Anti-Cancer Effects of α-Cubebenoate Derived from Schisandra chinensis in CT26 Colon Cancer Cells.

α-Cubebenoate derived from Schisandra chinensis has been reported to possess anti-allergic, anti-obesity, and anti-inflammatory effects and to exhibit anti-septic activity, but its anti-cancer effects have not been investigated. To examine the anti-cancer activity of α-cubebenoate, we investigated its effects on the proliferation, apoptosis, and metastasis of CT26 cells. The viabilities of CT26 cells (a murine colorectal carcinoma cell line) and HCT116 cells (a human colon cancer cell line) were remarkably and dose-dependently diminished by α-cubebenoate, whereas the viability of CCD-18Co cells (a normal human fibroblast cell line) were unaffected. Furthermore, α-cubebenoate treatment increased the number of apoptotic CT26 cells as compared with Vehicle-treated cells and increased Bax, Bcl-2, Cas-3, and Cleaved Cas-3 protein levels by activating the MAP kinase signaling pathway. α-Cubebenoate also suppressed CT26 migration by regulating the PI3K/AKT signaling pathway. Furthermore, similar reductions were observed in the expression levels of some migration-related proteins including VEGFA, MMP2, and MMP9. Furthermore, reduced VEGFA expression was found to be accompanied by the phosphorylations of FAK and MLC in the downstream signaling pathway of adhesion protein. The results of the present study provide novel evidence that α-cubebenoate can stimulate apoptosis and inhibit metastasis by regulating the MAPK, PI3K/AKT, and FAK/MLC signaling pathways.

Novel Characterization of Constipation Phenotypes in ICR Mice Orally Administrated with Polystyrene Microplastics.

Indirect evidence has determined the possibility that microplastics (MP) induce constipation, although direct scientific proof for constipation induction in animals remains unclear. To investigate whether oral administration of polystyrene (PS)-MP causes constipation, an alteration in the constipation parameters and mechanisms was analyzed in ICR mice, treated with 0.5 µm PS-MP for 2 weeks. Significant alterations in water consumption, stool weight, stool water contents, and stool morphology were detected in MP treated ICR mice, as compared to Vehicle treated group. Also, the gastrointestinal (GI) motility and intestinal length were decreased, while the histopathological structure and cytological structure of the mid colon were remarkably altered in treated mice. Mice exposed to MP also showed a significant decrease in the GI hormone concentration, muscarinic acetylcholine receptors (mAChRs) expression, and their downstream signaling pathway. Subsequent to MP treatment, concentrations of chloride ion and expressions of its channel (CFTR and CIC-2) were decreased, whereas expressions of aquaporin (AQP)3 and 8 for water transportation were downregulated by activation of the mitogen-activated protein kinase (MAPK)/nuclear factor (NF)-κB signaling pathway. These results are the first to suggest that oral administration of PS-MP induces chronic constipation through the dysregulation of GI motility, mucin secretion, and chloride ion and water transportation in the mid colon.

Laxative Effects of Phlorotannins Derived from Ecklonia cava on Loperamide-Induced Constipation in SD Rats.

This study investigated the laxative effects of phlorotannins (Pt) derived from Ecklonia cava (E. cave) on chronic constipation by evaluating alterations in stool parameters, gastrointestinal motility, histopathological structure, mucin secretion, gastrointestinal hormones, muscarinic cholinergic regulation, and fecal microbiota in SD rats with loperamide (Lop)-induced constipation subjected to Pt treatment. Stool-related parameters (including stool number, weight, and water contents), gastrointestinal motility, and length of intestine were significantly enhanced in the Lop+Pt-treated group as compared to the Lop+Vehicle-treated group. A similar recovery was detected in the histopathological and cytological structure of the mid-colon of Lop+Pt-treated rats, although the level of mucin secretion remained constant. Moreover, rats with Lop-induced constipation subjected to Pt treatment showed significant improvements in water channel expression, gastrointestinal hormone secretions, and expression of muscarinic acetylcholine receptors M2/M3 (mAChRs M2/M3) and their mediators of muscarinic cholinergic regulation. Furthermore, the Lop+Pt-treated group showed a significant recovery of Bifidobacteriaceae, Muribaculaceae, Clostridiaceae, and Eubacteriaceae families in fecal microbiota. Taken together, these results provide the first evidence that exposure of SD rats with Lop-induced constipation to Pt improves the constipation phenotype through the regulation of membrane water channel expression, GI hormones, the mAChR signaling pathway, and fecal microbiota.

