In VitroandIn VivoProtective Effects of Flavonoid-Enriched Lotus Seedpod Extract on Lipopolysaccharide-Induced Hepatic Inflammation

Abstract

Endotoxin lipopolysaccharide (LPS) plays an important role in the acceleration of hepatic inflammation. Natural medicinal plants that can prevent inflammation by targeting LPS have potential therapeutic clinical application. The aim of the study is to examine the anti-inflammatory effects of lotus seedpod extract (LSE), used as a traditional Chinese herbal medicine with hemostasis function and for eliminating bruise, on the LPS-induced hepatic inflammation and its underlying molecular mechanisms in vitro and in vivo. In vitro, LSE and its purified compound (-)-epigallocatechin (EGC) dose-dependently inhibited the expressions of pro-inflammatory cytokines and mediators, including tumor necrosis factor (TNF)-[Formula: see text], interleukin (IL)-6, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), without affecting cell viability in LPS-stimulated human hepatoma cell line HepG2. Molecular studies showed the anti-LPS effect of HLP or EGC might be mediated via downregulation of Toll-like receptor 4. (TLR4)-mediated both NF-[Formula: see text]B and p38 signaling, as demonstrated by the usage of pyrrolidine dithiocarbamate (PDTC), a specific NF-[Formula: see text]B inhibitor. In vivo, LPS-induced hepatic inflammation was significantly ameliorated in LSE-fed mice as gauged by dose-dependent inhibition of serum levels of biochemical markers of liver damage, the changes of hepatic lobular architecture and the secretion of pro-inflammatory mediators, as well as induction of anti-oxidant enzymes. As a result, our data presented the first evidence of EGC-enriched LSE as an anti-inflammatory agent in LPS-stimulated HepG2 cells and mice, and these findings may open interesting perspectives to the strategy in treatment for hepatic inflammation.

Lotus Seedpod Extracts Reduced Lipid Accumulation and Lipotoxicity in Hepatocytes

“…In addition, to demonstrate that LSE is an inhibitor of OA-induced cytotoxicity and lipid deposition, we excluded the effect of LSE alone on HepG2 cell growth by trypan blue dye exclusion assay showing that the cell viability was not significantly altered by the treatment of LSE at doses of <25 μg/mL ( Figure 1B). As shown in Figure 1C, the decreases were increased in the cells incubated with combinations of OA and increasing concentrations of LSE at 2.5, 5, and 10 μg/mL or EGC at 4 μM (the concentration of EGC in LSE at 10 μg/mL was approximately 1.26 μg/mL, which was equivalent to about 4 μM [25]) for 48 h, when compared to the OA alone group. It is worth noting the combination of OA and LSE indicated significant antagonistic efficacy, especially in the dose of 10 μg/mL of LSE, which almost completely blocked the OA-inhibited cell growth.…”

“…This course can have a lethal aftermath and lead to liver cirrhosis and carcinoma [40]. It has been informed that EGC enriched LSE as an anti-inflammatory agent in Lipopolysaccharides-stimulated HepG2 cells and mice [25]. Recent evidences have focused on the control for NAFLD pathogenesis using micronutrient antioxidants and/or dietary components, such as vitamins and polyphenols [41][42][43].…”

“…The dried lotus seedpods (100 g) were macerated with hot water (95 • C, 4000 mL) for 2 h and the aqueous extract was evaporated under vacuum at −85 • C. The extracted solution was filtered, then lyophilized to obtain approximately 27 g of LSE and stored at −20 • C before use. The concentrations of total polyphenols and flavonoids contents of LSE were measured as described previously [25].…”

“…The trypan blue dye exclusion assay was used to evaluate the effect of the test drugs on cell viability, as described previously [25], and to determine the non-cytotoxic concentrations. Cells were seeded at a density of 10 5 cells/mL and treated with OA (0.6 mM) in the presence or absence of LSE or EGC at various concentrations for 24 h or 48 h. Afterward, the cells were stained with trypan blue, and live cells were enumerated to determine % cell growth.…”

“…It has been reported that lotus seedpod is rich in polyphenols and an important natural source of oligomers and polymers of catechin, which are also denominated procyanidins [19,20]. In recent years, procyanidins from lotus seedpod have been shown to possess antioxidant, anti-aging, anti-memory impairment, radioprotective, anti-glycative, and anti-inflammatory activities [20][21][22][23][24][25]. However, little information is available on the anti-lipid accumulation and anti-lipotoxic effect of lotus seedpod extract (LSE) in hepatocytes.…”