Casticin Treatment Research Articles

Anti-osteoporosis activity of casticin in ovariectomized rats.

Postmenopausal osteoporosis (PMPO) is the most familiar type of osteoporosis, a silent bone disease. Casticin, a natural flavonoid constituent, improves osteoporosis in animal model. Nevertheless, the potential mechanism remains to be further explored. A model of PMPO was established in rats treated with ovariectomy (OVX) and RAW 264.7 cells induced with receptor activator of nuclear factor kappa-B ligand (RANKL). The effect and potential mechanism of casticin on PMPO were addressed by pathological staining, measurement of bone mineral density (BMD), three-point bending test, serum biochemical detection, filamentous-actin (F-actin) ring staining, TRAcP staining, reverse transcription quantitative polymerase chain reaction, western blot and examination of oxidative stress indicators. The casticin treatment increased the femoral trabecular area, bone maturity, BMD, elastic modulus, maximum load, the level of calcium and estrogen with the reduced concentrations of alkaline phosphatase (ALP) and tumor necrosis factor (TNF)-α in OVX rats. An enhancement in the F-actin ring formation, TRAcP staining and the relative mRNA expression of NFATc1 and TRAP was observed in RANKL-induced RAW 264.7 cells, which was declined by the treatment of casticin. Moreover, the casticin treatment reversed the reduced the relative protein expression of Nrf2 and HO-1 and the concentrations of superoxide dismutase and glutathione peroxidase, and the increased content of malondialdehyde both in vivo and in vitro. Casticin improved bone density, bone biomechanics, the level of calcium and estrogen, the release of pro-inflammatory factor and oxidative stress to alleviate osteoporosis, which was associated with the upregulation of Nrf2/HO-1 pathway.

Casticin promotes osteogenic differentiation of bone marrow stromal cells and improves osteoporosis in rats by regulating nuclear factor-κB/mitogen-activated protein kinase.

Osteoporosis has influenced millions of people, especially postmenopausal women, which has become a big burden to the whole world. Although the diverse roles of casticin (CAS) on different diseases were identified, whether it was implicated with osteoporosis was unknown. A rat model of osteoporosis was established through dexamethasone (DEX) treatment and a cell model reflecting the osteogenic and osteoclast induction was constructed in bone marrow stromal cells (BMSCs). The calcification at the late stage of induction was measured via Alizarin Red S staining. Western blot was applied to evaluate the levels of proteins. Hematoxylin and eosin staining revealed that the number of bone trabecular in DEX-induced osteoporosis rats was decreased, while increased doses of CAS treatment elevated the number of bone trabecular. CAS treatment alleviated DEX-induced osteoporosis in rats. Moreover, we found that CAS inhibited the nuclear factor-κB/mitogen-activated protein kinase (NF-κB/MAPK) pathway. In addition, CAS promoted osteogenic differentiation of BMSCs and reduced osteoclastogenesis of bone marrow monocytes. Finally, CAS was observed to retard the receptor activator of NFκ-B ligand-induced NF-κB/MAPK pathway. CAS promoted osteogenic differentiation of BMSCs and improved osteoporosis in rats by regulating the NF-κB/MAPK pathway. This might shed a light into using CAS as a drug treating osteoporosis in the future.

MiR-338-3p Plays a Significant Role in Casticin-Induced Suppression of Acute Myeloid Leukemia via Targeting PI3K/Akt Pathway.

