Effects Of 20-hydroxyecdysone Research Articles

The Effect of 20-Hydroxyecdysone on the Functioning of Isolated Mouse Skeletal Muscle Mitochondria

The Effect of 20-Hydroxyecdysone on the Functioning of Isolated Mouse Skeletal Muscle Mitochondria

The effects of 20-hydroxyecdysone on nuclear shape, heterochromatin quantity and gray-level co-occurrence matrix texture analysis of adrenal zona fasciculata cells in an obese gerbil (Gerbillus tarabuli) model for metabolic syndrome: a correlational study.

The aim of this study was to reveal the effects of obesity and phytotherapy with 20-hydroxyecdysone (20E) on the nuclei of adrenal zona fasciculata (ZF) in the gerbil Gerbillus tarabuli by analyzing nuclear shape and gray-level co-occurrence matrix (GLCM) texture characteristics and by quantifying heterochromatin. Twelve gerbils were divided into three groups: control (C), HC and HC-20E (animals receiving a high-calorie-diet without or with a supplement of 20E, respectively). The adrenals were removed and fixed for histological and statistical analysis. Principal component analysis showed a positive correlation of area, perimeter and textural correlation in C. Nevertheless, a negative correlation was recorded for contrast and entropy. The obesity caused a disorder in nuclear texture; negative correlation was noted with heterochromatin fraction, which may be related to increased ZF activity. However, administration of 20E seems to improve the nuclear state by preserving circularity, uniformity and homogeneity of nuclei as well as the proportion of heterochromatin, which could be a sign of a downregulation of cell activity.Our results suggest that new techniques of image processing could contribute to the understanding of nuclear changes associated with obesity and its possible therapy in this gerbil model for metabolic syndrome.

Effects of 20-hydroxyecdysone on UVB-induced photoaging in hairless mice

We recently reported that exposure of skin to ultraviolet B (UVB) irradiation for 2 weeks induces stress and accelerates skin aging. Interestingly, aldosterone synthase is known to be crucial in generating UVB-induced stress-related responses, suggesting that drugs that regulate its activity can be used as skin antiaging agents. Through extensive drug screening, we have identified 20-hydroxyecdysone (20E), a steroidal prohormone secreted by the prothoracic glands of insects, as a potent inhibitor of UVB-induced aging. Although 20E has been shown to exert antistress and anti-collagenase effects in vitro, its effects in vivo remain unexplored. Furthermore, the pharmacological and physiological effects of 20E on UVB-mediated photoaging are poorly understood. Therefore, in this study, we investigated the effects of 20E on aldosterone synthase and UVB-induced photoaging and skin lesions in hairless mice, focusing on the stress-related hypothalamic–pituitary–adrenal axis. We confirmed that 20E inhibited aldosterone synthase and reduced corticosterone levels. When applied to a UV-induced skin aging animal model, it ameliorated UV-induced stress and protected against the decrease in collagen levels. Importantly, when the aldosterone synthase inhibitor osilodrostat, an FDA-approved drug, was applied to the UV-induced skin aging model, the stress-reducing and antiaging effects of 20E were not observed. Thus, we conclude that 20E inhibits UVB-induced skin aging by blocking aldosterone synthase and is a potential candidate to prevent skin aging.

Effect of 20-hydroxyecdysone on the immune response following eccentric muscle injury in C57BL/6 mice