Molecular Characterization of Constipation Disease as Novel Phenotypes in CRISPR-Cas9-Generated Leptin Knockout Mice with Obesity.

(1) Background: We characterized a novel animal model with obesity-induced constipation because constipation is rarely known in genetically engineered mice (GEM); (2) Methods: The changes in the constipation parameters and mechanisms were analyzed in CRISPR-Cas9-mediated leptin (Lep) knockout (KO) mice from eight to 24 weeks; (3) Results: Significant constipation phenotypes were observed in the Lep KO mice since 16 weeks old. These mice showed a significant decrease in the gastrointestinal motility, mucosal layer thickness and ability for mucin secretion as well as the abnormal ultrastructure of Lieberkühn crypts in the transverse colon. The density or function of the enteric neurons, intestinal Cajal cells (ICC), smooth muscle cells, and the concentration of gastrointestinal (GI) hormones for the GI motility were remarkably changed in Lep KO mice. The downstream signaling pathway of muscarinic acetylcholine receptors (mAChRs) were activated in Lep KO mice, while the expression of adipogenesis-regulating genes were alternatively reduced in the transverse colon of the same mice; (4) Conclusions: These results provide the first strong evidence that Lep KO mice can represent constipation successfully through dysregulation of the GI motility mediated by myenteric neurons, ICC, and smooth muscle cells in the transverse colon during an abnormal function of the lipid metabolism.

Novel Function of α-Cubebenoate Derived from Schisandra chinensis as Lipogenesis Inhibitor, Lipolysis Stimulator and Inflammasome Suppressor.

The efficacy of α-cubebenoate isolated from Schisandra chinensis has been previously studied in three disease areas, namely inflammation, sepsis, and allergy, and its role in other diseases is still being explored. To identify the novel function of α-cubebenoate on lipid metabolism and related inflammatory response, alterations in fat accumulation, lipogenesis, lipolysis, and inflammasome activation were measured in 3T3-L1 preadipocytes and primary adipocytes treated with α-cubebenoate. Lipid accumulation significantly decreased in MDI (3-isobutyl-1-methylxanthine, dexamethasone, and insulin)-stimulated 3T3-L1 adipocytes treated with α-cubebenoate without any significant cytotoxicity. The mRNA levels of peroxisome proliferator-activated receptor (PPAR)γ and CCAAT-enhancer binding protein (C/EBP) α for adipogenesis, as well as adipocyte fatty acid binding protein 2 (aP2) and fatty acid synthetase (FAS) for lipogenesis, were reduced after α-cubebenoate treatment, while cell cycle arrest at G2/M stage was restored in the same group. α-cubebenoate treatment induced glycerol release in primary adipocytes and enhanced expression of lipolytic proteins (HSL, perilipin, and ATGL) expression in MDI-stimulated 3T3-L1 adipocytes. Inflammasome activation and downstream cytokines expression were suppressed with α-cubebenoate treatment, but the expression of insulin receptor signaling factors was remarkably increased by α-cubebenoate treatment in MDI-stimulated 3T3-L1 adipocytes. These results indicate that α-cubebenoate may play a novel role as lipogenesis inhibitor, lipolysis stimulator, and inflammasome suppressor in MDI-stimulated 3T3-L1 adipocytes. Our results provide the possibility that α-cubebenoate can be considered as one of the candidates for obesity management.

Therapeutic Effects of Cold-Pressed Perilla Oil Mainly Consisting of Linolenic acid, Oleic Acid and Linoleic Acid on UV-Induced Photoaging in NHDF Cells and SKH-1 Hairless Mice.

