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Abstracts: CryoLetters 41 (6), 2020

 

 

Volume 41, No. 6 November/December 2020

ISSN 0143-2044

 

 


PERSPECTIVE: Nanoparticle-mediated delivery of cryoprotectants for cryopreservation
Samantha Stewart, Alyssa Arminan and Xiaoming He

308-316

 

 


Difference in freezing tolerance between young potato plants derived from tissue culture plantlets and seed tubers
Kenta Kawamura, Jiwan P. Palta, Masahiko Mori
and Jun Kasuga

317-322

 

 


Oocytes of the endangered gray catfish (Surubim-do-Paraíba) submitted to different conservation protocols
Taís da Silva Lopes and Elizabeth Romagosa

323-329

 

 


Gene expression of fresh and frozen/thawed canine embryos
Carlos Renato de Freitas Guaitolini,
Rosiara Rosaria Dias Maziero,
Anthony César de Souza Castilho,
Ana Augusta Pagnano Derussi, Rodrigo Volpato,
Camila Louise Ackermann, Lídia Mitsuko Matsubara
and Maria Denise Lopes

330-336

 

 


Practical and safe method of long-term cryopreservation for clinical application of human adipose-derived mesenchymal stem cells without a programmable freezer or serum
Siqiang Gao, Mika Ogawa, Akiyoshi Takami,
Kyosuke Takeshita, Hidefumi Kato and Takayuki Nakayama

337-343

 

 


The protective effects of alpha lipoic acid on human sperm function during freezing-thawing
Erfaneh Shaygannia, Rana Ghandehari-Alavijeh,
Marziyeh Tavalaee and Mohammad

344-350

 

 


Effect of disaccharide inclusion in vitrification and warming solutions on developmental competence of vitrified/warmed germinal vesicle stage (GV) buffalo oocytes
Amr S. El-Shalofy, Sayed T. Ismail, Aly A.B. Badawy,
Gamal M. Darwish, Magdy R. Badr and Adel R. Moawad

351-357

 

 


Skimmed milk diluent promotes the sperm motility and conception rate of Dorper sheep compared to vitamin B12 diluent
Yuan-Yuan Bai, Xiu Xu, Xiao-Jie Yu, Jian Guo, Xiao-Xue Dong,
Xin-Yang Wang, Zhen-Ao Zhao and Jing Wang

358-364

 

 


Cryopreservation of bovine semen using extract of Bertholletia excelsa (Brazil nuts)
Franciellem Thaiuina de Souza Silva, Janaina Barros Luz,
Luis Rennan Sampaio Oliveira, Rafael Mezzomo,
Daiany Íris Gomes, Cláudia Siqueira Caldas,
Aller Silva Soares,  José Carlos Ferreira-Sila,
Marcos Antonio Lemos Oliveira and Kaliandra Souza Alves

365-370

 

 


Subject and species indices

371-374

 

 

 

 

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CryoLetters 41 (6), 308-316 (2020)
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PERSPECTIVE:
NANOPARTICLE-MEDIATED DELIVERY OF CRYOPROTECTANTS FOR CRYOPRESERVATION

Samantha Stewart1, Alyssa Arminan1 and Xiaoming He1-3*

1Fischell Department of Bioengineering, University of Maryland, College Park, MD 20742, United States
2Robert E Fischell Institute for Biomedical Devices, University of Maryland, College Park, MD 20742, United States
3Marlene and Stewart Greenebaum Comprehensive Cancer Center, University of Maryland, Baltimore, MD 21202, United States
*Corresponding author's E-mail:
shawnhe@umd.edu

Abstract

Nanotechnology research has continued to garner interest and is investigated across a number of fields and industries, ranging from water treatment to clinical and biomedical applications. In biomedical research, for example, polymeric nanoparticles can be leveraged for controlled delivery of drugs and chemical compounds into cells. In cryobiological applications, polymeric nanoparticles can be utilized to deliver cryoprotectants (CPAs) and other protective agents, particularly those impermeable to the cell membrane, into cells to study their effects on cells during cooling down and warming back and at low temperatures. This perspective will discuss how polymeric nanoparticles have been used in cryobiology, with particular focus on how delivery systems have been specifically developed for low temperature applications and the potential for these systems going forward.

