The effects of patchouli alcohol and combination with cisplatin on proliferation, apoptosis and migration in B16F10 melanoma cells.

Melanoma is a highly metastatic cancer with a low incidence rate, but a high mortality rate. Patchouli alcohol (PA), a tricyclic sesquiterpene, is considered the main active component in Pogostemon cablin Benth, which improves wound healing and has anti-tumorigenic activity. However, the pharmacological action of PA on anti-melanoma remains unclear. Thus, the present study aimed to investigate the role of PA in the proliferation, cell cycle, apoptosis and migration of melanoma cells. These results indicated that PA selectively inhibited the proliferation of B16F10 cells in a dose- and time-dependent manner. It induced cell cycle arrest at the G0 /G1 phase and typical morphological changes in apoptosis, such as chromatin condensation, DNA fragmentation and apoptotic bodies. In addition, PA reduced the migratory ability of B16F10 cells by upregulating E-cadherin and downregulating p-Smad2/3, vimentin, MMP-2 and MMP-9 expression. PA was also found to strongly suppress tumour growth in vivo. Furthermore, PA combined with cisplatin synergistically inhibited colony formation and migration of B16F10 cells and attenuated the development of resistance to treatment. Therefore, the results of this study indicate that PA may play a pivotal role in inducing apoptosis and reducing the migration of melanoma cells, and may thus be a potential candidate for melanoma treatment.

Angelica sinensis extract induces telomere dysfunction, cell cycle arrest, and mitochondria-mediated apoptosis in human glioblastoma cells.

Glioblastoma (GB) is one of the most aggressive and malignant tumors of the central nervous system. Conventional treatment for GB requires surgical resection followed by radiotherapy combined with temozolomide chemotherapy; however, the median survival time is only 12-15 months. Angelica sinensis Radix (AS) is commonly used as a traditional medicinal herb or a food/dietary supplement in Asia, Europe, and North America. This study aimed to investigate the effect of AS-acetone extract (AS-A) on the progression of GB and the potential mechanisms underlying its effects. The results indicated that AS-A used in this study showed potency in growth inhibition of GB cells and reduction of telomerase activity. In addition, AS-A blocked the cell cycle at the G0/G1 phase by regulating the expression of p53 and p16. Furthermore, apoptotic morphology, such as chromatin condensation, DNA fragmentation, and apoptotic bodies, was observed in AS-A-treated cells, induced by the activation of the mitochondria-mediated pathway. In an animal study, AS-A reduced tumor volume and prolonged lifespans of mice, with no significant changes in body weight or obvious organ toxicity. This study confirmed the anticancer effects of AS-A by inhibiting cell proliferation, reducing telomerase activity, altering cell cycle progression, and inducing apoptosis. These findings suggest that AS-A has great potential for development as a novel agent or dietary supplement against GB.

Cedrol restricts the growth of colorectal cancer in vitro and in vivo by inducing cell cycle arrest and caspase-dependent apoptotic cell death.

Background: Cedrol is a natural sesquiterpene alcohol found in Cedrus atlantica, which has been proven to have a broad spectrum of biological activities, such as antimicrobial, anti-inflammatory, analgesic, anxiolytic, and anti-cancer effects. However, the underlying anticancer mechanisms and in vivo inhibitory effects of cedrol on colorectal cancer (CRC) have not been elucidated. In the present study, we investigated the anti-CRC potential of cedrol using in vitro and in vivo models. Methods: The effects of cedrol on cell viability, cell cycle progression, and apoptosis of HT-29 and CT-26 cells were detected by MTT, flow cytometry, and TUNEL assays. Western blotting was used to measure protein expression for molecular signaling analyses. Results: Cedrol inhibited HT-29 and CT-26 cell proliferation in a time- and dose-dependent manner, with IC50 values of 138.91 and 92.46 µM, respectively. Furthermore, cedrol induced cell cycle arrest at the G0/G1 phase by regulating the expression of cell cycle regulators, such as CDK4 and cyclin D1, and triggered apoptosis through extrinsic (FasL/caspase-8) and intrinsic (Bax/caspase-9) pathways. In addition, cedrol in combination with the clinical drug 5-fluorouracil exhibited synergistic inhibitory effects on CRC cell growth. Importantly, cedrol treatment suppressed the progression of CRC and improved the survival rate of animals at a well-tolerated dose. Conclusion: These results suggest that cedrol has an anti-cancer potential via induction of cell cycle arrest and apoptosis, and it could be considered as an effective agent for CRC therapy.

