Inhibition of cyclooxygenase-2 and inducible nitric oxide synthase by silymarin in proliferating mesenchymal stem cells: comparison with glutathione modifiers

Silymarin, a mixture of flavonolignans, is extracted from milk thistle (Silybum marianum) and has a strong antioxidant activity and exhibits anticarcinogenic, anti-inflammatory, and cytoprotective effects. In this study we attempted to determine whether silymarin and the glutathione modifiers, buthionine sulfoxamine (BSO) and N-acetylcysteine (NAC), are involved in regulation of cyclooxygenase (COX)-2 and inducible nitric oxide synthase (iNOS) in proliferating mesenchymal stem cells (MSCs). Cellular glutathione was manipulated during a 14-day culture using BSO, NAC and silymarin. At intervals of 2, 7 and 14 days, cells were collected and COX-2 and iNOS levels were measured. In parallel, generation of cellular H₂O₂ and glutathione were measured. Supplementation of the culture media with BSO caused a dose-dependent decrease in MSC proliferation, whereas NAC or silymarin elevated the proliferation (p < 0.05). Treatment of MSC with NAC or silymarin caused a significant decrease in COX-2 levels. However, COX-2 levels in cells treated with high levels of NAC (1.0 mM) were significantly lower than those in MSCs treated with high levels of silymarin (100 μM). BSO (1.0 and 5.0 μM) caused a significant increase in COX-2 on days 2, 7 and 14. BSO caused a significant increase in iNOS, whereas NAC or silymarin decreased cellular iNOS. Overall result show that glutathione, iNOS and COX-2 in proliferating MSCs are affected by silymarin treatment. It appears that glutathione is the main target of silymarin, and in consequence iNOS and COX-2 are affected in response to silymarin treatment.     

Silymarin alleviates bleomycin-induced pulmonary toxicity and lipid peroxidation in mice

Context: The application of bleomycin is limited due to its side effects including lung toxicity. Silymarin is a flavonoid complex isolated from milk thistle [Silybum marianum L. (Asteraceae)] which has been identified as an antioxidant and anti-inflammatory compound.

Objective: This study evaluates the effect of silymarin on oxidative and inflammatory parameters in the lungs of mice exposed to bleomycin.

Materials and methods: BALB/c mice were divided into four groups of control, bleomycin (1.5?U/kg), bleomycin plus silymarin (50 and 100?mg/kg). After bleomycin administration, mice received 10?d intraperitoneal silymarin treatment. On 10th day, blood and lung samples were collected for measurement of oxidative and inflammatory factors.

Results: Silymarin led to a decrease in lung lipid peroxidation (0.19 and 0.17?nmol/mg protein) in bleomycin-injected animals. Glutathione-S-transferase (GST) which was inhibited by bleomycin (32.4?nmol/min/mg protein) induced by higher dose of silymarin (41?nmol/min/mg protein). Silymarin caused an elevation in glutathione (GSH): 2.6 and 3.1?µmol/g lung compare with bleomycin-injected animals 1.8?µmol/g lung. Catalase (CAT) was increased due to high dose of silymarin (65.7?µmol/min/ml protein) compare with bleomycin treated-mice. Myeloperoxidase (MPO) which was induced due to bleomycin (p?p?Conclusions: Silymarin attenuated bleomycin induced-pulmonary toxicity. This protective effect may be due to the ability of silymarin in keeping oxidant–antioxidant balance and regulating of inflammatory mediator release.     

Exposure to diesel exhaust upregulates COX-2 expression in ApoE knockout mice

Introduction: We have shown that diesel exhaust (DE) inhalation caused progression of atherosclerosis; however, the mechanisms are not fully understood. We hypothesize that exposure to DE upregulates cyclooxygenase (COX) expression and activity, which could play a role in DE-induced atherosclerosis.

Methods: ApoE knockout mice (30-week old) fed with regular chow were exposed to DE (at 200 µg/m³ of particulate matter) or filtered air (control) for 7 weeks (6?h/day, 5 days/week). The protein and mRNA expression of COX-1 and COX-2 were evaluated by immunohistochemistry analysis and quantitative real-time PCR, respectively. To examine COX activity, thoracic aortae were mounted in a wire myograph, and phenylephrine (PE)-stimulated vasoconstriction was measured with and without the presence of COX antagonists (indomethacin). COX-2 activity was further assessed by urine 2,3-dinor-6-keto PGF_1α level, a major metabolite of prostacyclin I₂ (PGI₂).