Keywords: nanotechnology, trehalose, CPA, cell banking, tissue banking

Download the paper: NANOPARTICLE-MEDIATED DELIVERY OF CRYOPROTECTANTS FOR CRYOPRESERVATION (PDF)

 

 

 

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CryoLetters 41 (6), 317-322 (2020)
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DIFFERENCE IN FREEZING TOLERANCE BETWEEN YOUNG POTATO PLANTS DERIVED FROM TISSUE CULTURE PLANTLETS AND SEED TUBERS

Kenta Kawamura1, Jiwan P. Palta2, Masahiko Mori1 and Jun Kasuga1*

1 Obihiro University of Agricultural and Veterinary Medicine, Nishi 2-11, Inada, Obihiro, Hokkaido 080-8555, Japan.
2 Department of Horticulture, University of Wisconsin-Madison, Madison, WI 53706, USA.
*Corresponding author's E-mail:
jkasuga@obihiro.ac.jp

Abstract

BACKGROUND: Although potato as a crop is commercially grown from seed tubers, plants grown from tissue culture plantlets are often used in physiological studies including freezing tolerance determination.  OBJECTIVE: This study aimed to examine the effects of the source of plants on freezing tolerance of potato plants at young developmental stages.  MATERIALS AND METHODS: We compared freezing tolerance and contents of soluble proteins and sugars of Solanum tuberosum plants derived from tissue culture with those derived from tubers before and after cold acclimation. RESULTS: Tuber-derived plants showed significantly higher freezing tolerance than tissue-culture-derived plants after cold acclimation, although non-acclimated plants did not show any marked differences. Soluble protein contents were higher in tuber-derived plants regardless of cold acclimation. Sucrose content increased to a higher level in tuber-derived plants after cold acclimation.  CONCLUSION: These results suggest that source of plant tissue can have a significant effect on the response of young potato plants to freezing stress and that the use of tissue culture plants in freezing tolerance studies may not accurately reflect the frost tolerance of commercially grown plants.

Keywords: frost tolerance, Solanum tuberosum, tissue culture, minituber, soluble protein, soluble sugar.

 

 

 

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CryoLetters 41 (6), 323-329 (2020)
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OOCYTES OF THE ENDANGERED GRAY CATFISH (SURUBIM-DO-PARAÍBA) SUBMITTED TO DIFFERENT CONSERVATION PROTOCOLS

Taís da Silva Lopes 1* and Elizabeth Romagosa2

1Department of Animal Science, Universidade Federal do Espírito Santo, Alegre, ES 29500-000, Brazil;
2Fishery Institute, APTA, SAA, São Paulo, SP 05001-970 Brazil.
*Corresponding author's E-mail:
tais.peixegen@gmail.com

Abstract

BACKGROUND: The gray catfish known as Surubim-do-Paraíba (Steindachneridion parahybae), which is endemic to the Paraíba do Sul river basin, is on the red list of Brazilian fauna threatened with extinction and the cryopreservation of germ cells of this fish is needed in support of conservation. OBJECTIVE: We aimed to assess the effect of storage temperature on S. parahybae mature and immature oocytes. METHODS: Two trials were carried out. Trial I (TI.1-3) used 30 mature oocytes (diameter >1.8 mm) placed in cryoprotectant solutions and submitted to three different techniques. Trial II (TII.1-3) used 30 immature oocytes (diameter <1.6 mm) placed in cryoprotectant solutions and  submitted to three storage temperatures (i.e., TII.1 at room temperature for 120 min; TII.2 in the freezer for 120 min; TII.3 in liquid nitrogen for 24 hours. RESULT: The mature oocytes were sensitive to every protocol used, including at room temperature. In contrast, the immature oocytes had increased sensitivity according to the temperature reduction to which they were submitted, with the treatment in liquid nitrogen causing greater damage. CONCLUSION: The immature stages exhibit more promising results, encouraging further studies using the combination of different CPSs, mainly penetrating ones, in oocyte cryopreservation protocols. 

 

Keywords: cryopreservation, endangered species, freezing protocol, storage temperature.

 

 

 

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CryoLetters 41 (6), 330-336 (2020)
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GENE EXPRESSION OF FRESH AND FROZEN/THAWED CANINE EMBRYOS

Carlos Renato de Freitas Guaitolini*1, Rosiara Rosaria Dias Maziero1, Anthony César de Souza Castilho2, Ana Augusta Pagnano Derussi3, Rodrigo Volpato4, Camila Louise Ackermann4,
Lídia Mitsuko Matsubara4 and Maria Denise Lopes4

1Paranaense University, Umuarama/PR, Brazil.
2Oeste Paulista University, Presidente Prudente/SP, Brazil.
3José do Rosário Vellano University, Alfenas/MG, Brazil.
4Department of Radiology and Animal Reproduction, FMVZ/UNESP, Botucatu/SP, Brazil.
*Corresponding author's E-mail:
carlosrfg@hotmail.com

