Differentiation of Human-induced Pluripotent Stem Cell-derived Dental Stem Cells through Epithelial-Mesenchymal Interaction.

Research on tooth regeneration using human-induced pluripotent stem cells (hiPSCs) is valuable for autologous dental regeneration. Acquiring mesenchymal and epithelial cells as a resource for dental regeneration is necessary because mesenchymal-epithelial interactions play an essential role in dental development. We reported the establishment of hiPSCs-derived dental epithelial-like cell (EPI-iPSCs), but hiPSCs-derived dental mesenchymal stem cells (MSCs) have not yet been reported. This study was conducted to establish hiPSCs-derived MSCs and to differentiate them into dental cells with EPI-iPSCs. Considering that dental MSCs are derived from the neural crest, hiPSCs were induced to differentiate into MSCs through neural crest formation to acquire the properties of dental MSCs. To differentiate hiPSCs into MSCs through neural crest formation, established hiPSCs were cultured and differentiated with PA6 stromal cells and differentiated hiPSCs formed neurospheres on ultralow-attachment plates. Neurospheres were differentiated into MSCs in serum-supplemented medium. Neural crest-mediated MSCs (NC-MSCs) continuously showed typical MSC morphology and expressed MSC markers. After 8 days of odontogenic induction, the expression levels of odontogenic/mineralization-related genes and dentin sialophosphoprotein (DSPP) proteins were increased in the NC-MSCs alone group in the absence of coculturing with dental epithelial cells. The NC-MSCs and EPI-iPSCs coculture groups showed high expression levels of amelogenesis/odontogenic/mineralization-related genes and DSPP proteins. Furthermore, the NC-MSCs and EPI-iPSCs coculture group yielded calcium deposits earlier than the NC-MSCs alone group. These results indicated that established NC-MSCs from hiPSCs have dental differentiation capacity with dental epithelial cells. In addition, it was confirmed that hiPSCs-derived dental stem cells could be a novel cell source for autologous dental regeneration.

Uncovering the colorectal cancer immunotherapeutic potential: Evening primrose (Oenothera biennis) root extract and its active compound oenothein B targeting the PD-1/PD-L1 blockade.

BACKGROUND: The emergence of immune checkpoint inhibitors, a novel class of immunotherapy drugs, represents a major breakthrough in cancer immunotherapy, substantially improving patient survival post-treatment. Blocking programmed death-ligand 1 (PD-L1) and programmed death protein-1 (PD-1) has demonstrated promising clinical results in various human cancer types. The US FDA has recently permitted only monoclonal antibody (mAb)-based PD-L1 or PD-1 blockers. Although these antibodies exhibit high antitumor efficacy, their size- and affinity-induced side effects limit their applicability. PURPOSE: As small-molecule-based PD-1/PD-L1 blockers capable of reducing the side effects of antibody therapies are needed, this study focuses on exploring natural ingredient-based small molecules that can target hPD-L1/PD-1 using herbal medicines and their components. METHODS: The antitumor potential of evening primrose (Oenothera biennis) root extract (EPRE), a globally utilized traditional herbal medicine, folk remedy, and functional food, was explored. A coculture system was established using human PD-L1-expressed murine MC38 cells (hPD-L1-MC38s) and CD8+ tumor-infiltrating T lymphocytes (CD8+ TILs) expressing humanized PD-1. The in vivo experiments utilized a colorectal cancer (CRC) C57BL/6 J mouse model bearing MC38 cells expressing humanized PD-L1 and PD-1 proteins. RESULTS: EPRE and its active compound oenothein B effectively hindered the molecular interaction between hPD-L1 and hPD-1. EPRE stimulated tumor-specific T lymphocytes of a hPD-L1/PD-1 CRC mice. This action resulted in the elevated infiltration of cytotoxic CD8+T lymphocytes and subsequent tumor growth reduction. Moreover, the combined therapy of oenothein B, a PD-1/PD-L1 blocker, and FOLFOX (5-fluorouracil plus oxaliplatin) cooperatively suppressed hPD-L1-MC38s growth in the ex vivo model through activated CD8+ TIL antitumor immune response. Oenothein B exhibited a high binding affinity for hPD-L1 and hPD-1. We believe that this study is the first to uncover the inhibitory effects of EPRE and its component, oenothein B, on PD-1/PD-L1 interactions. CONCLUSION: This study identified a promising small-molecule candidate from natural products that blocks the hPD-L1/PD-1 signaling pathway. These findings emphasize the potential of EPRE and oenothein B as effective anticancer drugs.