Surface Expression of Kynurenine 3-Monooxygenase Promotes Proliferation and Metastasis in Triple-Negative Breast Cancers.

Kynurenine 3-monooxygenase (KMO) is the pivotal enzyme in the kynurenine pathway and is located on the mitochondrial outer membrane. The dysregulation of KMO leads to various neurodegenerative diseases; however, it is rarely mentioned in cancer progression. Our previous study showed that KMO overexpression in canine mammary gland tumors (cMGT) is associated with poor prognosis in cMGT patients. Surprisingly, it was also found that KMO can be located on the cell membranes of cMGT cells, unlike its location in normal cells, where KMO is expressed only within the cytosol. Since cMGT and human breast cancer share similar morphologies and pathogenesis, this study investigated the possibility of detecting surface KMO in human breast cancers and the role of surface KMO in tumorigenesis. Using immunohistochemistry (IHC), flow cytometry (FC), immunofluorescence assay (IFA), and transmission electron microscopy (TEM), we demonstrated that KMO can be aberrantly and highly expressed on the cell membranes of breast cancer tissues and in an array of cell lines. Masking surface KMO with anti-KMO antibody reduced the cell viability and inhibited the migration and invasion of the triple-negative breast cancer cell line, MDA-MB-231. These results indicated that aberrant surface expression of KMO may be a potential therapeutic target for human breast cancers.

Juniperus indica Bertol. extract synergized with cisplatin against melanoma cells via the suppression of AKT/mTOR and MAPK signaling and induction of cell apoptosis.

Juniperus indica Bertol. is an herbal plant that belongs to the genus Juniperus, which is commonly used in traditional medicine to refresh the mind and for diuretic use. However, few studies have reported the function of J. indica Bertol. Hence, this study aimed to investigate the anti-tumor and synergistic potential of J. indica Bertol. extract (JIB extract) for melanoma cells. Our results indicated the anti-melanoma activity of JIB extract. JIB extract induced cell cycle arrest at the G0/G1 phase and decreased cyclin and cdk protein expressions. In addition, AKT/mTOR signaling and MAPK signaling were inhibited by JIB extract to suppress melanoma cell growth and proliferation. Additionally, JIB extract induced B16/F10 cell apoptosis via the caspase cascade. According to the JIB extract's anti-melanoma capacity, to assess the synergistic effects of cisplatin and JIB extract. The results demonstrated that JIB extract combined with cisplatin enhanced the inhibition of cell growth, proliferation, and survival through the obstruction of cell cycle progression and AKT/mTOR and MAPK signaling as well as the induction of cell apoptosis. Collectively, our results indicate that JIB extract showed anti-tumor effects and synergized with cisplatin against B16/F10 cells, indicating the possibility of JIB extract to be developed as adjuvant therapy for melanoma.

Enhancement of cytotoxicity and induction of apoptosis by cationic nano-liposome formulation of n-butylidenephthalide in breast cancer cells.

