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.     

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.     

Schizandra chinensis extracts induce apoptosis in human gastric cancer cells via JNK/p38 MAPK activation and the ROS-mediated/mitochondria-dependent pathway

Abstract

Context: Schizandra chinensis Baill (Magnoliaceae) fruit extract (SCE) is considered a traditional herbal medicine for the treatment and alleviation of various diseases. Gastric cancer is the second most common cause of cancer-related death worldwide, and the first most common in Korea.

Objectives: This study investigates the mechanism of SCE-induced apoptosis in AGS human gastric cancer cells.

Materials and methods: SCE concentrations from 100 to 400?µg/ml were used. Cell viabilities were determined using MTT assay. Members of the Bcl-2 family and Bax were detected by Western blotting. RT-PCR was performed to measure the expression level of the Fas/FasL pro-apoptotic genes.

Results: SCE inhibited the proliferation AGS cells for 24 or 72?h (inhibition by 3.1%?±?5.2% at 100?µg/ml and 87.3%?±?7.6% at 400?µg/ml at 24?h and by 40.2%?±?5.3% 100?µg/ml and 95.3%?±?1.3% 400?µg/ml at 72?h) and increased the sub-G1 phase (25.3%?±?5.2% at 100?µg/ml and 370.2%?±?7.2% at 400?µg/ml) and the mitochondrial membrane depolarization (11.2%?±?2.1% at 100?µg/ml and 311.5%?±?6.1% at 400?µg/ml). The SCE-induced apoptotic cell death showed the down-regulation of Bcl-2, but up-regulation of Bax. Subsequently, SCE increased the expression level of Fas/FasL, activated caspase-9 and -3, and increased reactive oxygen species generation. Also, JNK II inhibitor or a p38 MAPK inhibitor inhibited SCE-induced cell death.

Discussion and conclusion: These results indicate that SCE might be an effective chemotherapeutic for the treatment of human gastric cancer.     

Antioxidant activity and cytotoxicity of Cyrtosperma johnstonii extracts on drug sensitive and resistant leukemia and small cell lung carcinoma cells

Context: The number of patients with cancer is increasing. New therapeutic agents to overcome drug-resistant tumors are urgently needed. Cyrtosperma johnstonii N.E. Br. (Araceae) is used for treatment of cancer in Thai traditional medicine. This study aimed to evaluate antioxidant activity and cytotoxicity of C. johnstonii extracts on human cancer cells.

Materials and methods: Dried powder of C. johnstonii rhizomes was extracted with several solvents. The 0.1?mg/ml extract solution was tested for antioxidant activity by 2,2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and ferric reducing antioxidant power (FRAP) assays. Color formation from 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide was used to determine cell viability. Standardization of the extract was performed by high-performance liquid chromatography (HPLC) with photodiode array detector at 254 and 360?nm. Cell cycle arrest was evaluated by flow cytometry after 5?min, 12?h and 24?h treated with 20 µg/ml of the acetone extract.

Results: The acetone extract exhibited the highest phenolic content and antioxidant activity (TEAC and EC values = 19.2?±?0.14 and 19.2?±?0.31?mM/mg, respectively). The IC₅₀ values for leukemia ranged from 11?±?1 to 29?±?3 µg/ml and from 5?±?2 to 6?±?0 µg/ml for small cell lung carcinoma cells. Cell cycle arrest occurred at the G2/M phase followed by apoptosis. HPLC analysis revealed that rutin is the major constituents of the extract.

Discussion and conclusion: The acetone extract of C. johnstoni is a promising source of natural antioxidants and anticancer. The extract inhibits cancer cells effectively with less effect on normal cells.     

Synergistic antimicrobial profiling of violacein with commercial antibiotics against pathogenic micro-organisms

Context: Chromobacterium violaceum Bergonzini (Neisseriaceae), a Gram-negative bacterium, secretes a spectacular pigment called violacein. Violacein is a quorum-sensing metabolite and is also an active antimicrobial, anticancer agent. However, its efficiency as a potential drug, alone or in synergy with other active principles, has not being completely exploited. With the advent of different multi-drug resistant strains, it becomes essential to find a new natural product(s) that could be effectively used as a therapeutic agent.

