Amber (Succinite) Extract Enhances Glucose Uptake through the Up-Regulation of ATP and Down-Regulation of ROS in Mouse C2C12 Cells.

Traditionally, amber (Succinite) has been used to alleviate all types of pain, skin allergies, and headaches. However, no studies have been conducted on its antidiabetic and antioxidant effects. In this study, differentiated skeletal muscle C2C12 cells were used to demonstrate the protective effects of amber (AMB) against H2O2-induced cell death. In addition, the effects of AMB on glucose uptake and ATP production were investigated. Our results showed that AMB at 10, 25, and 50 µg/mL suppressed the elevation of ROS production induced by H2O2 in a dose-dependent manner. Moreover, AMB enhanced glucose utilization in C2C12 cells through the improvement of ATP production and an increase in PGC-1α gene expression resulting in an amelioration of mitochondrial activity. On the other hand, AMB significantly increased the gene expression of glucose transporters GLUT4 and GLUT1. Our finding suggests that AMB can be used as a natural supplement for diabetes treatment and for the promotion of skeletal muscle function.

Oxygen nanobubble water affects wound healing of fibroblast WI-38 cells.

This study investigated the usefulness of oxygen nanobubble water (O2NBW) for wound repair by analyzing its effect on the wound-healing process in human lung fibroblasts (WI-38 cells). The WI-38 cells were treated with 0%, 50%, and 100% O2NBW. The cell viability, reactive oxygen species (ROS) production, and wound healing following treatment were determined to elucidate the effects of O2NBW. Our findings revealed that O2NBW had no cytotoxic effects on WI-38 cells, but instead increased cell numbers. The production of ROS was inhibited in the presence of O2NBW. Further, O2NBW induced migration and wound closure in WI-38 cells. In addition, the mRNA expression levels of antioxidant enzymes and wound-healing-related genes were evaluated. The results demonstrated that O2NBW enhanced the expression levels of all representative genes. In conclusion, our findings suggest that O2NBW could affect ROS production and wound healing in WI-38 cells and genes associated with the antioxidant system and wound healing.

Ethyl pyruvate protects SHSY5Y cells against 6-hydroxydopamine-induced neurotoxicity by upregulating autophagy.

Parkinson disease is a chronic progressive neurodegenerative disorder with a prevalence that increases with age. The glycolytic end-product pyruvate, has antioxidant and neuroprotective feature. Here, we investigated the effects of ethyl pyruvate (EP), a pyruvic acid derivative, on 6-hydroxydopamine-induced SH-SY5Y cell apoptosis. Ethyl pyruvate decreased protein levels of cleaved caspase-3, phosphorylated endoplasmic reticulum kinase (pERK), and extracellular signal-regulated kinase (ERK), suggesting that EP reduces apoptosis via the ERK signaling pathway. Ethyl pyruvate also decreased oxygen species (ROS) and neuromelanin contents, suggesting that it suppresses ROS-mediated neuromelanin synthesis. Furthermore, increased protein levels of Beclin-1 and LC-II, and LC-II:LC-I ratios indicated that EP upregulates autophagy.

Low energy multiple blue light-emitting diode light Irradiation promotes melanin synthesis and induces DNA damage in B16F10 melanoma cells.

Visible light is present everywhere in our lives. Widespread use of computers and smartphones has increased the daily time spent in front of screens. What effect does this visible light have on us? Recent studies have shown that short-wavelength blue light (400-450nm) irradiation, similar to UV, inhibits the cell proliferation and differentiation, induces the intracellular oxidative stress, promotes the cell apoptosis and causes some other negative effects. However, it's unusual that directly face to such short-wavelength and high-energy blue light in daily life. Therefore, the effects of blue light with longer wavelength (470nm), lower energy (1, 2 J/cm2) and multiple times (simulated daily use) exposure on cells have been studied in this experiment. In our results, low energy density multiple blue light inhibited cell proliferation and metastatic capability with a weak phototoxicity. Blue light also promoted intracellular reactive oxygen species and caused DNA damage. Furthermore, the melanin synthesis was also promoted by low energy density multiple blue light exposure. Together, these results indicate that longer wavelength and low energy density blue light multiple exposure is still harmful to our cells. Furthermore, prolonged exposure to screens likely induces dull skin through induction of melanin synthesis. These results further mentioned us should paid more attention to controlling the daily use of digital device.

