Apigenin ameliorates imiquimod-induced psoriasis in C57BL/6J mice by inactivating STAT3 and NF-κB

Psoriasis is a chronic autoimmune disease featured by patches on the skin.It is caused by malfunction of immune cells and keratinocytes with inflammation as one of its key features.Apigenin(API)is a natural flavonoid with anti-inflammatory and immunoregulatory properties.Therefore,we speculated that API can ameliorate psoriasis,and determined its effect on the development of psoriasis by using imiquimod(IMQ)-induced psoriasis mouse model.Our results showed that API attenuated IMQ-induced phenotypic changes,such as erythema,scaling and epidermal thickening,and improved splenic hyperplasia.Abnormal differentiation of immune cells was restored in API-treated mice.Mechanistically,we revealed that API is a key regulator of signal transducer activator of transcription 3(STAT3).API regulated immune responses by reducing interleukin-23(IL-23)/STAT3/IL-17A axis.Moreover,it suppressed IMQ-caused cell hyperproliferation by inactivating STAT3 through regulation of extracellular signal-regulated kinase 1/2 and nuclear factor-κB(NF-κB)pathway.Furthermore,API reduced expression of inflammatory cytokines through inactivation of NF-κB.Taken together,our study demonstrates that API can ameliorate psoriasis and may be considered as a strategy for psoriasis treatment.

Apigenin is an anoikis sensitizer with strong anti-metastatic properties in experimental breast cancer

Loss of susceptibility to anoikis signals is a crucial step in metastasis.Anoikis resistance therefore represents a promising adjuvant therapeutic target for cancer management.In this study,we have conducted a rationalized screening to search for novel leading anoikis sensitizer from daily foods.Among 19 tested dietary phytochemicals,the best results were obtained with apigenin,a natural component of celery.Phenotypically,apigenin sensitized breast cancer cells to anoikis,lowered the number of circulating tumor cells,and protected against breast cancer metastasis to lung in mice.Mechanistically,we demonstrated that the thromboxane A_(2)(TXA_(2))-TXA_(2)receptor(TP)axis has a critical role in acquired anoikis resistance by activating PI3K-Akt signaling pathway.Blockage of TXA_(2)signaling up-regulated p53 as well as its target gene p21,caused a G1 phase arrest,and finally led to apoptosis in breast cancer cells.TXA_(2)level was positively correlated with breast cancer cell anoikis rate,and apigenin significantly inhibited TXA_(2)biosynthesis in vitro and in vivo.Collectively,we identified apigenin as a potent anoikis sensitizer with anti-metastatic properties in a mouse model of breast cancer,and these findings might provide a rationale for introducing apigenin supplementation to breast cancer patients.

Apigenin, a natural flavonoid, promotes autophagy and ferroptosis in human endometrial carcinoma Ishikawa cells in vitro and in vivo

Apigenin,a natural flavonoid has been reported against a variety of cancer types.However,it is unclear whether apigenin can promote autophagy and ferroptosis in Ishikawa cells.There are few reports on the mechanism of apigenin on autophagy and ferroptosis of endometrial cancer Ishikawa cells.We found that iron accumulation,lipid peroxidation,glutathione consumption,p62,HMOX1,and ferritin were increased,while,solute carrier family 7 member 11 and glutathione peroxidase 4 were decreased.Ferrostatin-1,an iron-death inhibitor could reverse the effects of apigenin in Ishikawa cells.On the other hand,apigenin could promote autophagy via up-regulating Beclin 1,ULK1,ATG5,ATG13,and LC3B and down-regulating AMPK,mTOR,P70S6K,and ATG4.Furthermore,apigenin could inhibit tumor tissue proliferation and restrict tumor growth via ferroptosis in vivo.