Breast cancer is the second most common malignancy in women. Current clinical therapy for breast cancer has many disadvantages, including metastasis, recurrence, and poor quality of life. Furthermore, it is necessary to find a new therapeutic drug for breast cancer patients to meet clinical demand. n-Butylidenephthalide (BP) is a natural and hydrophobic compound that can inhibit several tumors. However, BP is unstable in aqueous or protein-rich environments, which reduces the activity of BP. Therefore, we used an LPPC (Lipo-PEG-PEI complex) that can encapsulate both hydrophobic and hydrophilic compounds to improve the limitation of BP. The purpose of this study is to investigate the anti-tumor mechanisms of BP and BP/LPPC and further test the efficacy of BP encapsulated by LPPC on SK-BR-3 cells. BP inhibited breast cancer cell growth, and LPPC encapsulation (BP/LPPC complex) enhanced the cytotoxicity on breast cancer by stabilizing the BP activity and offering endocytic pathways. Additionally, BP and LPPC-encapsulated BP induced cell cycle arrest at the G0/G1 phase and might trigger both extrinsic as well as intrinsic cell apoptosis pathway, resulting in cell death. Moreover, the BP/LPPC complex had a synergistic effect with doxorubicin of enhancing the inhibitory effect on breast cancer cells. Consequently, LPPC-encapsulated BP could improve the anti-cancer effects on breast cancer in vitro. In conclusion, BP exhibited an anti-cancer effect on breast cancer cells, and LPPC encapsulation efficiently improved the cytotoxicity of BP via an acceleration of entrapment efficiency to induce cell cycle block and apoptosis. Furthermore, BP/LPPC exhibited a synergistic effect in combination with doxorubicin.

Cedrus atlantica Extract Suppress Glioblastoma Growth through Promotion of Genotoxicity and Apoptosis: In Vitro and In Vivo Studies.

Glioblastoma (GBM) is the most common malignant primary brain tumor in humans, exhibiting highly infiltrative growth and drug resistance to conventional chemotherapy. Cedrus atlantica (CAt) extract has been shown to decrease postoperative pain and inhibit the growth of K562 leukemia cells. The aim of this study was to assess the anti-GBM activity and molecular mechanism of CAt extract in vitro and in vivo. The results showed that CAt extract greatly suppressed GBM cells both in vitro and in vivo and enhanced the survival rate in subcutaneous and orthotopic animal models. Moreover, CAt extract increased the level of ROS and induced DNA damage, resulting in cell cycle arrest at the G0/G1 phase and cell apoptosis. Western blotting results indicated that CAt extract regulates p53/p21 and CDK4/cyclin D1 protein expression and activates extrinsic and intrinsic apoptosis. Furthermore, CAt extract enhanced the cytotoxicity of Temozolomide and decreased AKT/mTOR signaling by combination treatment. In toxicity assays, CAt extract exhibited low cytotoxicity toward normal cells or organs in vitro and in vivo. CAt extract suppresses the growth of GBM by induction of genotoxicity and activation of apoptosis. The results of this study suggest that CAt extract can be developed as a therapeutic agent or adjuvant for GBM treatment in the future.

Effects of Cedrus atlantica extract on acute myeloid leukemia cell cycle distribution and apoptosis.

This study investigated the anti-leukemic effects of Cedrus atlantica extract (CAt extract) on cell cycle distribution and apoptosis in human acute myeloid leukemia (AML) cells. AML often occurs in older adults, accounting for 60% of the cases, and is likely to be resistant to chemotherapy due to multidrug resistance, resulting in early death during cancer treatment. With the increasing focus on prevention medicine, natural plant components are being used as a major source for the development of therapeutic drugs or functional foods to cure or alleviate the disease. Cedrus species are known to have anti-inflammatory, antimicrobial, antiviral, and anticancer effects; however, the anticancer effects of CAt extract have not been elucidated. In this study, CAt extract demonstrated an inhibitory effect on human leukemia cells in a concentration-dependent manner; CAt extract induced G0/G1 phase arrest via restrained protein levels of p-Rb and cell cycle-related proteins. After CAt extract exposure, the extrinsic and intrinsic apoptotic pathways were activated through caspase-8, -9, and -3 cleavage. Additionally, CAt extract suppressed VEGF, MMP-2, and MMP-9 expression. This study demonstrated that CAt extract treatment significantly reduced cell growth, cell cycle arrest in the G0/G1 phase, and induction of apoptosis, leading to leukemia cell death.

Cedrol, a Sesquiterpene Alcohol, Enhances the Anticancer Efficacy of Temozolomide in Attenuating Drug Resistance via Regulation of the DNA Damage Response and MGMT Expression.

