scholarly journals Blastocyst complementation using Prdm14-deficient rats enables efficient germline transmission and generation of functional mouse spermatids in rats

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Toshihiro Kobayashi ◽  
Teppei Goto ◽  
Mami Oikawa ◽  
Makoto Sanbo ◽  
Fumika Yoshida ◽  
...  
Keyword(s):  
Pluripotent Stem Cells ◽  
Genetically Modified ◽  
Artificial Chromosome ◽  
Germline Transmission ◽  
Cell Competition ◽  
Somatic Development ◽  
Blastocyst Complementation ◽  
Genetically Modified Animals

AbstractMurine animal models from genetically modified pluripotent stem cells (PSCs) are essential for functional genomics and biomedical research, which require germline transmission for the establishment of colonies. However, the quality of PSCs, and donor-host cell competition in chimeras often present strong barriers for germline transmission. Here, we report efficient germline transmission of recalcitrant PSCs via blastocyst complementation, a method to compensate for missing tissues or organs in genetically modified animals via blastocyst injection of PSCs. We show that blastocysts from germline-deficient Prdm14 knockout rats provide a niche for the development of gametes originating entirely from the donor PSCs without any detriment to somatic development. We demonstrate the potential of this approach by creating PSC-derived Pax2/Pax8 double mutant anephric rats, and rescuing germline transmission of a PSC carrying a mouse artificial chromosome. Furthermore, we generate mouse PSC-derived functional spermatids in rats, which provides a proof-of-principle for the generation of xenogenic gametes in vivo. We believe this approach will become a useful system for generating PSC-derived germ cells in the future.

scholarly journals Transfer to the clinic: refining forward programming of hPSCs to megakaryocytes for platelet production in bioreactors

2021 ◽  
Vol 5 (7) ◽  
pp. 1977-1990
Author(s):  
Amanda L. Evans ◽  
Amanda Dalby ◽  
Holly R. Foster ◽  
Daniel Howard ◽  
Amie K. Waller ◽  
...  
Keyword(s):  
Small Molecules ◽  
Pluripotent Stem Cells ◽  
Thrombus Formation ◽  
Platelet Production ◽  
Hemostatic Effect ◽  
Physical Cues ◽  
Definitive Hematopoiesis

Abstract The production of in vitro–derived platelets has great potential for transfusion medicine. Here, we build on our experience in the forward programming (FoP) of human pluripotent stem cells (hPSCs) to megakaryocytes (MKs) and address several aspects of the complex challenges to bring this technology to the bedside. We first identify clinical-grade hPSC lines that generate MKs efficiently. We design a bespoke media to maximize both production and maturity of MKs and improve platelet output. Crucially, we transition the lentiviral-based FoP of hPSCs to a nonviral inducible system. We also show how small molecules promote a definitive hematopoiesis phenotype during the differentiation process, thereby increasing the quality of the final product. Finally, we generate platelets using a bioreactor designed to reproduce the physical cues that promote platelet production in the bone marrow. We show that these platelets are able to contribute to both thrombus formation in vitro and have a hemostatic effect in thrombocytopenic mice in vivo.

scholarly journals Neutrophils Derived from Genetically Modified Human Induced Pluripotent Stem Cells Circulate and Phagocytose Bacteria In Vivo

2019 ◽  
Vol 8 (6) ◽  
pp. 557-567 ◽  
Author(s):  
Lisa R. Trump ◽  
Ramesh C. Nayak ◽  
Abhishek K. Singh ◽  
Sana Emberesh ◽  
Ashley M. Wellendorf ◽  
...  
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Genetically Modified ◽  
Induced Pluripotent

Gradient biomimetic platforms for neurogenesis studies

2021 ◽  
Author(s):  
Laurissa Havins ◽  
Andrew Capel ◽  
Steven D Christie ◽  
Mark P Lewis ◽  
Paul Roach
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Scale Up ◽  
Cell Culture Techniques ◽  
Culture Techniques ◽  
Chemical Gradients ◽  
The Central Nervous System ◽  
3D Volume

Abstract There is a need for the development of new cellular therapies for the treatment of many diseases, with the central nervous system (CNS) currently an area of specific focus. Due to the complexity and delicacy of its biology, there is currently a limited understanding of neurogenesis and consequently a lack of reliable test platforms, resulting in several CNS based diseases having no cure. The ability to differentiate pluripotent stem cells into specific neuronal sub-types may enable scalable manufacture for clinical therapies, with a focus also on the purity and quality of the cell population. This focus is targeted towards an urgent need for the diseases that currently have no cure, e.g. Parkinson’s disease. Differentiation studies carried out using traditional 2D cell culture techniques are designed using biological signals and morphogens known to be important for neurogenesis in vivo. However, such studies are limited by their simplistic nature, including a general poor efficiency and reproducibility, high reagent costs and an inability to scale-up the process to a manufacture-wide design for clinical use. Biomimetic approaches to recapitulate a more in vivo-like environment are progressing rapidly within this field, with application of bio(chemical) gradients presented both as 2D surfaces and within a 3D volume. This review focusses on the development and application of these advanced extracellular environments particularly for the neural niche. We emphasise the progress that has been made specifically in the area of stem cell derived neuronal differentiation. Increasing developments in biomaterial approaches to manufacture stem cells will enable the improvement of differentiation protocols, enhancing the efficiency and repeatability of the process with a move towards up-scaling. Progress in this area brings these techniques closer to enabling the development of therapies for the clinic.

