Honokiol Suppresses Cell Proliferation and Tumor Migration through ROS in Human Anaplastic Thyroid Cancer Cells.

BACKGROUND: Honokiol is a natural polyphenolic compound extracted from Magnolia officinali, which is commonly used material in Chinese herbal medicine, has a variety of biological functions, including anti-tumor, anti-oxidant, anti-inflammation, anti-microbial and anti-allergy. Although honokiol has numerous beneficial effects on human diseases, the underlying mechanisms of tumor metastasis are still unclear. Previously, we reported that honokiol suppresses thyroid cancer cell proliferation with cytotoxicity through cell cycle arrest, apoptosis, and dysregulation of intracellular hemostasis. Herein, we hypothesized that the antioxidant effect of honokiol might play a critical role in thyroid cancer cell proliferation and migration. METHODS: The cell viability assays, cellular reactive oxygen species (ROS) activity, cell migration, and immunoblotting were performed after cells were treated with honokiol. RESULTS: Based on this hypothesis, we first demonstrated that honokiol suppresses cell proliferation in two human anaplastic thyroid carcinoma (ATC) cell lines, KMH-2 and ASH-3, within a dosage- and time-dependent manner by cell counting kit-8 (CCK-8) assay. Next, we examined that honokiol induced ROS activation and could be suppressed by pre-treated with an antioxidant agent, N-acetyl-l-cysteine (NAC). Furthermore, the honokiol suppressed cell proliferation can be rescued by pre-treated with NAC. Finally, we demonstrated that honokiol inhibited ATC cell migration by modulating epithelial-mesenchymal transition (EMT)-related markers by Western blotting. CONCLUSION: Taken together, we provided the potential mechanism for treating ATC cells with honokiol, which significantly suppresses tumor proliferation and inhibits tumor metastasis in vitro through reactive oxygen species (ROS) induction.

Piperlongumine, a Potent Anticancer Phytotherapeutic, Induces Cell Cycle Arrest and Apoptosis In Vitro and In Vivo through the ROS/Akt Pathway in Human Thyroid Cancer Cells.

Thyroid cancer (TC) is the most common endocrine malignancy, and its global incidence has steadily increased over the past 15 years. TC is broadly divided into well-differentiated, poorly differentiated, and undifferentiated types, depending on the histological and clinical parameters. Thus far, there are no effective treatments for undifferentiated thyroid cancers or advanced and recurrent cancer. Therefore, the development of an effective therapeutic is urgently needed for such patients. Piperlongumine (PL) is a naturally occurring small molecule derived from long pepper; it is selectively toxic to cancer cells by generating reactive oxygen species (ROS). In this study, we demonstrate the potential anticancer activity of PL in four TC cell lines. For this purpose, we cultured TC cell lines and analyzed the following parameters: Cell viability, colony formation, cell cycle, apoptosis, and cellular ROS induction. PL modulated the cell cycle, induced apoptosis, and suppressed tumorigenesis in TC cell lines in a dose- and time-dependent manner through ROS induction. Meanwhile, an intrinsic caspase-dependent apoptosis pathway was observed in the TC cells under PL treatment. The activation of Erk and the suppression of the Akt/mTOR pathways through ROS induction were seen in cells treated with PL. PL-mediated apoptosis in TC cells was through the ROS-Akt pathway. Finally, the anticancer effect and safety of PL were also demonstrated in vivo. Our findings indicate that PL exhibits antitumor activity and has the potential for use as a chemotherapeutic agent against TC. This is the first study to show the sensitivity of TC cell lines to PL.

Honeysuckle Aqueous Extracts Induced let-7a Suppress EV71 Replication and Pathogenesis In Vitro and In Vivo and Is Predicted to Inhibit SARS-CoV-2.