Glioblastoma (GBM) is a common and aggressive brain tumor with a median survival of 12-15 months. Temozolomide (TMZ) is a first-line chemotherapeutic agent used in GBM therapy, but the occurrence of drug resistance limits its antitumor activity. The natural compound cedrol has remarkable antitumor activity and is derived from Cedrus atlantica. In this study, we investigated the combined effect of TMZ and cedrol in GBM cells in vitro and in vivo. The TMZ and cedrol combination treatment resulted in consistently higher suppression of cell proliferation via regulation of the AKT and MAPK signaling pathways in GBM cells. The combination treatment induced cell cycle arrest, cell apoptosis, and DNA damage better than either drug alone. Furthermore, cedrol reduced the expression of proteins associated with drug resistance, including O6-methlyguanine-DNA-methyltransferase (MGMT), multidrug resistance protein 1 (MDR1), and CD133 in TMZ-treated GBM cells. In the animal study, the combination treatment significantly suppressed tumor growth through the induction of cell apoptosis and decreased TMZ drug resistance. Moreover, cedrol-treated mice exhibited no significant differences in body weight and improved TMZ-induced liver damage. These results imply that cedrol may be a potential novel agent for combination treatment with TMZ for GBM therapy that deserves further investigation.

Pogostemon cablin Triggered ROS-Induced DNA Damage to Arrest Cell Cycle Progression and Induce Apoptosis on Human Hepatocellular Carcinoma In Vitro and In Vivo.

The purpose of the study was to elucidate the anti-hepatoma effects and mechanisms of Pogostemon cablin essential oils (PPa extract) in vitro and in vivo. PPa extract exhibited an inhibitory effect on hepatocellular carcinoma (HCC) cells and was less cytotoxic to normal cells, especially normal liver cells, than it was to HCC cells, exerting a good selective index. Additionally, PPa extract inhibited HCC cell growth by blocking the cell cycle at the G0/G1 phase via p53 dependent or independent pathway to down regulated cell cycle regulators. Moreover, PPa extract induced the FAS-FASL-caspase-8 system to activate the extrinsic apoptosis pathway, and it increased the bax/bcl-2 ratio and reduced ΔΨm to activate the intrinsic apoptosis pathway that might be due to lots of reactive oxygen species (ROS) production which was induced by PPa extract. In addition, PPa extract presented to the potential to act synergistically with sorafenib to effectively inhibit HCC cell proliferation through the Akt/mTOR pathway and reduce regrowth of HCC cells. In an animal model, PPa extract suppressed HCC tumor growth and prolonged lifespan by reducing the VEGF/VEGFR axis and inducing tumor cell apoptosis in vivo. Ultimately, PPa extract demonstrated nearly no or low system-wide, physiological, or pathological toxicity in vivo. In conclusion, PPa extract effectively inhibited HCC cell growth through inducing cell cycle arrest and activating apoptosis in vitro and in vivo. Furthermore, PPa extract exhibits less toxicity toward normal cells and organs than it does toward HCC cells, which might lead to fewer side effects in clinical applications. PPa extract may be developed into a clinical drug to suppress tumor growth or functional food to prevent HCC initiation or chemoprotection of HCC recurrence.

Antitumor Effects of N-Butylidenephthalide Encapsulated in Lipopolyplexs in Colorectal Cancer Cells.

Colorectal cancer (CRC) is the third most common type of cancer and the second most common cause of cancer-related death in the world. N-Butylidenephthalide (BP), a natural compound, inhibits several cancers, such as hepatoma, brain tumor and colon cancer. However, due to the unstable structure, the activity of BP is quickly lost after dissolution in an aqueous solution. A polycationic liposomal polyethylenimine and polyethylene glycol complex (LPPC), a new drug carrier, encapsulates both hydrophobic and hydrophilic compounds, maintains the activity of the compound, and increases uptake of cancer cells. The purpose of this study is to investigate the antitumor effects and protection of BP encapsulated in LPPC in CRC cells. The LPPC encapsulation protected BP activity, increased the cytotoxicity of BP and enhanced cell uptake through clathrin-mediated endocytosis. Moreover, the BP/LPPC-regulated the expression of the p21 protein and cell cycle-related proteins (CDK4, Cyclin B1 and Cyclin D1), resulting in an increase in the population of cells in the G0/G1 and subG1 phases. BP/LPPC induced cell apoptosis by activating the extrinsic (Fas, Fas-L and Caspase-8) and intrinsic (Bax and Caspase-9) apoptosis pathways. Additionally, BP/LPPC combined with 5-FU synergistically inhibited the growth of HT-29 cells. In conclusion, LPPC enhanced the antitumor activity and cellular uptake of BP, and the BP/LPPC complex induced cell cycle arrest and apoptosis, thereby causing death. These findings suggest the putative use of BP/LPPC as an adjuvant cytotoxic agent for colorectal cancer.

