The timing of autophagy activation in humans during fasting remains unknown, and the widely cited “24–72 hours” likely comes from mouse experiments, review authors write

The timing of autophagy activation in humans during fasting remains unknown, and the widely cited “24–72 hours” likely comes from mouse experiments, review authors write

A review of how fasting and diets affect autophagy, the process by which cells break down worn-out components, was published on October 13 in Biochemical Society Transactions. Sanjna Singh and Timothy Sargent of the University of Adelaide compiled a table of 20 human studies and compared their findings with animal experiments.

In animals, several diets activate autophagy and are associated with longer lifespans. Videos about fasting give specific times for its activation. Mindy Pelz places the peak at 48–72 hours and acknowledges: “I cannot find the research on this 48-hour peak in autophagy, but I find many experts talking about it.”

The review authors suggest that the idea that 24–72 hours without food activates autophagy in humans most likely originated in mouse experiments. The laboratory of Yoshinori Ohsumi, who received the 2016 Nobel Prize for discovering the mechanisms of autophagy, developed mice with a fluorescent label on the vesicles into which cells package waste. After 24 and 48 hours without food, these vesicles became more numerous in many organs, with the exception of the brain. According to the review, however, mice begin to die after three days without food, whereas humans can survive at least three weeks, as demonstrated by 13 volunteers who fasted for that long. Levels of FGF21, a hormone released during fasting, rise in mice after 6 hours, whereas in humans they rise substantially only by days seven to ten. The authors conclude that the timing observed in mice cannot be applied directly to humans.

Even in animals, the outcome depends on the duration of fasting. In fruit flies, fasting for 20 hours every other day extended male lifespan by 13% and female lifespan by more than 18%, and only when autophagy genes were functional. Fasting for 24 hours shortened lifespan.

Testing this in humans is complicated by how autophagy is measured. Most studies measured autophagy proteins and the activity of their genes in blood, muscle or fat cells at the time of sampling. LC3B-II, a protein on the membranes of these vesicles, accumulates both when waste clearance accelerates and when its final stage stalls, so its level alone cannot distinguish between the two. To measure the rate of clearance, called autophagic flux, researchers divide fresh blood into two samples and add an inhibitor of lysosomes, the cellular compartments that digest waste, to one sample. The difference in LC3B-II accumulation indicates how much waste the cells would have broken down during that period. Blood is the only tissue that can be collected intact and kept alive outside the body, so flux is measured in blood. Yet in aging male mice, flux increased in blood while decreasing in the heart.

Of the six studies examining fasting for 24–72 hours, one measured flux. It found no change after a day without food, and LC3B-II increased only in cells that had spent an hour in a culture medium that itself reduces flux. Across the full table, six of the twenty studies measured flux, and four found no difference. In a trial involving 121 people with obesity, an exploratory analysis found higher flux after six months with intermittent fasting than with standard care. However, flux did not increase significantly from baseline; a decline in the standard-care group may have accounted for part of the difference. The review authors’ own group conducted three of the six studies that measured flux, and Sargent is listed as an inventor on a patent for the flux measurement method.

A registered trial plans to use the same method to measure flux during a multiday fast: 24 participants will consume only water for three days. The authors’ priorities are to establish whether muscle and fat biopsies are suitable for measuring flux and to identify a marker of autophagy in bodily fluids that could speed up the development of interventions targeting the process.