Exposure to cadmium during in vitro maturation at environmental nanomolar levels impairs oocyte fertilization through oxidative damage: A large animal model study

Abstract The COVID-19 pandemic outbreak led to a global ventilator shortage. Hence, different strategies to use a single ventilator to support multiple patients are considered. A mechatronic system Ventil divides and automatically controls gas volume pumped through two channels and was successfully validated in independent lung ventilation. We used Ventil in a series of experiments on a large animal model to verify its usability for ventilation in two patients using a single ventilator. The results of investigations on 12 pigs showed that the physiological level of respiratory parameters was maintained for 24 hours. Application of Ventil did not lead to injuries in the lungs, as indicated by CT scan analysis. We conclude that ventilation using Ventil can be considered safe in patients subjected to deep sedation without spontaneous breathing efforts.

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1 GENERATION OF A STABLE TRANSGENIC SWINE MODEL FOR CELL TRACKING AND CHROMOSOME DYNAMIC STUDIES

Reproduction Fertility and Development ◽

10.1071/rdv28n2ab1 ◽

2016 ◽

Vol 28 (2) ◽

pp. 130

Author(s):

R. Sper ◽

S. Simpson ◽

X. Zhang ◽

B. Collins ◽

J. Piedrahita

Keyword(s):

Cell Cycle ◽

Stem Cells ◽

Animal Model ◽

Regenerative Medicine ◽

Cancer Biology ◽

Large Animal Model ◽

Large Animal ◽

Fetal Fibroblasts ◽

Transgene Transmission

Transgenic pigs are an attractive research model in the field of translational research, regenerative medicine, and stem cell therapy due to their anatomic, genetic, and physiological similarities with humans. The development of a transgenic murine model with a fusion of green fluorescent protein (GFP) to histone 2B protein (H2B, protein of nucleosome core) resulted in an easier and more convenient method for tracking cell migration and engraftment levels after transplantation as well as a way to better understand the complexity of molecular regulation within cell cycle/division, cancer biology, and chromosome dynamics. Up to now the development of a stable transgenic large animal model expressing H2B-GFP has not been described. Our objective was to develop the first transgenic porcine H2B-GFP model via CRISPR-CAS9 mediated recombination and somatic cell nuclear transfer (SCNT). Porcine fetal fibroblasts were cotransfected with CRISPR-CAS9 designed to target the 3′ untranslated region of ACTB locus and a targeting vector containing 1Kb homology arms to ACTB flanking an IRES-H2B-GFP transgene. Four days after transfection GFP cells were fluorescence activated cell sorted. Single cell colonies were generated and analysed by PCR, and heterozygous colonies were used as donor cells for SCNT. The custom designed CRISPR-CAS9 knockin system demonstrated a 2.4% knockin efficiency. From positive cells, 119 SCNT embryos were generated and transferred to a recipient gilt resulting in three positive founder boars (P1 generation). Boars show normal fertility (pregnancies obtained via AI of wild type sows). Generated P1 clones were viable and fertile with a transgene transmission rate of 55.8% (in concordance with Mendel’s law upon chi-square test with P = 0.05). Intranuclear H2B-GFP expression was confirmed via fluorescence microscopy on 8-day in vitro cultured SCNT blastocysts and a variety of tissues (heart, kidney, brain, bladder, skeletal muscle, stomach, skin, and so on) and primary cultured cells (chondrocytes, bone marrow derived, adipocyte derived, neural stem cells, and so on) from P1 cloned boars and F1 42-day fetuses and viable piglets. In addition, chromosome segregation could be easily identified during cell cycle division in in vitro cultured stem cells. Custom designed CRISPR-CAS 9 are able to drive homologous recombination in the ACTB locus in porcine fetal fibroblasts, allowing the generation of the first described viable H2B-GFP porcine model via SCNT. Generated clones and F1 generation expressed H2B-GFP ubiquitously, and transgene transmission rates were with concordance of Mendel’s law. This novel large animal model represents an improved platform for regenerative medicine and chromosome dynamic and cancer biology studies.

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Comparative gonadotoxicity of the chemotherapy drugs cisplatin and carboplatin on prepubertal mouse gonads

MHR Basic science of reproductive medicine ◽

10.1093/molehr/gaaa008 ◽

2020 ◽

Vol 26 (3) ◽

pp. 129-140 ◽

Cited By ~ 6

Author(s):

Caroline M Allen ◽

Federica Lopes ◽

Rod T Mitchell ◽

Norah Spears

Keyword(s):

Animal Model ◽

Cell Population ◽

Parent Compound ◽

Chemotherapy Drugs ◽

Model Study ◽

Concomitant Reduction ◽

Animal Model Study ◽

Potential Benefits ◽

Reduced Toxicity

Abstract The treatment of childhood cancer with chemotherapy drugs can result in infertility in adulthood. Newer generations of drugs are developed to replace parent drugs, with the potential benefits of less toxic side effects. For platinum alkylating-like drugs, in contrast to the parent compound cisplatin, the newer-generation drug carboplatin is reported to have reduced toxicity in some respects, despite being administered at 5–15 times higher than the cisplatin dose. Whether carboplatin is also less toxic than cisplatin to the reproductive system is unknown. Here we compare the gonadotoxic impact of cisplatin and carboplatin on female and male mouse prepubertal gonads. In vitro cultured CD1 mouse ovaries or testis fragments were exposed to either cisplatin or carboplatin for 24 h on Day 2 of culture and analysed by Day 6. A dose response for each drug was determined for the ovary (0.5, 1 & 5 μg/ml cisplatin and 1, 5 & 10 μg/ml carboplatin) and the testis (0.01, 0.05 & 0.1 μg/ml cisplatin and 0.1, 0.5 & 1 μg/ml carboplatin). For the ovary, unhealthy follicles were evident from 1 μg/ml cisplatin (73% unhealthy, P = 0.001) and 5 μg/ml carboplatin (84% unhealthy, P = 0.001), with a concomitant reduction in follicle number (P = 0.001). For the testis, the proliferating germ cell population was significantly reduced from 0.05 μg/ml cisplatin (73% reduction, P = 0.001) and 0.5 μg/ml carboplatin (75% reduction, P = 0.001), with no significant impact on the Sertoli cell population. Overall, results from this in vitro animal model study indicate that, at patient equivalent concentrations, carboplatin is no less gonadotoxic than cisplatin.

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