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Abstracts: CryoLetters 36 (3), 2015

 

 

Volume 36, No. 3 May/June 2015

ISSN 0143-2044

 

 


Impact of liquid nitrogen exposure on selected biochemical and structural parameters of hydrated Phaseolus vulgaris L. seeds
Inaudis Cejas, Maribel Rivas, Lelurlys Nápoles,
Pedro Marrero, Lourdes Yabor, Carlos Aragón,
Aurora Pérez, Florent Engelmann,
Marcos Edel Martínez-Montero and José Carlos Lorenzo

149-157

 

 


Toxicity of cryoprotective agents and signaling of insulin-like growth factor in hen clam (Mactra chinensis) embryo
Youn Hee Choi and Taek Jeong Nam

158-164

 

 


Vitrification of Thai native cattle oocytes: effects of ethylene glycol concentrations and exposure time, linoleic acid albumin and cholesterol-loaded methyl-β-cyclodextrin
Jakkhaphan Chasombat, Thevin Vongpralub,
Saksiri Sirisathien, Yupin Phasuk and Pronjit Sonseeda

165-173

 

 


Extending the viability of spermatozoa and eggs of the sea urchin Lytechinus variegatus
Jéssica Malgarin and Charrid Resgalla Jr.

174-181

 

 


Effects of various semen extenders on semen parameters for the purpose of human male fertility preservation
Vickram A S, Kamini A Rao, Archana K, Jayaraman G,
Venkat Kumar S and Sridharan T.B

182-186

 

 


Evaluation of DMSO transport in human articular cartilage: vehicle solutions and effects on cell function
A.G. Kay, P. Rooney, J. Kearney and D.E. Pegg

187-194

 

 


Cryopreservation of in vitro-grown shoot tips of chinese medicinal plant Atractylodes macrocephala Koidz. Using a droplet-vitrification method
Jin-mei Zhang, Bin Huang, Xin-xiong Lu, Gayle M. Volk,
Xia Xin, Guang-kun Yin, Juan-juan He and Xiao-ling Chen

195-204

 

 


Seasonal changes of fatty acid composition in Thitarodes pui Larvae, a host of Ophiocordyceps sinensis
Jiequn Yi, Chenlin Guo, Zhiwen Zou and Guren Zhang

205-212

 

 


Numerical simulation on microwave rewarming of cryopreserved rabbit kidney with embedded superparamagnetic nanoparticles
Tao Wang and Gang Zhao

213-220

 

 


Development of a model to investigate red blood cell surface characteristics after cryopreservation
O.I. Gordiyenko, M.O. Anikieieva, S.L. Rozanova,
S.Ye. Kovalenko, I.F. Kovalenko and E.O.Gordiyenko

221-226

 

 

 

 

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CryoLetters 36 (3), 149-157 (2015)
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IMPACT OF LIQUID NITROGEN EXPOSURE ON SELECTED BIOCHEMICAL AND STRUCTURAL PARAMETERS OF HYDRATED PHASEOLUS VULGARIS L. SEEDS

Inaudis Cejas1, Maribel Rivas2, Lelurlys Nápoles2, Pedro Marrero1,
Lourdes Yabor2, Carlos Aragón2, Aurora Pérez2,
Florent Engelmann3, Marcos Edel Martínez-Montero2
and José Carlos Lorenzo2*

1Faculty of Agronomy, Universidad de Ciego de Ávila, Ciego de Ávila 69450, Cuba.
2Laboratory for Plant Breeding, Centro de Bioplantas, Universidad de Ciego de Ávila, Ciego de Ávila 69450, Cuba. E-mail:
jclorenzo@bioplantas.cu. URL: www.bioplantas.cu.
3RD, UMR DIADE, 911 Avenue Agropolis, BP 64501, 34394 Montpellier Cedex, 5, France, E-mail:
florent.engelmann@ird.fr
*Corresponding author email: jclorenzo@bioplantas.cu

