Induced Pluripotent Stem Cells in Asian Bats

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Questions and Answers

What is the primary purpose of introducing Yamanaka factors into somatic cells?

To enhance their immune response
To induce pluripotency (correct)
To increase their lifespan
To strengthen their viral resistance

Which technique was traditionally used to deliver reprogramming factors into target cells?

Microinjection
Nanoparticle delivery
Electroporation
Retroviruses or lentiviruses (correct)

What is a significant risk associated with using viral vectors for reprogramming?

Increased cost of the procedure
Reduced cell viability
Integration into the host genome (correct)
Lack of cell type specificity

Which of the following is NOT one of the Yamanaka factors used for reprogramming?

p53 (B) Signup and view all the answers

What advancement has been made in reprogramming techniques recently?

Development of non-integrating methods (A) Signup and view all the answers

Which of the following is an example of a non-integrating method for delivering reprogramming factors?

Protein delivery systems (C) Signup and view all the answers

What is the main function of the Yamanaka factors in the reprogramming process?

To reset gene expression patterns (D) Signup and view all the answers

What characteristic of non-integrating methods enhances the safety profile of iPSC generation?

Avoidance of permanent genetic modifications (C) Signup and view all the answers

What dye was used to counterstain the nuclei during the experiment?

DAPI (C) Signup and view all the answers

What was the dilution factor for the secondary antibodies used in the experiment?

1:200 (B) Signup and view all the answers

Which method was employed to assess the resultant teratoma histologically?

Haematoxylin and eosin staining (C) Signup and view all the answers

What is one of the indicators that would lead to the termination of the experiment?

Eating limitations due to tumor (D) Signup and view all the answers

What is the initial cell type used in the reprogramming process described?

Bat fetal fibroblast cells (C) Signup and view all the answers

Which of the following factors are carried by the Sendai viral particles used in the study?

Sox2, Oct3/4, Klf4, c-Myc (C) Signup and view all the answers

Which staining technique was used to examine muscle fibers or mesoderm derivatives?

Masson's trichrome stain (C) Signup and view all the answers

What was used for RNA extraction in the quantitative real-time PCR process?

TRIzol® reagent (B) Signup and view all the answers

What medium was used for culturing the transferred colonies?

Stempan E14 GMEM or TeSR™-E8™medium (B) Signup and view all the answers

What step follows the fixation of cell colonies in paraformaldehyde for alkaline phosphatase detection?

AP staining (B) Signup and view all the answers

How many BEFs were seeded onto the gelatin-coated wells prior to viral transduction?

3 × 10^5 (C) Signup and view all the answers

What was used to ensure the exclusion of autofluorescence in the experiment?

Using controls without primary antibodies (B) Signup and view all the answers

What solution was used to dissociate the colonies for passaging?

0.1% Trypsin/1 mM EDTA (A) Signup and view all the answers

At what confluency were individual cells expanded onto 10 cm dishes?

60–70% (D) Signup and view all the answers

Who approved the animal treatment and maintenance in the study?

Institutional Animal Care and Use Committee (IACUC) (B) Signup and view all the answers

Which chemical supplier is mentioned in the study for obtaining materials?

Sigma-Aldrich (Merck KGaA) (C) Signup and view all the answers

What type of cell lines were established using the integration-free CytoTune® iPS 2.0 Sendai Reprogramming Kit?

Bat-induced pluripotent stem cells (D) Signup and view all the answers

Which of the following factors had the highest amino acid identity compared to humans?

Sox2 (D) Signup and view all the answers

What potential outcome is anticipated from introducing BatiPSCs into nude mouse embryos?

Regeneration of thymic tissue (A) Signup and view all the answers

What is a significant advantage of the Sendai virus-based system for producing iPSCs?

It maintains genomic integrity (A) Signup and view all the answers

What was observed regarding chimeric formation in the study?

No chimeric embryos were formed (C) Signup and view all the answers

What specific stage of stem cells is necessary for creating chimeras between bats and mice?

Intermediate-stage differentiated stem cells (B) Signup and view all the answers

Which reprogramming factor had the lowest amino acid identity when compared to humans?

Myc (C) Signup and view all the answers

What is one of the risks associated with methods that integrate into the host genome?

Insertional mutagenesis (C) Signup and view all the answers

What method was used to dissociate BatiPSCs into single cells?

0.1% trypisin/1 mM EDTA solution (B) Signup and view all the answers

What was the primary purpose of washing the colonies three times with PBS?

To eliminate non-adherent cells (D) Signup and view all the answers

What was the incubation temperature and CO2 concentration specified for the humidified incubator?

37°C with 5% CO2 (B) Signup and view all the answers

What was used as a fluorescent signal to detect the target cDNA amounts during RT-qPCR?

TOPreal qPCR 2 PreMIX (SYBR green with low ROX) (B) Signup and view all the answers

What was the final step in the thermal cycling protocol for RT-qPCR?

40 seconds at 72°C (D) Signup and view all the answers

How much total RNA was isolated for conversion to cDNA?

200 ng (D) Signup and view all the answers

What was the purpose of replacing the medium with a fresh differentiation medium after 24 hours?

To eliminate non-adherent cells (A) Signup and view all the answers

What is the density at which the cells were plated onto the petri dishes?

2 × 10^6 cells per dish (B) Signup and view all the answers

What are induced pluripotent stem cells primarily used for?

Predictive toxicology as an alternative to animal testing (A) Signup and view all the answers

What is a potential advantage of using stem cell organoids in research?

They provide a more accurate representation of human diseases (C) Signup and view all the answers

Which of the following is a limitation of utilizing stem cell technologies?

High potential for ethical dilemmas (B) Signup and view all the answers

What does the data availability statement imply about the study's findings?

The data can be accessed by request from the author (B) Signup and view all the answers

Who authored the study on using stem cells as alternatives to animal experimentation?

Kim T-W, Che J-H, Yun J-W (A) Signup and view all the answers

Which publication discusses the availability and applications of induced pluripotent stem cells?

World J Stem Cells (D) Signup and view all the answers

What does the reference to bat-induced pluripotent stem cells suggest about the scope of stem cell research?

It includes research across diverse animal species (B) Signup and view all the answers

What is one of the primary goals of replacing animal testing with stem cell approaches?

To improve the predictive accuracy of toxicological assessments (A) Signup and view all the answers

Flashcards

Induced Pluripotent Stem Cells (iPSCs)

Somatic cells reprogrammed to an embryonic-like pluripotent state.

Reprogramming

The process of changing a cell's fate by altering its gene expression.