635 nm LED irradiation may prevent endoplasmic reticulum stress in MC3T3-E1 cells.

Although endoplasmic reticulum (ER) stress is thought to be involved in various diseases such as cancer, metabolic, and inflammatory disorders, the relationship between ER stress and bone diseases, are remains unclear. Tunicamycin-treated MC3T3-E1 osteoblasts were used as the ER stress model in this study. 635 nm light-emitting diode irradiation (635 nm-IR) was carried out for 1 h before and after inducing ER stress. To investigate the effects of 635 nm-IR on ER stress-induced MC3T3-E1 osteoblasts and the underlying mechanism, western blot, reverse transcription polymerase chain reaction, alkaline phosphatase and Alizarin red staining, 2',7'-dichlorodyhydrofluorescein diacetate assay, Fluo-3AM and immunocytochemistry were performed. Pretreatment with 635 nm-IR effectively prevented intracellular reactive oxygen species production and alleviated ER stress through the pancreatic ER kinase (PERK)-eukaryotic initiation factor 2 (eIF2)-activating transcription factor 4 (ATF4)-nuclear factor-like 2 (Nrf2) signaling pathway. Hence, 635 nm-IR may serve a protective role in the treatment of ER stress-related bone diseases.

Photochemical reaction to increase melanogenesis using Buddleja officinalis and blue light-emitting diode irradiation in B16F10.

Recently, the incidence of vitiligo has increased because of stresses induced by external environment. Ultraviolet (UV) light therapy is the most commonly used method of treating the disease; however, UV light therapy requires a long treatment period, and prolonged exposure to UV radiation has side effects. The purpose of the present study was to investigate the effects of natural products and LED irradiation (LED-IR) on the synthesis of melanin. It was not possible to effectively increase intracellular melanin production through individual applications of Buddleja officinalis (BO), which is a natural substance selected through screening, or blue light irradiation (Blue-IR). However, when used in combination, these two agents stimulated adenylyl cyclase (AC) and melanin production was induced in the stimulated cells via the CREB/MITF/TYR pathway. Furthermore, the combined treatment with BO and Blue-IR generated low levels of cellular reactive oxygen species (ROS) and induced p38 phosphorylation, which in turn activated MITF in ROS-stimulated synthetic melanocytes, resulting in the promotion of melanogenic pathways other than the CREB/MITF/TYR pathway. In addition, this treatment combination effected melanin transport. These results suggested that the combined therapies can be used to treat melanin-deficiency skin diseases such as vitiligo.

Raphanus sativus L. seed extracts induce apoptosis and reduce migration of oral squamous cell carcinoma KB and KBCD133+cells by downregulation of ß-catenin.

Although, oral cancer therapies have been developed for decades, patient survival rates have not changed. Side effects of chemotherapy and radiotherapy reduce quality of life of patients and it remains difficult to treat oral cancers due to the presence of cancer stem cells (CSCs) that cause recurrence and metastasis. Therefore, we search for natural products that affect oral cancer cells including oral cancer stem cells. In the present study, we investigated the anticancer effects of Raphanus sativus L. seed (RSLS) extracts on oral squamous cell carcinoma KB cells and CSC-like KBCD133+ cells. CD133 plays an important role in CSCs and physically binds to ß-catenin to activate the ß-catenin signaling targets. Therefore, a natural extract that can inhibit ß-catenin act in may be effective anticancer drug acquiring CSC. Of the natural product extract candidates, RSLS extracts induced apoptosis in KB and KBCD133+ cells and inhibited nuclear translocation of ß-catenin cell migration and invasion rates. Treatment of RSLS extracts resulted in increases of Axin and it leds to reductions of ß-catenin in KB and KBCD133+ cells. Hence, the result suggests that RSLS are potential candidate for anticancer drug against oral cancer cells and CSCs.AbbreviationsCSCcancer stem cellsOSCCsquamous cell carcinoma cellsRSLSRaphanus sativus L. seed.

Acanthopanacis Cortex extract: A novel photosensitizer for head and neck squamous cell carcinoma therapy.

