Protective Efficacy of Cholesterol-Loaded Cyclodextrin (CLC) Against Sperm Morphological Abnormalities in Marwari Stallions

Authors

  • Pramod Kumar Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, RAJUVAS, Bikaner, Rajasthan
  • J S Mehta Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, RAJUVAS, Bikaner, Rajasthan
  • S K Ravi Department of Veterinary Gynaecology and Obstetrics, Faculty of Veterinary Science and Animal Sciences, Rajiv Ghandhi South Campus, Barkachha-231001, Banaras Hindu University, Mirzapur, Uttar Pradesh
  • T R Talluri Division of Animal Reproduction, Equine Production Campus, ICAR-National Research Center on Equines, Bikaner, Rajasthan
  • B L Kumawat Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, RAJUVAS, Bikaner, Rajasthan
  • Tapendra Kumar Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, RAJUVAS, Bikaner, Rajasthan
  • Shivendra Kumar Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, RAJUVAS, Bikaner, Rajasthan
  • D Jhamb Department of Veterinary Gynaecology and Obstetrics, College of Veterinary and Animal Science, Navania, Udaipur, Rajasthan
  • Ramesh Kumar Dedar Division of Animal Reproduction, Equine Production Campus, ICAR-National Research Center on Equines, Bikaner, Rajasthan

DOI:

https://doi.org/10.48165/aru.2023.4.1.2

Keywords:

Cholesterol loaded cyclodextrins, cryopreservation, Marwari stallion, sperm abnormalities

Abstract

The present study aimed to examine the protective potential of cholesterol loaded  cyclodextrin (CLC) at varying concentrations against sperm morphological  abnormalities of cooled and frozen thawed semen of Marwari stallions. A total  of forty eight ejaculates (six stallions and eight ejaculates per stallion) were  promptly processed following semen collection and subjected to gross assessment  of semen quality attributes. The assessment of sperm morphologies in fresh  equine semen was conducted using Eosin and Nigrosine staining technique. The  semen samples from individual Marwari stallions were evenly divided into five  fractions and incubated in a water bath at 37°C for 15 min after dilution with  primary extender to obtain the spermatozoa concentration at 120 × 106 sperm/ml.  During incubation, the fractions of every semen sample were exposed to different  concentrations of CLC as 0, 1.0, 1.5, 2.0 and 3.0 mg/ml of extender (C, T1, T2,T3,  and T4, respectively). All the 5 aliquots were cooled and subsequently subjected  to cryopreservation. Each cooled and frozen- thawed (at 37°C for 30 sec) semen  samples were analyzed to assess the morphologies of the spermatozoa. Based on  the findings, it was concluded that none of the CLC treatment could render any  significant protection against the sperm morphological abnormalities at either  pre-freeze and/or post-thaw stage of semen samples from Marwari stallions. 

References

Allen WR. The development and application of the modern reproductive technologies to horse breeding. Reprod Domest Anim. 2005;40:(3)10–29. doi: 10.1111/j.1439- 0531.2005.00602.x.

Alvarez AL, Serres C, Torres P, Crespo F, Mateos E, Gomez Cuetara C. Effect of cholesterol-loaded cyclodextrin on the cryopreservation of donkey spermatozoa. Anim Reprod Sci. 2006;94:89–91. doi: 10.1016/j.anireprosci.2006.04.026

Alvarez AL, Torres P, Serres C, Mateos E, Graham JK, Gomez Cuetara C. Effect of cholesterol-loaded cyclodextrin on cooled semen from Zamorano-Leones donkey. Reprod Domest Anim. 2005;40:375.

Amorim EAM, Graham JK, Spizziri B, Meyers M, Torres CAA. Effect of cholesterol or cholesteryl conjugates on the cryo survival of bull sperm. Cryobiol. 2009;58(2):210-14. doi: 10.1016/j.cryobiol.2008.12.007.

Behera S, Harshan HM, Lekshmi BK, Ghosh AKN. Effect of cholesterol supplementation on cryosurvival of goat sper matozoa. Vet World. 2015;8(12):1386–91. doi: 10.14202/ vetworld.2015.1386-1391.