Stress Buffering and Longevity Effects of Amber Extract on Caenorhabditis elegans (C. elegans).

Amber is a fossilized tree resin historically used in wound healing and stress relief. Unfortunately, there is no concrete scientific evidence supporting such efficacy. Here, the stress buffering and longevity effect of Amber extract (AE) in Caenorhabditis elegans (C. elegans) was investigated. Survival assays, health span assays, Enzyme-Linked Immunosorbent Assay (ELISA), Stress biomarker detection assays, Green Fluorescence Proteins (GFP), Real Time quantitative PCR (RT-qPCR) and C. elegans mutants were employed to investigate the stress buffering and longevity effect of AE. In the study, it was observed that AE supplementation improved health span and survival in both normal and stressed worms. Additionally, AE positively regulated stress hormones (cortisol, oxytocin, and dopamine) and decreased fat and reactive oxygen species (ROS) accumulation. Through the Insulin/IGF-1 signaling (IIS) pathway, AE enhanced the nuclear localization of DAF-16 and the expression of heat shock proteins and antioxidant genes in GFP-tagged worms and at messenger RNA levels. Finally, AE failed to increase the survival of daf-16, daf-2, skn-1 and hsf-1 loss-of-function mutants, confirming the involvement of the IIS pathway. Evidently, AE supplementation relieves stress and enhances longevity. Thus, amber may be a potent nutraceutical for stress relief.

Protective Effect of Amber Extract on Human Dopaminergic Cells against 6-Hydroxydopamine-Induced Neurotoxicity.

Parkinson's disease (PD) is the second most common progressive neurodegenerative disease, after Alzheimer's disease. In our previous study, we found that amber-a fossilized plant resin-can protect cells from apoptosis by decreasing the generation of reactive oxygen species (ROS). In this study, we focused on the effect of amber on 6-hydroxydopamine-induced cell apoptosis in the human neuroblastoma cell line SHSY5Y (one model for PD). Initially, we determined the protective effect of amber on the PD model. We found that amber extract has a protective effect against 6-hydroxydopamine-induced cell apoptosis. The decrease in ROS, cleaved caspase-3, pERK, and extracellular signal-regulated kinase (ERK) protein levels confirmed that amber extract decreases apoptosis via the ROS-mediated ERK signaling pathway. Furthermore, we determined the effects of amber extract on autophagy. The results showed that amber extract increased the levels of LC3II and Beclin-1, suggesting that amber extract can protect neuronal cells against 6-hydroxydopamine-induced cell apoptosis by promoting autophagy.

Grape Extract Promoted α-MSH-Induced Melanogenesis in B16F10 Melanoma Cells, Which Was Inverse to Resveratrol.

Melanin is a natural pigment produced by cells to prevent damage caused by ultraviolet radiation. Previously, resveratrol was shown to reduce melanin synthesis. As a natural polyphenol with various biological activities, resveratrol occurs in a variety of beverages and plant foods, such as grapes. Therefore, we investigated whether grape extracts containing resveratrol also had the ability to regulate melanin synthesis. In this study, we used mouse B16F10 melanoma cells as a model for melanin synthesis with the melanogenesis-inducing α-melanocyte-stimulating hormone (α-MSH) as a positive control. Our results confirmed previous reports that resveratrol reduces melanin synthesis by reducing the activity of the rate-limiting enzyme tyrosinase. In contrast, the grape extract could not reduce melanin synthesis, and in fact promoted melanogenesis in the presence of α-MSH. The expression of genes related to melanin synthesis, such as tyrosinase, tyrosinase-related protein-1, tyrosinase-related protein-2, and microphthalmia-associated transcription factor, also supports these phenomena, which means that even in the presence of resveratrol, grape extract will strengthen the function of α-MSH in promoting melanin synthesis. Therefore, these results also provide a point of view for research on cosmetics.

