Figure 3. Elevation of systemic klotho levels occurs in human KL-VS carriers and enhances mouse survival, learning, and memory independent of age.

(A) Fasting morning serum klotho levels in individuals from Cohort 1 (55–85 years of age) that were non-carriers (n=118) or carriers (n=38) of a single KL-VS allele. Data were analyzed by a linear model, accounting for effects of age, sex, and education and testing for effects due to KL-VS genotype. APOEε4 carrier status had no effect (Table S7). (B) Hippocampal levels of klotho in NTG and KL mice (n=13–14 mice per genotype, age 3 months). Representative western blots for klotho and actin are shown above; images for each protein were from the same gel. (C) Kaplan-Meier curves show increased survival of heterozygous KL mice from line 46 (Kuro-o et al., 1997; Kurosu et al., 2005) compared to NTG littermates (n=22–29 mice per group, p<0.01 by log rank test). Proportional hazard testing revealed the KL effect was independent of age (p=0.76). (D-E) KL and NTG mice (n=8–9 mice per genotype) were tested in the Morris water maze at 10–12 months of age. (D) Spatial learning curves (platform hidden). Data represent the daily average of total distance traveled to the platform. Mixed model ANOVA: KL effect p<0.05. (E) Results of a probe trial (platform removed) 1 h after completion of hidden-platform training showing latency to reach the original platform location. (F–G) An independent cohort of mice (n=17–19 mice per genotype) was tested in the water maze at 4–7 months of age. (F) Spatial learning curves (platform hidden). Mixed model ANOVA: KL effect p<0.05. (G) Probe trial results. *p<0.05, **p<0.01, ***p<0.001 (t-test). See also Tables S7, S8 and Figure S2. Data are means ± SEM.