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Peter Stacey, Anne Mai Wassermann, Laura Kammonen, Emma Impey, Anna Wilbrey, Darren Cawkill
Screening against a disease-relevant phenotype to identify compounds that change the outcome of biological pathways, rather than just the activity of specific targets, offers an alternative approach to find modulators of disease characteristics. However, in pain research, use of in vitro phenotypic screens has been impeded by the challenge of sourcing relevant neuronal cell types in sufficient quantity and developing functional end-point measurements with a direct disease link. To overcome these hurdles, we have generated human induced pluripotent stem cell (hiPSC)-derived sensory neurons at a robust production scale using the concept of cryopreserved "near-assay-ready" cells to decouple complex cell production from assay development and screening...
March 1, 2018: SLAS Discovery
Sa Cai, Daisy K Y Shum, Ying-Shing Chan
Here we describe the in vitro derivation of sensory neurons for use in effecting fate commitment of Schwann cell-like cells derived from human bone marrow stromal cells (hBMSCs). We adopt a novel combination of small molecules in an 8-day program that induces the differentiation of human induced pluripotent stem cells into sensory neurons. In co-cultures, the derived sensory neurons present contact-dependent cues to direct hBMSC-derived Schwann cell-like cells toward the Schwann cell fate. These derived human Schwann cells survive passaging and cryopreservation, retain marker expression despite withdrawal of glia-inducing medium and neuronal cues, demonstrate capacity for myelination, and therefore promise application in autologous transplantation and re-myelination therapy...
2018: Methods in Molecular Biology
Y P Tsui, Graham K Shea, Y S Chan, Daisy K Y Shum
Our goal is to derive phenotypically stable Schwann cells from bone marrow stromal cells (BMSCs) for use in transplantation studies of central/peripheral nerve injuries. With the adult rat as model, here we describe steps that foster (1) expansion of the BMSC subpopulation of neural progenitors as neurosphere cells, (2) differentiation of the progenitors into Schwann cell-like cells in adherent culture supplemented with soluble factors, and (3) cell-intrinsic switch of Schwann cell-like cells to the Schwann cell fate following co-culture with sensory neurons purified from dorsal root ganglia...
2018: Methods in Molecular Biology
Adriana E Brooks, Gagani Athauda, Mary Bartlett Bunge, Aisha Khan
Cell-based therapies have become a major focus in preclinical research that leads to clinical application of a therapeutic product. Since 1990, scientists at the Miami Project to Cure Paralysis have generated extensive data demonstrating that Schwann cell (SC) transplantation supports spinal cord repair in animals with spinal cord injury. After preclinical efforts in SC transplantation strategies, efficient methods for procuring large, essentially pure populations of SCs from the adult peripheral nerve were developed for rodent and pig studies...
2018: Methods in Molecular Biology
Natalia D Andersen, Paula V Monje
Adult Schwann cell (SC) cultures are usually derived from nerves subjected to a lengthy step of pre-degeneration to facilitate enzymatic digestion and recovery of viable cells. To overcome the need for pre-degeneration, we developed a method that allows the isolation of adult rat sciatic nerve SCs immediately after tissue harvesting. This method combines the advantages of implementing a rapid enzymatic dissociation of the nerve fibers and a straightforward separation of cells versus myelin that improves both cell yield and viability...
2018: Methods in Molecular Biology
Mohamed Fathi, Adel R Moawad, Magdy R Badr
Cryopreservation of oocytes would serve as an alternative to overcome the limited availability of dromedary camel oocytes and facilitate improvements in IVP techniques in this species. Our goal was to develop a protocol for the vitrification of camel oocytes at the germinal vesicle (GV) stage using different cryoprotectant combinations: 20% EG and 20% DMSO (VS1), 25% EG plus 25% DMSO (VS2) or 25% EG and 25% glycerol (VS3) and various cryo-carriers; straws or open pulled-straw (OPS) or solid surface vitrification (SSV); and Cryotop...
2018: PloS One
F de Michele, J Poels, M G Giudice, F De Smedt, J Ambroise, M Vermeulen, D Gruson, C Wyns
STUDY QUESTION: How does the formation of the blood-testis barrier (BTB), as reflected by the expression of connexin 43 and claudin 11 proteins during the pubertal transition period, take place in vitro compared to samples from a large cohort of pre/peripubertal boys? SUMMARY ANSWER: The BTB connexin 43 and claudin 11 expression patterns appeared to be partially achieved in organotypic culture when compared to that in samples from 71 pre/peripubertal patients. WHAT IS KNOWN ALREADY: Although alterations in the protein expression patterns of the BTB, whose main components are connexin 43 and claudin 11, are known to be associated with impaired spermatogenesis in mice and adult men, there is a lack of knowledge on its formation in pre-peripubertal human tissue both in vitro and in vivo...
