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Abstracts: CryoLetters 41 (4), 2020

 

 

Volume 41, No. 4 July/August 2020

ISSN 0143-2044

 

 



PERSPECTIVE: Critical cooling and warming rates as a function of CPA concentration
Zonghu Han and John C. Bischof

185-193

 

 


Superoxide dismutase in extender improves the in vitro quality and in vivo fertility of cryopreserved water buffalo (Bubalus bubalis) spermatozoa
Zawar Ahmad, Liaqat Ali, Hussain Ahmed, S. Aftab H. Shah
and S. Murtaza H. Andrabi

194-201

 

 


Liquid storage of Geophagus brasiliensis semen in the presence of different extenders
Jôsie S. Caldas, Carine D. Corcini, Estela F. Silva,
Janaína C. Silva, Tainã F. Cardoso, Izani B. Acosta,
Mário R. C. Figueiredo and Antonio S. Varela Junior

202-208

 

 


Physical-mathematical model of substance redistribution between the cell and its hypertonic solution environment of penetrating cryoprotectants with relevance to membrane potential
A.F. Todrin, O.V. Timofeyeva, Ye.I. Smolyaninova,
L.I. Popivnenko and O.I. Gordienko

209-215

 

 


Depressed supercooling point and increased glycerol concentration in overwintering adult tiger beetles (Cicindelida)
McKenna Burns, Dan Herrera, Tierney Brosius
and Timothy J. Muir

216-222

 

 


Effects of cryolipolysis with plates in localized adiposity with the CrioPlace™ concept
Patrícia Froes Meyer, Rodrigo Marcel Valentim da Silva,
Felipe Rodrigues de Souza, Stephany Luanna Queiroga
Farias, Liliane Santos de Vasconcellos,
Eneida de Morais Carreiro and Fábio dos Santos Borges

223-229

 

 


Cryopreservation of adherent human pluripotent stem cells via a novel dish culture preservation method
Junta Hirayama

230-237

 

 


Effect of different permeable cryoprotectants on the quality of cat epididymal spermatozoa
Kakanang Buranaamnuay

238-245

 

 

 

 

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CryoLetters 41 (4), 185-193 (2020)
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PERSPECTIVE:
CRITICAL COOLING AND WARMING RATES AS A FUNCTION OF CPA CONCENTRATION

Zonghu Han1, John C. Bischof1,2*

1 Department of Mechanical Engineering, University of Minnesota, Minneapolis, USA
2 Department of Biomedical Engineering, University of Minnesota, Minneapolis, USA

Abstract

Cryoprotective agents (CPAs) are routinely applied in cryopreservation protocols to achieve the vitrified state thereby avoiding the damaging effects of ice crystals. Once the CPA has been added, the system needs to cool at a rate ≥ critical cooling rate (CCR) to avoid ice crystallization and successfully enter the vitrified state. Subsequently, upon warming the system needs to meet or exceed a critical warming rate (CWR), often one to two orders of magnitude higher than the CCR, to avoid ice formation and return the system to physiological temperatures for use.   Many experimental and theoretical studies have been published on CCRs and CWRs, and correlation for these rates as a function of concentration has been explored for some single component CPAs, but not the CPA cocktails which are commonly used in tissue and organ cryopreservation. In this paper, we summarize the available data of CCRs and CWRs for a variety of CPAs, and suggest a convenient mathematical expression for CCR and CWR that can guide general use for cryoprotective protocol, but also highlights the critical need for further study on CPA cocktails and tissue systems in which CPAs may behave differently and/or may not be fully equilibrated to the loaded CPA

Keywords: tissue and organ vitrification, plant vitrification, critical cooling rates, critical warming rates, cryoprotective agents.

Download the paper: CRITICAL COOLING AND WARMING RATES AS A FUNCTION OF CPA CONCENTRATION (PDF)

 

 

 

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CryoLetters 41 (4), 194-201 (2020)
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SUPEROXIDE DISMUTASE IN EXTENDER IMPROVES THE IN VITRO QUALITY AND IN VIVO FERTILITY OF CRYOPRESERVED WATER BUFFALO (Bubalus bubalis) SPERMATOZOA

Zawar Ahmad, Liaqat Ali, Hussain Ahmed, S. Aftab H. Shah
and S. Murtaza H. Andrabi*

Animal Reproduction Laboratory, Animal Sciences Institute, National Agricultural Research Center, Islamabad 45500, Pakistan.
* Corresponding author email:
andrabi123@yahoo.com