Extract of Juniperus indica Bertol Synergizes with Cisplatin to Inhibit Oral Cancer Cell Growth via Repression of Cell Cycle Progression and Activation of the Caspase Cascade.

Oral cancer-a type of head and neck cancer-is estimated to be the fifth most common cancer in Taiwan. However, efficacious therapies for oral cancer are still lacking due to drug resistance and recurrence. Consequently, the identification of new anticancer agents for clinical treatment is needed. Juniperus indica Bertol is a plant of the Juniperus genus often used as a treatment in traditional medicine due to its anti-inflammatory, antibacterial and diuretic functions. The biofunctions of Juniperus indica Bertol including its anticancer potential, have not been fully explored. As a result, the aim of this research was to investigate the anticancer activity of Juniperus indica Bertol extract (JIB extract) and determine whether JIB extract has synergistic effects with cisplatin in oral cancer. These results are the first to demonstrate that JIB extract exhibits anticancer capacity and synergizes with cisplatin to treat oral cancer. Our findings indicate that JIB extract has a potential to develop anticancer agent and chemo therapeutic adjuvant for oral cancer.

Extract Derived from Cedrus atlantica Acts as an Antitumor Agent on Hepatocellular Carcinoma Growth In Vitro and In Vivo.

Cedrus atlantica is widely used in herbal medicine. However, the anti-cancer activity of C. atlantica extract (CAt extract) has not been clarified in hepatocellular carcinoma. In the study, we elucidated the anti-hepatoma capacity of CAt extract on HCC in vitro and in vivo. To explore the anti-hepatoma mechanisms of the CAt extract in vitro, HCC and normal cells were treated with the CAt extract, which showed marked inhibitory effects on HCC cells in a dose-dependent manner; in contrast, the CAt extract treatment was less cytotoxic to normal cells. In addition, our results indicate that the CAt extract induced apoptosis via caspase-dependent and independent apoptosis pathways. Furthermore, the CAt extract inhibited HCC tumor cell growth by restraining cell cycle progression, and it reduced the signaling of the AKT, ERK1/2, and p38 pathways. In the xenograft model, the CAt extract suppressed HCC tumor cell growth and prolonged lifespan by inhibiting PCNA protein expression, repressing part of the VEGF-induced autocrine pathway, and triggering strong expression of cleaved caspase-3, which contributed to cell apoptosis. Moreover, the CAt extract did not induce any obvious changes in pathological morphology or body weight, suggesting it had no toxicity. CAt extract exerted anti-tumor effects on HCC in vitro and in vivo. Thus, CAt extract could be used as a potential anti-cancer therapeutic agent against HCC.

Juniperus Communis Extract Exerts Antitumor Effects in Human Glioblastomas Through Blood-Brain Barrier.

