Comparative proteome analysis of the embryo proper and yolk sac membrane of day 11.5 cultured rat embryos

Usami, Makoto; Mitsunaga, Katsuyoshi; Nakazawa, Ken

doi:10.1002/bdrb.20127

Cited by 7 publications

(8 citation statements)

References 33 publications

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“…Protein of the embryo proper and yolk sac membrane (50 µg/gel) was analyzed by 2‐DE as described previously (Usami et al, 2007a,b). Immobilized pH gradient (IPG) strips (Immobiline DryStrip pH 3–10 NL, 13 cm, Amersham Biosciences, Piscataway, NJ) were used in isoelectric focusing (IEF) for the first dimension, and 12.5% polyacrylamide gels (14 × 16 cm) were used in sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) for the second dimension.…”

Section: Methodsmentioning

confidence: 99%

“…Identification of protein spots by mass spectrometry was carried out as described previously (Usami et al, 2007a). 2‐DE gels were stained with mass‐analysis compatible ProteoSilver staining kit (Sigma, St. Louis, MO) and protein spots were excised into column‐shape pieces with a diameter of about 1 mm.…”

Section: Methodsmentioning

confidence: 99%

“…With this method, more than 800 and 1,000 protein spots were detected through the replicate 2‐DE gels each from one embryo in the embryo proper and yolk sac membrane, respectively. In addition, from protein spots specific to the embryo proper and yolk sac membrane, proteins involved in tissue characteristic functions, such as morphogenesis and nutritional transfer, were identified by mass spectrometric analysis (Usami et al, 2007a).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Proteomic analysis of selenium embryotoxicity in cultured postimplantation rat embryos

Usami

Mitsunaga

Nakazawa

et al. 2008

Birth Defects Research Pt B

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The identified proteins with quantitative changes by selenate or selenite were considered to be candidate proteins involved in Se embryotoxicity: the actin-binding proteins for selenate embryotoxicity, proteins with the similar changes for the common Se embryotoxicity and antioxidant proteins for modification of Se embryotoxicity by redox-related treatments. These proteins may also be used as biomarkers in developmental toxicity studies.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Proteomic analysis of selenium embryotoxicity in cultured postimplantation rat embryos

Usami

Mitsunaga

Nakazawa

et al. 2008

Birth Defects Research Pt B

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“…The binding of C3 to rat embryos explanted at day 9.5 or 10.5 corresponding to the cultured embryos was the binding to the visceral yolk sac, C3 seems to be degraded and not to be recycled, since many C3 fragments, but not unfragmented C3, are detected in the visceral yolk sac by proteome analysis (Usami et al, 2007).…”

Section: Binding Of C3 To Rat Embryosmentioning

confidence: 98%

Complement component C3 functions as an embryotrophic factor in early postimplantation rat embryos

Usami

Mitsunaga²,

Miyajima³

et al. 2010

Int. J. Dev. Biol.

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A presumed embryotrophic factor for early postimplantation rat embryos, partially purified from rat serum, was identified as complement component C3 (C3), the central component of the complement system, by sequence analysis of its N-terminal. Purified rat C3 showed embryotrophic activity for rat embryos cultured from day 9.5 of gestation for 48 h in the culture medium composed of rabbit serum. The maximum embryotrophic activity of C3 was observed around 0.5 mg/ml, a level which is lower than rat serum C3 levels. In the culture medium composed of rat serum, cultured rat embryos selectively consumed C3, and C3-depletion by cobra venom factor affected embryonic growth. Inactivation of the internal thiolester bond of C3, the critical functional site for its activity in the complement system, by methylamine had no effects on its embryotrophic activity. Purified rabbit C3 had only weak embryotrophic activity for cultured rat embryos, suggesting species specificity of the embryotrophic activity of C3. Immunochemical analyses showed the specific presence of C3 on the visceral yolk sac, but not on the embryo proper of day 9.5 or 10.5 rat embryos both in utero and in vitro. In analysis using fluorescein-labeled rat C3, unfragmented C3s bound to the visceral yolk sac stronger than C3b, the primary active fragment of C3 in the complement system. These results indicate that C3, which has always been considered to be detrimental to embryos, functions as an embryotrophic factor by novel mechanisms probably through the visceral yolk sac. The present study thus provides new insights into functions of C3 and postimplantation embryonic growth.

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“…The gender of rat embryos can be determined by polymerase chain reaction (PCR) of a sequence in Sry, a et al, 1998), in DNA samples from the embryonic tissues, i.e., the embryo proper or yolk sac membrane. There are, however, no reported methods applicable to comprehensive proteomic analyses of early postimplantation rat embryos, where both embryo proper and yolk sac membrane are Sexing of postimplantation rat embryos in stored twodimensional electrophoresis (2-DE) samples by polymerase chain reaction (PCR) of an Sry sequencees (Usami et al, 2007a(Usami et al, , 2008.…”

Section: Introductionmentioning

confidence: 99%

Sexing of postimplantation rat embryos in stored twodimensional electrophoresis (2-DE) samples by polymerase chain reaction (PCR) of an Sry sequence

et al. 2009

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-Proteomic analysis of developmental toxicity by two-dimensional electrophoresis (2-DE) may detect gender-related toxic effects in embryos without visible gender characteristics. In the present study, we explored sexing of rat embryo stored in frozen 2-DE samples by polymerase chain reacSry). The embryo proper and yolk sac membrane at gestation day 11 from Wistar rats were used for stored embryonic 2-DE samples. The embryonic 2-DE samples were desalted and their total DNA was extracted. The Sry sequence in the embryos with the PCR product of Sry were determined as male, and those without the product were determined as female. It was concluded that stored embryonic 2-DE samples could be used for retrospective examination of gender-related effects in proteomic analysis of developmental toxicity.

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Comparative proteome analysis of the embryo proper and yolk sac membrane of day 11.5 cultured rat embryos

Abstract: Proteomic analysis of cultured postimplantation rat embryos will be a new approach in developmental biology and toxicology at the protein level.

Cited by 7 publications

References 33 publications

Proteomic analysis of selenium embryotoxicity in cultured postimplantation rat embryos

Proteomic analysis of selenium embryotoxicity in cultured postimplantation rat embryos

Complement component C3 functions as an embryotrophic factor in early postimplantation rat embryos

Sexing of postimplantation rat embryos in stored twodimensional electrophoresis (2-DE) samples by polymerase chain reaction (PCR) of an Sry sequence

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