Anti-oxidative, anti-apoptotic, and anti-inflammatory activities of Connarus semidecandrus Jack ethanol extract in UVB-irradiated human keratinocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Connarus semidecandrus Jack (Family: connaraceae) is a medicinal plant known for its wide distribution throughout Southeast Asia. Renowned for its diverse therapeutic properties, it has been traditionally used for treating fever, skin irritation, and colic. AIM OF THE STUDY: Numerous individuals suffer from skin issues, including wrinkles, hyperpigmentation, and inflammation, due to environmental factors. Although many drugs are available to treat skin problems, chemical drugs have many shortcomings and side effects. Therefore, natural products are attractive potential medicines for alleviating skin troubles. We recently showed that Connarus semidecandrus Jack ethanol extract (Cs-EE) has anti-alopecia potential. This paper aims to explore the potential skin-protective effects and underlying molecular mechanisms of Connarus semidecandrus Jack in UVB-induced human keratinocytes (HaCaT). MATERIALS AND METHODS: Before utilization, Cs-EE was dissolved in dimethyl sulfoxide (DMSO) and was preserved at a temperature of -20 °C. The phytochemical constituents of Cs-EE were detected by gas chromatography-mass spectrometry analysis (GC-MS). Sequentially, HaCaT cells were exposed to varying concentrations of Cs-EE prior to ultraviolet B (UVB) irradiation. Evaluations of cellular responses in HaCaT cells, including assessments of cell viability, deoxyribonucleic acid (DNA) damage, and gene and protein expressions, were carried out. To explore the specific signaling pathway involved, we conducted a luciferase assay in addition to validating these pathways using Western blot analysis. RESULTS: Nitric oxide (NO) and intracellular reactive oxygen species were decreased. Melanin production through the activation of melanocytes by α-melanocyte-stimulating hormone (MSH) was also inhibited by Cs-EE. Furthermore, the mRNA expression levels of key factors such as cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), MMP-1, MMP-3, and MMP-9 exhibited a remarkable decrease. In addition, the phosphorylation of TAK1 within the signaling cascade exhibited a decline, and the activities of the transcription factor AP-1 were decreased according to a luciferase reporter assay. CONCLUSIONS: Taken together, these findings suggest that the anti-inflammatory, anti-aging, and anti-apoptotic effects of Cs-EE indicate the compound's potential usefulness as a natural component in pharmaceutical and cosmetic products.

Anti-Atopic Effect of Isatidis Folium Water Extract in TNF-α/IFN-γ-Induced HaCaT Cells and DNCB-Induced Atopic Dermatitis Mouse Model.

Isatidis folium or Isatis tinctoria L. is a flowering plant of the Brassicaceae family, commonly known as woad, with an ancient and well-documented history as an indigo dye and medicinal plant. This study aimed to evaluate the anti-atopic dermatitis (AD) effects of Isatidis folium water extract (WIF) using a 2,4-dinitrochlorobenzene (DNCB)-induced AD-like mouse model and to investigate the underlying mechanism using tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ)-activated HaCaT cells. Oral administration of WIF reduced spleen weight, decreased serum IgE and TNF-α levels, reduced epidermal and dermal thickness, and inhibited eosinophil and mast cell recruitment to the dermis compared to DNCB-induced control groups. Furthermore, oral WIF administration suppressed extracellular signal-regulated kinase and p38 mitogen-activated protein kinase protein expression levels, p65 translocation from the cytoplasm to the nucleus, and mRNA expression of TNF-α, IFN-γ, interleukin (IL)-6, and IL-13 in skin lesion tissues. In HaCaT cells, WIF suppressed the production of regulated upon activation, normal T cell expressed and secreted (RANTES), thymus and activation-regulated chemokine (TARC), macrophage-derived chemokine (MDC), MCP-1, and MIP-3a, which are inflammatory cytokines and chemokines related to AD, and inhibited the mRNA expression of RANTES, TARC, and MDC in TNF-α/IFN-γ-stimulated HaCaT cells. Overall, the results revealed that WIF ameliorated AD-like skin inflammation by suppressing proinflammatory cytokine and chemokine production via nuclear factor-κB pathway inhibition, suggesting WIF as a potential candidate for AD treatment.

Production of Secondary Metabolites from Cell Cultures of Sageretia thea (Osbeck) M.C. Johnst. Using Balloon-Type Bubble Bioreactors.