BACKGROUND/AIMS: Herbal materials derived from Juniperus communis (JCo) possess anticancer activity. In this study, we evaluated the efficacy of a JCo berry extract in suppressing glioblastoma growth. METHODS: The effects of JCo extract on the viability of normal and glioma cell lines was analyzed using a modified 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The synergistic therapeutic effect of JCo extract and temozolomide (TMZ) on glioma cells was examined by MTT analysis. Flow cytometry analysis, the terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) test, and western blotting were performed to identify the apoptotic pathway. To determine the in vivo efficacy of the JCo extract, rats were injected with 5 × 104 rat glioma RG2 cells in the back skin and brain hemisphere and then received a subcutaneous injection in the back skin that contained either JCo extract or vehicle. Finally, blood and histologic examinations were performed to evaluate JCo toxicity. RESULTS: The IC50 values of JCo extract were 57-69 µg/mL and 49-67 µg/mL in the glioblastoma cell lines after 24 and 48 h, respectively. However, in non-tumor cell lines, the respective IC50 values of JCo extract were 76-105 µg/mL and 77-108 µg/mL. The JCo extract had a stronger cytotoxicity and a larger range of IC50 values in glioma than in normal cells as compared to those effects caused by temozolomide (TMZ). In addition, the results of flow cytometry analysis, TUNEL test, and western blotting revealed that the JCo extract induced glioma cell cycle arrest through intrinsic and extrinsic apoptotic pathways. In the in vivo studies, a significant reduction of tumor size in JCo-treated rats, as measured by animal MRI, demonstrated that the JCo extract effectively inhibited glioma cell growth and successfully penetrated the blood-brain barrier. The immunohistochemical (IHC) staining detected positive signals of PCNA, VEGFR-1, and VEGFR -2 in 44.49%, 5.88%, and 5.85% of JCo-treated glioma cells, respectively. However, positive signals of PCNA, VEGFR-1, and VEGFR-2 were detected in 73.08%, 9.67%, and 11.70% of vehicle-treated glioma cells, respectively. The IHC examination of PCNA and VEGFR-1 and -2 indicated that JCo extract significantly decreased the degree of neovascularization. However, no significant differences in serum levels of blood cell count and hepatic enzymes, renal function index, and the histologic appearance of vital organs were detected between the JCo and vehicle-treated rats. CONCLUSION: The JCo extract penetrated the blood-brain barrier and significantly induced glioma cell apoptosis by reducing neovascularization via suppression of the PI3K/AKT/mTOR pathway. Furthermore, JCo extract was less cytotoxic to non-neoplastic vital organs than TMZ.

Antitumor Effect of n-Butylidenephthalide Encapsulated on B16/F10 Melanoma Cells In Vitro with a Polycationic Liposome Containing PEI and Polyethylene Glycol Complex.

Advanced melanoma can metastasize to distal organs from the skin and yield an aggressive disease and poor prognosis even after treatment with chemotherapeutic agents. The compound n-Butylidenephthalide (BP) is isolated from Angelica sinensis, which is used to treat anemia and gynecological dysfunction in traditional Chinese medicine. Studies have indicated that BP can inhibit cancers, including brain, lung, prostate, liver, and colon cancers. However, because BP is a natural hydrophobic compound, it is quickly metabolized by the liver within 24 h, and thus has limited potential for development in cancer therapy. This study investigated the anticancer mechanisms of BP through encapsulation with a novel polycationic liposome containing polyethylenimine (PEI) and polyethylene glycol complex (LPPC) in melanoma cells. The results demonstrated that BP/LPPC had higher cytotoxicity than BP alone and induced cell cycle arrest at the G0/G1 phase in B16/F10 melanoma cells. The BP/LPPC-treated cell indicated an increase in subG1 percentage and TUNEL positive apoptotic morphology through induction of extrinsic and intrinsic apoptosis pathways. The combination of BP and LPPC and clinical drug 5-Fluorouracil had a greater synergistic inhibition effect than did a single drug. Moreover, LPPC encapsulation improved the uptake of BP values through enhancement of cell endocytosis and maintained BP cytotoxicity activity within 24 h. In conclusion, BP/LPPC can inhibit growth of melanoma cells and induce cell arrest and apoptosis, indicating that BP/LPPC has great potential for development of melanoma therapy agents.

Protective Effect of Curcumin on the Tight Junction Integrity and Cellular Senescence in Human Retinal Pigment Epithelium of Early Diabetic Retinopathy.