Previously, we demonstrated that the phytoecdysteroid, 20-hydroxyecdysone (20E), accelerates the recovery of skeletal muscle function 7 days following eccentric contraction-induced injury. We hypothesized that 20E may influence the inflammatory response during recovery from eccentric muscle injury; therefore, male mice (3-6 mo) were subjected to eccentric damage of the anterior crural muscles via 150 eccentric contractions, then treated with either 20E (50mg/kg body mass) or vehicle daily until sacrifice (12 hours, 2 days or 4 days post-injury). Using immunohistochemical techniques, infiltration of neutrophils, M1-like, and M2-like macrophages were examined in sectioned tibialis anterior muscle. Our findings revealed that pro-inflammatory responses (infiltration of neutrophils and M1-like macrophages) appear to be attenuated in 20E-treated mice at 12 hours and 2 days post-injury, respectively; whereas anti-inflammatory responses (infiltration of M2-like macrophages) appeared to increase with 20E treatment at 4 days after eccentric muscle injury. Taken together, it is possible that 20E may attenuate the overall early pro-inflammatory response, while bolstering the later anti-inflammatory response to eccentric muscle injury, leading to accelerated muscle repair/regeneration. This research was partly funded by the Appalachian State University Office of Student Research. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Gene annotation of nuclear receptor superfamily genes in the kissing bug Rhodnius prolixus and the effects of 20-hydroxyecdysone on lipid metabolism.

The hormone 20-hydroxyecdysone is fundamental for regulating moulting and metamorphosis in immature insects, and it plays a role in physiological regulation in adult insects. This hormone acts by binding and activating a receptor, the ecdysone receptor, which is part of the nuclear receptor gene superfamily. Here, we analyse the genome of the kissing bug Rhodnius prolixus to annotate the nuclear receptor superfamily genes. The R. prolixus genome displays a possible duplication of the HNF4 gene. All the analysed insect organs express most nuclear receptor genes as shown by RT-PCR. The quantitative PCR analysis showed that the RpEcR and RpUSP genes are highly expressed in the testis, while the RpHNF4-1 and RpHNF4-2 genes are more active in the fat body and ovaries and in the anterior midgut, respectively. Feeding does not induce detectable changes in the expression of these genes in the fat body. However, the expression of the RpHNF4-2 gene is always higher than that of RpHNF4-1. Treating adult females with 20-hydroxyecdysone increased the amount of triacylglycerol stored in the fat bodies by increasing their lipogenic capacity. These results indicate that 20-hydroxyecdysone acts on the lipid metabolism of adult insects, although the underlying mechanism is not clear.

Effect of exogenously applied 20-hydroxyecdysone on the efficiency of primary photosynthetic processes substantially differs across plant species

The effects of 20-hydroxyecdysone on the primary photosynthetic processes (PPP) were examined across twelve plant species using the OJIP chlorophyll fluorescence analysis. Our experiments brought evidence that the ecdysteroids can function not only as protective compounds in plants but participate also in the regulation of photosynthesis. Exogenously applied 20-hydroxyecdysone positively affected the PPP particularly in French bean and canola. Contrary to this, the PPP were negatively influenced by 20-hydroxyecdysone in pea, sunflower, and wheat. The performace of the oxygen-evolving complex, the excitonic connectivity among PSII units, the electron transport between PSII and PSI, the size of the pool of the end electron acceptors, and the absorption and trapping of the excitation energy in the light-harvesting antennae were differently affected in individual plant species. Additionally, the older and younger leaves did not respond to 20-hydroxyecdysone in the same way and this was also a species-specific phenomenon.

Effect of 20-Hydroxyecdysone, a Phytoecdysteroid, on Development, Digestive, and Detoxification Enzyme Activities of Tribolium castaneum (Coleoptera: Tenebrionidae).

Plants present a delimited reservoir of biologically active compounds. Many plants synthesize several compounds of secondary metabolism, such as alkaloids, terpenoids, phenolics, steroids, etc. Such compounds are generally thought to be involved in plant–insect interactions. Phytoecdysteroids are a class of chemicals that plants synthesize; these compounds are analogues of molting hormones produced by insects. In this work, the effect of the 20-hydroxyecdysone, which is a molecule that belongs to the family of phytoecdysteroids, was tested on an insect pest, Tribolium castaneum (Herbst). Firstly, the effect of this molecule on post-embryonic development parameters was tested after ingestion at 300, 600, 900, and 1,200 ppm. Secondly, the effect of the 20-hydroxyecdysone was also tested on the biological parameters (proteins, alpha-amylase, detoxification enzymes). The results of the post-embryonic parameters test showed an important induction of larval mortality and a significant reduction of pupation and adult emergence rates. On the other hand, the test on the biological parameters showed that the 20-hydroxyecdysone caused a significant decrease in the levels of soluble proteins in treated larvae. In addition, the alpha-amylase activity was significantly inhibited by the ingestion of the phytoecdysteroid. And there was also a disruption of detoxification enzymes. The whole of the disturbances recorded in this work prove that phytoecdysteroids are thought to have potential value on T. castaneum control.

