PREreview of Senolytics enhance longevity inCaenorhabditis elegansby altering betaine metabolism

This manuscript investigates key metabolites in regulating aging and longevity. The authors found that the concentration of betaine in the serum of elderly humans and aged worms is lower than in their younger counterparts, as determined by NMR metabolomics. They also have examined several senolytics including metformin, quercetin, and minocycline, and found that the treated worms had higher betaine level and longer lifespan. Betaine supplementation with certain concentration alone was sufficient to prolong lifespan of worms via stimulating autophagy and improving antioxidant capacity. Overall, the textual content is well written, and the figures are clear. The results support the conclusions. However, there are several concerns that require further attention.

Major point:

1. How was the optimal concentration of senolytics decided? The manuscript didn’t include thorough analysis using different concentrations of senolytics. It is unclear whether the ones reported in the manuscript are most effective.

2. There are also changes in choline and glycine, which are important in betaine biosynthesis, in aged worms compared to young worms. What are the levels of choline and glycine in senolytics treated worms?

3. The current experiments were conducted using liquid culture, which is not the common procedure in longitudinal assays? If growing worms on plates, would senolytics and betaine treatments lead to the same lifespan extension?

4. The authors need to provide corresponding images of autophagosomes for young worms at day 1 and day 4 as comparison with aged worms.

5. Details about quantification of metabolites using NMR are missing, such as internal standard and normalization methods. So, please provide information on this quantification.

Minor point:

1) There is no explanation of handling progeneis in the method section, so, please provide information on how to manage the progenies in the lifespan assay and on how to measure ROS.

2) The authors mentioned the utilization of oviposition inhibitors. They should specify the name of the inhibitors, their concentration, and the timing of application.

3) Figure 2D needs more discussion, such as why micorcylcine group has lower percentage of fast body bends.

5) It would be better to indicate significance in images showing the lifespan results.

6) The label of the transmembrane protein is missing in Figure 5. The Figure 5 is also misleading like the readers may think betaine can be derived from these senolytics on the step of senolytics to betaine, please revise it.

8) There is no access to all the supplementary information, so we are unable to evaluate the related observations or conclusions.

Competing interests

The authors declare that they have no competing interests.