Activation or inhibition of NF-κB from the juvenile stage results in skewed sex ratios in common carp (Cyprinus carpio L.)

Due to sexual dimorphism in growth traits, all-female populations and populations with a high proportion of females hold significant economic value in common carp (Cyprinus carpio L.). However, the underlying mechanisms governing gonadal differentiation remain inadequately understood. NF-κB, as a crucial regulator of anti-apoptotic processes, plays a significant role in mammalian gonad formation and zebrafish oocyte maintenance. To investigate the potential roles of the NF-κB pathway in the gonad differentiation of common carp, juveniles with undifferentiated gonads were treated with an activator (DOC) or inhibitor (NAI) of NF-κB. Our findings revealed that juveniles exposed to DOC exhibited a significantly increased female-to-male ratio (from 0.97:1 to 4.30:1) and substantial increases in weight and body length. Conversely, inhibiting NF-κB with NAI resulted in a decreased female-to-male ratio (from 1.06:1 to 0.33:1). Furthermore, NAI also induced the occurrence of ovotestis. Additional in vivo and ex vivo experiments demonstrated that DOC activation of the NF-κB pathways led to inhibition of apoptosis, significant increases in serum 17β-estradiol (E2) levels and the E2/T ratio, and upregulation of the female-biased genes foxl2 and cyp19a1a. Simultaneously, it significantly suppressed serum 11-ketotestosterone (11-KT) levels and the expression of male-biased genes amh, dmrt1, and gsdf and induced VTG levels in the hepatopancreas of male carp. However, NAI exhibited contrasting effects. Both DOC and NAI disrupted the balance of the hypothalamus-pituitary-gonad (HPG) reproductive axis at multiple potential loci, including changes in the expression levels of steroid hormone synthesis-related genes in gonads. Transcriptomic analysis revealed comprehensive molecular changes in undifferentiated gonads during DOC-induced feminization and NAI-induced masculinization. The transcriptome results were consistent with the observed alterations in hormone content and the expression of genes involved in sex hormone synthesis, inflammation, and apoptosis following exposure to DOC or NAI. These findings suggest the involvement of NF-κB in the sex differentiation of common carp by regulating apoptosis and disrupting the HPG reproductive axis. Our study also explores a DOC-induced method of producing high ratios of females and discovers the underlying mechanism of NF-κB's contribution to sex differentiation in common carp.