Molecular cloning, gene expression analysis, and in silico characterization of UDP-N-acetylglucosamine pyrophosphorylase from Bombyx mori.

The present study was aimed to explore the molecular and structural features of UDP-N-acetylglucosamine pyrophosphorylase of Bombyx mori (BmUAP), an essential enzyme for chitin synthesis in insects. The BmUAP cDNA sequence was cloned and expression profiles were monitored during the molting and feeding stages of silkworm larvae. The effect of 20-hydroxyecdysone (20E) on BmUAP expression, and on silkworm molting was studied, which revealed that 20E regulates its expression. Multiple sequence alignment of various pyrophosphorylases revealed that the residues N223, G290, N327, and K407 of human UAP (PDB ID: 1JV1) were found to be highly conserved in BmUAP and all other eukaryotic UAPs considered for the study. Phylogenetic analysis inferred that the UAPs possess discrete variations in primary structure among different insect Orders while sharing good identity between species of the Order. The structure of BmUAP was predicted and its interactions with uridine triphosphate, N-acetylglucosamine-1-phosphate, and UDP-N-acetylglucosamine were analyzed. Virtual screening with a library of natural compounds resulted in five potential hits with good binding affinities. On further analysis, these five hits were found to be mimicking substrate and product, in inducing conformational changes in the active site. This work provides crucial information on molecular interactions and structural dynamics of insect UAPs.

Effects of 20-hydroxyecdysone on the development and morphology of the red flour beetle, Tribolium castaneum (Coleoptera: Tenebrionidae)

The red flour beetle, Tribolium castaneum, is a pest of stored products. It is also regarded as a model species for studying development, genetics, biology, physiology and biochemistry. Recently, it has become a model for use in RNA interference experiments. 20-hydroxyecdysone (20E) is involved in insect metamorphosis and its role in organ development in T. castaneum are based on hormonal treatment in conjunction with RNAi. However, information on the biological, morphological and physiological effects of 20E on T. castaneum is still limited. This study reveals the responses of T. castaneum larvae to injections with various doses of 20E (100, 200, 300, 400 and 500 ng / insect). The results show that larvae injected with 20E reached the prepupal, pupal and adult stages earlier than the control group. Different degrees of morphological change were observed in nine traits, including the appearance of pupal prothetelic organs in the larvae. Moreover, an injection of a high dose of 20E reduced the body weights of the resulting insects at each stage, as well as the length and width of elytra. The enzymatic activity of α-amylase in the resulting adults also decreased significantly. This indicates that injection of 20E caused precocious metamorphosis in T. castaneum by inducing changes in morphology and α-amylase activity, and the optimal concentrations that induce such phenomena were in the range of 100-200 ng / insect. Further investigations are needed to examine the roles of 20E in the regulation of α-amylase in T. castaneum.

Voltage-sensitive potassium channels expressed after 20-Hydroxyecdysone treatment of a mosquito cell line

