Coagulansin-A has beneficial effects on the development of bovine embryos invitro via HSP70 induction.

Imran Khan,Il-Keun Kong,Ihsan-Ul-Haq Ihsan-Ul-Haq,Chang Guo Yan,Kyeong-Lim Lee,Bushra Mirza,Md Fakruzzaman,Seok-Hwan Song

doi:10.1042/bsr20150222

Imran Khan, Il-Keun Kong + Show 6 more

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https://doi.org/10.1042/bsr20150222

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Abstract

Coagulansin-A (withanolide) is the steroidal lactone obtained from Withania coagulans which belong to Solanaceae family. The present study investigated the effects of coagulansin-A on bovine oocyte maturation and embryo development invitro. All these oocytes were aspirated from the ovaries obtained from Korean Hanwoo cows at a local abattoir. To determine whether coagulansin-A has beneficial effects on bovine oocyte maturation invitro, 355 oocytes per group (control and treated) in seven replicates were subjected with different concentrations (1, 2.5, 5, 7.5 and 10μM) of coagulansin-A. The coagulansin-A was added in the invitro maturation (IVM) media followed by invitro fertilization (IVF) and then invitro culture (IVC). Only treatment with 5μM coagulansin-A remarkably (P<0.05) improved embryos development (Day 8 blastocyst) having 27.30 and 40.01% for control and coagulansin-A treated groups respectively. Treatment with 5μM coagulansin-A significantly induced activation of heat shock protein 70 (HSP70) (P<0.05). Immunofluorescence analysis revealed that 5μM coagulansin-A treatment also significantly inhibited oxidative stress and inflammation during bovine embryo development invitro by decreasing 8-oxoguanosine (8-OxoG) (P<0.05) and nuclear factor-κB (NF-κB) (P<0.05). The expressions of HSP70 and NF-κB were also conformed through real-time PCR (RT-PCR). Additionally, the terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) assay confirmed that coagulansin-A treatment significantly improved the embryo quality and reduced bovine embryo DNA damage (P<0.05). The present study provides new information regarding the mechanisms by which coagulansin-A promotes bovine embryo development invitro.

Highlights

In assisted reproductive technology (ART), in vitro maturation (IVM) is an extremely important step, followed by fertilization and embryo culture
Cleavage and developmental rates of embryos generated from oocytes treated with various concentrations of coagulansin-A The percentage of cleaved embryos was determined on Day 3 of culture
This indicated that integral optical density (IOD) of heat shock protein 70 (HSP70) was significantly greater (P < 0.05) in the 5 μM coagulansin-A group than in control group (Figure 4)

Summary

Introduction

In assisted reproductive technology (ART), in vitro maturation (IVM) is an extremely important step, followed by fertilization and embryo culture. The increased O2 concentration leads to the production of large amounts of reactive oxygen species (ROS), which have detrimental effects on developing embryos [3]. These effects include DNA and protein damage and activation of various signalling pathways (P53 and P38), leading to apoptosis [4]. There are two Abbreviations: ART, assisted reproductive technology; COC, cumulus–oocyte complex; COX, cyclooxygenase; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HSP70, heat shock protein 70; IOD, integral optical density; IVC, in vitro culture; IVF, in vitro fertilization; IVM, in vitro maturation; NF-κB, nuclear factor-κB; 8-OxoG, 8-oxoguanosine; PVP, polyvinylpyrrolidone; ROS, reactive oxygen species; RT-PCR, real-time PCR; TUNEL, terminal deoxynucleotidyl transferase dUTP nick-end labelling.

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