Abstract

BACKGROUND: It is well known that cryopreserving seeds with high water content is detrimental to survival, but biochemical and structural parameters of cryostored hydrated common bean seeds have not been published.  OBJECTIVE: The objective of this work was to study the effect of liquid nitrogen exposure on selected biochemical and structural parameters of hydrated Phaseolus vulgaris seeds. MATERIALS AND METHODS: We cryopreserved seeds at various moisture contents and evaluated: germination; electrolyte leakage; fresh seed weight; levels of chlorophyll pigments, malondialdehyde, other aldehydes, phenolics and proteins; thickness of cotyledon epidermis, parenchyma, and starch storage parenchyma; and radicle and plumule lengths. RESULTS: Germination was totally inhibited when seeds were immersed in water for 50 min (moisture content of 38%, FW basis) before cryopreservation. The combined effects of seed water imbibition and cryostorage decreased phenolics (free, cell wall-linked, total), chlorophyll a and protein content. By contrast, electrolyte leakage and levels of chlorophyll b and other aldehydes increased as a result of the combination of these two experimental factors. These were the most significant effects observed during exposure of humid seed to liquid nitrogen.   CONCLUSION: Further studies are still required to clarify the molecular events taking place in plant cells during cryostorage.

Keywords: Common bean, cryopreservation, cryostorage

 

 

 

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CryoLetters 36 (3), 158-164 (2015)
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TOXICITY OF CRYOPROTECTIVE AGENTS AND SIGNALING OF INSULIN-LIKE GROWTH FACTOR IN HEN CLAM (MACTRA CHINENSIS) EMBRYOS

Youn Hee Choi and Taek Jeong Nam*

Institute of Fisheries Sciences, Pukyong National University, Busan 619-911, South Korea
*Corresponding author email:
namtj@pknu.ac.kr

Abstract

BACKGROUND: Signaling of Insulin-like growth factor-I (IGF-I) is involved in development, growth, reproduction and aging of organisms. OBJECTIVE: The work investigated the toxicity of glycerol, dimethyl sulfoxide (DMSO), and ethylene glycol (EG) to hen clam (Mactra chinensis) embryos, as well as the possible role of the insulin-like growth factor-I (IGF-I) during the development and growth of embryos after freeze. MATERIALS AND METHODS: Effects of glycerol, DMSO and EG at different concentrations were tested. The relationship between larval viability and signaling of IGF-I receptor after cryoprotective treatment and/or freezing was examined using immuno-blot analysis. RESULTS: Glycerol had the highest toxicity, followed by DMSO or EG. No embryo survived freeze and thaw without CPAs. After freeze, the activation of the IGF-I signaling pathway, including the IGF-I receptor (IGF-IR) ß-subunit, could be detected in freeze-thawed embryos. The level of IGF-IR expression was very weak in freeze-thawed embryos. CONCLUSION: The survival and developmental rate of embryos was closely related to CPA concentration. IGF-IR was activated and regulated the downstream IGF-I signaling in embryos. The reduced activation of IGF-IR could be related to the death of hen clam embryos.

Keywords: cryoprotective agent, embryo preservation, hen clam, IGF-I signaling

 

 

 

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CryoLetters 36 (3), 165-173 (2015)
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VITRIFICATION OF THAI NATIVE CATTLE OOCYTES: EFFECTS OF ETHYLENE GLYCOL CONCENTRATIONS AND EXPOSURE TIME, LINOLEIC ACID ALBUMIN AND CHOLESTEROL-LOADED METHYL-Β-CYCLODEXTRIN

Jakkhaphan Chasombat1, Thevin Vongpralub2*,
Saksiri Sirisathien3, Yupin Phasuk2 and Pronjit Sonseeda4

1Faculty of Agriculture, Princess of Naradhiwas University, Narathiwas 96000, Thailand,
2Department of Animal Science, Faculty of Agriculture, Khon Kaen University, Khon Kaen 40002, Thailand.,
3Department of Surgery and Theriogenology, Faculty of Veterinary Medicine, Khon Kaen University, Khon Kaen 40002, Thailand.,
4Faculty of Agriculture and Technology, Nakhon Phanom University, Nakhon Phanom 48000, Thailand,
*Corresponding author email:
vthevi@kku.ac.th.

Abstract  

The present study aimed to improve the oocyte vitrification procedure for preservation of Thai native cattle genetic resources. In Experiment I, oocytes were exposed to various doses (2%, 4% and 6%) of ethylene glycol (EG) in vitrification solution I (VS-I) for different equilibration times (10 or 20 min) before being exposed to VS-II and then subjected to vitrification. Experiment II was divided into two parts: (a) oocytes were matured in medium supplemented with linoleic acid albumin (LAA) (1% or 2%) and then vitrified; (b) matured oocytes were preincubated with cholesterol-loaded methyl- β-cyclodextrin (CLC) (1% or 2%) and then vitrified. Equilibration of oocytes by exposure to 6% EG in VS-I for 10 min (Experiment I), and in vitro maturation of immature oocytes in medium supplementation with 2% LAA (Experiment II) were the most effective methods; vitrified/thawed oocytes showed higher rates of survival and subsequent embryonic development compared with the other experimental groups.