Objective Casticin is generally used in traditional herbal medicine for its anti-inflammatory and anticarcinogenic pharmacological properties. Also, microRNAs are indispensable oncogenes or cancer suppressors being dysregulated in various diseases. In this study, we aimed to elucidate the mechanisms underlying effects of casticin on the progression of acute myeloid leukemia (AML). Methods CCK-8 and flow cytometry were utilized to measure the proliferation and apoptosis of AML cell lines, respectively, after treatment with different concentrations of casticin. The alteration of several microRNA expressions in response to casticin treatment was detected by performing qRT-PCR, and the activity of PI3K/Akt pathways was evaluated through immunoblotting. Afterwards, the potential target gene of miR-338-3p was investigated by dual-luciferase reporter assay. In order to evaluate the role of miR-338-3p in the casticin-induced cellular phenotype changes, AML cells were transfected with miR-338-3p mimics or inhibitor and then subjected to proliferation and apoptosis analysis. Finally, a mouse xenograft model system was employed to investigate the role of casticin in AML progression in vivo. Results Suppressed cellular proliferation and enhanced apoptosis were observed in HL-60 and THP-1 cells after exposure to casticin, accompanied by remarkable upregulation of the miR-338-3p expression as well as a decline in the phosphorylation of PI3K and Akt proteins. RUNX2 was identified as a direct target molecular of miR-338-3p, which might account for the findings that miR-338-3p knockdown enhanced the PI3K/Akt pathway activity, whereas the miR-338-3p overexpression inactivated this signaling pathway. In addition, the inhibition of the miR-338-3p expression attenuated severe cell apoptosis and suppressions of PI3K/Akt pathway induced by casticin. Furthermore, casticin treatment retarded tumor growth rate in mouse models, whilst elevating miR-338 expression and repressing the activity of PI3K/Akt pathway in vivo. However, miR-338-3p depletion could also abolish the phenotypic alterations caused by casticin treatment. Conclusion Casticin promotes AML cell apoptosis but inhibits AML cell proliferation in vitro and tumor growth in vivo by upregulating miR-338-3p, which targets RUNX2 and thereafter inactivates PI3K-Akt signaling pathway. Our results provide insights into the mechanisms underlying the action of casticin in the control of AML progression.

Antifibrotic and anthelminthic effect of casticin on Schistosoma mansoni-infected BALB/c mice.

Schistosomiasis is an important tropical disease caused by Schistosoma. Although the pathogenesis of liver fibrosis has been intensively studied, the choice of effective treatment is still inadequate. In this study, we aimed to investigate the potential of using Casticin to treat Schistosoma mansoni-induced liver fibrosis. BALB/c mice were divided into three groups - control, infection, and treatment group. The infection and treatment group were percutaneously infected with 100-120 cercariae. Mice from the treatment group were treated with 20mg/kg/day Casticin for 14 consecutive days to investigate the potential protective effects of Casticin. Mice were sacrificed and were used for histological, RNA, protein, and parasite burden analysis. Our results showed that hepatic fibrosis was significantly attenuated, as indicated by histology and reduction of fibrotic markers such as collagen AI, transforming growth factor β (TGF-β), and α-smooth muscle actin (α-SMA). Furthermore, Casticin treatment significantly reduced worm burden. Anthelmintic effect of Casticin was also observed by scanning electron microscopy. Collectively, our study suggested that Casticin may be a beneficial candidate in treating S. mansoni infection.

Combinational treatment of 5-fluorouracil and casticin induces apoptosis in mouse leukemia WEHI-3 cells in vitro.

Leukemia is one of the major diseases causing cancer-related deaths in the young population, and its cure rate is unsatisfying with side effects on patients. Fluorouracil (5-FU) is currently used as an anticancer drug for leukemia patients. Casticin, a natural polymethoxyflavone, exerts anticancer activity against many human cancer cell lines in vitro, but no other reports show 5-FU combined with casticin increased the mouse leukemia cell apoptosis in vitro. Herein, the antileukemia activity of 5-FU combined with casticin in WEHI-3 mouse leukemia cells was investigated in vitro. Treatment of two-drug combination had a higher decrease in cell viability and a higher increase in apoptotic cell death, the level of DNA condensation, and the length of comet tail than that of 5-FU or casticin treatment alone in WEHI-3 cells. In addition, the two-drug combination has a greater production rate of reactive oxygen species but a lower level of Ca2+ release and mitochondrial membrane potential (ΔΨm ) than that of 5-FU alone. Combined drugs also induced higher caspase-3 and caspase-8 activities than that of casticin alone and higher caspase-9 activity than that of 5-FU or casticin alone at 48 hours treatment. Furthermore, 5-FU combined with casticin has a higher expression of Cu/Zn superoxide dismutase (SOD [Cu/Zn]) and lower catalase than that of 5-FU or casticin treatment alone. The combined treatment has higher levels of Bax, Endo G, and cytochrome C of proapoptotic proteins than that of casticin alone and induced lower levels of B-cell lymphoma 2 (BCL-2) and BCL-X of antiapoptotic proteins than that of 5-FU or casticin only. Furthermore, the combined treatment had a higher expression of cleaved poly (ADP-ribose) polymerase (PARP) than that of casticin only. Based on these findings, we may suggest that 5-FU combined with casticin treatment increased apoptotic cell death in WEHI-3 mouse leukemia cells that may undergo mitochondria and caspases signaling pathways in vitro.

