Sunset Yellow protects against oxidative damage and exhibits chemoprevention in chemically induced skin cancer model.

Skin cancer and other skin-related inflammatory pathologies are rising due to heightened exposure to environmental pollutants and carcinogens. In this context, natural products and repurposed compounds hold promise as novel therapeutic and preventive agents. Strengthening the skin's antioxidant defense mechanisms is pivotal in neutralizing reactive oxygen species (ROS) and mitigating oxidative stress. Sunset Yellow (SY) exhibits immunomodulatory characteristics, evidenced by its capacity to partially inhibit the secretion of proinflammatory cytokines, regulate immune cell populations, and modulate the activation of lymphocytes. This study aimed to investigate the antioxidant and anti-genotoxic properties of SY using in-silico, in vitro, and physiochemical test systems, and to further explore its potential role in 7,12-dimethylbenz(a) anthracene (DMBA)/ 12-o-tetradecanoylphorbol-13-acetate (TPA)-induced two-stage skin carcinogenesis. In vitro experiments showed that pre-treatment of SY significantly enhanced the cell viability of HaCaT cells when exposed to tertiary-Butyl Hydrogen Peroxide (tBHP). This increase was accompanied by reduced ROS levels, restoration of mitochondrial membrane potential, and notable reduction in DNA damage in (SY + tBHP) treated cells. Mechanistic investigations using DPPH chemical antioxidant activity test and potentiometric titrations confirmed SY's antioxidant properties, with a standard reduction potential ( E o ) of 0.211 V. Remarkably, evaluating the effect of topical application of SY in DMBA/TPA-induced two-step skin carcinogenesis model revealed dose-dependent decreases in tumor latency, incidence, yield, and burden over 21-weeks. Furthermore, computational analysis and experimental validations identified GSK3ß, KEAP1 and EGFR as putative molecular targets of SY. Collectively, our findings reveal that SY enhances cellular antioxidant defenses, exhibits anti-genotoxic effects, and functions as a promising chemopreventive agent.

Alkaloids Extract from Linum usitatissimum Attenuates 12-OTetradecanoylphorbol- 13-Acetate (TPA)-induced Inflammation and Oxidative Stress in Mouse Skin.

BACKGROUND: In traditional medicine, Linum usitatissimum treats inflammatory, gastrointestinal, and cardiovascular diseases. OBJECTIVES: The present study aims to assess the anti-inflammatory and anti-oxidant effects of total alkaloid extract from Linum usitatissimum seeds (ALU) on the ear histological integrity and oxidant- antioxidant status in a mice model of a sub-chronic inflammation induced by multiapplication of TPA. METHODS: Topical TPA treatment induced various inflammatory changes, including edema formation, epidermal thickness, and the excess production of reactive oxygen species. Tissue samples were used for the measurement of reduced glutathione (GSH) and nitric oxide (NO) levels and Myeloperoxidase (MPO) and Catalase (CAT) activities. RESULTS: Oral administration of ALU (50, 100, and 200 mg/kg) produced anti-inflammatory and anti-oxidant effects. Also, ALU significantly reduced ear edema and inflammatory cell infiltration and restored the integrity of the ear. CONCLUSION: These findings suggest that the total alkaloid extract from Linum usitatissimum seeds presents significant anti-inflammatory and anti-oxidant effects on TPA-induced sub-chronic inflammation model in NMRI mice and can be used as an anti-inflammatory and anti-oxidant agent for the therapeutic management of inflammatory disorders.

Isolation of cancer stem cells from skin squamous cell carcinoma.