Keywords: cholesterol; cryoprotectant; linoleic acid albumin; oocyte cryopreservation; tropical cattle

 

 

 

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CryoLetters 36 (3), 174-181 (2015)
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EXTENDING THE VIABILITY OF SPERMATOZOA AND EGGS OF THE SEA URCHIN LYTECHINUS VARIEGATUS

Jéssica Malgarin and Charrid Resgalla Jr.*

Centro de Cięncias Tecnológicas da Terra e do Mar (CTTMar) - Univerdade do Vale do Itajaí (UNIVALI). Rua Uruguai, 458, Itajaí, SC, 88.302-901. Brazil.
*Corresponding author email:
cresgalla@univali.br

Abstract

BACKGROUND: The storage of spermatozoa and eggs of the sea urchin Lytecninus variegatus can meet the demand of different human activities. OBJECTIVE: To develop a protocol easy to reproduce for spermatozoa cryopreservation and cooling of the eggs of the sea urchin. MATERIALS AND METHODS: Different formulations of artificial sea water were tested for their effectiveness in the freezing of sea urchin spermatozoa and storage of the eggs. RESULTS: Protocol for freezing of spermatozoa in liquid nitrogen presented the positive results when the cryoprotectant solution was diluted in artificial seawater free of calcium and magnesium. For the conservation of the eggs by cooling, the calcium-free artificial sea water, the calcium- and magnesium-free sea water, and the low-sodium water proved more efficient in preserving the integrity of the eggs. CONCLUSION: The results showed success in the freezing protocol of spermatozoa and cooling of the eggs mainly in artificial calcium- and magnesium-free sea water.

Keywords: artificial seawater, echinodermata, cooling, freezing, gamic material, storage.

 

 

 

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CryoLetters 36 (3), 182-186 (2015)
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EFFECTS OF VARIOUS SEMEN EXTENDERS ON SEMEN PARAMETERS FOR THE PURPOSE OF HUMAN MALE FERTILITY PRESERVATION

Vickram A S, Kamini A Rao, Archana K, Jayaraman G,
Venkat Kumar S and Sridharan T.B*

Department of Industrial Biotechnology, School of Biosciences and Technology, VIT University, Vellore, India.
*Corresponding author email:
tbsridharan@vit.ac.in

Abstract

BACKGROUND: Cryopreservation enables semen to be preserved at subzero temperatures, usually at -196°C. There is a need in preparing good extender for the semen to be cryopreserved until use, especially in the field of assisted reproduction. OBJECTIVE: To elucidate the apt extender for preserving both infertile and fertile samples for a minimum period and to check the post thaw results for various extenders used.  MATERIALS AND METHODS: A total of 103 samples were collected for this research, and after semen analysis the semen samples were categorized into oligospermia (n=20), oligoasthenospermia (n = 22), asthenospermia (n=24), normospermia (n=28), and control (n = 9). RESULTS: The extender supplemented with various antioxidants yields better results when compared to all the other extenders in case of fertile and infertile samples. CONCLUSION: Supplementing semen extender with antioxidants and various ingredients is the concern in designing an apt semen extender recipe. This research prescribes antioxidant extender (E4) to preserve the infertile and fertile semen samples for the purpose of research and also for doing assisted reproduction.

Keywords: Antioxidants, assisted reproduction, cryoinjury, glycerol, semen extender.

 

 

 

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CryoLetters 36 (3), 187-194 (2015)
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EVALUATION OF DMSO TRANSPORT IN HUMAN ARTICULAR CARTILAGE: VEHICLE SOLUTIONS AND EFFECTS ON CELL FUNCTION

A.G. Kay1,2,3*, P. Rooney1, J. Kearney1 and D.E. Pegg3

1R&D Tissue Services, NHS Blood & Transplant, 14 Estuary Banks, Speke, Liverpool L24 8RB, UK.
2Leopold Muller Arthritis Research Centre, Robert Jones and Agnes Hunt Orthopaedic Hospital Foundation Trust, Oswestry, Shropshire SY10 7AG, UK
3Biology Department, University of York, York YO10 5DD, UK
*Corresponding author email:
A.G.Kay@keele.ac.uk

Abstract

Osteochondral allografting techniques are limited by the availability of suitable donor tissue; there is an urgent need for effective cryopreservation. A fundamental requirement is the need to establish initial conditions of exposure to cryoprotectant that the chondrocytes will tolerate and that load the tissue with an adequate concentration of cryoprotectant. Three vehicle solutions to transport DMSO into the tissue were studied. Knee joints were obtained from deceased donors with appropriate consent. Whole condyles were treated with 20% w/w DMSO in each of three vehicle solutions and chondrocyte function and tissue CPA content measured. The results showed that exposure to 20% DMSO in each vehicle solution for 2 hours at 0°C was tolerated without loss of GAG synthetic activity. It was observed that penetration of DMSO increased little after 1 hour of CPA exposure at 0°C but the final tissue concentration of CPA was markedly lower than that in the medium.

