Chrysin is a natural flavonoid currently under investigation due to its important biological anti-cancer properties. In most of the cancer cells tested, chrysin has shown to inhibit proliferation and induce apoptosis, and is more potent than other tested flavonoids in leukemia cells, where chrysin is likely to act via activation of caspases and inactivation of Akt signaling in the cells. Moreover, structure-activity relationships have revealed that the chemical structure of chrysin meets the key structural requirements of flavonoids for potent cytotoxicity in leukemia cells. It is possible that combination therapy or modified chrysin could be more potent than single-agent use or administration of unmodified chrysin. This study may help to develop ways of improving the effectiveness of chrysin in the treatment of leukemia and other human cancers in vitro.
Acyclic nucleosides have been of considerable interest since the approval of aciclovir by the FDA to be used as an antiviral agent in the 1990s. The acyclic moieties and the bases used in the experiment were either available commercially or synthesized using literature methods. Vorbruggen coupling method was utilized involving reaction of persilylated heterocyclic bases with the appropriate acyclic moiety in the presence of a Lewis acid catalyst. A series of novel 6-azapyrimidine acyclic oxosugar nucleosides was successfully synthesized with a promising yield (more than 50%). An efficient method of protection and deprotection was also investigated.
Phyllanthus is a traditional medicinal plant that has been used in the treatment of many diseases including hepatitis and diabetes. The main aim of the present work was to investigate the potential cytotoxic effects of aqueous and methanolic extracts of four Phyllanthus species (P.amarus, P.niruri, P.urinaria and P.watsonii) against skin melanoma and prostate cancer cells.
A challenge for studies involving microglia cultures is obtaining sufficient cells for downstream experiments. Macrophage colony-stimulating factor (M-CSF) has been used to improve yield of microglia in culture. However, the effects of M-CSF on activation profiles of microglia cultures are still unclear. Microglia activation is characterised by upregulation of co-stimulatory molecules and an inflammatory phenotype. The aim of this study is to demonstrate whether M-CSF supplementation alters microglial responses in resting and activated conditions. Microglia derived from mixed glia cultures and the BV-2 microglia cell line were cultivated with/without M-CSF and activated with lipopolysaccharide (LPS) and beta amyloid (Abeta). We show M-CSF expands primary microglia without affecting microglial responses to LPS and Abeta, as shown by the comparable expression of MHC class II and CD40 to microglia grown without this growth factor. M-CSF supplementation in BV-2 cells had no effect on nitric oxide (NO) production. Therefore, M-CSF can be considered for improving microglia yield in culture without introducing activation artefacts.
In a preliminary screen, Aaptos aaptos showed significant cytotoxic activity towards a panel of cell lines and was thus subjected to bioassay-guided isolation of the bioactive constituents. In addition to the known aaptamine, two new derivatives of the alkaloid were isolated from the bioactive chloroform fraction of the crude methanolic extract. Detailed analysis by NMR and mass spectroscopy enabled their identification to be 3-(phenethylamino)demethyl(oxy)aaptamine and 3-(isopentylamino)demethyl(oxy) aaptamine. The cytotoxic activities of the three alkaloids were further evaluated against CEM-SS cells.
Dendritic cells (DCs) have the potential for cancer immunotherapy due to their ability to process and present antigens to T-cells and also in stimulating immune responses. However, DC-based vaccines have only exhibited minimal effectiveness against established tumours in mice and humans. The use of appropriate adjuvant enhances the efficacy of DC based cancer vaccines in treating tumours.