Skin squamous cell carcinoma (skin SCC) is the most frequently occurring cancer. Skin is the first line of defense that provides protection from the external environment. Skin consists of epidermis, dermis, and hypodermis. The epidermis comprises of inter-follicular epidermis, hair follicles, sebaceous glands, and sweat glands. Stem cells within these epidermal compartments play crucial role in epidermal regeneration and repair. Various factors such as higher exposure to ultraviolet light (UV) of sun, genetic predisposition, exposure to carcinogens, etc. that give rise to skin cancer. Within the skin SCC, there exists a pool of cancer stem cells (CSCs) that are highly quiescent with self-renewal capacity. Further, isolation and molecular characterization of CSCs would enable to unravel mechanism involved in tumor progression, metastasis, relapse, and resistance to chemotherapeutic agents. To understand the sequential events of carcinogenesis, the two-stage skin carcinogenesis murine model is proposed, which employs the topical application of a chemical carcinogen, DMBA that causes several activating mutations occurring in the genes responsible for cell proliferation and growth. Further, initiation is followed by tumor promotion, which is induced by repeated application of tumor-promoting agent, TPA, which fixes the activating mutations resulting in the formation of a benign papilloma. Subsequently, papilloma further progresses to highly malignant SCC. Here, using the two-stage skin carcinogenesis murine model, we provide a detailed protocol for the isolation of CSCs from murine skin SCC. FACS sorting of CSCs is followed by assays such as invitro-spheroid assay, in vivo-tumorigenesis-limiting dilution and in vivo-tumorigenesis-serial transplantation assay and expression profiling.

PINCH-1 promotes IGF-1 receptor expression and skin cancer progression through inhibition of the GRB10-NEDD4 complex.

Background: Insulin-like growth factor 1 receptor (IGF-1R) expression and signaling play important roles in promotion of skin cancer progression. Identification of signaling pathways that regulate IGF-1R is crucial for understanding the pathogenesis and therapeutic treatment of skin cancer. Methods: Molecular, cellular and genetic approaches were used to investigate the function of PINCH-1 in regulation of IGF-1R expression and skin cell behavior. Furthermore, conditional PINCH-1 knockout mouse and carcinogen (7, 12-dimethylbenz[a]anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA))-induced skin cancer model were employed to determine the function of PINCH-1 in regulation of IGF-1R expression and skin carcinogenesis in vivo. Results: Knockdown of PINCH-1 from HaCaT keratinocytes or A431 squamous carcinoma cells diminished IGF-1R levels, suppressed cell proliferation and increased apoptosis. Re-expression of PINCH-1 in PINCH-1 knockdown cells restored IGF-1R expression, cell proliferation and survival. Furthermore, depletion of NEDD4 effectively reversed PINCH-1 deficiency-induced down-regulation of IGF-1R expression, cell proliferation and survival. Conditional knockout of PINCH-1 from keratin 5 (K5) positive keratinocytes in mice, like depletion of PINCH-1 from keratinocytes in culture, reduced the IGF-1R level. Using a mouse model of DMBA/TPA-induced skin cancer, we show that the levels of both PINCH-1 and IGF-1R were significantly increased in response to treatment with the carcinogens. Genetic ablation of PINCH-1 from the epidermis markedly reduced the IGF-1R expression and cell proliferation despite stimulation with DMBA/TPA, resulting in resistance to chemical carcinogen-induced skin cancer initiation and progression. Conclusions: Our results reveal a PINCH-1-NEDD4-IGF-1R signaling axis that is critical for promotion of skin tumorigenesis and suggest a new strategy for therapeutic control of skin cancer progression.

In Vitro and Clinical Evaluation of Cannabigerol (CBG) Produced via Yeast Biosynthesis: A Cannabinoid with a Broad Range of Anti-Inflammatory and Skin Health-Boosting Properties.

Cannabigerol (CBG) is a minor non-psychoactive cannabinoid present in Cannabis sativa L. (C. sativa) at low levels (<1% per dry weight) that serves as the direct precursor to both cannabidiol (CBD) and tetrahydrocannabinol (THC). Consequently, efforts to extract and purify CBG from C. sativa is both challenging and expensive. However, utilizing a novel yeast fermentation technology platform, minor cannabinoids such as CBG can be produced in a more sustainable, cost-effective, and timely process as compared to plant-based production. While CBD has been studied extensively, demonstrating several beneficial skin properties, there are a paucity of studies characterizing the activity of CBG in human skin. Therefore, our aim was to characterize and compare the in vitro activity profile of non-psychoactive CBG and CBD in skin and be the first group to test CBG clinically on human skin. Gene microarray analysis conducted using 3D human skin equivalents demonstrates that CBG regulates more genes than CBD, including several key skin targets. Human dermal fibroblasts (HDFs) and normal human epidermal keratinocytes (NHEKs) were exposed in culture to pro-inflammatory inducers to trigger cytokine production and oxidative stress. Results demonstrate that CBG and CBD reduce reactive oxygen species levels in HDFs better than vitamin C. Moreover, CBG inhibits pro-inflammatory cytokine (Interleukin-1ß, -6, -8, tumor necrosis factor α) release from several inflammatory inducers, such as ultraviolet A (UVA), ultraviolet B (UVB), chemical, C. acnes, and in several instances does so more potently than CBD. A 20-subject vehicle-controlled clinical study was performed with 0.1% CBG serum and placebo applied topically for 2 weeks after sodium lauryl sulfate (SLS)-induced irritation. CBG serum showed statistically significant improvement above placebo for transepidermal water loss (TEWL) and reduction in the appearance of redness. Altogether, CBG's broad range of in vitro and clinical skin health-promoting activities demonstrates its strong potential as a safe, effective ingredient for topical use and suggests there are areas where it may be more effective than CBD.

