Skip to main content
SpringerLink
Log in

Generation of reactive oxygen species during cryopreservation may improve Lilium × siberia pollen viability

  • Physiology
  • Published:
In Vitro Cellular & Developmental Biology - Plant Aims and scope Submit manuscript

Abstract

During biotic and abiotic stress in plants, reactive oxygen species (ROS) may play two very different roles: high ROS concentrations can exacerbate damage, whereas low concentrations can activate defense responses. The aim of this study was to investigate the relationship between ROS generation and pollen viability after cryopreservation. ROS generation was detected from ‘Siberia’ (Lilium × siberia) pollen using flow cytometry with 2′,7′-dichlorodihydrofluorescein diacetate as a fluorescent probe. Pollen viability was determined by 2,3,5-triphenyltetrazolium chloride staining. ROS generation was slightly increased by rapid cooling (26.13 ± 4.74 vs. 15.80 ± 2.30 for fresh pollen) and significantly increased by vitrification (49.74 ± 1.43; P < 0.01). Pollen viabilities after rapid cooling and vitrification were significantly increased (58.88 ± 3.76% and 70.35 ± 2.90%, respectively) over that of fresh pollen (46.65 ± 1.61%; P < 0.01). No significant differences in ROS generation were associated with cold acclimation at different temperatures before rapid cooling. However, sharp decreases in viability were observed with cold acclimation at 4°C and −20°C relative to rapid cooling without acclimation (P < 0.01). We observed nonsignificant decreases in ROS generation among vitrification treatments that omitted different steps and a significant decrease when the unloading step was omitted (P < 0.05). Pollen viabilities were significantly reduced when the loading or dehydration steps were omitted (P < 0.01). No significant differences were observed in ROS generation or pollen viability among the treatments when 200 U/ml catalase was added to different solutions used in the vitrification process. Comprehensive analysis of all data indicated a positive correlation between ROS generation and pollen viability (r = 0.651, P < 0.001). Therefore, increasing ROS generation during cryopreservation may improve the viability of ‘Siberia’ pollen.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Figure 1.
Figure 2.
Figure 3.
Figure 4.
Figure 5.
Figure 6.

Similar content being viewed by others

References

  • Ahmad P, Sarwat M, Sharma S (2008) Reactive oxygen species, antioxidants and signaling in plants. J Plant Biol 51:167–173

    Article  CAS  Google Scholar 

  • Barnabas B, Rajki E (1976) Storage of maize (Zea mays L.) pollen at −196°C in liquid nitrogen. Euphytica 25:747–752

    Article  Google Scholar 

  • Bathgate R (2011) Antioxidant mechanisms and their benefit on post-thaw boar sperm quality. Reprod Domest Anim 46:23–25

    Article  PubMed  Google Scholar 

  • Baumber J, Ball BA, Linfor JJ (2005) Assessment of the cryopreservation of equine spermatozoa in the presence of enzyme scavengers and antioxidants. Am J Vet Res 66:772–779

    Article  CAS  PubMed  Google Scholar 

  • Benson EE, Harding K (2012) Cryopreservation of shoot tips and meristems: an overview of contemporary methodologies. In: Loyola-Vargas VM, Ochoa-Alejo N (eds) Plant cell culture protocols. Humana Press, Totowa, pp 191–226

    Chapter  Google Scholar 

  • Elkins EC (2006) Cryopreservation of domestic cat (Felis catus) epididymal spermatozoa in the presence of antioxidants. Dissertation, American University, Washington

    Google Scholar 

  • Ganeshan S (1986) Cryogenic preservation of papaya pollen. Sci Hortic (Amsterdam) 28:65–70

    Article  Google Scholar 

  • Ganeshan S, Rajasekharan PE (2005) Conservation and management of haploid genetic diversity through pollen cryopreservation. J Palynol 41:39–48

    Google Scholar 

  • Garg N, Manchanda G (2009) ROS generation in plants: boon or bane? Plant Biosyst 143:81–96

    Article  Google Scholar 

  • Gechev TS, Van Breusegem F, Stone JM, Denev I, Laloi C (2006) Reactive oxygen species as signals that modulate plant stress responses and programmed cell death. Bioessays 28:1091–1101

    Article  CAS  PubMed  Google Scholar 

  • Goes PAA, Nichi M, Silva ROC, Perez EGA, Dalmazzo A, Gurgel JRC, Rocha CC, Simoes R, Peres MA, Assumpcao MEOA, Barnabe RC, Barnabe VH (2010) 75 influence of cryopreservation on the susceptibility of goat sperm against different reactive oxygen species. Reprod Fertil Dev 23:143