Belmonte SA, Lopez CI, Roggero CM, De Blas GA, Tomes CN, Mayorga LS. Cholesterol content regulates acroso mal exocytosis by enhancing Rab3A plasma membrane

association. Dev Biol. 2005;285(2):393-408. doi: 10.1016/j. ydbio.2005.07.001.

Blanch E, Tomas C, Graham JK, Moce E. Response of boar sperm to the treatment with cholesterol-loaded-cyclodex trins added prior to cryopreservation. Reprod Dom Anim. 2012;47(6):959-964. doi: 10.1111/j.1439-0531.2012.01999.x.

Boscarato AG, Martins LF, Pinho RO, Silva YFRS, Papa FO, Macedo GG. Effect of adding cyclodextrin loaded with cho lesterol prior to semen frozen/tawed of adult Nellore bulls. Revist Bras de Reprod Anim. 2016;40(3):105-110.

Combes GB, Varner DD, Schroeder F, Burghardt RC, Blanchard TL. Effect of cholesterol on the motility and plasma mem brane integrity of frozen equine spermatozoa after thawing. J Reprod Fertil. 2000; 56:127–132.

Cox R, Evans LE, Youngs CR. The effect of 2-hydroxypro pyl-β-cyclodextrin on post-thaw parameters of cryopre served jack and stallion semen. J Equine Vet Sci. 2013;33(4): 272–8.

Crichton EG, Pukazhenthi BS, Billah M, Skidmore JA. Cholesterol addition aids the cryopreservation of drom edary camel (Camelus dromedarius) spermatozoa. Theriogenology. 2015;83(2):168-74. doi: 10.1016/j.theriog enology.2014.09.005.

Darin-Bennett A, White IG. Influence of the cholesterol con tent of mammalian spermatozoa on susceptibility to cold-shock. Cryobiol. 1977;14:466–70. doi: 10.1016/0011- 2240(77)90008-6.

Gemeda AE, Ghosh SK, Patra MK, Gupta SK, Srivastava N, Kujur A. Physico-morphological Characteristics of Semen Ejaculates in Vrindavani Crossbred Bulls. Anim. Reprod. Update 2022, 2 (2), 31-37. Doi:10.48165/aru.2022.2.2.7.

Guthrie HD, Liu J, Critser JK. Osmotic tolerance limits and effects of cryoprotectants on motility of bovine spermato zoa. Biol Reprod. 2002;67:1811–1816. doi: 10.1095/biolre prod67.6.1811.

Hartwig FP, Lisboa FP, Monteiro GA, Maziero RRD, Freitas Dell Aqua CP, Alvarenga MA, Dell’AquaJr JA. Use of cho lesterol-loaded cyclodextrin: An alternative for bad cooler stallions. Theriogenology. 2014;81(2):340-6. doi: 10.1016/j. theriogenology.2013.10.003.

Inanc ME, Uysal O, Ata A. Cryopreservation and evaluation of Akkaraman ram semen with 7-dehydrocholesterol. Ankara Univ Vet Fak Derg. 2018;65:187-192.

Khan J, Tahir MZ, Khalid A, Sattar A, Ahmad N. Effect of cho lesterol-loaded cyclodextrins on cryosurvival of dog sper matozoa. Reprod Domest Anim. 2017;52(2):265-8. doi: 10.1111/rda.12893.

Kiso WK, Asano A, Travis AJ, Schmitt DL, Brown JL, Pukazhenthi BS. Pretreatment of Asian elephant (Elephas maximus)

spermatozoa with cholesterol-loaded cyclodextrins and glycerol addition at 4°C improves cryosurvival. Reprod Fertil Dev. 2012;24(8):1134–42. doi: 10.1071/RD11266.

Konyali C, Tomas C, Blanch E, Gomez EA, Graham JK, Moce E. Optimizing conditions for treating goat semen with cho lesterol-loaded cyclodextrins prior to freezing to improve cryosurvival. Cryobiol. 2013;67(2):124-31. doi: 10.1016/j. cryobiol.2013.06.001.

Kumar P, Mehta JS, Ravi SK, Dedar RK, Purohit GN, Legha RA, Tripathi BN, Talluri TR. Cholesterol loaded cyclodextrin supplementation enhances the cholesterol-to-phospholipid ratio and diminishes oxidative stress in Jack spermatozoa during cryopreservation. J Equine Vet Sci. 2020;914:1-10. doi: 10.1016/j.jevs.2020.103237.