Conventional cytotoxic anticancer drugs that target all rapidly dividing cells are nonselective in their mechanism of action, because they disrupt essential components that are crucial to both malignant and proliferating normal cells. Instead, targeting cellular functions that are distinctly different between normal and cancer cells may provide a basis for selective killing of tumor cells. One such strategy that is still largely unexplored is to utilize the relatively higher negative mitochondrial membrane potential in carcinoma cells compared with adjacent normal epithelial cells to enhance accumulation and retention of cytotoxic lipophilic cations in the former. In this study, the anticancer activities of a new class of rosamines with cyclic amine substituents and their structure-activity relationships were investigated. From an in-vitro cell growth inhibition assay, 14 of the rosamines inhibited the growth of human leukemia HL-60 cells by 50% at micromolar or lower concentrations. Derivatives containing hydrophilic substituents had less potent activity, whereas aryl substitution at the meso position conferred extra activity with thiofuran and para-iodo aryl substitutions being the most potent. In addition, both compounds were at least 10-fold more cytotoxic than rhodamine 123 against a panel of cell lines of different tissue origin and similar to rhodamine 123, exhibited more cytotoxicity against cancer cells compared with immortalized normal epithelial cells of the same organ type. In subsequent experiments, the para-iodo aryl substituted rosamine was found to localize exclusively within the mitochondria and induced apoptosis as the major mode of cell death. Our results suggest that these compounds offer potential for the design of mitochondria-targeting agents that either directly kill or deliver cytotoxic drugs to selectively kill cancer cells.
Our current interest in searching for natural anti-cancer lead compounds from plants has led us to the discovery that the stem and roots of Garcinia mangostana can be a source of such compounds. The stem furnished 2,8-dihydroxy-6-methoxy-5-(3-methylbut-2-enyl)-xanthone (1), which is a new xanthone. Meanwhile, the root bark of the plant furnished six xanthones, namely alpha-mangostin (2), beta-mangostin (3), gamma-mangostin (4), garcinone D (5), mangostanol (6), and gartanin (7). The hexane and chloroform extracts of the root bark of G. mangostana as well as the hexane extract of the stem bark were found to be active against the CEM-SS cell line. gamma-Mangostin (4) showed good activity with a very low IC(50) value of 4.7 microg/ml, while alpha-mangostin (2), mangostanol (6), and garcinone D (5) showed significant activities with IC(50) values of 5.5, 9.6, and 3.2 microg/ml, respectively. This is the first report on the cytotoxicity of the extracts of the stem and root bark of G. mangostana and of alpha-mangostin, mangostanol, and garcinone D against the CEM-SS cell line.
Tioman virus (TioPV) and Menangle virus (MenPV) are two antigenically and genetically related paramyxoviruses (genus: Rubulavirus, family: Paramyxoviridae) isolated from Peninsular Malaysia (2001) and Australia (1997), respectively. Both viruses are potential zoonotic agents. In the present study, the infectivity, growth kinetics, morphology and morphogenesis of these two paramyxoviruses in a human neuronal cell (SK-N-SH) line were investigated. Sub-confluent SK-N-SH cells were infected with TioPV and MenPV at similar multiplicity of infection. These cells were examined by conventional and immunoelectron microscopy, and virus titres in the supernatants were assayed. Syncytia were observed for both infections in SK-N-SH cells and were more pronounced during the early stages of TioPV infection. The TioPV titre increased consistently (10(1)) every 12 h after infection. In MenPV-infected cells, cellular material was frequently observed within budding virions, and microfilaments and microtubules were abundant. Viral budding was common, and extracellular MenPVs tended to be more pleomorphic compared to TioPVs, which appeared to be more spherical in appearance. The MenPV cytoplasmic viral inclusion appeared to be comparatively smaller, loose and interspersed with randomly scattered circle-like particles, whereas huge tubule-like cytoplasmic inclusions were observed in TioPV-infected cells. Both viruses also displayed different cellular pathology in the SK-N-SH cells. The intracellular ultrastructural characteristics of these two viruses in infected neuronal cells may allow them to be differentiated by electron microscopy.
The effect of mild hypothermic (32 degrees C) conditions on cell growth, cell-cycle progress, and antibody production of hybridoma C2E7 cells was investigated in the present study. The growth of hybridoma cells was slower during the mild hypothermic condition compared to that at 37 degrees C; this led to about 10% decrease in maximum viable cell density and volumetric antibody productivity. However, under mild hypothermic growth conditions, the culture viability was substantially improved and the specific antibody productivity was enhanced compared to that at 37 degrees C. The average specific productivity for the entire batch culture at 32 degrees C is about 5% higher than that at 37 degrees C. Cell-cycle analysis data showed that there was no growth arrestment during the mild hypothermic growth of hybridoma cells. The G1-phase cells were increased, while the S-phase cells were decreased gradually as the culture time progressed. Further analysis showed that the specific antibody productivity of hybridoma cells was correlated to the fraction of S-phase cells.