Positive physiological benefits of several plant oils on the UV-induced photoaging have been reported in some cell lines and model mice, but perilla oil collected from the seeds of Perilla frutescens L. has not been investigated in this context. To study the therapeutic effects of cold-pressed perilla oil (CPO) on UV-induced photoaging in vitro and in vivo, UV-induced cellular damage and cutaneous photoaging were assessed in normal human dermal fibroblasts (NHDFs) and HR-1 hairless mice. CPO contained five major fatty acids including linolenic acid (64.11%), oleic acid (16.34%), linoleic acid (11.87%), palmitic acid (5.06%), and stearic acid (2.48%). UV-induced reductions in NHDF cell viability, ROS production, SOD activity, and G2/M cell cycle arrest were remarkably improved in UV + CPO treated NHDF cells as compared with UV + Vehicle treated controls. Also, UV-induced increases in MMP-1 protein and galactosidase levels were remarkably suppressed by CPO. In UV-radiated hairless mice, topical application of CPO inhibited an increase in wrinkle formation, transepidermal water loss (TEWL), erythema value, hydration and melanin index on dorsal skin of UVB-irradiated hairless mice. CPO was observed to similarly suppress UV-induced increases in epidermal thickness, mast cell numbers, and galactosidase and MMP-3 mRNA levels. These results suggest CPO has therapeutic potential in terms of protecting against skin photoaging by regulating skin morphology, histopathology and oxidative status.

α-Linolenic Acid-Enriched Cold-Pressed Perilla Oil Suppress High-Fat Diet-Induced Hepatic Steatosis through Amelioration of the ER Stress-Mediated Autophagy.

Perilla oil has been considered to have excellent potential for treating various diseases due to its contents of beneficial fatty acids, such as α-linolenic acid, oleic acid and linoleic acid. The therapeutic effects and molecular mechanism of an α-linolenic acid-enriched cold-pressed perilla oil (LEP) on hepatic steatosis of an obesity model were investigated by analyzing alterations in fat accumulation and endoplasmic reticulum (ER) stress-mediated autophagy, in high-fat diet (HFD)-induced obesity C57BL/6N mice treated with LEP for 16 weeks. Although no significant alterations were detected in body weight and most organ weights, the liver weight and accumulation of lipid droplets in the liver section were significantly lower in HFD + LEP treated group as compared to the HFD + Vehicle treated group. Reduced mRNA expression levels of adipogenesis and lipogenesis regulating factors, including the peroxisome proliferator-activated receptor (PPAR)γ, CCAAT/enhancer-binding protein (C/EBP)α, fatty acid synthase (FAS), and adipocyte fatty acid-binding protein 2 (aP2) were observed after LEP treatment for 16 weeks, while the levels of lipolysis were remarkably increased in the same group. Moreover, the LEP-treated groups showed suppression of ER stress-regulating factors, such as the C/EBP homologous protein (CHOP), eukaryotic translation initiation factor 2α (eIF2α), inositol-requiring protein 1 (IRE1)α, and Jun-N-terminal kinase (JNK) during anti-hepatic steatosis effects. The expression level of the microtubule-associated protein 1A/1B-light chain 3 (LC3) protein and phosphatidylinositol-3-kinase (PI3K)/AKT/ mammalian target of rapamycin (mTOR) pathway for the autophagy response showed a significant decrease in the HFD+LEP-treated group. Furthermore, ER stress-mediated autophagy was accompanied with enhanced phosphorylation of extracellular signal-regulated kinase (ERK), JNK, and p38 protein in the mitogen-activated protein (MAP) kinase signaling pathway. Taken together, the results of the present study indicate that treatment with LEP inhibits hepatic steatosis in the HFD-induced obese model through regulation of adipogenesis and lipolysis. We believe our results are the first to show that the anti-hepatic steatosis activity of α-linolenic acid from cold-pressed perilla oil might be tightly correlated with the amelioration of ER stress-mediated autophagy.

Anti-inflammatory effects of Capparis ecuadorica extract in phthalic-anhydride-induced atopic dermatitis of IL-4/Luc/CNS-1 transgenic mice.