scholarly journals piggyBac-Based Non-Viral In Vivo Gene Delivery Useful for Production of Genetically Modified Animals and Organs

Pharmaceutics ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 277 ◽  
Author(s):  
Masahiro Sato ◽  
Emi Inada ◽  
Issei Saitoh ◽  
Satoshi Watanabe ◽  
Shingo Nakamura
Keyword(s):  
Gene Delivery ◽  
Mammalian Cells ◽  
Viral Vectors ◽  
Genetically Modified ◽  
Basic Research ◽  
Sleeping Beauty ◽  
In Vivo Gene Delivery ◽  
Genetically Modified Animals

In vivo gene delivery involves direct injection of nucleic acids (NAs) into tissues, organs, or tail-veins. It has been recognized as a useful tool for evaluating the function of a gene of interest (GOI), creating models for human disease and basic research targeting gene therapy. Cargo frequently used for gene delivery are largely divided into viral and non-viral vectors. Viral vectors have strong infectious activity and do not require the use of instruments or reagents helpful for gene delivery but bear immunological and tumorigenic problems. In contrast, non-viral vectors strictly require instruments (i.e., electroporator) or reagents (i.e., liposomes) for enhanced uptake of NAs by cells and are often accompanied by weak transfection activity, with less immunological and tumorigenic problems. Chromosomal integration of GOI-bearing transgenes would be ideal for achieving long-term expression of GOI. piggyBac (PB), one of three transposons (PB, Sleeping Beauty (SB), and Tol2) found thus far, has been used for efficient transfection of GOI in various mammalian cells in vitro and in vivo. In this review, we outline recent achievements of PB-based production of genetically modified animals and organs and will provide some experimental concepts using this system.

Shall I trust you? From child human-robot interaction to trusting relationships

2019 ◽  
Author(s):  
Cinzia Di Dio ◽  
Federico Manzi ◽  
Giulia Peretti ◽  
Angelo Cangelosi ◽  
Paul L. Harris ◽  
...  
Keyword(s):  
Behavioral Pattern ◽  
Human Robot Interaction ◽  
School Age Children ◽  
Social Relevance ◽  
Robot Interaction ◽  
Cognitive Correlates ◽  
The Social ◽  
The Relationship

Studying trust within human-robot interaction is of great importance given the social relevance of robotic agents in a variety of contexts. We investigated the acquisition, loss and restoration of trust when preschool and school-age children played with either a human or a humanoid robot in-vivo. The relationship between trust and the quality of attachment relationships, Theory of Mind, and executive function skills was also investigated. No differences were found in children’s trust in the play-partner as a function of agency (human or robot). Nevertheless, 3-years-olds showed a trend toward trusting the human more than the robot, while 7-years-olds displayed the reverse behavioral pattern, thus highlighting the developing interplay between affective and cognitive correlates of trust.

scholarly journals Influence of calcium ion-modified implant surfaces in protein adsorption and implant integration

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Eduardo Anitua ◽  
Andreia Cerqueira ◽  
Francisco Romero-Gavilán ◽  
Iñaki García-Arnáez ◽  
Cristina Martinez-Ramos ◽  
...  
Keyword(s):  
Protein Adsorption ◽  
Titanium Implant ◽  
Volume Density ◽  
Bone Implant ◽  
The Mean ◽  
Intervention Costs ◽  
Medial Section ◽  
Post Implantation

Abstract Background Calcium (Ca) is a well-known element in bone metabolism and blood coagulation. Here, we investigate the link between the protein adsorption pattern and the in vivo responses of surfaces modified with calcium ions (Ca-ion) as compared to standard titanium implant surfaces (control). We used LC–MS/MS to identify the proteins adhered to the surfaces after incubation with human serum and performed bilateral surgeries in the medial section of the femoral condyles of 18 New Zealand white rabbits to test osseointegration at 2 and 8 weeks post-implantation (n=9). Results Ca-ion surfaces adsorbed 181.42 times more FA10 and 3.85 times less FA12 (p<0.001), which are factors of the common and the intrinsic coagulation pathways respectively. We also detected differences in A1AT, PLMN, FA12, KNG1, HEP2, LYSC, PIP, SAMP, VTNC, SAA4, and CFAH (p<0.01). At 2 and 8 weeks post-implantation, the mean bone implant contact (BIC) with Ca-ion surfaces was respectively 1.52 and 1.25 times higher, and the mean bone volume density (BVD) was respectively 1.35 and 1.13 times higher. Differences were statistically significant for BIC at 2 and 8 weeks and for BVD at 2 weeks (p<0.05). Conclusions The strong thrombogenic protein adsorption pattern at Ca-ion surfaces correlated with significantly higher levels of implant osseointegration. More effective implant surfaces combined with smaller implants enable less invasive surgeries, shorter healing times, and overall lower intervention costs, especially in cases of low quantity or quality of bone.

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