Part 2 of a 3 part series on the recent JDRF/University of Geneva study of α-to-β-cell conversion in mice. Read Part 1 and Part 3.
So what do the researchers do with a bunch of diphtheria-infused, pseudo-diabetic mice? Watch and wait.
Immediately after the diphtheria injection, more than 99% of the mice’s β-cells had been ablated (killed off). In other words, there may have been an occasional, stray β-cell left behind, but nothing that would allow the mouse pancreas to create enough insulin to maintain glycemic control.
Fifteen days after the injection, though, the researchers euthanized a subset of the mice and measured the β-cell mass in the pancreas; they found three times as many β-cells as immediately after injection, and ten times as much creation of insulin. In other words, the mice were regenerating β-cells and regaining the ability to manufacture insulin on their own.
The researchers continued to measure and monitor β-cell mass for up to ten months after killing off the initial set of β-cells. They kept the mice alive by injecting insulin when necessary.
But how cool is this: after six months, none of the mice needed insulin injections anymore. All of the mice had experienced β-cell regeneration, and the scientists found ten- to forty-four times as many β-cells in the pancreatic islands, meaning some mice were creating enough insulin to be comparable to recent-onset Type 1 diabetics, and others were actually creating enough insulin to have normal blood glucose levels, without any insulin treatment.
Yes, really. The question is, how? And how could the researchers know?
One hypothesis as to where the new β-cells were coming from would be that the left-over β-cells were replicating. β-cells, to my chagrin as a diabetic, are known for being slow to reproduce, and the normal pancreas functions on the assumption that β-cells are long-lasting cells that don’t die often. Perhaps, though, in the mice in this experiment, β-cells were replicating more to compensate for their ablated brethren.
To test this hypothesis, the researchers created a strain of the diphtheria-toxin-receptive mice that only had diphtheria receptors in half their β-cells. The researchers also tagged the mice’s existing β-cells, modifying the genetic makeup such that when the mice were injected with tamoxifen, a yellow fluorescent protein in the β-cells reacted. As a result, when diphtheria was injected, only half of the β-cells in the pancreas were killed off, and the remaining half were traceable as the original β-cells.
Immediately after the diphtheria injection, all the β-cells were marked by the yellow fluorescent protein; fifteen days after, 80% were; and thirty days after, only 7.6% were. What does this mean? Firstly, that β-cell regeneration happens fairly rapidly after ablation. Secondly, that the left-over β-cells are likely not responsible for the new cells– if they were replicating, the proportion of β-cells with the yellow fluorescent protein would stay the same or increase. The fact that the proportion of β-cells with the yellow fluorescent protein dropped off so rapidly implies that the vast majority of the new cells were coming from some other source.
But Where did the New β-Cells Come from?
One notable aspect of the new cells that appeared shortly after the diphtheria injection was that they were not β-cells at first; they were bihormonal cells, that both produced glucagon like α-cells and produced insulin like β-cells. These cells had some of the genetic markers of β-cells, and eventually some stopped producing glucagon and became full β-cells, producing only insulin. None of these bihormonal cells carried the yellow fluorescent protein when the original β-cells were marked with it, so the researchers looked for a different source for these cells. Perhaps they came from α-cells, or some other precursor cells that learn new tricks when the original β-cells die off.
To determine where the glucagon/insulin cells were coming from, the researchers tagged the α-cells in the pancreas with a yellow fluorescent protein. When they did so, they found that, indeed, the bihormonal cells that appeared after killing the β-cells were also tagged with the yellow fluorescent protein, implying they were from the line of tagged α-cells. Also, when the researchers tried killing α-cells and β-cells simultaneously, they found that no bihormonal cells appeared, implying that it was not another, third type of cell that was responsible for the new glucagon/insulin cells. It was the α-cells that were being reprogrammed and retrained to produce insulin.