Anti-inflammatory Effect of Cannabidiol and Palmitoylethanolamide Containing Topical Formulation on Skin in a 12- O -Tetradecanoylphorbol-13-Acetate-Induced Dermatitis Model in Mice.

BACKGROUND: Chronic inflammatory skin disorders, such as atopic dermatitis, have significant disease burden worldwide. Although efficacious, the adverse effect profile of topical corticosteroids limits long-term use. As an alternative, cannabinoids have been shown to have anti-inflammatory therapeutic effects. OBJECTIVE: The aim of this study was to assess the effects of a topical cannabinoid product using dermatitis mouse model. METHODS: Thirty-five mice were randomized into treatment groups. 12- O -tetradecanoylphorbol-13-acetate was used as an irritant on 1 ear with the contralateral ear serving as a control. Ear edema was calipered. The test product containing 0.9% cannabidiol and palmitoylethanolamide was compared with a potent topical corticosteroid. RESULTS: Treatment with topical cannabinoid formulation reduced ear edema by 51.27% at 24 hours' and 65.69% at 48 hours' postapplication. Alternatively, mometasone reduced ear edema by 89.82% at 24 hours and 98.25% at 48 hours. Natural reduction (control) in ear edema was 26.32% at 24 hours and 44.21% at 48 hours. Both test groups resulted in significantly decreased edema when compared with baseline ( P < 0.05), as well as compared with the negative control group ( P < 0.05). CONCLUSIONS: Significant reduction in ear edema, a marker for localized cutaneous inflammation, could be attributed to anti-inflammatory properties of cannabinoids. Although effects were less robust than topical corticosteroid use, cannabinoid formulations have therapeutic promise for dermatitis.

Phorbol 12-Myristate 13-Acetate Induced Toxicity Study and the Role of Tangeretin in Abrogating HIF-1α-NF-κB Crosstalk In Vitro and In Vivo.

Phorbol 12-myristate 13-acetate (PMA) is a potent tumor promoter and highly inflammatory in nature. Here, we investigated the toxic effects of PMA on different model system. PMA (10 µg) caused chromosomal aberrations on the Allium cepa root tip and induced mitotic dysfunction. Similarly, PMA caused embryonic and larval deformities and a plummeted survivability rate on zebrafish embryo in a dose-dependent manner. Persistently, PMA treatment on immortalized human keratinocyte human keratinocyte (HaCaT) cells caused massive inflammatory rush at 4 h and a drop in cell survivability at 24 h. Concomitantly, we replicated a cutaneous inflammation similar to human psoriasis induced by PMA. Herein, we used tangeretin (TAN), as an antagonist to counteract the inflammatory response. Results from an in vivo experiment indicated that TAN (10 and 30 mg/kg) significantly inhibited PMA stimulated epidermal hyperplasia and intra-epidermal neutrophilic abscesses. In addition, its treatment effectively neutralized PMA induced elevated reactive oxygen species (ROS) generation on in vitro and in vivo systems, promoting antioxidant response. The association of hypoxia-inducible factor 1-alpha (HIF-1α)-nuclear factor kappa-light-chain-enhancer of activated b cells (NF-κB) crosstalk triggered by PMA enhanced PKCα-ERK1/2-NF-κB pathway; its activation was also significantly counteracted after TAN treatment. Conclusively, we demonstrated TAN inhibited the nuclear translocation of HIF-1α and NF-κB p65. Collectively, TAN treatment ameliorated PMA incited malignant inflammatory response by remodeling the cutaneous microenvironment.