Sageretia thea is used in the preparation of herbal medicine in China and Korea; this plant is rich in various bioactive compounds, including phenolics and flavonoids. The objective of the current study was to enhance the production of phenolic compounds in plant cell suspension cultures of Sageretia thea. Optimum callus was induced from cotyledon explants on MS medium containing 2,4-dichlorophenoxyacetic acid (2,4-D; 0.5 mg L-1), naphthalene acetic acid (NAA, 0.5 mg L-1), kinetin (KN; 0.1 mg L-1) and sucrose (30 g L-1). Browning of callus was successfully avoided by using 200 mg L-1 ascorbic acid in the callus cultures. The elicitor effect of methyl jasmonate (MeJA), salicylic acid (SA), and sodium nitroprusside (SNP) was studied in cell suspension cultures, and the addition of 200 µM MeJA was found suitable for elicitation of phenolic accumulation in the cultured cells. Phenolic and flavonoid content and antioxidant activity were determined using 2,2 Diphenyl 1 picrylhydrazyl (DPPH), 2,2'-azino-bis (3-ethybenzothiazoline-6-sulphonic acid (ABTS), ferric reducing antioxidant power (FRAP) assays and results showed that cell cultures possessed highest phenolic and flavonoid content as well as highest DPPH, ABTS, and FRAP activities. Cell suspension cultures were established using 5 L capacity balloon-type bubble bioreactors using 2 L of MS medium 30 g L-1 sucrose and 0.5 mg L-1 2,4-D, 0.5 mg L-1 NAA, and 0.1 mg L-1 KN. The optimum yield of 230.81 g of fresh biomass and 16.48 g of dry biomass was evident after four weeks of cultures. High-pressure liquid chromatography (HPLC) analysis showed the cell biomass produced in bioreactors possessed higher concentrations of catechin hydrate, chlorogenic acid, naringenin, and other phenolic compounds.

In vitro anti-cancer effects of beauvericin through inhibition of actin polymerization and Src phosphorylation.

BACKGROUND: Beauvericin (BEA) is a depsipeptide with antimicrobial, anti-inflammatory, and anti-cancer activities isolated from Beauveria bassiana. However, little is understood on its anti-cancer activities and mechanism. PURPOSE: Aim of this study was to explore the anti-cancer activity of BEA and its underlying molecular mechanism to provide a theoretical basis for its role as a candidate natural drug in cancer diseases. STUDY DESIGN: Various cancer cells such as C6 glioma, U251, MDA-MB-231, HeLa, HCT-15, LoVo cells, and HEK293T cells were used to the anti-cancer activity of BEA. METHODS: To evaluate the anti-cancer activity of BEA, cell viability test (MTT assay), morphological change check, confocal microscopy, actin polymerization assay, flow cytometry, and Western blotting analysis. To check the target enzyme of BEA, overexpression and site-directed mutagenesis was employed. RESULTS: BEA inhibited the viability of cancer cells including C6, MDA-MB-231, HeLa, HCT-15, LoVo, and U251 cells. Treatment of BEA in C6 glioma cells induced cell membrane blebbing and apoptosis. Caspase-3 and -9 were dose-dependently activated by BEA, and the mRNA expression of Bcl-2 was inhibited by BEA. According to confocal microscopy, actin polymerization and actin-actin interaction were interrupted by BEA in C6 cells. BEA regulated the apoptosis of C6 cells depending on the protein phosphorylation of Src and Signal transducer and activator of transcription (STAT3). Moreover, c-terminal amino acids in Src directly interacted with BEA in C6 cells, and the binding of Src and BEA suppressed the kinase activity of Src. CONCLUSIONS: These results suggest that BEA may be a critical candidate or substitute drug for cancer treatment via suppression of the Src/STAT3 pathway.

Inhibitory Effects of Grewia tomentosa Juss. on IgE-Mediated Allergic Reaction and DNCB-Induced Atopic Dermatitis.

