Quercetin-3- O-rutinoside from Moringa oleifera Downregulates Adipogenesis and Lipid Accumulation and Improves Glucose Uptake by Activation of AMPK/Glut-4 in 3T3-L1 Cells

Abstract

Natural product-based therapeutic alternatives have drawn immense interest to deal with growing incidence of metabolic disorders. Rutin (quercetin-3-O-rutinoside) is found in a variety of fruits, vegetables, and plant beverages. In the present study, rutin was isolated from Moringa oleifera Lam., leaves and its anti-lipidemic and anti-adipogenic activity was evaluated through inhibition of key digestive enzymes and in vitro cell culture experiments using 3T3-L1 adipocytes. Rutin treatment substantially reduced α-glucosidase and pancreatic lipase activities with IC₅₀ values of 40 and 35 μg/ml, respectively. MTT assay with 3T3-L1 cells demonstrated the non-toxic effect of rutin up to 160 μg/ml. Oil Red O-stained images of rutin-treated 3T3-L1 cells depicted that rutin considerably reduced lipid content and adipogenesis (79.9%), and enhanced glycerol release in 3T3-L1 cells when compared to untreated cells. Rutin significantly (p < 0.05) enhanced glucose uptake in 3T3-L1 adipocytes and also led to reduced levels of leptin but enhanced levels of adiponectin. Western blot analysis of rutin-treated (40 µg/ml) adipocytes showed phosphorylation of AMPK, upregulated expression of Glut-4 (1.31-fold) and UCP-1 (1.47-fold), but downregulated expression of PPAR-γ by 0.73-fold. At transcriptional level, similar trends were observed in the mRNA expression of the above genes, except AMPK. Our results demonstrate that rutin isolated from M. oleifera significantly alleviates lipid content and adipogenesis, and improves glucose uptake through regulating PPAR-γ and AMPK signaling pathways; thus, rutin can be considered as a potential therapeutic agent against adiposity and glucose intolerance.

Keywords: Adipogenesis; Adipokines; Glucose uptake; Lipase; Rutin; α-Glucosidase.

Fig. 1

Effect on key digestive enzymes. a α-Glucosidase inhibition. b Pancreatic lipase inhibition (*p < 0.01). Acarbose and orlistat were the standards used for glucosidase and lipase respectively

Fig. 2

Effect on 3T3-L1 adipocytes’ differentiation. a Percentage of cell viability at 24 and 48 h by using MTT assay. b Total lipid content estimated from the lysate of ORO-stained 3T3-L1 cells. c ORO-stained adipocytes’ images illustrating reduced lipid droplets in a concentration-dependent manner (magnification 40 ×). d Estimated glycerol released into the culture media by lipolysis assay. *p < 0.05 and **p < 0.01 significant difference between control and treated adipocytes, respectively

Fig. 3

Effect on glucose uptake. Ins. D, insulin deficient. *p < 0.05 and **p < 0.01 significant difference between control and treated adipocytes, respectively

Fig. 4

Effect on mRNA expression of a UCP-1, b PPAR-γ, c AMPK, and d GLUT-4. *p < 0.05 and **p < 0.01 significant difference between control and treated adipocytes, respectively

Fig. 5

Effect on protein expression. a Protein expression analysis of UCP-1, PPAR-γ, p-AMPK, AMPK, and GLUT-4. b Graphical representation of their protein expression levels. *p < 0.05 and **p < 0.01 significant difference between control and treated adipocytes, respectively

Fig. 6

Effect on adipokines. DMI, differentiation medium of induction. *p < 0.05 and **p < 0.01 indicate significant difference between DMI and rutin-treated adipocytes, respectively