Results: Immunohistochemistry analysis demonstrates that DE exposure enhanced COX-2 expression in both thoracic aorta (p < 0.01) and aortic root (p < 0.03), with no modification of COX-1 expression. The increased COX-2 expression was positively correlated with smooth muscle cell content in aortic lesions (R² = 0.4081, p < 0.008). The fractional changes of maximal vasoconstriction in the presence of indomethacin was attenuated by 3-fold after DE exposure (p < 0.02). Urine 2,3-dinor-6-keto PGF_1α level was 15-fold higher in DE group than the control (p < 0.007). The mRNA expression of COX-2 (p < 0.006) and PGI synthase (p < 0.02), but not COX-1, was significantly augmented after DE exposure.

Conclusion: We show that DE inhalation enhanced COX-2 expression, which is also associated with phenotypic changes of aortic lesion.     

Silymarin selectively protects human renal cells from cisplatin-induced cell death

Context: Cisplatin-induced nephrotoxicity has been accepted as an important obstacle for efficient cisplatin-based chemotherapy. Silymarin from seeds of milk thistle [Silybum marianum L. (Asteraceae)] has been shown to possess various potential pharmacological properties; however, whether or not this agent selectively protects renal cells from cisplatin-induced cell death with no interfering effect on cancer cells is not clear.

Objective: Potential of silymarin in protection of cisplatin-induced renal cell death without compromising effect on anticancer activity of cisplatin was demonstrated in this study.

Materials and methods: Cisplatin-induced cell death was evaluated in human proximal tubular HK-2, lung carcinoma H460, and melanoma G361 cells using MTT, Hoechst 33342, and propidium iodide assays.

Results: Cisplatin induced both apoptosis and necrosis in HK-2 cells and caused a decrease in cell viability by ~40% and 60% at the doses of 25 and 100 µM, respectively. Pretreatment with 25–200 µM of silymarin significantly protected against cisplatin-induced cell death in a dose-dependent manner. In contrast, pretreatment of silymarin (25–100 µM) caused no significant change on cisplatin-induced cell death in H460 cells but significantly potentiated cisplatin-induced apoptosis in G361 cells.

Discussion and conclusion: These findings reveal the selectivity of silymarin in protection of renal cells from cisplatin-induced cell death and could be beneficial for the development of this considerately safe compound as a renoprotective agent.     

Phytochemical investigation and radical scavenging activities of Melia azedarach and its DNA protective effect in cultured lymphocytes

Abstract

Context: Melia azedarach Linn (Meliaceae) is an Ayurvedic medicinal plant which is native to India. It is traditionally used for the treatment of leprosy, inflammation, scrofula, anthelmintic, antilithic, diuretic, deobstruent and cardiac disorders.

Objective: To evaluate the phytochemical constituents and antioxidant activities of the ethanol leaf extract of Melia azedarach (MA) and its protective effect against H₂O₂-induced cellular damage in cultured lymphocytes.

Materials and methods: The dose-dependent study of MA (20, 40, 60, 80, 100?µg/ml) was used to study in vitro radical scavenging assays. The effective dose of MA (60?µg/ml) was further used to study the H₂O₂-induced DNA damage (comet assay and DNA fragmentation assay) in cultured lymphocytes.

Results: The ethanol extract of MA (20, 40, 60, 80, 100?µg/ml) exhibited a significant dose-dependent inhibition of in vitro radical scavenging assays and their corresponding IC₅₀ values as follows: hydroxyl radical (26.50?±?0.26?µg/ml), superoxide anion (30.00?±?0.32?µg/ml), nitric oxide radical (48.00?±?0.48?µg/ml), DPPH radical (30.55?±?0.32?µg/ml) and reducing power (22.00?±?0.22?µg/ml). The increase in the severity of DNA damage and TBARS was increased significantly (p?<?0.05) at 500?µM H₂O₂-treated cultured lymphocytes and RBC cellular membranes. The phytochemical screening studies identified 13 chemical constituents present in the leaf extract of MA.

