Tephrosia purpurea alleviates phorbol ester-induced tumor promotion response in murine skin.

In recent years, considerable emphasis has been placed on identifying new cancer chemopreventive agents, which could be useful for the human population. Tephrosia purpurea has been shown to possess significant activity against hepatotoxicity, pharmacological and physiological disorders. Earlier we showed that Tephrosia purpurea inhibits benzoyl peroxide-mediated cutaneous oxidative stress and toxicity. In the present study, we therefore assessed the effect of Tephrosia purpurea on 12-O-tetradecanoyl phorbal-13-acetate (TPA; a well-known phorbol ester) induced cutaneous oxidative stress and toxicity in murine skin. The pre-treatment of Swiss albino mice with Tephrosia purpurea prior to application of croton oil (phorbol ester) resulted in a dose-dependent inhibition of cutaneous carcinogenesis. Skin tumor initiation was achieved by a single topical application of 7,12-dimethyl benz(a)anthracene (DMBA) (25 microg per animal per 0.2 ml acetone) to mice. Ten days later tumor promotion was started by twice weekly topical application of croton oil (0.5% per animal per 0.2 ml acetone, v /v). Topical application of Tephrosia purpurea 1 h prior to each application of croton oil (phorbol ester) resulted in a significant protection against cutaneous carcinogenesis in a dose-dependent manner. The animals pre-treated with Tephrosia purpurea showed a decrease in both tumor incidence and tumor yield as compared to the croton oil (phorbol ester)-treated control group. In addition, a significant reduction in TPA-mediated induction in cutaneous ornithine decarboxylase (ODC) activity and [3H]thymidine incorporation was also observed in animals pre-treated with a topical application of Tephrosia purpurea. The effect of topical application of Tephrosia purpurea on TPA-mediated depletion in the level of enzymatic and non-enzymatic molecules in skin was also evaluated and it was observed that topical application of Tephrosia purpurea prior to TPA resulted in the significant recovery of TPA-mediated depletion in the level of these molecules, namely glutathione, glutathione S-transferase, glutathione reductase and catalase. From these data we suggest that Tephrosia purpurea can abrogate the tumor-promoting effect of croton oil (phorbol ester) in murine skin.     

Modulatory effect of gentisic acid on the augmentation of biochemical events of tumor promotion stage by benzoyl peroxide and ultraviolet radiation in Swiss albino mice

The present study was carried out to study the effect of gentisic acid (2,5-dihydroxybenzoic acid (2,5-DHBA)) on the tumor promotion related events of carcinogenesis in murine skin. Benzoyl peroxide (BPO) (20 mg/0.2 ml/animal) and ultraviolet radiations (UVR) (0.420 J/m2/s) were used to induce tumor promotion response and oxidative stress and caused significant depletion in the detoxification and antioxidant enzyme armory with concomitant elevation in malondialdehyde (MDA) formation, hydrogen peroxide (H2O2) generation, ornithine decarboxylase (ODC) activity and unscheduled DNA synthesis. However, gentisic acid pretreatment at two different doses restored the levels of the above said parameters (P < 0.05) in a dose-dependent manner except in the case of ODC activity. Therefore, we propose that it might suppress the promotion stage via inhibition of oxidative stress but may not affect the polyamine biosynthetic pathway.     

HPLC法测定祛痘类化妆品中的过氧化苯甲酰

目的建立高效液相色谱法测定祛痘除螨类化妆品中的过氧化苯甲酰的方法。方法采用ZORBAX C18柱,4．6×250mm．5μm;以甲醇-0．02mol·L^-1的醋酸铵溶液（80：20）为流动相;检测波长为230nm。结果过氧化苯甲酰在0．224μg·mL^-1～44．8μg·mL^-1范围内呈良好线性关系,r=0．9996．过氧化苯甲酰的平均回收率为95．94％．RSD％为2．5％。结论上述建立的方法简便易行、准确、重现性好,可用于祛痘类化妆品中禁用物质过氧化苯甲酰的检测。     

Role of the benzoyloxyl radical in DNA damage mediated by benzoyl peroxide.