Keywords: Cartilage, cryoprotectant, vehicle solutions, Liquidus Tracking

 

 

 

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CryoLetters 36 (3), 195-204 (2015)
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CRYOPRESERVATION OF IN VITRO-GROWN SHOOT TIPS OF CHINESE MEDICINAL PLANT ATRACTYLODES MACROCEPHALA KOIDZ. USING A DROPLET-VITRIFICATION METHOD

Jin-mei Zhang1, Bin Huang2, Xin-xiong Lu1, Gayle M. Volk3,
Xia Xin1, Guang-kun Yin1, Juan-juan He1 and Xiao-ling Chen1*

1National Genebank, Institute of Crop Science, Chinese Academy of Agricultural Sciences, Beijing 100081, China;
2Fujian Agriculture and Forestry University, Fuzhou 350002, China;
3USDA-ARS National Center for Genetic Resources Preservation, 1111 S. Mason St., Fort Collins, Colorado 80521, U.S.A.
*Corresponding author email:
chenxiaoling@caas.cn

Abstract

BACKGROUND: Atractylodes macrocephala Koidz. is an important medicinal species from China that has been used for thousands of years for its special pharmacological antioxidant, hepatoprotective, anti-inflammatory, anti-allergic, antithrombotic, antiviral, and anticarcinogenic activities. OBJECTIVE: The aim of this research was to develop an efficient droplet-vitrification protocol for A. macrocephala shoot tips which could be used as a strategy for long-term conservation within gene banks. MATERIALS AND METHODS: The duration of preculture, loading, and PVS2 steps, as well as the recovery medium formulation, were optimized to achieve high levels of survival and regrowth for A. macrocephala shoot tips after liquid nitrogen exposure. RESULTS: Survival and regrowth levels after cryopreservation in the cultivar 'Baizhu' were as high as 76% and 62%, respectively. Thermal analysis using differential scanning calorimetry suggested that the PVS2 treatment plays a critical role for successful cryopreservation. CONCLUSION: The droplet-vitrification method established in this study could be used to cryopreserve A. macrocephala.

Keywords: Atractylodes macrocephala Koidz., droplet-vitrification, regrowth, shoot tips, survival, thermal analysis

 

 

 

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CryoLetters 36 (3), 205-212 (2015)
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SEASONAL CHANGES OF FATTY ACID COMPOSITION IN Thitarodes pui Larvae, A HOST OF Ophiocordyceps sinensis

Jiequn Yi1, Chenlin Guo1, Zhiwen Zou1* and Guren Zhang2*

1 School of Life Sciences, Nanchang University, Nanchang;
2 State Key Laboratory for Biocontrol & Institute of Entomology, Sun Yat-Sen University, Guangzhou, China.
*Corresponding author email:
zouzhiwen@ncu.edu.cn or zhanggr@mail.sysu.edu.cn

Abstract

BACKGROUND: Thitarodes larvae are the host of Ophiocordyceps sinensis and exist in the permafrost region of the Tibetan Plateau. OBJECTIVE: To understand the adaptation mechanism of Thitarodes larvae to seasonal fluctuations of ambient temperatures in the Tibetan Plateau by studying seasonal changes of the fatty acids composition in the larvae of T. pui. MATERIALS AND METHODS: The profile of fatty acids in the 6th instar T. pui larvae collected at the mid-month in a whole year were examined by GC-MS. RESULTS: There was a negative correlation between the total lipid and ambient (soil) temperature. Further study indicated that oleic, palmitic, linoliec, palmitoleic, stearic were the major fatty acids. The ratio of unsaturated fatty acids to saturated fatty acids (U/S) and the unsaturated index (UI) in triacylglycerols remain stable during the whole year, while the U/S and UI in phospholipids vary dramatically in response to soil temperature. CONCLUSION: The fluctuations in phospholipids were attributed to seasonal changes of oleic and linoleic. The changes of the fatty acid composition may result from their adaptation to the vairation of temperature in different seasons.

