 20E in a daily subcutaneous injection to the left leg for 7 days. Group RC, the control received 0.9% NaCl instead of 20E. On the 12th day after the snake venom injection which constituted for the 8th day of new muscle formation, the regenerating soleus and the contralateral contralateral /con·tra·lat·er·al/ (-lat´er-al) pertaining to, situated on, or affecting the opposite side. con·tra·lat·er·al adj. normal soleus and the EDL muscles were dissected and the animals were killed with an overdose of chloralhydrate. [FIGURE 2 OMITTED] Collection of muscles and staining for HE or immunocytochemistry im·mu·no·cy·to·chem·is·try n. The study of cell constituents by immunologic methods, such as the use of fluorescent antibodies. immunocytochemistry The dissected regenerated soleus, normal soleus and EDL muscles were frozen in isopentane cooled with liquid nitrogen and kept at--70 [degrees]C. In all, 15 [micro]m thick cryosections were taken from the central part of each frozen muscles and stained with haematoxylin-eosin, Immunostaining was carried out as in Zador et al. (1998). We identified IIx fibers as non-stained ones with a combination of BA-D5 (mouse, 1:50), SC-71 (mouse, 1:20) and BF-F3 (mouse, 1:10) antibodies. Fiber cross-sectional area and myonuclear domain The cross-sectional area (CSA) of 150 fibers of each muscle was measured by Olympus DP-soft, version 3.2 program (Olympus, Hamburg, Germany) on haematox-ylin-eosin stained or on immunostained sections. The number of myonuclei in the fibers (in more than 100 fibers of each muscle) was counted on haematoxyli n-eosin stained sections with the 40 x objective of the light microscope. The accuracy of this method was controlled by comparing the results of two independent experienced persons. The myonuclear domains in each muscle were calculated from the average myonuclear number per fiber divided by the average fiber CSA. In the EDL the large fibers were counted for myonuclei and divided by the average CSA of the IIx and IIB IIB Institute for Independent Business IIB Institute of International Business IIB Institute of International Bankers IIB International Investment Bank IIB Indian Institute of Banking & Finance IIB Included in Bankruptcy IIB Ice, Ice, Baby fibers, as these fiber types were responsible for the size increase in response to the 20E treatment. We also counted the number of myonuclei of the small fibers in EDL and divided by the average CSA of the type I and IIA fibers. Statistics The cumulative data of muscle fiber CSA's obtained from four muscles (150 from each) were compared among groups by using t-test for unpaired samples or one-way analysis of variance followed by Newman--Keuls post-hoc test. The number of myonuclei and the size of myonuclear domain were also compared as cumulative data of four muscles (from 100 fibers of each). All tests were performed by using the GraphPad Prism version 3.00. Results were considered significant at p < 0.05. All data are expressed as means [+ or -] SE. Results Body and muscle mass The body and muscle mass increased significantly in group N20E compared to group C. However these two parameters did not change in group R20E-1 and only the muscle mass of the right EDL increased in group R20E-2 compared to group RC (Table 1).
Table 1. Body weight and muscle mass of the 20E-treated rats
Bofy weight (g) m. soleus (mg)
Start of End of Change in Right Left
treatment experiment weight
C
324.5 336.5 12.0 140.3 143.5
[+ or -] [+ or -] [+ or -] [+ or -] [+ or -]
2.50 6.55 4.16 5.69 4.09
(318-330) (320-352) (2-22) (128-155) (135-151)
N20E
309.0 373.0 64.0 160.0 175
[+ or -] [+ or -] [+ or -] [+ or -] [+ or -]
3.82 5.26 4.32 *** 4.77 *** 7.14 *
(304-316) (364-388) (52-72) (150-170) (160-194)
RC
285.3 353.8 68.5 158.0 --
[+ or -] [+ or -] [+ or -] [+ or -]
3.82 7.96 10.65 3
(272-292) (340-376) (49-94) (150-160)
R20E-1
293.5 352.5 59.0 168.0 --
[+ or -] [+ or -] [+ or -] [+ or -]
2.06 4.78 3.11 7.5
(290-298) (340-360) (50-64) (150-180)
R20E-2
314.5 387.0 73.0 174.0 --
[+ or -] [+ or -] [+ or -] [+ or -]
8.29 5.80 4.04 6.96
(296-336) (376-402) (66-80) (158-189)
m. EDL (mg) Regeneration m.
