Compounds from Geijera parviflora with prostaglandin E2 inhibitory activity may explain its traditional use for pain relief.

ETHNOPHARMACOLOGICAL RELEVANCE: Australian Aboriginal people used crushed leaves of Geijera parviflora Lindl. both internally and externally for pain relief, including for toothache (Cribb and Cribb, 1981). This study tested the hypothesis that this traditional use might be at least in part explained by the presence of compounds with anti-inflammatory activity. MATERIALS AND METHODS: A crude extract (95% EtOH) was prepared from powdered dried leaves. From the CH3Cl fraction of this extract compounds were isolated by bioassay-guided fractionation and tested for: (1) cytotoxicity in RAW 264.7 murine leukemic monocyte-macrophages, (2) prostaglandin E2 (PGE2) inhibitory activity in 3T3 Swiss albino mouse embryonic fibroblast cells, as well as (3) nitric oxide (NO) and (4) tumour necrosis factor alpha (TNFα) inhibitory activity in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Isolated compounds were also tested for (5) antibacterial activity against a panel of Gram-positive (Staphylococcus aureus ATCC 29213 and ATCC 25923, Staphylococcus epidermidis ATCC 35984, biofilm-forming) and Gram-negative (Escherichia coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853) strains by broth microdilution. RESULTS: Eleven compounds were isolated, including one new flavone and one new natural product, with a further four compounds reported from this species for the first time. Some of the compounds showed good anti-inflammatory activity in vitro. In particular, flindersine (1) and N-(acetoxymethyl) flindersine (3) inhibited PGE2 release with IC50 values of 5.0µM and 4.9µM, respectively, without any significant cytotoxicity. Several other compounds showed moderate inhibition of NO (5, 6, 7) and TNF-α (6), with IC50 in the low micromolar range; however much of this apparent activity could be accounted for by the cytotoxicity of these compounds. None of the compounds showed anti-bacterial activity. CONCLUSIONS: The inhibition of PGE2, an important mediator of inflammation and pain, by flindersine and a derivative thereof, along with the moderate anti-inflammatory activity shown by several other compounds isolated from Geijera parviflora leaf extract, support the traditional use of this plant for pain relief by Australian Aboriginal people.

Biologically active dibenzofurans from Pilidiostigma glabrum, an endemic Australian Myrtaceae.

In an effort to identify new anti-inflammatory and antibacterial agents with potential application in wound healing, five new dibenzofurans, 1,3,7,9-tetrahydroxy-2,8-dimethyl-4,6-di(2-methylbutanoyl)dibenzofuran (1), 1,3,7,9-tetrahydroxy-2,8-dimethyl-4-(2-methylbutanoyl)-6-(2-methylpropionyl)dibenzofuran (2), 1,3,7,9-tetrahydroxy-2,8-dimethyl-4,6-di(2-methylpropionyl)dibenzofuran (3), 1,3,7,9-tetrahydroxy-4,6-dimethyl-2-(2-methylbutanoyl)-8-(2-methylpropionyl)dibenzofuran (4), and 1,3,7,9-tetrahydroxy-4,6-dimethyl-2,8-di(2-methylpropionyl)dibenzofuran (5), were isolated from the leaves of Pilidiostigma glabrum together with one previously described dibenzofuran. Structure elucidation was achieved by way of spectroscopic measurements including 2D-NMR spectroscopy. Compounds with 2,8-acyl substitutions had potent antibacterial activity against several Gram-positive strains (MIC in the low micromolar range), while compounds with 4,6-acyl substitutions were less active. All compounds except 3 inhibited the synthesis of nitric oxide in RAW264 macrophages with IC(50) values in the low micromolar range. Compounds with 2,8-acyl substitutions also inhibited the synthesis of PGE(2) in 3T3 cells, whereas 4,6-acyl-substituted compounds were inactive. None of the compounds inhibited the synthesis of TNF-α in RAW264 cells. The compounds showed variable but modest antioxidant activity in the oxygen radical absorbance capacity assay. These findings highlight that much of the Australian flora remains unexplored and may yet yield many new compounds of interest. Initial clues are provided on structure/activity relationships for this class of bioactives, which may enable the design and synthesis of compounds with higher activity and/or selectivity.

A novel method for screening viral interferon-resistance genes.

Many viruses have evolved mechanisms to antagonize the interferon (IFN) system, targeting all the major components involved in receptor binding and signaling. Although a number of these vital proteins are homologous to cellular proteins involved in IFN downregulation (e.g., viral IFN regulatory factors [vIRFs]), many share little resemblance to known proteins. To determine the IFN-blocking properties of these proteins, functional assays are required. Here, we present a new and rapid functional screening method, based on the 2fTGH cell line, which is able to determine viral gene products that inhibit the IFN-alpha/Jak-Stat signaling pathway. Expression cloning of viral IFN-blocking genes into 2fTGH and consequent selection with IFN-alpha and 6-thioguanine result in the outgrowth of cells that are no longer responsive to IFN-alpha. We also demonstrate that selection occurs if members of the Jak-Stat signaling pathway are lost. To show the utility of our system, we have used a known suppressor of IFN signaling, the human papillomavirus (HPV) E7 gene. Expression of E7 causes the loss of ability of 2fTGH cells to respond to IFN-alpha treatment because of a functional disruption of the signaling pathway. This approach offers a new strategy for identifying novel viral genes or new functions of already described viral genes that have a role in IFN-alpha signaling inhibition.