cKit+ cells Do Make Heart Muscle After All

In the heart lies a population of cells that contains a protein called cKit, and are, therefore, called cKit+ cells. cKit+ cells have been the subject of a good deal of attention by researchers, but unfortunately, these have become the focus of a good deal of controversy.

When cKit+ cells were first discovered, there was a good deal of excitement about them, since they seemed to be able to make heart muscle cells and replace damaged heart muscle cells in the heart of living creatures (Beltrami AP, et al., Cell. 2003 Sep 19;114(6):763-76). In 2012, the results of a Phase I clinical trial with cKit+ cells (SCIPIO) were published (Chugh AR et al., Circulation 2012 Sep 11;126(11 Suppl 1):S54-64). This trial seemed to show that patients who received their own cKit+ cells had significant increases in heart function after a heart attack. Follow-up work in pigs even appeared to confirm that infused cKit+ cells could differentiate into heart muscle and integrate into the walls of the heart (Bolli R et al., Circulation 2013 Jul 9;128(2):122-31). So the cells were able to regenerate heart muscle in mice, pigs, and humans. It is not an understatement to say that cKit+ cells were once thought to be the key to cardiac regeneration.

The first trouble in paradise came from mouse experiments. While cKit+ cells could indeed improve the function of damaged hearts, the evidence for engraftment of the cells into the walls of the heart was wanting. Scientists in the laboratory of Jeff Molkentin Cincinnati Children’s Hospital Medical Center reported in a high-profile paper in the journal Nature that cKit+ cells can readily produce cardiac blood vessel cells, they rarely make heart muscle cells (cardiomyocytes). Because Molkentin and his team had carefully marked and traced the cells that they implanted into mice, the result was pretty devastating to the status of cKit+ cells. Molkentin’s results, however, conflicted with data from the laboratory of Bernardo Nadal-Ginard from King’s College London, who showed that heart regeneration in laboratory rodents depends on cKit+ cells and depleting cKit+ populations from the heart abolishes the ability of the heart to repair itself (Ellison GM, et al., Cell. 2013 Aug 15;154(4). Technical differences between the two papers, however, made comparisons between them difficult.

The next issues came with the SCIPIO publication itself. Two of the figures appeared to have some mistakes in them. Piero Anversa from Brigham and Women’s Hospital’s, the senior author of the SCIPIO study, admitted that there might be problems with the figures but insisted that the clinical data of the trial were sound. Other concerns about SCIPIO were expressed as well in print.  Add to that the fact that Anversa had to retract one of his earlier papers, and the whole edifice of SCIPIO and cKit+ cells seemed to totter.  These issues knocked cKit+ cells off their pedestal. At the very least, they put a hold on the SCIPIO trial until other questions had been resolved.

A new study by Joshua Hare and his group from the University of Miami Miller School of Medicine has stirred up the controversy pot once again. Hare and his team have published a paper in the journal PNAS in which they showed that cKit+ cells can readily form heart muscle cells in culture. However, apparently the cKit+ cells are finicky and only form heart muscle conditions if the conditions are just right. These results from Hare’s group might (and oh what a big might) explain why other groups have not been able to replicate the results of either Anversa or Nadal-Ginard. In Hare’s own words, “It’s not that the [cKit+] cells don’t have the capacity [to form heart muscle], but they’re entering the heart at a time that’s nonpermissive for them to become cardiac myocytes.”

In a nutshell, Hare and his team used mouse induced pluripotent stem cells (iPSCs) and differentiated them into heart muscle cells. They found that if you inhibited bone morphogenetic protein (BMP) signaling in these cells, an integral signaling event in the development of the heart; the iPSCs would express cKit and differentiate into heart muscle cells. The Hare group also used fate-mapping techniques to trace the developmental origin of cKit+ cells in the heart and they discovered that cKit+ cells are derived from the neural crest cells that delaminate from the closing neural tube during the formation of the central nervous system and migrate throughout the body to form a whole host of cell types and contribute to many different tissues.

Unlike Molkentin’s group, Hare and his crew did not observe an increased tendency for cKit+ cells to form heart blood vessel (endothelium) cells. Hare was somewhat unsure why this might be the case, but suggested that the different ways that the two teams labeled their cells for fate mapping purposes might be at least part of the issue.

Despite his success at showing that cKit+ cells can become heart muscle cells, Hare does not think that his work necessarily explains the results of the SCIPIO clinical trial, but he does think that his work might suggest how the regenerative capacities of cKit+ cells might be augmented.

Bernardo Nadal-Ginard found Hare’s work “convincing,” but added that “the paper claims the quandary and the dispute is over. But, unfortunately, it is not.” I think we can say “Amen” to that, since more work almost certainly needs to be done. Nadal-Ginard also brought up a very good point when he added that no one really knows the frequency with which cKit+ cells differentiate into heart muscle cells or other cells types or even the rate with which they replace dead or dying cells. Hare’s paper did not focus on quantitating such events, and since it did not examine the ability of cKit+ cells to repopulate a living heart, these are still questions that must be addressed.

Cornell University’s Michael Kotlikoff also made an excellent point by noting that Hare’s team did not show that cKit+ cells have the same ability to regenerate a living heart in laboratory animals as they do in culture. In an article in The Scientist by Kerry Grens, Kotlikoff said, “They never show the myogenic potential of those cells and don’t show them giving rise to cardiomyogensis” in vivo. Kotlikoff continued: “The expression of [cKit], per se, is not sufficient to identify cells as precursors and the further presumption that signaling processes observed in in vitro differentiation experiments limit such cells from undergoing myogenesis in the adult heart, the stage at which clinical regenerative efforts are focused, is not supported by data,” he added.

Hare almost certainly is either planning or is presently carrying out such experiments with laboratory mice. Presently, however, Hare has founded a company called Vestion, whose goal is to establish off-the-shelf regenerative heart therapies. According the Kerry Grens, Hare is also a part of two planned clinical trials that will administer cKit+ cells to patients with heart failure.

Piero Anversa, who remains a big fan of cKit+ cells despite their knocks, spoke approvingly of Hare’s paper and added, “To say human trials should be stopped because the experiment didn’t work in the mouse is a bit aggressive. The answer is going to be in the trial. If the trial goes well we win, if the trial doesn’t go well, we lose.”