CONTEXT: The natural products derived from Capparis ecuadorica H.H. Iltis (Capparaceae) could have great potential for anti-inflammation since they inhibited the inflammatory response in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. OBJECT: This study investigated the anti-inflammatory effects and related mechanism of methanol extract of C. ecuadorica leaves (MCE) during atopic dermatitis (AD) responses. MATERIALS AND METHODS: Alterations in the phenotypical markers for AD, luciferase signal, iNOS-mediated COX-2 induction pathway, and inflammasome activation were analysed in non-Tg (n = 5) and 15% phthalic anhydride (PA) treated IL-4/Luc/CNS-1 transgenic (Tg) HR1 mice (n = 5 per group), subsequent to treatment with acetone-olive oil (AOO), vehicle (DMSO) and two dose MCE (20 and 40 mg/kg) three times a week for 4 weeks. RESULTS: MCE treatment reduced the intracellular ROS level (48.2%), NO concentration (7.1 mmol/L) and inflammatory cytokine expressions (39.1%) in the LPS-stimulated RAW264.7 cells. A significant decrease was detected for ear thickness (16.9%), weight of lymph node (0.7 mg), IgE concentration (1.9 µg/mL), and epidermal thickness (31.8%) of the PA + MCE treated Tg mice. MCE treatment induced the decrease of luciferase signal derived from the IL-4 promoter and the recovery of the IL-4 downstream regulator cytokines. PA + MCE treated Tg mice showed decreasing infiltration of mast cells (42.5%), iNOS-mediated COX-2 induction pathway, MAPK signalling pathway and inflammasome activation in the ear tissue. CONCLUSIONS: These findings provide the first evidence that MCE may have great potential to suppress chemical-induced skin inflammation through the suppression of IL-4 cytokine and the iNOS-mediated COX-2 induction pathway, and activation of inflammasome.

Comparison of immunophenotypes between Rag2 knockout mice derived from two different sources.

BACKGROUND: Recombination activating gene2 (Rag2) knockout (KO) mice are used widely in various research fields, including vaccine development, transplantation studies, and hematopoiesis research, but few studies have compared their phenotypes. This study examined whether there were differences in the immunophenotypes between Rag2 KO mice derived from different sources. In particular, the changes in the organ weight, histological structure, and subpopulation of T and B cells were compared in the spleen and thymus of C57BL/6-Rag2em1hwl/Korl (Rag2/Korl KO) and B6.Cg-Rag2tm1.1Cgn/J (Rag2/J KO) mice. RESULTS: The weight of the spleen and thymus similarly decreased in the Rag2/Korl and Rag2/J KO mice compared to their wild type (WT) mice, even though the other organs were kept at the same weight. A slight difference between the Rag2/Korl and Rag2/J KO group were detected in the number of white blood cells (WBC), lymphocytes (LYM), red cell distribution width (RDW), and platelets (PLT). In addition, the white pulp of the spleen and the cortex region of the thymus decreased in both Rag2 KO mice compared to WT mice. On the other hand, significant differences in the number of CD8+ T and B cell subpopulations between WT and Rag2 KO mice were observed between Rag2/Korl and Rag2/J KO group, while the CD4+ T subpopulation was maintained similarly in both groups. CONCLUSIONS: These results suggest that Rag2/Korl and Rag2/J KO mice exhibit similar immunophenotypes in the spleen and thymus except for the differences in the number of CD8+ T and B cell subpopulations.

Anti­tumor effects of an aqueous extract of Ecklonia cava in BALB/cKorl syngeneic mice using colon carcinoma CT26 cells.

Ecklonia cava (E. cava) is well known as one of edible alga that contains various unique polyphenols. The anti­tumor activity of an aqueous extract of E. cava (AEC) against colon carcinoma was evaluated by analyzing the alterations in tumor growth, histopathological structure and molecular mechanisms in CT26 tumor­bearing BALB/cKorl syngeneic mice after administrating AEC for five weeks. AEC contained high total phenolic contents and demonstrated significant scavenging activity against 2,2­diphenyl­1­picrylhydrazyl radicals. Marked anti­tumor effects were demonstrated in the AEC­treated CT26 cells. In the in vivo syngeneic model, the AEC treatment decreased the volume and weight of CT26 tumors, and expanded the necrotic region in the hematoxylin and eosin stained tumor sections. The inhibitory effects of AEC on tumor growth were reflected by the increased level of apoptotic proteins, inhibition of cell proliferation, suppression of metastasis ability and increase in tumor­suppressing activity in CT26 tumor­bearing BALB/cKorl syngeneic mice. The potential function of phlorotannin (PT), one of the primary active compounds in AEC, was demonstrated by the increased cytotoxicity, apoptosis and suppression of cell proliferation in PT­treated CT26 cells. Overall, the results of the present study provide novel scientific evidence that AEC can suppress the growth of CT26 colon cancer by activating apoptosis, suppressing cell proliferation, inhibiting cell migration and enhancing the tumor­suppressing activity.