The goal of this research was to express receptors and ion channels in hormone-treated insect cell lines. Treatment of Anopheles gambiae Sua1B cells with 20-hydroxyecdysone showed an inhibition of cell growth over a time course of three days, with no change in cellular morphology. The effect of 20-hydroxyecdysone was enhanced in the presence of the potassium channel blocker 4-aminopyridine, but not tetraethylammonium. Concentration-response curves of 4-aminopyridine in the presence of 42 μM (1 mg/ml) 20-hydroxyecdysone showed similar IC50 values (6–10 μM) across 3 day exposures. Whole cell patch clamp confirmed the expression of delayed-rectifier (Kv2) potassium channels in hormone-supplemented Sua1B cells, whereas untreated Sua1B cells showed no evidence of Kv2 expression. The hormone-induced expression of Kv2 channels occurred in as little as 4 h after treatment, but were not observed after 24 h of exposure to 20-hydroxyecdysone, suggesting they played a role in cell death. The expressed channels had current-voltage relationships diagnostic for the Kv2 subtype, and were inhibited with an IC50 = 13 mM of tetraethylammonium. Overall, these parameters were similar to Anopheles gambiae Kv2 potassium channels expressed in HEK-293 cells. The induced presence of ion channels (and possibly receptors) in these cells has potential utility for high throughput screening and basic neuroscience research.

Effect of 20-Hydroxyecdysone on Proteolytic Regulation in Skeletal Muscle Atrophy.

20-Hydroxyecdystone (20E) is an ecdysteroid hormone which controls molting and reproduction in arthropods. 20E also produces a variety of effects in vertebrates, including enhancing protein synthesis and skeletal muscle regeneration. The effect of 20E on disuse muscle atrophy has not been reported to date. This study examined the proteolytic regulation of 20E in tenotomized rat slow soleus and fast plantaris muscles. Male Wistar rats were randomly divided into three groups: sedentary control (CON), tenotomy without 20E treatment (TEN), and tenotomy with treatment of 5 mg/kg BW of 20E (TEN+20E). The TEN+20E group was administered 20E via subcutaneous injection to the right thigh for 7 days after tenotomy. 20E treatment tended to attenuate disuse muscle atrophy and reduced ubiquitination only in soleus muscle. 20E treatment alleviates skeletal muscle atrophy partially mediated by ubiquitinate pathway, dependent on the muscle phenotype.

The effect of 20-hydroxyecdysone on the susceptibility of Escherichia coli to different antibiotics.

The aim of this study was to investigate the influence of 20-hydroxyecdysone (20E) on the susceptibility of growing Escherichia coli to antibiotics. Susceptibility of E.coli to antibiotics in the presence of 20E was estimated by determination of the colony-forming ability and the specific growth rate. Pretreatment with 20E decreased the bactericidal effect of ciprofloxacin (0·3 and 3·0μgml-1 ), streptomycin (10 and 40μgml-1 ) and kanamycin (10μgml-1 ) and increased the bactericidal action of 0·03μgml-1 ciprofloxacin. To study the influence of 20E on gene expression, we used strains of E.coli carrying fusions of promoters of relevant genes with the structural gene of β-galactosidase. 20E had no marked effect on the expression of antioxidant genes katG, katE, sodA and the rpoS gene (general stress response), while it (alone or combined with ciprofloxacin) markedly stimulated expression of the sulA gene belonging to the SOS regulon (DNA damage response). In growing E.coli, 20E modulated the bactericidal action of antibiotics and stimulated the SOS response. The results of this study may be used to enhance the efficiency of antibacterial therapies.

Roles of ecdysteroids for progression of reproductive cycle in the fresh water crustacean Daphnia magna