Abstract

BACKGROUND: Canine embryo cryopreservation and subsequent transfer are relevant in the use of reproductive technologies. OBJECTIVE: The purpose of this study is the identification and quantification of the gene expression BAX and Bcl2, AQP3, Na+/K+ ATPase α1 and β1 and LIFr in canine embryos obtained in vivo and after freezing. MATERIALS AND METHODS: For the collection of embryos, the bitches were identified at pro-estrous until the detection of 80-90% superficial cells. After that, they were artificially inseminated with fresh semen. The embryos were collected after ovariohysterectomy. RNA was extracted and amplified, and embryos were randomly distributed into fresh (Fr) and frozen/thawed (Ft) groups. RESULTS: Eighteen blastocysts were collected from three bitches. Genes BAX, AQP3 and LIFr did not differ among the studied groups. CONCLUSION: We suggest, through these results, that the genes BAX, Bcl2, AQP3, Na + / K + ATPase α1 and β1 and LIFr were expressed in canine blastocysts collected in vivo and after slow freezing cryopreservation.

Keywords: genes, embryos, slow freezing.

 

 

 

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CryoLetters 41 (6), 337-343 (2020)
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PRACTICAL AND SAFE METHOD OF LONG-TERM CRYOPRESERVATION FOR CLINICAL APPLICATION OF HUMAN ADIPOSE-DERIVED MESENCHYMAL STEM CELLS WITHOUT A PROGRAMMABLE FREEZER OR SERUM

Siqiang Gao1*, Mika Ogawa2, Akiyoshi Takami3, Kyosuke Takeshita4, Hidefumi Kato1 and Takayuki Nakayama2

1Department of Transfusion Medicine, Aichi Medical University, Nagakute, Aichi 480-1195, Japan.
2Clinical Laboratory, Aichi Medical University, Nagakute, Aichi 480-1195, Japan.
3Department of Hematology, Aichi Medical University, Nagakute, Aichi 480-1195, Japan.
4Department of Clinical Laboratory, Saitama Medical Center, Kawagoe, Saitama 350-8550, Japan.
*corresponding author's E-mail:
sgao0303@aichi-med-u.ac.jp

Abstract

BACKGROUND: Adipose-derived mesenchymal stem cells (ADSCs) have emerged as a promising modality for cellular therapy. However, techniques of ADSC cryopreservation, which can facilitate their clinical application, haven't been established yet. OBJECTIVE: To determine optimal conditions for ADSC cryopreservation.  MATERIALS AND METHODS: We used three cryoprotectants [serum containing 10% dimethyl sulfoxide; CP-1™ (5% dimethyl sulfoxide, serum-free); Stem-CellBanker™ (dimethyl sulfoxide and serum-free)], two storage temperatures (−80°C, −150°C) and two cell densities (1 × 106, 7 × 106 cells/mL). Storage was up to 18 months using cryovials. We didn't use a rate-controlled freezer or liquid nitrogen storage. RESULTS: We found that CP-1™ was a suitable cryoprotectant. Storage at −150°C and higher cell density (7×106 cells/mL) kept the best viability of ADSCs, but storage at −80°C and a lower cell density (1×106 cells/mL) is acceptable for up to 9 months. We also confirmed large quantities of ADSCs, stored with CP-1 in a cryobag, were still viable after −150°C cryopreservation for 24 months. CONCLUSION: We have developed a safe, cost-effective way to cryopreserve ADSCs that could be used in the clinical setting.

Keywords: adipose-derived mesenchymal stem cells, cryopreservation, serum-free cryoprotectant.

 

 

 

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CryoLetters 41 (6), 344-350 (2020)
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THE PROTECTIVE EFFECTS OF ALPHA LIPOIC ACID ON HUMAN SPERM FUNCTION DURING FREEZING-THAWING

Erfaneh Shaygannia1, Rana Ghandehari-Alavijeh1, Marziyeh Tavalaee1 and Mohammad Hossein Nasr-Esfahani1,2 *

1Department of Animal Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran.
2Isfahan Fertility and Infertility Center, Isfahan, Iran.
*Corresponding author's E-mail:
nasr.royan@gmail.com and mh.nasr-esfahani@royaninstitute.org

