Introduction to organoids
Organoids are three-dimensional cell cultures in which stem cells are embedded in a matrix gel. Under the influence of chemical small-molecule inhibitors/activators, cytokines, and medium additives, these cultures develop into organ-like tissue structures.
Characterization of organoids
Organoids have the ability to self-renew, maintain the physiological structure and function of the source tissue, and are known as "micro-organs in a petri dish". Using the self-renewal, differentiation and self-organization abilities of stem cells, organoids can be cryopreserved for use as biobanks and expanded indefinitely. Organoids exhibit high complexity and more closely resemble the in vivo state compared to traditional 2D cell cultures.
![Figure 1. Organoid culture of human colon adenocarcinoma cells [1]](https://cdn.shopify.com/s/files/1/0803/9419/1166/files/1_5935b867-79e5-4a23-859d-b65fcd8e4c44_1024x1024.png?v=1740728801)
Figure 1. Organoid culture of human colon adenocarcinoma cells [1]
Application of organoids
Organoids enable superior simulation of the in vivo environment, bridging the gap between animal models and cellular studies. They serve as powerful tools for tumor research, drug screening, regenerative medicine, and other fields, with applications ranging from functional tissue induction to clinical translational studies.and have been widely used in a variety of aspects of research such as functional tissue induction, disease modeling, drug screening, anti-inflammatory tests, clinical end-studies and other research, and have been used in basic research and translational applications.It has great application prospects in basic research and translational applications.
With the continuous development of organoid culture systems and experimental techniques, organoid cultures have been used for a variety of tissues and organs, including intestines(small intestine/colon), stomach, liver, heart, lungs, prostate, pancreas, kidneys, mammary glands, brain, retina, and inner ear.
Tumor stem cell-derived organoids have also begun to show great potential in helping to understand the mechanism of tumor development, screening drug sensitivity, and promoting precision medicine and personalized diagnosis. Several papers from Cell and Science have shown that tumor organoids have high sensitivity and specificity in predicting the effectiveness of anticancer drugs. Recently, tumor organoids have been shown to be useful in predicting patients' responses to cancer drugs and helping to develop personalized drug regimens.
1.Study of developmental mechanisms: the differentiation capacity possessed by organoids can be used in the study of embryonic developmental processes and their mechanisms.
Organoids regulate processes induced by signaling pathways such as Wnt and BMP, which can be used to study the development of organs such as the brain, pancreas and stomach.[2][3][4]
2.Modeling of disease damage: organoid-induced specific tissues or organs that can be used for research in specific disease models.
The team of Bing Zhao and Xinhua Lin applied a human-like organ infection model to study the molecular mechanism of SARS-CoV-2 (the novel coronavirus) infection and damage to the liver, which provides an important tool for the study of neocoronavirus pathogenesis and subsequent drug development.[5]
Deng Hongkui's research group in the School of Biological Sciences of Peking University has constructed a novel small intestinal organoid-Hyper organoid with injury regeneration characteristics in vitro by using small molecules and cytokines for stimulation. This organoid has the characteristics of being able to amplify and maintain the genome in long-term transmigration, promote damage repair of colon tissues, and alleviate pathological symptoms in animal models of acute colitis.[6]
3.Regenerative medicine: stem cell-derived organoids, which are capable of repairing or replacing damaged or diseased tissues to restore normal tissue function, have a wide range of applications in cellular therapy, including for other neurodegenerative diseases, diabetes, cardiovascular diseases, retinopathy and spinal injuries.
As a novel therapeutic modality in the field of regenerative medicine-DA01 utilizes the small molecules SB-431542 (Cat#53004ES), LDN193189 (Cat#53012ES), CHIR-99021
(Cat#53003ES), and Y-27632 (Cat#53006ES, Cat#52604ES), and Sonic Hedgehog (Shh) protein (Cat#92566ES, Cat#92589ES), to stimulate the differentiation of pluripotent stem cells into dopaminergic neurons and transplantation into the injured areas of the brains of patients with advanced Parkinson's disease, to provide new directions and ideas for the treatment of the disease.[7]
4.Drug toxicity and efficacy testing: use of organoids to verify the pharmacokinetics toxicity of new drugs in specific organs or tissues, providing data support for new drug development.
The nephrotoxicity of Cisplatin (cisplatin) was verified using the Hyman renal organoid.[8]
5.Drug screening: stem cell-derived organoids can be used for in vitro testing of drug responses, providing theoretical support for drug screening
Colonic organs can be used to study dosing regimens in patients with CFTR mutations, and tumor organs can be used to assess individualized dosing in patients.[9]
The evolution of organoids
Sources of organoids
Normal organoids are mainly derived from stem cells, which include pluripotent stem cells (PSCs) and adult stem cells (ASCs), of which pluripotent stem cells include embryonic stem cells (ESCs), induced pluripotent stem cells ( Induced pluripotent stem cells (iPSCs). Compared with pluripotent stem cells, adult stem cells have the advantage of easy and fast modeling, but the disadvantage that the constructed organoid structures are relatively simple. The organoid structures constructed by pluripotent stem cells are more complex.