Role of apigenin in high glucose-induced retinal microvascular endothelial cell dysfunction via regulating NOX4/p38 MAPK pathway in vitro

AIM:To investigate the retinoprotective role of Apigenin(Api)against high glucose(HG)-induced human retinal microvascular endothelial cells(HRMECs),and to explore its regulatory mechanism.METHODS:HRMECs were stimulated by HG for 48h to establish the in vitro cell model.Different concentrations of Api(2.5,5,and 10μmol/L)were applied for treatment.Cell counting kit-8(CCK-8),Transwell,and tube formation assays were performed to examine the effects of Api on the viability,migration,and angiogenesis in HG-induced HRMECs.Vascular permeability was evaluated by Evans blue dye.The inflammatory cytokines and oxidative stress-related factors were measured using their commercial kits.Protein expression of nicotinamide adenine dinucleotide phosphate(NADPH)oxidase 4(NOX4)and p38 mitogen-activated protein kinase(MAPK)was measured by Western blot.RESULTS:Api prevented HG-induced HRMECs viability,migration,angiogenesis,and vascular permeability in a concentration-dependent manner.Meanwhile,Api also concentration-dependently inhibited inflammation and oxidative stress in HRMECs exposed to HG.In addition,HG caused an elevated expression of NOX4,which was retarded by Api treatment.HG stimulation facilitated the activation of p38 MAPK signaling in HRMECs,and Api could weaken this activation partly via downregulating NOX4 expression.Furthermore,overexpression of NOX4 or activation of p38 MAPK signaling greatly weakened the protective role of Api against HG-stimulated HRMECs.CONCLUSION:Api might exert a beneficial role in HGstimulated HRMECs through regulating NOX4/p38 MAPK pathway.

Apigenin ameliorates diabetic neuropathy in rats by modulating the TLR4/MyD88 signaling pathway

Objective:To determine the neuroprotective effects of apigenin against streptozotocin(STZ)-induced diabetic neuropathy(DN).Methods:To induce DN,Wistar rats(150-200 g)were administered with STZ(55 mg/kg,i.p.).Then they were randomly assigned to various groups,viz.,normal,diabetic control,insulin(10 IU/kg,s.c.),apigenin(5,10,and 20 mg/kg,p.o.),and insulin(10 IU/kg)plus apigenin(20 mg/kg,p.o.).Various behavioral,biochemical,and molecular markers[tumor necrosis factor-alpha(TNF-α),interleukin(IL)-1β,IL-6,Toll-like receptor 4(TLR4),myeloid differentiation primary response 88(MyD88),and nuclear factor erythroid 2-related factor 2(Nrf2)]were assessed.Results:Apigenin(10 and 20 mg/kg,p.o.)substantially reduced plasma glucose levels,lipid profile,aspartate transaminase,alanine transaminase,glycated hemoglobin,and neural advanced glycation end products in STZ-induced DN rats(P<0.05).After apigenin intervention,STZ-induced changes in food and water intake,body weight,urine output,allodynia,hyperalgesia,and insulin levels were markedly improved(P<0.05).Neural antioxidant enzymes(superoxide dismutase and glutathione)and Na+K+ATPase activity were also considerably elevated(P<0.05)while the level of lipid peroxidation was diminished following apigenin therapy(P<0.05).Furthermore,apigenin markedly upregulated the Nrf2 mRNA level while downregulating the mRNA expressions of TNF-αand ILs and the protein expressions of TLR4 and MyD88(P<0.05).STZ-induced histological abnormalities in the sciatic nerve were also improved by apigenin treatment.Conclusions:Apigenin exerts its neuroprotective effect by modulating the inflammatory and oxidative stress pathways via regulating the TLR4-MyD88 signaling pathway.

Anti-diabetic potential of apigenin,luteolin,and baicalein via partially activating PI3K/Akt/GLUT-4 signaling pathways in insulin-resistant HepG2 cells

Dietary flavonoids are abundant in natural plants and possess multiple pharmacological and nutritional activities.In this study,apigenin,luteolin,and baicalein were chosen to evaluate their anti-diabetic effect in high-glucose and dexamethasone induced insulin-resistant(IR)HepG2 cells.All flavonoids improves the glucose consumption and glycogen synthesis abilities in IR-HepG2 cells via activating glucose transporter protein 4(GLUT4)and phosphor-glycogen synthase kinase(GSK-3β).These fl avonoids signifi cantly inhibited the production of reactive oxygen species(ROS)and advanced glycation end-products(AGEs),which were closely related to the suppression of the phosphorylation form of NF-κB and P65.The expression levels of insulin receptor substrate-1(IRS-1),insulin receptor substrate-2(IRS-2)and phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)pathway in IR-HepG2 cells were all partially activated by the fl avonoids,with variable effects.Furthermore,the intracellular metabolic conditions of the fl avonoids were also evaluated.