Daphnia magna exhibits a parthenogenetic reproductive cycle linked to a moulting cycle, but regulatory mechanisms of neither moulting nor reproductive cycle are understood in daphnids. Moulting is regulated by ecdysteroids in insects. A previous study showed that a titre of ecdysteroids changed during the reproductive cycle in D. magna; however, no clear correlation among titre, moulting and reproductive cycles has been proved in daphnids. To understand endocrine mechanisms underlying the coordinated reproductive cycle, we analysed the expression of genes coding for enzymes in ecdysteroids synthesis or inactivation pathways, and the effects of 20-hydroxyecdysone (20E) on moulting and ovulation in D. magna. We cloned orthologues of neverland (nvd) and shade (shd) in the ecdysteroids synthesis pathway, and Cyp18a1 in the ecdysteroids inactivation pathway previously identified in insects. Gene expression of Cyp18a1 changed conversely with the fluctuation in ecdysteroids titre during the intermoulting period. Tissue-specific expression analysis of nvd showed a prominent expression in the gut. Furthermore, treatment of adult female D. magna with 20E inhibited moulting and/or ovulation. Our cloning and phylogenetic analyses showed that nvd and shd as well as Cyp18a1 are evolutionary conserved in D. magna, suggesting that these genes appeared in arthropods before the radiation of insects. The gene expression analysis during the reproductive cycle indicated that Cyp18a1 possibly regulates the decline of ecdysteroid titre before moulting and ovulation. Furthermore, the expression of nvd in the gut suggested that ecdysone might be synthesised in the gut. Exogenous 20E-treatment resulted in the failure of not only moulting, but also ovulation, suggesting that a low level of ecdysteroids before moulting is required for moulting and ovulation in D. magna.

Protective effects of 20-hydroxyecdysone on H2O2-induced cytotoxicity in human neuroblastoma SH-SY5Y cells

Protective effects of 20-hydroxyecdysone on H2O2-induced cytotoxicity in human neuroblastoma SH-SY5Y cells

Molecular characterization of four actin cDNAs and effects of 20-hydroxyecdysone on their expression in swimming crab, Portunus trituberculatus (Miers, 1876)

Crustacean muscle fibers exhibit a high level of plasticity in various growth and developmental stages and understanding its mechanism is one of the important factors to improve the commercial value of the decapod products including shrimp, lobsters, and crabs. In the present study, four cDNAs encoding actins (PotActinSK1, PotActinSK2, PotActinHT, and PotActinCT) were identified from the swimming crab (Portunus trituberculatus) using a PCR-based cloning strategy. Although the four cDNA encoding actins exhibit similarities in size and deduced amino acid sequences, their tissue expression profiles differ significantly from one another. Phylogenetic analysis revealed that muscular and cytoplasmic actins have been diverged before vertebrates were evolved and cardiac and skeletal muscular actins were branched out after the vertebrates were evolved. Based on the phylogenetic analysis and expression patterns, PotActinSK1 and SK2 belong to the slow-type skeletal muscle actins and the fast-type skeletal muscle actin has not been identified. PotActinHT showed the highest sequence similarity to Ha-ActinHT from H. americanus and appeared to be an ortholog of heart muscle actins in decapod crustaceans. PotActinCT exhibited the highest sequence similarity to the cytoplasmic actin of L. vannamei ActinT1. End-point RT-PCR results showed that PotActinSK1 and PotActinSK2 were strongly expressed in skeletal muscular tissues including chela propodus muscle, chela merus muscle, leg muscle, thoracic muscle, and there was no significant difference in the expression ratio between the two genes of those four tissues. PotActinCT was ubiquitously expressed in all the tissues, whereas PotActinHT was exclusively expressed in the heart. Interestingly, a considerable level of PotActinCT was expressed in the heart, which is similar to those of PotActinHT. The transcriptional response to 20-hydroxyecdysone (20E) injection was different in each PotActin suggesting that effects of muscle plasticity by 20E may differ in each tissue of decapod crustaceans.

The glutathione-related detoxication responses to juvenile and ecdysone hormones in Galleria mellonella

