Vitellogenin in Amur sturgeon (Acipenser schrenckii): induction, purification and changes during the reproductive cycle

Abstract

Summary The objective of the study was to assess the changes of vitellogenin (Vg) during the course of the reproductive cycle in Amur sturgeon (Acipenser schrenckii). Vg was purified from the serum of vitellogenic female Amur sturgeon by distilled water and gel filtration. Vg had an apparent molecular mass of 410 kDa and appeared as one band corresponding to 205 kDa after SDS-PAGE under reducing conditions. These bands were immunoreacted in Western blotting using antiserum against amur sturgeon lipovitellin (anti-Lv) which is an egg yolk protein derived from Vg. Lv was purified from egg extracts by ammonium sulfate solution and gel filtration. Vg was confirmed to be a lipoglycophosphoprotein by staining with Red oil, Molecular Proes’ Pro-Q® Emerald 300 Glycoprotein Gel stain and Pro-Q® Diamond Phosphoprotein Gel Stain. Vg induction was detected after injection of E2 at a concentration of at least 0.5 mg kg−1, and the plasma Vg concentration was increased by injection of the animals to 0.5 mg kg−1 for 5 day late. Immature sturgeons at 1015 g weight do not naturally synthesize Vg, but strongly responded to exogenously added E2. In addition, if they were not stimulated with exogenous E2, the youngest sturgeons did not show any detectable amount of Vg in their plasma, suggesting that Amur sturgeons with a body mass range of 903–1015 g could be used for the induction test, irrespective of their sex. The Enzyme-linked immuno-sorbant assay for sturgeon vg was developed to quantify serum Vg, using purified sturgeon Vg and anti-Vg. The measurable range was from 16.25 to 1000 ng ml−1. The dilution curve in ELISA of vitellogenic female serum was parallel to the standard curve of purified Vg. The coefficient variations of intra- and inter-assay were less than 5%, respectively. Vg levels varied throughout natural vitellogenesis from 0 μg ml−1 (1–3 years old) to approximately 200 μg ml−1 (7–8 years old). We observed an early transitory peak of serum Vg levels 200 μg ml−1 (for 10 months) at the time of early vitellogenesis and high Vg levels 218 μg ml−1 (for 3 months) in spring period before ovulation. It appears that the duration of vitellogenesis in Amur sturgeon is lasting more than 2 years.