Abstract

BACKGROUND: Superoxide dismutase (SOD) as an antioxidant in semen extender may be used for the cryopreservation of buffalo spermatozoa and in vivo fertility. OBJECTIVE: This study was aimed to evaluate the effects of SOD (SOD1, 100 IU/mL; SOD2, 200 IU/mL; SOD3, 300 IU/mL) and control (0.0) in Tris citric acid extender on in vitro quality and in vivo fertility of cryopreserved water buffalo bull spermatozoa. MATERIALS AND METHODS: Semen collection was carried out on a weekly basis (four bulls, three replicates, and n = 24 ejaculates). The conventional freezing of semen loaded straws (0.5 mL) was undertaken by placing them horizontally on a steel rack inside a Styrofoam box for 10 min containing liquid nitrogen (LN2) vapours, and plunging into a liquid nitrogen tank (-196 °C) for storage, followed by thawing at 37 °C for 30 s and analysis by computer-assisted sperm analyzer (CASA) and other assays. RESULTS: At post-dilution, the acrosome integrity (ACR-I, %) was significantly improved (P < 0.05) in extender supplemented with SOD3 as compared to other experimental groups. In addition, DNA integrity (DNA-I, %) was significantly higher (P < 0.05) in SOD1 and SOD3 compared to SOD2 and control. At post-thawing, the mean values of sperm progressive motility (PM, %), average path velocity (VAP, µm/s) and straight line velocity (VSL, µm/s) were significantly higher (P < 0.05) in extender supplemented with SOD3 compared to the control. At post-thawing, mean values of subjective motility (SM, %), plasma membrane integrity (PMI, %) and ACR-I were significantly higher (P < 0.05) in extender supplemented with SOD3 compared to the control. At post-thawing, sperm DNA-I was significantly higher (P < 0.05) in extender supplemented with all SOD doses compared to the control in a dose-dependent manner. The in vivo fertility rate (%) was significantly higher with SOD3 compared to the control (68.2 % vs. 49.5 %). CONCLUSION: The supplementation of SOD3 (300 IU/mL) in Tris citric acid extender improves both in vitro quality and in vivo fertility of buffalo bull spermatozoa.

Keywords: buffalo spermatozoa; superoxide dismutase; CASA; in vitro quality; in vivo fertility.

 

 

 

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CryoLetters 41 (4), 202-2081 (2020)
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LIQUID STORAGE OF Geophagus brasiliensis SEMEN IN THE PRESENCE OF DIFFERENT EXTENDERS

Jôsie S. Caldas1, Carine D. Corcini3, Estela F. Silva2, Janaína C. Silva2,
Tainã F. Cardoso2, Izani B. Acosta3, Mário R. C. Figueiredo2
and Antonio S. Varela Junior4*

1 BAAC, Aquatic Environments Continental Biology, Institute of Biological Sciences, Federal University of Rio Grande, Rio Grande, RS, Brazil.
2 RAC - Compared Animal Reproduction, Institute of Biological Sciences, Federal University of Rio Grande, Rio Grande, RS, Brazil.
3 ReproPel, Faculty of Veterinary Medicine, Campus Capão do Leão, Federal University of Pelotas, Pelotas, Brazil.
4 Institute of Biological Sciences, Federal University of Rio Grande, Rio Grande, RS, Brazil.
*Corresponding author email:
antoniovarela@furg.br

Abstract

BACKGROUND: In order to preserve the genetic diversity of cichlid fish in gene banks, it is necessary to use certain extenders to maintain the integrity of spermatozoa cells during cooling. OBJECTIVE: To evaluate the effects of different extenders on the quality parameters of cooled semen of Geophagus brasiliensis. MATERIALS AND METHODS: Semen samples were collected from seven adult fish and diluted with five extenders: Beltsville Thawing Solution (BTS™), Hanks' Balanced Salt Solution (HBSS), Tris-glucose, Ginsburg's Fish Ringers, and Phosphate buffered Saline. All parameters were evaluated in fresh semen samples and after cooling at 4°C at 0, 24, 48, and 96 h to evaluate cell viability (membrane integrity, DNA, and mitochondrial functionality) and motility rate and weather motility. RESULTS: The BTS and Tris-glucose resulted in the best outcomes (P<0.05) in terms of membrane integrity assessments (35.1% and 30.9 %, respectively), DNA integrity (71.6%; 75.7%), mitochondrial function (26.9%; 28.0 %) and motility rate (8.6%; 8.6%), respectively, for semen cooled to 4°C for 96 h. However, the 48-h period motility after cooling in BTS was superior to all other treatments.  CONCLUSION: BTS and Tris-glucose can be considered as the best extenders for the cold storage of Geophagus brasiliensis spermatozoa.

