Antimicrobial and Cytotoxic Activity of Extracts of Ferula heuffelii Griseb. ex Heuff. and Its Metabolites

The antimicrobial and cytotoxic activities of isolates (CHCl₃ and MeOH extracts and selected metabolites) obtained from the underground parts of the Balkan endemic plant Ferula heuffelii Griseb . ex Heuff . were assessed. The CHCl₃ and MeOH extracts exhibited moderate antimicrobial activity, being more pronounced against Gram‐positive than Gram‐negative bacteria, especially against Staphylococcus aureus (MIC=12.5 μg/ml for both extracts) and Micrococcus luteus (MIC=50 and 12.5 μg/ml, resp.). Among the tested metabolites, (6E)‐1‐(2,4‐dihydroxyphenyl)‐3,7,11‐trimethyl‐3‐vinyldodeca‐6,10‐dien‐1‐one ( 2 ) and (2S*,3R*)‐2‐[(3E)‐4,8‐dimethylnona‐3,7‐dien‐1‐yl]‐2,3‐dihydro‐7‐hydroxy‐2,3‐dimethylfuro[3,2‐c]coumarin ( 4 ) demonstrated the best antimicrobial activity. Compounds 2 and 4 both strongly inhibited the growth of M. luteus (MIC=11.2 and 5.2 μM , resp.) and Staphylococcus epidermidis (MIC=22.5 and 10.5 μM , resp.) and compound 2 additionally also the growth of Bacillus subtilis (MIC=11.2 μM ). The cytotoxic activity of the isolates was tested against three human cancer cell lines, viz., cervical adenocarcinoma (HeLa), chronic myelogenous leukemia (K562), and breast cancer (MCF‐7) cells. The CHCl₃ extract exhibited strong cytotoxic activity against all cell lines (IC₅₀<11.0 μg/ml). All compounds strongly inhibited the growth of the K562 and HeLa cell lines. Compound 4 exhibited also a strong activity against the MCF‐7 cell line, comparable to that of cisplatin (IC₅₀=22.32±1.32 vs. 18.67±0.75μM ).     

An insertion element of the extremely thermophilic archaeon Sulfolobus solfataricus transposes into the endogenous β-galactosidase gene

Three phenotypically stable mutants of the extremely thermophilic archaeon Sulfolobus solfataricus have been isolated by screening for β-galactosidase negative colonies on plates with X-Gal (5-bromo-4-chloro-3-indolyl-(3-d-galactopyranoside). From one of these mutants an insertion element, designated ISC1217, was isolated and characterized. Sequence analysis of ISC1217 and of the regions adjacent to the insertion site in the β-galactosidase gene revealed features typical of a transposable element: ISC1217 contained terminal inverted repeats and was flanked by a direct repeat of 6 bp. The 1147 by sequence contained an open reading frame encoding a putative protein of 354 amino acid residues and, overlapping this, two smaller open reading frames on the opposite strand. There were approximately 8 copies of the insertion element in the S. solfataricus genome. ISC1217 did not cross-hybridize with DNA of other Sulfolobus species. All three independently isolated β-galactosidase mutants of S. solfataricus arose by transposition of ISC1217 or a related element.     

Fluctuations of Serum Neuron Specific Enolase and Protein S-100B Concentrations in Relation to the Use of Shunt during Carotid Endarterectomy

Objective

To evaluate the changes in serum neuron specific enolase and protein S-100B, after carotid endarterectomy performed using the conventional technique with routine shunting and patch closure, or eversion technique without the use of shunt.

Materials and Methods

Prospective non-randomized study included 43 patients with severe (>80%) carotid stenosis undergoing carotid endarterectomy in regional anesthesia. Patients were divided into two groups: conventional endarterectomy with routine use of shunt and Dacron patch (csCEA group) and eversion endarterectomy without the use of shunt (eCEA group). Protein S-100B and NSE concentrations were measured from peripheral blood before carotid clamping, after declamping and 24 hours after surgery.

Results

Neurologic examination and brain CT findings on the first postoperative day did not differ from preoperative controls in any patients. In csCEA group, NSE concentrations decreased after declamping (P<0.01), and 24 hours after surgery (P<0.01), while in the eCEA group NSE values slightly increased (P=ns), accounting for a significant difference between groups on the first postoperative day (P=0.006). In both groups S-100B concentrations significantly increased after declamping (P<0.05), returning to near pre-clamp values 24 hours after surgery (P=ns). Sub-group analysis revealed significant decline of serum NSE concentrations in asymptomatic patients shunted during surgery after declamping (P<0.05) and 24 hours after surgery (P<0.01), while no significant changes were noted in non-shunted patients (P=ns). Decrease of NSE serum levels was also found in symptomatic patients operated with the use of shunt on the first postoperative day (P<0.05). Significant increase in NSE serum levels was recorded in non-shunted symptomatic patients 24 hours after surgery (P<0.05).

Conclusion

Variations of NSE concentrations seemed to be influenced by cerebral perfusion alterations, while protein S-100B values were unaffected by shunting strategy. Routine shunting during surgery for symptomatic carotid stenosis may have the potential to prevent postoperative increase of serum NSE levels, a potential marker of brain injury.     

Antibody-bound amyloid precursor protein upregulates ornithine decarboxylase expression

Alzheimer's disease is a neurodegenerative disorder characterised by extracellular accumulation of the Abeta peptide, derived from the amyloid precursor protein (APP). The function of APP as a cell surface receptor was examined by ligand-mimicking using an antibody against the APP extracellular domain. Alterations in gene expression evoked by antibody-bound APP were analysed using human pathway-finder gene arrays and the largest change in expression levels was found for ornithine decarboxylase (ODC). These results were confirmed by Western blotting which showed even higher upregulation on the protein level. APP knockdown by RNAi verified that upregulation of ODC was APP-mediated. This APP signalling event did not require gamma-secretase cleavage, as it was independent of the presence of presenilin-1 or -2. The induced ODC expression was rapid and biphasic, resembling growth-factor stimulated signalling events. This study shows that antibody-bound APP leads to altered gene expression that may be relevant to AD.     

Discrepancies between flow cytometric analysis and [3H]thymidine incorporation in stimulated lymphocytes

The DNA synthesis system of freshly isolated tonsillar lymphocytes and those stimulated by phytohaemagglutinin were compared by different methods. Both cell populations had high DNA polymerase α and thymidine kinase activities, as well as a high rate of incorporation of [³H]thymidine into DNA. However, the two cell populations differed when their DNA distributions were compared by flow cytometry. Freshly isolated cells contained many less (6%) cells in S phase than were found in phytohaemagglutinin-stimulated lymphocytes (18%) as detected by flow cytometry. The labelling of different subpopulations of lymphocytes was studied by sorting them electrically with a fluorescence-activated cell sorter. Analysis of the radioactivity of [³H]thymidine pulse-labelled cells, sorted according to their DNA content, showed that cells in the G₁ peak of DNA distribution had a significant amount of incorporated [³H]thymidine. Sorting of cells according to their size (i.e., by light scattering) revealed that only large cells were labelled with [³H]thymidine.