March 12, 2018: Molecular Human Reproduction
Tzong-Ming Shieh, Chi-Yuan Chen, Chuen Hsueh, Cheng-Chia Yu, Chin-Chuan Chen, Tong-Hong Wang
The quality of biological samples greatly affects the accuracy of scientific results. However, RNA in cryopreserved tissues gradually degrades during storage, leading to errors in the results of subsequent experiments. A suitable sample preservative solution can prolong storage and enhance the research value of samples. Here, we developed a sample preservative solution using the properties of the ribonucleoside vanadyl complex (RVC) and compared its effects on RNA and DNA quality, protein activity, and tissue morphology with the commercially available and widely used RNAlater® Stabilization Solution...
2018: PloS One
Sellappan Selvaraju, Sivashanmugam Parthipan, Lakshminarayana Somashekar, B Krishnan Binsila, Atul P Kolte, Arunachalam Arangasamy, Janivara Parameshwaraiah Ravindra, Stephen A Krawetz
With artificial insemination (AI) and other precision dependent assisted reproductive technologies (ART) being followed in large scale in human and animal reproduction, assessing semen quality and fertilizability is under continuous scrutiny. Various tests have been developed to predict semen quality, but so far no single, highly reliable test is available. In this regard, transcriptomic profiling of spermatozoa assumes significance as it carries the information about spermatogenesis, sperm function, and paternal roles in post-fertilization events...
March 14, 2018: Systems Biology in Reproductive Medicine
Tomáš Štětina, Petr Hůla, Martin Moos, Petr Šimek, Petr Šmilauer, Vladimír Koštál
Physiological adjustments accompanying insect cold acclimation prior to cold stress have been relatively well explored. In contrast, recovery from cold stress received much less attention. Here we report on recovery of drosophilid fly larvae (Chymomyza costata) from three different levels of cold stress: supercooling to -10 °C, freezing at -30 °C, and cryopreservation at -196 °C. Analysis of larval CO2 production suggested that recovery from all three cold stresses requires access to additional energy reserves to support cold-injury repair processes...
March 13, 2018: Scientific Reports
A Mostek, M Slowinska, S Judycka, H Karol, A Ciereszko, M A Dietrich
During semen cryopreservation, spermatozoa are exposed to physical and chemical stressors that result in their functional and structural damage. Growing evidence suggests that most cryoinjuries result from oxidative stress accompanying sperm cryopreservation. Elevated amounts of reactive oxygen species (ROS) generated during cryopreservation can react with sperm macromolecules, including proteins. The goal of this study was to investigate the oxidative modifications (measured as carbonylation level changes) of carp spermatozoa proteins triggered by the cryopreservation process...
March 9, 2018: Journal of Animal Science
Karin de Ruiter, Selma van Staveren, Bart Hilvering, Edward Knol, Nienke Vrisekoop, Leo Koenderman, Maria Yazdanbakhsh
Upon activation granulocytes upregulate several adhesion molecules (CD11b) and granule proteins (CD35, CD66b) and shed surface l-selectin (CD62L). These changes in expression, as assessed by flow cytometry, can be used as markers for activation. Whereas these markers are usually studied in fresh blood samples, a new method is required when samples are collected at a field site with no direct access to a flow cytometer. Therefore, we developed and tested a field-applicable method in which fixed leukocytes were cryopreserved...
March 13, 2018: Cytometry. Part A: the Journal of the International Society for Analytical Cytology
Jessica R Zolton, Toral P Parikh, Dennis D Hickstein, Steven M Holland, Micah J Hill, Alan H DeCherney, Erin F Wolff
PURPOSE: To describe controlled ovarian stimulation (COS) in a population of women with GATA2 deficiency, a genetic bone marrow failure syndrome, prior to allogeneic hematopoietic stem cell transplant METHODS: This is a retrospective case series of nine women with GATA2 deficiency who underwent oocyte preservation at a research institution. Main outcomes measured include baseline fertility characteristics ((antimullerian hormone (AMH) and day 3 follicle-stimulating hormone (FSH) and estradiol (E2)) and total doses of FSH and human menopausal gonadotropins (HMG), E2 on day of trigger, and total number of metaphase II oocytes retrieved...