Keywords: Thitarodes pui, total lipid, fatty acid composition

 

 

 

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CryoLetters 36 (3), 213-220 (2015)
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NUMERICAL SIMULATION ON MICROWAVE REWARMING OF CRYOPRESERVED RABBIT KIDNEY WITH EMBEDDED SUPERPARAMAGNETIC NANOPARTICLES

Tao Wang1,2 and Gang Zhao1,3*

1Centre for Biomedical Engineering, Department of Electronic Science & Technology, University of Science and Technology of China, Hefei;
2School of Electronic Science & Applied Physics, Hefei University of Technology, Hefei;
3Anhui Provincial Engineering, Technology Research Center for Biopreservation and Artificial Organs, Hefei, China.
*Corresponding author email:
zhaog@ustc.edu.cn

Abstract

BACKGROUND: Rewarming cryopreserved organs without detrimental damages is a task full of challenging, since devitrification-associated thermal stresses can cause uncontrollable injuries. The ideal rewarming method should obtain a uniform thermal field with a rapid warming rate enough to avoid devitrification. Microwave rewarming is considered to be the most promising method to rewarm cryopreserved organs safely. However, it is difficult to accurately predict the rewarming rates and temperature gradients in cryopreserved organs since the coupling of electromagnetic field and temperature field is changing during microwave rewarming process.  OBJECTIVE: This study is to evaluate the feasibility of microwave rewarming of cryopreserved rabbit kidney embedded with superparamagnetic nanoparticles in a single-mode resonant cavity (434MHz, TE101 mode). MATERIALS AND METHODS: The Finite Element Method (FEM) was used to calculate the coupling of the electromagnetic field and temperature field in a microwave system composed of a rectangular resonant cavity, an antenna source, and a frozen rabbit kidney with temperature-dependent properties. Heat generated by water molecules and nanoparticles in the electromagnetic field of microwave cavity was calculated. RESULTS: The simulation results showed that, during the rewarming process of the sample phantom without nanoparticles, the rewarming rate was 29.45° C/min and the maximum temperature gradient in the sample was 2.23°C/mm. With nanoparticles embedded in the sample at the same power input to the microwave cavity, the rewarming rate was increased to be 41.38°C/min and the maximum temperature gradient in the sample was 1.93°C/mm. CONCLUSION: The study indicates that the use of nanoparticles increases the rewarming rate and temperature uniformity.

Keywords: microwave resonant cavity, superparamagnetic nanoparticles, finite element method.

 

 

 

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CryoLetters 36 (3), 221-226 (2015)
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DEVELOPMENT OF A MODEL TO INVESTIGATE RED BLOOD CELL SURFACE CHARACTERISTICS AFTER CRYOPRESERVATION

O.I. Gordiyenko1, M.O. Anikieieva2*, S.L. Rozanova1, S.Ye. Kovalenko1, I.F. Kovalenko1 and E.O.Gordiyenko1

1Institute for Problems of Cryobiology and Cryomedicine NAS of Ukraine, Kharkiv, Ukraine.
2National University of Pharmacy, Kharkiv, Ukraine.
*Corresponding author email: 
Maryano4ka2008@ukr.net

Abstract

BACKGROUND: Maintaining cell surface properties after freezing and thawing, characterized in particular by the surface potential and associated with it cell ability to intercellular adhesion, could be used as a characteristic of successful cryopreservation.  OBJECTIVE: This study was conducted to research applying different erythrocytes freezing modes and analyses the regimes cryopreservation effect on the cell surface charge and adhesion to microorganisms. MATERIALS AND METHODS: Human erythrocytes frozen by three modes. In order to determine adhesion index was used dried bacterial cells of S. thermophilus. The surface charge of erythrocytes was evaluated using Alcian blue cationic dye.  RESULTS: The results showed the significant decrease in the lactobacillus adhesion to erythrocytes frozen glycerol and 1,2-propanediol. After erythrocytes were freezen with glycerol and 1,2-propanediol, the cationic dye binding to erythrocytes significantly reduced. AB binding to erythrocytes frozen with PEG-1500 does not differ from control data. CONCLUSION: Erythrocytes frozen with PEG-1500 mantained surface properties after thawing better, compared to erythrocytes cryopreserved by other methods.

Keywords: Erythrocytes, S. thermophilus, cryopreservation, adhesion, surface charge.

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