Grewia tomentosa Juss. is a deciduous shrub that mainly grows in Asia. Despite studies of other Grewia species for treatment of various diseases, Grewia tomentosa Juss. has not been studied as a medicinal herb. This study evaluates the anti-allergic and anti-topic dermatitis activity of Grewia tomentosa Juss. ethanol extract (Gt-EE). The results show that Gt-EE suppressed IgE-antigen-induced ß-hexosaminidase release. The mRNA expression of IL-1ß, IL-4, IL-5, IL-6, IL-13, TNF-α, MCP-1, and TSLP, which are involved in allergic responses, was inhibited by Gt-EE in IgE-stimulated RBL-2H3 cells. In addition, the phosphorylation of Syk, PLCγ1, PKCδ, PI3K, AKT, NF-κB p65, NF-κB p50, p38, JNK, and ERK1/2 was decreased by Gt-EE in these cells. Gt-EE also showed anti-inflammatory effects in in vivo mouse models. In passive cutaneous anaphylaxis (PCA), a commonly used mouse model, Gt-EE decreased the allergic response, infiltration of mast cells, and mRNA level of IL-4. Furthermore, Gt-EE ameliorated symptoms of DNCB-induced atopic dermatitis (AD). In DNCB-induced AD, Gt-EE suppressed the increase in mast cells, serum IgE level, expression of allergic mediators (IL-1ß, IL-4, IL-5, IL-6, TNF-α), and phosphorylation of proteins (IκBα, NF-κB p65, NF-κB p50, p38, JNK, and ERK1/2) implicated in allergic reactions.

Vitamin C Suppresses Pancreatic Carcinogenesis through the Inhibition of Both Glucose Metabolism and Wnt Signaling.

Cumulative studies have indicated that high-dose vitamin C has antitumor effects against a variety of cancers. However, the molecular mechanisms underlying these inhibitory effects against tumorigenesis and metastasis, particularly in relation to pancreatic cancer, are unclear. Here, we report that vitamin C at high concentrations impairs the growth and survival of pancreatic ductal adenocarcinoma (PDAC) cells by inhibiting glucose metabolism. Vitamin C was also found to trigger apoptosis in a caspase-independent manner. We further demonstrate that it suppresses the invasion and metastasis of PDAC cells by inhibiting the Wnt/ß-catenin-mediated epithelial-mesenchymal transition (EMT). Taken together, our results suggest that vitamin C has therapeutic effects against pancreatic cancer.

Fluoroscopic criteria for on-site evaluation of failed intussusception reduction during air enema technique.

Background: There is no reliable fluoroscopic criteria for failed intussusception reduction during air enema technique. Methods: This retrospective case-control study included 373 episodes of ileocolic intussusceptions who had undergone air enema under fluoroscopy. All procedures were initially classified by conventional fluoroscopic criteria: presumptive successful procedures (PSP) vs. presumptive failed procedures (PFP). PFP were divided into true failure, false failure, and undetermined groups. The configuration and size of the residual mass were evaluated on fluoroscopic images. Statistical analyses included Mann-Whitney U-test, Fisher's exact test, receiver operating characteristic (ROC) analysis, logistic regression analyses, and Kruskal-Wallis rank sum test with a post hoc Tukey test. Results: PSP was 264 episodes (71%) and PFP was 109 episodes (29%). The true failure was 40 (37%) and false failure was 48 (44%). The true failure group commonly showed a larger size and round configuration for the residual mass than false failure (P<0.001). Multivariable analysis revealed configuration (P=0.004) and transverse diameter (P=0.007) as significant parameters that differentiated true and false failure. The optimal cut-off value of the transverse diameter of the residual mass was 2.3 cm. The sensitivity and specificity of conventional fluoroscopic criteria for failed reduction was 100% and 85%, respectively. The combination of new fluoroscopic findings and conventional criteria increased the specificity to 100%. Conclusions: Fluoroscopic finding of round-shape and larger size residual mass combined with conventional criteria may be useful for differentiating false failure from truly failed enema reduction in children with intussusception.

Inhibitory effect of Isatis tinctoria L. water extract on DNCB-induced atopic dermatitis in BALB/c mice and HaCaT cells.

BACKGROUND: Isatis tinctoria L (PLG) is a medicinal herb from the roots of Isatis indigotica Fort (Family Cruciferae). Previous studies have shown that PLG has anti-inflammatory and therapeutic effects against conditions such as acute and chronic hepatitis, various respiratory inflammations, and cancer. The purpose of this study was to define the pharmacological effects of PLG on inflammatory reactions and skin hyperkeratosis, which are the main symptoms of atopic dermatitis (AD), in vivo and in vitro. METHODS: For the AD in vivo experiment, 2,4-dinitrochlorobenzene (DNCB) induction and oral administration of PLG were performed on male BALB/c mice for four weeks. For in vitro experiments, keratinocytes were activated using TNF-α/IFN-γ in cultured human keratinocyte (HaCaT) cells. PLG inhibited inflammatory chemokine production and blocked the nuclear translocation of NF-κB p65 in activated keratinocytes. RESULTS: As a result of oral administration of PLG, dermis and epidermis thickening, as well as eosinophil and mast cell infiltration, were attenuated in AD skin lesions. In addition, the levels of immunoglobulin E (IgE), pro-inflammatory cytokines, and the MAPK/NF-κB signaling pathway were decreased in serum and dorsal skin tissues. Furthermore, PLG inhibited inflammatory chemokine production and blocked the nuclear translocation of NF-κB p65 in activated keratinocytes. In addition, epigoitrin and adenosine, the standard compounds of PLG, were identified as candidate AD compounds. CONCLUSIONS: These results indicate that PLG is a potent therapeutic agent for attenuating symptoms of AD.

