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A new oxidative stress model, 2,2-azobis(2-amidinopropane) dihydrochloride induces cardiovascular damages in chicken embryo.

He RR, Li Y, Li XD, Yi RN, Wang XY, Tsoi B, Lee KK, Abe K, Yang X, Kurihara H - PLoS ONE (2013)

Bottom Line: At this concentration, AAPH was found to significantly reduce the density of blood vessel plexus that was developed in the chorioallantoic membrane (CAM) of HH 35 chick embryos.These results implied that AAPH-induced oxidative stress could impair the whole developmental processes associated with vasculogenesis and angiogenesis.In conclusion, oxidative stress, induced by AAPH, could lead to damage of the cardiovascular system in the developing chick embryo.

View Article: PubMed Central - PubMed

Affiliation: Pharmacy College, Jinan University, Guangzhou, People's Republic of China.

ABSTRACT
It is now well established that the developing embryo is very sensitive to oxidative stress, which is a contributing factor to pregnancy-related disorders. However, little is known about the effects of reactive oxygen species (ROS) on the embryonic cardiovascular system due to a lack of appropriate ROS control method in the placenta. In this study, a small molecule called 2,2-azobis(2-amidinopropane) dihydrochloride (AAPH), a free radicals generator, was used to study the effects of oxidative stress on the cardiovascular system during chick embryo development. When nine-day-old (stage HH 35) chick embryos were treated with different concentrations of AAPH inside the air chamber, it was established that the LD50 value for AAPH was 10 µmol/egg. At this concentration, AAPH was found to significantly reduce the density of blood vessel plexus that was developed in the chorioallantoic membrane (CAM) of HH 35 chick embryos. Impacts of AAPH on younger embryos were also examined and discovered that it inhibited the development of vascular plexus on yolk sac in HH 18 embryos. AAPH also dramatically repressed the development of blood islands in HH 3+ embryos. These results implied that AAPH-induced oxidative stress could impair the whole developmental processes associated with vasculogenesis and angiogenesis. Furthermore, we observed heart enlargement in the HH 40 embryo following AAPH treatment, where the left ventricle and interventricular septum were found to be thickened in a dose-dependent manner due to myocardiac cell hypertrophy. In conclusion, oxidative stress, induced by AAPH, could lead to damage of the cardiovascular system in the developing chick embryo. The current study also provided a new developmental model, as an alternative for animal and cell models, for testing small molecules and drugs that have anti-oxidative activities.

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Effects of AAPH on yolk-sac blood vessels of chick embryo.(A) Representative appearance of yolk-sac blood vessels of HH 18 embryo treated with saline, (B) 4 µmol AAPH and (C) 5 µmol AAPH for 12 hours. The silastic rings in A–C had inside diameter at 9 mm and outside diameter at 11 mm. The yolk-sac blood vessel images were taken by a stereomicroscope (Olympus MVX10 with OPTPRO 2007 image acquisition system) with resolution ratio at 1024×768 (Scale bar = 1 mm). (D) Statistical chart showing the yolk-sac blood vessel density from AAPH treated and untreated embryos. The results represent the mean ± S.D (n = 10). Statistical significances were determined using SPSS13.5 software, **p<0.01 compared with control group.
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pone-0057732-g003: Effects of AAPH on yolk-sac blood vessels of chick embryo.(A) Representative appearance of yolk-sac blood vessels of HH 18 embryo treated with saline, (B) 4 µmol AAPH and (C) 5 µmol AAPH for 12 hours. The silastic rings in A–C had inside diameter at 9 mm and outside diameter at 11 mm. The yolk-sac blood vessel images were taken by a stereomicroscope (Olympus MVX10 with OPTPRO 2007 image acquisition system) with resolution ratio at 1024×768 (Scale bar = 1 mm). (D) Statistical chart showing the yolk-sac blood vessel density from AAPH treated and untreated embryos. The results represent the mean ± S.D (n = 10). Statistical significances were determined using SPSS13.5 software, **p<0.01 compared with control group.

Mentions: The effect of AAPH-induced oxidation was also investigated in earlier stage chick embryos. Stage HH 18 chick embryos were mounted with silastic rings and treated with 4 or 5 µmol of AAPH for 12 hours. The effects of AAPH on yolk-sac blood vessel were then examined. The results revealed that AAPH exposure significantly inhibited the development of newly formed vascular plexus (Figures 3B and C) compared with the control (Figure 3A). There were fewer vessels (Figure 3B) and shrinkages in the leading edge of the vascular plexus (Figure 3C). The yolk-sac blood vessel density was also significantly decreased after AAPH treatment (Figure 3D). These results implied that AAPH inhibits angiogenesis in yolk sac of chick embryo.