Apigenin对人乳腺癌细胞体外侵袭及诱导血管生成能力的影响

目的 证实Apigenin可抑制乳腺癌细胞侵袭能力及诱导血管生成的能力 ,从而达到抑制乳腺癌的作用。方法 建立肿瘤细胞体外侵袭模型和体外血管形成模型 ,观察乳腺癌细胞株ZR 75 3 0在Apigenin作用下其侵袭能力和诱导血管生成能力的改变 ;免疫细胞化学染色和RT PCR检测Apigenin对ZR 75 3 0细胞MMP 9基因蛋白表达的影响。结果 在Apigenin作用下 ,ZR 75 3 0细胞的侵袭能力和诱导血管生成的能力明显受到抑制 ,MMP 9基因和蛋白的表达下降。结论 Apigenin可能通过抑制MMP 9表达 ,进而抑制ZR 75 3 0细胞侵袭和诱导血管生成能力 ,达到抗乳腺癌的作用。

Apigenin/GCV对C_6-TK细胞株的杀伤效应及作用机制

目的探讨Apigenin/GCV对体外培养鼠胶质瘤C6细胞的杀伤效应及作用机制。方法对含有导入pcDNA3-TK质粒的C6细胞进行培养、传代:通过MTT法、TUNEL实验法及光镜技术检测GCV/Apigenin对检测不同组别C6细胞C6-TK细胞株的杀伤效应。结果给GCV+Apigenin后:转染组、空载体组和对照组中两两比较:转染组的细胞增殖水平显著低于空载体组和对照组(P<0.05);相对未处理组,用10μmol/LApigenin预先处理6h后,GCV对C6混合细胞的杀伤效应显著增强(P<0.01)。结论缝隙连接功能上调剂Apigenin可显著提高HSV-TK/GCV肿瘤自杀基因系统的"旁观者效应"及显著促凋亡作用。GCV的作用效能对细胞间通讯功能有一定的依赖性。

Autophagy inhibition enhances apigenin-induced apoptosis in human breast cancer cells

Apigenin （4＇,5,7-trihydroxyflavone） is a member of the flavone subclass of flavonoids present in fruits and vegetables. The involvement of autophagy in the apigenin-induced apoptotic death of human breast cancer cells was investigated. Cell proliferation and viability were assessed by 3-（4,5-dimethylthiazol- 2-yl）-2,5-diphenyltetrazolium bromide （MTT） and clonogenic assays. Flow cytometry, fluorescent staining and Western blot analysis were employed to detect apoptosis and autophagy, and the role of autophagy was assessed using autophagy inhibitors. Apigenin dose- and time-dependently repressed the proliferation and clonogenic survival of the human breast cancer T47D and MDA-MB-231 cell lines. The death of T47D and MDA-MB-231 cells was due to apoptosis associated with increased levels of Caspase3, PARP cleavage and Bax/Bcl-2 ratios. The results from flow cytometry and fluorescent staining also verified the occurrence of apoptosis. In addition, the apigenin-treated cells exhibited autophagy, as characterized by the appearance of autophagosomes under fluorescence microscopy and the accumulation of acidic vesicular organelles （AVOs） by flow cytometry. Furthermore, the results of the Western blot analysis revealed that the level of LC3-Ⅱ, the processed form of LC3-Ⅰ, was increased. Treatment with the autophagy inhibitor, 3-methyladenine （3-MA）, significantly enhanced the apoptosis induced by apigenin, which was accompanied by an increase in the level of PARP cleavage. Similar results were also confirmed by flow cytometry and fluorescence microscopy. These results indicate that apigenin has apoptosis- and autophagy-inducing effects in breast cancer cells. Autophagy plays a cyto-protective role in apigenin-induced apoptosis, and the combination of apigenin and an autophagy inhibitor may be a promising strategy for breast cancer control.