Discussion and conclusion: The results of this study demonstrate that MA offers protection against H₂O₂-induced cellular damage and it can be developed as an effective antioxidant during oxidative stress.     

Altered oxidative status and integrin expression in cyclosporine A-treated oral epithelial cells

Abstract

Background and objective: Cyclosporine A (CsA) is an immunosuppressive agent administered to transplant patients. A well-known reported oral side effect of CsA consumption is gingival overgrowth (GO). Changes in the expression of integrins occurring in the gingiva following CsA treatment have been reported but these reports are mainly concerned with the connective tissue of the gingiva. In this study we targeted the alterations in the oral epithelium using KB cells, an oral epithelial cell line.

Methods: Cultured oral epithelial cells were treated with increasing concentrations of CsA (0.1, 1 and 10?µg/mL) and the molecular changes involving antioxidant enzymes [glutathione peroxidase (GPx) and glutathione reductase (GR)] and the level of reactive oxygen species (ROS) were measured. Quantitative real-time PCR was used to assess the expression of selected integrins (α₂, α₅ and β₁).

Results: At CsA concentration above 0.1?µg/mL GPx demonstrated an increase in activity while GR activity and the level of reduced glutathione were diminished (p?<?0.05). α₅ and β₁ integrin were downregulated at all treatment concentrations of CsA while α₂ integrin presented this effect at concentrations above 1?µg/mL (p?<?0.05).

Conclusion: The results suggest a possible role for oxidative stress and the altered expression of integrins in the pathology of CsA-induced gingival overgrowth.     

Apoptotic role of natural isothiocyanate from broccoli (Brassica oleracea italica) in experimental chemical lung carcinogenesis

Abstract

Context: Sulforaphane (SFN) [1-isothiocyanato-4-(methylsulfinyl)butane] is a naturally occurring isothiocyanate found in cruciferous vegetables such as broccoli [Brassica oleracea L. var. italica Plenck. (Brassicaceae)]. Since it is among the most potent bioactive components with antioxidant and antitumor properties, it has received intense attention in the recent years for its chemopreventive properties.

Objective: The present work determined the rehabilitating role in alleviating the oxidative damage caused by benzo(a)pyrene [B(a)P] to biomolecules and the apoptotic cascade mediated by orally administered isothiocyanate-SFN (9?µmol/mouse/day) against B(a)P (100?mg/kg body weight, i.p.) induced pulmonary carcinogenesis in Swiss albino mice.

Materials and methods: Oxidative damage was assessed by measuring lipid peroxidation, 8-hydroxydeoxyguanosine, hydrogen peroxide (H₂O₂) production, glycoprotein components, protein carbonyl levels and DNA-protein crosslinks. DNA fragmentation by agarose gel electrophoresis and caspase-3 activity by ELISA proved apoptotic induction by SFN along with the protein expression of Bcl-2, Bax and Cyt c.

Results: SFN treatment was found to decrease the H₂O₂ production (p?<?0.001) in cancer induced animals, proving its antioxidant potential. Apoptosis was induced by increasing the release of Cyt c (p?<?0.001) from mitochondria, decreasing and increasing the expression of Bcl-2 (p?<?0.01) and Bax (p?<?0.001), respectively. Caspase-3 activity was also enhanced (p?<?0.001) which leads to DNA fragmentation in SFN treated groups.

Conclusion: Our results reflect the rehabilitating role of SFN in B(a)P induced lung carcinogenesis.     

Anti-inflammatory effects of Cynanchum taiwanianum rhizome aqueous extract in IL-1β-induced NRK-52E cells

Context: Cynanchum taiwanianum T. Yamaza (Asclepiadaceae) is a medicinal herb used in folk medicine for the treatment of several inflammation-related diseases such as hepatitis and dermatitis in Taiwan.

Objective: In the present study, we investigated the anti-inflammatory effect of C. taiwanianum T. Yamaza rhizome aqueous extract (CTAE).

Materials and methods: The present study investigated the anti-inflammatory effect of CTAE using IL-1β-induced NRK-52E cells. Production of NO and PGE₂ by ELISA, the mRNA and protein expression of iNOS and COX-2, phosphorylation of IκBα, and activation of NF-κB by RT-PCR and western blotting were determined.