Abstract

BACKGROUND: Sperm cryopreservation is presently used for conservation of male gametes in assisted reproduction technologies (ART). Despite the benefits of sperm banking, freeze-thawing process is injurious to sperm integrity due to induced oxidative stress by cold stress. Oxidative stress reduces sperm motility, viability and DNA integrity. OBJECTIVE: To investigate the effect of alpha lipoic acid (ALA) on human sperm function during the freeze-thawing process. MATERIALS AND METHODS: Thirty semen samples were collected and different concentrations (0, 0.05, 0.1, 0.2, 0.4, 0.8, and 8mM) of ALA were added to a sperm freeze medium and its effects on sperm motility, DNA damage, and lipid peroxidation of frozen–thawed spermatozoa were assessed. RESULTS: The addition of 0.2 mM ALA to the sperm freeze medium resulted in significant improvement in percentage of sperm motility, less DNA damage and decreased lipid peroxidation during freeze-thawing process (p<0.05). CONCLUSION: ALA improves the cryo-protective capacity of sperm freeze medium used for human semen by protecting the sperm from ROS attack induced by the freezing–thawing process. We suggest that sperm freeze medium supplemented with 0.2 mM ALA would be beneficial for the cryopreservation of male gametes in ART.

Keywords: alpha lipoic acid, freezing–thawing, sperm motility, DNA damage.

 

 

 

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CryoLetters 41 (6), 352-357 (2020)
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EFFECT OF DISACCHARIDE INCLUSION IN VITRIFICATION AND WARMING SOLUTIONS ON DEVELOPMENTAL COMPETENCE OF VITRIFIED/WARMED GERMINAL VESICLE STAGE BUFFALO OOCYTES

Amr S. El-Shalofy 1, Sayed T. Ismail1, Aly A.B. Badawy1,
Gamal M. Darwish 2, Magdy R. Badr 2 and Adel R. Moawad1*

1Department of Theriogenology, Faculty of Veterinary Medicine, Cairo University, PO BOX 12211, Giza, Egypt.
2Department of AI and ET, Animal Reproduction Research Institute, Agriculture Research Centre, Giza, Egypt.
*For correspondence:
adelreda902@hotmail.com.

Abstract

BACKGROUND: Cryopreservation of immature oocyte is a potential strategy for preserving the female germline, providing a non-seasonal, easily accessible source for reproduction and science. Exposure of oocytes to high concentrations of cryoprotectants during vitrification is toxic and can negatively impact the fertilization ability and development of vitrified/warmed oocytes. OBJECTIVE: 1) to evaluate the effects of exposure of buffalo germinal vesicle (GV) oocytes to different vitrification solutions (VS), either supplemented with or without sucrose, on cumulus expansion and nuclear maturation following IVM; and 2) to compare the effects of sucrose and trehalose in the warming solution on developmental competence of buffalo oocytes vitrified at the GV-stage. MATERIALS AND METHODS: Cumulus oocyte complexes (COCs) obtained at slaughter from mature buffalo ovaries were randomly assigned into five groups: control - directly subjected to IVM); VS1 group – exposed to 20% ethylene glycol (EG) + 20% glycerol (GLY) + 0.5 M sucrose;  VS2 group - exposed to 20% EG + 20% GLY; VS3 group - subjected to 20% EG+20% dimethyl sulfoxide (DMSO) + 0.5 M sucrose; and VS4 group - subjected to 20% EG+20% DMSO. Following cryoprotectant dilution, viable oocytes were matured in vitro for 22 h; cumulus expansion and nuclear maturation were then evaluated (Experiment 1). COCs were vitrified by solid surface vitrification (SSV) in a solution composed of 20% EG + 20% DMSO (VS4). Following vitrification, COCs were warmed in a solution composed of either sucrose or trehalose in decreasing concentrations (1 M, 0.5 M and 0.25 M). Morphologically viable oocytes were matured, fertilized and cultured in vitro. Cleavage and blastocyst rates were evaluated at 30 h and day 7 post-insemination (p.i.), respectively (Experiment 2). RESULTS: Exposure of GV-buffalo oocytes to different cryoprotectant combinations did not significantly affect cumulus expansion following IVM. However, nuclear maturation rate (oocytes at MII) was significantly higher (P< 0.05) in the groups exposed to sucrose-free vitrification solutions (VS2 and VS4) and not significantly different from the control. Compared with the control group, the cleavage and blastocyst rates were significantly (P<0.05) lower in oocytes vitrified and then warmed in a solution containing trehalose; whilst this was not the case when sucrose was present in the solution. CONCLUSION: Our results suggest that exposure of buffalo GV-oocytes to sucrose-free vitrification solutions improved nuclear maturation after IVM. Moreover, warming of vitrified buffalo oocytes in sucrose-based solution improved preimplantation development following IVM and IVF compared to trehalose based media.