Casticin elicits inflammasome-induced pyroptosis through activating PKR/JNK/NF-κB signal in 5-8F cells

Casticin is one of the effective ingredients of fructus viticis. Most studies have shown that casticin has a strong anti-proliferation activity against various tumor cells. However, its anti-tumor effect and molecular mechanism in nasopharyngeal carcinoma remain unclear. In this study, we demonstrated that the casticin selectively inhibited the proliferation of 5–8F cells in vitro. Further analysis revealed that casticin treatment significantly increased sub-G2 phase and incited pyroptotic process. Moreover, we demonstrated that PKR participated in in regulating the process of GSDMD-dependent pyroptotic tumor cell death. PKR knockdown alleviated the activation of JNK pathway and the expression of its downstream proteins, including cleaved caspase-1, GSDMD-N, interleukin-1β. These findings indicate that PKR/JNK/NF-κB signal is essential for casticin-induced caspase-1 inflammasome formation and inflammatory cytokines release in 5–8F cell.

Casticin Attenuates Osteoarthritis-Related Cartilage Degeneration by Inhibiting the ROS-Mediated NF-κB Signaling Pathway in vitro and in vivo.

Casticin, a flavonoid isolated from Vitex trifolia, has been shown to have anti-inflammatory and antitumor effects in previous studies. Osteoarthritis (OA) is a disease based on degenerative pathological changes. The disease process is often accompanied by inflammatory pathological changes. However, there is no safe and effective drug for prevention and treatment. In the present study, we aimed to clarify the role of casticin in the murine model of destabilization of the medial meniscus (DMM). Male BALB/c mice were randomly divided into three groups: Sham, DMM-induced OA treated with vehicle, and DMM-induced OA treated with casticin. Our results indicated that the casticin treatments markedly reduced the destruction of cartilage and OARSI grades compared with those of the vehicle-treated mice. The levels of matrix metalloproteinase-13 (MMP13) in cartilage were also significantly reduced in the casticin-treated mice. Casticin also significantly regulated oxidative stress and reduced inflammation in the cartilage of mice with OA. These results suggest that casticin prevents the development of posttraumatic OA in mice. Consequently, decreased reactive oxygen species levels and suppressed proinflammatory cytokine production were confirmed in casticin-treated IL-1β-stimulated ADTC5 cells. After casticin treatment, the NF-κB signaling pathway was significantly inhibited in the cells. It can be concluded that casticin can alleviate arthritis-related cartilage degeneration by inhibiting ROS-mediated NF-κB signaling pathway in vitro and in vivo.

Casticin: A Promising Candidate to Develop a Stem Cell Targeted Strategy in AML Treatment

Purpose: Acute myeloid leukemia is the most common form of acute leukemia with genetic and epigenetic heterogeneity. Although current therapeutic agents provide successful remission, 5-year survival rates are still low. Insufficiency of targeting leukemia stem cells is considered as the main obstacle that causes drug resistance and relapse. Phytochemicals remain a promising source for targeted drug research. Studies showed that Casticin has antiproliferative effects on leukemic cells, but its effects on leukemic stem cells are still unclear. In this study, we aimed to investigate the antiproliferative capacity of Casticin on acute myeloid leukemia stem-like KG1a cells and its relatively mature parental KG1 cells in comparison with healthy peripheral blood mononuclear cells PBMC . Method: The antiproliferative effects of Casticin on cells and IC50 values were determined by MTT test. The effects of Casticin on caspase 3/7 activity, apoptosis and necrosis in cells were evaluated by flow cytometry and TUNEL assays. Results: 2 μM Casticin treatment for 24 h was increased apoptotic cell death and caspase 3/7 activation in KG1 27.2%; 17.30%; p

Casticin inhibits nasopharyngeal carcinoma growth by targeting phosphoinositide 3-kinase