Benzoyl peroxide (BzPO) is both a tumor promoter and progressor in mouse skin; however, BzPO is neither an initiator nor a complete carcinogen in this tissue. Although not mutagenic, BzPO has been observed to produce strand breaks in DNA of exposed cells. These actions are presumed to be mediated by free-radical derivatives of BzPO. Previous studies suggested that the metabolism of BzPO in keratinocytes proceeds via the initial cleavage of the peroxide bond, yielding benzoyloxy radicals which, in turn, can either fragment to form phenyl radicals and carbon dioxide or abstract H atoms from biomolecules to yield benzoic acid. Benzoic acid is the major stable metabolite of BzPO produced by keratinocytes. In the present study we have investigated the role of BzPO and its metabolites in the generation of strand scissions in a cell-free system using phi X-174 plasmid DNA. In this system BzPO produced DNA damage that was dose-dependent over a concentration range of 0.1-1 mM and required the presence of copper but not other transition metals. By contrast, benoic acid did not produce DNA damage in this system, either in the presence or in the absence of copper. The inclusion of spin trapping agents, such as N-tert-butyl-alpha-phenylnitrone (PBN), 3,5-dibromo-4-nitrosobenzenesulfonate, and nitrosobenzene, in incubations was found to significantly reduce the extent of DNA damage generated via the copper-mediated activation of BzPO. Electron paramagnetic resonance spectroscopy studies suggested that the primary radical trapped by PBN following copper-mediated decomposition of BzPO was the benzoyloxy radical.(ABSTRACT TRUNCATED AT 250 WORDS)     

Selective assay of benzoyl peroxide in lotions and creams.

A selective titrimetric method for the determination of benzoyl peroxide in lotions and creams was developed. It is based on the work of Horner and Jürgens, in which diacylperoxides, dialkylperoxides, peracids, and alkylhydroperoxides can be determined selectively by iodometry and acidimetry. The proposed assay is stability indicating with respect to peracids. Good recovery data were obtained.     

Terminoside A,a new triterpene glycoside from the bark of Terminalia arjuna inhibits nitric oxide production in murine macrophages

Terminoside A (1), a new oleanane-type triterpene was isolated from the acetone fraction of the ethanolic extract of stem bark of Terminalia arjuna. The structure was established as olean-1alpha,3beta,22beta-triol-12-en-28-oic acid-3beta-D-glucopyranoside. On the basis of spectral data and chemical reactions, terminoside A, potently inhibited nitric oxide (NO) production and decreased inducible nitric oxide synthase (iNOS) levels in lipopolysaccharide-stimulated macrophages.     

Terminoside A,a new triterpene glycoside from the bark of Terminalia arjuna inhibits nitric oxide production in murine macrophages

Terminoside A (1), a new oleanane-type triterpene was isolated from the acetone fraction of the ethanolic extract of stem bark of Terminalia arjuna. The structure was established as olean-1α,3β,22β-triol-12-en-28-oic acid-3β-D-glucopyranoside. On the basis of spectral data and chemical reactions, terminoside A, potently inhibited nitric oxide (NO) production and decreased inducible nitric oxide synthase (iNOS) levels in lipopolysaccharide-stimulated macrophages.     

Inhibitory effect of euphol, a triterpene alcohol from the roots of Euphorbia kansui, on tumour promotion by 12-O-tetradecanoylphorbol-13-acetate in two-stage carcinogenesis in mouse skin

The anti-inflammatory activity of euphol, twelve other triterpene alcohols and sitosterol-beta-D-glucopyranoside, isolated from the dichloromethane extract of the roots of Euphorbia kansui, has been evaluated in mice with inflammation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). TPA (1.7 nmol; 1.0 microg/ear) was dissolved in acetone and 10 microL delivered to the inner and outer surfaces of the right ear of ICR mice. A triterpene alcohol, sterol glucoside or vehicle (20 microL; chloroform-methanol 1:1), was applied topically approximately 30 min before each TPA treatment. The ear thickness was measured before treatment and then oedema was measured 6 h after TPA treatment. For the two-stage carcinogenesis experiment, initiation was accomplished by administration of a single topical application of 7,12-dimethylbenz[a]anthracene (DMBA; 195 nmol; 50 microg/mouse) to the shaved backs of mice. Promotion was with 1.7 nmol (1.0 microg) TPA, applied twice weekly to the same shaved area, begun one week after the initiation. Euphol (2.0 micromol; 853 microg), or its vehicle (acetone-dimethylsulphoxide, 9:1; 100 microL), was applied topically 30 min before each TPA treatment. The number and diameter of skin tumours were measured every other week for 20 weeks. All the compounds were found to possess marked inhibitory activity and their 50% inhibitory dose for TPA-induced inflammation was 0.2-1.0 mg/ear. Topical application of euphol (2.0 micromol; 853 microg/mouse) markedly suppressed the tumour-promoting effect of TPA (1.7 nmol; 1.0 microg/mouse) in mouse skin initiated with DMBA.     

