Gon Sup Kim

Citrus aurantium flavonoids inhibit adipogenesis through the Akt signaling pathway in 3T3-L1 cells

BackgroundObesity is a health hazard that is associated with a number of diseases and metabolic abnormalities, such as type-2 diabetes, hypertension, dyslipidemia, and coronary heart disease. In the current study, we investigated the effects of Citrus aurantium flavonoids (CAF) on the inhibition of adipogenesis and adipocyte differentiation in 3T3-L1 cells.MethodsDuring adipocyte differentiation, 3T3-L1 cells were treated with 0, 10, and 50 μg/ml CAF, and then the mRNA and protein expression of adipogenesis-related genes was assayed. We examined the effect of CAF on level of phosphorylated Akt in 3T3-L1 cells treated with CAF at various concentrations during adipocyte differentiation.ResultsThe insulin-induced expression of C/EBPβ and PPARγ mRNA and protein were significantly down-regulated in a dose-dependent manner following CAF treatment. CAF also dramatically decreased the expression of C/EBPα, which is essential for the acquisition of insulin sensitivity by adipocytes. Moreover, the expression of the aP2 and FAS genes, which are involved in lipid metabolism, decreased dramatically upon treatment with CAF. Interestingly, CAF diminished the insulin-stimulated serine phosphorylation of Akt (Ser473) and GSK3β (Ser9), which may reduce glucose uptake in response to insulin and lipid accumulation. Furthermore, CAF not only inhibited triglyceride accumulation during adipogenesis but also contributed to the lipolysis of adipocytes.ConclusionsIn the present study, we demonstrate that CAF suppressed adipogenesis in 3T3-L1 adipocytes. Our results indicated that CAF down-regulates the expression of C/EBPβ and subsequently inhibits the activation of PPARγ and C/EBPα. The anti-adipogenic activity of CAF was mediated by the inhibition of Akt activation and GSK3β phosphorylation, which induced the down-regulation of lipid accumulation and lipid metabolizing genes, ultimately inhibiting adipocyte differentiation.

Induction of the cell cycle arrest and apoptosis by flavonoids isolated from Korean Citrus aurantium L. in non-small-cell lung cancer cells

This study investigated the anti-proliferative and apoptotic effect of flavonoids isolated from Korean Citrus aurantium L. using A549 lung cancer cells. Flavonoids potently inhibited of A549 cells in a dose-dependent manner, whereas flavonoids had a weak inhibitory effect on proliferation of WI-38 cells. Flow cytometry and Western blot analysis showed that flavonoids induced cell cycle arrest at the G2/M checkpoint by controlling the proteins expression level of cyclin B1, cdc2, cdc25c and p21(WAF1/CIP1). Also, flavonoids induced apoptosis through the regulation of the expression of caspases, cleaved PARP and Bax/Bcl-xL ratio. The activity of caspase-3 on A549 cells increased in a dose-dependent manner. These results clearly indicated that the anti-cancer effect of flavonoids on A549 cells follows multiple cellular pathways through G2/M arrest and the induction of apoptosis.

Flavonoids Isolated from Korea Citrus aurantium L. Induce G2/M Phase Arrest and Apoptosis in Human Gastric Cancer AGS Cells

Aim of the Study. Citrus species is used in traditional medicine as medicinal herb in several Asian countries including Korea. Flavonioids became known as various properties, such as anti-oxidants, anti-inflammation and anti-cancer, and so forth. The present study, the anti-cancer effect of flavonioids isolated from Citrus aurantium L. in human gastric cancer AGS cells has been investigated. Materials and Methods. The anti-proliferative activity was assayed using MTT assay. Cell cycle analysis was done using flow cytometry and apoptosis detection was done using by hoechst fluorescent staining and Annexin V-propidium iodide double staining. Western blot was used to detect the expression of protein related with cell cycle and apoptosis. Results. Flavonoids isolated from Citrus aurantium L. have the effect of anti proliferation on AGS cells with IC50 value of 99 μg/mL. Flavonoids inhibited cell cycle progression in the G2/M phase and decrease expression level of cyclin B1, cdc 2, cdc 25c. Flavonoids induced apoptosis through activate caspase and inactivate PARP. Conclusions. Flavonoids isolated from Citrus aurantium L. induced G2/M phase arrest through the modulation of cell cycle related proteins and apoptosis through activation caspase. These finding suggest flavonoids isolated from Citrus aurantium L. were useful agent for the chemoprevention of gastric cancer.