Results: The CTAE significantly (P?<?0.05) inhibited NO and PGE₂ production (decreased by 46.1% and 51%, respectively), and also significantly (P?<?0.05) attenuated protein and mRNA expression of iNOS and COX-2 (decreased by 90% and 55% for iNOS and by 72% and 74%% for COX-2, respectively) in IL-1β-induced NRK-52E cells, in a dose-dependent manner, without obvious cytotoxic effects. Furthermore, the CTAE suppressed the NF-κB nuclear translocation, in terms of inhibition of IκBα phosphorylation.

Discussion and conclusion: Our results provided evidence for its folkloric uses and suggest that the anti-inflammatory activities of CTAE may result from the inhibition of inflammatory mediators, such as NO and PGE₂, and an upstream suppression of a NF-κB-dependent mechanism, might be involved.     

Emodin induces cytotoxic effect in human breast carcinoma MCF-7 cell through modulating the expression of apoptosis-related genes

Abstract

Context: The poor prognostic outcome of breast cancer is largely due to its resistance to cancer therapies. Development of therapeutic agents that can inhibit growth and induce apoptosis in breast cancer cells can help solve the problem. Emodin is an active anthraquinone that has been reported to have diverse biological effects.

Objective: In this study, the anticancer effects of emodin on growth inhibition, apoptosis induction and the expression of apoptosis-related genes in MCF-7 cells were investigated.

Materials and methods: Growth inhibition induced by emodin was investigated by the MTS assay and the colony formation assay; while emodin-induced apoptosis was determined by the COMET assay and DNA fragmentation detection. Emodin (35?μM)-induced alterations in the expression of apoptotic-related genes were detected by using real-time PCR.

Results: Emodin had significant growth inhibitory effects on MCF-7 cells with IC₅₀?=?7.22?µg/ml (～30?μM). It also exerted a concentration-dependant inhibitory effect on the colony-forming ability of MCF-7 cells with IC₅₀?=?7.60?µg/ml (～30?µM). Hallmarks of apoptosis, such as single-strand DNA breakage and DNA fragmentation, were observed in emodin-treated MCF-7 cells. The gene expression of Fas ligand (FASL) was up-regulated (p?<?0.01) but those of MCL1, CCND1 and C-MYC were down-regulated (p?<?0.05) in emodin-treated MCF-7 cells.

Discussion and conclusion: This study indicated that emodin could induce growth inhibition and apoptosis in MCF-7 cells through the modulation of the expression of apoptosis-related genes. The growth inhibitory effects of emodin might involve both the intrinsic and the extrinsic apoptotic pathways and cell cycle arrest.     

24-hour bronchodilator efficacy of single doses of indacaterol in subjects with COPD: comparison with placebo and formoterol

ABSTRACT

Objective: To assess the bronchodilator efficacy, safety and tolerability of indacaterol, a novel, once-daily inhaled β₂-agonist bronchodilator, in patients with chronic obstructive pulmonary disease (COPD).

Methods: This crossover, double-blind, double-dummy study was conducted to evaluate the 24-h bronchodilator effect of a range of single doses of indacaterol (150?µg, 300?µg and 600?µg), given in the morning via single-dose dry powder inhaler (SDDPI) in subjects with COPD, compared with placebo and with the daily therapeutic dose of formoterol (two 12?µg doses 12?h apart, via an SDDPI). Tolerability and safety were also assessed.

Results: Fifty-one subjects with moderate-to-severe COPD received each of the five treatments on separate study days in randomised sequence. The 24-h trough FEV₁ (primary endpoint; mean [95%?CI]) was 1.46 (1.43, 1.49)?L with indacaterol 600?µg (p?<?0.001 vs. placebo, p?<?0.01 vs. formoterol, p?<?0.05 vs. indacaterol 150?µg), 1.45 (1.42, 1.48)?L with indacaterol 300?µg (p?<?0.001 vs. placebo, p?<?0.05 vs. formoterol), 1.42 (1.39, 1.45)?L with indacaterol 150?µg (p?<?0.001 vs. placebo), 1.41 (1.38, 1.43)?L with formoterol (p?<?0.001 vs. placebo) and 1.28 (1.25, 1.31)?L with placebo. All treatments were well tolerated and there was little effect on serum potassium, blood glucose or QTc interval.