Objective: This work focused on the extraction of violacein from an isolated strain of C. violaceum and determined the combinatory effect of violacein with commercial antibiotics against various pathogens.

Materials and methods: Violacein production was optimized and was later extracted using ethanol and characterized by liquid chromatography-mass spectrometry and infrared spectroscopy. Then, individual minimum inhibitory concentration (MIC) values for each of the antibiotics were determined followed by violacein–commercial antibiotics (1:1) combinations, tested at different concentrations starting from 500 to 1?µg/ml against major pathogens.

Results and discussion: The individual MIC data for violacein was found to be 5.7?µg/ml (Staphylococcus aureus), 15.6?µg/ml (Klebsiella pneumoniae), 18.5?µg/ml (Pseudomonas aeruginosa), 20.0?µg/ml (Vibrio cholerae) and 5.7?µg/ml (Salmonella typhi). Violacein–gentamicin and violacein-cefadroxil combinations had MIC of 1.0?µg/ml against S. aureus. Most violacein–macrolide and violacein--aminoglycoside class combinations revealed fractional inhibitory concentration indices (FICI) of <0.5, thus exhibiting synergism. Furthermore, violacein–azithromycin and violacein–kanamycin combination, exhibited significant synergy (FICI-0.3) against S. typhi.

Conclusion: Violacein works synergistically with most commercial antibiotics and could be used as drug in combination with other antimicrobial agents.     

Magnolia dealbata seeds extract exert cytotoxic and chemopreventive effects on MDA-MB231 breast cancer cells

Context: Cancer prevention remains a high priority for the scientific world. Magnolia dealbata Zucc (Magnoliaceae), a Mexican endemic species, is used for the empirical treatment of cancer.

Objective: To evaluate the cytotoxic and cancer chemopreventive effects of an ethanol extract of Magnolia dealbata seeds (MDE).

Materials and methods: The cytotoxic effect of MDE, at concentrations ranging from 1 to 200?µg/ml, on human cancer cells and human nontumorigenic cells was evaluated using the MTT assay for 48?h. The apoptotic activities of MDE 25?μg/ml on MDA-MB231 breast cancer cells were evaluated using the TUNEL assay and the detection of caspase 3 using immunofluorescence analysis for 48?h, each. The chemopreventive effect was evaluated by administrating different doses of MDE, between 1 and 50?mg/kg, injected intraperitoneally daily into athymic mice which were implanted with MDA-MB231 cells during 28 days. The growth and weight of tumors were measured.

Results: MDE showed cytotoxic effects on MDA-MB231 cells (IC₅₀?=?25?µg/ml) and exerted pro-apoptotic activities as determined by DNA fragmentation in MDA-MB231 cells. MDE 25?µg/ml also induces the activation of caspase 3 in MDA-MB231 cells. These results suggest that Magnolia dealbata may be an optimal source of the bioactive compounds: honokiol (HK) and magnolol (MG). MDE 50?mg/kg i.p. exerted chemopreventive effects by inhibiting the growth of MDA-MB231 tumor by 75% in athymic mice, compared to the control group.

Conclusions: MDE exerts cytotoxic, apoptotic and chemopreventive activities on MDA-MB231 human cancer cells.     

Cytotoxic effects of Eryngium kotschyi and Eryngium maritimum on Hep2, HepG2, Vero and U138 MG cell lines

Abstract

Context: Eryngium maritimum L. and the endemic Eryngium kotschyi Boiss. of the Apiaceae family are used for antiinflammatory, antivenom, antinociceptive and diuretic purposes in folk medicine in Turkey.

Objective: This study investigated the cytotoxic effects of the plant extracts belonging to Eryngium L. genus on various cell lines.

Materials and methods: Cytotoxic activites of the lyophilized aqueous aereal and root parts of the plant extracts on human hepatocellular carcinoma (HepG2), human laryngeal epidermoid carcinoma (Hep2), human glioma (U138-MG) and African green monkey kidney epithelial (Vero) cell lines at 8.33–266.62?µg/ml concentrations were analyzed by lactate dehydrogenase (LDH) leakage and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) (MTT) cell viability assays.