A Glycosaminoglycan-Rich Fraction from Sea Cucumber Isostichopus badionotus Has Potent Anti-Inflammatory Properties In Vitro and In Vivo.

Sea cucumber body wall contains several naturally occurring bioactive components that possess health-promoting properties. Isostichopus badionotus from Yucatan, Mexico is heavily fished, but little is known about its bioactive constituents. We previously established that I. badionotus meal had potent anti-inflammatory properties in vivo. We have now screened some of its constituents for anti-inflammatory activity in vitro. Glycosaminoglycan and soluble protein preparations reduced 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammatory responses in HaCaT cells while an ethanol extract had a limited effect. The primary glycosaminoglycan (fucosylated chondroitin sulfate; FCS) was purified and tested for anti-inflammatory activity in vivo. FCS modulated the expression of critical genes, including NF-ĸB, TNFα, iNOS, and COX-2, and attenuated inflammation and tissue damage caused by TPA in a mouse ear inflammation model. It also mitigated colonic colitis caused in mice by dextran sodium sulfate. FCS from I. badionotus of the Yucatan Peninsula thus had strong anti-inflammatory properties in vivo.

Anti-inflammatory effect of lanoconazole on 12-O-tetradecanoylphorbol-13-acetate- and 2,4,6-trinitrophenyl chloride-induced skin inflammation in mice.

BACKGROUND: Lanoconazole (LCZ) is a topical antifungal agent clinically used to treat fungal infections such as tinea pedis. LCZ has not only antifungal effects but also anti-inflammatory effects, which have the potential to provide additional clinical benefits. However, the characteristic features of the inhibitory effects of LCZ on skin inflammation remain unclear. OBJECTIVE: We evaluated the inhibitory effects of topical application of LCZ, and compared the effects of LCZ with those of other antifungal agents including liranaftate, terbinafine and amorolfine. METHODS: Each antifungal agent was topically applied on 12-O-tetradecanoylphorbol-13-acetate-induced irritant dermatitis and 2,4,6-trinitrophenyl chloride-induced contact dermatitis in mice (BALB/c). The ear thickness, myeloperoxidase activity and inflammatory mediator contents were evaluated. RESULTS: LCZ dose-dependently suppressed 12-O-tetradecanoylphorbol-13-acetate-induced irritant dermatitis, suppressed the production of neutrophil chemotactic factors such as keratinocyte-derived chemokine and macrophage inflammatory protein-2, and inhibited neutrophil infiltration to the inflammation site. Moreover, 1% LCZ reduced the ear swelling in mice with 2,4,6-trinitrophenyl chloride-induced contact dermatitis in accordance with the inhibition of interferon-γ production. The inhibitory potency of LCZ on these types of dermatitis in mice was stronger than that of other types of antifungal agents. CONCLUSION: The anti-inflammatory effects of LCZ were exerted through the inhibition of inflammatory mediator production. These effects may contribute to the relief of dermatitis symptoms in patients with tinea pedis.

Ubiquitylation and endocytosis of the human LAT1/SLC7A5 amino acid transporter.

The human L-type amino acid transporter 1 (LAT1), also known as SLC7A5, catalyzes the transport of large neutral amino acids across the plasma membrane. As the main transporter of several essential amino acids, notably leucine, LAT1 plays an important role in mTORC1 activation. Furthermore, it is overexpressed in various types of cancer cells, where it contributes importantly to sustained growth. Despite the importance of LAT1 in normal and tumor cells, little is known about the mechanisms that might control its activity, for example by promoting its downregulation via endocytosis. Here we report that in HeLa cells, activation of protein kinase C by phorbol 12-myristate 13-acetate (PMA) triggers efficient endocytosis and degradation of LAT1. Under these conditions we found LAT1 downregulation to correlate with increased LAT1 ubiquitylation. This modification was considerably reduced in cells depleted of the Nedd4-2 ubiquitin ligase. By systematically mutagenizing the residues of the LAT1 cytosolic tails, we identified a group of three close lysines (K19, K25, K30) in the N-terminal tail that are important for PMA-induced ubiquitylation and downregulation. Our study thus unravels a mechanism of induced endocytosis of LAT1 elicited by Nedd4-2-mediated ubiquitylation of the transporter's N-terminal tail.