Hymenocallis littoralis ameliorates inflammatory responses in LPS-stimulated RAW264.7 cells and HCl/EtOH-induced gastric mucosal injury via targeting the MAPK pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Hymenocallis littoralis (Jacq.) Salisb. Also known as Pancratium littorale Jacq. And Hymenocallis panamensis Lindl., is a medicinal plant from the family Amarylideceae used for emetic and wound healing and has manifested anti-neoplastic, anti-oxidant, and anti-viral properties. AIM OF THE STUDY: The aim of this paper is to investigate the anti-inflammatory potential and molecular mechanism of H. littoralis against lipopolysaccharide (LPS)-induced macrophages and in vivo HCl/EtOH-induced gastritis mucosal injury models. MATERIALS AND METHODS: The production of pro-inflammatory cytokines and mediators was evaluated by Griess assay, RT-PCR, and real-time PCR. Moreover, the relevant proteins of mitogen-activated protein kinases (MAPKs) including ERK, JNK, p38, c-Jun, and c-Fos were detected using immunoblotting. RESULTS: We demonstrated that H. littoralis prominently dampened production of nitric oxide (NO) in LPS-, poly I:C-, or pam3CSK-stimulated RAW264.7 cells; down-regulated the expression levels of interleukin 6 (IL-6) and inducible nitric oxide synthase; and markedly attenuated the luciferase activities of AP-1 reporter promoters. Moreover, H. littoralis administration prominently downregulated c-Fos and c-Jun phosphorylation as well as JNK1, ERK2, and MKK7 overexpression in HEK 293T cells. Furthermore, H. littoralis displayed anti-inflammatory effects in the HCl/EtOH-induced gastritis mice model. CONCLUSIONS: Cumulatively, these results demonstrated that H. littoralis exerts eminently anti-inflammatory activities in LPS-stimulated RAW264.7 cells in vitro and in HCl/EtOH-induced gastritis mice models in vivo. These activities could be attributed to its modulatory effects on the MAPK signaling pathway.

Synergistic effects of novel herbal decoctions from Panax ginseng and Morus alba on tyrosinase activity and melanogenesis in vitro.

Many medicinal plants such as a Panax ginseng and Morus alba (mulberry tree) have been widely used as depigmenting agents in Asia. To maximize their synergistic effects on melanogenesis, new herbal decoctions were created by mixing Ginseng Radix Alba (GR) and Mori Radicis Cortex (MC) at a ratio of 3:2 which called GMC decoction. A decoction of GR and Mori Ramulus (MR), which called GMR, was also formulated in order to compare the anti-melanogenic capacity. Combined decoctions, GMC and GMR, significantly decreased mushroom tyrosinase activity in vitro; however, single extracts, including MC and MR, showed weaker inhibitory activity. Melanin content assay and Fontana-Masson staining confirmed that two decoctions showed stronger inhibitory effects on the forskolin-induced melanin level in B16 cells, without cytotoxicity. Our findings suggest that ginseng in combination with mulberry tree enhances the anti-melanogenic effect in vitro.

Indigo Pulverata Levis (Chung-Dae, Persicaria tinctoria) Alleviates Atopic Dermatitis-like Inflammatory Responses In Vivo and In Vitro.

Atopic dermatitis (AD) is a chronic inflammatory skin disease associated with a type 2 T helper cell (Th2) immune response. The IndigoPulverata Levis extract (CHD) is used in traditional Southeast Asian medicine; however, its beneficial effects on AD remain uninvestigated. Therefore, we investigated the therapeutic effects of CHD in 2,4-dinitrochlorobenzene (DNCB)-induced BALB/c mice and tumor necrosis factor (TNF)-α- and interferon gamma (IFN)-γ-stimulated HaCaT cells. We evaluated immune cell infiltration, skin thickness, and the serum IgE and TNF-α levels in DNCB-induced AD mice. Moreover, we measured the expression levels of pro-inflammatory cytokines, mitogen-activated protein kinase (MAPK), and the nuclear factor-kappa B (NF-κB) in the mice dorsal skin. We also studied the effect of CHD on the translocation of NF-κB p65 and inflammatory chemokines in HaCaT cells. Our in vivo results revealed that CHD reduced the dermis and epidermis thicknesses and inhibited immune cell infiltration. Furthermore, it suppressed the proinflammatory cytokine expression and MAPK and NF-κB phosphorylations in the skin tissue and decreased serum IgE and TNF-α levels. In vitro results indicated that CHD downregulated inflammatory chemokines and blocked NF-κB p65 translocation. Thus, we deduced that CHD is a potential drug candidate for AD treatment.