Results: Inhibitory concentration 50 (IC₅₀) values were found <100?µg/ml in most cases varying around 16.33–125.66?µg/ml. IC₅₀ values for E. kostchyi and E. maritimum root parts on Hep2 cells (32.86 and 30.25?µg/ml, respectively), E. kotschyi aereal, E. maritimum aereal and root parts on HepG2 cells (31.75, 32.42 and 35.01?µg/ml, respectively) by MTT assay were found to be close to the US National Cancer Institute (NCI) recommendations (IC₅₀?<?30?µg/ml) to define the antivity aganist cancer cells. The lowest IC₅₀ values according to the LDH method were observed in Hep2 cells and the highest in U138-MG cells. Root parts were found to be more toxic than aereal parts for both plants in both methods in general.

Discussion and conclusion: Both plant extracts exerted cytotoxic activity aganist Hep2 and HepG2 cells, with low IC₅₀ values defining their promising anticancer property according to NCI; however, further analysis are needed to confirm their activity.     

Asiatic acid exerts anticancer potential in human ovarian cancer cells via suppression of PI3K/Akt/mTOR signalling

Context Asiatic acid, a triterpenoid compound extracted from the tropical medicinal plant Centella asiatica (Family: Apiaceae), has exhibited various biological activities.

Objective This study was performed to investigate the cytotoxic effects of asiatic acid on human ovarian cancer cells.

Materials and methods SKOV3 and OVCAR-3 ovarian cancer cells were exposed to different concentrations of asiatic acid (10–100?μg/mL) for 72 or 48?h. Cell viability, colony formation, cell cycle distribution, apoptotic response were examined. Involvement of the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway was tested.

Results At the concentration of 40?μg/mL, asiatic acid caused about 50% reduction in the viability of ovarian cancer cells, but had little effect on the viability of normal human ovarian epithelial cells. Asiatic acid at 10?μg/mL reduced colony formation of ovarian cancer cells by 25–30%. Asiatic acid-treated cells showed a cell cycle arrest at the G0/G1 phase and 7- to 10-fold increase in apoptosis. The phosphorylation levels of PI3K, Akt and mTOR were remarkably lower in asiatic acid-treated cells. Overexpression of constitutively active Akt partially reversed the cytotoxic effects of asiatic acid, as evidenced by increased cell viability and colony formation. Furthermore, knockdown of Akt mimicked the growth-suppressive activity of asiatic acid.

Discussion and conclusion These results provide first the evidence for the anticancer potential of asiatic acid in ovarian cancer cells, partially via inactivation of the PI3K/Akt/mTOR pathway. Asiatic acid may represent a potential therapeutic agent for ovarian cancer.     

Novel anticancer alkene lactone from Persea americana

Abstract

Context: Persea americana Mill (Lauraceae) root bark is used in ethnomedicine for a variety of diseases including cancer.

Objective: To isolate and characterize the chemical constituent in P. americana, and also to determine the anticancer property of a new alkene lactone from the root bark of P. americana.

Materials and methods: The MCF-7 cells were treated with different concentrations of the pure compound for 48?h. The percentage of cells in the various phases, online monitoring of metabolic changes and integrin receptor expression determined by flow cytometry.

Results: One novel alkene lactone (4-hydroxy-5-methylene-3-undecyclidenedihydrofuran-2 (3H)-one) (1) was isolated and characterized using 1D-NMR, 2D-NMR, infrared, UV and MS. At a concentration of 10?µg/mL, significant reduction of proliferation of MCF-7 was induced while MCF-12?A cell was significantly stimulated by 10?µg/mL. The IC₅₀ value for MCF-7 cells is 20.48?µg/mL. Lower concentration of 1 harbor no significant effect on either MCF-7 or MCF-12A. The apoptotic rates of MCF-7 cells were increased significantly. At the final concentration 10?µg/mL, up to 80% of all breast cancer cells were dead. On the non-tumorigenic cell line MCF-12A, the same concentrations (1 and 10?µg/mL) of compound 1 caused significant enhanced apoptotic rates. A total of 1?µg/mL of 1 caused a decrease of α4-, α6-, β1- and β3-integrin expression.