Inhibition of skin tumor promotion by TPA using a combination of topically applied ursolic acid and curcumin.

Prevention remains an important strategy to reduce the burden of cancer. One approach to prevent cancer is the use of phytochemicals in various combinations as safe and effective cancer preventative agents. The purpose of this study was to examine the effects of the combination of ursolic acid (UA) and curcumin (Curc) for potential combinatorial inhibition of skin tumor promotion using the mouse two-stage skin carcinogenesis model. In short-term experiments, the combination of UA + Curc given topically prior to 12-O-tetradecanoylphorbol-13-acetate (TPA) significantly inhibited activation of epidermal EGFR, p70S6K, NF-κB p50, Src, c-Jun, Rb, and IκBα. Levels of c-Fos, c-Jun, and Cox-2 were also significantly reduced by the combination compared to the TPA treated group. The alterations in these signaling pathways by the combination of UA + Curc were associated with decreased epidermal proliferation as assessed by measuring BrdU incorporation. Significant effects were also seen with the combination on epidermal inflammatory gene expression and dermal inflammation, with the greatest effects on expression of IL-1ß, IL-6, IL-22, and CXCL2. Furthermore, results from skin tumor experiments demonstrated that the combination of UA + Curc given topically significantly inhibited mouse skin tumor promotion by TPA to a greater extent than the individual compounds given alone. The greatest effects were seen on tumor free survival, tumor size, and tumor weight, although tumor incidence and multiplicity were also further reduced by the combination. These results demonstrate the potential cancer chemopreventive activity and mechanism(s) for the combination of UA + Curc.

Three Constituents of Moringa oleifera Seeds Regulate Expression of Th17-Relevant Cytokines and Ameliorate TPA-Induced Psoriasis-Like Skin Lesions in Mice.

As a folk medicine, Moringa oleifera L. is used effectively to treat inflammatory conditions and skin diseases. However, its mechanism of action is not well understood, limiting its medical use. We isolated and identified three compounds, namely niazirin, marumoside A and sitosterol-3-O-ß-d-glucoside, from the seeds of Moringa oleifera, and studied their effects on the expression of Th17-relevant cytokines (IL-12/IL-23 p40, IL-17A, IL-22 and IL-23 p19) using lipopolysaccharide-stimulated THP-1 cells. Additionally, as Th17 plays a critical role in the pathogenesis of psoriasis, we used a 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced psoriasis-like skin lesion mouse model to study their potential therapeutic application in vivo. The compounds suppressed the expression of IL-12/IL-23 p40, IL-17A, IL-22 and IL-23 p19 in vitro, and in vivo they ameliorated psoriasis-like skin lesions, decreased IL-17A mRNA expression, and increased the expression of keratinocyte differentiation markers. To our knowledge, this is the first report regarding the mechanism and therapeutic application of Moringa oleifera seeds to treat psoriasis-like lesions in vivo.

Salidroside prevents skin carcinogenesis induced by DMBA/TPA in a mouse model through suppression of inflammation and promotion of apoptosis.

Salidroside (SR) is a main component of Rhodiola rosea L. and exhibits a variety of pharmacologic properties. The present study was carried out to explore the potential effect of SR against skin cancer induced by 7,12-dimethylbenz(a)anthracene (DMBA) and 12-O-tetradecanoylphorbol-13­acetate (TPA) in female Institute for Cancer Research (ICR) mice and to reveal the underlying molecular targets regulated by SR. The mice were randomly divided into 4 groups: control, DMBA/TPA, DMBA/TPA+SR (20 mg/kg) and DMBA/TPA+SR (40 mg/kg). SR was administered to mice five times a week after DMBA treatments. In our study, we found that SR dose-dependently ameliorated skin cancer incidence and the multiplicity in the animal models by reducing the release of inflammation-related cytokines, including tumor necrosis factor α (TNF-α), interleukin-1ß (IL-1ß), interleukin-18 (IL-18), interleukin-6 (IL-6), cyclooxygenase 2 (COX2) and transforming growth factor ß-1 (TGF-ß1). Suppression of the nuclear factor (NF)-κB signaling pathway by SR was effective to prevent skin carcinogenesis. Furthermore, TUNEL analysis indicated that compared to the DMBA/TPA group, enhanced apoptosis was observed in the DMBA/TPA+SR group. In addition, p53 expression levels were increased by SR in the DMBA/TPA-induced mice. Therefore, SR was effective for inducing apoptosis during skin cancer progression triggered by DMBA/TPA. Consistently, p21, p53 upregulated modulator of apoptosis (PUMA), Bax and caspase-3 were highly induced by SR to enhance the apoptotic response for preventing skin cancer. Moreover, in vitro, we found that SR dramatically reduced the inflammatory response, while enhancing the aoptotic response by blocking NF-κB and activating caspase-3 pathways, respectively. In addition, flow cytometric analysis further confirmed the induction of apoptosis by SR in DMBA-treated cells in vitro. Taken together, the in vivo and in vitro studies illustrated that SR might be a promising compound to reduce skin cancer risk.