The effect of 20-hydroxyecdysone (20E) and juvenile hormone (JH) on the glutathione pathway of the greater wax moth Galleria mellonella (Lepidoptera: Pyralidae) was determined by investigating glutathione peroxidase (GSH-Px), glutathione S-transferases (GST), and glutathione reductase (GR) activities as well as reduced and oxidized glutathione (GSH and GSSG) content with respect to developmental stage. The continuous decreases of GSH-Px and GST activities dependent on the growth period of G. mellonella occurred in JH and 20E groups over and under their controls, respectively. While the GR activities of G. mellonella showed increases in young pupa (YP) for both control and in old larvae (OL) for the 20E groups after the minimum at these periods, they also increased after old pupa (OP) for the JH group with a maximum in OL period. Although GR activity levels in the JH group were significantly higher compared with controls and 20E groups up to OP period, the activity levels for the control and 20E groups were higher than those of the JH group at adult (AD) and old pupa (OP) periods, respectively. In spite of increases in the GR activity of 20E and control groups of G. mellonella, decreased GSH and increased GSSG levels were observed at aging period. GSH levels in the JH group reached a maximum at prepupa (PP) and then decreased with non-significant changes from OL to AD period. According to the results, GSH and GSSG levels, as well as GSH/GSSG ratios, were below and over control levels in 20E and JH groups, respectively, during all of the investigated developmental stages. On the contrary, the LPO levels were higher than the control for 20E and lower for the JH groups during the developmental period. These results show that while ecdysone hormone has a negative effect on the glutathione-related detoxication capacity of G. mellonella, the juvenile hormone has a positive effect on this process.

Effect of 20-hydroxyecdysone on glucose and lipid metabolism in vitro

Ecdysteroids such as 20-hydroxyecdysone (20HE) are commonly found in the highly nutritious Andean pseudocereal quinoa (Chenopodium quinoa Willd.). 20HE and an ecdysteroid-enriched extract from quinoa have been shown to improve insulin sensitivity and decrease adiposity in diet-induced obese mice. One proposed mechanism of action for this in vivo effect is 20HE's negative impact on hepatic glucose output. However, associated anti-diabetic and anti-obesity mechanims, including enhanced insulin sensitivity in peripheral tissues and changes to adipogenesis or lipid metabolism in adipose tissue, have not been reported. Here, we tested the effect of this molecule on these parameters by measuring (a) glucose uptake and insulin sensitivity in L6 rat myotubes, (b) lipid accumulation in 3T3-L1 mouse adipocytes, and (c) lipolysis in 3T3-L1 adipocytes. We also confirmed the impact of 20HE on glucose production in HII4E rat hepatocytes and showed that several fractions of a crude ethanolic extract from quinoa containing ecdysteroids similarly decrease glucose output in this model. Our data suggest that the complex mixture of secondary metabolites found in quinoa may play a synergistic role in the improvement of physiologic parameters associated with metabolic syndrome, and quinoa may be an important functional food for the treatment and prevention of diabetes and obesity.

Effects of 20-hydroxyecdysone, Leuzea carthamoides extracts, dexamethasone and their combinations on the NF-κB activation in HeLa cells

The plant steroid 20-hydroxyecdysterone (20E) and 20E-containing extracts from Leuzea carthamoides (Willd.) DC are sold with claims of anabolic and immunomodulatory effects. Yet their effect on the activation of nuclear factor kappa B (NF-κB), a key player in immune response and cell fate, and their influence on the NF-κB-inhibiting activity of steroidal anti-inflammatory drugs is still unknown. The ability of 20E, Leuzea extracts and selected steroidal/non-steroidal anti-inflammatory drugs to influence the activation of NF-κB was explored using, as the experimental model, human cervical cancer HeLa-IL-6 cells stably transfected with an IL-6-bound reporter gene. Effects on cell viability and proliferation were monitored (MTT assay). HPLC-DAD was used to establish links between chemical patterns of Leuzea extracts and their bioactivities. 20E inhibited NF-κB activation (IC50 31.8 µm) but was less active than other plant metabolites (xanthohumol 3.8 µm, withaferin A 1.4 µm). Leuzea extracts with high content in 20E had a fair activating effect, but in contrast, some extracts with low 20E content significantly inhibited NF-κB activation at IC50s ranging from 3.5 to 6.2 µg/ml. Combination tests confirmed that 20E does not explain the NF-κB modulation achieved by Leuzea extracts. The extracts but not 20E itself showed a significant modulation of the NF-κB inhibitory effect of dexamethasone. 20E is unlikely a major player in the NF-κB inhibitory effects displayed by some Leuzea extracts in vitro. If confirmed in vivo, caution should prevail towards marketed Leuzea extracts that are non-standardised or standardised on 20E only, since different starting materials and extracts may even cause opposite effects. More importantly, our results indicate the interaction potential of Leuzea with steroidal anti-inflammatory drugs.