Conclusion: All doses of indacaterol were effective in providing 24-h bronchodilation and were well-tolerated in subjects with COPD. The bronchodilator efficacy of indacaterol (150, 300 and 600?μg) at 24?h post-dose was at least as efficacious as formoterol 12?µg twice daily.     

Antidiabetic and antioxidant potency evaluation of different fractions obtained from Cucumis prophetarum fruit

Abstract

Context: Cucumis prophetarum Linn. (Cucurbitaceae) fruit is used for inflammatory-related problems and is proved to be possessing anticancer and hepatoprotective effects.

Objective: The present investigation was to study the effect of different fractions of C. prophetarum on antidiabetic and antioxidant activity.

Materials and methods: Aqueous crude extract (CE) of C. prophetarum fruits was fractionated into water soluble fraction 1 (F1), chloroform fraction 2 (F2), basic fraction 3 (F3), and neutral fraction 4 (F4) by acid–base extraction. CE and its fractions at different doses (0.02–0.1?mg/mL) were subjected to antidiabetic (α-amylase and α-glucosidase inhibition assays) and antioxidant (DPPH, superoxide radical scavenging (SO) and metal chelation) evaluation.

Results: F1 exhibited effective antidiabetic activity (p?<?0.05) with an IC₅₀ value of 20.6 and 59.9?µg/mL. The activity decreased in the order of CE?>?F4?>F3?>?F2, according to α-amylase assay, which were the same, with the exception of the rank order of F4 and CE, as the α-glucosidase assay. Furthermore, F1 (IC₅₀?=?73?µg/mL) showed better reducing ability than CE?>F4?>F2?>?F3 (IC₅₀?=?78–272?µg/mL), according to the DPPH assay. In SO and metal chelation assays, F1 showed the highest activity (IC₅₀?=?101 and 147?µg/mL), respectively; the activity decreased in the order of CE?>F4?>F3?>?F2 (IC₅₀?=?126–469?µg/mL) for SO and 194–944?µg/mL for metal chelation assay.

Conclusion: The results indicate that F1 possesses potent in vitro antidiabetic and antioxidant activities.     

Hepatoprotective effect of withanolide-rich fraction in acetaminophen-intoxicated rat: decisive role of TNF-α, IL-1β, COX-II and iNOS

Context: Overdose of acetaminophen (APAP) is common in humans and is often associated with hepatic damage. Withania somnifera (L.) Dunal (Solanaceae) shows multiple pharmacological activities including antioxidant and anti-inflammatory potential.

Objective: To evaluate the possible mechanism of hepatoprotective activity of withanolide-rich fraction (WRF) isolated from a methanolic extract of Withania somnifera roots.

Materials and methods: Hepatotoxicity was induced by oral administration of APAP (750?mg/kg, p.o.) for 14 d. The control group received the vehicle. APAP-treated animals were given either silymarin (25?mg/kg) or graded doses of WRF (50, 100 and 200mg/kg) 2?h prior to APAP administration. Animals were killed on 15th day and blood and liver tissue samples were collected for the further analysis.

Results: In WRF-treated group, there was significant and dose-dependent (p?<?0.01 and p?<?0.001) decrease in serum bilirubin, ALP, AST and ALT levels with significant and dose-dependent (p?<?0.01 and p?<?0.001) increase in hepatic SOD, GSH and total antioxidant capacity. The level of MDA and NO decreased significantly (p?<?0.01) by WRF treatment. Up-regulated mRNA expression of TNF-α, IL-1β, COX-II and iNOS was significantly down-regulated (p?<?0.001) by WRF. Histological alternations induced by APAP in liver were restored to near normality by WRF pretreatment.

Conclusion: WRF may exert its hepatoprotective action by alleviating inflammatory and oxido-nitrosative stress via inhibition of TNF-α, IL-1β, COX-II and iNOS.     