Organoid culture
|
Organoid Type |
Required Small Molecule |
Required Growth Factors |
Media and Additives |
|
Small intestine |
Y-27632, SB-202190, A 83-01, Gastrin, Nicotinamide |
EGF, Noggin, R-Spondin 1, Wnt-3a |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin, N-2 |
|
Gastric |
Y-27632, SB-202190, A 83-01, Gastrin I, Nicotinamide |
FGF-10, EGF, Noggin, R-Spondin 1, Wnt-3a |
HEPES, L-alanyl-L- glutamine solution, penicillin- streptomycin |
|
Liver disease |
Y-27632, A 83-01, DAPT, Forskolin, Gastrin, Nicotinamide, Prostaglandin E2 |
BMP-4, EGF, FGF-basic, FGF-10, HGF, Noggin, Wnt-3a |
N-2, HEPES, L-alanyl-L-glutamine solution, penicillin- streptomycin |
|
kidney |
CHIR-99021、Retinoic Acid |
BMP-2、BMP-4、BMP-7、FGF- basic 、FGF-9 |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Lungs |
CHIR-99021、SB-431542 |
Activin A、FGF- basic、FGF-4、Noggin |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Pancreas |
Gastrin I、A 83-01、Nicotinamide |
FGF-10、 EGF、Noggin、R-Spondin 1、Wnt-3a |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Prostate |
Y-27632、SB-202190、A 83-01、Nicotinamide、Prostaglandin E2、Testosterone |
EGF、Activin A、FGF- basic、FGF-10、Noggin、R-Spond in 1、Wnt-10b |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Mammary gland |
Y-27632 |
Here gulin β-1、R-Spondin 1、R-Spondin 2、Noggin、EGF、 FGF- basic、FGF-10、Wnt-3a、Prolactin |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Retina |
CHIR-99021、Y-27632 |
SHH、Wnt-3a |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Inner ear |
SB-431542、A 83-01 |
BMP-4、 FGF- basic |
HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
|
Brain |
Y-27632、MK-2206、GDC-0068、Dorsomorphin |
FGF- basic、Noggin、DKK-1、 EGF、BDNF、GDNF |
B-27, HEPES, L-alanyl-L- glutamine solution, penicillin-streptomycin |
Summary of small molecules commonly used in organoid culture: super useful, don't forget to bookmark it!
❶ Y-27632 (Cat#53006ES, Cat#52604ES): It is a potent inhibitor of Rock, which inhibits p160ROCK (Ki=140 nM) and ROCK-II (IC50=800 nM) competitively with ATP, and also inhibits PRK2 (IC50= 600 nM), and it is usually added at the first time of culture in the seed plate, but not in subsequent culture. Y-27632 (10 µM) can inhibit stem cell apoptosis, improve cloning efficiency and prolong cell progeny by treating human embryonic stem cells with Y-27632 (10 µM) for 1 h.
The recommended working concentration is 10 μM
❷ SB-202190 (Cat#53005ES): A potent p38 MAPK kinase inhibitor targeting p38α/β. SB202190 induces the differentiation of human embryonic stem cells to cardiomyocytes, promotes the self-renewal of neural stem cells, and can be used in gastrointestinal and mammary organoid cultures.
Recommended dissolution concentration: 10 mg dissolved to 10 mM by adding 3.018 mL of DMSO solution and stored at -20°C in portions.
The recommended working concentration is 10 μM
❸ CHIR-99021 (Cat# 53003ES): An aminopyrimidine derivative that acts as an inhibitor of GSK-3 (GSK3α/β), which induces the differentiation of human embryonic stem cells towards the endoderm, and is used in renal and retinal organoid cultures. CHIR-99021, in combination with other reagents, stimulates the reprogramming of somatic cells into stem cells.
Recommended Dissolution Concentration: 5 mg dissolved to 3 mM by adding 3.58 mL of DMSO solution and stored at -20°C in portions.
The recommended working concentration: 3 μM
❹ A 83-01 (Cat#53002ES): is an Activin/NODAL/TGF-β pathway inhibitor that inhibits ALK5/4/7 kinase activity and is generally used in liver, prostate, and mammary organoid cultures, and is often used to inhibit the differentiation of iPSCs and to maintain self-renewal of cells in vitro. Recommended dissolution concentration: 5 mg added to 5.93 mL of DMSO solution to dissolve to 2 mM, -20 ℃ divided and stored. (Special Note: This product is unstable in solution state, it is recommended to use it now.)
