Binary Effects of Gynostemma Gold Nanoparticles on Obesity and Inflammation via Downregulation of PPARγ/CEPBα and TNF-α Gene Expression

Abstract

Nanoscience is a multidisciplinary skill with elucidated nanoscale particles and their advantages in applications to various fields. Owing to their economical synthesis, biocompatible nature, and widespread biomedical and environmental applications, the green synthesis of metal nanoparticles using medicinal plants has become a potential research area in biomedical research and functional food formulations. Gynostemma pentaphyllum (GP) has been extensively used in traditional Chinese medicine to cure several diseases, including diabetes mellitus (DM). This is the first study in which we examined the efficacy of G. pentaphyllum gold nanoparticles (GP-AuNPs) against obesity and related inflammation. GP extract was used as a capping agent to reduce Au²⁺ to Au⁰ to form stable gold nanoparticles. The nanoparticles were characterized by using UV-VIS spectroscopy, and TEM images were used to analyze morphology. In contrast, the existence of the functional group was measured using FTIR, and size and shape were examined using XRD analysis. In vitro analysis on GP-AuNPs was nontoxic to RAW 264.7 cells and 3T3-L1 cells up to a specific concentration. It significantly decreased lipid accumulation in 3T3-L1 obese and reduced NO production in Raw 264.7 macrophage cells. The significant adipogenic genes PPARγ and CEPBα and a major pro-inflammatory cytokine TNF-α expression were quantified using RT-PCR. The GP-AuNPs decreased the face of these genes remarkably, revealing the antiadipogenic and anti-inflammatory activity of our synthesized GP-AuNPs. This study represents thorough research on the antiobesity effect of Gynostemma pentaphyllum gold nanoparticles synthesized using a green approach and the efficacy instead of related inflammatory responses.

Keywords: Gynostemma pentaphyllum; gold nanoparticles; inflammation; obesity; obesity-induced inflammation.

Figure 1

UV–VIS spectra of GP-AuNPs (black) and GP-Ex (red).

Figure 2

Optimization of GP-AuNP synthesis using different plant extract (a), time (b), pH (c).

Figure 3

Field emission transmission characterization of GP-AuNPs on electron micrographs showed that GP-AuNPs are predominantly spherical in shape (a,b,d), and the selected area electron diffraction pattern revealed the crystalline nature of the biosynthesized nanoparticles (c). Elemental mapping and energy dispersive X-ray spectroscopy confirmed the purity of the nanoparticles (e,f).

Figure 4

FTIR analysis of synthesized GP-AuNPs and GP-Ex.

Figure 5

XRD analysis of GP-AuNPs.

Figure 6

Cell viability of 3T3-L1 cell line on treatment with (a) GP extract and GP-AuNPs for 24 h. (b) GP extract and GP-AuNPs on Raw 264.7 cell viability for 24 h. (c) Trypan blue cell viability images before and after treatment with GP-AuNPs on 3T3-L1 cells. Each set of data represents the mean of triplicate experiment ± standard deviation. A significant difference between the groups was calculated using a two-tailed Student’s t-test. * < 0.05, ** p < 0.01 vs. control is used to represent a significant difference in cell viability of the sample compared with a nontreated control group.

Figure 7

Inhibitory effect of the lipid accumulation for GP-AuNPs on MDI-induced 3T3-L1 adipocytes. (A) Fat droplets were measured by oil red O staining and observed using a microscope (at ×40). (B) The absorbance of lipid accumulation, which was oil red O dye, was dissolved in isopropyl alcohol (520 nm). The data are mean values of three experiments ± SEM; ### <0.001 compared with control, ** p < 0.01, *** p < 0.001 compared with the MDI.

Figure 8

Inhibitory Effect of the lipid accumulation for GP extract on MDI-induced 3T3-L1 adipocytes. (A) Fat droplets were measured by oil red O staining and observed using a microscope (at ×40). (B) The absorbance of lipid accumulation, which was oil red O dye, was dissolved in isopropyl alcohol (520 nm). The data are mean values of three experiments ± SEM; ### <0.01 compared with control, ** p < 0.01 compared with the MDI.

Figure 9

Dose-dependent in vitro DPPH radical scavenging and antioxidant activity of biosynthesized GP-AuNPs. Approximately 60% of DPPH inhibition was recorded at the 200 and 400 µg/mL. The data shown represent the mean values of three experiments ± SD. ** p < 0.01, *** p < 0.001 as compared with the Vit-C.2.5. Effect of GP-AuNPs on the LPS-induced NO production.

Figure 10

Effects on the production of NO GP-AuNPs (a) and GP extract (b). Macrophage RAW 264.7 cells were pretreated in both samples for 1 h and then stimulated with LPS (1 μg/mL) for 24 h. The concentrations of nitrite were measured as described in the materials and methods. The data shown represent the mean values of three experiments ± SD. ** p < 0.01, *** p < 0.001 as compared with the group treated with LPS.

Figure 11

Inhibition of reactive oxygen species (ROS) generation by GP-AuNPs in MDI-induced 3T3-L1 adipocytes was determined by DCFDA method. GP-AuNP treatment reduced intracellular ROS production dose dependently. Data are expressed as a percentage of control. *** p < 0.001 MDI vs. control, while ## p < 0.01 and ### p < 0.001 GP-AuNPs vs. MDI.

Figure 12

Effects of GP-AuNPs on mRNA expression levels of adipogenesis-related genes in 3T3-L1 cells. GP-AuNPs inhibit the RNA level of adipogenesis-related genes. ### <0.01 compared with control, * p < 0.05, *** p < 0.001 compared with the MDI.

Figure 13

Effect of GP-AuNPs on the transcriptional activation of the inflammatory gene TNF-α in RAW 264.7 cells. RAW 264.7 cells were pretreated with the indicated concentrations GP-AuNPs for 1 h, and then stimulated with LPS (1 μg/mL) for 24 h. Subsequently, total RNAs were extracted, and the mRNA expression levels were determined by RT-PCR analysis and compared with those of β-actin. The data shown are representative of the mean values of three independent experiments ± SD. ** p < 0.01, *** p < 0.001 as compared with the group treated with LPS, and ### p < 0.001 as compared with the control.