Rutin inhibits human leukemia tumor growth in a murine xenograft model in vivo

Numerous studies have shown that rutin has anticancer effects. We have previously reported that rutin induced cell cycle arrest and apoptosis in murine leukemia WEHI‐3 cells in vitro and in vivo. However, there are no data showing that rutin inhibits human leukemia HL‐60 cells in vivo in a murine xenograft animal model. Human leukemia HL‐60 cells were implanted into mice and treated with vehicle (1% DMSO), rutin (120 mg/kg of body weight) or vinblastine (120 μg/kg of body weight). Compounds and agents were injected once every four days intraperitoneally (i.p.) for 36 days. Treatment with 120 mg/kg of rutin or with 120 μg/kg of vinblastine resulted in a reduction of tumor weight and volume when compared with the control groups. Tumor size in xenograft mice treated with 120 mg/kg of rutin was significantly smaller than that in the untreated‐control group. These novel findings indicate that rutin inhibits tumor growth in a xenograft animal model. Rutin may be useful in treating leukemia but certainly much more research is needed. © 2011 Wiley Periodicals, Inc. Environ Toxicol, 2012.     

Low iron state is associated with reduced tumor promotion in a two-stage mouse skin carcinogenesis model.

The purpose of this study was to investigate the effect of low iron state in a two-stage mouse skin carcinogenesis model using 7,12-dimethylbenz[a]anthracene (DMBA)-initiated and benzoyl peroxide (BPO)-promoted cutaneous tumorigenesis. All mice were treated with DMBA. Low iron state was achieved by injection with phenylhydrazine hydrochloride and feeding low iron diet. A low iron state resulted in a decrease in tumor incidence (papillomas and carcinomas) and number of tumors/mouse. Also, the conversion of papillomas to carcinomas was lower in mice on a low iron state. BPO treatment enhanced epidermal lipid peroxidation (LPO) and was accompanied by a depletion in the level of epidermal reduced glutathione (GSH) and decrease in the activities of antioxidant enzymes. BPO treatment also increased ornithine decarboxylase (ODC) activity and [3H]thymidine incorporation into cutaneous DNA. Mice in a low iron state were less susceptible to the effects of BPO treatment, as was apparent from a partial recovery of GSH levels and the activities of antioxidant enzymes, as well as a lower induction in ODC activity, [3H]thymidine incorporation into cutaneous DNA and lesser epidermal LPO. As expected, cutaneous iron levels were lower in mice on a low iron state. Thus, our data show that the tumor-promoting potential of BPO is reduced by low iron state in a two-stage mouse skin carcinogenesis model.     

Effect of Hibiscus rosa sinensis extract on hyperproliferation and oxidative damage caused by benzoyl peroxide and ultraviolet radiations in mouse skin

The present study was conducted to investigate the ameliorative potential of Hibiscus rosa sinensis extract in mice skin. Combination of a single topical application of benzoyl peroxide (20 mg/0.2 ml/animal) followed by ultraviolet radiations (0.420 J/m2/s) was used to induce hyperproliferation and oxidative stress. Single benzoyl peroxide application prior to ultraviolet B radiations exposure caused significant depletion in the detoxification and antioxidant enzymes, while malondialdehyde formation, hydrogen peroxide content, ornithine decarboxylase activity and DNA synthesis were raised significantly. However, pretreatment of H. rosa sinensis extract (3.5 mg and 7 mg/ kg b.wt.) partly restored the levels of cellular protective enzymes (P<0.05). Besides, malondialdehyde formation and hydrogen peroxide content (P<0.05) were statistically significantly reduced at both doses. The ornithine decarboxylase activity and thymidine incorporation in DNA were also reduced dose dependently (P<0.05) by the plant extract. Therefore, we propose that H. rosa sinensis extract exerts a protective effect against the tumour promotion stage of cancer development.     

High-pressure liquid chromatographic assay of benzoyl peroxide in dermatological gels and lotions.

A high-pressure liquid chromatographic method for the assay of benzoyl peroxide in dermatological preparations is described. Degradation products such as benzoic acid and perbenzoic acid do not interfere. The method is simple, precise, accurate, and stability indicating.     