Increased Anti-Allergic Effects of Secretome of Low-Level Light Treated Tonsil-Derived Mesenchymal Stem Cells in Allergic Rhinitis Mouse Model.

BACKGROUND: Low-level light therapy (LLLT) is widely used for the photobiomodulation of cell behavior. Recent studies have shown that LLLT affects the proliferation and migration of various types of mesenchymal stem cells (MSCs). However, there is a lack of studies investigating the effect of LLT on enhancing the immunomodulatory properties of tonsil-derived MSCs (T-MSCs). OBJECTIVE: The aim of this study was to investigate the immunomodulatory effects of conditioned media from T-MSCs (T-MSCs-CM) treated with LLLT in allergic inflammation. METHODS: We isolated T-MSCs from human palatine tonsils and evaluated the ingredients of T-MSCs-CM. The effect of T-MSCs-CM treated with LLLT was evaluated in a mouse model of allergic rhinitis (AR). We randomly divided the mice into four groups (negative control, positive control, T-MSCs-CM alone, and T-MSCs-CM treated with LLLT). To elucidate the therapeutic effect, we assessed rhinitis symptoms, serum immunoglobulin (Ig), the number of inflammatory cells, and cytokine expression. RESULTS: We identified increased expression of immunomodulatory factors, such as HGF, TGF-ß, and PGE, in T-MSCs-CM treated with LLLT, compared to T-MSCs-CM without LLLT. Our animal study demonstrated reduced allergic symptoms and lower expression of total IgE and OVA-specific IgE in the LLLT-treated T-MSCs-CM group compared to the AR group and T-MSCs-CM alone. Moreover, we found that T-MSCs-CM treated with LLLT showed significantly decreased infiltration of eosinophils, neutrophils, and IL-17 cells in the nasal mucosa and reduced IL-4, IL-17, and IFN-γ expression in OVA-incubated splenocytes compared to the AR group. CONCLUSIONS: The present study suggests that T-MSCs-CM treated with LLLT may provide an improved therapeutic effect against nasal allergic inflammation than T-MSCs-CM alone.

Sanguisorbae Radix Suppresses Colorectal Tumor Growth Through PD-1/PD-L1 Blockade and Synergistic Effect With Pembrolizumab in a Humanized PD-L1-Expressing Colorectal Cancer Mouse Model.

Immune checkpoints such as programmed death-1 (PD-1) have been proven as antitumor targets by enhancing cytotoxic T cell activity. All immune checkpoint blockades are antibody therapeutics that have large size and high affinity, as well as known immune-related side effects and low responses. To overcome the limitation of antibody therapeutics, we have explored PD-1/PD-L1 (programmed death-ligand 1) blockades in traditional oriental medicine, which has a long history but has not yet studied PD-1/PD-L1 blockades. Sanguisorbae Radix extract (SRE) blocked PD-1 and PD-L1 binding in competitive ELISA. SRE effectively inhibited the PD-1/PD-L1 interaction, thereby improving T cell receptor (TCR) signaling and the NFAT-mediated luciferase activity of T cells. SRE treatment reduced tumor growth in the humanized PD-L1 MC38 cell allograft humanized PD-1 mouse model. Additionally, the combination of SRE and pembrolizumab (anti-PD-1 antibody) suppressed tumor growth and increased infiltrated cytotoxic T cells to a greater extent did either agent alone. This study showed that SRE alone has anticancer effects via PD-1/PD-L1 blockade and that the combination therapy of SRE and pembrolizumab has enhanced immuno-oncologic effects.

The Effects of Korean Red Ginseng on Biological Aging and Antioxidant Capacity in Postmenopausal Women: A Double-Blind Randomized Controlled Study.