The flavonolignan‐silymarin protects enzymatic,hematological, and immune system against γ‐radiation‐induced toxicity

The main focus of this study is evaluation of radioprotective efficacy of silymarin, a flavonolignan, against γ‐radiation‐induced damage to hematological, vital organs (liver and intestine), and immune system. Survival studies revealed that silymarin (administered orally for 3 days) provided maximum protection (67%) at 70 mg/kg body weight (b.wt.) against lethal 9 Gy γ‐irradiation (dose reduction factor = 1.27). The study revealed significant (p < 0.05) changes in levels of catalase (12.57 ± 2.58 to 30.24 ± 4.89 units), glutathione peroxidase (6.23 ± 2.95 to 13.26 ± 1.36 µg of reduced glutathione consumed/min/mg protein), glutathione reductase (0.25 ± 5.6 to 11.65 ± 2.83 pM NADPH consumed/min/mg protein), and superoxide dismutase (11.74 ± 0.2 to 16.09 ± 3.47 SOD U/mg of protein) activity at 30th day. Silymarin pretreated irradiated group exhibited increased proliferation in erythrocyte count (1.76 ± 0.41 × 10⁶ to 9.25 ± 0.24 × 10⁶), hemoglobin (2.15 ± 0.48g/dL to 14.77 ± 0.25g/dL), hematocrit (4.55 ± 0.24% to 37.22 ± 0.21%), and total leucocyte count (1.4 ± 0.15 × 10⁶ to 8.31 ± 0.47 × 10⁶) as compared with radiation control group on 15th day. An increase in CD4:CD8 ratio was witnessed (0.2–1%) at 30th day time interval using flow cytometry. Silymarin also countered radiation‐induced decrease (p < 0.05) in regulatory T‐cells (T_regs) (11.23% in radiation group at 7th day versus 0.1% in pretreated silymarin irradiated group at 15th day). The results of this study indicate that flavonolignan‐silymarin protects enzymatic, hematological, and immune system against γ‐radiation‐induced toxicity and might prove useful in management of nuclear and radiological emergencies. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 641–654, 2016.     

In vitro demonstration of enhanced prostate cancer toxicity: pretargeting with Bombesin bispecific complexes and targeting with polymer-drug-conjugates

Abstract

Background: Bombesin has been used to target Bombesin receptor, a growth receptor, which is over-expressed in many cancers, including prostate cancer. Polymer-anti-neoplastic-drug-conjugates (PDC) were also developed to reduce non-specific toxicity and increase tumor toxicity utilizing the enhanced permeability and retention effect, benefitting treatment of large tumors with well-established vasculature.

Purpose: If PDCs were delivered by targeted delivery to cancer cells, tumor toxicity would be enhanced and non-specific toxicity decreased.

Methods: Cardiocyte toxicity was assessed in H9c2 cardiocytes with doxorubicin (Dox) or N-terminal DTPA-modified-Doxorubicin-loaded-polyglutamic acid polymers (D-Dox-PGA). Therapeutic efficacy of targeted D-Dox-PGA after pretargeting with Bombesin-conjugated anti-DTPA-antibody Bispecific Complexes (Bom-BiSpCx) was compared to that of Dox in PC3 cells. Bom-BiSpCx was generated by thioether bond between Bombesin to Anti-DTPA antibody.

Results: D-Dox-PGA was demonstrated to have less cardiocyte toxicity (IC50?=?20?µg/ml) than free Dox (1.55?µg/ml, p?<?0.001). However, after pre-targeting of human prostate cancer PC3 cells with Bom-BiSpCx and targeting with D-Dox-PGA, IC₅₀ (13.2?µg/ml) was about two times less than that of Dox (28.5?µg/ml, p?<?0.0001).

Discussion: Targeted delivery of PDCs having lower cardiocyte toxicity enabled higher efficiency cancer cell therapy.

Conclusion: This study may allow development of very efficient targeted prostate cancer pro-drug therapy.     

Antioxidant,anti-collagenase and anti-elastase activities of Phyllanthus emblica,Manilkara zapota and silymarin: an in vitro comparative study for anti-aging applications

Context Phyllanthus emblica L. (Euphorbiaceae) (amla), Manilkara zapota L.P. Royen (Sapotaceae) (sapota) and silymarin are reported to contain antioxidant effects. However, information on other biological activities relating to the anti-aging properties is limited.

Objective To compare in vitro antioxidants, anti-collagenase (MMP-1 and MMP-2) and anti-elastase properties as well as the phenolic and flavonoid contents of amla, sapota and silymarin as potential anti-aging ingredients.