Conclusions: The compound caused a stimulatory effect on non-tumorigenic MCF-12A cells with respect to cell adhesion while tumorigenic MCF-7 cells detached continuously. This is the first report on the anticancer effects of this class of compound.     

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.     

Assessment of bendamustine-induced genotoxicity in eukaryotic cells

Bendamustine, an anticancer drug with alkylating properties, is widely used to treat hematological malignancies. Since the nitrogen mustard family alkylators induce DNA damages and have been associated with an elevated risk of second malignancy, current study evaluates the cytotoxic, mutagenic, and recombinogenic effects of bendamustine by using, respectively the mitotic index assay, the in vitro mammalian cell micronucleus test (Mnvit) and the chromosome aberration (CA) test in human peripheral lymphocytes, and the in vivo homozygotization assay in Aspergillus nidulans, which detects the loss of heterozygosity (LOH) due to somatic recombination. Bendamustine (6.0?µg/ml, 9.0?µg/ml, and 12.0?µg/ml) induced a statistically significant concentration-related increase in the frequencies of micronuclei and a significant reduction in the cytokinesis block proliferation index (CBPI) rates when compared to negative control. In the CA test, bendamustine significantly increased the frequencies of structural aberrations at the three tested concentrations when compared to the negative control. Aspergillus nidulans diploids, obtained after bendamustine treatment (6.0?µg/ml, 12.0?µg/ml, and 24.0?µg/ml), produced, after haploidization, homozygotization index (HI) rates higher than 2.0 and significantly different from the negative control. Since bendamustine showed genotoxic effects in all tested concentrations, two of them corresponding to the peak plasma concentrations observed in cancer patients treated with bendamustine, data provided in the current research work may be useful to identify the most appropriate dosage regimen to achieve the efficacy and safety of this anticancer medication.     

Terpenoids of methanol extract of Clerodendrum infortunatum exhibit anticancer activity against Ehrlich’s ascites carcinoma (EAC) in mice

Context: Clerodendrum infortunatum Linn. is a widely used plant in the Indian indigenous system of medicine for the treatment of tumors.

Objective: The present study evaluated the anticancer activity of methanol extract of C. infortunatum (MECI) against Ehrlich’s ascites carcinoma (EAC) bearing Swiss albino mice and isolation of bioactive terpenoids from it.

Methods: HPLC analysis of the methanol extract showed the presence of three major components. Out of those, two compounds were isolated and characterized as oleanolic acid and clerodinin A. The anticancer activity of MECI was assessed by measuring the tumor growth response, percentage increase of life span, study of hematological parameters, lipid peroxidation, antioxidant enzyme activity like glutathion and CAT. In vitro cytotoxicity assay was also performed using EAC cell lines.

Result and conclusion: Treatment with MECI causes significant decrease in the tumor cell volume and increase in the life span. The median survival time (MST) of EAC control group was found as 19.42?±?0.91 d, whereas the MST was increased to 23.44?±?2.69 d and 27.57?±?2.57 d for the groups treated with MECI at 100 and 200?mg/kg, respectively. All the hematological parameters, malonaldehyde content and antioxidant enzymes’ activity were restored towards the normal level. IC₅₀ value of MECI was found as 498.33 µg/mL in cytotoxicity study. The experimental results suggested that MECI has significant anticancer activity, which can be attributed to the presence of oleanolic acid and clerodinin A.     

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.     

In vitro inhibitory effects of thymol and carvacrol on dendritic cell activation and function

Context: Thyme has been used in traditional medicine for medicinal purposes since ancient times.

Objective: The objective of this study was to investigate the effects of thymol and carvacrol as two major constituents of thyme on dendritic cells (DCs) maturation and T cell activation.