Postmenopausal women are vulnerable to aging and oxidative stress due to reduced estrogen. Previous studies have shown that Korean red ginseng (KRG) has beneficial effects on aging and antioxidant capacity. Therefore, we evaluated the effects of KRG on biological aging and antioxidant capacity in postmenopausal women. This study conducted a double-blinded, placebo-controlled clinical trial. The participants were randomly administered KRG or a placebo, and the following metrics were measured: mitochondria DNA (mtDNA) copy number as an indicator of biological aging and, total antioxidant status (TAS) as a marker of antioxidant capacity. Clinical symptoms of fatigue, as measured by the fatigue severity scale, were assessed before and after KRG administration. There were 63 participants, of whom 33 received KRG and 30 received a placebo. The mtDNA copy number (KRG group: 1.58 ± 2.05, placebo group: 0.28 ± 2.36, p = 0.023) and TAS (KRG group: 0.11 ± 0.25 mmol/L, placebo group: -0.04 ± 0.16 mmol/L, p = 0.011) increased and the fatigue severity scale (KRG group: -7 ± 12, placebo group: -1 ± 11, p = 0.033) decreased significantly more in the KRG group than the placebo group. KRG significantly increased the mtDNA copy number, total antioxidant status, and improved symptoms of fatigue in postmenopausal women.

Matricaria chamomilla (Chamomile) Ameliorates Muscle Atrophy in Mice by Targeting Protein Catalytic Pathways, Myogenesis, and Mitochondrial Dysfunction.

Muscle atrophy, or loss of skeletal muscle, is caused by aging, malnutrition, immobility through injury, or diseases such as cancer. Chamomile (Matricaria chamomilla L.) contains various active components, including flavonoids, sesquiterpenes, polyacetylenes, and coumarins, and is used in various herbal medicines in the European Pharmacopoeia. In this study, we investigated the effects of ethanol extract of chamomile [Formula: see text](MC) on muscle wasting and its mechanism of action. Mice with dexamethasone (DEX)-induced muscle atrophy were orally administered MC (100, 200, and 300 mg/kg) for 4 weeks. Micro-computed tomography analysis showed that MC (200 and 300 mg/kg) significantly recovered DEX-induced loss of muscle volume, density, and weight and MC-treated DEX-induced mice also showed increased moving distance and grip strength. MC suppressed the mRNA level of muscle RING finger 1 (MuRF1) while increasing the expression of mitochondrial transcription factor A (TFAM), MyoD, and Myogenin-1. We found 25 peaks in MC samples through HPLC analysis and identified 6 peaks by comparison with a profile of standard compounds: chlorogenic acid (CGA), luteolin-7-O-glucoside (L7G), patulitrin, apigenin-7-O-glucoside (A7G), herniarin, and (E)-tonghaosu. Of these components, the gene expression of MyoD was significantly augmented by patulitrin, herniarin, CGA, and L7G in C2C12 cells, while Myogenin-1 gene expression was increased by A7G, patulitrin, herniarin, CGA, and L7G. Moreover, TFAM gene expression and phosphorylation of AKT were increased by all six ingredients. Based on our results, we suggest MC for use as a supplement or remedy for muscle wasting, including cachexia and sarcopenia.

Antioxidative and Skin Protective Effects of Canarium subulatum Methanol Extract on Keratinocytes.

Canarium subulatum is a traditional medical herb used in South Asia. Recently, the anti-inflammatory effects of C. subulatum methanol extract (Cs-ME) have been reported; however, the effect of Cs-ME on skin physiology has not yet been elucidated. Therefore, in this study, we evaluated the protective effect of Cs-ME on UV-induced skin aging and cell death as well as the reinforcing effect on the skin barrier. According to viable cell counting and MTT assays, Cs-ME significantly reduced UV-evoked HaCaT cell death. Cs-ME blocked reactive oxygen species (ROS) generation in UV-irradiated HaCaT cells and showed radical scavenging activity against DPPH and ABTS. In addition, H2O2-induced cell death was inhibited by Cs-ME, indicating that Cs-ME protects cells from UV-derived cell death through the suppression of ROS. PCR analysis revealed that Cs-ME diminished the expression of aging-related HYAL-1 and MMP-1 genes in UV-treated HaCaT cells. Elevated HYAL-1 and MMP-1 mRNA expression in H2O2-stimulated HaCaT cells was also decreased by Cs-ME, suggesting that Cs-ME exerts antiaging activity via the inhibition of ROS. Expression of skin barrier components including filaggrin and hyaluronic acid synthase-1 was increased by Cs-ME and was modulated by ERK/p38-AP-1 signaling. Collectively, our data show that Cs-ME has cytoprotective and antiaging activity based on antioxidant properties. Furthermore, Cs-ME exerts skin barrier protective ability by regulating the AP-1 signaling pathway. Therefore, Cs-ME has the potential for use as an ingredient in cosmetics to protect the skin from UV irradiation, prevent photoaging, and strengthen the skin barrier.