Anti-chikungunya activity of luteolin and apigenin rich fraction from Cynodon dactylon

Objective:To obtain Iuteolin and apigenin rich fraction from the ethanolic extract of Cynodon dactylon(L.)(C.dactylon) Pers and evaluate the fraction's cytotoxicity and anti-Chikungunya potential using Vero cells.Methods:The ethanolic extract of C.dactylon was subjected to silica gel column chromatography to obtain anti-chikungunya virus(CHIKV) fraction.Reverse phase-HPLC and GC-MS studies were carried out to identily the major phytochemicals in the fraction using phylochemical standards.Cytotoxicity and the potential of the fraction against CHIKV were evaluated in vitro using Vero cells.Reduction in viral replication was assessed by reverse transcriptase-polymerase chain reaction(RT-PCR) after treating the viral infected Vero cells with the fraction.Results:Reverse Phase-HPLC and GC-MS studies confirmed the presence of flavonoids,luteolin and apigenin as major phytochemicals in the anti-CHIKV ethanolic fraction of C.dactylon- The fraction was found to exhibit potent viral inhibitory activity(about 98%) at the concentration of 50 μg/mL as observed by reduction in cytopathic effect,and the cytotoxic concentration of the fraction was found to be 250 μg/mL.RT-PCR analyses indicated that the reduction in viral mRNA synthesis in fraction treated infected cells was much higher than the viral infected control cells.Conclusions:Luteolin and apigenin rich ethanolic fraction from C.dactylon can be utilized as a potential therapeutic agent against CHIKV infection as the fraction does not show cytotoxicity while inhibiting the virus.

Inhibitory effects of apigenin on the growth of gastric carcinoma SGC-7901 cells

AIM： To explore the growth inhibition and apoptosisinducing effect of apigenin on human gastric carcinoma SGC-7901 cells. METHODS： The effects of apigenin on the growth, clone formation and proliferation of human gastric carcinoma SGC-7901 cells were observed by N-rr, done-forming assay, and morphological observation. Fluorescent staining and flow cytometry analysis were used to detect apoptosis of cells. RESULTS： Apigenin obviously inhibited the growth, clone formation and proliferation of SGC-7901 cells in a dosedependent manner. Inhibition of growth was observed on d 1 at the concentration of 80 μmol/L, while after 4 d, the inhibition rate （IR） was 90%. The growth IRs at the concentration of 20, 40, and 80 μmol/L were 38%, 71%, and 99% respedJvely on the 7^th d. After the cells were treated with apigenin for 48 h, the number of clone-forming in control, 20, 40, and 80 μmol/L groups was 217±16.9, 170±11.1 （P〈0.05）, 98±11.1 （P〈0.05）, and 25±3.5 （P〈0.05） respectively. Typical morphological changes of apoptosis was found by fluorescent staining. The cell nuclei had lost its smooth boundaries, chromatin was condensed, and cell nuclei were broken. Flow cytometry detected typical apoptosis peak. After the cells were treated with apigenin for 48 h, the apoptosis rates were 5.76%, 19.17%, and 29.30% respectively in 20, 40, and 80 μmol/L groups. CONCLUSION： Apigenin shows obvious inhibition on the growth and clone formation of SGC-7901 cells by inducing apoptosis.