soleus (mg)
Right Left
142.8 [+ or -] 3.64 146.0 [+ or -] 3.03 -
(136-152) (137-150)
166.0 [+ or -] 6.38 * 172.0 [+ or -] 9.58 * -
(156-184) (154-197)
155.0 [+ or -] 5 - 113.0 [+ or -] 6
(150-170) (100-130)
150.0 [+ or -] 4.08 - 113.0 [+ or -] 7.5
(140-160) (100-130)
178.0 [+ or -] 4.05 * - 116.0 [+ or -] 10.49
(169-188) (90-136)
*, **, *** p < 0.05, 0.01, 0.001 compared to the correspondent control
(0.9% NaCl and notexin + 0.9% NaCl). The mass range is in parenthesis.
CSA of MyHC fiber types The size of muscle fibers increased significantly in response to the 20E treatment in the left soleus of group N20E (5011 [+ or -] 67.44 [micro][m.sup.2]) compared to group C (4261[+ or -]49.68 [micro][m.sup.2]). The rat soleus muscle consists of slow-oxidative type I (90%) and fast-oxidative type IIA (9%) fibers expressing MyHCl and MyHC2a isoforms, respectively. The rate of IIx/d (mentioned as IIx) fibers is only 1%. The CSA of both type I and type IIA fibers in the soleus muscle of the treated left hindlimb increased (p < 0.001) in group N20E compared to control group C (Fig. 3.A, black and empty columns). [FIGURE 3 OMITTED] 20E also increased the size of muscle fibers in the EDL (3137 [+ or -]56.98 vs. 2488 [+ or -] 48.07 [micro][m.sup.2]) and the fiber size distribution of the treated muscle showed a bimodular pattern, suggesting a distinct effect on the different fiber types. In the EDL four fiber types (I, IIA, IIx and IIB) are present expressing correspondingly the MyHC1, MyHC2a MyHC2x and MyHC2b isoforms. However in the left EDL the CSA of the type IIx fibers increased (p < 0.001), while the type I and type IIB fibers did not change, but the CSA of type IIA fibers decreased (p < 0.001) compared to those of group C (Fig. 3B, black and empty columns). This shows that 20E affects fiber size in a muscle-specific fashion instead of a fiber type-dependent manner. The effect of 20E on the contralateral hindlimb The fiber sizes in the treated (left) and non-treated (right) hindlimbs of group N20E were compared with those of the corresponding hindlimbs in the control. Interestingly, the size of the same fiber types was different in the left than in the right hindlimb muscles even in the control group (Fig. 3). In group N20E, the type I fibers were larger (p < 0.05) in the right than in the left soleus, while the IIA fibers became larger in the left than in the right soleus (p < 0.01) compared to those in group C (Fig. 3A). In the EDL, the CSA of type I fibers were not different from the control, but the IIA fibers became smaller after the 20E treatment in the left than in the right muscle. Moreover the size of IIx fibers increased in the left (p0.001) but not in the right EDL, while the size of the IIB fibers was higher in the right EDL of the control and it was decreased by 20E in the right but not in the left EDL compared to the controls (Fig. 3B). This shows that the distance from the site of the 20E treatment blunted the decrease of IIA fibers and the increase of IIx fibers in the EDL. 20E stimulates the fiber growth in the regenerating soleus The soleus muscle consisted of entirely new myofibers after 5 days of notexin treatment, later 98% of them expressed slow myosin (Whalen et al. 1990). The size of the regenerating fibers has been increased (p < 0.001) by 20E in group R20E-1 (Fig. 4A). [FIGURE 