Amber Extract Reduces Lipid Content in Mature 3T3-L1 Adipocytes by Activating the Lipolysis Pathway.

Amber-the fossilized resin of trees-is rich in terpenoids and rosin acids. The physiological effects, such as antipyretic, sedative, and anti-inflammatory, were used in traditional medicine. This study aims to clarify the physiological effects of amber extract on lipid metabolism in mouse 3T3-L1 cells. Mature adipocytes are used to evaluate the effect of amber extract on lipolysis by measuring the triglyceride content, glucose uptake, glycerol release, and lipolysis-related gene expression. Our results show that the amount of triacylglycerol, which is stored in lipid droplets in mature adipocytes, decreases following 96 h of treatment with different concentrations of amber extract. Amber extract treatment also decreases glucose uptake and increases the release of glycerol from the cells. Moreover, amber extract increases the expression of lipolysis-related genes encoding perilipin and hormone-sensitive lipase (HSL) and promotes the activity of HSL (by increasing HSL phosphorylation). Amber extract treatment also regulates the expression of other adipocytokines in mature adipocytes, such as adiponectin and leptin. Overall, our results indicate that amber extract increases the expression of lipolysis-related genes to induce lipolysis in 3T3-L1 cells, highlighting its potential for treating various obesity-related diseases.

Niclosamide affects intracellular TDP-43 distribution in motor neurons, activates mitophagy, and attenuates morphological changes under stress.

Amyotrophic lateral sclerosis (ALS) is characterized by progressive motor neuron loss in the brain and spinal cord; however, its etiology is unknown, and no curative treatment exists. TAR DNA-binding protein 43 (TDP-43), encoded by TARDBP, is a genetic mutation observed in 2-5% of familial ALS, and TDP is known to be mislocalized in the cytoplasm. This study aimed to identify compounds that inhibited the nuclear to cytoplasmic localization of TDP-43 in human induced pluripotent stem (iPS) cells-derived neurons. TDP-43 transgenic human iPS cells were constructed, differentiated into motor neurons, and then treated with MG-132 and sodium arsenite (stressors) to induce nuclear to cytoplasmic localization of TDP-43. STAT3 inhibitors, such as niclosamide, prevented TDP-43 mislocalization and degraded TDP-43 aggregates, and attenuated morphological changes under stress. Furthermore, niclosamide activated mitophagy via the PINK1-parkin-ubiquitin pathway. These findings suggest niclosamide may be a therapeutic candidate for ALS.

Kaempferol ameliorates symptoms of metabolic syndrome by improving blood lipid profile and glucose tolerance.

Kaempferol (KPF) is a dietary polyphenol reported to have various beneficial effects on human health. However, its molecular mechanisms in regulating lipid and glucose metabolism are not fully understood. This study examined the effects of KPF on obesity, dyslipidemia, and diabetes in Tsumura, Suzuki, Obese Diabetes mice. The 6-week administration of KPF decreased fat weight, serum total cholesterol, and low-density lipoproteins (LDLs); increased high-density lipoproteins (HDLs); and improved glucose tolerance. Additionally, KPF increased LDL receptor (LDLR) and apolipoprotein A1 (ApoA1) gene expression and decreased serum resistin levels. These findings suggest that the decrease in LDL and the increase in HDL caused by KPF may be due to increases in hepatic LDLR and ApoA1 expression, respectively. Furthermore, it is possible that the improvement in glucose tolerance by KPF may occur via resistin reduction. These mechanisms may be parts of complex mechanism by which KPF improves metabolic syndrome.