Evaluation of Deodorizing Effects of Saccharina japonica in 10-Month-Old ICR Mice Using a Novel Odor Marker Associated with Aging.

The potential deodorizing effects of Saccharina japonica have been evaluated by determining their deodorizing performance, but they are yet to be validated in experimental animals. The deodorizing effects of S. japonica were examined in an animal model using a novel odor marker associated with aging by comparing the concentration of odor component in urine obtained from two- and 10-month-old ICR mice using gas chromatography-mass spectrometry (GC-MS), and the changes in the trimethylamine (TMA) concentration, ammonia level, and structure of sweat gland were determined after exposing 10-month-old ICR mice to 70% ethanol extract of S. japonica (EESJ) for four weeks. In vitro analysis was performed to confirm the composition of EESJ with respect to the total flavonoid contents (TFC, 28.6 ± 2.5 mg/g), total polyphenol contents (TPC, 107.3 ± 8.9 mg/g), and total condensed tannin contents (TTC, 65.7 ± 5.2 mg/g) contents, as well as to the deodorizing performance to ammonia and acetic acid (91.2 ± 7.8% and 54.8 ± 6.3%, respectively). In vivo analysis revealed TMA to be the novel odor marker associated with aging among the 19 odor components evaluated, considering the higher concentration in the urine of 10-month-old ICR mice. The peak area of TMA on the gas chromatogram was significantly lower in the 10-month-old ICR mice treated with EESJ than in the two-month-old mice. A similar decrease was observed in the level of ammonia obtained from the dirty bedding of the EESJ-treated group. Moreover, tissues obtained from the mouse foot of the group exposed to EESJ showed a dose-dependent decrease in the gland tube number of sweat glands and the TMA dehydrogenase transcription level. Overall, these results provide novel evidence that the administration of EESJ helps reduce the body TMA and ammonia concentrations, resulting in reduced odor and a decrease in the number of sweat glands and the expression of TMA dehydrogenase in the ICR mouse feet.

Promotion of the inflammatory response in mid colon of complement component 3 knockout mice.

To determine whether complement component 3 (C3) deficiency affects its receptor downstream-mediated inflammatory response, the current study was undertaken to measure alterations in the inducible nitric oxide synthase (iNOS)­mediated cyclooxygenase­2 (COX­2) induction pathway, inflammasome pathway, nuclear factor-κB (NF-κB) activation, and inflammatory cytokine expressions in the mid colon of C3 knockout (KO) mice. Significant enhancement was observed in expressions of key components of the iNOS­mediated COX­2 induction pathway, and in the phosphorylation of mitogen­activated protein (MAP) kinase members. A similar pattern of increase was also observed in the expression levels of inflammasome proteins in C3 KO mice. Moreover, compared to WT mice, C3 KO mice showed remarkably enhanced phosphorylation of NF-κB and Inhibitor of κB-α (IκB-α), which was reflected in entirety as increased expressions of Tumor necrosis factor (TNF), IL-6 and IL-1α. However, the levels of E-cadherin, tight junction channels and ion channels expressions were lower in the C3 KO mice, although myeloperoxidase (MPO) activity for neutrophils was slightly increased. Taken together, results of the current study indicate that C3 deficiency promotes inflammatory responses in the mid colon of C3 KO mice through activation of the iNOS­mediated COX­2 induction pathway, Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC)-inflammasome pathway and NF-κB signaling pathway, and the enhancement of inflammatory cytokine expressions.

