I am trying to culture liver organoids by encapsulating mouse primary cells in a photocrosslinked polymer-based hydrogel. Even with sufficient RGD and optimized medium composition, the cells only cluster right after seeding and show no further growth.
Has anyone successfully cultured organoids in polymer-based or photocrosslinkable hydrogels? Any tips or advice would be greatly appreciated!
From my experience I can say that it looks that crosslinking is too strong. Too much crosslinking – your cells are constrained, thus YAP signalling reduced and cells stop to divide. Try to optimize your crosslinking.
If it doesn't work share more info on cell source and polymer/cross-linking.
Thank you for the good advice! I think I should try an experiment adjusting the ratio of the crosslinker.
If your experiment went well, did you consider the gel’s optimal mechanical properties? I heard the requirements differ between polymers and Matrigel. Thank you again for your response.@Alex Mitriashkin
@Dongchang Kim
Not sure what you mean but I try to answer your question.
The thing with hydrogels is – what you are trying to model/replicate. Biological question comes first. Do you want to model normal, pathologic tissue or specific ECM composition? Do you focus on mechanical properties (stiffness) or specific ligand interactions? Do you model isotropic/anisotropic conditions? Do you model hidrostatic pressure?
First answer what you model... And remember that often simpler model with less complexity gives you better answer to your biological question. Dissect your biological problem to simple system.
If you require specific advise on your model feel free to contact me.
I sincerely appreciate your thoughtful reply. I'm currently conducting the experiment as you suggested.
My previous question was mainly focused on the mechanical properties (stiffness) of the gel or specific ligand (RGD peptide) interactions.
I asked because I found that while the storage modulus of Matrigel is usually around 100 Pa, organoids can still be cultured in polymers with a modulus ranging from 500 to 1500 Pa. @Alex Mitriashkin
Photocrosslinked hydrogels offer precise control over the 3D microenvironment for organoid culture, with key strategies focusing on material selection, mechanical properties, and biofunctionalization. Gelatin methacryloyl (GelMA) is a popular choice due to its natural ECM components, tunable stiffness, and cell-adhesive RGD motifs, making it suitable for intestinal, hepatic, and cancer organoids. Synthetic hydrogels like PEGDA or PEG-4MAL provide reproducibility and modularity but require supplementation with adhesion peptides (e.g., RGD) or ECM proteins (e.g., laminin) to support cell-matrix interactions. Hybrid systems combining GelMA with hyaluronic acid (MeHA) or collagen further enhance bioactivity while maintaining mechanical tunability.
Mechanical properties are critical, with stiffness typically adjusted between 0.5–5 kPa to mimic native tissues. This is achieved by modulating photocrosslinking parameters such as light intensity (365–405 nm), exposure time (
Alex Mitriashkin My dear friend, you are misunderstanding... I wanted to help you with this response.
Abdulhamid Dehghani me? You don't see who is topicstarter? And why you just post CharGPT answer? Q&A section is for sharing experience, not just inserting ChatGPT answers.
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