The Effects of Kaempferol-Inhibited Autophagy on Osteoclast Formation

Abstract

Kaempferol, a flavonoid compound, is derived from the rhizome of Kaempferia galanga L., which is used in traditional medicine in Asia. Autophagy has pleiotropic functions that are involved in cell growth, survival, nutrient supply under starvation, defense against pathogens, and antigen presentation. There are many studies dealing with the inhibitory effects of natural flavonoids in bone resorption. However, no studies have explained the relationship between the autophagic and inhibitory processes of osteoclastogenesis by natural flavonoids. The present study was undertaken to investigate the inhibitory effects of osteoclastogenesis through the autophagy inhibition process stimulated by kaempferol in murin macrophage (RAW 264.7) cells. The cytotoxic effect of Kaempferol was investigated by MTT assay. The osteoclast differentiation and autophagic process were confirmed via tartrate-resistant acid phosphatase (TRAP) staining, pit formation assay, western blot, and real-time PCR. Kaempferol controlled the expression of autophagy-related factors and in particular, it strongly inhibited the expression of p62/SQSTM1. In the western blot and real time-PCR analysis, when autophagy was suppressed with the application of 3-Methyladenine (3-MA) only, osteoclast and apoptosis related factors were not significantly affected. However, we found that after cells were treated with kaempferol, these factors inhibited autophagy and activated apoptosis. Therefore, we presume that kaempferol-inhibited autophagy activated apoptosis by degradation of p62/SQSTM1. Further study of the p62/SQSTM1 gene as a target in the autophagy mechanism, may help to delineate the potential role of kaempferol in the treatment of bone metabolism disorders.

The Role of Autophagy and Mitophagy in Bone Metabolic Disorders

“…During RANKL-induced osteoclast differentiation, the expression and localization of p62/SQSTM1 negatively correlated with LC3 accumulation and F-actin ring formation for regulating autophagic activation 95 . Kaempferol, a flavonoid compound, can inhibit autophagy by degradation of p62/SQSTM1 and activate cell apoptosis during RANKL-induced osteoclastogenesis in murine macrophage (RAW264.7) cells 96 . Paget's disease of bone is a bone disorder characterized by focal areas of abnormal, excessive bone turnover, specifically increased bone resorption and resulting in disorganized bone formation 97 .…”

Rhoifolin ameliorates titanium particle‐stimulated osteolysis and attenuates osteoclastogenesis via RANKL‐induced NF‐κB and MAPK pathways

“…In recent years, an increasing number of studies have been carried out on the role of flavonoids in the prevention and treatment of bone diseases. For example, kaempferol, genistein, quercetin, and naringin have been proven to be capable of protecting bones by inhibiting osteoclastic differentiation and function (Forte et al, 2016;Kim et al, 2018;Phan et al, 2010;X. C. Wang et al, 2014;T.…”

Erigeron annuus Protects PC12 Neuronal Cells from Oxidative Stress Induced by ROS‐Mediated Apoptosis

“…Homogentisic and salicylic acids, which were abundant in both EAA and EAF, confer beneficial effects such as the inhibition of oxidation, anti-inflammation [36][37][38], and disease tolerance in plants [39]. Kaempferol, which was significantly present in EAF, exerts other beneficial effects such as anticancer cell Evidence-Based Complementary and Alternative Medicine proliferation, in vitro antioxidation, and the inhibition of autophagy [40][41][42][43]. ese phenolic and flavonoid compounds produced by photosynthesis are stored in plant leaves and accumulate in the vacuoles of flowers [44].…”