Spontaneous transformation of stem cells versus cross-contamination in culture

by Alexey Bersenev on September 6, 2012 · 0 comments

in cancer, cell culture, neural

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A year ago we’ve written about the study, which demonstrates spontaneous transformation of human fetal neural stem cells in long-term culture. It turns out that, more likely, it was not spontaneous transformation but simple cross-contamination by HeLa – cancer cell line.

The story unfolded after another group of researchers (from Norway) did some data mining experiment with published data and came to conclusion:

In the article by Wu and colleagues, they have characterized the T1 cells by DNA fingerprinting. Most interesting, the DNA fingerprint of the transformed cells (T1) did not match the “cell of origin”, and the authors explain this by genetic instability. However, we have compared the T1 fingerprint published by Wu et al., with public available cell line STR profiles, and find that the T1 STR profile published by Wu et al. is surprisingly similar to HeLa cells.

The degree of overlap in fingerprint results between published T1 line and HeLa was as high as 97%. This results allow to conclude that genetically these 2 types of cells are identical.

After this independent validation, Shen’s group went on and did the same experiment as Norwegian group. They got exactly the same results! The fact of cross-contamination was undeniable!

Intriguingly, Shen’s group tested their T1 cell line versus HeLa in in vivo model and observed that:

Two weeks later, subcutaneous xenografts were formed in HeLa cell-implanted mice (Fig. 1H). In contrast, T1 cells did not result in xenografts subcutaneously. This is consistent with our previous results we reported in the paper. On the other hand, intracranial neoplasias were found in T1 cell-implanted mice (Fig.1E) after 6 weeks.
Approximately 10 days after HeLa cells were injected into the mice brain, all the animals died, but no obvious neoplasia was found.

So, even though, fingerprinting clearly showed HeLa genome in T1, the neural stem cell line retained specific neurotropic tumorigenicity and expressed neural and stem cell markers (nestin+/ CD133+).

So, Shen’s group agrees on cross-contamination event, but also stands on possible spontaneous transformation event. I wonder how these 2 events interplay. Could spontaneous transformation even occur without HeLa contamination? Was HeLa kind of contagious for neural stem cells? Why neural stem cell genome didn’t pop up at any degree in DNA fingerprinting analysis? I also wonder what editors will decide on the fate of Shen’s original paper.

The most important lesson from this story is a necessity to check your expanded stem cells genome versus HeLa. Use DNA fingerprint analysis:

A number of scientists have pointed at the problem of cross-contamination for decades, and it is now highly recommended to authenticate cells by DNA fingerprinting [7, 9]. Several databases for checking the fingerprinted profiles are available, such as STR profile databases at ATCC (www.lgcstandards-atcc.org) and DSMZ (www2.dsmz.de). Also a list of 360 cross-contaminated cell lines is available to help researchers quality-check their work [10], and HeLa is still the most frequent cross-contaminating cell line [10]. In conclusion, it is highly questionable if the article presented by Wu et al., actually describes a transforming event of hsNSCs.

So, when we expand stem cells in culture, we have to make sure that (i) there are no spontaneous transformation events and (ii) our cells are not contaminated by HeLa (or any other cancer cell line). Injection of HeLa or other cancer cell line into patient could lead to serious unwanted complications. Seem like DNA fingerprint will answer both of these questions. I wonder if applying clinical-grade (GMP) manufacturing will help to avoid issue of cross-contamination.

This is not a new problem in cancer cell biology and cell therapy. I’d like to remind you that cancer cell biologists struggle with contamination of virtually everything by HeLa for more than 20 years. Two years ago, the initial findings for spontaneous transformation of human mesenchyma stromal cells, were refuted by 2 independent groups. The reason was exactly the same – cross-contamination by cancer cell lines.

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