Fractional extraction phenolics from C. oleifera seed kernels exhibited anti-inflammatory effect via PI3K/Akt/NF-κB signaling pathway under Caco-2/RAW264.7 co-culture cell model

Camellia oleifera Abel (C. oleifera) is a multifunctional oilseed, which is rich in many biological active substances with health-promoting properties, especially Polyphenols. Previous research revealed that camellia oil phenolics exhibited anti-inflammatory effect, which originated from seed. Thus, we aimed to explore the components of camellia seed phenolics and its potential mechanism of anti-inflammation. Initially, fractional extraction was processed to prepare the phenolics from camellia seed kernels, and we compare four different fractions of phenolics under the LPS-induced Caco-2/RAW264.7 coculturing model. Results showed that free phenolics (FP) had best effect on alleviating pro-inflammatory cytokines (IL-1β, IL-6, IL-8 and TNF-α) compared to esterified-bound phenolics (EP), glycosylated-bound phenolics (GP) and insoluble-bound phenolics (IP). Furthermore, FP reduced inflammation by suppressing the PI3K/Akt/NF-κB signaling pathway and effectively inhibited LPS-induced intestinal permeability increase, tight junction related proteins loss (ZO-1, claudin-1). Same results obtained, as the transepithelial electrical resistance (TEER) and Alkaline Phosphatase (AKP) activity of high-dose FP treated group was high than model group. Finally, molecular docking was used for evaluating the anti-inflammatory effect for phenolic monomer. KGRG (kaempferol -3-O-(2-O-glucopyranosyl-6-O-rhamnopyranosyl)-glucopyranoside), KXR (kaempferol 3-O-(2''-xylopyranosyl)-rutinoside) and leucoside (kaempferol 3-O-sambubioside) show lower binding energy docking with NF-κB, PI3K and Akt protein, indicating better interactions, which might be effective constituents against inflammation. Subsequently, five major Polyphenols were obtained to validate the docking results, especially, indicating the best anti-inflammatory activities of KGRG. Overall, this research sheds insights on the therapy of phenolics from C. oleifera seed towards LPS-induced intestinal inflammation model in vitro and its related mechanism.

C.oleifera seed kernel; Caco-2/RAW264.7 co-culture cell model; PI3K/Akt/NF-κB; Phenolics; Tight junction proteins.