Materials and methods: Splenic DCs were treated with non-cytotoxic concentrations of the components and then analyzed for MHC II, CD86, and CD40 expression by flow cytometry. The effects of compounds on mitogenic, as well as allogenic T cell responses in mixed lymphocyte culture (MLR) and the release of cytokines were investigated.

Results: At 0.1?µg/ml, reduced mean fluorescent intensity (MFI) of CD86 for thymol (80.3?±?0.2% of untreated control) and CD40 for carvacrol (79.5?±?0.14%) was observed (p?<?0.001). Decreased mitogenic T cell proliferation by thymol [proliferation index (PI) from 0.93?±?0.11 at 1?µg/ml to 0.42?±?0.16 at 100?µg/ml (p?<?0.01)] and carvacrol [PI from 1.08?±?0.3 at 1?µg/ml to 0.28?±?0.1 at 100?µg/ml (p?<?0.001)] was seen. Ten micrograms/ml thymol (PI, 0.85?±?0.04) and carvacrol (PI, 0.89?±?0.03) inhibited allogenic T cell response (p?<?0.05). Decreased IFN-γ level in MLR supernatant from 1441?±?27.7?pg/ml in untreated cells to 944?±?32.1 at 10?µg/ml of thymol and of carvacrol (886?±?31.7?pg/ml) (p?<?0.01) was found. IL-4 levels were decreased in the presence of both compounds (p?<?0.01).

Conclusion: These data showed the suppressive effects of thymol and carvacrol on DCs maturation and function, as well as T cell responses.     

Bioactivity-guided isolation of cytotoxic constituents from three medicinal plants

Abstract

Context: The ethanol extracts and their fractions of three Indian medicinal plants, Ervatamia coronaria (Jacq.) Stapf, (Apocynaceae), Mimosa pudica L. (Mimosaceae) and Caesalpinia bonduc (L.) Roxb. (Caesalpiniaceae) were tested for their cytotoxic activity in the brine shrimp lethality (BSL) bioassay and in various cancer cell lines. The plants were selected based on their traditional use in the treatment of cancer/tumors.

Objectives: To investigate the in vitro cytotoxicity of Ervatamia coronaria, Mimosa pudica and Caesalpinia bonduc.

Materials and methods: Ethanolic extracts and their fractions of E. coronaria, M. pudica and C. bonduc were subjected to cytotoxicity studies using BSL bioassay method with concentrations of 10, 50, 100, 500 and 1000?µg/ml. The alkaloid fraction of E. coronaria with significant cytotoxicity in BSL bioassay was subjected to in vitro cytotoxicity studies with HT-29, A-549, HepG-2, MCF-7 and L-6 cell lines at concentrations of 12.5, 25, 50, 100 and 200?µg/ml and a DNA fragmentation study using the HT-29 cell line.

Results: The alkaloid fractions of E. coronaria and M. pudica showed significant cytotoxicity with LC₅₀ values of 65.83 and 85.10?µg/ml in the BSL bioassay, respectively. The purified alkaloid fraction of E. coronaria exhibited highest cytotoxicity in HT-29, A-549 and MCF-7 cell lines with IC₅₀ values of 32.5, 47.5 and 72.5?µg/ml, respectively, and induced DNA fragmentation in the HT-29 cell line at a concentration of 65?µg/ml.

Conclusion: The alkaloid fraction of E. coronaria exhibited significant cytotoxicity. Alkaloids such as ervatamine, apparicine and coronaridine that were earlier reported may be responsible for this activity.     

Antioxidant,antimicrobial and wound healing properties of Struthanthus vulgaris

Context: Struthanthus vulgaris (Vell.) Mart. (Loranthaceae) has been widely used in traditional medicine in Brazil to bathe wounds.

Objective: The objective of this study is to investigate the in vitro wound healing effects, together with the antioxidant and antimicrobial activities of S. vulgaris leaf and branch extracts.

Material and methods: Ethanol leaf and branch extracts of S. vulgaris were investigated at 1–100?µg/ml concentrations in the scratch assay after 14?h. Antioxidant activity was investigated using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical-scavenging assay, and the antibacterial activity was tested at concentrations up to 1000?µg/ml against Gram-positive and Gram-negative bacteria by the microdilution test after 24?h. The total phenolic and flavonoid contents were determined by colorimetric methods.