Growth inhibition and apoptosis induced by lupeol, a dietary triterpene, in human hepatocellular carcinoma cells

Hepatocellular carcinoma (HCC) is the fifth most malignant tumor worldwide and is known to be resistant to conventional chemotherapy. New therapeutic strategies are urgently needed for treating HCC. Lup-20(29)-en-3H-ol (Lupeol), a novel dietary triterpene, is found in fruits, vegetables, and medicinal plants and possesses multiple bio-activities with very low toxicity. In the current study, we investigated its growth-inhibitory effects in HCC cell lines SMMC7721 and HepG2. In the in vitro studies, lupeol treatment alone caused decrease of cell viability in two HCC cell lines in a dose-dependent manner. It also induced apoptosis and caused cell accumulation in S phase. Further analysis revealed the induction of active caspase-3 and poly(ADP-ribose)polymerase (PARP) cleavage by lupeol treatment. In the in vivo studies, nude mice implanted with SMMC7721 cells subcutaneously were treated with lupeol three times a week and tumor development was significantly inhibited. We further investigated the combination anti-tumor effect of lupeol and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in HCC, considering TRAIL treatment alone could not achieve high level of anti-tumor effect. The results demonstrated that lupeol could exert a combinational effect with TRAIL, resulting in chemosensitization of HCC. Our results suggested that lupeol alone or as an adjuvant to therapeutic agents could be developed as a potential agent for treating HCC.     

Comparison of clindamycin/benzoyl peroxide, tretinoin plus clindamycin, and the combination of clindamycin/benzoyl peroxide and tretinoin plus clindamycin in the treatment of acne vulgaris: a randomized, blinded study

In the treatment of mild to moderate acne vulgaris, the combination of an antibiotic and benzoyl peroxide provides enhanced efficacy over the individual agents, with the potential to decrease the emergence of resistant strains of P. acnes. To evaluate treatment regimens combining the daily use of a clindamycin/benzoyl peroxide gel, a tretinoin gel, and a clindamycin gel, the current randomized, evaluator-blind study was conducted. Results demonstrate that once-daily administration of clindamycin/benzoyl peroxide gel (combination formulation) was as effective as clindamycin/benzoyl peroxide gel + tretinoin gel + clindamycin gel. Both of these regimens provided greater efficacy than tretinoin + clindamycin. Treatment with clindamycin/benzoyl peroxide demonstrated a significant benefit over other treatments at Week 2, highlighting its rapid onset of action. All regimens were safe and generally well tolerated, with less severe peeling seen in patients who received clindamycin/benzoyl peroxide. In conclusion, the regimens that included clindamycin/benzoyl peroxide were more effective than tretinoin + clindamycin in the treatment of acne vulgaris, with no clinical advantage of adding tretinoin + clindamycin to once-daily clindamycin/benzoyl peroxide treatment.     

Dose response of early effects related to tumor promotion of 2-acetylaminofluorene.

The genotoxic effects of 2-acetylaminofluorene (AAF) alone cannot explain the formation of rat liver tumors. It has been proposed that mitochondrial effects are associated with its tumor-promoting properties. These mitochondrial effects are thought to trigger apoptosis and regenerative proliferation, which alters the liver lobule in a cirrhosis-like manner. A situation is generated which favors the growth of initiated cells. To test this sequence of events, the dose dependence of early effects with time was studied. Male Wistar rats received 50, 100, 200, 400, or 800 ppm AAF in the diet and the following endpoints were analyzed at 2, 4, 8, and 16 weeks of feeding: apoptotic cell death, cell proliferation, GST-P-positive foci (placental form of glutathione S-transferase), and morphological alterations. Hydrolyzable hemoglobin adducts were used as a dosimeter for metabolic activation after 2 weeks of feeding. The hemoglobin adduct levels increased linearly with dose. With the conventional carcinogenic concentration of 200-ppm AAF in the diet, the number of apoptoses increased first, predominantly in the periportal area (2 weeks). Cell proliferation followed with some delay (4 weeks). This reflects regenerative tissue response and not the growth of initiated cells, because the number of enzyme-altered foci was still extremely low at that time. Foci developed only later when the morphology had changed. With 50 ppm AAF in the diet, a no-effect level had not been reached for any of the endpoints, but foci developed much more gradually than with higher doses. Unexpectedly, the proliferative response stabilized at 8 weeks with a labeling index of 12-17, with all AAF concentrations. The observed sequence of events supports the hypothesis. It is concluded that (1) The proliferation of initiated cells-defined as promotable cells-is largely determined by the cellular environment, such as morphologically altered liver. (2) The morphological alterations in rat liver result from imperfect regeneration from toxic effects. (3) Imperfect regeneration results from limitations in the possibilities to adapt to chemical stress. (4) If these limits are overwhelmed and morphology has changed to a certain extent, preneoplastic foci develop; this occurs much faster, at least, than without these changes.     

Effect of scutellarin on nitric oxide production in early stages of neuron damage induced by hydrogen peroxide.

