Quinic acid could be a potential rejuvenating natural compound by improving survival of Caenorhabditis elegans under deleterious conditions

Abstract

Quinic acid (QA) is an active ingredient of Cat's Claw (Uncaria tomentosa), which is found to be active in enhancing DNA repair and immunity in model systems and able to generate neuroprotective effects in neurons. However, QA's role in improving survival is not well studied. Here we report that QA can provide protection in Caenorhabidits elegans and improve worm survival under stress. Under heat stress and oxidative stress, QA-treated wild-type C. elegans N2 (N2) survived 17.8% and 29.7% longer, respectively, than the control worms. Our data suggest that under heat stress, QA can upregulate the expression of the small heat shock protein hsp-16.2 gene, which could help the worms survive a longer time. We also found that QA extended the C. elegans mutant VC475 [hsp-16.2 (gk249)] life span by 15.7% under normal culture conditions. However, under normal culture conditions, QA did not affect hsp-16.2 expression, but upregulated the expression of daf-16 and sod-3 in a DAF-16-dependent manner, and downregulated the level of reactive oxygen species (ROS), suggesting that under normal conditions QA acts in different pathways. As a natural product, QA demonstrates great potential as a rejuvenating compound.

FIG. 1.

Effect of quinic acid (QA) on the life span of N2 worms under normal culture conditions. At 20°C, QA exhibited different effects at different concentrations: 0.05 mg/mL (3.2%, n=258), 0.1 mg/mL (6.9%, n=193), 0.2 mg/mL (2.2%, n=179), 0.4 mg/mL (−3.6%, n=194); resveratrol (100 μM, positive control, 5.2%, n=212). Rates of increase were calculated in comparison with the control (n=183). Worms were fed on live E. coli strain OP50. Please refer to Table 1 for statistical data. (*) p<0.05; (**) p<0.01.

FIG. 2.

Quinic acid (QA) provides protection for C. elegans under heat stress. (A) Under heat stress (35°C), QA at 0.1 mg/mL significantly improves the worm survival time by 17.8% (n=267), compared to the control (n=271). Please refer to Table 2 for statistical data. (B) There was no significant difference between the QA (0.1 mg/mL)-treated group and the control before heat stress and 4 hr after heat stress. (C) Representative heat shock protein-16.2::green fluorescent protein (HSP-16.2::GFP) picture of the control group, after 24-hr recovery following heat stress. (D) Representative HSP-16.2::GFP picture of the QA (0.1 mg/mL)-treated group, after 24-hr recovery following heat stress, stronger than C. (E) HSP-16.2::GFP intensity comparison between the control and QA (0.1 mg/mL)-treated group, after 24-hr recovery following heat stress. There was a significant increase of HSP-16.2::GFP intensity in the QA-treated group. Results were averaged from four single experiments, with 20 worms in every experiment, and were detected on a Thermolabsystems Fluoroskan Ascent microplate reader. (F) QA significantly extended the life span of mutant VC475 [hsp-16.2 (gk249)] by 15.7%. QA-treated group, n=173. Control group, n=170. Please refer to Table 3 for statistical data. The error bars indicate±standard error SE). (*) p<0.05; (**) p<0.01. OP50, E. coli strain OP50.

FIG. 3.

Quinic acid (QA) upregulates the expression of the sod-3 gene in a daf-16–dependent manner. (A) Representative green fluorescent protein (GFP) photograph demonstrating the fluorescence intensity of superoxide dismutase-3::green fluorescent protein (SOD-3::GFP) in adult mutant CF1553 (muIs84) in the control group. (B) Representative GFP photograph demonstrating the fluorescence intensity of SOD-3::GFP in adult mutant CF1553 (muIs84) in QA (0.1 mg/mL)-treated group (2 days treatment), stronger than A. (C) QA (0.1 mg/mL) could significantly upregulate fluorescence intensity of SOD-3::GFP in CF1553 (muIs84), but not in mutant CF1588 [daf-16(mu86) I; daf-2(e1370) III; muIs84] when daf-16 was mutated. (D) SOD-3::GFP intensity (±SE) in adult CF1553 (muIs84) with or without 0.1 mg/mL QA over a period of 9 days. QA could keep the SOD-3::GFP fluorescence intensity at a higher level than the control. (C and D) Data were averaged from four single experiments, with 20 worms in every experiment, and were detected on a Thermolabsystems Fluoroskan Ascent microplate reader. (E) QA at 0.1 mg/mL could significantly upregulate the expression of sod-3 and daf-16 in N2 worms. Gene expression was detected with quantitative real-time PCR. (F) QA (0.1 mg/mL) could not extend the life span of mutant CF1038 [daf-16(mu86) I], when daf-16 was mutated. For the QA-treated group, n=359; for the control group, n=320. Please refer to Table 4 for statistical data. The error bars indicate±standard error (SE). (*) p<0.05.

FIG. 4.

Quinic acid (QA) scavenges free radicals directly and indirectly. (A) In vitro free radical scavenging effect of QA (0.1 mg/mL). (B) QA (0.1 mg/mL) increased the reduced glutathione to oxidized glutathione ratio (GSH/GSSG) in N2 worms. (C) In vivo free radical scavenging effect of QA (0.1 mg/mL) under normal culture conditions, detected after 30 min of incubation at 37°C. (D) In vivo free radical scavenging effect of QA (0.1 mg/mL) under oxidative stress generated by 500 μM juglone, detected after 30 min of incubation at 37°C, and lasted 90 min. (E) Under oxidative stress (500 μM juglone), QA at 0.1 mg/mL significantly improved the N2 worm survival time by 29.7% (n=269), compared to the control (n=201). Please refer to Table 5 for statistical data. Every data point is the average value of three trials. The error bars indicate±standard error (SE). (*) p<0.05; (**) p<0.01.