The recommended working concentration: 2 μM
❺ Gastrin I (Cat#53007ES): Gastrin is an endogenous gastrointestinal peptide hormone that stimulates the secretion of gastric acid from gastric lining cells. Gastrin binds to the cholecystokinin B receptor (CCKBR), which increases intracellular Ca2+, promotes the production of phosphatidylinositol, and protein kinase C activation. Gastrin is also involved in the proliferation and differentiation of gastric epithelial cells and is used in the study of gastrointestinal-like organs. The addition of gastrin is required to prolong the survival of intestinal and liver organoids in culture.
Recommended dissolution concentration: 1 mg dissolved to 0.2 mM by adding 2.38 mL of 1% ammonia solution and stored at -20°C in portions.
The recommended working concentration: 10 nM
❻ Nicotinamide Nicotinamide (Cat# 51402ES): A B3 vitamin involved in a variety of enzymatic redox reactions and is used in gastrointestinal, liver, and mammary organoid cultures.
Nicotinamide is used in conjunction with cytokines and other biochemicals to exert anti- inflammatory properties and to promote the differentiation of MSCs into insulin-producing cells, to inhibit the activity of sirtuins, and to promote the formation of organoids and prolong organoid lifespan.
Recommended dissolution concentration: 100 mg dissolved to 100 mM by adding 8.19 mL of H2O (or DMSO) and stored at -20°C in portions.
The recommended working concentration: 10 mM
❼ Forskolin trichothecene (Cat#51001ES): Activates adenylyl cyclase and is commonly used to increase intracellular cAMP levels.Forskolin induces differentiation of many cell types and activates PXR and FXR.It has platelet anticoagulant and antihypertensive effects and, when combined with other small molecules, can induce fibroblast reprogramming to iPSCs.This substance must be added to liver organoid cultures. reprogrammed into iPSCs. must be added during liver-like organ culture. Recommended working concentration: 1-10 μM
❽ Prostaglandin E2 (Cat#60810ES): Prostaglandin E2 (PGE2) regulates many physiological systems, mediates cell proliferation and differentiation upon binding to specific receptors, is required to be added to liver-like and prostate-like organ cultures, and has been implicated in smooth muscle sparing, inflammation, fertility, sleep cycle regulation and gastric mucosal integrity.
Recommended dissolution concentration: 1 mg added to 0.28 mL of DMSO solution dissolved to 10 mM and stored at -20°C in portions.
Recommended working concentration: 500 nM
❾ N-acetyl-L-Cysteine (Cat#50303ES): N-acetyl-L-cysteine (NAC) is a precursor to the antioxidant glutathione, which has antioxidant properties, and a ROS inhibitor that inhibits apoptosis in neuronal cells, and which is required to be added in most organoid cultures.
Recommended dissolution concentration: 2 g added to 24.51 mL of H2O (or DMSO) dissolved to 500 mM and stored at -20°C in portions.
Recommended working concentration: 1 mM
Product Recommendation
Small molecule compounds (Inhibitors/Activators)
|
Product name |
Catalog number |
Specification |
|
Retinoic acid (retinoic acid, retinoic acid) |
100 mg/500 mg/1 g |
|
|
A 83-01 |
1 mg/5 mg/10 mg |
|
|
CHIR-99021 |
2 mg/5 mg/10 mg |
|
|
SB-431542 |
5 mg/10 mg/50 mg |
|
|
SB-202190 |
5 mg/10 mg/25 mg |
|
|
Y-27632 |
1 mg/5 mg/10 mg |
|
|
Gastrin I (human) |
1 mg/5 mg |
|
|
MK-2206 2HCl |
1 mg/5 mg/10 mg |
|
|
GDC-0068 (Ipatasertib; RG7440; GDC0068) |
53009ES |
1 mg/5 mg/10 mg |
|
(Dorsomorphin 2HCl) |
53010ES |
1 mg/5 mg/10 mg |
|
Y-27632 dihydrochloride |
5 mg/10 mg/25 mg |
|
|
Nicotinamide |
1 g/5 g |
|
|
DAPT (GSI-IX, LY-374973) |
5 mg/25 mg |
|
|
Forskolin Hair Throat |
10 mg/50 mg/100 mg/1 g |
|
|
Prostaglandin (PG) E2 Prostaglandin E2 |
60810ES |
1 mg |
|
Testosterone |
60803ES |
1 g/5 g |
|
NAC (N-acetyl-L-cysteine) |
50303ES |
2 g |
|
RepSox |
5 mg/10 mg/25 mg |
|
|
LDN193189 (DM-3189) |
53012ES |
5 mg/10 mg |
Media and supplements