March 13, 2018: Journal of Assisted Reproduction and Genetics
R Poverini, R Lisi, F Lisi, V Berlinghieri, W Bielli, P Carfagna, A Costantino, D Iacomino, G Nicodemo
Granulocyte-macrophage colony-stimulation factor plays different crucial roles during embryo implantation and subsequent development. Here we aimed to evaluate the effects of embryo cell culture medium, with the inclusion of granulocyte-macrophage colony-stimulation factor (GM-CSF), on embryo development and pregnancy rate. To this end, we took advantage of our retrospective observational study to correlate the outcomes from two different culture media. We included in this study 25 unselected patient from our IVF Center that underwent heterologous IVF cycle with crypreserved oocytes...
December 2018: Cell Death Discovery
Tamás Müller, Hajime Matsubara, Yuki Kubara, Ákos Horváth, Balázs Kolics, János Taller, Viktor Stéger, Balázs Kovács, László Horváth, Juan F Asturiano, David S Peñaranda, Béla Urbányi
The objective of this study was to assess impact of cryopreserved European eel sperm and Japanese eel native sperm on early fertilization, hatch, survival, and malformation rates of larvae, as well as develop molecular techniques to distinguish different eel species. Eggs from Japanese eel females (Anguilla japonica) were artificially fertilized with sperm of Japanese eel males and cryopreserved sperm from European eel (A. anguilla, extender was modified Tanaka solution and methanol as cryoprotectant). There were no statistical differences (p > 0...
March 2, 2018: Theriogenology
Ram V Devireddy
To model the cryobiological responses of cells and tissues, the cellular membrane permeability characteristics are often measured at suprazero temperatures as well as at subzero temperatures with and without the presence of extra-cellular ice. These measured membrane permeability characteristics are then used to predict the responses of cells and tissues for a given thermal insult with the ultimate aim of mitigating the damage caused during the freeze-thaw process. This brief review articles summarizes efforts from my research group over the past 15 years as related to cryobiology of mammalian ovarian tissue sections, i...
March 8, 2018: Minerva Ginecologica
Avi Ben-Haroush, Irit Ben-Aharon, Yechezkel Lande, Benjamin Fisch
BACKGROUND: Controlled ovarian hyperstimulation (COH) followed by oocyte retrieval is a leading option for fertility preservation before chemotherapy, yet this procedure causes excessive serum levels of estradiol (E2), which are often detrimental for cancer patients. Aromatase inhibitors are often used in breast cancer patients during COH to prevent elevated levels of E2. OBJECTIVES: To describe our experience with COH for oocyte cryopreservation in non-breast cancer patients using aromatase inhibitors...
March 2018: Israel Medical Association Journal: IMAJ
Jaime Guerrero, Miguel Gallardo, Adoración Rodríguez-Arnedo, Jorgen Ten, Rafael Bernabeu
The survival of human blastocysts to vitrification with two different carriers is compared. Both vitrification carriers used in this study are in the category of closed carriers, as they completely isolate the samples from direct contact with liquid nitrogen or its vapours during cooling and storage, until warming. This characteristic is appealing because it reduces or eliminates the theoretical risk of cross-contamination during that period of time. The two closed vitrification systems used present very different design and features: in the High Security Vitrification device, the carrier straw containing the embryos is encapsulated inside an external straw before plunging in liquid nitrogen, resulting in thermal insulation during cooling...
March 8, 2018: Cryobiology
Malinda S Lee, Eden R Cardozo, Anatte E Karmon, Diane L Wright, Thomas L Toth
OBJECTIVE: To identify the impact of embryo transfer time (total seconds from the loading of the transfer catheter to the expulsion of the embryo(s) into the uterine cavity) on clinical pregnancy (CPR), implantation (IR), and live birth (LBR) rates. DESIGN: Retrospective cohort study. SETTING: Academic hospital practice. PATIENT(S): A total of 465 women undergoing 571 frozen-embryo transfers with the use of cryopreserved blastocysts in a single academic institution from 2007 through 2014...
March 7, 2018: Fertility and Sterility
Clara Malo, Elizabeth G Crichton, Jane M Morrell, Budhan S Pukazhenthi, Anders Johannisson, Rebecca Splan, Julian A Skidmore
Colloids have been successfully used in a number of species to improve sperm populations for IVF and for cryopreservation The usefulness of Single Layer Centrifugation (SLC) for freezing dromedary camel spermatozoa in two different extenders was evaluated by examining the motility, viability, acrosome status, DNA integrity, and ability of cryopreserved sperm to penetrate oocytes in vitro in a heterologus IVF system. Two ejaculates from each of five males were divided into four aliquots: two were processed by SLC (selected) while two were centrifuged without colloid (control)...
February 12, 2018: Animal Reproduction Science
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