Mechanisms of Apigenin-7-glucoside As a Hepatoprotective Agent

Ixeris chinesis (Thunb.) Ankai has been used as a Chinese folk medicine, but only scanty information is available on the physiological and biochemical functions of the compounds extracted from I. chinesis. In the present study the effects of apigenin -7-glucoside (APIG) isolated from I. chinesis against liver injury caused by carbon tetrachloride (CCl4) were investigated. Methods The contents of malondialdehyde (MDA), glutamic pyruvic transaminase (GPT), glutamic oxaloacetic transaminase (GOT), and reduced glutathione (GSH) were evaluated by spectrophotography. The content of 8-Hydroxydeoxyguanosine (8-OHdG) was measured with high-performance liquid chromatography (HPLC) equipped with electrochemical and UV detection methods. The antioxidant activity of APIG was evaluated using chemiluminescence single photon counting technology. Results CCl4 significantly increased the enzyme activities of GPT and GOT in blood serum, as well as the level of MDA and 8-OHdG in liver tissue, and decreased the levels of GSH. Pretreatment with APIG was able not only to suppress the elevation of GPT, GOT, MDA and 8-OHdG, and inhibit the reduction of GSH in a dose-dependent manner in vivo, but also to reduce the damage of hepatocytes in vitro. On the other hand, we also found that APIG had strong antioxidant activity against reactive oxygen species (ROS) in vitro in a concentration-dependent manner. Conclusion The hepatoprotective activity of APIG is possibly due to its antioxidant properties, acting as scavengers of ROS. These results obtained in vivo and in vitro suggest that APIG has protective effects against hepatic oxidative injury induced by chemicals. Further studies on the pharmaceutical functions and immunological responses of APIG may help its clinical application.

VEGF EXPRESSION IS INHIBITED BY APIGENIN IN HUMAN BREAST CANCER CELLS

Objective： To study the effects of apigenin on vascular endothelial growth factor （VEGF） in human breast cancer cells （MDA-MB-231. Methods： MTT assay was used to detect the cell proliferation inhibitory effect of apigenin on MDA-MB-231 cell. ELISA was used to determine the protein level of VEGF secreted by MDA-MB-231 cells. RT-PCR was used to detect mRNA levels of VEGF in MDA-MB-231 cells. The protein levels of HIF-1α, p-AKT, p-ERK1/2, and p53 were detected by Western Blotting. Results： Apigenin did not inhibit the cell viability of MDA-MB-231 cell. Apigenin reduced the secretion and mRNA levels of VEGF in MDA-MB-231 cells. Additionally, apigenin decreased the expressions of HIF-1α, p-AKT and p-ERK1/2, but induced the expression of p53. Conclusion： Apigenin can inhibit VEGF expression in human breast cancer cells, and this may be achieved through decreasing HIF-1α.

Effect of apigenin on the reproductive system in male mice

This study aimed to characterize the effect of apigenin on the reproductive system in male mice. Adult male mice were treated with intraperitoneal injection of apigenin at the dose levels of 5, 10, 15, 20 and 25 mg/kg.bw, 0.05% DMSO and 0.9% normal saline daily for seven days. Then, testis and epididymis sperms in sperm motility, sperm morphology, the percentages of ploidy cells and seminiferous epithelium cells at the cell-circle phase, and the ratio of ploidy cells were evaluated. The results showed that sperm density significantly reduced in the 25 mg/kg group compared with the solvent control group. The abnormal sperms were mainly amorphous;non-hook sperms took the second largest group;and banana, double-tail and folded-tail sperms were rare. Abnormal sperms were mainly in the head sperm. Moreover, after intraperitoneal injection of 5 mg/kg apigenin, the percentage of 1C population increased, and the percentage of 4C declined, leading to a significant increase of the 1C:4C ratio, compared with the solvent and negative control groups. The percentage of seminiferous epithelium cells at the cell-circle phase of G0/G1 exhibited a significant increase in the 25 mg/kg group compared with the control groups. Taken together, that apigenin has adverse effects on the reproductive system in adult male mice is demonstrated.

Analysis of apigenin in <i>Blumea balsamifera</i>Linn DC. and its inhibitory activity against aldose reductase in rat lens

To investigate the therapeutic potentials of na- tural sources, stepwise polarity fractions of Blumea balsamifera were tested for their ability to inhibit aldose reductase (AR) activity in rat lenses. Of these, the ethyl acetate (EtOAc) fraction exhibited a unique AR inhibitory activity (IC50 value, 0.11 μg/mL). Apigenin was identified from the active EtOAc fraction and exhibited high AR inhibitory activity (IC50 value, 4.03 μM). The content of apigenin was measured in B. balsamifera (0.47 mg/g) by HPLC/UV analysis. Our result suggests that B. balsamifera could be a useful natural source for the development of a novel AR inhibitory agent against diabetic complications.

