Lesson 6 Review Organoids

Applied Molecular Cellular Biology

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🌟 What Are Organoids?

Organoids = tiny, 3D “mini-organs” grown in the lab.
Built from stem cells that self-organize into tissue-like structures.
Two main flavors:
1. PSC-derived (pluripotent stem cells): mimic embryonic development. Good for studying how organs form 🧠🫁.
2. ASC-derived (adult stem cells): mimic tissue repair. Great for modeling adult organs and diseases 🧑‍⚕️.

🔬 How Do They Compare to Older Models?

2D cell lines: cheap and simple, but often mutated, lack diversity, and don’t mimic real tissue well.
Patient-derived xenografts (PDXs): human tumors transplanted into mice. Better than 2D, but slow, costly, and tricky.
Organoids: faster, more faithful, expandable, and usable from both healthy and diseased tissue. ✅

🧩 Building Organoids

Need extracellular matrix support (often Matrigel).
Use specific growth factor cocktails (e.g., Wnt, R-spondin, EGF, Noggin).
PSC-organoids = long protocols (months), complex tissues.
ASC-organoids = faster (days to weeks), stable, expandable long term.

🏥 Applications of Organoids

1. Studying Physiology 🧬

Organoids allow culture of previously impossible cell types (e.g., hepatocytes that make albumin + detox enzymes).
Reveal mechanisms like BMP gradients controlling gut hormone expression.

2. Modeling Diseases 🦠🧑‍⚕️

Infectious disease:
- Cryptosporidium life cycle in gut organoids 💩
- Helicobacter pylori in stomach organoids 🍲
- Norovirus replication only works in intestinal organoids 🚽
- RSV infection in airway organoids → syncytia + neutrophil attraction 🫁
- Influenza strains tested for infectivity 🦠
Genetic diseases:
- Cystic fibrosis → organoids swell in forskolin assay only if CFTR works. Used to test drugs + CRISPR repair 💨
- Alagille syndrome, α1-antitrypsin deficiency, microvillus inclusion disease, etc. modeled.
Cancer:
- Tumor-derived organoids capture heterogeneity 🎭
- CRISPR-engineered normal organoids can model stepwise mutations (e.g., APC, KRAS, TP53).
- Used to study metastasis, drug resistance, DNA repair failures.

📦 Organoid Biobanks

Living libraries of organoids from patients.
Cover colon, pancreas, liver, prostate, breast, ovary, bladder, etc.
Store both tumor and matched normal tissue → great for drug testing and biology.
Example: breast cancer biobank retained hormone receptor status (ER/PR/HER2).
Drug responses in organoids often mirror patient responses (e.g., pancreatic, ovarian, GI cancers). 🔑 for personalized therapy.

🧑‍⚕️ Organoids in Medicine

Genetic Repair

CRISPR correction in cystic fibrosis organoids restored function 🛠️.

Transplantation

Mouse studies show transplanted colon or liver organoids can integrate and restore function (colitis, liver failure) 🐭➡️❤️.

Immunotherapy

Organoids cocultured with immune cells help study tumor–immune interactions ⚔️.
Tumor organoids used to activate patient T cells → potential personalized immunotherapy.

Cystic Fibrosis in the Netherlands

First clinical use: forskolin swelling assay determines which drugs work for each patient.
Therapy choices (like Kalydeco, Orkambi) now guided by organoid response instead of just genotype. 🚀

⚠️ Limitations

Dependence on animal-based matrices (Matrigel) 🐭.
High cost 💰.
Lack of full tissue complexity (no blood vessels, immune cells, or nerves).
Some cancers/tissues still hard to grow.

🔮 Future Perspectives

Expand to hard-to-culture cancers (sarcomas, melanomas).
Develop synthetic, defined culture systems.
Use for rare genetic diseases with no large clinical trials.
Scale up for regenerative medicine and personalized drug screening. 🌍

✨ Key Takeaways

Organoids are 3D mini-organs from PSCs (development) or ASCs (repair).
They’re genetically stable, expandable, and versatile.
Useful for modeling infections, genetic diseases, cancers, and physiology.
Biobanks + CRISPR + drug screens make them powerful for personalized medicine.
Already clinically applied in cystic fibrosis treatment decisions.
Challenges: cost, lack of non-epithelial components, reliance on animal matrices.

Quiz

Score: 0/30 (0%)

Q0. What defines an organoid in biological research?

A 2D culture derived from fibroblasts

A 3D structure of organ-specific cell types grown from stem cells

A xenotransplant of tumor cells into mice

A colony of bacteria forming tissue-like layers

Q1. What is the main role of laminin in organoid culture?

