Clerodendranthus spicatus (Thunb.) Water Extracts Reduce Lipid Accumulation and Oxidative Stress in the Caenorhabditis elegans

Abstract

Clerodendranthus spicatus (Thunb.) (Kidney tea) is a very distinctive ethnic herbal medicine in China. Its leaves are widely used as a healthy tea. Many previous studies have demonstrated its various longevity-promoting effects; however, the safety and specific health-promoting effects of Clerodendranthus spicatus (C. spicatus) as a dietary supplement remain unclear. In order to understand the effect of C. spicatus on the longevity of Caenorhabditis elegans (C. elegans), we evaluated its role in C. elegans; C. spicatus water extracts (CSw) were analyzed for the major components and the effects on C. elegans were investigated from physiological and biochemical to molecular levels; CSw contain significant phenolic components (primarily rosmarinic acid and eugenolinic acid) and flavonoids (primarily quercetin and isorhamnetin) and can increase the lifespan of C. elegans. Further investigations showed that CSw modulate stress resistance and lipid metabolism through influencing DAF-16/FoxO (DAF-16), Heat shock factor 1 (HSF-1), and Nuclear Hormone Receptor-49 (NHR-49) signalling pathways; CSw can improve the antioxidant and hypolipidemic activity of C. elegans and prolong the lifespan of C. elegans (with the best effect at low concentrations). Therefore, the recommended daily use of C. spicatus should be considered when consuming it as a healthy tea on a daily basis.

Keywords: Caenorhabditis elegans; Clerodendranthus spicatus; anti-ageing; antioxidant; kidney tea; resistance.

Figure 1

Effect of CSw with different concentrations on the life span of C. elegans. (A) Survival curves of C. elegans treated with different concentrations of CSw (0, 20, 50, 100, 500, and 1000 μg/mL); (B) qualitative observation of fluorescence intensity of lipofuscin; (C) representative pictures of lipofuscin. Different letters (a, b) represent significant (p < 0.05) differences among different groups; the same letters indicate no significant differences between the two groups. The same subscript number (a₁, b₁, c₁) indicates that the significant differences analysis is performed within the same group.

Figure 2

Effect of CSw with different concentrations on the health parameters of C. elegans. (A) Spawning; (B) pharyngeal pumping rate was counted for 30 s; (C) body length; (D) body width; (E) head swing; and (F) body bends counted under a dissecting microscope for 20 s. Different letters (a, b, c) represent significant (p < 0.05) differences among different groups; the same letters indicate no significant differences between the two groups. The same subscript number (a₁, b₁, c₁) indicates that the significant differences analysis is performed within the same group.

Figure 3

CSw activated the antioxidant system of C. elegans. Survival rates of C. elegans under (A) heat stress; (B) oxidative stress; and (C) UV stress; (D) representative picture of reactive oxygen species (ROS) in C. elegans; (E) ROS and Malondialdehyde (MDA) content; (F) Superoxide dismutase (SOD) and Catalase (CAT) activity. Different letters (a, b, c) represent significant (p < 0.05) differences among different groups; the same letters indicate no significant differences between the two groups. The same subscript number (a₁, b₁, c₁) indicates that the significant differences analysis is performed within the same group.

Figure 4

CSw reduce the fat content of C. elegans. (A) Photographs of C. elegans stained with oil red O; (B) quantitative results of oil red O; (C) triglyceride (TG) content; and (D) free fatty acid (FFA) content in C. elegans. Different letters (a, b, c) represent significant (p < 0.05) differences among different groups; the same letters indicate no significant differences between the two groups.

Figure 5

CSw reduce body fat content in C. elegans by NHR-49. (A) mRNA levels of genes related to insulin signalling (IIS) pathway; (B) mRNA levels of genes related to SBP-1/MDT-15 signalling pathway; (C) mRNA levels of genes related to TOR and hexosamine signalling pathway; (D) mRNA levels of genes related to NHR-49 signalling pathway; (E) expression of acs-2 in nhr-49 mutants after CSw treatment; (F,G) transgenic representative pictures and quantitative fluorescence intensity of acs-2::GFP in C. elegans. Different letters (a, b, c) represent significant (p < 0.05) differences among different groups; the same letters indicate no significant differences between the two groups. The same subscript number (a₁, b₁, c₁, a₂, b₂, a₃, b₃, a₄, b₄) indicates that the significant differences analysis is performed within the same group.

Figure 6

Life-prolonging effect of CSw depends on DAF-16 and HSF-1. (A–C) mRNA levels of daf-16, hsf-1, and skn-1 and their target genes; (D,E) representative pictures and distribution of daf-16 and skn-1. * p < 0.05, ** p < 0.01.

Figure 7

Life-prolonging effect of CSw depends on DAF-16 and HSF-1. (A–D) Survival curves of mutants daf-16, hsf-1, skn-1, and mev-1; (E) ROS levels of mutants treated with different concentrations of CSw; (F,G) transgenic representative pictures and quantitative fluorescence intensities of sod-3::GFP and hsp-16.2::GFP in C. elegans. Different letters (a, b, c, d) represent significant (p < 0.05) differences among different groups; the same letters indicate no significant differences between the two groups. The same subscript number (a₁, b₁) indicates that the significant differences analysis is performed within the same group.