    Article  Google Scholar 

  • Houdan A, Véron B, Claquin P, Lefebvre S, Poncet JM (2005) Cryopreservation of the coccolithophore, Emiliania huxleyi (Haptophyta, Prymnesiophyceae). J Appl Phycol 17:413–422

    Article  Google Scholar 

  • Juránek I, Bezek Š (2005) Controversy of free radical hypothesis: reactive oxygen species–cause or consequence of tissue injury? Gen Physiol Biophys 24:263–278

    PubMed  Google Scholar 

  • Karun A, Sajini KK, Nair M, Kumaran PM, Samsudheen K (2006) Cryopreservation of coconut (Cocos nucifera.L) pollen. J Plant Crops 34:568–571

    Google Scholar 

  • Li B (2010) Studies on differentially expressed protein of pollen cryopreservation and cryobank construction of Paeonia spp. Dissertation, Beijing Forestry University, Beijing (in Chinese)

    Google Scholar 

  • Li G (2005) Studies on cryopreservation of Camellia pollen. Dissertation, Beijing Forestry University, Beijing (in Chinese)

    Google Scholar 

  • Li Z, Lin Q, Liu R, Xiao W, Liu W (2010) Protective effects of ascorbate and catalase on human spermatozoa during cryopreservation. J Androl 31:437–444

    Article  CAS  PubMed  Google Scholar 

  • Luz H, Wanderley LS, Faustino LR, Silva CD, Figueiredo JD, Rodrigues A (2011) Role of antioxidants agents in germ cells and embryos cryopreservation. Acta Sci Vet 39:956–969

    Google Scholar 

  • Luza JG, Polito VS (1985) In vitro germination and storage of English walnut pollen. Sci Hortic (Amsterdam) 27:303–316

    Article  Google Scholar 

  • Martinez-Montero ME, Mora N, Quinones J, Gonzalez-Arnao MT, Engelmann F, Lorenzo JC (2002) Effect of cryopreservation on the structural and functional integrity of cell membranes of sugarcane (Saccharum sp.) embryogenic calluses. CryoLetters 23:237–244

    CAS  PubMed  Google Scholar 

  • McCarthy MJ, Meyers SA (2011) Antioxidant treatment in the absence of exogenous lipids and proteins protects rhesus macaque sperm from cryopreservation-induced cell membrane damage. Theriogenology 76:168–176

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Mori IC, Schroeder JI (2004) Reactive oxygen species activation of plant Ca2+ channels. A signaling mechanism in polar growth, hormone transduction, stress signaling, and hypothetically mechanotransduction. Plant Physiol 135:702–708

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Parfitt DE, Almehdi AA (1983) Cryogenic storage of grape pollen. Am J Enol Vitic 34:227–228

    Google Scholar 

  • Parton E, Vervaeke I, Delen R, Vandenbussche B, Deroose R, De Proft M (2002) Viability and storage of bromeliad pollen. Euphytica 125:155–161

    Article  CAS  Google Scholar 

  • Prasad PV, Djanaguiraman M (2011) High night temperature decreases leaf photosynthesis and pollen function in grain sorghum. Funct Plant Biol 38:993–1003

    Article  CAS  Google Scholar 

  • Roca J, Rodríguez MJ, Gil MA, Carvajal G, Garcia EM, Cuello C, Vazquez JM, Martinez EA (2005) Survival and in vitro fertility of boar spermatozoa frozen in the presence of superoxide dismutase and/or catalase. J Androl 26:15–24

    CAS  PubMed  Google Scholar 

  • Rossi T, Mazzilli F, Delfino M, Dondero F (2001) Improved human sperm recovery using superoxide dismutase and catalase supplementation in semen cryopreservation procedure. Cell Tissue Bank 2:9–13

    Article  CAS  PubMed  Google Scholar 

  • Sakai A, Engelmann F (2007) Vitrification, encapsulation-vitrification and droplet-vitrification: a review. CryoLetters 28:151–172

    CAS  PubMed  Google Scholar 

  • Sandermann H (2000) Active oxygen species as mediators of plant immunity: three case studies. Biol Chem 381:649–653

    Article  CAS  PubMed  Google Scholar 

  • Shang X (2005) Studies on cryopreservation of peony pollen. Beijing Forestry University, Dissertation (in Chinese)

    Google Scholar 

  • Sicherle CC, Maia MS, Bicudo SD, Rodello L, Azevedo HC (2011) Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen supplemented with catalase or Trolox. Small Rumin Res 95:144–149