Syk/NF-κB-targeted anti-inflammatory activity of Melicope accedens (Blume) T.G. Hartley methanol extract.

ETHNOPHARMACOLOGICAL RELEVANCE: Melicope accedens (Blume) Thomas G. Hartley is a plant included in the family Rutaceae and genus Melicope. It is a native plant from Vietnam that has been used for ethnopharmacology. In Indonesia and Malaysia, the leaves of M. accedens are applied externally to decrease fever. AIM OF THE STUDY: The molecular mechanisms of the anti-inflammatory properties of M. accedens are not yet understood. Therefore, we examined those mechanisms using a methanol extract of M. accedens (Ma-ME) and determined the target molecule in macrophages. MATERIALS AND METHODS: We evaluated the anti-inflammatory effects of Ma-ME in lipopolysaccharide (LPS)-stimulated RAW264.7 cells and in an HCl/EtOH-triggered gastritis model in mice. To investigate the anti-inflammatory activity, we performed a nitric oxide (NO) production assay and ELISA assay for prostaglandin E2 (PGE2). RT-PCR, luciferase gene reporter assays, western blotting analyses, and a cellular thermal shift assay (CETSA) were conducted to identify the mechanism and target molecule of Ma-ME. The phytochemical composition of Ma-ME was analyzed by HPLC and LC-MS/MS. RESULTS: Ma-ME suppressed the production of NO and PGE2 and the mRNA expression of proinflammatory genes (iNOS, IL-1ß, and COX-2) in LPS-stimulated RAW264.7 cells without cytotoxicity. Ma-ME inhibited NF-κB activation by suppressing signaling molecules such as IκBα, Akt, Src, and Syk. Moreover, the CETSA assay revealed that Ma-ME binds to Syk, the most upstream molecule in the NF-κB signal pathway. Oral administration of Ma-ME not only alleviated inflammatory lesions, but also reduced the gene expression of IL-1ß and p-Syk in mice with HCl/EtOH-induced gastritis. HPLC and LC-MS/MS analyses confirmed that Ma-ME contains various anti-inflammatory flavonoids, including quercetin, daidzein, and nevadensin. CONCLUSIONS: Ma-ME exhibited anti-inflammatory activities in vitro and in vivo by targeting Syk in the NF-κB signaling pathway. Therefore, we propose that Ma-ME could be used to treat inflammatory diseases such as gastritis.

Sauropus brevipes ethanol extract negatively regulates inflammatory responses in vivo and in vitro by targeting Src, Syk and IRAK1.

CONTEXT: Sauropus brevipes Müll. Arg. (Phyllanthaceae) has been used as an effective ingredient in a decoction for the treatment of diarrhoea. However, there was no report on its modulatory role in inflammation. OBJECTIVE: This study investigates anti-inflammatory effect of S. brevipes in various inflammation models. MATERIALS AND METHODS: The aerial part of S. brevipes was extracted with 95% ethanol to produce Sb-EE. RAW264.7 cells pre-treated with Sb-EE were stimulated by lipopolysaccharide (LPS), and Griess assay and PCR were performed. High-performance liquid chromatography (HPLC) analysis, luciferase assay, Western blotting and kinase assay were employed. C57BL/6 mice (10 mice/group) were orally administered with Sb-EE (200 mg/kg) once a day for five days, and peritonitis was induced by an intraperitoneal injection of LPS (10 mg/kg). ICR mice (four mice/group) were orally administered with Sb-EE (20 or 200 mg/kg) or ranitidine (positive control) twice a day for two days, and EtOH/HCl was orally injected to induce gastritis. RESULTS: Sb-EE suppressed nitric oxide (NO) release (IC50=34 µg/mL) without cytotoxicity and contained flavonoids (quercetin, luteolin and kaempferol). Sb-EE (200 µg/mL) reduced the mRNA expression of inducible NO synthase (iNOS). Sb-EE blocked the activities of Syk and Src, while inhibiting interleukin-1 receptor associated kinases (IRAK1) by 68%. Similarly, orally administered Sb-EE (200 mg/kg) suppressed NO production by 78% and phosphorylation of Src and Syk in peritonitis mice. Sb-EE also decreased inflammatory lesions in gastritis mice. DISCUSSION AND CONCLUSIONS: This study demonstrates the inhibitory effect of Sb-EE on the inflammatory response, suggesting that Sb-EE can be developed as a potential anti-inflammatory agent.