A new oxidative stress model, 2,2-azobis(2-amidinopropane) dihydrochloride induces cardiovascular damages in chicken embryo.

He RR, Li Y, Li XD, Yi RN, Wang XY, Tsoi B, Lee KK, Abe K, Yang X, Kurihara H - PLoS ONE (2013)

Effects of AAPH on yolk-sac blood vessels of chick embryo.(A) Representative appearance of yolk-sac blood vessels of HH 18 embryo treated with saline, (B) 4 µmol AAPH and (C) 5 µmol AAPH for 12 hours. The silastic rings in A–C had inside diameter at 9 mm and outside diameter at 11 mm. The yolk-sac blood vessel images were taken by a stereomicroscope (Olympus MVX10 with OPTPRO 2007 image acquisition system) with resolution ratio at 1024×768 (Scale bar = 1 mm). (D) Statistical chart showing the yolk-sac blood vessel density from AAPH treated and untreated embryos. The results represent the mean ± S.D (n = 10). Statistical significances were determined using SPSS13.5 software, **p<0.01 compared with control group.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585800&req=5

pone-0057732-g003: Effects of AAPH on yolk-sac blood vessels of chick embryo.(A) Representative appearance of yolk-sac blood vessels of HH 18 embryo treated with saline, (B) 4 µmol AAPH and (C) 5 µmol AAPH for 12 hours. The silastic rings in A–C had inside diameter at 9 mm and outside diameter at 11 mm. The yolk-sac blood vessel images were taken by a stereomicroscope (Olympus MVX10 with OPTPRO 2007 image acquisition system) with resolution ratio at 1024×768 (Scale bar = 1 mm). (D) Statistical chart showing the yolk-sac blood vessel density from AAPH treated and untreated embryos. The results represent the mean ± S.D (n = 10). Statistical significances were determined using SPSS13.5 software, **p<0.01 compared with control group.
Mentions: The effect of AAPH-induced oxidation was also investigated in earlier stage chick embryos. Stage HH 18 chick embryos were mounted with silastic rings and treated with 4 or 5 µmol of AAPH for 12 hours. The effects of AAPH on yolk-sac blood vessel were then examined. The results revealed that AAPH exposure significantly inhibited the development of newly formed vascular plexus (Figures 3B and C) compared with the control (Figure 3A). There were fewer vessels (Figure 3B) and shrinkages in the leading edge of the vascular plexus (Figure 3C). The yolk-sac blood vessel density was also significantly decreased after AAPH treatment (Figure 3D). These results implied that AAPH inhibits angiogenesis in yolk sac of chick embryo.

Bottom Line: At this concentration, AAPH was found to significantly reduce the density of blood vessel plexus that was developed in the chorioallantoic membrane (CAM) of HH 35 chick embryos.These results implied that AAPH-induced oxidative stress could impair the whole developmental processes associated with vasculogenesis and angiogenesis.In conclusion, oxidative stress, induced by AAPH, could lead to damage of the cardiovascular system in the developing chick embryo.

View Article: PubMed Central - PubMed

Affiliation: Pharmacy College, Jinan University, Guangzhou, People's Republic of China.

ABSTRACT
It is now well established that the developing embryo is very sensitive to oxidative stress, which is a contributing factor to pregnancy-related disorders. However, little is known about the effects of reactive oxygen species (ROS) on the embryonic cardiovascular system due to a lack of appropriate ROS control method in the placenta. In this study, a small molecule called 2,2-azobis(2-amidinopropane) dihydrochloride (AAPH), a free radicals generator, was used to study the effects of oxidative stress on the cardiovascular system during chick embryo development. When nine-day-old (stage HH 35) chick embryos were treated with different concentrations of AAPH inside the air chamber, it was established that the LD50 value for AAPH was 10 µmol/egg. At this concentration, AAPH was found to significantly reduce the density of blood vessel plexus that was developed in the chorioallantoic membrane (CAM) of HH 35 chick embryos. Impacts of AAPH on younger embryos were also examined and discovered that it inhibited the development of vascular plexus on yolk sac in HH 18 embryos. AAPH also dramatically repressed the development of blood islands in HH 3+ embryos. These results implied that AAPH-induced oxidative stress could impair the whole developmental processes associated with vasculogenesis and angiogenesis. Furthermore, we observed heart enlargement in the HH 40 embryo following AAPH treatment, where the left ventricle and interventricular septum were found to be thickened in a dose-dependent manner due to myocardiac cell hypertrophy. In conclusion, oxidative stress, induced by AAPH, could lead to damage of the cardiovascular system in the developing chick embryo. The current study also provided a new developmental model, as an alternative for animal and cell models, for testing small molecules and drugs that have anti-oxidative activities.

Show MeSH
Related in: MedlinePlus