Effect of in ovo zinc injection on the embryonic development, tissue zinc contents, antioxidation, and related gene expressions of broiler breeder eggs

Two experiments were conducted to investigate the effect of in ovo zinc （Zn） injection on the embryonic development, tissue Zn contents, antioxidation and related gene expressions of fertilized eggs of Arbor Acres broiler breeders. Experiment 1 was conducted to determine an optimal embryonic age for early in ovo injection. A total of 720 fertilized eggs with similar weights were randomly allotted to 4 treatments with 6 replicates per treatment and 30 eggs per replicate in a completely randomized design. The eggs were injected with 0.1 mL sterilized water at 3, 6 and 9 embryonic days of incubation （E3, E6 and E9） or non-injection （the control）, respectively. The results from experiment 1 showed that E3 and E6 injections increased （P〈0.05） the embryonic mortalities, and decreased （P〈0.05） hatchabilities compared to the non-injected control, but no differences （P〉0.05） between E9 injection and the non-injected control were observed in either embryonic mortality or hatchability. The findings suggest that the E9 is the optimal embryonic age for early in ovo injection. In experiment 2, a total of 672 fertilized eggs with similar weights were randomly allocated to 7 treatments with 6 replicates per treatment and 16 eggs per replicate in a completely randomized design. The eggs were injected with 0 （the negative control）, 50, 100, 150, 200, or 250 μg Zn/egg as reagent grade ZnSO4.7H20 in a 0.1-mL solution, or non-injection （the positive control）, respectively at E9-10. The results from the experiment 2 demonstrated that no differences （P〉0.05） among 50 and 100 μg Zn/egg groups and the negative control were observed in the embryonic mortality and hatchability, however, the injection of 200 μg Zn/egg increased （P〈0.05） the embryonic mortality, and injections of 150 and 200 μg Zn/egg decreased （P〈0.05） hatchabilities compared with the controls. The embryonic tibia Zn contents at E20 were increased （P〈0.05） by injections of 150, 200 and 250μg Zn/egg. Zinc injection did not affect （P〉0.05） malonaldehyde （MDA） contents, copper- and Zn- containing superoxide dismutase （CuZnSOD） activities and mRNA expression levels in the liver and heart of chick embryos at E15 and E20. Compared with the negative control, injections of 50, 150 and 200 μg Zn/egg up-regulated （P〈0.05） themetallothionein （MT） mRNA expression levels in the embryonic liver at E20. These results indicated that in ovo Zn injections increased Zn contents in the embryonic tibia and MT mRNA expression levels in the embryonic liver at E20, however, injections of 150-200 μg Zn/egg were harmful to the embryonic development.

The in ovo injection of methionine improves intestinal cell proliferation and differentiation in chick embryos by activating the JAK2/STAT3 signaling pathway

The intestinal health of chick embryos is vital for their life-long growth,and exogenous nutrition intervention may provide sufficient nutrition for embryonic development.In the present study,we investigated the effect of in ovo injection of L-methionine(L-Met)on the intestinal structure and barrier function of chick embryos.There were 4 groups of treatments:the control(CON)group injected with phosphate-buffered saline(PBS)and the other 3 groups injected with 5,10,and 20 mg L-Met/egg,respectively.The injection was performed on embryonic day 9(E9),and intestinal samples were collected on the day of hatching for analysis.The results showed that,compared with the CON group,the groups administered an in ovo injection of L-Met increased relative weights of the duodenum,jejunum,and ileum(P<0.05).Hematoxylin and eosin(H&E)staining showed that the groups injected with 5,10,and 20 mg L-Met significantly increased villus height and crypt depth(P<0.05).Moreover,in ovo injection of 10 mg L-Met also increased the transepithelial electrical resistance(TEER)of the jejunum(P<0.05).Injection with 10 and 20 mg L-Met increased the expression of the tight junction proteins(ZO-1 and claudin-1)and the fluorescence signal intensity of Ki67 and villin proteins(P<0.05).Further,the protein expression of phospho-Janus kinase 2(p-JAK2)and phospho-signal transducer and activator of transcription 3(p-STAT3)was significantly increased by 10 or 20 mg L-Met injection(P<0.05).In conclusion,the injection of L-Met,especially at a dose of 10 mg,showed beneficial effects on the intestinal integrity of chick embryos due to the activation of the JAK2/STAT3 signaling pathway.Our results may provide new insights for regulating the intestinal development of embryonic chicks and the rapid growth of chicks after hatching.

Methionine sources and genotype affect embryonic intestinal development,antioxidants,tight junctions,and growth-related gene expression in chickens

Methionine(Met)is an essential and first limiting amino acid in the poultry diet that plays a significant role in chicken embryonic development and growth.The present study examined the effect of in ovo injection of DL-Met and L-Met sources and genotypes on chicken embryonic-intestinal development and health.Fertilized eggs of the two genotypes,TETRA-SL layer hybrid(TSL)—commercial layer hybrid and Hungarian Partridge colored hen breed(HPC)—a native genotype,were randomly distributed into four treatments for each genotype.The treatment groups include the following:1)control non-injected eggs(NoIn);2)saline-injected(SaIn);3)DL-Met injected(DLM);and 4)L-Met injected(LM).The in ovo injection was carried out on 17.5 d of embryonic development;after hatching,eight chicks per group were sacrificed,and the jejunum was extracted for analysis.The results showed that both DLM and LM groups had enhanced intestinal development as evidenced by increased villus width,villus height,and villus area(P<0.05)compared to the control.The DLM group had significantly reduced crypt depth,glutathione(GSH)content,glutathione S-transferase 3 alpha(GST3),occludin(OCLN)gene expression and increased villus height to crypt depth ratio in the TSL genotype than the LM group(P<0.05).The HPC genotype has overexpressed insulin-like growth factor 1(IGF1)gene,tricellulin(MD2),occludin(OCLN),superoxide dismutase 1(SOD1),and GST3 genes than the TSL genotype(P<0.05).In conclusion,these findings showed that in ovo injection of Met enhanced intestinal development,and function,with genotypes responding differently under normal conditions.Genotypes also influenced the expression of intestinal antioxidants,tight junction,and growth-related genes.