Keywords: fish semen; liquid storage; Tris-glucose; BTS

 

 

 

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CryoLetters 41 (4), 209-215 (2020)
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PHYSICAL-MATHEMATICAL MODEL OF SUBSTANCE REDISTRIBUTION BETWEEN THE CELL AND ITS HYPERTONIC SOLUTION ENVIRONMENT OF PENETRATING CRYOPROTECTANTS WITH RELEVANCE TO MEMBRANE POTENTIAL

A.F. Todrin*, O.V. Timofeyeva, Ye.I. Smolyaninova, L.I. Popivnenko
and O.I. Gordienko

Institute for Problems of Cryobiology and Cryomedicine, The National Academy of Sciences of Ukraine, Kharkiv, Ukraine.
*Corresponding author:
altodrin@gmail.com

Abstract

BACKGROUND: The redistribution of basic ions between the cell cytoplasm and its surrounding medium due to osmotic action affects transmembrane potential and plasma membrane integrity at all stages of low temperature preservation. OBJECTIVE: To develop a physical-mathematical model describing the redistribution of osmotically active solutes between the cell and its hypertonic solutions of penetrating cryoprotectants that enables the calculation of kinetic changes in cell volume, cryoprotectant and ion concentrations, as well as the cell transmembrane potential during cell equilibration with cryoprotectant solutions. MATERIALS AND METHODS: The study has modeled the mass transfer process of mouse oocytes upon exposure to 1.5 M DMSO and 1,2-Propanediol (1,2-PD) solutions. RESULTS: Equations for changes of the normalized volume as well as intracellular concentrations of DMSO, 1,2-PD, potassium, sodium, chlorine and transmembrane potential have been obtained in a dimensionless form. The membrane permeability coefficients for DMSO and 1,2-propanediol have been determined and compared with the data of Paynter et al (5). CONCLUSION: The study shows that the incorporation of transmembrane ion movement and electrical potential change in the mathematical model leads to lower values of mouse oocyte membrane permeability coefficients for water and cryoprotectants in comparison with data determined by the traditional model.

Keywords:  mouse oocyte, water transport, ion transport, transmembrane electrical potential.

 

 

 

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CryoLetters 41 (4), 216-222 (2020)
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DEPRESSED SUPERCOOLING POINT AND INCREASED GLYCEROL CONCENTRATION IN OVERWINTERING ADULT TIGER BEETLES (Cicindelida)

McKenna Burns1,2*, Dan Herrera1, Tierney Brosius1
and Timothy J. Muir1

1Department of Biology, Augustana College IL, Rock Island, IL 61201, USA
2Department of Biology, Miami University, Oxford, OH 45056, USA
*Corresponding author:
burnsmp3@miamioh.edu

Abstract

BACKGROUND: Tiger beetles are a widely distributed group including species that may be exposed to sub-freezing temperature overwinter. Despite being well studied, little is known about tiger beetle cold tolerance. OBJECTIVE: We investigated seasonal changes in cold hardiness of two northerly distributed tiger beetle species (Cicindela repanda and Cicindela limbalis). MATERIALS AND METHODS: We monitored the supercooling point (SCP), glycerol concentration, and hemolymph osmolality of adult tiger beetles during a 3.5-month acclimation to winter.  RESULTS: SCP decreased during winter acclimation for C. repanda, but not for C. limbalis. Both species modestly increased glycerol concentration, and C. repanda increased hemolymph osmolality by 38%. CONCLUSION: This initial investigation into the cold-hardiness of adult tiger beetles suggests that they are capable of lowering their SCP as winter approaches, which may help them survive sub-freezing winter temperatures. Further assessment of their chill and freeze tolerance and of their overwintering conditions in the field is needed to better understand their winter physiology.

Keywords: tiger beetle, overwintering, supercooling point, glycerol, chill-tolerance.

 

 

 

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CryoLetters 41 (4), 223-229 (2020)
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EFFECTS OF CRYOLIPOLYSIS WITH PLATES IN LOCALIZED ADIPOSITY WITH THE CRIOPLACE™ CONCEPT

Patrícia Froes Meyer1; Rodrigo Marcel Valentim da Silva1*;
Felipe Rodrigues de Souza 2, Stephany Luanna Queiroga Farias2;
Liliane Santos de Vasconcellos1; Eneida de Morais Carreiro2;
Fábio dos Santos Borges3

1 Federal University of Rio Grande do Norte (UFRN), Brazil.
2 Potiguar University, Natal/RN, Brazil.
3 Estácio de Sá University, Rio de Janeiro/RJ, Brazil.
*Corresponding author:
rodrigomarcelvalentim@gmail.com

