A new role for RU486 (mifepristone): it protects sperm from premature capacitation during cryopreservation in buffalo

Dalal, Jasmer; Kumar, Pradeep; Chandolia, R.K.; Pawaria, Shikha; Rajendran, Rasika; Sheoran, Suman; Adnot, Jérôme; Kumar, Dharmendra

doi:10.1038/s41598-019-43038-4

Cited by 26 publications

(39 citation statements)

References 46 publications

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“…Intracellular calcium is an important modulator for capacitation (Darszon et al, 2006), but its concentration should not be high before reaching the female reproductive tract, otherwise it may induce capacitation-like change before fertilization. To confirm the capacitation-like changes induced by cryopreservation, we performed immunoblotting of protein tyrosine phosphorylation, which is an ultimate biomarker of capacitation (Dalal et al, 2019).…”

Section: Discussionmentioning

confidence: 99%

“…The ethanol extract of EY, LDL, and seminal plasma was prepared as described by Dalal et al (2019). Briefly, 1 ml of phosphate-buffered saline (PBS) was added to 1 ml of sample and homogenized for 5 min.…”

Section: Estimation Of Progesterone In Ey Ldl and Seminal Plasmamentioning

confidence: 99%

“…To assess the status of premature capacitation of sperm during the cryopreservation, the CTC assay was performed. The CTC assay was performed as described by Dalal et al (2019).…”

Section: Ctc Assaymentioning

confidence: 99%

“…analyzed as per the method described by Dalal et al (2019). The stock solution (1 mM) of Fluo-3 AM esters in dimethyl sulfoxide was prepared and stored at −20°C.…”

mentioning

confidence: 99%

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Low‐density lipoproteins protect sperm during cryopreservation in buffalo: Unraveling mechanism of action

Dalal

Chandolia

Pawaria

et al. 2020

Molecular Reproduction Devel

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This study was carried out to reveal factors and the mechanism of action by which lowdensity lipoproteins (LDLs) protect sperm better than egg yolk (EY) during cryopreservation. We extracted LDL from EY and compared the amount of calcium, progesterone, and antioxidants in EY and LDL. We found a very high concentration of progesterone (1423.95 vs. 10.46 ng/ml) and calcium (29.19 vs. 0.47 mM) in EY as compared with LDL. Antioxidant assays like DPPH (2,2-diphenyl-1-picrylhydrazyl) and the ferric reducing antioxidants power assay revealed that the LDL extender had almost double ability to lose hydrogen than the EY extender. For sperm cryopreservation, 20 ejaculates from four Murrah buffalo bulls were collected. Each ejaculate was divided into four aliquots and extended in 10%, 12%, and 14% LDL (w/v) and EY-based extenders, followed by cryopreservation. The LDL-based extender prevented excessive cholesterol efflux, and its high content of antioxidants minimized reactive oxygen species generated during cryopreservation, resulting in a functional CatSper channel. The EY-based extender promoted excess cholesterol efflux due to the presence of highdensity lipoprotein, resulting in a compromised CatSper channel. High intracellular calcium in a cryopreserved sperm in the EY group as compared with the LDL group indicates that progesterone present in EY activates the CatSper channel, resulting in a heavy calcium influx into the sperm. The greater tyrosine phosphorylation and increased number of F-pattern in the sperm cryopreserved in the EY extender indicate that high intracellular calcium triggers more capacitation-like changes in the sperm cryopreserved in EY than LDL extender. In conclusion, we demonstrated the new facts and understandings about LDL and EY for semen cryopreservation.

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Section: Discussionmentioning

confidence: 99%

Section: Estimation Of Progesterone In Ey Ldl and Seminal Plasmamentioning

confidence: 99%

“…To assess the status of premature capacitation of sperm during the cryopreservation, the CTC assay was performed. The CTC assay was performed as described by Dalal et al (2019).…”

Section: Ctc Assaymentioning

confidence: 99%

“…analyzed as per the method described by Dalal et al (2019). The stock solution (1 mM) of Fluo-3 AM esters in dimethyl sulfoxide was prepared and stored at −20°C.…”

mentioning

confidence: 99%

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Low‐density lipoproteins protect sperm during cryopreservation in buffalo: Unraveling mechanism of action

Dalal

Chandolia

Pawaria

et al. 2020

Molecular Reproduction Devel

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“…Intracellular calcium of spermatozoa was analyzed as per the method described earlier(Dalal et al, 2019) with slight modifications. Stock solution (1 mM) of Fluo-3 AM esters (#73881; Sigma-Aldrich) in DMSO was prepared and stored at −20°C.…”

mentioning

confidence: 99%

Escherichia coli membrane‐derived oxygen‐reducing enzyme system (Oxyrase) protects bubaline spermatozoa during cryopreservation

Dalal

Chandolia

Jan

et al. 2020

Molecular Reproduction Devel

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The objective of this study was to determine the effectiveness of deoxygenation of semen extender using Escherichia coli membrane-derived oxygen scavenger (Oxyrase) on post-thaw quality of buffalo (Bubalus bubalis) spermatozoa. Sixteen semen ejaculates, four each from four bulls, were each divided into five equal fractions, diluted using Tris-egg yolk extender supplemented with different concentrations of Oxyrase (0, 0.3, 0.6, 0.9, and 1.2 U/ml), designated as treatments T1, T2, T3, T4, and T5, respectively, and cryopreserved. Immediately after thawing, Oxyrase did not improve sperm kinetics and motility; however, it improved the keeping quality (significantly lower deterioration of post-thaw sperm motility after incubation for 120 min) in T3. Further, T3 reduced (p < .05) cholesterol efflux and protected the intactness of the sperm plasma membrane. Flow cytometry with Fluo-3 AM/propidium iodide (PI) dual staining revealed the highest (p < .05) proportion of live spermatozoa with low intracellular calcium in T3. Oxyrase supplementation protected spermatozoa from premature capacitation which was confirmed by low expression of tyrosine-phosphorylated proteins (32, 75, and 80 kDa) and a relatively lower percentage of F-pattern (uncapacitated spermatozoa) in chlortetracycline assay. Importantly, the Oxyrase fortification decreased superoxide anion in a dose-dependent manner indicating reduced availability of oxygen at sperm mitochondrial level. Similarly, in Oxyrase-fortified sperm, malondialdehyde concentration, an index of lipid peroxidation, is also reduced in a dose-dependent manner. In conclusion, we demonstrate that deoxygenation of buffalo semen by Oxyrase has the potential of improving post-thaw sperm quality by overcoming the problem of cryocapacitation and oxidative damage during cryopreservation process.

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Buffalo Semen Cryopreservation: An Update

Kumar

Adnot

2022

Current Concepts in Bovine Reproduction

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A new role for RU486 (mifepristone): it protects sperm from premature capacitation during cryopreservation in buffalo

Cited by 26 publications

References 46 publications

Low‐density lipoproteins protect sperm during cryopreservation in buffalo: Unraveling mechanism of action

Low‐density lipoproteins protect sperm during cryopreservation in buffalo: Unraveling mechanism of action

Escherichia coli membrane‐derived oxygen‐reducing enzyme system (Oxyrase) protects bubaline spermatozoa during cryopreservation

Buffalo Semen Cryopreservation: An Update

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