4 OMITTED] Injection of a 10 times lower dose of 20E (0.5 mg/kg BW) for 7 days into rats (group R20E-2) increased the fiber size (p < 0.001) only in the regenerating soleus, but this increase was less pronounced than in the animals treated with 5 mg/kg BW of 20E (group R20E-1) (Fig. 4B). This showed that the effect of 20E was dose dependent and the 0.5 mg/kg BW was still an effective dose on the regenerating soleus but not on the fibers of normal muscles in the contralateral leg (data not shown). The regenerating soleus modified the effect of 20E on fiber size of normal muscles The relative changes in CSA of fiber types in soleus and EDL of the right hindlimb of group R20E-1 and N20E were different compared to the corresponding controls, RC and C (Fig. 5). The increase of type I fiber sizes in the right soleus was smaller (p < 0.001) in the R20E-1 than in the N20E group. A smaller but still significant difference was also found between the relative changes of the type IIA fibers of the right soleus muscles of the two groups (p < 0.05). In the right EDL the type I fibers were not changed compared to controls. The IIA fibers were more decreased in R20E-1 than in the N20E group (p < 0.001). Interestingly, the size of IIx fibers was increased only in the R20E- 1 group, while the size of IIB fibers increased in the R20E-1 group and decreased in the N20E group. This showed that the presence of a regenerating soleus in the rat modifies the effect of 20E on the size of fiber types. [FIGURE 5 OMITTED] Myonuclear number The numbers of myonuclei followed the increase of cross-sectional areas in most muscles after the 20E treatment. This meant that the size of myonuclear domains did not change, except in group N20E when an increase (p < 0.01) in the right soleus and decreases in large fibers (IIx and IIB) in EDL on both sides were found (Table 2). The myonuclear domains also decreased in small fibers of the EDL in R20E-1 group. The positions of myonuclei did not change in response to the 20E treatment, i.e. they remained peripheral in normal and central in regenerating muscles.
Table 2. Myonuclear numbers and domains in 20E-treated muscles
Myonuclei/fiber
m. soleus
Right Left
0.88[+ or -]0.06 0.91[+ or -]0.03
0.87[+ or -]0.01 1.09[+ or -]0.03 *
0.92[+ or -]0.05 -
0.96[+ or -]0.04 -
- -
Myonuclear domain ([mu][m.sup.2])
4405[+ or -]81.73 4481[+ or -]179.9
5532[+ or -]306.0 * 4493[+ or -]145.1
4011[+ or -]167.1 -
4378[+ or -]89.0 -
- -
m. EDL Regenerating
m. soleus
Lage fibers Small fibers
Right Left Right Left
C
1.590 1.536 0.45 0.52 -
[+ or -] [+ or -] [+ or -] [+ or -]
0.05 0.03 0.017 0.013
N20E
1.96 2.04 0.44 0.45 -
[+ or -] [+ or -] [+ or -] [+ or -]
0.07 * 0.08 ** 0.02 0.02
RC
1.67 - 0.42 - 1.57
[+ or -] [+ or -] [+ or -]
0.05 0.018 0.06
R20E-1
2.00 - 0.52 - 1.91
[+ or -] [+ or -] [+ or -]
0.13 0.01 ** 0.06 **
R20E-2
- - - - 1.82
[+ or -]
0.03 **
C
1808 1759 3072 2713 -
[+ or -] [+ or -] [+ or -] [+ or -]
96.41 55.42 197.6 168.4
N20E
1447 1539 2860 2613 -
[+ or -] [+ or -] [+ or -] [+ or -]
81.16 * 42.02 * 81.22 6.4
RC
1774 - 3449 - 1234
[+ or -] [+ or -] [+ or -]
221.4 260.30 55.63
R20E-1
1596 - 2590 - 1082
[+ or -] [+ or -] [+ or -]
105.7 134.20 * 32.75
R20E-2
- - - - 1028
[+ or -]
104.2
*, **, ***p < 0.05, 0.01, 0.001 compared to the correspondent control
(0.9% NaCl and notexin + 0.9% NaCl).