Loperamide-induced Constipation Activates Inflammatory Signaling Pathways in the Mid Colon of SD Rats Via Complement C3 and its Receptors.

BACKGROUND: Complement component 3 (C3) receptors play an important role as inflammatory mediators in the innate immune system, although their mechanisms were not well studied during constipation. OBJECTIVE: The aim of this study is to investigate the regulatory role of C3 and its receptors' downstream signaling during constipation. METHODS: Alterations in the C3, C3a receptor (C3aR), and C3b receptor (C3bR) expressions, PI3K/AKT pathway, RhoA/MLC pathway, MAP kinase pathway, and inflammatory cytokine expressions were measured in the mid colon of loperamide (Lop) treated SD rats. RESULTS: Lop treatment successfully induced constipation phenotypes, including decreased stool parameters and histological structure alterations. The expression levels of C3 were significantly increased, whereas expressions of C3aR and C3bR were decreased during Lop-induced constipation. Moreover, significant upregulation was observed in the phosphorylation levels of PI3K, AKT, and GSK3ß in mid colons of Lop treated SD rats. The expression of RhoA and phosphorylation of MLC were also enhanced in the Lop treated group. Furthermore, a similar pattern was detected in the MAP kinase pathway and inflammatory cytokine expressions. Subsequent to the Lop treatment, the phosphorylation of ERK and p38, as well as the mRNA levels of NF-κB, TNF-α, IL-6 and IL-1α were remarkably increased in the mid colon. CONCLUSION: These results indicate that Lop-induced constipation is tightly linked to the downregulation of C3aR and C3bR expressions, and upregulation of the C3 and C3Rs downstream signaling pathway, including PI3K/AKT, RhoA/MLC, and MAP kinase pathways as well as inflammatory cytokine expressions in the mid colon of SD rats.

Novel role of Dipterocarpus tuberculatus as a stimulator of focal cell adhesion through the regulation of MLC2/FAK/Akt signaling pathway.

To investigate a novel function of Dipterocarpus tuberculatus on focal cell adhesion stimulation, alterations to the regulation of focal cell adhesion-related factors were analyzed in NHDF cells and a calvarial defect rat model after treatment with methanol extracts of D. tuberculatus (MED). MED contained gallic acid, caffeic acid, ellagic acid, and naringenin in high concentrations. The proliferation activity, focal cell adhesion ability, adhesion receptors-mediated signaling pathway in NHDF cells were increased by MED. Also, a dense adhered tissue layer and adherent cells on MED-coated titanium plate (MEDTiP) surfaces were detected during regeneration of calvarial bone. The results of the present study provide novel evidence that MED may stimulate focal cell adhesion in NHDF cells and a calvarial defect rat model.

Dysbiosis of Fecal Microbiota From Complement 3 Knockout Mice With Constipation Phenotypes Contributes to Development of Defecation Delay.

Significant phenotypes for constipation were detected in complement 3 (C3) knockout (KO) mice, although no research has been conducted on an association with alteration of gut microbiota. To investigate the effects of dysbiosis on fecal microbiota from C3 KO mice with constipation, the composition of fecal microbiota was characterized in mid-colons of 16-week-old C3 KO mice, and their function for defecation delay development was examined after fecal microbiota transplantation (FMT) of C3 KO mice. Some significant alterations in constipation phenotypes, including stool parameters and histopathological structure, were detected in 16-week-old C3 KO mice compared to those of wild-type (WT) mice. Fecal microbiota of C3 KO mice exhibited decreases in Anaerocolumna, Caecibacterium, Christensenella, Kineothrix, and Oscillibacter populations and increases in Prevotellamassilia, Reuthenibacterium, Prevotella, Eubacterium, Culturomica, Bacteroides, and Muribaculum populations. In FMT study, key stool parameters, including weight and water content, were remarkably declined in a transplanted KO (KFMT) group of antibiotics-induced depletion of microbiota (AiDM)-WT and AiDM-KO mice, and a similar change was observed in fecal morphology. However, intestine length decreased in only the KFMT group of AiDM-WT mice compared with that of AiDM-KO mice. The mucosal layer and muscle thickness were commonly decreased in the KFMT group of AiDM-WT and AiDM-KO mice, and significant alterations in the crypt structure of Lieberkuhn and molecular regulators, including AQP8, C-kit, and 5-HT, were observed in the same group. Taken together, results of the present study indicate that dysbiosis of fecal microbiota from C3 KO mice with constipation phenotypes has a key role in the induction and regulation of defecation delay.