Diabetic retinopathy (DR) is a secondary complication of diabetes that can lead to visual impairment and blindness. The retinal pigment epithelium (RPE) is a monolayer of pigment cells that forms the blood-retinal barrier (BRB) via tight junction (TJ) proteins and plays a crucial role in the physiological function of the retina. Hyperglycemia induces RPE death and BRB breakdown, which accelerates the process of DR. Curcumin, an active extract of Curcuma longa , has anti-inflammatory, antioxidant, antiapoptotic, and neuroprotective properties. However, the effect of Curcumin on the BRB under high glucose conditions remains unknown. This study aimed to investigate the protective effects of Curcumin on RPE physiology in vitro and in vivo . Curcumin significantly alleviated cell viability inhibition under high glucose conditions. Moreover, high glucose reduced extracellular signal-regulated kinase and Akt pathways activation to diminish RPE cell growth but reversed by Curcumin treatment. Curcumin protected not only TJ integrity but also retinoid regeneration through TJ proteins and isomerase modulation in diabetic retina. Furthermore, Curcumin decreased the expression of angiogenic factor to inhibit retinal neovascularization. Finally, Curcumin treatment markedly reduced apoptosis during hyperglycemia. In conclusion, Curcumin can alleviate the progression of DR by promoting RPE survival, TJ integrity, retinoid isomerase activity, RPE senescence inhibition, and neovascularization. Therefore, Curcumin exhibits high potential for use as a therapeutic agent for early DR.

Downregulation of VEGFR2 signaling by cedrol abrogates VEGF­driven angiogenesis and proliferation of glioblastoma cells through AKT/P70S6K and MAPK/ERK1/2 pathways.

Cedrol is a sesquiterpene alcohol isolated from Cedrus atlantica, which has been traditionally used in aromatherapy and has anticancer, antibacterial and antihyperalgesic effects. One characteristic of glioblastoma (GB) is the overexpression of vascular endothelial growth factor (VEGF), which induces a high degree of angiogenesis. Although previous studies have reported that cedrol inhibits GB growth by inducing DNA damage, cell cycle arrest and apoptosis, its role in angiogenesis remains unclear. The aim of the present study was to investigate the effects of cedrol on VEGF-induced angiogenesis of human umbilical vein endothelial cells (HUVECs). HUVECs were treated with 0-112 µM cedrol and 20 ng/ml VEGF for 0-24 h, and then anti-angiogenic activation of cedrol was determined by MTT assay, wound healing assay, Boyden chamber assay, tube formation assay, semi-quantitative reverse transcription-PCR and western blotting. These results demonstrated that cedrol treatment inhibited VEGF-induced cell proliferation, migration and invasion in HUVECs. Furthermore, cedrol prevented VEGF and DBTRG-05MG GB cells from inducing capillary-like tube formation in HUVECs and decreased the number of branch points formed. Moreover, cedrol downregulated the phosphorylation of VEGF receptor 2 (VEGFR2) and the expression levels of its downstream mediators AKT, ERK, VCAM-1, ICAM-1 and MMP-9 in HUVECs and DBTRG-05MG cells. Taken together, these results demonstrated that cedrol exerts anti-angiogenic effects by blocking VEGFR2 signaling, and thus could be developed into health products or therapeutic agents for the prevention or treatment of cancer and angiogenesis-related diseases in the future.

Patchouli alcohol induces G0 /G1 cell cycle arrest and apoptosis in vincristine-resistant non-small cell lung cancer through ROS-mediated DNA damage.