Materials and methods The ethanol amla and sapota fruit extracts were prepared by three cycles of maceration with 24 h duration each. The total phenolic (TPC) and flavonoid (TFC) contents were determined. The antioxidant capacity was evaluated by DPPH and ABTS assays. The effects of MMP-1, MMP-2 and elastase inhibitions were determined by using the EnzChek® assay kits (Molecular-Probes, Eugene, OR).

Results Amla exhibited the highest in TPC (362.43?±?11.2?mg GAE/g) while silymarin showed the highest in TFC (21.04?±?0.67?mg QE/g). Results of antioxidant activity by DPPH and ABTS methods showed that amla possessed the most potent capacity with IC₅₀ values of 1.70?±?0.07 and 4.45?±?0.10?μg/mL, respectively. Highest inhibitions against MMP-1, MMP-2 and elastase were detected for sapota with IC₅₀ values of 89.61?±?0.96, 86.47?±?3.04 and 35.73?±?0.61?μg/mL, respectively.

Discussion and conclusion Test extracts offered anti-aging properties in different mechanisms. Amla showed the highest phenolic content and antioxidant property with moderate anti-collagenase. Silymarin exhibited measurable flavonoid content with anti-elastase effect. Sapota showed the highest collagenase and elastase inhibitions with moderate antioxidant effect. Thus, extracts might be added as a mixture to gain the overall anti-aging effects.     

Habitat influence on antioxidant activity and tannin concentrations of Spondias tuberosa

Context: Different habitat conditions can be responsible for the production of secondary metabolites and for the antioxidant properties of plant products.

Objective: Thus, the aim of this study was to evaluate whether the antioxidant activity and tannin concentrations in the stem bark of Spondias tuberosa Arruda (Anacardiaceae) varied with collection site.

Material and methods: The bark was collected from 25 individual trees, distributed in five different landscape units, as follows: agroforestry gardens, areas of pastures, maize cultivation areas, mountain areas and mountain bases, with the former 3 being considered as anthropogenic habitats, and the latter 2 considered as habitats with native coverage. The study was conducted in the rural area of the city of Altinho, Pernambuco State (Northeast Brazil). The DPPH (1,1-diphenyl-2-picrylhydrazyl) method was used to measure the antioxidant activity and tannin concentrations were evaluated by using the radial diffusion method.

Results: The results demonstrated that there were no significant differences among the tannin concentrations of the individuals from the native (6.27% ± 1.75) or anthropogenic areas (4.63% ± 2.55), (H?=?2.24; p?>?0.05). In contrast, there were significant differences (H?=?5.1723; p?<?0.05) among the CE₅₀ means of the antioxidant activities of the individuals from the native (32.10 µg/ml ± 5.27) and anthropogenic areas (27.07 µg/ml ± 2.29). However, correlations between the tannin concentrations and antioxidant activity of the extracts were not observed in the native (r?=?0.39; p?>?0.05) or in the anthropogenic areas (r?=?0.38; p?>?0.05).

Discussion and conclusion: Because the variation of the antioxidant capacity of S. tuberosa bark was not accompanied by a variation in the tannin concentration, this property may be related to the presence of other metabolite(s).     

Amelioration of experimental colitis by a novel nanoselenium–silymarin mixture

Background: Silymarin has intracellular antioxidant property and inhibits activation of nuclear factor-κB (NF-κB) in low concentrations and reduces tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6 levels, cyclooxygenase (COX), and angiogenesis. Selenium is one of the necessary trace element nutrients for human and animals. Selenium nanoparticles (nano-Se) have more bioavailability with less toxicity.

Aims: To investigate the combination effect of silymarin and nano-Se on inhibition of NF-κB, proinflammatory cytokines, and oxidative stress biomarkers in the experimental colitis.

Methods: Trinitrobenzene sulfonic acid (TNBS) was used to induce colitis. After TNBS instillation, rats were distributed into six groups, containing silymarin and nano-Se alone or in combination, dexamethasone, negative control with no treatment and the last one was normal sham rats. All drugs were administered for 7 days. Colon samples were scored macroscopically and microscopically. The levels of activated NF-κB, IL-1β, TNF-α, myeloperoxidase (MPO), lipid peroxidation, protein carbonyl (PC), and the antioxidant power of the colon homogenates were determined.