Photodynamic therapy (PDT) is a promising cancer treatment which involves activation of a photosensitizing drug with light to produce reactive oxygen species that kill tumors without causing damage to unirradiated normal tissues. To date, only Photofrin, Foscan and Levulan have been approved for clinical treatment of cancer. Tropical habitats such as those found in Malaysia are attractive sources of new therapeutic compounds as tremendous chemical diversity is found in a large number of plants, animals, marine- and micro-organisms. In our screening program for novel photosensitizers from nature, colorful strains of fungi (from Aspergillus and Penicillium genus) and bacteria (including actinomycetes and photosynthetic bacteria) were collected from various habitats in Peninsular Malaysia, such as coastal soil, peat soil, marine sponges and wastewater ponds. Methanolic extracts from a total of 85 different species were evaluated with a short-term cell viability assay for photo-cytotoxicity, where a promyelocytic leukemia cell-line, HL60 incubated with 20 microg/ml of extracts was irradiated with 9.6 J/cm(2) of a broad spectrum light. Two of these extracts, one from Rhodobacter sphaeroides (PBUM003) and one from Rhodopseudomonas palustris (PBUM001) showed moderate to strong photo-cytotoxicity. Subsequent bioassay guided isolation of the PBUM001 extract yielded known photosensitisers that are based on bacteriochlorophyll-a by comparing their molecular weight data, HPLC profiles and UV-vis absorption spectra with literature values, thereby demonstrating the validity of our screening approach.
Polyhydroxyalkanoates (PHA) are naturally occurring biopolyesters that have great potential in the medical field. However, the leachables resulting from sterilization process of the biomaterials may exert toxic effect including genetic damage. Here, we demonstrate that although gamma-irradiation of poly(3-hydroxybutyrate-co-50 mol % 4-hydroxybutyrate) [P(3HB-co-4HB)] did not cause any change in the morphology by scanning electron microscopy, there was a significant degradation of this copolymer where the molecular weight was reduced by 37% after sterilization indicating the generation of leachables. Therefore, further investigation on the ability of the extract of this poststerilized copolymer to induce mutagenic effect was performed using Ames test (S. typhimurium strains TA1535 and TA1537) and umu test (S. typhimurium strain TA1535/pSK1002). Additionally, the capability of the extract to induce clastogenic effect was determined using Chinese hamster lung V79 fibroblast cells. Our results showed that with and without the presence of S9 metabolic activation, no mutagenic effects were observed in both Ames and umu tests when treated with P(3HB-co-4HB) extract. Similarly, treatment of P(3HB-co-4HB) extract in V79 fibroblast cells showed no significant production of micronuclei when compared with the positive control (Mitomycin C). Together, these results indicate that leachables of poststerilized P(3HB-co-4HB) cause no mutagenic and clastogenic effects.
A series of ternary metal(ii) complexes {M(phen)(edda); 1a (Cu), 1b (Co), 1c (Zn), 1d (Ni); H(2)edda = N,N(')-ethylenediaminediacetic acid} of N,N'-ethylene-bridged diglycine and 1,10-phenanthroline were synthesized and characterized by elemental analysis, FTIR, UV-visible spectroscopy and magnetic susceptibility measurement. The interaction of these complexes with DNA was investigated using CD and EPR spectroscopy. MTT assay results of 1a-1c , screened on MCF-7 cancer cell lines, show that synergy between the metal and ligands results in significant enhancement of their antiproliferative properties. Preliminary results from apoptosis and cell cycle analyses with flow cytometry are reported. seems to be able to induce cell cycle arrest at G(0)/G(1). The crystal structure of 1a is also included.
