Skeletal Softening in Cyprinus carpio rubrofuscus: Insights from Mineral Metabolism, Histology, and Autophagy

C. c. rubrofuscus is an economically important species in South China. During breeding, some individuals develop skeletal softening, but the underlying mechanisms remain unclear. In this study, five-month-old C. c. rubrofuscus were classified into a hard-bone group and a softened-bone group based on X-ray radiography. A systematic comparison was then conducted between the two groups, including analyses of external morphology, skeletal morphology, vertebral bone mineral density (BMD), calcium (Ca) and phosphorus (P) levels in serum and bone tissue, bone histology, and the expression of autophagy-related genes and proteins. The prevalence of the softened-bone phenotype was approximately 14% in the cultured population. Compared with the hard-bone group, the softened-bone group showed significantly lower vertebral BMD, significantly increased serum Ca levels, significantly decreased serum P levels, and markedly reduced Ca and P contents in bone tissue. In addition, varying degrees of deformity were observed in the ribs, caudal intermuscular spines, and urostyle. Histological examination further revealed severe skeletal malformations in the softened-bone group, characterized by irregular cortical bone thickness in the ribs and pterygiophores, together with a significantly reduced osteocyte density. Meanwhile, microtubule-associated protein 1 light chain 3 (LC3) levels were significantly increased, whereas sequestosome 1 (p62) levels were significantly decreased. The upregulated expression of autophagy-related genes suggested dysregulated autophagy, which may contribute to osteocyte loss. Collectively, these results indicate that the softened-bone phenotype in C. c. rubrofuscus is not associated with obvious changes in external body morphology. However, disrupted Ca-P homeostasis, together with altered autophagy, may impair osteocyte viability and bone mineralization, ultimately leading to bone softening and skeletal deformity. These findings provide a theoretical basis for further investigation of the mechanisms underlying bone softening and deformity in C. c. rubrofuscus and for the genetic improvement and selective breeding of cyprinid fish to reduce the occurrence of the softened-bone trait.