Soy Phosphatidylglycerol Reduces Inflammation in a Contact Irritant Ear Edema Mouse Model In Vivo.

We have previously shown that phosphatidylglycerol (PG) regulates the function of keratinocytes, the predominant cells that compose the epidermis, inhibiting the proliferation of rapidly dividing keratinocytes. In particular, soy PG, a PG mixture with a high proportion of polyunsaturated fatty acids, is efficacious at inhibiting these proliferating keratinocytes. Psoriasis is a skin disorder characterized by hyperproliferation of keratinocytes and inflammation. Data in the lung suggest that PG in pulmonary surfactant inhibits inflammation. To investigate the possibility of using PG containing polyunsaturated fatty acids for the treatment of psoriasis, we examined the effect of soy PG on inflammation induced by the application of 12-O-tetradecanoylphorbol 13-acetate (TPA), a contact irritant, to mouse ears in vivo. We monitored ear thickness and weight as a measure of ear edema, as well as CD45-positive immune cell infiltration. Our results indicate that soy PG when applied together with 1,25-dihydroxyvitamin D3 (vitamin D), an agent known to acutely disrupt the skin barrier, suppressed ear edema and inhibited the infiltration of CD45-positive immune cells. On the other hand, neither PG nor vitamin D alone was effective. The combination also decreased tumor necrosis factor-α (TNFα) levels. This result suggested the possibility that PG was not permeating the skin barrier efficiently. Therefore, in a further study we applied PG in a penetration-enhancing vehicle and found that it inhibited inflammation induced by the phorbol ester and decreased CD45-positive immune cell infiltration. Our results suggest the possibility of using soy PG as a topical treatment option for psoriasis.

MAPKs and NF­κB pathway inhibitory effect of bisdemethoxycurcumin on phorbol­12­myristate­13­acetate and A23187­induced inflammation in human mast cells.

Inflammation­associated damage may occur in any tissue following infection, exposure to toxins, following ischemia, and in allergic and auto­immune reactions. Inflammation may also result from mast cell degranulation induced by the intracellular calcium concentration. The inflammatory process may be inhibited by compounds that affect mast cells. Bisdemethoxycurcumin [1,7­bis(4­hydroxyphenyl) hepta­1,6­diene­3,5­dione, BDCM] is the active component of turmeric. It has anticancer, antioxidant and antibacterial properties. To investigate the molecular mechanism associated with the anti­inflammatory activity of BDCM, human mast cell line 1 (HMC­1) cells were treated with phorbol­12­myristate­13­acetate (PMA) and calcium ionophore A23187 (A23187) to induce the inflammatory process. Various HMC­1 cells were pretreated with BDCM prior to stimulation of inflammation. BDCM inhibited the inflammation­triggered production of cytokines including interleukin (IL)­6, IL­8, and tumor necrosis factor (TNF)­α. BDCM inhibition extended to the gene level. In activated HMC­1 cells, phosphorylation levels of extracellular signal­regulated kinase, c­jun N­terminal kinase and p38 mitogen­activated protein kinase were decreased by treatment with BDCM. BDCM also inhibited nuclear factor­(NF)­κB activation and IκB degradation. In conclusion, BDCM suppresses the expression of TNF­α, IL­8, and IL­6 by inhibiting the NF­κB and mitogen activated protein kinase signaling pathways.

Bryostatin 1 causes attenuation of TPA­mediated tumor promotion in mouse skin.

