Population doublings in clinical expansion of mesenchymal stromal cells

by Alexey Bersenev on May 6, 2014 · 2 comments

in cell culture,mesenchymal

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Mesenchymal stromal cells (MSC) is becoming a major cell type in regenerative medicine clinic. There are many consideration, which should be taken in account during clinical-grade ex vivo expansion of MSCs, such us:

  • starting material/ population (type of donor tissue, processing on day 0, sorted cells or total mononuclears);
  • seeding density;
  • growth kinetics (number of passages);
  • culture media supplements (serum or alternatives, growth factors);
  • culture surface (monolayer versus 3D);
  • quality tests: phenotype, genomic stability, potency…

One of the most important parameter of clinical cell culture is population doubling level (PDL). This is a precise way to measure cell growth kinetics. PDL = number of times of two-fold increase (doubling) in the total number of cells in culture. Importance of PDL in clinical MSC culture was emphasized many times in past conferences and consensus papers. As of now, PDL data is frequently not present in the time of IND submission to FDA. In order to conclude “MSC health”, professional community is highly recommend to cell product developers to switch from passage number to PDL.

Why PDL is so important for MSC culture and why should it be used instead of passage number?

  1. MSC growth vary significantly between donors and between preparations, therefore more precise tool should be used to measure it;
  2. PDL number of MSC directly correlates with replicative senescence, which linked to loss of potency;
  3. PDL number of MSC directly correlates with genomic instability;
  4. PDL as a standard for all clinical MSC preparations will allow to compare and analyze different studies.

Based on these considerations, PDL could be a single most important predictor of MSC senescence, genomic stability and loss of potency. However, developers should remember, that exact PDL number, associated with replicative senescence could vary between different preparations. For example, MSC senescence was reported in cultures with PDL from ~10 to 40. So, you have to define optimal PDL number for your particular MSC – know your process, know your product!

Right now PDL is becoming a standard for MSC reference material. It could, and probably should become a standard for reporting of clinical MSC culture parameters. You can read more about definition of PDL here and here and use calculators here and here.

{ 2 comments… read them below or add one }

Augustine May 12, 2014 at 2:21 am

What is the difference between POPULATION DOUBLING TIME and POPULATION DOUBLING RATE?

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Priya Baraniak May 14, 2014 at 2:01 pm

Population DT is the amount of time it takes for the population to double (e.g. to go from 10 million cells to 20 million cells). The doubling rate (since cells undergo exponential growth and each mother cell divides to produce 2 daughter cells, it can be called doubling rate) or growth rate is a measure of the number of doublings that occur per a unit of time. Doubling time is calculated as: DT = {ln(2)/growth rate}. Both of these will change with cell cycle times and with cell aging/reaching senescence.

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