BackgroundCasticin, an isoflavone compound extracted from the herb Fructus Viticis, has demonstrated anti-inflammatory and anticancer activities and properties. The aim of this study was to investigate the effects and mechanisms of casticin in nasopharyngeal carcinoma (NPC) cells and to determine its potential for targeted use as a medicine.MethodsNPC cells were used to perform the experiments. The CCK‑8 assay and colony formation assays were used to assess cell viability. Flow cytometry was used to measure the cell cycle and apoptosis analysis (annexin V/PI assay). A three-dimensional (3D) tumour sphere culture system was used to characterize the effect of casticin on NPC stem cells. In silico molecular docking prediction and high-throughput KINOME scan assays were used to evaluate the binding of casticin to phosphoinositide 3-kinase (PI3K), including wild-type and most of mutants variants. We also used the SelectScreen assay to detect the IC50 of ATP activity in the active site of the target kinase. Western blotting was used to evaluate the changes in key proteins involved cell cycle, apoptosis, stemness, and PI3K/protein kinase B (AKT) signalling. The effect of casticin treatment in vivo was determined by using a xenograft mouse model.ResultsOur results indicate that casticin is a new and novel selective PI3K inhibitor that can significantly inhibit NPC proliferation and that it induces G2/GM arrest and apoptosis by upregulating Bax/BCL2 expression. Moreover, casticin was observed to affect the self-renewal ability of the nasopharyngeal carcinoma cell lines, and a combination of casticin with BYL719 was observed to induce a decrease in the level of the phosphorylation of mTORC1 downstream targets in BYL719-insensitive NPC cell lines.ConclusionCasticin is a newly emerging selective PI3K inhibitor with potential for use as a targeted therapeutic treatment for nasopharyngeal carcinoma. Accordingly, casticin might represent a novel and effective agent against NPC and likely has high potential for combined use with pharmacological agents targeting PI3K/AKT.

Casticin inhibits breast cancer cell migration and invasion by down-regulation of PI3K/Akt signaling pathway.

Casticin is one of the major active components isolated from Fructus viticis. Increasing studies have revealed that casticin has potential anticancer activity in various cancer cells, but its effects on breast cancer cell migration and invasion are still not well known. Therefore, the ability of cell migration and invasion in the breast cancer MDA-MB-231 and 4T1 cells treated by casticin was investigated. The results indicated that casticin significantly inhibited cell migration and invasion in the cells exposed to 0.25 and 0.50 µM of casticin for 24 h. Casticin treatment reduced matrix metalloproteinase (MMP) 9 (MMP-9) activity and down-regulated MMP-9 mRNA and protein expression, but not MMP-2. Casticin treatment suppressed the nuclear translocation of transcription factors c-Jun and c-Fos, but not nuclear factor-κB (NF-κB), and decreased the phosphorylated level of Akt (p-Akt). Additionally, the transfection of Akt overexpression vector to MDA-MB-231 and 4T1 cells could up-regulate MMP-9 expression concomitantly with a marked increase in cell invasion, but casticin treatment reduced Akt, p-Akt, and MMP-9 protein levels and inhibited the ability of cell invasion in breast cancer cells. Additionally, casticin attenuated lung metastasis of mouse 4T1 breast cancer cells in the mice and down-regulated MMP-9 expression in the lung tissues of mice treated by casticin. These findings suggest that MMP-9 expression suppression by casticin may act through inhibition of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, which in turn results in the inhibitory effects of casticin on cell migration and invasion in breast cancer cells. Therefore, casticin may have potential for use in the treatment of breast cancer invasion and metastasis.

Casticin protects against IL-1β-induced inflammation in human osteoarthritis chondrocytes

Casticin, an active compound isolated from Vitex rotundifolia L., was reported to possess anti-inflammatory activity. However, the effect of casticin on inflammatory response in human osteoarthritis (OA) chondrocytes remains unclear. In the current study, we examined the anti-inflammatory effects of casticin on chondrocytes exposed to interleukin-1β (IL-1β). Our results demonstrated that casticin treatment significantly improved cell viability in chondrocytes exposed to IL-1β. Casticin significantly inhibited IL-1β-induced NO and PGE2 production, and iNOS and COX-2 expression in human OA chondrocytes. It also suppressed the levels of TNF-α and IL-6, as well as decreased production of MMP-3, MMP-13, ADAMTS-4 and ADAMTS-5 in IL-1β-stimulated chondrocytes. Furthermore, casticin prevented IL-1β-induced NF-κB activation in chondrocytes. Taken together, these findings showed that casticin attenuates inflammatory responses in chondrocytes stimulated with IL-1β, possibly through the NF-κB signaling pathway. Thus, casticin may serve as a potential anti-inflammatory agent in the treatment of OA.