F16 is a plant-derived pharmacologically active fraction extracted from Eurycoma longifolia Jack. Previously, we have reported that F16 inhibited the proliferation of MCF-7 human breast cancer cells by inducing apoptotic cell death while having some degree of cytoselectivity on a normal human breast cell line, MCF-10A. In this study, we attempted to further elucidate the mode of action of F16. We found that the intrinsic apoptotic pathway was invoked, with the reduction of Bcl-2 protein. Then, executioner caspase-7 was cleaved and activated in response to F16 treatment. Furthermore, apoptosis in the MCF- 7 cells was accompanied by the specific proteolytic cleavage of poly(ADP-ribose) polymerase-1 (PARP-1). Surprisingly, caspase-9 and p53 were unchanged with F16 treatment. We believe that the F16-induced apoptosis in MCF-7 cells occurs independently of caspase-9 and p53. Taken together, these results suggest that F16 from E. longifolia exerts anti-proliferative action and growth inhibition on MCF-7 cells through apoptosis induction and that it may have anticancer properties.
The study of bioactivity of natural product is one of the major researches for drug discovery. The aim of this finding was to study the proliferation effect of Rhaphidophora korthalsii methanol extract on human PBMC and subsequently the cytotoxic effect of activated PBMC toward HepG2 human hepatocellular carcinoma. In this present study, MTT assay, cell cycle study and Annexin 5 binding assay were used to study the immunomodulatory and cytotoxic effects. In vitro cytotoxic screening of Rhaphidophora korthalsii methanol extract showed that the extract was non-toxic against hepatocellular carcinoma (HepG2). In contrast, the extract was able to stimulate the proliferation of human PBMC at 48 h and 72 h in MTT assay and cell cycle progress study. The application of immunomodulator in tumor research was studied by using MTT microcytotoxicity assay and flow cytometric Annexin V. Results indicated that pre-treated PBMC with Rhaphidophora korthalsii methanol extract induced the highest cytotoxicity (44.87+/-6.06% for MTT microcytotoxicity assay and 51.51+/-3.85% for Annexin V) toward HepG2. This finding demonstrates that Rhaphidophora korthalsii methanol extract are potent to stimulate the cytotoxic effect of immune cells toward HepG2.
The plant Andrographis paniculata found throughout Southeast Asia contains Andrographolide 1, a diterpenoid lactone, which has antitumour activities against in vitro and in vivo breast cancer models. In the present study, we report on the synthesis of andrographolide derivatives, 3,19-isopropylideneandrographolide (2), 14-acetyl-3,19-isopropylideneandrographolide (3) and 14-acetylandrographolide (4), and their in vitro antitumour activities against a 2-cell line panel consisting of MCF-7 (breast cancer cell line) and HCT-116 (colon cancer cell line). Compounds 2 and 4 were also screened at the US National Cancer Institute (NCI) for their activities against a panel of 60 human cancer cell lines derived from nine cancer types. Compound 2 was found to be selective towards leukaemia and colon cancer cells, and compound 4 was selective towards leukaemia, ovarian and renal cancer cells at all the dose-response parameters. Compounds 2 and 4 showed non-specific phase of the cell cycle arrest in MCF-7 cells treated at different intervals with different concentrations. NCI's COMPARE and SOM mechanistic analyses indicated that the anticancer activities of these new class of compounds were not similar to that of standard anticancer agents, suggesting novel mechanism(s) of action.
A new carbazole alkaloid, 3-carbomethoxy-2-hydroxy-7-methoxycarbazole, Clausine-TY (1), together with two known carbazole alkaloid, Clausine-H (2) and Clausine-B (3), were isolated from the ethyl acetate extract of the stem bark of the Malaysian Clausena excavata. The structures of these compounds were elucidated by spectroscopic analyses. The new carbazole alkaloid shows significant cytotoxicity against CEM-SS cell line.
Coral matrix of Porites sp. has the suitable properties for bone cell growth. This study was aimed to study the gene expression levels of osteoblast specific genetic markers; RUNX2, osteopontin, alkaline phosphatase and osteocalcin from osteoblasts seeded in coral scaffold, which are important in determining the feasibility of osteoblasts. Human osteoblasts were inoculated onto the processed coral in Dulbecco's Minimum Essential Medium. The cells were trypsinized on day 1, 7, 14, 18, and 21 and added with RNALater for preservation of RNA in cells. The RNA was extracted using commercial RNA extraction kit and the respective genes were amplified using RT-PCR kit and analyzed qualitatively on 1.5% agarose gel. The expressions were evaluated with the Integrated Density Value based on the intensity of band for different periods of cell harvest. Increased expressions of the RUNX2, osteopontin, alkaline phosphatase and osteocalcin genes in the present study proved that coral is a favorable carrier for osteogenetically competent cells to attach and remain viable.