Kumar P, Mehta JS, Ravi SK, Purohit GN, Chaudhary AK, Dedar RK, Talluri TR. Effect of addition of cholesterol loaded cyclo dextrin (CLC) on sperm abnormalities during cryopreser vation of Poitou jack semen. Vet Prac. 2019b;20(2):218-222.

Kumar P, Mehta JS, Ravi SK, Talluri T Rao, Purohit GN, Chaudhary AK. Cholesterol loaded cyclodextrin increases cryopreserve ability of Marwari stallion (Equus ferus caballus) spermatozoa by increasing cholesterol to phos

pholipid ratio. J Anim Res. 2019a;9(2):227-235.

Lee YS, Lee S, Lee SH, Yang BK, Park CK. Effect of cholester ol-loaded-cyclodextrin on sperm viability and acrosome reaction in boar semen cryopreservation. Anim Reprod Sci. 2015;159:124-30. doi: 10.1016/j.anireprosci.2015.06.006.

Lone SA, Prasad JK, Ghosh SK, Das GK, Balamurugan B, Katiyar R, Verma MR. Effect of incubation on freezability of cholesterol-loaded cyclodextrin treated buffalo (Bubalus bubalis) spermatozoa. Vet World. 2016;9(2):182–5. doi: 10.14202/vetworld.2015.182-185.

Moce E, Graham JK. Cholesterol-loaded cyclodextrins added to fresh bull ejaculates improve sperm cryosurvival. J Anim Sci. 2006;84(4):826-33. doi: 10.2527/2006.844826x.

Moce E, Purdy PH, Graham JK. Treating ram sperm with cho lesterol-loaded cyclodextrins improves cryosurvival. Anim Reprod Sci. 2010;118: 236-247. doi: 10.1016/j.anirepro sci.2009.06.013.

Moore AI, Squires EL, Graham JK. Adding cholesterol to the stallion sperm plasma membrane improves cryosur vival. Cryobiol. 2005;51(3):241-249. doi: 10.1016/j.cryo biol.2005.07.004.

Moraes EA, Matos WC, Graham JK, Ferrari WD Jr. Cholestanol loaded-cyclodextrin improves the quality of stallion spermatozoa after cryopreservation. Anim Reprod Sci. 2015;158:19-24. doi: 10.1016/j.anireprosci.2015.04.004.

Moran DM, Jasko DJ, Squires EL, Amann RP. Determination of temperature and cooling rate which induce cold shock

in stallion spermatozoa. Theriogenology. 1992;38(6):999- 1012. doi: 10.1016/0093-691x(92)90114-7.

Navratil AM, Bliss SP, Berghorn KA, Haughian JM, Farmerie TA, Graham JK, Roberson MS. Constitutive localization of the gonadotropin-releasing hormone (GnRH) receptor to low density membrane microdomains is necessary for GnRH signaling to ERK. J Biol Chem. 2003;278(34):31593-602. doi: 10.1074/jbc.M304273200.

Oliveira RR, Rates DM, Pugliesi G, Carvalho GR. Use of choles terol-loaded cyclodextrin in Donkey semen cryopreserva tion improves sperm viability but results in low fertility in mares. Reprod Dom Anim. 2014;40:845-50. doi: 10.1111/ rda.12379.

Parks JE, Lynch DV. Lipid composition and thermotropic phase behaviour of boar, bull, stallion and rooster sperm mem branes. Cryobiol. 1992;29:255–66. doi: 10.1016/0011- 2240(92)90024-v.

Parks JE, Meachan TN, Saacke RG. Cholesterol and phospho lipids of bovine spermatozoa. II. Effect of liposomes pre pared from egg phosphatidylcholine and cholesterol on sperm cholesterol, phospholipids, and viability at 40C and 370 C. Biol Reprod. 1981;24:399–404. doi: 10.1095/biolre prod24.2.399.

Partyka A, Bonarska-Kujawa D, Spornjak M, Strojecki M, Nizanski W. Modification of membrane cholesterol and its impact on frozen–thawed chicken sperm char acteristics. Zygot. 2016;24(5):714-23. doi: 10.1017/ S0967199416000022.