Casticin prevents DSS induced ulcerative colitis in mice through inhibitions of NF-κB pathway and ROS signaling.

Casticin, a compound purified from the Chinese herb Viticis Fructus, has been proven effective in preventing tumor progression in previous studies. Ulcerative colitis (UC) is a common inflammatory bowel disease that affects millions of people worldwide, but no effective and safe drugs are available. In this study, we aimed to study how did casticin affect UC by evaluating its effects on dextran sulfate sodium (DSS)-induced colitis in mice. Our data suggested that casticin attenuated body weight loss, colon length shortening, and pathological damage in the colon of DSS-treated mice. Casticin decreased reactive oxygen species level and chemocytokines (IL-1β, IL-6, TNF-α) productions in colon tissue. The decreased reactive oxygen species level and suppressed proinflammatory cytokines productions were also confirmed in casticin-treated LPS-stimulated RAW264.7 cells and hydrogen peroxide-treated CACO-2 cells in vitro. Mechanistically, casticin treatment prevented the profound activation of AKT signaling caused by DSS administration. And casticin inhibited the productions of proinflammatory chemocytokines through downregulating AKT/NF-κB pathway in macrophages. Meanwhile, data revealed that casticin increased expressions of endogenous antioxidants peroxiredoxin 3 and MnSOD were through activation in FOXO3α signaling by downregulating AKT signaling in colon epithelium cells. Our findings demonstrated that casticin alleviated DSS-induced UC by increasing the antioxidant enzyme peroxiredoxin 3 and MnSOD expressions, and decreasing the production of proinflammatory chemocytokines through inhibition of AKT signaling.

Casticin alleviates lipopolysaccharide-induced inflammatory responses and expression of mucus and extracellular matrix in human airway epithelial cells through Nrf2/Keap1 and NF-κB pathways.

Asthma is one of the most common chronic inflammatory diseases of childhood, characterized by airway inflammation, mucus hypersecretion, and accumulation of extracellular matrix proteins. Casticin is an active compound that possesses broad biological activities including anti-inflammatory effect. However, the effect of casticin on asthma remains unknown. The aim of the present study was to evaluate the effect and mechanism of casticin on inflammatory responses and expression of mucus and extracellular matrix in human airway epithelial cells. The results showed that lipopolysaccharide induced the mRNA and protein levels of IL-6, IL-8, MUC5AC, collagen type I, and fibronectin in 16-HBE cells, whereas casticin treatment significantly inhibited the induction of lipopolysaccharide. Casticin induced Nrf2/Keap1 and inhibited nuclear factor κB pathways in 16-HBE cells. Knockdown of Nrf2 attenuated the effect of casticin on production of IL-6 and IL-8, expression of MUC5AC, collagen type I, and fibronectin in 16-HBE cells. In conclusion, the results indicated that casticin might be a novel therapeutic strategy for the treatment of asthma.

Casticin induces apoptosis and G0/G1 cell cycle arrest in gallbladder cancer cells

BackgroundCasticin, the flavonoid extracted from Vitex rotundifolia L, exerts various biological effects, including anti-inflammatory and anti-cancer activity. The aim of this study is to investigate the effects and mechanisms of casticin in human gallbladder cancer cells.MethodsHuman NOZ and SGC996 cells were used to perform the experiments. CCK-8 assay and colony formation assay were performed to evaluate cell viability. Cell cycle analyses and annexin V/PI staining assay for apoptosis were measured using flow cytometry. Western blot analysis was used to evaluate the changes in protein expression, and the effect of casticin treatment in vivo was experimented with xenografted tumors.ResultsIn this study, we found that casticin significantly inhibited gallbladder cancer cell proliferation in a dose- and time-dependent manner. Casticin also induced G0/G1 arrest and mitochondrial-related apoptosis by upregulating Bax, cleaved caspase-3, cleaved caspase-9 and cleaved poly ADP-ribose polymerase expression, and by downregulating Bcl-2 expression. Moreover, casticin induced cycle arrest and apoptosis by upregulating p27 and downregulating cyclinD1/cyclin-dependent kinase4 and phosphorylated protein kinase B. In vivo, casticin inhibited tumor growth.ConclusionCasticin induces G0/G1 arrest and apoptosis in gallbladder cancer, suggesting that casticin might represent a novel and effective agent against gallbladder cancer.

