Assessing Health Span in Caenorhabditis elegans: Lessons From Short-Lived Mutants

Abstract

Genetic changes resulting in increased life span are often positively associated with enhanced stress resistance and somatic maintenance. A recent study found that certain long-lived Caenorhabditis elegans mutants spent a decreased proportion of total life in a healthy state compared with controls, raising concerns about how the relationship between health and longevity is assessed. We evaluated seven markers of health and two health-span models for their suitability in assessing age-associated health in invertebrates using C elegans strains not expected to outperform wild-type animals. Additionally, we used an empirical method to determine the transition point into failing health based on the greatest rate of change with age for each marker. As expected, animals with mutations causing sickness or accelerated aging had reduced health span when compared chronologically to wild-type animals. Physiological health span, the proportion of total life spent healthy, was reduced for locomotion markers in chronically ill mutants, but, surprisingly, was extended for thermotolerance. In contrast, all short-lived mutants had reduced "quality-of-life" in another model recently employed for assessing invertebrate health. Results suggest that the interpretation of physiological health span is not straightforward, possibly because it factors out time and thus does not account for the added cost of extrinsic forces on longer-lived strains.

Keywords: Health; Invertebrate; Life-span measurement; Phenotype; Sarcopenia.

Figure 1.

Short-lived Caenorhabditis elegans mutants demonstrate deficiencies in health during development and adulthood. (A) Representative survival curves of N2 wild-type and short-lived mutants from four biological repeats (Supplementary Table 1). Survival curves were compared using Mantel-Cox log-rank tests and all short-lived mutants had significantly (p < .05) shorter median life spans than wild-type N2 (mean survival difference for daf-16(mu86) was −33%, mev-1(kn1) was −29%, hsf-1(sy441) was −37%, and sir-2.1(ok434) was −26%). (B) Mean survival at 35°C on Day 6 of adulthood was reduced in daf-16, mev-1 and hsf-1, but not in sir-2.1 (p = 8.0E-8, 2.4E-11, 2.4E-11, and 0.79, respectively, calculated using Tukey’s test). Data were pooled from three independent experiments, n = 300/strain. (C) Mean survival was reduced in all short-lived strains in response to 7.5-µM hydrogen peroxide at Day 5 of adulthood (p < .0001, Tukey’s test). Data were pooled from three independent experiments, n = 138/strain. (D) The size of selected strains based on imaged area on Day 5 of adulthood showed that only mev-1 were smaller compared with wild type (p = 2.8E-13, Tukey’s test). Data were pooled from 3 independent experiments, n = 45/strain. Data were pooled from 3 independent experiments. E) Time to development for each strain as measured from time to egg laying to first day of adulthood. Compared with wild type, mev-1 and hsf-1 had a longer development time while daf-16 and sir-2.1 were similar (p = 1E-16, p = 1.0E-16, p = .088 and p =.049, respectively Tukey’s test). Data were pooled from 4 independent experiments, n = 70/strain. F) Fecundity of each selected strain as measured by the number of eggs hatched. The mutants mev-1 and sir-2.1 had reduced hatched progeny compared with wild type (p = 1.0E-12 and 1.4E-12, respectively in Tukey’s test). Data were pooled from four independent experiments, n = 64/strain. Error bars indicate SEM. ***p < .001.

Figure 2.

Locomotion and thermotolerance were reduced sooner chronologically in short-lived mutants. Mean and maximum speed, maximum bending amplitude, and thermotolerance throughout life are shown in A–D. Error bars indicate SEM. Data were pooled from three independent experiments. n = 45/strain for locomotion parameters. n = 300/strain for thermotolerance. *p < .05, ***p < .001, as compared with N2 wild type on the same day by analysis of variance with post hoc Tukey’s test.

Figure 3.

Autofluorescence and pharyngeal pumping were not altered sooner in short-lived mutants. (A) DAPI channel autofluorescence, (B) GFP channel autofluorescence, and (C) pharyngeal pumping of wild-type and short-lived mutants throughout life. Insufficient numbers of hsf-1 mutants survived to Day 19 to take complete measurements for that time point. Pumping was not significantly different at any time point for any strain compared with wild type. Error bars indicate SEM. Data were pooled from three independent experiments. n = 45/strain for autofluorescence measurements, n = 200/strain for pharyngeal pumping. *p < .05, **p < .01, ***p < .001, as compared with wild type on the same day by analysis of variance with post hoc Tukey’s test.

Figure 4.

Physiological health span is decreased for locomotion markers and increased for thermotolerance in short-lived mutants. Chronological health span and gerospan for movement speed, maximum speed, maximum bending amplitude, and thermotolerance markers were calculated in A–D, respectively. (E,H) Same as in A–D, but for physiological health span and gerospan. Health span and gerospan are indicated by lighter and darker hues, respectively. Error bars indicate SEM for the prediction of the onset of gero span. The cumulative quality-adjusted survival, at metric sensitive to both the total quality and duration of life was calculated for speed, maximum speed, maximum amplitude, and thermotolerance in I–L.