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Polyadenylated tail length variation pattern in ultra-rapid vitrified bovine oocytes.

ABSTRACT: AIM:Thecurrent study aims at investigating the polyadenylated (poly[A]) tail length of morphologically high and low competent oocytes at different developmental stages. Furthermore, effect of ultra-rapid vitrification on the poly(A) tail length was studied. MATERIALS AND METHODS:Fresh bovine cumulus oocyte complexes from abattoir originated ovaries were graded based on morphological characters and matured in vitro. Cryopreservation was done by ultra-rapid vitrification method. mRNA was isolated from different categories of oocyte and subjected to ligation-mediated poly(A) test followed by polymerase chain reaction for determining the poly(A) tail length of ? actin, gap junction protein alpha 1 (GJA1), poly(A) polymerase alpha (PAPOLA), and heat shock 70 kDa protein (HSP70) transcripts. RESULTS:GJA1, PAPOLA, and HSP70 showed significantly higher poly(A) in immature oocytes of higher competence irrespective of vitrification effects as compared to mature oocytes of higher competence. CONCLUSION:mRNA poly(A) tail size increases in developmentally high competent immature bovine oocytes. There was limited effect of ultra-rapid vitrification of bovine oocytes on poly(A).

SUBMITTER: Dutta DJ

PROVIDER: S-EPMC5104714 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Polyadenylated tail length variation pattern in ultra-rapid vitrified bovine oocytes.

Dutta D J DJ Raj Himangshu H Dev And Hiramoni AH

Veterinary world 20161014 10

<h4>Aim</h4>Thecurrent study aims at investigating the polyadenylated (poly[A]) tail length of morphologically high and low competent oocytes at different developmental stages. Furthermore, effect of ultra-rapid vitrification on the poly(A) tail length was studied.<h4>Materials and methods</h4>Fresh bovine cumulus oocyte complexes from abattoir originated ovaries were graded based on morphological characters and matured <i>in vitro</i>. Cryopreservation was done by ultra-rapid vitrification meth ...[more]

PMID: 27847415

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Project description:BackgroundWe had previously demonstrated that vitrification reduces the levels of certain phospholipid classes, and that oocytes from aged mice show a similar lipidome alteration, even without vitrification. In the current investigation, we examined if vitrification-warming of mouse oocytes from young and aged mice causes any changes in molecular aspects of lipid-associated features.MethodsMetaphase II (MII) stage oocytes were harvested from young (10-14-week-old) and aged (45-54-week-old) mice by a superovulation regime with PMSG followed by hCG. We examined the status of the intracellular lipid pool and the integrity of the plasma membrane by staining oocytes with BODIPY 500/510 and CellMask live dyes. Expression of lipid uptake- and necroptosis-associated genes was assessed by quantitative PCR analyses, in oocytes from young and old mice, before and after vitrification. Localization patterns of two crucial necroptosis proteins, phosphorylated MLKL (pMLKL) and phosphorylated RIPK1 (pRIPK1) were examined in mouse oocytes by immunofluorescence staining. Necrostain-1 (Nec1), an inhibitor of RIPK1, was used to examine if RIPK1 activity is required to maintain oocyte quality during vitrification.ResultsWe confirmed that vitrified-warmed oocytes from aged mice showed noticeable decrease in both CellMask and BODIPY 500/510 dyes. Among the lipid uptake-associated genes, Cd36 expression was higher in oocytes from aged mice. Necroptosis is a type of programmed cell death that involves damage to the plasma membrane, eventually resulting in cell rupture. The expression of necroptosis-associated genes did not significantly differ among groups. We observed that localization patterns of pMLKL and pRIPK1 were unique in mouse oocytes, showing association with microtubule organizing centers (MTOCs) and spindle poles. pMLKL was also localized on kinetochores of MII chromosomes. Oocytes treated with Nec1 during vitrification showed a decreased survival rate, indicating the importance of RIPK1 activity in oocyte vitrification.ConclusionsWe report that oocytes from aged mice show differential expression of CD36, which suggests that CD36-mediated lipid uptake may be influenced by age. We also show for the first time that pMLKL and pRIPK1 exhibit unique localization pattern in mouse oocytes and this may suggest role(s) for these factors in non-necroptosis-associated cellular processes.

Dataset Information

Polyadenylated tail length variation pattern in ultra-rapid vitrified bovine oocytes.

Publications

Polyadenylated tail length variation pattern in ultra-rapid vitrified bovine oocytes.

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