The Influence Of Embryo And Bone Development Treated With Norfloxacin In Rare Minnow Gobiocypris Rarus Offspring

Fluoroquinolones antibiotics(FQs)have become a widely used antibacterial drug in clinic nowadays,due to its good pharmacokinetic properties and therapeutic effects.However,there are many adverse reactions in clinic,such as neurotoxicity,cardiac toxicity,phototoxicity,chondrotoxicity and so on.The specificity of chondrotoxicity limits the use of FQs in pediatric patients,so more attentions have been paid to chondrotoxicity of FQs.In recent years,animal studies have shown that long-term administration of FQs can cause cartilage damage in young dogs,mice,rabbits,pigs and other animals.At present,there is no report on whether long-term FQs treatment can cause skeletal dysplasia in fish,and there is no study on whether FQs treatment in animal parents causes cartilage toxicity of offspring.Therefore,the purpose of this study is to explore the effects of FQs on bone development and molecular mechanism of rare minnow.The study is divided into three parts: First,the female rare minnow of four months old was exposed to 0.1,10 ?g/L NOR solution for three months and then propagated with the normal male to produce offspring,to explore the effects of maternal long-term exposure on embryonic development and skeletal development of the larvae.Second,the newly fertilized eggs were exposed to 0.1,10 ?g/L NOR solution for 72 hours to explore the effects of short-term embryo exposure on embryonic development and skeletal development of the larvae.Third,after 72 hours fertilization,the hatched larvae began to be exposed to 0.1 ?g/L NOR to explore the effects of direct exposure to larvae on bone development.This experiment has tested statistical analysis of embryo fertilization rate,mortality rate,hatching rate,deformity rate,heart rate,body length and autonomic movement,and stain cartilage,detection calcium content,detection lysine oxidase(LOX)activity and qRT-PCR detection larvae cartilage development related genes expression of bmp 2,bmp 4,bmp 6,sox9 a,runx2,col2?1 and lox1.The main results were as follows:1.Compared with the control group,the number of spawning parent fish decreased,the rate of embryo fertilization and hatching decreased,the rate of deformity and mortality increased,the heart rate and the frequency of spontaneous movement of tail slowed down,and the larval body length was shorter.These results suggest that NOR exposure may affect the reproductive ability of the parentage and inhibit the development of embryo and nervous system of Carassius auratus.The length of the hyoid,mandible and mandibular arch were shortened,the width of mandible was enlarged,and the whole mandible expanded to both sides.The results showed that NOR exposure significantly inhibited the cartilage development of the offspring.The overall calcium content of larvae decreased significantly,suggested that NOR exposure may inhibit cartilage development by reducing the calcium level of larvae.The expression of bmp2,bmp4 and sox9 a genes regulated the proliferation and differentiation of chondrocytesand and the related genes bmp6,lox1 and runx2 expanded osteogenesis in larvae decreased significantly.The expression of col2?1 gene encoding collagen,a key component of cartilage,and the expression of lox1,an important gene in the process of cartilage cross-linking,were also significantly inhibited.2.Compared with the control group,after the fertilized eggs of rare minnow was treated with 0.1 ?g/L and 10 ?g/L NOR,the incubation period was delayed.It is concluded that direct NOR exposure to fertilized eggs will affect the embryonic development of the rare minnow.The overall calcium content of larvae decreased significantly.The expression levels of bmp2,bmp4,bmp6,runx2,col2?1,lox1 and sox9 a were significantly decreased.3.After the larvae of rare minnow was treated with 0.1 ?g/L NOR,the length of the hyoid,mandible and mandibular arch were shortened,the width of mandible was enlarged,and the whole mandible expanded to both sides.Lysine oxidase activity decreased and the expression levels of bmp2,bmp4,bmp6,runx2,col2?1,lox1 and sox9 a were significantly decreased.The results indicated that NOR treatment significantly inhibited the skeletal development of rare minnow larvae.In this study,the inhibitory effects of NOR treatment on embryonic and skeletal development of the rare minnow were studied from three parts: the female parent,the fertilized eggs and the larvae.The results showed that NOR had obvious inhibitory effect on embryo development of rare minnow and provide a theoretical basis for the application of fluoroquinolones.