Honeysuckle (Lonicera japonica Thunb) is a traditional Chinese medicine (TCM) with an antipathogenic activity. MicroRNAs (miRNAs) are small non-coding RNA molecules that are ubiquitously expressed in cells. Endogenous miRNA may function as an innate response to block pathogen invasion. The miRNA expression profiles of both mice and humans after the ingestion of honeysuckle were obtained. Fifteen overexpressed miRNAs overlapped and were predicted to be capable of targeting three viruses: dengue virus (DENV), enterovirus 71 (EV71) and SARS-CoV-2. Among them, let-7a was examined to be capable of targeting the EV71 RNA genome by reporter assay and Western blotting. Moreover, honeysuckle-induced let-7a suppression of EV71 RNA and protein expression as well as viral replication were investigated both in vitro and in vivo. We demonstrated that let-7a targeted EV71 at the predicted sequences using luciferase reporter plasmids as well as two infectious replicons (pMP4-y-5 and pTOPO-4643). The suppression of EV71 replication and viral load was demonstrated in two cell lines by luciferase activity, RT-PCR, real-time PCR, Western blotting and plaque assay. Furthermore, EV71-infected suckling mice fed honeysuckle extract or inoculated with let-7a showed decreased clinical scores and a prolonged survival time accompanied with decreased viral RNA, protein expression and virus titer. The ingestion of honeysuckle attenuates EV71 replication and related pathogenesis partially through the upregulation of let-7a expression both in vitro and in vivo. Our previous report and the current findings imply that both honeysuckle and upregulated let-7a can execute a suppressive function against the replication of DENV and EV71. Taken together, this evidence indicates that honeysuckle can induce the expression of let-7a and that this miRNA as well as 11 other miRNAs have great potential to prevent and suppress EV71 replication.

Honeysuckle (Lonicera japonica) and Huangqi (Astragalus membranaceus) Suppress SARS-CoV-2 Entry and COVID-19 Related Cytokine Storm in Vitro.

COVID-19 is threatening human health worldwide but no effective treatment currently exists for this disease. Current therapeutic strategies focus on the inhibition of viral replication or using anti-inflammatory/immunomodulatory compounds to improve host immunity, but not both. Traditional Chinese medicine (TCM) compounds could be promising candidates due to their safety and minimal toxicity. In this study, we have developed a novel in silico bioinformatics workflow that integrates multiple databases to predict the use of honeysuckle (Lonicera japonica) and Huangqi (Astragalus membranaceus) as potential anti-SARS-CoV-2 agents. Using extracts from honeysuckle and Huangqi, these two herbs upregulated a group of microRNAs including let-7a, miR-148b, and miR-146a, which are critical to reduce the pathogenesis of SARS-CoV-2. Moreover, these herbs suppressed pro-inflammatory cytokines including IL-6 or TNF-α, which were both identified in the cytokine storm of acute respiratory distress syndrome, a major cause of COVID-19 death. Furthermore, both herbs partially inhibited the fusion of SARS-CoV-2 spike protein-transfected BHK-21 cells with the human lung cancer cell line Calu-3 that was expressing ACE2 receptors. These herbs inhibited SARS-CoV-2 Mpro activity, thereby alleviating viral entry as well as replication. In conclusion, our findings demonstrate that honeysuckle and Huangqi have the potential to be used as an inhibitor of SARS-CoV-2 virus entry that warrants further in vivo analysis and functional assessment of miRNAs to confirm their clinical importance. This fast-screening platform can also be applied to other drug discovery studies for other infectious diseases.

Sesamin, a Naturally Occurring Lignan, Inhibits Ligand-Induced Lipogenesis through Interaction with Liver X Receptor Alpha (LXRα) and Pregnane X Receptor (PXR).

Liver X receptor (LXR) is a nuclear receptor that regulates various biological processes, including de novo lipogenesis, cholesterol metabolism, and inflammation. Selective inhibition of LXR may aid the treatment of nonalcoholic fatty liver disease (NAFLD). Sesamin is a naturally occurring lignan in many dietary plants and has a wide range of beneficial effects on metabolism. The mechanism underlying sesamin action especially on the regulation of LXR remains elusive. Reporter assays, mRNA and protein expression, and in silico modeling were used to identify sesamin as an antagonist of LXRα. Sesamin was applied to the hepatic HepaRG and intestinal LS174T cells and showed that it markedly ameliorated lipid accumulation in the HepaRG cells, by reducing LXRα transactivation, inhibiting the expression of downstream target genes. This effect was associated with the stimulation of AMP-activated protein kinase (AMPK) signaling pathway, followed by decreased T0901317-LXRα-induced expression of SREBP-1c and its downstream target genes. Mechanistically, sesamin reduced the recruitment of SRC-1 but enhanced that of SMILE to the SREBP-1c promoter region under T0901317 treatment. It regulated the transcriptional control exerted by LXRα by influencing its interaction with coregulators and thus decreased mRNA and protein levels of genes downstream of LXRα and reduced lipid accumulation in hepatic cells. Additionally, sesamin reduced valproate- and rifampin-induced LXRα and pregnane X receptor (PXR) transactivation. This was associated with reduced expression of target genes and decreased lipid accumulation. Thus, sesamin is an antagonist of LXRα and PXR and suggests that it may alleviate drug-induced lipogenesis via the suppression of LXRα and PXR signaling.