The anti-proliferative effects of the aqueous and methanol extracts of leaves of Pereskia bleo (Kunth) DC (Cactaceae) against a mouse mammary cancer cell line (4T1) and a normal mouse fibroblast cell line (NIH/3T3) were evaluated under an optimal (in culture medium containing 10% foetal bovine serum (FBS)) and a sub-optimal (in culture medium containing 0.5% FBS) conditions. Under the optimal condition, the aqueous extract showed a significant (p<0.05) anti-proliferative effect at 200 microg/mL and 300 microg/mL in 4T1 cells and 300 microg/mL in NIH/3T3 cells, whereas the methanol extract did not show any notable anti-proliferative effect in these cell lines, at any of the concentrations tested. Under the sub-optimal condition, the aqueous extract showed a significant (p<0.05) anti-proliferative effect at 200 microg/mL and 300 microg/mL in NIH/3T3 cells, whilst the methanol extract showed a significant (p<0.05) anti-proliferative effect at 200 microg/mL and 300 microg/mL in both cell lines. An upward trend of apoptosis was observed in both 4T1 and NIH/3T3 cells treated with increasing concentrations of the aqueous extract. The level of apoptosis observed at all the concentrations of the aqueous extract tested was consistently higher than necrosis. There was a significant (p<0.05) increase in the level of necrosis observed in the 4T1 cells treated with 300 microg/mL of the methanol extract. Generally, the level of necrosis was noted to be higher than that of apoptosis in the methanol extract-treated cells. The mutagenicity assay performed showed that in the absence of S-9 liver metabolic activation, the extract was not mutagenic up to the concentration of 165 microg/mL . However, in the presence of S-9 liver metabolic activation, the aqueous extract was mutagenic at all the concentrations tested. This study shows that both the aqueous and methanol extracts of the leaves from Pereskia bleo (Kunth) DC (Cactaceae) do not have appreciable anti-proliferative effect on the 4T1 and NIH/3T3 cells as the EC(50) values obtained are greater than 50 microg/mL when tested under optimal culture condition. Moreover, the aqueous extract may form mutagenic compound(s) upon the metabolisation by liver enzymes.
The aim of the present study was to determine the effect of nitric oxide (NO) on the production of prostaglandin E2 (PGE2) by a human osteoblast cell line (HOS cells) stimulated with hydroxyapatite. Cells were cultured on the HA surfaces with or without the presence of NO donors (SNAP and NAP) for 3 days. The effect of NO scavenger, carboxy PTIO, or endothelial nitric oxide synthase (eNOS) inhibitor, L-NIO, was assessed by adding this scavenger in the cultures of HA-stimulated HOS cells with or without the presence of SNAP. Furthermore, HOS cells were pre-treated with anti-human integrin alphaV antibody, indomethacin, a non-specific inhibitor, aspirin, a COX-1 inhibitor, or nimesulide, a COX-2 inhibitor, prior to culturing on HA surfaces with or without the presence of SNAP. The levels of PGE2 were determined from the 3 day culture supernatants. The results showed that the production of PGE2 by HA-stimulated HOS cells was augmented by SNAP. Carboxy PTIO suppressed but L-NIO only partially inhibited the production of PGE2 by HA-stimulated HOS cells with or without the presence of exogenous NO. Pre-treatment of the cells with anti-human integrin alphaV antibody, indomethacin or nimesulide but not aspirin suppressed the production of PGE2 by HA-stimulated HOS cells with or without the presence of NO. Therefore, the results of the present study suggest that NO may up-regulate the production of PGE2 by augmenting the COX-2 pathway initiated by the binding between HOS cell-derived integrin alphaV and HA surface.