Therapeutic Effects of Dipterocarpus tuberculatus with High Antioxidative Activity Against UV-Induced Photoaging of NHDF Cells and Nude Mice.

To investigate the therapeutic effects of methanol extracts of Dipterocarpus tuberculatus Roxb. (MED) against UV-induced photoaging, we assessed for alterations in the antioxidant activity, anti-apoptotic effects, ECM modulation, skin appearances, and anti-inflammatory response in normal human dermal fibroblast (NHDF) cells and nude mice orally treated with MED. High levels of tannin content and high free radical scavenging activity to DPPH were determined in MED, while seven active components, namely, gallic acid, bergenin, ellagic acid, ε-viniferin, asiatic acid, oleanolic acid, and 2α-hydroxyursolic acid, were identified using LC-MS analyses. UV-induced alterations in the NO concentration, SOD activity, and Nrf2 expression were remarkably recovered in MED-treated NHDF cells. Moreover, the decreased number of apoptotic cells and G2/M phase arrest were observed in the UV + MED-treated groups. Similar recoveries were detected for ß-galactosidase, MMP-2/9 expression, and intracellular elastase activity. Furthermore, MED treatment induced suppression of the COX-2-induced iNOS mediated pathway, expression of inflammatory cytokines, and inflammasome activation in UV-radiated NHDF cells. The anti-photoaging effects observed in NHDF cells were subsequently evaluated and validated in UV + MED-treated nude mice through skin phenotypes and histopathological structure analyses. Taken together, these results indicate that MED exerts therapeutic effects against UV-induced photoaging and has the potential for future development as a treatment for photoaging.

Comparison of cisplatin-induced anti-tumor response in CT26 syngeneic tumors of three BALB/c substrains.

BACKGROUND: To determine whether the background of BALB/c substrains affects the response to anti-tumor drugs, we measured for alterations in tumor growth, histopathological structure of the tumor, and expressions of tumor-related proteins in three BALB/c substrains derived from different sources (BALB/cKorl, BALB/cA and BALB/cB), after exposure to varying concentrations of cisplatin (0.1, 1 and 5 mg/kg). RESULTS: Cisplatin treatment induced similar responses for body and organ weights, serum analyzing factors, and blood analyzing factors in all BALB/c substrains with CT26 syngeneic tumor. Few differences were detected in the volume and histopathological structure of the CT26 tumor. Growth inhibition of CT26 tumors after exposure to cisplatin was greater in the BALB/cB substrain than BALB/cKorl and BALB/cA substrains, and a similar pattern was observed in the histopathological structure of tumors. However, the expression levels of other tumor-related factors, including Ki67, p27, p53, Bcl-2-associated X protein (Bax), B-cell lymphoma 2 (Bcl-2), caspase-3 (Cas-3), matrix metallopeptidase 2 (MMP2) and vascular endothelial growth factor (VEGF) proteins, were constantly maintained in the tumors of all three substrains after cisplatin treatment. A similar decrease pattern was observed for the expressions of inflammatory cytokines, including interleukin (IL)-1ß, IL-6 and IL-10, in the CT26 tumors of the three BALB/c substrains. CONCLUSIONS: Taken together, results of the present study indicate that the genetic background of the three BALB/c substrains has no major effect on the therapeutic responsiveness of cisplatin, except growth and histopathology of the CT26 syngeneic tumor.

Anti-Obesity Effect of α-Cubebenol Isolated from Schisandra chinensis in 3T3-L1 Adipocytes.

