Depletion of chop suppresses procedural apoptosis and enhances innate immunity in loach Misgurnus anguillicaudatus under ammonia nitrogen stress.

Abstract

Ammonia nitrogen is highly toxic to fish, and it can easily cause fish poisoning or even high mortality. So far, many studies have been conducted on the damages to fish under ammonia nitrogen stress. However, there are few studies of ammonia tolerance improvement in fish. In this study, the effects of ammonia nitrogen exposure on apoptosis, endoplasmic reticulum (ER) stress, and immune cells in loach Misgurnus anguillicaudatus were investigated. Loaches (60 d post fertilization) were exposed to different concentrations of NH4Cl, and their survival rates were examined every 6 h. The results showed that high-concentration and long-time NH4Cl exposure (20 mM + 18 h; 15 mM + 36 h) induced apoptosis and gill tissue damages, finally causing a decline in survival. chop plays an important role in ER stress-induced apoptosis, and thus we constructed a model of chop-depleted loach by using CRISPR/Cas9 technology to investigate its response to ammonia nitrogen stress. The results showed that ammonia nitrogen stress down-regulated the expressions of apoptosis-related genes in chop+/- loach gills, while wildtype (WT) exhibited an opposite gene expression regulation pattern, suggesting that the depletion of chop suppressed apoptosis level. In addition, chop+/- loach showed a larger number of immunity-related cells and higher survival rate than WT under the NH4Cl exposure, indicating that the inhibition of chop function strengthened the innate immune barrier in general, thus increasing survival. Our findings provide the theoretical basis for developing high ammonia nitrogen-tolerant germplasm with aquaculture potential.