Prediction of Apigenin Targets in Lung Cancer Using Network Pharmacology and Molecular Docking

[Objectives]To elucidate the mechanism of action of apigenin against lung cancer.[Methods]First,drug-target pathways and networks were constructed to predict potential protein targets of apigenin and their main interactions with the drug.Then,the ingenuity pathway analysis(IPA)was carried out to identify enriched gene pathways and networks,and the candidate protein targets of apigenin were predicted using networks and pathways that overlapped between proteins associated with lung cancer and proteins targeted by apigenin.[Results]Docking studies with apigenin indicated that BCL-2,CASP9,CDK2,CYCLIND1,PI3K,NF-κB,Rb1,p53,and AKT are the top candidate targets of apigenin,suggesting that the drug acts against lung cancer by regulating proteins involved in molecular mechanisms of cancer,as well as small cell lung cancer,pancreatic adenocarcinoma,glucocorticoid receptor,and p53 signaling.It was also found that apigenin affects networks mainly involved in the cell cycle,cell-to-cell signaling and interactions,hematological system development and function,cell death and survival,and organismal injury and abnormalities.[Conclusions]This study shows that network pharmacology is useful in generating hypotheses about how apigenin exerts therapeutic effects in lung cancer,as well as in discovering new multitarget drugs against other complex diseases.

STUDY ON THE NON-COVALENT INTERACTION OF APIGENIN MOLECULARLY IMPRINTED POLYMERS BY SPECTROMETRY

The non-covalent interaction between apigenin (API) and different functional monomers (α-methylacrylic acid (MAA), acrylamide (AM), 2-vinylpyridine (2-Vpy) and combined functional monomers (AM/2-Vpy)) was determined by UV spectrometry, and a series of apigenin molecularly imprinted polymers (API-MIPs) was synthesized with different functional monomers through molecular imprinting technology. The relationship between the non-covalent interaction of template/functional monomer and absorption of MIPs also was studied. The results showed that the order of the strength of the non-covalent interaction between API and different functional monomers in tetrahydrofuran (THF) is as follows: 2-Vpy> AM/2-Vpy>AM>MAA, which is positive correlation to the absorption capability of corresponding MIPs, and 2-Vpy is the optimum functional monomer among the used monomer for preparing API- MIPs.

Apigenin对奥氮平诱导肥胖大鼠模型的影响

目的观察Apigenin对奥氮平诱导肥胖大鼠模型的影响。方法将40只健康雄性SD大鼠随机分为4组:对照组(0.5%CMC-Na 1.0 mL·100 g^(-1))、模型组(奥氮平1.2 mg·kg^(-1))、高剂量组(Apigenin 200 mg·kg^(-1)+奥氮平1.2 mg·kg^(-1))、低剂量组(Apigenin 100 mg·kg^(-1)+奥氮平1.2 mg·kg^(-1))。连续灌胃35 d,药物处理期间,每隔2 d测1次体质量,奥氮平组大鼠体质量超过对照组20%为模型建立成功。计算Lee’s指数、脏器系数和脂肪系数;测定血浆总胆固醇(TC)、甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)、空腹血糖(FBS); HE染色检测肾周脂肪组织形态学改变。结果与正常组比较,模型组大鼠体质量、Lee’s指数、脂肪系数均明显增加,血浆中FBS、TG、LDL-C、TC水平显著升高,HDL-C水平显著下降,脂肪细胞体积明显增大,差异均有统计学意义(碷P<0.05)。与模型组比较,Apigenin高、低剂量组均可降低大鼠体质量、Lee’s指数、脂肪系数,呈剂量依赖性,显著降低血浆FBS、TG、LDL-C及TC水平,升高HDL-C水平,并减小脂肪细胞体积。除Apigenin低剂量组对体质量的影响无统计学意义外,其他差异均具有统计学意义(碷P<0.05)。结论 Apigenin对奥氮平诱导的大鼠肥胖及代谢紊乱有一定的改善作用。