Anti-inflammatory activities of amber extract in lipopolysaccharide-induced RAW 264.7 macrophages.

Amber is a type of fossil tree resin with several bioactive properties and has been traced in traditional medicines used in Russia and China. However, its anti-inflammatory activities are poorly characterized. Here, the anti-inflammatory effects of the extract of amber mined from Kaliningrad, Russia was investigated in lipopolysaccharide (LPS)-induced RAW 264.7 cells. The effect of the amber extract on cell viability was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. Further, its effects on the production of intracellular reactive oxygen species (ROS), NO, and inflammatory cytokines were assessed by 2',7'-dichlorodihydrofluorescein diacetate staining, Griess test, and cytokine enzyme-linked immunosorbent assays, respectively. Western blotting and real-time reverse transcription-polymerase chain reaction analysis were performed to assess the mRNA and protein expression levels of the inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α) interleukin-6 (IL-6), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS). The translocation of the nuclear factor-kappa B (NF-κB) p65 subunit was observed by immunofluorescent staining. Amber extract negatively regulated the LPS-induced differentiation of RAW 264.7 cells to dendritic-like cells and reduced the LPS-induced increase in ROS and NO levels. It also reduced the level of mRNA and protein expressions of TNF-α, IL-6, COX-2, and iNOS in LPS-induced RAW 264.7 macrophages, in a dose-dependent manner. Furthermore, amber extract suppressed the nuclear translocation of the NF-κB p65 subunit. These findings suggest that the potent anti-inflammatory effect of the amber extract is mediated by the inhibition of the NF-κB p65 signaling pathway. Collectively, this study renders amber extract as a potential pharmacological alternative to treat inflammation-related diseases.

Role of amber extract in protecting SHSY5Y cells against amyloid ß1-42-induced neurotoxicity.

Alzheimer disease (AD) is an irreversible, progressive brain disease. Amyloid ß plays a critical role in AD development. Some Chinese traditional medicines, such as the fossilized plant resin, amber, have been applied as mental stabilizers. However, the effects of amber on AD pathogenesis remain unknown. Therefore, we aimed to determine the potential of amber extract for treating AD by evaluating its effects on amyloid-ß (1-42) (Aß (1-42))-induced neuronal cell death. We measured levels of ROS, Bcl-2, and Bax mRNA, and found that amber extract decreased Aß (1-42)-induced cell apoptosis via the reactive oxygen species (ROS)-mediated mitochondrial pathway. Amber extract also decreased ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) and increased microtubule-associated proteins 1A/1B light chain 3B (LC3II) and Beclin 1. These findings suggested that amber extract protects neuronal cells against Aß (1-42)-induced cell apoptosis by upregulating autophagy and downregulating BACE1.

α-Mangostin suppressed melanogenesis in B16F10 murine melanoma cells through GSK3ß and ERK signaling pathway.

Mangosteen (Garcinia mangostana L) fruit contains many xanthones in its pericarp, such as α-mangostin. Here, we aimed to elucidate the physiological effect of α-mangostin and the mechanism on melanogenesis in mouse B16F10 cells. The melanin production in B16F10 cells was decreased by α-mangostin treatment. α-Mangostin also suppressed the enzymatic activity of tyrosinase, the critical enzyme for melanin synthesis. Furthermore, Western blot analysis revealed that α-mangostin down-regulated the protein quantity of tyrosinase, tyrosinase relative protein (TRP)-2, and microphthalmia-associated transcription factor (MITF). We also used inhibitors of the extracellular signal-regulated kinase (ERK), and glycogen synthase kinase 3 (GSK-3ß) to identify the upstream signaling cascade of MITF. Results showed us GSK3ß plays a more important role in α-mangostin regulated melanogenesis. Further, the de-pigmentation effect on normal human epidermal melanocytes (NHEMs) of α-mangostin was also confirmed. These results suggested that α-mangostin is a reagent for depigmentation and it has the potential to be applied as a component of cosmetics or pharmaceuticals for the therapy of spots, chloasma, or melanosis.

