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Forkhead box H1 (FoxH1) is a sexually dimorphic gene in Oreochromis niloticus, Oplegnathus fasciatus, and Acanthopagrus latus, indicating that it is essential for gonadal development. In the present study, the molecular characteristics and potential function of FoxH1 and the activation of the cyp19a1a promoter in vitro were evaluated in Monopterus albus. The levels of foxh1 in the ovaries were three times higher than those in the testes and were regulated by gonadotropins (Follicle-Stimulating Hormone and Human Chorionic Gonadotropin). FoxH1 colocalized with Cyp19a1a in the oocytes and granulosa cells of middle and late vitellogenic follicles. In addition, three FoxH1 binding sites were identified in the proximal promoter of cyp19a1a, namely, FH1 (−871/−860), FH2 (−535/−524), and FH3 (−218/−207). FoxH1 overexpression significantly attenuated the activity of the cyp19a1a promoter in CHO cells, and FH1/2 mutation increased promoter activity. Taken together, these results suggest that FoxH1 may act as an important regulator in the ovarian development of M. albus by repressing cyp19a1a promoter activity, which provides a foundation for the study of FoxH1 function in bony fish reproductive processes.

tRNA-derived small RNAs (tsRNAs) are derived from tRNA and include tRNA halves (tiRNAs) and tRNA fragments (tRFs). tsRNAs have been implicated in a variety of important biological functions, such as cell growth, transcriptional regulation, and apoptosis. Emerging evidence has shown that Ago1-guided and Ago2-guided tsRNAs are expressed at 3 and 30 days in Drosophila and that tRF biogenesis in fruit flies affects tRNA processing and tRNA methylation. However, a wide analysis of tsRNA patterns in different ages of Drosophila have not been reported via the small RNA sequencing method. In the present study, tsRNAs of young (7 days) and old (42 days) Drosophila were sequenced and their expression characteristics were analysed. Then, a specific tRF (named tRF-Trp-CCA-014) was determined and was found to be conserved in fruit flies, mice, and humans. The expression patterns of tRF-Trp-CCA-014 in different tissues and stages of fruit flies and mice, and mouse NIH/3T3 cells were detected. Furthermore, mouse embryonic fibroblast NIH/3T3 cells were used as a model to analyse the function and targets of tRF-Trp-CCA-014. The RNA-seq data of six groups (Mimics, Mimic NC, Inhibitors, Inhibitor NC, Aging (adriamycin), and Control (Normal)) in mouse NIH3T3 cells were analysed. The results showed that the number of tsRNAs at 42 days (417) was more than at 7 days (288); thus, it was enriched with age. tRFs-1 were the most enriched, followed by 5′-tRFs and 3′-tRFs. Twenty-one differentially expressed tsRNAs were identified between 7 days and 42 days. Then, the conserved tRF tRF-Trp-CCA-014 was identified and found to accumulate in aged fruit flies and aged mouse NIH3T3 cells. RNA-seq data showed that most differentially expressed genes were involved in the immune system, cancer: overview, and signal translation. Furthermore, tRF-Trp-CCA-014 was found to bind to the 3′UTR of H3C4 in a dual-luciferase reporter gene assay. tRF-Trp-CCA-014 and H3C4 were detected in the cytoplasm of aged NIH3T3 cells by RNA in situ hybridization. These results suggest that the H3C4 gene is the target of tRF-Trp-CCA-014. This study will advance the current understanding of tRF roles and their implication in Drosophila and mouse studies.

