Effects of salinity acclimation on histological characteristics and miRNA expression profiles of scales in juvenile rainbow trout (Oncorhynchus mykiss).

BACKGROUND: The scales serve as an ideal model for studying the regulatory mechanism of bone homeostasis in fish. To explore the effect of salinity acclimation on bone metabolism of juvenile rainbow trout (Oncorhynchus mykiss), three sampling time points during salinity acclimation (7D, 14D and 21D) were selected to detect variations in histological characteristics. In the histological analysis, osteoblast marker enzymes alkaline phosphatase (ALP), osteoclast marker tartrate-resistant acid phosphatase (TRAcP) and calcium salt deposit areas (Von Kossa's) were detected. Changes in calcium (Ca), phosphorus (P) and the molar mass ratio of calcium to phosphorus (Ca/P) in the scales were also detected by Inductively Coupled Plasma Mass Spectrometry (ICP-MS). In addition, the global MicroRNA (miRNA) expression profiles during salinity acclimation were examined using Illumina sequencing platform because of their important regulatory roles in teleost biological processes. RESULTS: Twelve independent miRNA libraries were constructed, a total of 664 known and 92 putative novel miRNAs were identified. A total of 290 differentially expressed (DE) miRNAs were found in clusters with significant trends in the cluster analysis, and five types of clustering patterns were obtained; 22,374 DE predicted target genes of the aforementioned 290 DE miRNAs were obtained, 5957 of which clustered in six types of clustering patterns with a significant trend. To better understand the functions of the DE miRNAs, GO and KEGG analysis was performed on the 5957 target genes, as a result, they were significantly enriched in bone metabolism related signaling pathways such as MAPK signaling pathway, Calcium signaling pathway, Wnt signaling pathway, Mineral absorption and NF-kappa B signaling pathway. Six DE miRNAs were randomly selected and their expression were verified by quantitative real-time PCR (qRT-PCR), the expression trends were consistent with the results of transcriptome sequencing. CONCLUSIONS: The DE miRNAs and DE target genes identified in this study might play an important role in regulation of bone metabolism during salinity acclimation, relative genes or pathways could serve as key candidates for further studies to elucidate molecular mechanism of teleost bone metabolism, and help performing salinity acclimation and developing marine culture of salmonid species.

Tetramethylpyrazine protects palmitate-induced oxidative damage and mitochondrial dysfunction in C2C12 myotubes.

AIMS: Tetramethylpyrazine (TMP), one of the active ingredients isolated from a Chinese herbal prescription, possesses protective effects against oxidative stress caused by high glucose in endothelial cells. In this study, the role of TMP in preventing muscle cells from palmitate-induced oxidative damage was investigated and the possible mechanisms of action elucidated. MAIN METHODS: Mitochondrial reactive oxygen species (ROS) were measured in C2C12 myotubes, a palmitate-induced oxidative stress cell model, with or without TMP. Both mitochondrial membrane potential (MMP) and oxygen consumption were assessed in conjunction with quantification of mitochondrial DNA and mitochondrial biogenesis-related factors, such as peroxisome proliferator-activated receptor-γ coactivator 1 α (PGC1α), nuclear respiratory factor 1 (NRF1) and mitochondrial transcription factor A (Tfam), by real-time polymerase chain reaction. Expression of mitochondrial respiratory chain complex III as an index of mitochondrial function was evaluated by immunoblotting, and glucose transport into the C2C12 myotube examined by analyzing 2-deoxy-[(3)H]glucose uptake. KEY FINDINGS: TMP significantly alleviated palmitate-induced mitochondrial ROS production, mitigated mitochondrial dysfunction and increased D-loop mRNA expression as compared with the control. This was accompanied by a marked reversal of palmitate-induced down-regulation in the expression of mitochondrial biogenesis-related factors (PGC1α, NRF1 and Tfam) and decreased glucose uptake in C2C12 myotubes. As a result, cell respiration, as reflected by the elevated expression of mitochondrial respiratory chain complex III and oxygen consumption, was enhanced. SIGNIFICANCE: TMP is capable of protecting C2C12 myotubes against palmitate-induced oxidative damage and mitochondrial dysfunction, and improving glucose uptake in muscle cells partially through the up-regulation of mitochondrial biogenesis.