The present study was designed to investigate the tumor inhibitory potential of bryostatin 1 in a 12­O­tetradecanoylphorbol­13­acetate (TPA)­induced mouse model of skin cancer. The radical inhibition potential of various doses of bryostatin 1 was investigated against 2,2­diphenyl­1­picrylhydrazyl (DPPH) bleach in vitro. The DPPH radical potential was observed compared with treatment with 5, 10, 15, 20, 25 and 30 µM doses of bryostatin 1. In vivo, bryostatin 1 prevented the TPA­mediated increase in the level of H2O2 and myeloperoxidase in mouse epidermal tissue. Pretreatment of the mice with bryostatin 1 (30 µM) followed by administration of TPA reduced the edema, as demonstrated via punched­out mouse ear tissue, to 7.2 mg, compared with 14 mg in the TPA­treated group. Treatment with bryostatin 1 prior to TPA administration markedly prevented the inflammation of the skin by inhibiting hyperplasia in the epidermal layer and the aggregation of inflammatory cells. The results demonstrated that treatment of mice with bryostatin 1 at a 30 µM dose prior to TPA administration significantly (P<0.005) inhibited the TPA­mediated increase in the level of COX­2. The activity of ornithine decarboxylase, increased by TPA, was additionally inhibited following pretreatment of the mice with bryostatin 1. In the mice treated with bryostatin 1 at 30 µM doses prior to the administration of TPA, the appearance of papillomas was 20%, compared with 100% in the TPA group. Mice pretreated with bryostatin 1 at 30 µM doses prior to TPA administration exhibited the appearance of 0.4 mean papillomas in each animal, compared with 5.2 in the TPA group. Therefore, the results of the present study demonstrated that bryostatin 1 inhibited the development and progression of tumors of skin in the mice, through the prevention of inflammation­inducing processes and the quenching of radicals. Therefore, bryostatin 1 maybe considered to be adrug of importance in the treatment of skin tumor.

Aqueous and Ethanol Extracts of Australian Cane Toad Skins Suppress Pro-Inflammatory Cytokine Secretion in U937 Cells via NF-κB Signaling Pathway.

Toad skin extracts, such as aqueous extracts (AE) of Chinese toad skins, have demonstrated therapeutic benefits for a range of diseases including pain, inflammation, swelling, heart failure, and various types of cancers. In this study, we investigated the anti-inflammatory potential of an AE (0.1-10 µg/mL) and a 60% ethanol extract (EE; 0.1-10 µg/mL) from Australian cane toad (Bufo marinus) skins and the known bioactive compound, bufotenine (BT; 0.1-10 nM). The assay employed a model of the human monocyte cell line U937 stimulated with lipopolysaccharide (LPS) and phorbol 12-myristate 13-acetate (PMA) for the release of tumor necrosis factor (TNF)-α and interleukin (IL)-6. We demonstrated that AE, EE, and BT significantly inhibited the release and expression of TNF-α and IL-6 in a dose-dependent manner when the cells were pre-treated at non-cytotoxic concentrations. Further investigation revealed that the inhibition of TNF-α and IL-6 release and expression was associated with the suppression of nuclear factor (NF)-kappa (κ)B activation. These results indicate that AE, EE, and BT are strong inflammation inhibitors, thus have the potential for further development as anti-inflammatory therapeutic agents from a natural source regarded as a feral pest in Australia. J. Cell. Biochem. 117: 2769-2780, 2016. © 2016 Wiley Periodicals, Inc.

Low dose triterpene-quinone fraction from Ardisia crispa root precludes chemical-induced mouse skin tumor promotion.