Fiber type proportion The relative fiber type proportion did not change in the muscles of any of the groups. The 20E also did not influence the formation of predominantly slow type fibers in the regenerated soleus. Discussion Our study demonstrated that 20E affects the size of fiber types in a different manner in the soleus than in the EDL muscles. This suggests that this compound affects fiber type size in a muscle-specific fashion. A muscle-specific effect on fiber size has also been reported in case of anabolic-androgenic steroids (AAS). Treatment with a low (1.5 m/g/kg BW/week) and a high (7.5 mg/kg BW/week) therapeutic dose of nandrolone decanoate nandrolone decanoate Endocrinology A long-acting parenteral 17β-hydroxyl ester of 19-nortestosterone with weak androgenic activity, used as an androgen–eg, for refractory anemia. Cf Androgen replacement therapy. over a 5 weeks period increased the dimension of type IIx and IIB fibers in the diaphragm and of type IIa fibers in the gastrocnemius gastrocnemius /gas·troc·ne·mi·us/ (gas?tro-ne´me-?s) (gas?trok-ne´me-us) see under muscle. gas·troc·ne·mi·us n. pl. , while the other fiber types remained unchanged (Bisschop et al., 1997). In human, where the MyHC2b is hardly expressed in type II fibers, a long-term AAS administration (power lifters taking anabolic supplements for nearly 10 years) increased the CSA of type I and type II fibers both in m. vastus lateralis vas·tus lat·e·ra·lis n. A muscle with origin from the posterior ridge of the femur as far as the greater trochanter, with insertion into the tibia, with nerve supply from the femoral nerve, and whose action extends the leg. and m. trapezius tra·pe·zi·us n. A muscle with origin from the superior nuchal line, the external occipital protuberance, the nuchal ligament, the spinous processes of the seventh cervical and thoracic vertebrae, with insertion into the lateral third of the posterior , while no alteration in the fiber type proportions was found (Smerdu et al., 1994; Eriksson et al., 2005; Kadi Ka´di n. 1. A Turkish judge. See Cadi. et al., 1999). Similar observations were made in healthy young men when the diameter of both type I and II fibers were increased after 20 weeks supplementation of gonadotrop-releasing hormone agonist (to suppress endogenous testosterone release) and of exogene testosterone (Sinha-Hikim et al., 2002). Although the effect of 20E on these muscles is to be determined, it appears that both anabolic steroids and ecdysteroids are acting on the size of fibers in a multiple, muscle-specific fashion. However, it should be noted that AAS are effective in a lower dose than 20E. It is another difference that 20E did not alter the fiber type proportion in the studied muscles as the anabolic steroids (Holmang et al., 1990). The fast anabolic effect of steroids in skeletal muscle is exerted through signal transduction pathways and not via intracellular steroid receptors (Estrada et al. 2003). The overexpression of calcineurin, a central player in signalling in muscle, influences fiber phenotype and differentiation in a muscle-specific fashion (Talmadge et al. 2004). This is in line with the muscle-specific effects of AAS and ecdysteroids on fiber types and it implies that ecdysteroids might act on skeletal muscle via signal pathways. In accordance with this, ecdysteroids have been shown to act on signal mechanisms of mammalian hematopoietic cells (Constantino et al. 2001). The 20E showed a more pronounced effect in the treated left hindlimb (on IIx and IIb fibers of the EDL) than in the untreated right hindlimb, probably because of the distance from the site of administration. This shows that the 20E acts locally and via the circulation. 