Hi,

Generally, it is not more difficult than reprogramming PBMCs from young people. However, reprogramming PBMCs is more difficult than reprogramming fibroblasts. Reprogramming fibroblasts is definitely more efficient. Sometimes differentiation of PBMCs requires the use of a few additional "tricks". A lot also depends on the efficiency of the method of reprogramming chosen by you.

Best regards,

Dr Justyna Augustyniak

Dear Dr. Rituparna Chaudhuri

Yes, it is true. PBMCs form older individuals may exhibit slower proliferation rates and a tendency to form clumps in culture. It is the effect of aging that can impact the cellular environment and the ability of the cells to respond to reprogramming signals. With age, cells undergo epigenetic modifications and experience decreased cellular plasticity, making them less amenable to reprogramming. You may overcome these hurdles by optimizing culture conditions or using specific growth factors.

You may supplement the culture media with vitamin C that can enhance the activity of Tet enzymes, which play a vital role in DNA demethylation and epigenetic remodeling during reprogramming. This can help overcome the epigenetic memory and promote a more permissive epigenetic landscape for pluripotency.

You may also add Rho kinase (ROCK) inhibitor like Y-27632 in culture medium which can improve the survival and proliferation of iPSCs, particularly during the initial stages of reprogramming when cells are fragile.

Supplementation with compounds like nicotinamide riboside can increase NAD+ levels, potentially influencing cellular health and reprogramming.

The below attached article may help.

Article Influence of donor age on induced pluripotent stem cells

Hope the above suggestions help!

Regards,

Malcolm Nobre

Thank you so much. I will see if any media additions can help with the growth of colonies.