Naringin induces autophagy-mediated growth inhibition by downregulating the PI3K/Akt/mTOR cascade via activation of MAPK pathways in AGS cancer cells

Naringin, one of the major bioflavonoid of Citrus, has been demonstrated as potential anticancer agent. However, the underlying anticancer mechanism still needs to be explored further. This study investigated the inhibitory effect of Naringin on human AGS cancer cells. AGS cell proliferation was inhibited by Naringin in a dose- and time-dependent manner. Naringin did not induce apoptotic cell death, determined by no DNA fragmentation and the reduced Bax/Bcl-xL ratio. Growth inhibitory role of Naringin was observed by western blot analysis demonstrating downregulation of PI3K/Akt/mTOR cascade with an upregulated p21CIPI/WAFI. Formation of cytoplasmic vacuoles and autophagosomes were observed in Naringin-treated AGS cells, further confirmed by the activation of autophagic proteins Beclin 1 and LC3B with a significant phosphorylation of mitogen activated protein kinases (MAPKs). Collectively, our observed results determined that anti-proliferative activity of Naringin in AGS cancer cells is due to suppression of PI3K/Akt/mTOR cascade via induction of autophagy with activated MAPKs. Thus, the present finding suggests that Naringin induced autophagy- mediated growth inhibition shows potential as an alternative therapeutic agent for human gastric carcinoma.

Hesperidin Induces Paraptosis Like Cell Death in Hepatoblatoma, HepG2 Cells: Involvement of ERK1/2 MAPK

Hesperidin, a natural flavonoid abundantly present in Citrus is known for its anti-cancer, anti-oxidant and anti-inflammatory properties. In this study we examined the effect of hesperidin on HepG2 cells. HepG2 cells treated with various concentration of hesperidin undergo a distinct type of programed cell death. Cytoplasmic vacuolization, mitochondria and endoplasmic reticulum swelling and uncondensed chromatin were observed in hesperidin treated cells. DNA electrophoresis show lack of DNA fragmentation and western blot analysis demonstrates lack of caspase activation and PARP cleavage. It was observed that hesperidin induced cell death is nonautophagic and also activate mitogen activated protein kinase ERK1/2. Taken together, the data indicate that hesperidin induces paraptosis like cell death in HepG2 cells with the activation of ERK1/2. Thus our finding suggests that hesperidin inducing paraptosis may offer an alternative tool in human liver carcinoma therapy.

Regulation of Proinflammatory Mediators via NF-κB and p38 MAPK-Dependent Mechanisms in RAW 264.7 Macrophages by Polyphenol Components Isolated from Korea Lonicera japonica THUNB

Lonicera japonica THUNB., which abundantly contains polyphenols, has been used as a traditional medicine for thousands of years in East Asian countries because of the anti-inflammation properties. This study aimed to investigate the anti-inflammatory mechanism of polyphenol components isolated from Korea L. japonica T. by nuclear factor-kappaB (NF-κB) and mitogen-activated protein kinases (MAPKs) pathway. Polyphenols significantly decreased lipopolysaccharide- (LPS-) induced mRNA and protein expression of inducible nitric oxide synthase and cyclooxygenase-2, as well as mRNA expression of tumor necrosis factor-alpha, interleukin- (IL-) 1β, and IL-6. Moreover, polyphenols inhibited nuclear translocation of NF-κB p65, phosphorylation/degradation of the inhibitor of κB, and phosphorylation of p38 MAPK, whereas the extracellular signal-regulated kinase and Janus N-terminal kinase were not affected. These results indicate that polyphenol components isolated from Korea L. japonica T. should have anti-inflammatory effect on LPS-stimulated RAW 264.7 cells through the decrease of proinflammatory mediators expression by suppressing NF-κB and p38 MAPK activity.

Mitochondrial Dysfunction and Ca(2+) Overload Contributes to Hesperidin Induced Paraptosis in Hepatoblastoma Cells, HepG2.

Paraptosis is a programmed cell death which is morphologically and biochemically different from apoptosis. In this study, we have investigated the role of Ca2+ in hesperidin‐induced paraptotic cell death in HepG2 cells. Increase in mitochondrial Ca2+ level was observed in hesperidin treated HepG2 cells but not in normal liver cancer cells. Inhibition of inositol‐1,4,5‐triphosphate receptor (IP3R) and ryanodine receptor also block the mitochondrial Ca2+ accumulation suggesting that the release of Ca2+ from the endoplasmic reticulum (ER) may probably lead to the increase in mitochondrial Ca2+ level. Pretreatment with ruthenium red (RuRed), a Ca2+ uniporter inhibitor inhibited the hesperidin‐induced mitochondrial Ca2+ overload, swelling of mitochondria, and cell death in HepG2 cells. It has also been demonstrated that mitochondrial Ca2+ influxes act upstream of ROS and mitochondrial superoxide production. The increased ROS production further leads to mitochondrial membrane loss in hesperidin treated HepG2 cells. Taken together our results show that IP3R and ryanodine receptor mediated release of Ca2+ from the ER and its subsequent influx through the uniporter into mitochondria contributes to hesperidin‐induced paraptosis in HepG2 cells. J. Cell. Physiol. 231: 1261–1268, 2016.