Bmpr2 plays a central role in the regulation of reproductive development in mammals, but its role during ovarian development in fish is still unclear. To ascertain the function of bmpr2 in ovarian development in the ricefield eel, we isolated and characterized the bmpr2 cDNA sequence; the localization of Bmpr2 protein was determined by immunohistochemical staining; and the expression patterns of bmpr2 in ovarian tissue incubated with FSH and hCG in vitro were analyzed. The full-length bmpr2 cDNA was 3311 bp, with 1061 amino acids encoded. Compared to other tissues, bmpr2 was abundantly expressed in the ovary and highly expressed in the early yolk accumulation (EV) stages of the ovary. In addition, a positive signal for Bmpr2 was detected in the cytoplasm of oocytes in primary growth (PG) and EV stages. In vitro, the expression level of gdf9, the ligand of bmpr2, in the 10 ng/mL FSH treatment group was significantly higher after incubation for 4 h than after incubation for different durations. However, bmpr2 expression in the 10 ng/mL FSH treatment group at 2 h, 4 h and 10 h was significantly lower. Importantly, the expression level of bmpr2 and gdf9 in the 100 IU/mL hCG group had similar changes that were significantly decreased at 4 h and 10 h. In summary, Bmpr2 might play a pivotal role in ovarian growth in the ricefield eel, and these results provide a better understanding of the function of bmpr2 in ovarian development and the basic data for further exploration of the regulatory mechanism of gdf9 in oocyte development.

Apoptosis plays a key role in the effective removal of excessive and defective germ cells, which is essential for sequential hermaphroditism and sex change in vertebrates. The ricefield eel, Monopterus albus is a protogynous hermaphroditic fish that undergoes a sequential sex change from female to male. Previous studies have demonstrated that apoptosis is involved in sex change in M. albus. However, the apoptotic signaling pathway is unclear. In the current study, we explored the underlying mechanism of apoptosis during gonadal development and focused on the role of the mitochondrial apoptosis signaling pathway in sex change in M. albus. Flow cytometry was performed to detect apoptosis in gonads at five sexual stages and ovary tissues exposed to hydrogen peroxide (H2O2) in vitro. Then the expression patterns of key genes and proteins in the mitochondrial pathway, death receptor pathway and endoplasmic reticulum (ER) pathway were examined. The results showed that the apoptosis rate was significantly increased in the early intersexual stage and then decreased with the natural sex change from female to male. Quantitative real-time PCR revealed that bax, tnfr1, and calpain were mainly expressed in the five stages. ELISA demonstrated that the relative content of cytochrome-c (cyt-c) in the mitochondrial pathway was significantly higher than that of caspase8 and caspase12, with a peak in the early intersexual stage, while the levels of caspase8 and caspase12 peaked in the late intersexual stage. Interestingly, the Pearson’s coefficient between cyt-c and the apoptosis rate was 0.705, which suggests that these factors are closely related during the gonadal development of M. albus. Furthermore, the cyt-c signal was found to be increased in the intersexual stage by immunohistochemistry. After incubation with H2O2, the mRNA expression of mitochondrial pathway molecules such as bax, apaf-1, and caspase3 increased in ovary tissues. In conclusion, the present results suggest that the mitochondrial apoptotic pathway may play a more important role than the other apoptotic pathways in sex change in M. albus.

Many studies have revealed that circulating long noncoding RNAs (lncRNAs) regulate gene and protein expression in the process of hepatic fibrosis. Liver fibrosis is a reversible wound healing response followed by excessive extracellular matrix accumulation. In the development of liver fibrosis, some lncRNAs regulate diverse cellular processes by acting as competing endogenous RNAs (ceRNAs) and binding proteins. Previous investigations demonstrated that overexpression of lncRNAs such as H19, maternally expressed gene 3 (MEG3), growth arrest-specific transcript 5 (GAS5), Gm5091, NR_002155.1, and HIF 1alpha-antisense RNA 1 (HIF1A-AS1) can inhibit the progression of liver fibrosis. Furthermore, the upregulation of several lncRNAs [e.g., nuclear paraspeckle assembly transcript 1 (NEAT1), hox transcript antisense RNA (Hotair), and liver-enriched fibrosis-associated lncRNA1 (lnc-LFAR1)] has been reported to promote liver fibrosis. This review will focus on the functions and mechanisms of lncRNAs, the lncRNA transcriptome profile of liver fibrosis, and the main lncRNAs involved in the signalling pathways that regulate hepatic fibrosis. This review provides insight into the screening of therapeutic and diagnostic markers of liver fibrosis.