Results: Struthanthus vulgaris leaf and branch extracts at 100?µg/ml concentration stimulated migration and proliferation of fibroblasts and enhanced cell numbers by 56.2% and 18.6%, respectively. Antioxidant activity exhibited IC₅₀ values of 24.3 and 18.9?µg/ml for the leaf and branch extracts, respectively. The ethanol leaf extract showed antimicrobial activity against the Gram-positive Staphylococcus mutans and Staphylococcus aureus bacteria, exhibiting minimum inhibitory concentration values of 125 and 500?µg/ml, respectively. An appreciable total phenolic content in the leaves (813.6?±?2.7?mg/g) and branches (462.8?±?9.6?mg/g), and relatively low concentration of flavonoids in the leaves (13.3?±?4.3?mg/g) and branches (1.9?±?0.2?mg/g), was detected.

Discussion and conclusion: The antioxidant and antibacterial activities, together with the strong ability to stimulate proliferation and migration of fibroblasts, provide some support for the traditional use of S. vulgaris.     

In vitro antioxidant and anti-cholinesterase activities of Rhizophora mucronata

Context: Rhizophora mucronata Lam. (Rhizophoraceae), commonly known as Asiatic mangrove, has been used traditionally among Asian countries as folk medicine.

Objective: This study investigates the cholinesterase inhibitory potential and antioxidant activities of R. mucronata.

Materials and method: Rhizophora mucronata leaves were successively extracted using solvents of varying polarity and a dosage of 100–500?µg/ml were used for each assay. Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibitory activities were assessed according to the method of Ellman. In vitro antioxidant activity was assessed using free radical scavenging, reducing power, and metal-chelating activity (duration – 3 months). Total phenolic and flavonoid content were quantified spectrophotometrically. Compound characterization was done using column chromatography, NMR, FTIR, and LC-MS analysis.

Results: Methanolic leaf extract (500?µg/ml) exhibited the highest inhibitory activity against AChE (92.73?±?0.54%) and BuChE (98.98?±?0.17%), with an IC₅₀ value of 59.31?±?0.35 and 51.72?±?0.33?µg/ml, respectively. Among the different solvent extracts, methanolic extract exhibited the highest antioxidant activity with an IC₅₀ value of 47.39?±?0.43, 401.45?±?18.52, 80.23?±?0.70, and 316.47?±?3.56?µg/ml for DPPH, hydroxyl, nitric oxide radical, and hydrogen peroxide, respectively. Total polyphenolic and flavonoid contents in methanolic extract were observed to be 598.13?±?1.85?µg of gallic acid equivalent and 48.85?±?0.70?μg of rutin equivalent/mg of extract. Compound characterization illustrated (+)-catechin as the bioactive compound responsible for cholinesterase inhibitory and antioxidant activities.

Conclusion: The presence of rich source of flavonoids, in particular catechin, might be responsible for its cholinesterase inhibitory and antioxidant activities.     

Chemical profile and biological activities of Veronica thymoides subsp. pseudocinerea

Abstract

Context: In Turkey, Veronica species (Plantaginaceae) have been used as a diuretic and for wound healing in traditional medicine.

Objective: To examine the fatty acid and essential oil profiles, the antioxidant, anticholinesterase, antimicrobial, and DNA damage effects of Veronica thymoides P.H. Davis subsp. pseudocinerea M.A. Fischer as a potential source of natural active compounds.

Materials and methods: GC/MS was used to analyze essential oil and fatty acid obtained from whole plant. The antioxidant activity was evaluated by the β-carotene-linoleic acid test system, DPPH-free and ABTS cation radicals scavenging, and cupric reducing antioxidant capacity assays. The anticholinesterase and antimicrobial activities were determined by Ellman and broth macrodillution methods, respectively. The effect of the methanol extract on DNA cleavage was investigated.