Acts as an energy source

Triggers apoptosis in differentiated cells

Forms part of the extracellular matrix to support cell attachment and organization

Regulates transcription of stem cell genes

Q2. Which growth factors are essential for maintaining intestinal organoids in culture?

EGF, R-spondin, and Noggin

BMP4, VEGF, and FGF

TGF-β, PDGF, and SHH

IL-6, IL-8, and TNF-α

Q3. Which statement best distinguishes PSC-derived from ASC-derived organoids?

PSC-derived organoids model adult tissue repair

ASC-derived organoids require differentiation cocktails mimicking embryogenesis

PSC-derived organoids recapitulate development; ASC-derived model adult regeneration

ASC-derived organoids contain mesenchymal and endothelial cells

Q4. Why are adult stem cell (ASC)-derived organoids limited to epithelial tissues?

Epithelial tissues have immune privilege

Only epithelial tissues contain stem cells with regenerative capacity

Other tissues lack transcriptional activation

They require neural and vascular inputs to grow

Q5. What role does the Wnt signaling pathway play in intestinal organoid cultures?

Promotes cell differentiation

Suppresses apoptosis in differentiated cells

Maintains stemness and proliferation of Lgr5+ stem cells

Inhibits BMP signaling during apoptosis

Q6. What does the Forskolin-Induced Swelling assay measure in cystic fibrosis organoids?

Cell division rate

CFTR-dependent chloride and water transport

Calcium signaling activity

Apoptotic response to DNA damage

Q7. What is meant by 'isogenic controls' in CRISPR-modified organoid studies?

Controls derived from different species

Cultures genetically identical except for one specific mutation

Organoids created from distinct patients

Cultures exposed to varying growth media

Q8. Which infectious agent was modeled in kidney tubuloids to mimic transplant-related nephropathy?

Hepatitis B virus

BK virus

Zika virus

Influenza virus

Q9. In the study, what does 'loss of niche factor dependency' indicate in tumor organoids?

Tumor cells can grow without external stem-cell supporting signals

Tumor cells require increased growth factors to survive

Tumor cells undergo apoptosis under stress

Tumor cells become senescent and non-dividing

Q10. What is a biobank in the context of organoid research?

A chemical storage for antibiotics

A collection of patient data records

A living repository of organoid lines derived from patient tissues

A computational model for organoid simulation

Q11. How do PARP inhibitors selectively kill BRCA-deficient tumor cells?

By inducing oxidative stress

By blocking single-strand repair, causing double-strand breaks that BRCA-mutant cells cannot fix

By activating p53 signaling

By increasing telomere length

Q12. What does 'xenotransplantation' refer to in organoid experiments?

Transplanting organoids between humans

Transplanting organoids into a different species, often mice

Transferring organoids into artificial matrices

Transplanting tissues into 2D culture plates

Q13. Why are animal-based matrices like Matrigel used in organoid culture?

They provide hormones that trigger differentiation

They serve as a scaffold mimicking the extracellular matrix

They supply nutrients to immune cells

They prevent microbial contamination

Q14. What are the main limitations of ASC-derived organoids?

Uncontrolled genetic instability and high tumorigenicity

Lack of non-epithelial components, use of animal matrix, and high cost

Inability to model infectious diseases

Short culture lifespan

Q15. Pluripotent stem cell–derived organoids are best suited for studying organ development.

True

False

Q16. Adult stem cell–derived organoids include stromal and vascular components by default.

True

False

Q17. Organoids can be derived from both healthy and diseased tissue.

True

False

Q18. The Forskolin-induced swelling assay directly measures gene expression changes.

True

False

Q19. Helicobacter pylori infection in stomach organoids activates inflammatory signaling similar to that seen in vivo.

True

False

Q20. Loss of TP53 in tumor organoids increases sensitivity to Nutlin-3.

True

False

Q21. Organoids derived from metastatic tumors generally retain the genetic profile of the original tumor.

True

False

Q22. Wnt proteins are secreted signaling molecules that regulate stem cell proliferation.

True

False

Q23. Basal-type pancreatic tumors are more dependent on Wnt signaling than classical-type tumors.

True

False

Q24. The term 'carcinoma' refers to cancers derived from epithelial cells.

True

False

Q25. Organoid biobanks are used solely for basic research and not for drug testing.

True

False

Q26. Engraftment refers to successful attachment and functional integration of transplanted organoids.

True

False

Q27. PARP inhibitors enhance the DNA repair capacity of cancer cells.

True

False

Q28. Ethical discussions around organoid biobanking include ownership and consent issues.

True

False

Q29. Loss of niche factor dependency generally reduces a tumor’s metastatic potential.

True

False