MTT assay based inhibition analysis of A2780 cells proliferation using Mollugo nudicaulis Lam. extract with CXCR4 and HER2 expression

It is of interest to document the inhibition of A2780 cell proliferation using Mollugo nudicaulis Lam.(M.nudicaulis) extract by MTT assay and by monitoring the CXCR4 and HER2 expression through RT-PCR analysis. Results shown that the n-hexane extract of M.nudicaulis have anticancer activity IC₅₀ values of 32.46±0.92 µg/mL on A2780 cell lines. It is further found that the CXCR4 and HER2 mRNA and protein expression were significantly reduced in M.nudicaulis treated A2780 cell lines. Thus, the n-hexane extract of M.nudicaulis is a natural source of bioactive compounds as potential anticancer agents.     

Crystallization of the arginine-dependent repressor/activator AhrC from Bacillus subtilis

The arginine-dependent repressor/activator AhrC from Bacillus subtilis has been crystallized in space group C222(1), with unit cell dimensions a = 229.8 A, b = 72.8 A, c = 137.7 A and one aporepressor hexamer per asymmetric unit. Preliminary X-ray photographs show measurable intensities beyond 3.0 A.     

The effect of catalase and methionine-S-oxide reductase on oxidised alpha 1-proteinase inhibitor

Oxidative damage to alpha 1-proteinase inhibitor (alpha 1-PI) may be important in the pathogenesis of emphysema. We have studied the ability of 2 enzymes (catalase and methionine-S-oxide reductase) to prevent and reverse oxidation of alpha 1-PI by hydrogen peroxide. Pre-incubation of catalase with H2O2 protected alpha 1-PI from oxidation, but the enzyme could not reverse prior oxidation of alpha 1-PI. In contrast, methionine-S-oxide reductase fully restored activity to H2O2-oxidised alpha 1-PI. Sputum sol-phase from smokers and non-smokers contained alpha 1-PI that was only about 30% active. Functional activity increased in both smokers (p less than 0.025) and non-smokers (p less than 0.05) approximately 2-fold following incubation with the reductase. Western blotting of the samples showed that about 20% of the alpha 1-PI was present as an enzyme-inhibitor complex and 20% was proteolytically cleaved. These observations suggest proteolysis, complexing with enzyme and oxidation are mechanisms of inactivation of alpha 1-PI in lung secretions.     

Hyperreactivity of adenines and conformational flexibility of a translational repression site.

We have used a diethylpyrocarbonate (DEPC) modification [(1976) Prog. Nucl. Acids Res. 16, 189-262] to probe the accessibility of adenines essential for coat protein binding in the MS2 translational operator [(1983) Biochemistry 22, 2601-2610, 2610-2615, 4723-4730; (1987) Biochemistry 26, 1563-1568]. The essential adenines are apparently hyperreactive with this reagent relative to other sites within the same molecule. Variation of ionic strength, pH and divalent cation concentrations reveal the existence of two distinct conformers of the RNA operator as judged by DEPC reactivity. We propose that the hyperreactivity observed is due to the participation of neighbouring bases in the DEPC modification reaction and can be used as a novel structural probe.     

Probing the molecular mechanism of action of co-repressor in the E. coli methionine repressor-operator complex using surface plasmon resonance (SPR).

We have studied quantitatively the effect of the corepressor, S-adenosylmethionine (SAM), on the interaction between the E. coli methionine repressor, MetJ, and an idealised operator fragment, by recording measurements of surface plasmon resonance using a BIAcore instrument. We have recorded kinetic binding data in the presence of SAM, which carries a net positive charge, and two corepressor analogues, adenosylornithine (AO) and aza-SAM, which differ in the location of the atom carrying the positive charge. Our data support the hypothesis that the effect of the corepressor is electrostatic in origin. The difference in electrostatic interaction energy between the SAM- and AO-repressor-operator complexes of approximately 3.5 kJ/mol calculated from the known three-dimensional structure is within the range of our experimentally determined values of 2.8-4.3 kJ/mol. These results illustrate the potential of SPR measurements for studying protein-nucleic acid interactions.     

Cell density-dependent proliferation of the murine bone marrow-derived stromal cell lines

Proliferation of three murine marrow-derived stromal cell lines, LC1, LC2, and LC3, depended on initial cell density. For LC2 and LC3, the cell density-dependence was negated by conditioned-media, indicating growth dependence on a soluble growth factor. For LC1, conditioned-media failed to stimulate proliferation, suggesting growth dependence on direct cell-cell contact.     

Television epilepsy and pattern sensitivity.

Properly functioning domestic television sets may induce seizures in epileptic patients (TV epilepsy). We investigated the effects of different types of visual stimuli on paroxysmal electroencephalographic (EEG) activity in 32 patients known to be sensitive to intermittent photic stimulation (stroboscopic light). We monitored sensitivity to patterns of horizontal and vertical lines, both stationary and vibrated (pattern sensitivity), and to normal broadcasts on a domestic, black and white (405- or 625-line) TV receiver (TV sensitivity). Twenty-three of the 32 patients were sensitive to pattern. Twenty-two were sensitive to vibrated patterns, and 11 to static patterns (P less than 0-01), All patients sensitive to pattern were also sensitive to TV; The association between sensitivity to pattern and to TV was significant. Clinical history of TV epilepsy (16 out of 32 patients) and laboratory evidence of pattern or TV sensitivity were not significantly associated. The high incidence of pattern sensitivity among flicker-sensitive patients and its association with TV sensitivity suggests that linear patterns produced by the raster of a black and white set as it scans, or "line-jitter" produced by the raster in areas of low TV-signal strength may contribute to the epileptogenic effect of TV.     

Choline acetyltransferase activity of spinal cord cell cultures increased by co-culture with muscle and by muscle-conditioned medium

Activity of the enzyme choline acetyltransferase (CAT), which mediates the synthesis of the neurotransmitter, acetylcholine, was increased up to 20- fold in spinal cord (SC) cells grown in culture with muscle cells for 2 wk. This increase was directly related to the duration of co-culture as well as to the cell density of both the SC and muscle involved and was not affected by the presence of the acetylcholine receptor blocking agent, α-bungarotoxin. Glutamic acid decarboxylase (GAD) activity was often markedly decreased in SC-muscle cultures while the activities of acetylcholinesterase and several other enzymes were little changed. Increased CAT activity was also observed when SC cultures were maintained in medium which had been conditioned by muscle cells or by undifferentiated cells from embryonic muscle. Muscle-conditioned medium (CM) did not affect the activities of SC cell GAD or acetylcholinesterase. Dilution or concentration of the CM directly affected its ability to increase SC CAT activity , as did the duration and timing of exposure of the SC cells to the CM. The medium could be conditioned by muscle cells in the presence or absence of serum, and remained effective after dialysis or heating to 58 degrees C. Membrane filtration data were consistent with the conclusion that the active material(s) in CM had a molecular weight in excess of 50,000 daltons. We conclude that large molecular weight material that is released by muscle cells is capable of producing a specific increase in CAT activity of SC cells.