OBJECTIVES: The aim of this study was to develop a novel photosensitizer from traditional plant extracts and to investigate the photodynamic therapy (PDT) effect and mechanism of action of the novel photosensitizer on KB and Hep-2 cells. METHODS: Fluorescence emission, cell viability, and intracellular distribution of candidates were analyzed to screen potential photosensitizers from traditional plant extracts. Cellular reactive oxygen species (ROS) quantification, Annexin V-FITC/PI staining, and western blotting were performed to explore the mechanism of cell death in KB and Hep-2 cells. RESULT: Of 289 traditional plant extracts, 13 plant extracts with strong fluorescence were initially screened by fluorescence emission analysis. The cell viability assay and intracellular distribution of candidates showed that Acanthopanacis Cortex (AC) extract is a potential photosensitizer. Under optimal PDT conditions, high levels of ROS were produced in KB and Hep-2 cells, followed by cell death. However, there was no significant damage to HaCaT cells. Moreover, apoptosis induced by AC extract with 625 nm irradiation (IR) down-regulated the expression of Bcl-2 protein and up-regulated the expression of Bax protein, as well as that of cleaved PARP-1 protein in both KB and Hep-2 cells. CONCLUSION: The fluorescence intensity of AC extract at 420 nm is similar to that of the commercial Hematoporphyrin (HP). AC extract with 625 nm IR could enhance the PDT effect, induce ROS generation, and trigger apoptotic pathways in KB and Hep-2 cells. Therefore, we suggest that AC is a potential novel photosensitizer for PDT in head and neck squamous cell carcinoma.

Sanguisorba officinalis L. extracts activate Wnt/ß-catenin pathway, and subsequently control adipo-osteogenic differentiation.

Wnt/ß-catenin pathway plays an important role in adipogenesis and osteogenesis. To search for novel activators of the Wnt/ß-catenin pathway, we screened plant extracts and found that Sangusorba officinalis L. extracts (SOE) could increase ß-catenin expression level and nuclear accumulation in 3T3-L1 and MC3T3-E1 cells. It was confirmed that SOE could effectively control adipo-osteogenic differentiation. SOE inhibited adipocyte differentiation of 3T3-L1 cells, markedly decreased intracellular lipid accumulation, and decreased expression levels of key adipogenic transcription factors including PPARγ, C/EBPα, and SREBP-1c. SOE enhanced ALP activity and terminal osteoblast differentiation which was confirmed by Alizarin Red S staining of MC3T3-E1 cells. In ex vivo culture, SOE significantly increased the thickness of calvarial bone in mice. Taken together, our results indicate that SOE could be used as a potential therapeutic agent for dual-treatment of anti-obesity and anti-osteoporosis via activation of Wnt/ß-catenin pathway.

Development of Novel Photosensitizer Using the Buddleja officinalis Extract for Head and Neck Cancer.

Photodynamic therapy (PDT) is generally safer and less invasive than conventional strategies for head and neck cancer treatment. However, currently available photosensitizers have low selectivity for tumor cells, and the burden and side effects are so great that research is needed to develop safe photosensitizers. In this study, it was confirmed that the Buddleja officinalis (BO) extract, used in the treatment of inflammation and vascular diseases, shows fluorescence when activated by LED light, and, based on this, we aimed to develop a new photosensitive agent suitable for PDT. MTT, Diff-Quick® staining, and DCF-DA were performed to measure the effects of treating head and neck cancer cells with BO extract and 625 nm LED light (BO-PDT). Cell cycle, TUNEL, and western blot assays, as well as acridine orange staining, were performed to explore the mechanism of BO-PDT-induced cell death. We found that when the BO extract was irradiated with 625 nm LED light, it showed sufficient fluorescence and stronger intracellular toxicity and ROS effect than the currently commercially available hematoporphyrin. BO-PDT resulted in a decrease of mTOR activity that was correlated with an increase in the levels of ATG5, beclin-1, and LC3-II, which interfere with the formation of autophagosomes. In addition, BO-PDT induced the activation of PARP and led to an increase in the expression of proapoptotic protein Bax and a decrease in the expression of the antiapoptotic protein Bcl-2. Moreover, BO-PDT has been shown to induce the autophagy pathway 4 h after treatment, while apoptosis was induced 16 h after treatment. Finally, we confirmed that BO-PDT caused cell death of head and neck cancer cells via the intrinsic pathway. Therefore, we suggest that BO extract can be used as a new photosensitizer in PDT of head and neck cancer.