The efficacy of α-cubebenol isolated from Schisandra chinensis has been studied in several diseases, including cecal ligation, puncture challenge-induced sepsis, and degranulation of neutrophils. To identify the novel functions of α-cubebenol on lipid metabolism, alterations on the regulation of lipogenesis, lipolysis, and inflammatory response were observed in 3T3-L1 adipocytes treated with α-cubebenol. Most lipogenic targets, including lipid accumulation, level of lipogenic transcription factors, and expression of lipogenic regulators, were suppressed in MDI (3-isobutyl-1-methylxanthine, dexamethasone, and insulin)-stimulated 3T3-L1 adipocytes treated with α-cubebenol without significant cytotoxicity. In addition, similar inhibition effects were observed in the iNOS-induced COX-2 mediated pathway and NLRP3 inflammasome pathway of MDI-stimulated 3T3-L1 cells treated with α-cubebenol. Lipolytic targets, such as cAMP concentration, expression of adenylyl cyclase and PDE4, and their downstream signaling pathway, in MDI-stimulated 3T3-L1 cells were stimulated by the α-cubebenol treatment. The levels of transcription factors and related proteins for ß-oxidation were significantly higher in the MDI + α-cubebenol treated group than in the MDI + Vehicle treated group. These results show that α-cubebenol has a novel role as a lipogenesis inhibitor, lipolysis and ß-oxidation stimulator, and inflammasome suppressor in MDI-stimulated 3T3-L1 adipocytes.

Inflammatory response in the mid colon of ICR mice treated with polystyrene microplastics for two weeks.

BACKGROUND: The oral administration of polystyrene-microplastics (PS-MPs) causes chronic constipation of ICR mice, but there are no reports on their effects on the inflammatory response in the colon. To determine if the oral administration of MPs causes inflammation in the colon, the changes in the apoptosis-associated speck like protein containing a caspase recruitment domain (ASC)-inflammasome pathway, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway, and inflammatory cytokine expression were evaluated in the mid colon of ICR mice treated with 0.5 µm size PS-MPs for two weeks. RESULTS: The thicknesses of the mucosa, muscle, flat luminal surface, and crypt layer were decreased significantly (p < 0.01) in the mid colon of the MPs treated group compared to the Vehicle treated group. On the other hand, a remarkable increase in the expression levels of NOD-like receptor pyrin domain-containing protein (NLRP) 3, ASC, and Cleaved Caspase (Cas)-1 protein was observed in the MPs treated group. In addition, similar increasing pattern in the levels of p-NF-κB and phospho-inhibitory subunit of NF-κB (p-IkB) α protein was detected. Four inflammatory cytokines, including NF-κB, interleukin (IL)-6, tumor necrosis factor (TNF)-α, and IL-1ß, showed an increased expression level after the MPs treatment. CONCLUSIONS: Therefore, the present study suggests that PS-MPs can be a novel cause of an inflammatory response in the mid colon of ICR mice.

Antioxidant activity and laxative effects of tannin-enriched extract of Ecklonia cava in loperamide-induced constipation of SD rats.

To investigate the role of tannin-enriched extracts of Ecklonia cava (TEE) on the regulation of oxidative balance and laxative activity in chronic constipation, we investigated alterations after exposure to TEE, on constipation phenotypes, muscarinic cholinergic regulation, and oxidative stress responses in the transverse colons of SD rats with loperamide (Lop)-induced constipation. This extract contains high levels of total condensed tannin content (326.5 mg/g), and exhibited high inhibitory activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) radicals. TEE treatment induced significant improvements in reactive oxygen species (ROS) production, superoxide dismutase (SOD) expression and nuclear factor erythroid 2-related factor 2 (Nrf2) phosphorylation in primary smooth muscles of rat intestine cells (pRISMCs) and transverse colon of constipation model. Also, Lop+TEE treated groups showed alleviated outcomes for the following: most stool parameters, gastrointestinal transit, and intestine length were remarkably recovered; a similar recovery pattern was observed in the histopathological structure, mucin secretion, water channel expression and gastrointestinal hormones secretion in the transverse colon; expressions of muscarinic acetylcholine receptors M2/M3 (mAChR M2/M3) and their mediators on muscarinic cholinergic regulation were significantly recovered. Taken together, these results provide the first evidence that TEE stimulates oxidative stress modulation and muscarinic cholinergic regulation when exerting its laxative effects in chronic constipation models.