The aims of the present study were to investigate the regulatory function of scutellarin on production of nitric oxide (NO) as well as activities of constitutive NO synthase (cNOS) and inducible NO synthase (iNOS) in early stages of neuron damage induced by hydrogen peroxide. Direct detection of NO production was performed on primary cultures of living rat neuronal cells with an electrochemical sensor. Hydrogen peroxide significantly increased culture supernatant levels of NO, the total integral value of the defined areas (500-6500 sxpA) reached 3.68 x 10(6). Pre-treatment with scutellarin, caused the total integral value to decrease in a dose-dependent fashion (3.24 x 10(6), 2.15 x 10(6), 1.84 x 10(6) for groups 10, 50, and 100 uM scutellarin, respectively). After exposure to 2.0mM hydrogen peroxide for 2h, malondialdehyde (MDA) level, a marker of lipid peroxidation, was remarkably increased. The elevation can be suppressed by scutellarin. Hydrogen peroxide also caused significant loss of neuron viability. In comparison with the control group, scutellarin significant attenuated the loss. Results also showed that hydrogen peroxide increased activity of cNOS, which was markedly inhibited by scutellarin. However, exposure of neuronal cells to hydrogen peroxide did not lead to an increase in iNOS activity. In conclusion, our results suggest that NO production, which increased in early stages of neuron damage induced by hydrogen peroxide can be effectively inhibited by scutellarin. Moreover, our results indicate that increase in NO production is mediated by cNOS.     

Investigation of degradation products in a topical gel containing erythromycin and benzoyl peroxide by liquid chromatography-mass spectrometry

Benzamycin, combining benzoyl peroxide and erythromycin, is a topical gel used in the treatment of acne vulgaris. Because of the reactivity of benzoyl peroxide, preparations containing both erythromycin and benzoyl peroxide might be unstable and degradation products could be formed. To investigate and identify these latter products, a gradient-based liquid chromatographic method using volatile mobile phase constituents was developed. Mass spectrometry data were acquired on solutions containing erythromycin and benzoyl peroxide and on freshly prepared, 2-month-old and 18-month-old samples of Benzamycin. With the reference spectra as interpretative templates, it was concluded that erythromycin undergoes oxidation, followed by benzoylation.     

A calcitonin gene-related peptide (CGRP) antagonist (CGRP8-37) inhibits microvascular responses induced by CGRP and capsaicin in skin.

1. The effect of the calcitonin gene-related peptide (CGRP) antagonist CGRP8-37 on responses to CGRP and other mediators was investigated in rabbit dorsal skin. 2. Blood flow changes at intradermally-injected sites were measured by a multiple site 133xenon clearance technique. CGRP8-37 had little effect on blood flow at doses up to 0.3 nmol/site, when injected alone, although a significant increase in blood flow was observed at the highest dose tested (1 nmol/site). 3. CGRP8-37 dose-dependently inhibited the increased blood flow induced by human alpha CGRP and human beta CGRP, but had no effect on equivalent vasodilator responses induced by vasoactive intestinal peptide (VIP) and prostaglandin E1 (PGE1). CGRP8-37 showed a preferential ability to inhibit alpha CGRP (IC50 0.04 nmol), when compared with beta CGRP (IC50 greater than or equal to 0.3 nmol). 4. Capsaicin, which selectively activates sensory nerves, caused a dose-dependent increase in blood flow when injected intradermally into rabbit skin. The effects of capsaicin (0.01-0.1 mumol/site) were inhibited by CGRP8-37 (0.3 nmol/site), with a partial but significant attenuation of blood flow induced by the highest dose of capsaicin. 5. Oedema formation, induced by intradermal histamine injection (3 nmol/site), was measured in rabbit skin by the local accumulation of intravenously-injected 125I-labelled albumin. Vasodilator doses of CGRP, PGE1 and capsaicin potentiated, in a synergistic manner, oedema formation induced by histamine. GRP8-37 totally inhibited the potentiating effect of CGRP, partially inhibited the synergistic effect of capsaicin, but did not affect PGE1-induced responses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ruthenium red selectively inhibits oedema formation and increased blood flow induced by capsaicin in rabbit skin.

It has been suggested that ruthenium red has a selective inhibitory effect on capsaicin-induced nociceptor stimulation. We have investigated the effect of ruthenium red on oedema formation and vasodilatation induced by intradermal (i.d.) injection of capsaicin in the rabbit in vivo. Responses induced by capsaicin were inhibited by ruthenium red, but responses induced by bradykinin, N-formyl-methionyl-leucyl-phenylalanine (FMLP), platelet activating factor (PAF), histamine and calcitonin gene-related peptide (CGRP) were not affected. These results suggest that ruthenium red selectively inhibits capsaicin-induced local plasma protein leakage and vasodilatation in the rabbit skin microvasculature.