Red light-emitting diode irradiation regulates oxidative stress and inflammation through SPHK1/NF-κB activation in human keratinocytes.

Oxidative stress, in which the amount of oxidants exceeds the capacity of antioxidant defense system, is a well-accepted pathogenesis of several human diseases. Light-emitting diode irradiation (LEDI) is an efficient strategy to counteract this condition. The biological effect of phototherapy, using visible light, has attracted recent attention especially in dermatological practice. However, little is known about the molecular mechanism of the anti-oxidant and anti-inflammatory effects of red light irradiation. We evaluated these effects of LEDI in HaCaT human keratinocyte cells under phorbol-12-myristate-13-acetate (PMA) induced reactive oxygen species (ROS). Microarray analysis revealed changes in 309 genes after LEDI. LEDI at 625 nm produced ROS scavenging and anti-inflammatory effects. One of the most important genes identified by microarray analysis was sphingosine kinase-1 (SPHK1), which is a key molecule in sphingolipid metabolism. SPHK1 knock-down drastically reduced ROS scavenging efficiency as well as expression levels of inflammation-related proteins in PMA-treated HaCaT cells. These results not only indicate the potential for the clinical application of 625-nm LEDI in treating skin disorders via ROS and/or inflammation, but also suggest SPHK1 as a potential therapeutic target in phototherapy.

Cell proliferation and migration mechanism of caffeoylserotonin and serotonin via serotonin 2B receptor in human keratinocyte HaCaT cells.

Caffeoylserotonin (CaS), one derivative of serotonin (5-HT), is a secondary metabolite produced in pepper fruits with strong antioxidant activities. In this study, we investigated the effect of CaS on proliferation and migration of human keratinocyte HaCaT cells compared to that of 5-HT. CaS enhanced keratinocyte proliferation even under serum deficient condition. This effect of CaS was mediated by serotonin 2B receptor (5-HT2BR) related to the cell proliferation effect of 5-HT. We also confirmed that both CaS and 5-HT induced G1 progression via 5-HT2BR/ERK pathway in HaCaT cells. However, Akt pathway was additionally involved in upregulated expression levels of cyclin D1 and cyclin E induced by CaS by activating 5-HT2BR. Moreover, CaS and 5-HT induced cell migration in HaCaT cells via 5-HT2BR. However, 5-HT regulated cell migration only through ERK/AP-1/MMP9 pathway while additional Akt/NF-κB/MMP9 pathway was involved in the cell migration effect of CaS. These results suggest that CaS can enhance keratinocyte proliferation and migration. It might have potential as a reagent beneficial for wound closing and cell regeneration. [BMB Reports 2018; 51(4): 188-193].

Feruloylserotonin inhibits hydrogen peroxide-induced melanogenesis and apoptosis in B16F10 and SK-Mel-2 melanoma cells.

Feruloylserotonin (FS) is a major bioactive component of safflower seeds, with documented strong antibacterial, anti-inflammatory, and free radical scavenging activities. Reactive oxygen species (ROS) can strongly induce melanogenesis and cell apoptosis. The present study aimed to investigate the ability of FS in preventing hydrogen peroxide (H2O2)-induced melanogenesis and cell apoptosis. Melanogenesis and apoptotic cell death were induced by transient exposure to H2O2 in B16F10 and SK-Mel-2 melanoma cells. FS significantly inhibited melanogenesis and cell death in both cell lines. FS inhibited H2O2-induced melanin production by down-regulating CREB/MITF/TYR signaling via inhibited intracellular cAMP accumulation. Additionally, FS induced extracellular regulated kinase activation, which led to the degradation of MITF and consequently decreased TYR expression and melanin production in H2O2-stimulated cells. Furthermore, FS inhibited H2O2-induced apoptotic cell death by maintaining mitochondrial membrane potential. Therefore, FS might have potential use for cosmetic whitening and as a therapeutic agent for hyperpigmentation disorder.