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QNZ is a quinazoline-type NF-κB inhibitor and is one of the hot anti-inflammatory drug candidates in recent years. With its development and application, QNZ will inevitably enter the aquatic environment posing a threat to aquatic organisms. To investigate the potential toxicity of QNZ in the early life stages of the organism, this study exposed embryos of large-scale loach (Paramisgurnus dabryanus) to 0, 20, 40, 60, and 80 nM of QNZ. The hatching of embryos was significantly inhibited and hatching time was delayed. We explored the mechanism of hatching delay and failure. The results suggested that QNZ exposure reduced the number of hatching gland cells (HGCs) and hatching enzyme activity. Also, the frequency of spontaneous movements was inhibited by interfering with the expression of genes related to the cholinergic system and skeletal muscle development. Further, QNZ exposure induces a series of morphological changes (spine deformation, pericardial edema, tail deformation, and yolk sac edema) in embryos and newly-hatched larvae, and finally increased the deformity rate and mortality rate of newly-hatched larvae. The information presented in this study will provide a scientific basis for further studies into the potential toxicity of QNZ on aquatic organisms. [Display omitted] ● QNZ causes increased embryo mortality, hatching inhibition, ultimately abnormal larval development. ● QNZ inhibited the hatching of embryos through decreasing the number of HGCs and activity of hatching enzyme. ● QNZ affected the spontaneous movements of embryos by interference related systems, further inhibiting the hatching. [ABSTRACT FROM AUTHOR]

Sox4 belonged to the SoxC subfamily of the Sox family, which play important roles in the development of the vertebrate gonad and nervous system. A Sox4 homologue was cloned from brain of Paramisgurnus dabryanus by using homologous cloning and rapid amplification of cDNA ends (RACE), designated as PdSox4 . The full-length cDNA was 2163bp, containing the 759bp 5'-untranslated region, 267bp 3'-untranslated region and encoding a putative protein of 378 amino acids with a characteristic high mobility group box (HMG-box) DNA-binding domain of 79 amino acids with the specific motif (RPMNAFMVW). Alignment and phylogenetic analyses indicated that PdSox4 shares highly identical sequence with Sox4 homologues from different species. The signal peptide analysis predicted that PdSox4 is a non-secretory protein. The hydropathy profile of PdSox4 protein revealed that this protein is hydrophilic in nature. The expression profiles of PdSox4 in different developmental stages and various adult tissues of sexs were analyzed by quantitative real-time RT-PCR (qRT-PCR) and In situ hybridization (ISH). The results showed that PdSox4 was ubiquitously expressed during embryogenesis and various adult tissues, especially in central nervous system. Tissue distribution analyses revealed that PdSox4 was expression in developing germ cells. Taken together, these preliminary findings suggested that PdSox4 is highly conserved during vertebrate evolution and involved in a wide range of developmental processes including embryogenesis, neurogenesis and gonad development. [ABSTRACT FROM AUTHOR]

Teleost fins can regenerate accurate position-matched structure and function after amputation. However, we still lack systematic transcriptional profiling and methodologies to understand the molecular basis of fin regeneration. After histological analysis, we established a suppression subtraction hybridization library containing 418 distinct sequences expressed differentially during the process of blastema formation and differentiation in caudal fin regeneration. Genome ontology and comparative analysis of differential distribution of our data and the reference zebrafish genome showed notable subcategories, including multi-organism processes, response to stimuli, extracellular matrix, antioxidant activity, and cell junction function. KEGG pathway analysis allowed the effective identification of relevant genes in those pathways involved in tissue morphogenesis and regeneration, including tight junction, cell adhesion molecules, mTOR and Jak-STAT signaling pathway. From relevant function subcategories and signaling pathways, 78 clones were examined for further Southern-blot hybridization. Then, 17 genes were chosen and characterized using semi-quantitative PCR. Then 4 candidate genes were identified, including F11r , Mmp9 , Agr2 and one without a match to any database. After real-time quantitative PCR, the results showed obvious expression changes in different periods of caudal fin regeneration. We can assume that the 4 candidates, likely valuable genes associated with fin regeneration, deserve additional attention. Thus, our study demonstrated how to investigate the transcript profiles with an emphasis on bioinformatics intervention and how to identify potential genes related to fin regeneration processes. The results also provide a foundation or knowledge for further research into genes and molecular mechanisms of fin regeneration. [ABSTRACT FROM AUTHOR]