Proteomic analysis of honokiol-induced cytotoxicity in thyroid cancer cells.

AIMS: Honokiol is a natural product extracted from herbal plants such as the Magnolia species which have been shown to exhibit anti-tumor and anti-metastatic properties. However, the effects of honokiol on thyroid cancers are largely unknown. MATERIALS AND METHODS: To determine whether honokiol might be useful for the treatment of thyroid cancer and to elucidate the mechanism of toxicity of honokiol, we analyzed the impact of honokiol treatment on differential protein expression in human thyroid cancer cell line ARO using lysine-labeling two-dimensional difference gel electrophoresis (2D-DIGE) combined with mass spectrometry (MS). KEY FINDINGS: This study revealed 178 proteins that showed a significant change in expression levels and also revealed that honokiol-induced cytotoxicity in thyroid cancer cells involves dysregulation of cytoskeleton, protein folding, transcription control and glycolysis. SIGNIFICANCE: Our work shows that combined proteomic strategy provides a rapid method to study the molecular mechanisms of honokiol-induced cytotoxicity in thyroid cancer cells. The identified targets may be useful for further evaluation as potential targets in thyroid cancer therapy.

Gold lotion from citrus peel extract ameliorates imiquimod-induced psoriasis-like dermatitis in murine.

BACKGROUND: Gold lotion (GL), a natural mixed product made from the peels of six citrus fruits, has recently been identified as possessing anti-oxidative, anti-inflammatory, and immunomodulatory effects. GL has been used to protect skin against UV-induced damage, but its activity against psoriasis, a chronic autoimmune skin disease caused by dysregulation between immune cells and keratinocytes, is not known. We therefore evaluated the effect of GL on imiquimod (IMQ)-induced psoriasis-like inflammation in mice. RESULTS: GL treatment significantly attenuated IMQ-induced psoriasis-like symptoms in mice. The inflammatory cytokines upregulated by IMQ in skin lesions were also inhibited by feeding GL. In addition, GL treatment reduced the infiltration of CD4+ T cells/neutrophils in skin lesions and the percentage of IL-17-/IL-22-producing T cells in lymph nodes. Furthermore, GL impaired IMQ-induced type I interferon production by plasmacytoid dendritic cells (pDCs) in vitro. CONCLUSION: Our results indicate GL can act to suppress the initiation of psoriasis and strongly suggest that GL may have potential to be applied to the treatment of psoriasis. © 2018 Society of Chemical Industry.

Modulation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR) activation by ursolic acid (UA) attenuates rifampin-isoniazid cytotoxicity.

BACKGROUND: Interactions between transcriptional inducers of cytochrome P450 (CYP450) enzymes and therapeutic drugs may be prevented by antagonizing the activation of a nuclear receptor (NR), pregnane X receptor (PXR, NR1I2), thus improving therapeutic efficacy. PURPOSE: In the present study, we aim to identify that ursolic acid (UA), a widely distributed pentacyclic triterpene, may act as an effective antagonist of PXR and its sister NR receptor, constitutive androstane receptor (CAR, NR1I3). METHODS: The hepatocellular carcinoma cell line, HepG2, was used to evaluate the promoter activity of PXR and CAR target genes, CYP3A4 and CYP2B6, respectively. Catalytic activities, mRNA, and protein expression of CYP3A4 and CYP2B6 were evaluated in a differentiated HepaRG cell line. Coregulation of PXR with coregulators on CYP3A4 promoter response elements was also been characterized. RESULTS: Transient transfection assays showed that UA effectively attenuated CYP3A4 and CYP2B6 promoter activities mediated by rifampin (RIF, human PXR agonist) and CITCO (human CAR agonist). These inhibitory effects were well correlated with the expression and catalytic activities of CYP3A4 and CYP2B6. Furthermore, the interaction of co-regulators with PXR and the transcriptional complexes in the CYP3A4 promoter activity and CYP3A4 promoter xenobiotic response element (everted repeat 6, ER6), respectively, were disrupted in the presence of UA. UA showed an antagonistic effect against PXR, and reversed the cytotoxic effects of isoniazid (INH) induced by RIF. Taken together, these results show that UA inhibits the transactivation effects of PXR and CAR, and reduces the expression and function of CYP3A4 and CYP2B6. CONCLUSION: The present study suggests that UA could be a powerful agent for reducing potentially dangerous interactions between transcriptional inducers of CYP enzymes and therapeutic drugs.