Cortisol promotes stress tolerance via DAF-16 in Caenorhabditis elegans.

In this study, we studied the effects of cortisol and cortisone on the age-related decrease in locomotion in the nematode Caenorhabditis elegans and on the tolerance to heat stress at 35 °C and to oxidative stress induced by the exposure to 0.1% H2O2. Changes in mRNA expression levels of C. elegans genes related to stress tolerance were also analyzed. Cortisol treatment restored nematode movement following heat stress and increased viability under oxidative stress, but also shortened worm lifespan. Cortisone, a cortisol precursor, also restored movement after heat stress. Additionally, cortisol treatment increased mRNA expression of the hsp-12.6 and sod-3 genes. Furthermore, cortisol treatment failed to restore movement of daf-16-deficient mutants after heat stress, whereas cortisone failed to restore the movement of dhs-30-deficient mutants after heat stress. In conclusion, the results suggested that cortisol promoted stress tolerance via DAF-16 but shortened the lifespan, whereas cortisone promoted stress tolerance via DHS-30.

Citric acid promoted melanin synthesis in B16F10 mouse melanoma cells, but inhibited it in human epidermal melanocytes and HMV-II melanoma cells via the GSK3ß/ß-catenin signaling pathway.

Melanin, a pigment synthesized by melanocytes in the skin, resists the damage caused by ultraviolet rays to cells. Citric acid, a well-known food additive, is commonly used as an antioxidant and is an important part of the tricarboxylic acid (TCA) cycle for energy production during cellular metabolism. Here, we aimed to investigate whether the addition of excess citric acid regulates melanin synthesis, and to delineate the underlying mechanism. First, we observed that citric acid exerts opposite redox effects on mouse and human cells. Interestingly, treatment with excess citric acid increased the melanin content in mouse cells but decreased it in human cells. Furthermore, the expression of factors important for melanin synthesis, such as microphthalmia-associated transcription factor (MITF), was also regulated by citric acid treatment-it was promoted in mouse cells and suppressed in human cells. Citric acid also impacted the upstream regulators of MITF, glycogen synthase kinase 3ß (GSK3ß), and ß-catenin. Second, we determined the importance of GSK3ß in the citric acid-mediated regulation of melanin synthesis, using a GSK3ß inhibitor (BIO). To the best of our knowledge, this is the first study to show that citric acid regulates melanin synthesis via the GSK3ß/ß-catenin signaling pathway, and that equal amounts of exogenous citric acid exert opposing effects on mouse and human cells.

Quercetin enhances motility in aged and heat-stressed Caenorhabditis elegans nematodes by modulating both HSF-1 activity, and insulin-like and p38-MAPK signalling.

Quercetin is a yellow pigment that is found in many common dietary plants, and that protects against oxidative stress, inflammation, and arteriosclerosis. It has also been suggested to prolong the lifespan of, and enhance heat-stress tolerance in nematodes; thus, the present study investigated its effects on both the nematode life- and health span by assessing its capacity to promote nematode motility after aging and/or heat stress, as well as the mechanisms underlying these effects. The results of the conducted analyses showed that quercetin feeding prolonged lifespan, suppressed age-related motility retardation, improved motility recovery after heat stress, and decreased the production of both intercellular and mitochondrial reactive oxygen species in the analysed Caenorhabditis elegans strains, likely by modulating the insulin-like signalling (ILS) pathway and p38-mitogen-activated protein kinase (MAPK) pathway. In particular, the transcription factors DAF-16 and SKN-1 were found to mediate the observed quercetin-induced effects, consistent with their previously demonstrated roles as regulators of aging. Furthermore, we demonstrated, for the first time, that quercetin induced heat-stress tolerance in C. elegans by modulating HSF-1 expression and/or activity. Thus, the present study provides valuable insights into the mechanisms by which quercetin inhibit aging and enhance heat-stress tolerance via ILS and MAPK pathway in C. elegans.