The effects of 20-hydroxyecdysone and 17β-estradiol on the skin of ovariectomized rats

20-hydroxyecdysone has numerous favorable effects on a variety of organs, including the skin, where it improves wound healing. It is devoid of estrogenic and androgenic effects. Therefore, application of 20-hydroxyecdysone might be a new approach to improve skin conditions in postmenopausal women, and this was investigated in ovariectomized (OVX) rats. After ovariectomy, rats received Ecd (18, 57, or 116 mg/animal/day) or 17β-estradiol (E₂)-3-benzoate (60 μg/kg body weight) in food for 12 weeks, and skin samples were evaluated histologically to quantify two dermal layers, the subcutaneous fat and muscle layers. Epidermal thickness was lowest in the OVX animals, slightly higher in the E₂-treated animals, and significantly higher in the Ecd-treated animals. Dermal thickness was lowest in the intact and E₂-treated animals and highest in the Ecd-treated animals. The subcutaneous fat layer was thickest in the OVX animals, thinner in the intact animals, and intermediate in the Ecd-treated animals. The muscle layer was smallest in the OVX and intact animals and significantly larger in the E₂- and Ecd-treated animals. The number of proliferating cell nuclear antigen antibody-positive cells was lowest in OVX controls and significantly higher in all other groups. The Ecd-induced increases in epidermal and dermal thickness are suggestive of functional changes of the skin. The decreased amounts of subcutaneous fat in the E₂- and Ecd-treated animals point to either a fat catabolic or an antianabolic effect. The ovariectomy-induced decrease in subcutaneous musculature was prevented by Ecd but not by E₂. The stimulatory effects of Ecd on epidermal and dermal thickness and the muscle-increasing effects in the skin of OVX rats may indicate functional changes of the skin.

Identification and expression analysis of the Broad-Complex core protein isoform 6 (BR-C Z6) gene in the giant tiger shrimp Penaeus monodon (Penaeidae: Decapoda)

Broad-Complex (BR-C) is an early ecdysone-responsive gene encoding a family of zinc-finger transcription factors that function during metamorphosis in insects. We identified two full-length cDNAs of BR-C Z6 in the giant tiger shrimp (Penaeus monodon). They were 2422 and 2060 bp in length, containing open reading frames of 1440 and 1443 bp, corresponding to polypeptides of 479 and 480 amino acids, respectively. Tissue distribution analysis indicated that PmBR-C Z6 was abundantly expressed in hemocytes and ovaries in juveniles. In broodstock, PmBR-C Z6 was constitutively expressed in all tissues examined, and the highest expression was observed in ovaries. The expression of PmBR-C Z6 in ovaries was significantly greater than in testes in both juveniles and broodstock of P. monodon. Quantitative real-time PCR indicated that the expression level of PmBR-C Z6 was significantly down-regulated in stages II and III of ovaries in intact wild broodstock and returned to the basal level in stage IV ovaries and after spawning. In eyestalk-ablated broodstock, PmBR-C Z6 was significantly up-regulated in stage IV (mature) ovaries. Moreover, the expression level of PmBR-C Z6 in vitellogenic, early cortical rod and mature (stages II-IV) ovaries of eyestalk-ablated broodstock was greater than that of the same ovarian stages in intact broodstock. In situ hybridization revealed that PmBR-C Z6 transcripts were localized in oogonia and cytoplasm of previtellogenic and vitellogenic oocytes of both wild intact and eyestalk-abated broodstock. The effects of 20-hydroxyecdysone on expression of PmBR-C Z6 were examined. The expression level of PmBR-C Z6 in ovaries of juvenile P. monodon was significantly increased at 168 h post-injection. Taken together, these findings indicate that PmBR-C Z6 plays an important role in ovarian development of P. monodon.