Hsp70 is the most strongly induced in response to various cellular stresses and a good candidate for investigating its role in tissue injury. We firstly cloned full-length cDNA of hsp70 from Paramisgurnus dabryanus ( PdHsp70 ) by RACE method (GenBank: KP402408.1 ). Then regeneration and inflammation of fin were established by amputation and scratch respectively. Quantitative RT-PCR detected the PdHsp70 began to increase rapidly its expression at 1 days post amputation (dpa) and reached the peak at 2 dpa during fin regeneration. Its expression was also up-regulated at 2 days post scratch (dps) of inflammation but still significant weaker in comparison with it in regenerated fin at 2 dpa. Next, immunohistochemistry analysis of PdHsp70 showed that PdHsp70 located mainly in the deeper epidermis of regenerated fin and was stronger than its expression in the scratched inflammatory fin which was involved in whole epidermal. SDS-PAGE and Western blotting confirmed that the PdHsp70 protein expressed efficiently in Escherichia coli BL21. These findings have implied that PdHsp70 are implicated in different regulation of regeneration and inflammation in response to injury stimulation. During the regeneration, it is involved in the formation of wound epidermis by mediating cellular protection whereas it can modulate inflammatory by activating the innate immune response. [ABSTRACT FROM AUTHOR]

Pollution of microplastics (MPs) has become a global environmental issue due to the difficulty in its degradation and may cause unexpected ecological effects. Nevertheless, little is known about the potential effects of MPs on reproduction toxicity in aquatic species. In this study, adult loach (Paramisgurnus dabryanus, F0 generation) were exposed to two concentrations (1 and 10 mg/L) of polyethylene MPs (PE-MPs) for 15 or 30 days, and the toxic effects in parental loach and the offspring (F1 generation) were examined. Our results showed that PE-MPs exposure could change the indicators content of antioxidant system in the brain, liver, and gonad. PE-MPs can accumulate in the gonads, disrupt the transcription of HPG-axis related genes, alter sex hormone levels, increase cell apoptosis and gonadal pathological lesions, lead to the damage of biological characteristics of semen, and affect the reproduction in F0 generation. PE-MPs remaining in the parental gonads can be transferred to the F1 generation embryos and accumulated on the embryonic chorionic membrane, increasing mortality and malformation rates, accelerating hatching time, and decreasing hatching rate and body length. These results suggest that PE-MPs leads to a potential adverse influence on reproduction and serious impacts on population sustainability. This work provides a new perspective into the effects of MPs on reproductive damage and cross-generational effects in teleost fish, which have implications in fields of freshwater ecology and environmental toxicology. [Display omitted] • PE-MPs could remain in the gonads and destroyed the reproductive organs. • PE-MPs could disturb the antioxidant, immune and endocrine system. • PE-MPs caused germ cell apoptosis and gamete quality decline. • PE-MPs transferred to the embryo chorionic membrane by parental transmission. • Parental exposure resulted in developmental toxicity of F1 larvae. [ABSTRACT FROM AUTHOR]

Abstract: Heat shock protein 60 (Hsp60) is a kind of highly conserved immunogenic molecule involved in a wide range of biochemical processes in response to external stressors. Its multifunction in regulating immune responses and modulating signal pathway interests us in investigating its role in fin regeneration that has become an excellent and interesting model for studying the molecular basis of morphogenesis. We firstly clarified basical process and crucial period of caudal fins regeneration in Paramisgurnus dabryanus by histological analysis. Then we cloned full-length cDNA of hsp60 from P. dabryanus (designated as PdHsp60) by RACE method. The cDNA contains a 124 bp 5′UTR, a 1731 bp open reading frame (ORF) encoding 576 amino acids and a 510bp 3′UTR (Accession no.: KF544774). The phylogenetic tree shows that the PdHsp60 fits within the hsp60 clade. And quantitative RT-PCR detected the PdHsp60 began to increase rapidly its expression at 1 dpa and reached its peak at 2 dpa. Next, spatial distribution analysis of PdHsp60 in fins showed that PdHsp60 located mainly in the deeper lay of regenerated epidermis when PdHsp60 expressed most. After the PdHsp60 had been cloned into the pET-32a vector, SDS-PAGE and Western blotting analysis confirmed that the PdHsp60 protein was efficiently expressed in Escherichia coli BL21. These findings have revealed that PdHsp60, a highly conserved gene related to the innate immune system and stress response during vertebrate evolution, is involved in response to wounding stimulation—in the formation of wound epidermis which occurs as the first phase of fin regeneration after fin amputation in caudal fin regeneration. [Copyright &y& Elsevier]