α-Pinene odor exposure enhances heat stress tolerance through Daf-16 in Caenorhabditis elegans.

Aromatherapy has been widely used as complementary and alternative medicine to reduce pain and induce sleep. However, the scientific evidence regarding the biological effects of odor is scarce and the underlying molecular mechanisms have not been clarified. We treated worms with contactless S-(-)- and R-(+)-α-pinene and analyzed heat stress tolerance. Odor stimulation induced motility recovery after incubation at 35 °C for 4 h. This increase in heat stress tolerance was not present in odr-3 mutants and daf-16 mutants. S-(-)- and R-(+)-α-pinene expanded health span and increased fat accumulation. Moreover, S-(-)- and R-(+)-α-pinene modulated the expression of 84 and 54 genes, respectively. These results show that α-pinene odor stimulation is related to stress tolerance, lipid metabolism, and health span via some specific signaling pathways. This study may provide a potential target for antiaging and disease prevention.

Alpha Mangostin promotes myogenic differentiation of C2C12 mouse myoblast cells.

Mangosteen, a fruit mainly produced in Southeast Asia, has been used as food and as an antipyretic and for treating skin diseases. The xanthones contained in mangosteen have many physiological activities including melanin suppression and anticancer activities, but little is known about the physiological effects of the most abundant xanthone, α-mangostin (α-MG) on myoblasts. In this study, we applied α-MG to C2C12 cells that had been induced to differentiate using 2% HS, and analyzed the physiological action of α-MS and the underlying mechanism in the context of myogenic differentiation. α-MG increased the survival rate of C2C12 cells in a concentration-dependent manner. Analysis of the morphological changes in the cells showed that α-MG significantly enhanced the myogenic differentiation of C2C12 myoblasts, whereas the mitochondrial number was only slightly affected. Expression analysis of differentiation-related proteins in C2C12 cells revealed that α-MG promoted the expression of MyoD and Myogenin. Thus, the present study revealed that α-MG improves the survival and myogenic differentiation of C2C12 myoblasts.

Effect of inactivated Bifidobacterium longum intake on obese diabetes model mice (TSOD).

Obesity and diabetes have been increasing at an alarming rate worldwide. Studies have shown the futility of chemical drugs in the treatment of obesity and diabetes. Bifidobacterium longum (BL), a common member of the gut microbiota throughout the human lifespan, has been widely reported to play a role in host health and disease. Here, we evaluated the effects of inactivated cells of BL (IBL) on obesity and blood glucose levels in TSOD mice by administering IBL orally for 5 weeks. The treated mice showed a significant decrease of body weight gain, adipose tissue mass and blood glucose levels, as well as a significant reduction in blood glucose during an oral glucose tolerance test. The treatment also resulted in reduced levels of cholesterol, triglycerides, and NEFA. Moreover, serum and urine analysis showed low creatinine levels in IBL-treated mice. These data demonstrate that IBL may have the potential to prevent obesity and diabetes.

Oxytocin promotes heat stress tolerance via insulin signals in Caenorhabditis elegans.

Oxytocin, has various physiological functions that have been well studied and many that remain unknown. Here, we aimed to determine new physiological functions of oxytocin using Caenorhabditis elegans. Oxytocin treatment promoted the restoration of movement after heat stress and enhanced the viability under heat stress. However, oxytocin had no effect on the life span and only little effect on the oxidative stress tolerance. In contrast, oxytocin treatment didn't promote the restoration of movement or enhance the viability of deficient mutants of ntr-1/2, which is the gene encoding the oxytocin receptor. In addition, for mutants of daf-16, daf-2, tax-4, and some insulin-like peptides, the heat stress tolerance effect by oxytocin was canceled. Furthermore, oxytocin increased the expression levels of the DAF-16 target genes. Our results suggest that oxytocin treatment promoted the heat stress tolerance of C. elegans via the insulin/IGF-1 signaling pathway.