BACKGROUND: Drastic increment of skin cancer incidence has driven natural product-based chemoprevention as a promising approach in anticancer drug development. Apart from its traditional usages against various ailments, Ardisia crispa (Family: Myrsinaceae) specifically its triterpene-quinone fraction (TQF) which was isolated from the root hexane extract (ACRH) was recently reported to exert antitumor promoting activity in vitro. This study aimed at determining chemopreventive effect of TQF against chemically-induced mouse skin tumorigenesis as well as elucidating its possible pathway(s). METHODS: Mice (n = 10) were initiated with single dose of 7,12-dimethylbenz[α]anthracene (DMBA) (390 nmol/100 µl) followed by, a week later, repeated promotion (twice weekly; 20 weeks) with 12-O-tetradecanoylphorbol-13-acetate (TPA) (1.7 nmol/100 µl). TQF (10, 30 and 100 mg/kg) and curcumin (10 mg/kg; reference) were, respectively, applied topically to DMBA/TPA-induced mice 30 min before each TPA application. Upon termination, histopathological and biochemical analysis, as well as Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and transcription factor enzyme-linked immunosorbent assay (ELISA) assays were performed to elucidate the potential mechanism of TQF. RESULTS: With comparison to the carcinogen control, results revealed that lower dose of TQF (10 mg/kg) conferred antitumor promoting effect via significant (P < 0.05) suppression against lipid peroxidation (LPO), apoptotic index (cell death) and nuclear factor-kappa B (NF-κB), along with reduction of keratinocyte proliferation; whilst its higher dose (100 mg/kg) was found to promote tumorigenesis by significantly (P < 0.05) increasing LPO and apoptotic index, in addition to aggravating keratinocyte proliferation. CONCLUSIONS: This study evidenced that TQF, particularly at its lower dosage (10 mg/kg), ameliorated DMBA/TPA-induced mouse skin tumorigenesis. Though, future investigations are warranted to determine the lowest possible therapeutic dose of TQF in subsequent in vivo chemopreventive studies.

Inhibition of Breast Cancer Cell Proliferation and In Vitro Tumorigenesis by a New Red Apple Cultivar.

PURPOSE: The aim of this study was to evaluate the antiproliferative activity in breast cancer cells and the inhibition of tumorigenesis in pre-neoplastic cells of a new apple cultivar with reddish pulp, called the Pelingo apple. METHODS: The antiproliferative activity was evaluated in MCF-7 and MDA-MB-231 human breast cancer cells. The inhibition of tumorigenesis was performed in JB6 promotion-sensitive (P+) cells. RESULTS: Results showed that Pelingo apple juice is characterized by a very high polyphenol content and strongly inhibited breast cancer cell proliferation. Its antiproliferative activity was found to be higher than the other five apple juices tested. Pelingo juice induced cell accumulation in the G2/M phase of the cell cycle and autophagy through overexpression of p21, inhibition of extracellular signal-regulated kinases 1/2 (ERK1/2) activity and an increase in lipidated microtubule-associated protein-1 light chain-3 beta (LC3B). Remarkably, Pelingo juice inhibited the 12-o-tetra-decanoyl-phorbol-13-acetate (TPA)-induced tumorigenesis of JB6 P+ cells, suppressing colony formation in semi-solid medium and TPA-induced ERK1/2 phosphorylation. CONCLUSIONS: Our data indicate that the Pelingo apple is rich in food components that can markedly inhibit in vitro tumorigenesis and growth of human breast cancer cells and could provide natural bioactive non-nutrient compounds, with potential chemopreventive activity.

Cancer-promoting effect of capsaicin on DMBA/TPA-induced skin tumorigenesis by modulating inflammation, Erk and p38 in mice.

Epidemiologic and animal studies revealed that capsaicin (8-methyl-N-vanillyl-6-noneamide) can act as a carcinogen or cocarcinogen. However, the influence of consumption of capsaicin-containing foods or vegetables on skin cancer patients remains largely unknown. In the present study, we demonstrated that capsaicin has a cocarcinogenic effect on 9, 10-dimethylbenz[a]anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin tumorigenesis. Our results showed that topical application of capsaicin on the dorsal skin of DMBA-initiated and TPA-promoted mice could significantly accelerate tumor formation and growth and induce more and larger skin tumors than the model group (DMBA + TPA). Moreover, capsaicin could promote TPA-induced skin hyperplasia and tumor proliferation. Mechanistic study found that inflammation-related factors cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) were highly elevated by pretreatment with capsaicin, suggesting an inflammation-dependent mechanism. Furthermore, mice that were administered capsaicin exhibited significant up-regulation of phosphorylation of nuclear factor kappaB (NF-κB), Erk and p38 but had no effect on JNK. Thus, our results indicated that inflammation, Erk and P38 collectively played a crucial role in cancer-promoting effect of capsaicin on carcinogen-induced skin cancer in mice.