20E increased fiber CSA in the regenerating soleus muscle, suggesting that the 20E exerted a beneficial effect on muscle regeneration. A similar effect of AAS (nandrolone nandrolone /nan·dro·lone/ (nan´dro-lon) an anabolic steroid with lesser androgenic effects; used as n. decanoate and n. ) has been reported on the regenerating soleus but not on the EDL (Ferry et al., 1999). However, in our experiments, the regenerating soleus muscle in the left hindlimb influenced the effect of 20E in the contralateral hindlimb compared to the animals without regenerating soleus. This cannot be explained by the overload of the right hindlimb (caused by the retained use of the left hindlimb), since overload increases the size and the number of type I fibers (Zador et al., 1999), unlike it happened in the right hindlimb. It is more likely that the regenerating soleus influences the growth factor environment which interacts with the effect of 20E on the right EDL. In support of this, one example is known when ecdysteroids altered the signalling by interleukin-3 in mammalian hematopoetic cells (Constantino et al., 2001). A 10 times lower dose (0.5 mg/kg BW) of 20E induced a lower increase of the fiber's CSA in the regenerating soleus but had no effect on the fibers in the contralateral normal muscles. However the low dose of 20E increased the weight of the EDL, suggesting dose dependence and a difference in the mechanism of action of a low and high dose of 20E. The 20E increased the myonuclear number in most of the affected muscles. Muscle fibers also grow in response to AAS with increasing myonuclear number (Eriksson et al., 2005). The myonuclei derive from the accretion of myoblasts produced by the activated satellite cells (muscle resident mesenchymal stem cells), and help to maintain gene expression within the fiber. The 20E increased the myonuclear number in proportion to the fiber growth, therefore maintained the size of the myonuclear domains (the sarcoplasm volume around the myonuclei). This suggests that the 20E stimulation of fibers size involves the activation of satellite cells similar to AAS (reviewed by Chen et al., 2005). In conclusion, 20E modifies muscle fiber size in normal and regenerating muscles even after 7 days administration in a slightly higher dose than the anabolic steroids. This effect is dose dependent and similarly to that of anabolic steroids, it influences the size of fiber types in a muscle-specific fashion. The 20E probably acts in concert with other growth factors because its effect on normal muscles is modified by the presence of a regenerating soleus. This suggests that 20E may provide an opportunity for substitution of anabolic-androgenic steroids in therapeutic treatments against muscle atrophy. Acknowledgements This work was partially supported by the ETT ETT Empresa de Trabajo Temporal (Spain) ETT European Transactions on Telecommunications ETT Exercise Treadmill Test ETT Embedded Training Team ETT Exercise Tolerance Test (cardiology) 168/2003 grant from the Hungarian Ministry of Health. Thanks to professor Maria Bathori for the 20-hydroxyecdysone. Thanks to Dr. Arpad Marki for technical assistance in the initial phase of this study. References Bathori, M., Toth, I., Szendrei, K., Reisch, J., 1982. Ecdysteroids in Spinacia oleracea and Chenopodium bonus-Henricus. 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Motil. 20, 395-402. Noemi Toth (a), Andras Szabo (c), Peter Kacsala (b), Julia Heger (c), Erno Zador (c), * (a) Department of Pharmacognosy, Szeged, Hungary (b) Faculty of Pharmacy, Department of Pharmacodynamics pharmacodynamics /phar·ma·co·dy·nam·ics/ (-di-nam´iks) the study of the biochemical and physiological effects of drugs and the mechanisms of their actions, including the correlation of their actions and effects with their chemical and Biopharmacy, Szeged, Hungary (c) Faculty of General Medicine, Institute of Biochemistry, University of Szeged, H-6720 Szeged, Hungary Abreviations: 20E, 20-hydroxyecdysone; AAS, anabolic-androgenic steroid; BW, body weight; CSA, cross-sectional area; M. EDL, muscle extensor digitorum longus; MyHC, myosin heavy chain. * Corresponding author. Tel.: +3662 545 096; fax: +3662 545 097. E-mail address: erno@biochem.szote.u-szeged.hu (E. Zador). |
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