1,720,988 research outputs found
Long-term culture of muscle explants from Sparus aurata
No full textAlthough there are mammalian myoblast cell lines, no fish myoblast cell line has been developed so far. The aim of this study was to develop a culture system of muscle explants for fish, as explants provide an approximation of the in vivo conditions for cell proliferation and differentiation, and enable a close comparison with events in muscle regenerating in vivo. Here we describe the main features of a long-term in vitro culture system for muscle explants from Sparus aurata fry. At the time of sampling, the original fibres were damaged and subsequently degenerated as shown by the loss of parvalbumin (PV) and presence of apoptotic nuclei. This mechanical damage provoked a myogenic response by activation of myogenic precursor cells. After a few days, new mononucleate cells aligned with the original fibres were seen in the explants, some with proliferating cell nuclear antigen (PCNA-) and Myf-5-positive nuclei, indicating proliferation and their myogenic fate. By 1 week, multinucleate cells with desmin immunoreactivity but PCNA- and Myf5-negative nuclei were present, equivalent to differentiated, postmitotic myotubes. Some of these myotubes were also immunoreactive for PV and insulin-like growth factors (IGFs). By 11 days, many of the myotubes were also immunoreactive for myostatin (MSTN). By 23 days, many of the myotubes had increased in diameter, were packed with myofibrils, and were strongly PV-positive and immunoreactive for MSTN, IGF-I and IGF-I receptor. This study shows that a proliferative process occurs in the explants despite the death of the original muscle fibres, and new muscle fibres expressing growth regulators are formed by regeneration from myogenic precursors present in the explants at the time of sampling
Cellular localization of insulin-like growth factor-II protein in the sea bass (Dicentrarchus labrax) from hatching to adult
No full textThe cellular localization of IGF-II protein was investigated during larval and postlarval developmental stages of sea bass (Dicentrarchus labrax) by immunohistochemistry using antisera raised against Sparus aurata IGF-II. At hatching, IGF-II immunoreactivity was already present in the skin, developing intestine and skeletal muscle. During larval life IGF-II protein was also observed in heart musculature, in kidney and gill epithelia as well as in liver. In fry skeletal muscle a moderate IGF-II immunostaining was detected in red fibres, whereas white muscle fibres exhibited a faint immunoreactivity. In adults, a marked IGF-II immunostaining was observed in red muscle fibres. A moderate immunoreactivity was also present in white fibres as well as in heart striated myocardial fibres. These results are in agreement with previous findings on the spatial localization of IGF-II and IGF type 1 receptor in S. aurata and Umbrina cirrosa, confirming the role of IGF system during development and growth of fish
Characterization and expression of Insulin-like Growth Factors (IGF-I and IGF-II) in the marine teleost, Umbrina cirrosa
No full textGenomic cloning and promoter functional analysis of myostatin-2 in shi drum, Umbrina cirrosa: conservation of muscle-specific promoter activity.
No full textMyostatin (MSTN) is a member of the transforming growth factor-ß superfamily, known as a negative regulator of skeletal muscle development and growth in mammals. In contrast to mammals, fish possess at least two paralogs of MSTN: MSTN-1 and MSTN-2. Here we describe the cloning and sequence analysis of spliced and precursor (unspliced) transcripts as well as the 5' flanking region of MSTN-2 from the marine fish Umbrina cirrosa (ucMSTN-2). In silico analysis revealed numerous putative cis regulatory elements including several E-boxes known as binding sites to myogenic transcription factors. Transient transfection experiments using non-muscle and muscle cell lines showed high transcriptional activity in muscle cells and in differentiated neural cells, in accordance with our previous findings in MSTN-2 promoter from Sparus aurata. Comparative informatics analysis of MSTN-2 from several fish species revealed high conservation of the predicted amino acid sequence as well as the gene structure (exon length) although intron length varied between species. The proximal promoter of MSTN-2 gene was found to be conserved among Perciforms. In conclusion, this study reinforces our conclusion that MSTN-2 promoter is a very strong promoter, especially in muscle cells. In addition, we show that the MSTN-2 gene structure is highly conserved among fishes as is the predicted amino acid sequence of the peptide
Structure, gene expression, and production of recombinant follistatin from the marine fish Sparus aurata.
No full textMyostatin cellular localization in several tissues of teleost fish: an immunohistochemical study
No full text
Expression and cellular localization of insulin-like growth factor-II protein and mRNA in Sparus aurata during development
No full textThe spatial localization of IGF-II protein and mRNA was investigated during larval and postlarval developmental stages of the gilthead sea bream (Sparus aurata) by immunohistochemistry and in situ hybridization, using specific antisera and riboprobes. Steady-state levels of IGF-II mRNA in larvae were determined by Northern blot analysis and were found to be increased. Immunoreactivity towards IGF-II was found in larval skin, muscle, gills, gut, olfactory epithelium and kidney. After metamorphosis, the strongest immunoreactivity was found in red skeletal muscle. Positive reaction with IGF-II antibodies was also found in the olfactory epithelium and in the epithelia of pharynx, oesophagus, stomach and kidney. In the adult, the most intense signal was observed in the red and pink musculature and in heart musculature. Immunostaining was also found in saccus vasculosus, thymus, spleen and ovary. IGF-II mRNA was detected by in situ hybridization in the brain, olfactory epithelium, eye, pharynx, skeletal musculature and liver. The spatial distribution of IGF-II shown in this study is consistent with previous findings on the cellular localization of IGF type 1 receptor in the sea bream and supports a role for IGF-II during development and growth of sea bream. Furthermore, these results suggest that IGF-II acts in an autocrine/paracrine manner
Insulin-like growth factor-I and other neuromediators in the gut of Umbrina cirrosa (Pisces: Osteichthyes)
No full text- …