|
Catalog number |
Specification |
|
|
HEPES Free Acid Cell Culture Grade |
60110ES |
100 g/500 g |
|
L-alanyl-L-glutamine solution, 200 mM L-alanyl-L-glutamine, 200 mM |
60701ES |
20 mL/100 mL/500 mL |
|
Penicillin-Streptomycin (100×), Suitable for Cell Cu lture Penicillin-Streptomycin (Dual Antibody) for C ell Culture |
100 mL |
|
|
B-27 serum-free medium, 50 x |
60703ES |
10 mL |
|
N-2 supplement, serum free, 100X N-2 serum free additive, 100× |
60706ES |
5 mL |
Cytokine
|
Product name |
Catalog number |
Specification |
|
Recombinant Human Activin A |
10 μg/100 μg/500 μg |
|
|
10 μg/100 μg/500 μg |
||
|
Recombinant Human DKK-1 |
20 μg/100 μg/500 μg |
|
|
Recombinant Human EGF |
100 μg/500 μg |
|
|
Recombinant Human bFGF |
10 μg/100 μg/500 μg |
|
|
Recombinant Human FGF-4 |
5 μg/100 μg/500 μg |
|
|
Recombinant Human FGF-9 |
91305ES |
5 μg/100 μg/500 μg |
|
Recombinant Human FGF-10 |
91306ES |
5 μg/100 μg/500 μg |
|
Recombinant Human GDNF |
10 μg/100 μg/500 μg |
|
|
Recombinant Human Noggin |
5 μg/100 μg/500 μg |
|
|
Recombinant Human NRG1-beta1 |
10 μg/100 μg/500 μg |
|
|
Recombinant Human R-Spondin 1 |
92274ES |
100 μg/1 mg |
|
Recombinant Human Shh |
5 μg/100 μg/500 μg |
|
|
Recombinant Human BAFF |
5 μg/100 μg/500 μg |
|
|
Recombinant Human sCD40 Ligand |
10 μg/100 μg/500 μg |
|
|
Recombinant Human BMP-4 |
10 μg/100 μg/500 μg |
|
|
Recombinant Human HGF |
5 μg/100 μg/500 μg |
|
|
Recombinant Human BDNF |
5 μg/20 μg/100 μg |
Mini-classroom: Small molecule compounds are compounds with molecular weight less than 1000 Da (especially less than 500 Da) and have biological activity. Unlike cytokines and proteins, small molecules can enter the cell through the cell membrane and fulfill their biological functions. Small molecule compounds have a wide range of targets and have been widely used in many important research fields in life sciences, such as stem cells, organoids, immunology, neurobiology, epigenetics, apoptosis, ion channels, oncology and signal transduction.
Yeasen small molecule product line covers a full range and variety of products, with high purity, strict quality inspection, quality assurance, professional pre- sale and after-sale technical team, and excellent price and high price-performance ratio of the products. We insist on serving our customers with high quality products to help your scientific research!
References
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[2] Lancaster MA, Renner M, et al. Cerebral organoids model human brain development and microcephaly. Nature. 2013;501(7467):373-379. doi:10.1038/nature12517.
[3] Greggio C, et al. Artificial three-dimensional niches deconstruct pancreas development in vitro. Development. 2013.140(21):4452-4462. http://dx.doi.org/10.1242/dev.096628.
[4] McCracken KW, et al. Modelling human development and disease in pluripotent stem-cell-derived gastric organoids. nature. 2014.516(7531):400- 404. http://dx.doi.org/10.1038/nature13863.
[5] Zhao B, Ni C, et al. Recapitulation of SARS-CoV-2 infection and cholangiocyte damage with human liver ductal organoids. Protein Cell. 2020 Oct;11(10 ):771-775. doi: 10.1007/s13238-020-00718-6. PMID: 32303993; PMCID: PMC7164704.
[6] Qu M, Xiong L, et al. Establishment of intestinal organoid cultures modeling injury-associated epithelial regeneration. Cell Res. 2021 Mar;31(3). 259-271. doi: 10.1038/s41422-020-00453-x. Epub 2021 Jan 8. PMID:
[7] 33420425; PMCID: PMC8027647.
[8] BlueRock Therapeutics Announces First Patient Dosed with DA01 in Phase 1 Study in Patients with Advanced Parkinson's Disease. BlueRock Therapeutics Press Release: June 8, 2021.
[9] Takasato M, Er PX, et al. Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis. nature. 2015.526(7574):564 -568. http://dx.doi.org/10.1038/nature15695.
[10] Spence JR, Mayhew CN, et al. Directed differentiation of human pluripotent stem cells into intestinal tissue in vitro. Nature. 2011.470(7332):105- 109. http://dx.doi.org/10.1038/nature09691.