Abstract

BACKGROUND: Plate cryolipolysis is a method of applying cooling without a vacuum system, which can be used in regions with less possibility of skin suction or fibrosis.  OBJECTIVE: To investigate the effects of cryolipolysis with the use of plate-shaped applicators (CrioPlace™) for localized fat treatment. METHODS: The sample consisted of men aged 20 to 45 years with complaints of localized adiposity in the abdominal region and flanks. Two plates were positioned in the flank and abdomen regions, respectively. They received two 60-min applications in the temperature of -2°C. The anthropometric, thermographic and ultrasound assessments were performed, and a satisfaction questionnaire was applied after treatment. The re-evaluations occurred 30 and 60 days after the first intervention. RESULT: A reduction in adiposity was observed in flank region plicometry (p<0.05) and abdominal and flank ultrasound (p <0.05). About 66.7% of the volunteers reported less water retention, about 41.7% reported that their clothes were looser, and 100% reported overall satisfaction. Fifty percent rated the treatment as excellent and 58.3% felt improvement in overall aesthetics. CONCLUSION: The CrioPlace™ method was effective in reducing localized adiposity, with clinical satisfaction of measurement reduction, both in plicometry and ultrasound analyses, with highlights to the flank region results.

Keywords: adipose tissue, cryotherapy, ultrasonography.

 

 

 

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CryoLetters 41 (4), 230-237 (2020)
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CRYOPRESERVATION OF ADHERENT HUMAN PLURIPOTENT STEM CELLS VIA A NOVEL DISH CULTURE PRESERVATION METHOD

Junta Hirayama

Mayekawa MFG Co, Ltd, 2000 Tatsuzawa, Moriya, Ibaraki 302-0118, Japan
For correspondence:
hirayama-junta@mayekawa.co.jp

Abstract

BACKGROUND: Human induced pluripotent stem cells (hiPSCs) are valuable resources for cell therapy and drug discovery. Cryopreservation is a key technique used to realize these applications, which require a large number of cells. However, standard protocols for the preservation of the adherent cells involve multiple complicated steps, which can lead to technical difficulties. OBJECTIVE: To develop a more efficient method for cryopreservation of adherent cells using culture dishes. MATERIALS AND METHODS: Ice-seeding treatment was employed to avoid intercellular freezing, and rapid warming used to improve cell viability. RESULTS: The immediate survival rate after thawing was 48%. The recovery period of cells cryopreserved by the dish culture method was shortened upon subsequent passage culture, and the time for re-cultured cells to reach the appropriate confluency was reduced by two days. CONCLUSION: The hiPSCs can be successfully cryopreserved in culture dishes with improved viability and faster recovery. The optimization of the ice-seeding temperature and cooling rate increased the survival rate.

Keywords: human pluripotent stem cell, ice-seeding, adherent cells, dish culture cryopreservation.

 

 

 

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CryoLetters 41 (4), 238-245 (2020)
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EFFECT OF DIFFERENT PERMEABLE CRYOPROTECTANTS ON THE QUALITY OF CAT EPIDIDYMAL SPERMATOZOA

Kakanang Buranaamnuay

Molecular Agricultural Biosciences Cluster, Institute of Molecular Biosciences (MB), Mahidol University, Nakhon Pathom 73170, Thailand
For correspondence:
ningkakanang@yahoo.com

Abstract

BACKGROUND: Permeable cryoprotectants (CPAs) are required for successful sperm cryopreservation. OBJECTIVE: To investigate the efficacy of adding different CPAs to a freezing extender on cat epididymal sperm quality. METHODS: Epididymal spermatozoa were suspended in Tris-glucose-citrate-egg yolk extender supplemented either with glycerol, methanol, formamide, ethylene glycol (EG), propylene glycol (PG) and dimethylsulfoxide (DMSO), all at 5% (v/v), and then cryopreserved. Sperm motility, viability, functional membrane integrity, morphology and acrosome integrity were examined at post-thaw. RESULTS: Glycerol, formamide, EG and DMSO exhibited good, comparable cryoprotective effects, whereas PG showed moderate cryoprotection. Sperm viability in PG was lower than that in glycerol and EG, but was not different in formamide and DMSO. Contrarily, the least efficacy was observed in methanol. Interestingly, the CPA type has no effect on functional membrane integrity and morphology. CONCLUSION: Using Tris-glucose-citrate-egg yolk extender, formamide, EG and DMSO could substitute glycerol as permeable CPAs for cat epididymal sperm cryopreservation.

Keywords: alcohols, amides, cryoprotectants, felids, sperm freezing.

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