Effect of different vitrification protocols on post thaw viability and gene expression of ovine preantral follicles

Gupta, P. S. P.; Kaushik, Kalpana; Johnson, Patricia A.; Krishna, K. M.; Nandi, S.; Mondal, Sukanta; Tej, J. Nikhil Kumar; Somoskői, Bence; Cseh, Sándor

doi:10.1016/j.theriogenology.2021.10.024

Cited by 8 publications

(12 citation statements)

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“…APAF1 then triggers Caspase-9 and Caspase-3 and leads to apoptosis of cell [ 57 ]. This pathway is mainly regulated by apoptotic (Bax) and anti-apoptotic (BCL-2) genes [ 58 , 59 , 60 ]. mRNA expression levels of Bax and BCL-2 were quantified in the present study to check the impact of cryopreservation on apoptosis.…”

Section: Discussionmentioning

confidence: 99%

Antioxidant Activity and Oxidative Stress-Oriented Apoptosis Pathway in Saccharides Supplemented Cryopreserved Sperm of Pacific Abalone, Haliotis discus hannai

et al. 2022

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The Pacific abalone Haliotis discus hannai is a highly commercialized seafood in Southeast Asia. The aim of the present study was to determine the antioxidant activity and oxidative stress-oriented apoptosis pathway in saccharides supplemented cryopreserved sperm of Pacific abalone. Cryopreserved sperm showed impaired antioxidant defenses due to the reduced mRNA abundance of antioxidant genes (CAT, Cu/Zn-SOD, Mn-SOD, GPx, GR, and BCL-2), apoptosis inhibitor (HSP70, and HSP90) gene, and enzymatic antioxidant activity compared to fresh sperm. Such impaired antioxidant defenses caused an increase in the mRNA expression of apoptosis genes (Bax, and Caspase-3), finally leading to apoptosis. The impaired antioxidant defense also increased O2•− production and lipid peroxidation (MDA) levels, which further accelerated apoptosis. Considering all the experimental findings, an apoptosis pathway of cryopreserved sperm has been adopted for the first time. Specifically, sperm cryopreserved using 3% sucrose combined with 8% dimethyl sulfoxide (DMSO) showed improved mRNA stability, enzymatic activity, and DNA integrity with reduced O2•− production and MDA levels compared to sperm cryopreserved with the other types of examined cryoprotectants (8% ethylene glycol + 1% glucose, 6% propylene glycol + 2% glucose, 2% glycerol + 3% glucose, and 2% methanol + 4% trehalose). The present study suggests that 3% sucrose combined with 8% DMSO is suitable to cryopreserve the sperm of this valuable species for molecular conservation.

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Section: Discussionmentioning

confidence: 99%

Antioxidant Activity and Oxidative Stress-Oriented Apoptosis Pathway in Saccharides Supplemented Cryopreserved Sperm of Pacific Abalone, Haliotis discus hannai

et al. 2022

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“…The in vitro culture of preantral follicles was adapted from an earlier study (Gupta et al, 2022). The preantral follicles was incubated in a CO 2 incubator at 38 °C, 5% CO 2 , and 95% RH for 6 or 12 days.…”

Section: In Vitro Culture Of Secondary Folliclesmentioning

confidence: 99%

“…Quantitative PCR was performed using 10 µL reactions mixture containing 5.0 µL of iTaq™ Universal 2x SYBR Green Supermix (Biorad), 0.5 μL (0.5 pmol each) primer mix (sense and antisense primers), 1.5 µL of cDNA and 2.5 µL of nuclease free water. Ribosomal protein S18 (RPS18) was used as the internal reference (Gupta et al, 2022), and the order-specific primer SOD1 and SOD2 were designed by a NCBI website and Primer 3 software. The sequences of primer, product length and GenBank accession numbers are mentioned in Table 1.…”

Section: Rna Isolation and Quantitative Real-time Pcr Analysis (Qpcr)mentioning

confidence: 99%

“…Procedure of Gupta et al (2022 ) was followed for Isolation of preantral follicle (200-300 um). After isolation, preantral follicles with a visible oocyte, surrounded by at least two granulosa cell layers, an intact basement membrane and with no antral cavity were selected for this study (Ferreira et al , 2018).…”

Section: Isolation and Selection Of Preantral Folliclesmentioning

confidence: 99%

“…The vitrification of preantral follicles was performed using the protocol as described by Gupta et al (2022). A total of 480 preantral follicles were selected for the study and distributed into four groups: (1) Fresh preantral follicles (n =121), (2) Vitrified preantral follicles (n=118) without any supplementation of α-tocopherol, (3) Vitrified follicles (n=122) with 10 mM of alpha tocopherol supplementation and (4) Vitrified follicles (n=119) with 20 mM of α-tocopherol supplementation.…”

Section: Vitrification and Thawing Proceduresmentioning

confidence: 99%

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Effect of α-Tocopherol on Viability, Lipid Peroxidation and Oxidative Stress of Cryopreserved Ovine Preantral Follicle

Kaushik

Pulukuri

Tej

et al. 2022

ijvsbt

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Vitrification of preantral follicles is a promising technique to preserve female fertility. The aim of the present study was to evaluate the effect of supplementation of α-tocopherol in the vitrification solution on the viability, lipid peroxidation and mRNA expression of superoxide dismutases (SOD1 and SOD2) in vitrified cultured ovine preantral follicles at day-6 and day-12. Preantral follicles (200-300 µm) were isolated from the ovine ovaries by the mechanical method and were distributed separately to the vitrification medium supplemented with 10 mM and 20 mM of α-tocopherol. After a week, the follicles were thawed and analyzed for follicular viability by trypan blue dye exclusion method and subjected to in vitro culture (IVC) for 6 and 12 days. Our results revealed that the significant increase (p<0.05) of viability in 20 mM α-tocopherolsupplemented vitrified group when compared to the vitrified without α-tocopherol group. On day-6 of IVC, malondialdehyde (MDA) concentration was significantly(p<0.05) higher in vitrified group without α-tocopherol in comparison to vitrified supplemented with 20 mM of α-tocopherol and control fresh groups. However, no significant difference in MDA content was found among the groups at day-12. The mRNA expression level of SOD1 at day-6 was significantly (p<0.05) higher in vitrified with 20 mM of α-tocopherol and control fresh groups compared to the vitrified without α-tocopherol and with10 mM α-tocopherol groups. The expression pattern of SOD2 was significantly (p<0.05) higher in control fresh group compared to the other groups at day-6 and day-12 of IVC.We conclude that lowering the vitrification-induced lipid peroxidation of preantral follicles by α-tocopherol at 20 mM concentration may be mediated by increasing SOD expression during IVC.

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Effects of different cryopreservation methods on canine isolated preantral follicles

Somoskői,

Bordás,

Uno

et al. 2023

Animal Reproduction Science

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Effect of different vitrification protocols on post thaw viability and gene expression of ovine preantral follicles

Cited by 8 publications

References 58 publications

Antioxidant Activity and Oxidative Stress-Oriented Apoptosis Pathway in Saccharides Supplemented Cryopreserved Sperm of Pacific Abalone, Haliotis discus hannai

Antioxidant Activity and Oxidative Stress-Oriented Apoptosis Pathway in Saccharides Supplemented Cryopreserved Sperm of Pacific Abalone, Haliotis discus hannai

Effect of α-Tocopherol on Viability, Lipid Peroxidation and Oxidative Stress of Cryopreserved Ovine Preantral Follicle

Effects of different cryopreservation methods on canine isolated preantral follicles

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