BACKGROUND: Lung cancer, especially non-small cell lung cancer (NSCLC), is one of the leading causes of cancer-related deaths worldwide. Vincristine (VCR) is a chemotherapeutic agent for lung cancers; however, its effectiveness is limited by side effects and the development of drug resistance. Patchouli alcohol (PA), from Pogostemon cablin extract, is known to possess anti-inflammatory and anticancer properties. In this study, we investigated the role of PA in inducing reactive oxygen species (ROS)-mediated DNA damage in A549 and VCR-resistant A549/V16 NSCLC cells. METHODS: The anticancer potential of PA was studied using the MTT assay, colony formation, flow cytometry analysis, western blotting, DCFDA staining, immunofluorescence staining, and TUNEL assay techniques. RESULTS: The intracellular ROS levels were enhanced in PA-treated cells, activating the CHK1 and CHK2 signaling pathways. PA further inhibited proliferation and colony-forming abilities and induced cell cycle arrest at the G0 /G1 phase by regulating p53/p21 and CDK2/cyclin E1 expression. Moreover, PA increased the percentage of cells in the subG1 phase and induced apoptosis by activating the Bax/caspase-9/caspase-3 intrinsic pathway. In addition, drug resistance (p-glycoprotein) and cancer stem cell (CD44 and CD133) markers were downregulated after PA treatment. Furthermore, combining PA and cisplatin exhibited synergistic inhibitory activity in A549 and A549/V16 cells. CONCLUSIONS: PA induced ROS-mediated DNA damage, triggered cell cycle arrest and apoptosis, attenuated drug resistance and cancer stem cell phenotypes, and synergistically inhibited proliferation in combination with cisplatin. These findings suggest that PA has the potential to be used for the treatment of NSCLC with or without VCR resistance.

The alleviation effects of n-butylidenephthalide on apoptosis, senescence, and tight junction impairment of retinal pigment epithelium by activating Nrf-2/HO-1 signaling pathway in early diabetic retinopathy.

AIMS: Diabetic retinopathy (DR) is a common complication of diabetes that causes visual impairment and blindness in adults. This study aimed to explore the protective effects of n-Butylidenephthalide (BP) on hyperglycemia-treated RPE in vitro and in vivo. MAIN METHODS: C57BL/6 mice were injected with STZ by intraperitoneal to induce early DR and orally administrated with 2 mg/kg BP every day for twelve weeks. Body weight and blood glucose were measured once a week. The level of retina damage was determined by TUNEL assay and H&E staining. The outer blood-retinal barrier integrity and RPE65 expression of retina were evaluated by immunofluorescence. In in vitro study, ARPE-19 cells were long-term cultured with high glucose and BP for 8 days and studied for cell survival, tight junction integrity, RPE65 expression, angiogenic factors, mitochondria membrane potential (MMP), and ROS by MTT assay, Western blot, ß-galactosidase staining, immunofluorescence, JC-1, or DCFH-DA. KEY FINDINGS: The results indicate that BP suppressed the hyperglycemic effect and maintained retina anatomy normalization, as well as protected RPE cell survival, tight junction integrity, and RPE65 expression in vitro and in vivo. In vitro results showed BP stimulated high glucose-treated ARPE-19 cell proliferation and suppressed senescence via ERK pathway. Numerous ROS production and MMP imbalance were prevented by BP through Nrf-2/HO-1 pathway. BP inhibited high glucose-induced RPE neovascularization by VEGF dysregulation. SIGNIFICANCE: BP significantly protected tight junction integrity and RPE cellular physiology through ERK/Nrf-2/HO-1 pathway to prevent DR progression. Thus, BP has great potential to be developed therapeutic agents or adjuvants for DR.

Juniperus communis extract ameliorates lipopolysaccharide-induced acute kidney injury through the adenosine monophosphate-activated protein kinase pathway.

Septic shock can aggravate organ dysfunction and even lead to death. Juniperus communis (JCo) extract has been experimentally demonstrated to have anti-inflammatory and antioxidant effects. We investigated the anti-inflammatory and antioxidant mechanism of JCo extract in vivo and in vitro. In a lipopolysaccharide (LPS)-induced acute kidney injury rat model, JCo extract improved animal survival, reduced kidney injury scores, suppressed kidney injury molecule-1, and preserved E-cadherin expression from LPS damage, as demonstrated by the immunohistochemistry examinations of the rat kidneys. In LPS-stimulated NRK-52E cells, JCo extract inhibited nuclear factor-κB (NF-κB) and increased adenosine monophosphate-activated protein kinase (AMPK) expression, prompting the activation of the antioxidant nuclear factor erythroid 2-related factor-2/heme oxygenase-1 pathway against oxidative stress. JCo extract ameliorated LPS-induced acute kidney injury by suppressing NF-κB signaling and stimulating the release of tumor necrosis factor-α and interleukin-1ß through the AMPK pathway.