Unexplained abnormalities in stem cells prompt Columbia researchers to pull diabetes paper

Researchers at Columbia University have retracted a 2013 paper in The Journal of Clinical Investigation, after uncovering abnormalities in the stem cell lines that undermined the conclusions in the paper.

Last year, corresponding author Dieter Egli discovered he could not reproduce key data in the 2013 paper because almost all the cell lines first author Haiqing Hua used contained abnormalities, casting doubt on the overall findings. When Egli reached out to Hua for answers, Hua could not explain the abnormalities. As a result, Hua and Egli agreed the paper should be retracted.

Since some of the details of how the paper ended up relying on abnormal cells remain unclear, the university confirmed to us that it is investigating the matter.

Here’s the retraction notice for “iPSC-derived β cells model diabetes due to glucokinase deficiency,” cited 42 times:

The corresponding authors were made aware of karyotype abnormalities through a routine quality control test of pluripotent stem cells used in the studies reported in this paper. After extensive internal review and genetic analysis, they found that the karyotypes of some of the cells used for the experiments reported were abnormal and that the normal karyotypes shown in Figure 1 and Supplemental Figure 2 were not from cell lines used in the study. They also cannot confirm the endonuclease-mediated correction of the mutant GCK G299R allele. H. Hua takes responsibility for the characterization and presentation of cell line karyotypes and the genetic manipulations. Because of these discrepancies, the authors wish to retract the article. They apologize for these errors and for any inconvenience caused to others.

In the fall of 2009, Hua joined Rudolph Leibel’s diabetes and nutrition lab at Columbia University, under the co-supervision of Egli, who brought an expertise in stem cell biology. Hua told us:

The aim of my research project was to leverage the expertise of both Dr. Egli (on stem cell biology) and Dr. Leibel (on diabetes) and to demonstrate the concept that the islet cells generated in the lab from diabetic patients through stem cell technology would present comparable dysfunction as the islet cells in the patients’ body. Because we chose patients with genetic mutations that cause diabetes, we were hoping to demonstrate that correction of the mutations would restore the normal function of the islet cells.

But, Hua noted, he wasn’t — and still isn’t — an expert in stem cell biology, so he had to learn on the job:

When I began the project, I never worked with cells before and had no experience or understanding of cell line karyotype.

Hua started by generating several cell lines from a diabetic patient. To check that the genetic makeup of these cell lines were the same, he sent several for analysis to a contracted service, which examines 20 cells per cell line and generates a report:

I did karyotype analysis for the cell lines right after I derived them, probably in 2011, before I started to do any experiments on them. The reports came back with some cells being normal and some being abnormal. To be fair, I thought what I learned from Dr. Egli was that it is a normal phenomenon that some cells are abnormal as long as the number is not high.

Indeed, Egli, an assistant professor of stem cell biology at Columbia University Medical Center, confirmed that pluripotent stem cells are often prone to undergo abnormalities:

Karyotypic abnormalities are common, and occur in many cells upon extended cultures, so this is not in and of itself a concern. Often one can go back to earlier cultures that are normal.

Hua published the work in 2013, along with a related paper in Diabetes in 2014, “β-Cell Dysfunction Due to Increased ER Stress in a Stem Cell Model of Wolfram Syndrome.” Hua believes, at a conceptual level, both papers achieved the goal of demonstrating that the “correction of the mutations would restore the normal function of the islet cells.”

In 2014, Hua told us that he moved back to China for “family reasons.”

Last year, other investigators asked Egli to share the cells lines from the 2013 study. To ensure he was providing high quality material, Egli sent what he believed to be normal cell lines from the study for quality control testing. Egli said that’s when he learned many of the cell lines contained abnormalities.

To suss out the problem, Egli went back to the cell lines stored in the lab to look for normal cells:

Dr. Hua had already left the University at that time and so I personally started to look for karyotypically normal cells. There were no normal cells to be found.

Egli explained what the abnormalities meant for the study results:

You could best describe the abnormalities of the [cell] lines [Hua] used as ‘mumbo-jumbo.’ There were multiple rearrangements in the chromosomes in the cell lines and thus you wouldn’t know if the effects you saw were due to gene modifications or simply due to those rearrangements. Essentially, the abnormal cell lines question the entire paper, and it’s very unlikely the paper would have been accepted at the journal.

When Egli failed to reproduce the data from the 2013 paper, he contacted Hua to find out where the normal cell lines were. But Hua was not sure — in fact, he told us it was a “surprise” to learn that most of the cell lines he had used contained abnormalities, adding:

…another layer of complication is that when cells became karyotype abnormal, they could behave like cancer cells, namely they could start as minor portion in the culture but later on took over and became majority. So another mistake we made was that we didn’t perform karyotype analysis at the end of the study to make sure that after all the experiments we did, the cells were still normal.

A spokesperson at Columbia University verified that the university is conducting an investigation into the issues:

I could confirm that there is an ongoing investigation.

When Hua was informed of these issues, he suggested the study be retracted:

Immediately, I proposed to Dr. Egli and Dr. Leibel that we should retract the publication because we were not certain about the conclusion any more.

Hua takes responsibility for what happened, adding:

So this was done at very early phase of my research, and I was busy with a lot of parallel projects since I was the first post-doc of Dr. Egli…Because I wasn’t understanding the problem correctly, I put up the figures with normal karyotype as first figure for the publication and continued my research with one particular cell line.

Egli also talked about the experience of retracting a paper:

Retracting a paper is not a rewarding process, and often reports stay in the literature even if they should not. Retracting the paper exposes us to the possibility of damage. I took proactive steps to investigate and retract because I wanted to correct the record. This would not have happened without my initiative involving 2-3 months of benchwork.

Hua described this as a “truly unfortunate and painful” chapter, which he hopes others can learn from:

The health of academic world and advance of science really depends on correction of previous mistakes and clearance of uncertainties. … [A]voiding overwhelming multitasking is important. At the first year of my research, I was setting the lab together with Dr. Egli and meanwhile performed more than 100 experiments. Each of them would took more than 10 days and I was really stacking all the experiments. This particular project was about one fourth of my effort at that time. … My biggest recommendation or reflection would be that it is very very very important to quality control and characterize starting materials of a project. Many people, including myself, are more focused on rushing the project forward and do not realized that if the starting materials are flawed, anything built on them has no solid foundation.

Hat tip: Rolf Degen

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