Casticin inhibits the activity of transcription factor Sp1 and the methylation of RECK in MGC803 gastric cancer cells.

The present study investigated the effect of casticin on reversion-inducing-cysteine-rich protein with kazal motifs (RECK) gene expression and intracellular methylation levels in MGC803 gastric cancer cells. Cells were treated with 1, 10 and 30 µmol/l casticin. Western blotting and reverse transcription-quantitative polymerase chain reaction assays were performed to determine the protein expression and mRNA levels of RECK and DNA methyltransferase 1 (DNMT1), respectively. High-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry was used to detect RECK methylation. In addition, MGC803 cell proliferation was measured by an MTT assay and the DNA-binding activity of transcription factor Sp1 was determined using an enzyme-linked immunosorbent assay. The results demonstrated that treatment with 1, 10 and 30 µmol/l casticin significantly increased RECK protein expression and mRNA levels. In addition, casticin (30 µmol/l) decreased RECK promoter methylation levels by 31%, global DNA methylation levels by 39% and nuclear methylation activity by 71.6%. Furthermore, casticin downregulated the mRNA levels and protein expression of DNMT1. The MTT assay demonstrated that MGC803 cell proliferation was inhibited by casticin treatment and DNA binding assays indicated that casticin reduced the DNA-binding activity of Sp1. The present study therefore indicated that casticin inhibits the proliferation of gastric cancer MGC803 cells by upregulating RECK gene expression and reducing intracellular methylation levels.

RIP kinase-mediated ROS production triggers XAF1 expression through activation of TAp73 in casticin-treated bladder cancer cells.

The p53 family protein p73 plays an important role in apoptosis induced by chemotherapeutic drugs. Transcriptionally active (TA) p73 (TAp73) substitutes for p53 in the response to stress. XIAP associated factor 1 (XAF1) is a novel predictive and prognostic factor in patients with bladder cancer, but the association between TAp73 and XAF1 expression in bladder cancer cells is poorly understood. Here, we investigated the status of TAp73 and XAF1 in T24 bladder cancer cells to identify molecular mechanisms in casticin‑exposed T24 cells. Casticin induced activation of JNK/p38 MAPK that preceded activation of the caspase cascade and disruption of the mitochondria membrane potential (∆ψm). Expression of XAF1 and TAp73 was also upregulated in casticin-treated T24 cells. Casticin treatment of T24 cells induced receptor-interacting protein (RIP) kinase expression and increased intracellular production of reactive oxygen species (ROS). Casticin-mediated ROS induced an increase in phosphorylated JNK/p38 MAPK, resulting in progressive upregulation of TAp73, which in turn led to XAF1 expression. Our data suggest that the apoptotic activity of casticin in T24 cells is mediated by activation of the TAp73-XAF1 signaling pathway through RIP kinase-mediated ROS production.

Casticin Inhibits A375.S2 Human Melanoma Cell Migration/Invasion through Downregulating NF-κB and Matrix Metalloproteinase-2 and -1.

Casticin is one of the main components from Fructus Viticis, which is widely used as an anti-inflammatory agent. The mechanism of how casticin affects melanoma cell migration and invasion is still not well known. Here we studied the anti-metastasis effects of casticin on A375.S2 melanoma cells by using a non-lethal concentration. First; we used an adhesion assay to test the A375.S2 cells’ adhesion ability after treatment with casticin. We next investigated the cell migration ability after casticin treatment by using a wound healing assay to prove that the migration of A375.S2 cells can be inhibited by casticin and double checked the results using the transwell-migration assay. The suppressive effects on matrix metalloproteinase-2; and -9 (MMP-2; and -9) activities were examined by gelatin zymography. Furthermore, western blotting was used to investigate the protein level changes in A375.S2 cells. We found that p-EGFR; Ras and p-ERK1/2 are decreased by casticin, indicating that casticin can down-regulate the migration and invasion ability of A375.S2 cells via the p-EGFR/Ras/p-ERK pathway. The NF-κB p65 and p-ERK levels in nuclear proteins are also decreased by treatment with casticin. An EMSA assay also discovered that the NF-κB p65 and DNA interaction is decreased. NF-κB p65 protein level was examined by immunofluorescence staining and also decreased. Our findings suggest that casticin has anti-metastatic potential by decreasing the invasiveness of A375.S2 cells. We also found that casticin suppressed A375.S2 cell proliferation and cell adhesion ability, but did not affect cell death, as examined using cytometry and a collagen adhesion assay. Based on these observations, casticin could be used as an inhibitor of migration and invasion of human melanoma cells in the future.