Pereira R, Rosalia S, Barros A, Sousa M. Major regulatory mechanisms involved in sperm motility. Asian J Androl. 2017;19(1):5-14. doi: 10.4103/1008-682X.167716.

Purdy PH, Graham JK. Effect of cholesterol-loaded cyclo dextrin on the cryosurvival of bull sperm. Cryobiology. 2004;48(1):36-45. doi: 10.1016/j.cryobiol.2003.12.001.

Quinn PJ. Principles of membrane stability and phase behavior under extreme conditions. J Bioenerg Biomembr.1989;21:3-19. doi: 10.1007/BF00762209.

Rajoriya JS, Prasad JK, Ramteke SS, Perumal P, Ghosh SK, Singh M, Srivastava N. Enriching membrane cholesterol improves stability and cryosurvival of buffalo spermatozoa. Anim Reprod Sci. 2016;164:72-81. doi: 10.1016/j.anirepro

sci.2015.11.014.

Salmon VM, Leclerc P, Bailey JL. Cholesterol-loaded cyclodextrin increases the cholesterol content of goat sperm to improve cold and osmotic resistance and maintain sperm function after cryopreservation. Biol Reprod. 2016;94(4):85. doi: 10.1095/biolreprod.115.128553.

Samper JC, Pycock JF, McKinnon AO. Current therapy in equine reproduction. Mossouri, USA. Sunders Elseveir. 2007;2:492. Sieme H, Katila T, Klug E. Effect of semen collection practices on sperm characteristics before and after storage and on fer tility of stallions. Theriogenology. 2004;61(4):769-84. doi: 10.1016/s0093-691x(03)00251-6.

Snedecor GW, Cochran WG. Statistical methods, 8th edition, Lowa State University Press. 1994.

Talluri Rao T, Ravi SK, Singh J, Legha RA, Yash Pal, Gupta AK, Singh RK, Tripathi BN. Some reproductive parameters of Zanskari mares reared in an organized farm under tropical climate. Indian J Anim Sci. 2016;86:163–167.

Tomas C, Gomez-Fernandez J, Gomez-Izquierdo E, Moce E, de Mercado E. Addition of cholesterol-loaded cyclodextrins to the thawing extender: effects on boar sperm quality. Reprod Domest Anim. 2014;49(3):427-32. doi: 10.1111/rda.12291.

Ucan U, Kuçuk N, Ahmad E, Naseer Z, Aksoy M, Serin L, Ceylan A. Effect of different sugars supplemented to the extender in combination with cholesterol-loaded cyclodex trin (CLC) on post-thaw quality of ram spermatozoa. Small Rumi Res. 2016;136:243-6.

Yadav HP, Kumar A, Shah N, Chauhan DS, Saxena A, Yadav S, Swain DK. Effect of cholesterol loaded cyclodextrin sup plementation on tyrosine phosphorylation and apoptosis like changes in frozen thawed Hariana bull spermatozoa. Theriogenology. 2017;96:164-171. doi: 10.1016/j.theriog enology.2017.04.016.

Zahid N, Ahmad E, Aksoy M, Niyazi K,Serin I, Ceylan A, Murat B, Cavit K. Protective effect of cholesterol-loaded cyclodex trin pretreatment against hydrogen peroxide induced oxi dative damage in ram sperm. Cryobiol. 2015;71:18–23. doi: 10.1016/j.cryobiol.2015.06.007.

Zahn FS, Papa FO, Dell Aqua JA Jr. Cholesterol incorporation on equine sperm membrane: effects on post-thaw sperm parameters and fertility. Theriogenology. 2002;58:237–240.

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Published

2023-08-23

How to Cite

Kumar, P., Mehta, J.S., Ravi, S.K., Talluri, T.R., Kumawat, B.L., Kumar, T., … Dedar, R.K. (2023). Protective Efficacy of Cholesterol-Loaded Cyclodextrin (CLC) Against Sperm Morphological Abnormalities in Marwari Stallions . Animal Reproduction Update , 4(1), 3–10. https://doi.org/10.48165/aru.2023.4.1.2