    Article  Google Scholar 

  • Smirnova AV, Matveyeva NP, Polesskaya OG, Yermakov IP (2009) Generation of reactive oxygen species during pollen grain germination. Russ J Dev Biol 40:345–353

    Article  CAS  Google Scholar 

  • Smirnova AV, Timofeyev KN, Breygina MA, Matveyeva NP, Yermakov IP (2012) Antioxidant properties of the pollen exine polymer matrix. Biophysics 57:174–178

    Article  CAS  Google Scholar 

  • Soh N (2006) Recent advances in fluorescent probes for the detection of reactive oxygen species. Anal Bioanal Chem 386:532–543

    Article  CAS  PubMed  Google Scholar 

  • Speranza A, Crinelli R, Scoccianti V, Geitmann A (2012) Reactive oxygen species are involved in pollen tube initiation in kiwifruit. Plant Biol 14:64–76

    CAS  PubMed  Google Scholar 

  • Sukhvibul N, Considine JA (1993) Medium and long term storage of Anigozanthos manglesii (D. Don) pollen. N Z J Crop Hortic 21:343–347

    Article  Google Scholar 

  • Suzuki N, Koussevitzky S, Mittler R, Miller G (2012) ROS and redox signalling in the response of plants to abiotic stress. Plant Cell Environ 35:259–270

    Article  CAS  PubMed  Google Scholar 

  • Tatone C, Di Emidio G, Vento M, Ciriminna R, Artini PG (2010) Cryopreservation and oxidative stress in reproductive cells. Gynecol Endocrinol 26:563–567

    Article  PubMed  Google Scholar 

  • Thomson LK, Fleming SD, Aitken RJ, De Iuliis GN, Zieschang JA, Clark AM (2009) Cryopreservation-induced human sperm DNA damage is predominantly mediated by oxidative stress rather than apoptosis. Hum Reprod 24:2061–2070

    Article  CAS  PubMed  Google Scholar 

  • Uchendu EE, Leonard SW, Traber MG, Reed BM (2010) Vitamins C and E improve regrowth and reduce lipid peroxidation of blackberry shoot tips following cryopreservation. Plant Cell Rep 29:25–35

    Article  CAS  PubMed  Google Scholar 

  • Wang S, Xie B, Yin L, Duan L, Li Z, Egrinya Eneji A, Tsuji W, Tsunekawa A (2010) Increased UV-B radiation affects the viability, reactive oxygen species accumulation and antioxidant enzyme activities in maize (Zea mays L.) pollen. Photochem Photobiol 86:110–116

    Article  CAS  PubMed  Google Scholar 

  • Zhang Y (2007) Pollen cryopreservation of Prunus mume Sieb. et. Zucc. and the construction of pollen bank. Dissertation, Beijing Forestry University, Beijing (in Chinese)

    Google Scholar 

  • Zhang Y, Chen R, Huang C, Liu Y (2009) Cryo-banking of Prunus mume pollen and its application in cross-breeding. CryoLetters 30:165–170

    CAS  PubMed  Google Scholar 

  • Zhao S, Wu L, Yao M, Cheng A (1993) The cryopreservation of tomato pollen. Acta Hortic Sin 20:66–70 (in Chinese)

    Google Scholar 

Download references

Acknowledgments

This work was supported by the National Natural Science Foundation of China (grant no. 31370693). We thank Prof. Jia Guixia for her help in identifying ‘Siberia’ and Prof. Hugh W. Pritchard for his help in revising this paper.

Author information

Authors and Affiliations

  1. Beijing Key Laboratory of Ornamental Plants Germplasm Innovation & Molecular Breeding, National Engineering Research Center for Floriculture, College of Landscape Architecture, Beijing Forestry University, Beijing, 100083, People’s Republic of China

    Jin Xu, Qian Liu, Mengxue Jia, Yan Liu, Bingling Li & Yin Shi

Authors
  1. Jin Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Qian Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mengxue Jia
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yan Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Bingling Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yin Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Yan Liu.

Additional information

Editor: Barbara Reed

Rights and permissions

Reprints and permissions

About this article

Cite this article

Xu, J., Liu, Q., Jia, M. et al. Generation of reactive oxygen species during cryopreservation may improve Lilium × siberia pollen viability. In Vitro Cell.Dev.Biol.-Plant 50, 369–375 (2014). https://doi.org/10.1007/s11627-014-9615-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11627-014-9615-3

Keywords

Search

Navigation