Evodiamine Induces Cell Growth Arrest, Apoptosis and Suppresses Tumorigenesis in Human Urothelial Cell Carcinoma Cells.

BACKGROUND/AIM: Evodiamine, an indole alkaloid derived from Evodia rutaecarpa, exhibits pharmacological activities including vasodilatation, analgesia, anti-cardiovascular disease, anti-Alzheimer's disease, anti-inflammation, and anti-tumor activity. MATERIALS AND METHODS: This study analyzes the anti-tumor effects of evodiamine on cellular growth, tumorigenesis, cell cycle and apoptosis induction of human urothelial cell carcinoma (UCC) cells. RESULTS: The present study showed that evodiamine significantly inhibited the proliferation of UCC cells in a dose- and time-dependent manner. Also, evodiamine suppressed the tumorigenesis of UCC cells in vitro. Moreover, evodiamine caused G2/M cell-cycle arrest and induced caspase-dependent apoptosis in UCC cells. Finally, we demonstrated that evodiamine exhibits better cytotoxic than 5-fluorouracil, a clinical chemotherapeutic drug, for UCC cells. CONCLUSION: Evodiamine induces growth inhibition, tumorigenesis suppression, cell-cycle arrest, and apoptosis induction in human UCC cells. Therefore, this agent displays a therapeutic potential for treating human UCC cells and is worthy for further investigation.

Honeysuckle aqueous extract and induced let-7a suppress dengue virus type 2 replication and pathogenesis.

ETHNOPHARMACOLOGICAL RELEVANCE: Honeysuckle (Lonicera japonica Thunb.), a traditional Chinese herb, has widely been used to treat pathogen infection. However, the underlying-mechanism remains elusive. AIMS OF THE STUDY: To reveal the host microRNA (miRNA) profile with the anti-viral activity after honeysuckle treatment. MATERIALS AND METHODS: Here we reveal the differentially expressed miRNAs by Solexa® deep sequencing from the blood of human and mice after the aqueous extract treatment. Among these overexpressed innate miRNAs both in human and mice, let-7a is able to target the NS1 region (nt 3313-3330) of dengue virus (DENV) serotypes 1, 2 and 4 predicated by the target predication software. RESULTS: We confirmed that let-7a could target DENV2 at the predicated NS1 sequence and suppress DENV2 replication demonstrated by luciferase-reporter activity, RT-PCR, real-time PCR, Western blotting and plaque assay. ICR-suckling mice consumed honeysuckle aqueous extract either before or after intracranial injection with DENV2 showed decreased levels of NS1 RNA and protein expression accompanied with alleviated disease symptoms, decreased virus load, and prolonged survival time. Similar results were observed when DENV2-infected mice were intracranially injected with let-7a. CONCLUSION: We reveal that honeysuckle attenuates DENV replication and related pathogenesis in vivo through induction of let-7a expression. This study opens a new direction for prevention and treatment of DENV infection through induction of the innate miRNA let-7a by honeysuckle.

Cortex Moutan Induces Bladder Cancer Cell Death via Apoptosis and Retards Tumor Growth in Mouse Bladders.

Cortex Moutan is the root bark of Paeonia suffruticosa Andr. It is the herbal medicine widely used in Traditional Chinese Medicine for the treatment of blood-heat and blood-stasis syndrome. Furthermore, it has been reported that Cortex Moutan has anticancer effect. In this study, the Cortex Moutan extract was evaluated in bladder cancer therapy in vitro and in vivo. Cortex Moutan extract reduces cell viability with IC50 between 1~2 mg/ml in bladder cancer cells, and it has lower cytotoxicity in normal urotheliums. It arrests cells in G1 and S phase and causes phosphatidylserine expression in the outside of cell membrane. It induces caspase-8 and caspase-3 activation and poly(ADP-ribose) polymerase degradation. The pan caspase inhibitor z-VAD-fmk reverses Cortex Moutan-induced cell death. Cortex Moutan also inhibits cell invasion activity in 5637 cells. In mouse orthotopic bladder cancer model, intravesical application of Cortex Moutan decreases the bladder tumor size without altering the blood biochemical parameters. In summary, these results demonstrate the antiproliferation and anti-invasion properties of Cortex Moutan in bladder cancer cells and its antibladder tumor effect in vivo. Cortex Moutan may provide an alternative therapeutic strategy for the intravesical therapy of superficial bladder cancer.