Casticin treatment inhibits squamous cell carcinoma cell proliferation and arrests cell cycle in S phase

<p class="Abstract">The present study demonstrates the effect of casticin on esophageal squamous cell carcinoma cell lines, TE-1 and TE-15. The cells were treated with various concentrations (10-50 μM) of casticin for different time periods. The results revealed that casticin treatment significantly inhibited the rate of cell proliferation in both TE-1 and TE-15 cell lines after 48 hours. Casticin treatment induced cell cycle arrest in S phase, enhanced the expression of proapoptotic gene, Bax and activation of caspase-3. Moreover, the morphological features of the cells were altered resulting in apoptosis. Casticin also inhibited the migration potential of TE-1 cells. Thus, casticin exhibits inhibitory effect on the esophageal squamous cell carcinoma cell lines by inhibiting cell proliferation, arresting cell cycle, inducing apoptosis and inhibiting migration. Therefore, casticin can be of therapeutic importance for the treatment of esophageal squamous cell carcinoma.</p><p> </p>

Activation of the apoptosis signal-regulating kinase 1/c-Jun N-terminal kinase pathway is involved in the casticin-induced apoptosis of colon cancer cells.

Casticin is one of the main components of the fruits of Vitex rotundifolia L. Studies have shown that casticin inhibits the growth of various cancer cells, including colon cancer. In the present study, the anti-carcinogenic effects of casticin on human colon cancer and the underlying mechanisms were investigated. The results revealed that casticin significantly induced apoptosis of HT-29, HCT-116, SW480 and Caco-2 cells, induced the accumulation of reactive oxygen species (ROS) and increased the protein levels of apoptosis signal-regulating kinase 1 (ASK1), c-Jun N-terminal kinase (JNK) and B-cell lymphoma 2-interacting mediator of cell death (Bim) in HT-29 cells. Pretreatment with N-acetylcysteine, an antioxidant chemical compound, inhibited the activation of ASK1, JNK and Bim, as well as the apoptosis induced by casticin. Small interfering RNA targeting ASK1 significantly attenuated the induction of JNK and Bim activation and apoptotic cell death by casticin treatment. SP600125, a specific JNK inhibitor, attenuated Bim activation and apoptosis, but did not alter ASK1 phosphorylation levels. In addition, casticin treatment resulted in apoptosis by the same mechanism in HCT-116, SW480 and Caco-2 cells. These results suggest that casticin significantly induced apoptosis by the activation of the ASK1-JNK-Bim signaling cascade and the accumulation of ROS in colon cancer cells.

Casticin induces breast cancer cell apoptosis by inhibiting the expression of forkhead box protein M1.

Casticin is an active ingredient derived from Fructus Viticis, a traditional Chinese medicine. This study aimed to investigate the role of forkhead box O3 (FOXO3a) in breast cancer cells and examine the regulatory mechanisms of FOXO3a in response to casticin treatment of the cells by ELISA, flow cytometry, small interfering RNA (siRNA) transfection and western blot analysis. Casticin treatment induced apoptosis and reduced the expression of the transcription factor forkhead box protein M1 (FOXM1). In addition, FOXM1 repression induced by casticin treatment was associated with the activation of FOXO3a via increased dephosphorylation. Notably, silencing FOXO3a expression by siRNA-mediated gene knockdown attenuated casticin-mediated apoptosis. Collectively, these findings suggest that FOXO3a is a critical mediator of the inhibitory effects of casticin on apoptosis in breast cancer cells.