Morin, a Flavonoid from Moraceae, Induces Apoptosis by Induction of BAD Protein in Human Leukemic Cells

Evidence suggests that phytochemicals can safely modulate cancer cell biology and induce apoptosis. Here, we investigated the anti-cancer activity of morin, a flavone originally isolated from members of the Moraceae family in human leukemic cells, focusing on apoptosis. An anti-cancer effect of morin was screened with several human leukemic cell lines. U937 cells were most sensitive to morin, where it induced caspase-dependent apoptosis in a dose-dependent manner. It also induced loss of MMP (ΔΨm) along with cytochrome c release, down-regulated Bcl-2 protein, and up-regulated BAX proteins. The apoptotic activity of morin was significantly attenuated by Bcl-2 augmentation. In conclusion, morin induced caspase-dependent apoptosis through an intrinsic pathway by upregulating BAD proteins. In addition, Bcl-2 protein expression is also important in morin-induced apoptosis of U937 cells. This study provides evidence that morin might have anticancer properties in human leukemic cells.

Centipede grass exerts anti-adipogenic activity through inhibition of C/EBPβ, C/EBPα, and PPARγ expression and the AKT signaling pathway in 3T3-L1 adipocytes

BackgroundCentipede grass (CG) originates from China and South America and is reported to contain several C-glycosyl flavones and phenolic constituents, including maysin and luteolin derivatives. This study aimed to investigate, for the first time, the antiobesity activity of CG and its potential molecular mechanism in 3T3-L1 cells.MethodsTo study the effect of CG on adipogenesis, differentiating 3T3-L1 cells were treated every day with CG at various concentrations (0–100 μg/ml) for six days. Oil-red O staining and triglyceride content assay were performed to determine the lipid accumulation in 3T3-L1 cells. The expression of mRNAs or proteins associated with adipogenesis was measured using RT-PCR and Western blotting analysis. We examined the effect of CG on level of phosphorylated Akt in 3T3-L1 cells treated with CG at various concentration s during adipocyte differentiation.ResultsDifferentiation was investigated with an Oil-red O staining assay using CG-treated 3T3-L1 adipocytes. We found that CG suppressed lipid droplet formation and adipocyte differentiation in 3T3-L1 cells in a dose-dependent manner. Treatment of the 3T3-L1 adipocytes with CG resulted in an attenuation of the expression of adipogenesis-related factors and lipid metabolic genes. The expression of C/EBPα and PPARγ, the central transcriptional regulators of adipogenesis, was decreased by the treatment with CG. The expression of genes involved in lipid metabolism, aP2 were significantly inhibited following the CG treatment. Moreover, the CG treatment down-regulated the phosphorylation levels of Akt and GSK3β.ConclusionsTaken collectively, these data indicated that CG exerts antiadipogenic activity by inhibiting the expression of C/EBPβ, C/EBPα, and PPARγ and the Akt signaling pathway in 3T3-L1 adipocytes.

Flavonoids Identified from Korean Scutellaria baicalensis Georgi Inhibit Inflammatory Signaling by Suppressing Activation of NF-κB and MAPK in RAW 264.7 Cells

Scutellaria baicalensis Georgi has been used as traditional medicine for treating inflammatory diseases, hepatitis, tumors, and diarrhea in Asia. Hence, we investigated the anti-inflammatory effect and determined the molecular mechanism of action of flavonoids isolated from Korean S. baicalensis G. in lipopolysaccharide- (LPS-) stimulated RAW 264.7 macrophages. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was performed to examine cytotoxicity of the flavonoids at various concentrations of 10, 40, 70, and 100 µg/mL. No cytotoxicity was observed in RAW 264.7 cells at these concentrations. Furthermore, the flavonoids decreased production of inflammatory mediators such as inducible nitric oxide synthase, cyclooxygenase-2, interleukin-6, and tumor necrosis factor-alpha and inhibited phosphorylation of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPKs) in LPS-induced RAW 264.7 cells. Moreover, to identify the differentially expressed proteins in RAW 264.7 cells of the control, LPS-treated, and flavonoid-treated groups, two-dimensional gel electrophoresis and mass spectrometry were conducted. The identified proteins were involved in the inflammatory response and included PRKA anchor protein and heat shock protein 70 kD. These findings suggest that the flavonoids isolated from S. baicalensis G. might have anti-inflammatory effects that regulate the expression of inflammatory mediators by inhibiting the NF-κB signaling pathway via the MAPK signaling pathway in RAW 264.7 cells.