Result: A significant decrease in NF-κB activity in treated groups was observed. The levels of TNF-α, IL-1β, MPO, lipid peroxidation, and PC were reduced and an improvement in antioxidant power of treated groups was seen. Combination of silymarin and nano-Se were more effective than each one alone in improvement of NF-κB, TNF-α, antioxidant power, and lipid peroxidation values, although this difference was not significant in other factors.

Conclusion: Co-administration of silymarin and nano-Se with a good antioxidant profile and inhibition of NF-κB is a possible candidate for better management of inflammatory bowel disease.     

Ethyl acetate extract from Glycosmis parva leaf induces apoptosis and cell-cycle arrest by decreasing expression of COX-2 and altering BCL-2 family gene expression in human colorectal cancer HT-29 cells

Abstract

Context: Glycosmis parva Craib (Rutaceae) is reported to have cytotoxic and anti-inflammatory activities by decreasing COX-2 expression.

Objective: To investigate the effect of G. parva on human colorectal cancer cells expressing COX-2, HT-29 cells.

Materials and methods: HT-29 cells were treated with ethyl acetate extract from the leaves of G. parva (GPE 6.25–100?µg/ml) for 24–72?h. Cell viability was evaluated by the resuzurin reduction assay. An apoptotic study was performed using annexinV/FITC-PI staining. The cell-cycle pattern was investigated by PI staining. The expression of BCL-2 family genes was analyzed by quantitative RT-PCR and expression of cyclins and COX-2 were done by RT-PCR.

Results: GPE at 6.25–100?µg/ml reduced HT-29 cell viability with IC₅₀ values of 69.49, 55.89, and 48.94?µg/ml at 24, 48, and 72?h, respectively. HT-29 apoptosis was induced by 18.23% at 100?µg/ml. Cells in S phase decreased by 5.22% and 13.28% at 50 and 100?µg/ml, respectively, causing G0/G1 (10.6% at 50?µg/ml) and G2/M (15.67% at 100?µg/ml) accumulation. GPE at 50?µg/ml downregulated cyclin A (11.46%), cyclin E (17.98%), BCL-2 (0.32-fold), and COX-2 (29.06%) expression with an increased BAK expression (1.79-fold).

Discussion and conclusion: GPE reduced HT-29 cell viability, inhibited cell proliferation, induced apoptosis, and arrested the cell cycle. Underlying mechanisms may involve decreases in COX-2, cyclin A, and cyclin E expression in addition to changes in BCL-2 family gene expression. Fundamental knowledge of GPE anticancer effects found in this study could lead to future use of this compound for colorectal cancer treatment.     

Antidiabetic potential of salvianolic acid B in multiple low-dose streptozotocin-induced diabetes

Abstract

Context: Salvianolic acids are the most abundant water-soluble compounds extracted from the herb Salvia miltiorrhiza L. (Lamiaceae) with antioxidant and protective effects.

Objective: This study evaluates the antidiabetic effect of salvianolic acid B (Sal B) in multiple low-dose streptozotocin (MLDS)-induced diabetes in rat.

Materials and methods: Rats were divided into control, Sal B40-treated control, diabetic, Sal B20-, and Sal B40-treated diabetic groups. Sal B was daily administered at doses of 20 or 40?mg/kg (i.p.), started on third day post-STZ injection for 3 weeks. Serum glucose and insulin level and some oxidative stress markers in pancreas were measured in addition to the oral glucose tolerance test (OGTT), histological assessment, and apoptosis determination.

Results: After 3 weeks, treatment of diabetic rats with Sal B20 and Sal B40 caused a significant decrease of the serum glucose (p?<?0.05–0.01) and improvement of OGTT. Meanwhile, serum insulin was significantly higher in Sal B20- and Sal B40-treated diabetics (p?<?0.01) and treatment of diabetics with Sal B40 significantly lowered malondialdehyde (MDA) (p?<?0.05), raised glutathione (GSH) (p?<?0.05), and activity of catalase (p?<?0.01) with no significant change of nitrite. Furthermore, the number of pancreatic islets (p?<?0.05) and their area (p?<?0.01) was significantly higher and apoptosis reactivity was significantly lower (p?<?0.05) in the Sal B40-treated diabetic group versus diabetics.

Discussion and conclusion: Three-week treatment of diabetic rats with Sal B exhibited antidiabetic activity which is partly exerted via attenuation of oxidative stress and apoptosis and augmentation of antioxidant system.