Keywords: buffalo, GV-oocytes, sucrose, trehalose, vitrification, warming.

 

 

 

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CryoLetters 41 (6), 358-364 (2020)
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SKIMMED MILK DILUENT PROMOTES THE SPERM MOTILITY AND CONCEPTION RATE OF DORPER SHEEP COMPARED TO VITAMIN B12 DILUENT

Yuan-Yuan Baia,#, Xiu Xub,#, Xiao-Jie Yua, Jian Guob, Xiao-Xue Donga, Xin-Yang Wanga, Zhen-Ao Zhaoc, * and Jing Wanga,*

aCollege of Animal Science and Technology, Hebei North University, Zhangjiakou, 075000, Hebei, P.R. China.
bZhangjiakou Animal Husbandry Technology Extension Station, Zhangjiakou, 075000, Hebei, P.R. China.
cInstitute of Microcirculation & Department of Pathophysiology of Basic Medical College, Hebei North University, Zhangjiakou, 075000, Hebei, P.R. China.
*Corresponding authors' E-mails:
wangjing197410@163.com and zhaozhenao@hebeinu.edu.cn
#
Co-first authors: Yuan-Yuan Bai and Xiu Xu

Abstract

BACKGROUND: Dorper sheep is an ideal breed for improvement, with higher meat production and increased adaptability. Artificial insemination is an efficient technique for Dorper genetic improvement and reproduction management. However, there is no uniform diluent for Dorper semen dilution. OBJECTIVE: To compare the effects of vitamin B12 (VB12) and skimmed milk diluents on sperm motility at different ratios and time points, and the effects on conception rate. MATERIALS AND METHODS: We detected the effect of diluents on sperm density, deformity, motility and conception rate of Dorper sheep. RESULTS: We found the optimal dilution ratio of skimmed milk is 1:3. Compared to VB12, skimmed milk at 1:3 ratio prolonged semen storage time (48 h vs. 18 h, storage at a low temperature of 4°C) and increased the survival index of sperm (44.7 ± 2.8 vs. 18.5 ± 0.6, P<0.01). CONCLUSION: Skimmed milk is more effective, nutritious and convenient than vitamin B12, representing a more advantageous diluent.

Keywords: ram, semen, motility, survival time, survival index.

 

 

 

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CryoLetters 41 (6), 365-370 (2020)
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CRYOPRESERVATION OF BOVINE SEMEN USING EXTRACT OF Bertholletia excelsa (BRAZIL NUTS)

Franciellem Thaiuina de Souza Silva1, Janaina Barros Luz1*,
Luis Rennan Sampaio Oliveira1, Rafael Mezzomo1, Daiany Íris Gomes1,
Cláudia Siqueira Caldas1, Aller Silva Soares1,
José Carlos Ferreira-Silva2, Marcos Antonio Lemos Oliveira2
and Kaliandra Souza Alves1

1Laboratory of Biotechnology in Animal Reproduction of Carajás, Federal Rural University of Amazonia, Parauapebas-PA, Brazil.
2Laboratory of Reproductive Biotechniques, Federal Rural University of Pernambuco, Recife-PE, Brazil.
*Corresponding author's E-mail:
janaina.ufra@hotmail.com

Abstract

BACKGROUND: Semen cryopreservation is essential in animal breeding programs for improving the availability of genetic resources from animals with high breeding value. OBJECTIVE: To evaluate the addition of Brazil nut extract as a replacement for egg yolk in bovine semen cryopreservation. MATERIALS AND METHODS: Semen was collected from five Nelore bulls and cryopreserved with the addition (treatments) of 0, 25, 50, 75, or 100% Brazil nut extract in the cryoprotectant medium. After thawing, spermatic cells were evaluated for morphology, plasma membrane integrity, spermatic kinetics, and in vitro fertilization. The experimental design was in randomized blocks, and the data were submitted to regression analysis. RESULTS: The minor-type and total defects, and plasma membrane integrity were affected (P < 0.05) as a function of egg yolk substitution with Brazil nut extract. There was a significant effect (P < 0.05) of Brazil nut extract addition on the spermatic kinetics and cleavage rate. CONCLUSION: The addition of Brazil nut extract in the cryoprotective medium as a substitute of egg yolk for freezing bovine semen negatively affects sperm quality and fertility.

Keywords: Antioxidant, freezing, cryoprotectant, plasma membrane.

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