Results: Hexatriacontene (21.0%) was found to be the main constituent in essential oil, and linoleic acid (25.2%) and palmitic acid (20.6%) in fatty acid. Methanol extract demonstrated the best IC₅₀ values in lipid peroxidation (49.81?±?0.31?µg/ml) and DPPH-free radical scavenging activity (15.32?±?0.17?µg/ml). Methanol and water extracts possessed strong ABTS cation radical scavenging activity with IC₅₀ values 9.15?±?0.28 and 8.90?±?0.14?µg/ml, respectively. The acetone extract exhibited moderate butyrylcholinesterase inhibitory activity. The highest antimicrobial activity was determined in methanol extract against Escherichia coli with 31.25?µg/ml MIC value. Inhibition of methanol extract on plasmid DNA cleavage by OH radicals was found to be 93.32% at 500?µg/ml.

Conclusion: The methanol extract having strong antioxidant and DNA damage effects could be investigated phytochemically to find natural active compounds.     

Effect of Musca domestic maggot polypeptide extract on HUVEC dysfunction induced by early-activated macrophages

Context: Musca domestica Linn. maggot is a traditional Chinese medicine. In our previous studies, Musca domestica maggot protein-enriched fraction and polypeptide extract (molecular weight <30?kD) were found to reverse endothelial cell dysfunction in atherosclerotic lesions.

Objective: This study determines whether and how M. domestica maggot polypeptide extract affects the dysfunction of human umbilical vein endothelial cells (HUVEC) induced by macrophages (M?).

Materials and methods: HUVEC and early-activated THP-1?M? (incubated with LPS of 1?μg/ml for 2?h) were co-cultured in a Transwell system. The effects of Musca domestica maggot polypeptide extract (with increasing concentrations, i.e., 1.0, 2.5, 5.0, 10.0, 20.0, and 40.0?µg/ml) on the proliferation and migration HUVEC and their secretion of vascular endothelial growth factor (VEGF) were determined by flow cytometry, modified Boyden chamber assay, and enzyme-linked immunosorbent assay (ELISA) after incubation for 24?h.

Results: Musca domestica maggot polypeptide extract decreased the proliferation of HUVEC in a concentration-dependent manner, with a 50% effective concentration (EC₅₀) of 22.16?±?1.48?µg/ml, and effectively inhibited HUVEC migration (EC₅₀?=?35.15?±?2.03?µg/ml) and VEGF secretion (EC₅₀?=?28.64?±?1.29?µg/ml).

Discussion and conclusion: Musca domestica maggot polypeptide extract can inhibit the dysfunction of HUVEC induced by early-activated THP-1?M?.     

Anticancer activity of Saussurea lappa extract by apoptotic pathway in KB human oral cancer cells

Abstract

Context: Saussurea lappa Dence (Compositae) is used as a traditional herbal medicine to treat abdominal pain and tenesmus in East Asia. Current studies have shown that S. lappa has anticancer activity in divergent of cancer cells. However, the effects of S. lappa on oral cancer and its mechanisms of action have yet to be elucidated.

Objective: To explore its potential chemotherapeutic effects and mechanism of cell growth inhibition on human oral cancer cells.

Materials and methods: The dried roots of S. lappa were used in this study. Cell viability of KB cells was evaluated by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay after treatment with 30?µg/ml of methanol extract from the dried roots of S. lappa. To understand whether its effect on cell death is related with apoptosis pathway, we performed DNA fragmentation assay, western blot, caspase activity assay and fluorescence-activated cell sorting (FACS) analysis.

Results: Treatment of S. lappa extract onto KB cells reduced cell viability significantly with an IC₅₀ value of 30?µg/ml. The formation of a DNA ladder was observed starting at the 24?h treatment. In western blotting analysis, the S. lappa extract induced the proteolytic processing of caspase-3, -9 and poly (ADP-ribose) polymerase, a significant increase of Bax and marked reduction of Bcl-2. We also confirmed the activation of caspase-3/-7 in living KB cells by fluorescence microscopy.

Conclusion: These results suggested that S. lappa extract inhibited cell proliferation through the apoptosis pathway in KB human oral cancer cells.