[Two-dimensional electrophoresis of proteins from breast cancer cells MCF-7. Changes in synthesis induced by FGF-2]. / Analyse par electrophorèse bidimensionnelle des protéines de cellules de cancer du sein MCF-7. Modifications de synthèse induites par le FGF-2.

Two-dimensional electrophoresis analysis of proteins from breast cancer cells MCF-7. Modifications of synthesis induced by FGF-2. Using high resolution two-dimensional electrophoresis, we have separated more than 1,000 proteins from the breast cancer cell line MCF-7. Computer assisted analysis of gels allowed us to classify these proteins in function of their isoelectric point, molecular weight and relative quantity. This data-base will now be used as a powerful tool to identified proteins which synthesis is regulated in various experimental or pathological situations. Thus we studied modifications induced by FGF-2. This growth factor induces the synthesis of four polypeptides which are not detected in cells not stimulated by this factor. In addition, intensity of nine other polypeptides was found increased in presence of FGF-2.

Acetylcholinesterase activity of the polychaete Nereis diversicolor: effects of temperature and salinity.

In order to estimate the potential use of the mean wholebody acetylcholinesterase (AChE) activity from the ragworm Nereis diversicolor for the biological assessment of pollution by anticholinesterase agents in estuarine areas, we measured the effects of the main abiotic factors (i.e. temperature and salinity) on AChE activity. We report here that AChE activity tends to decrease in individuals sampled in tanks at a salinity of 30 per thousand as temperature increases. No tendencies in the evolution of AChE activity were observed in individuals sampled in tanks at a salinity of 15 per thousand. In contrast, salinity seems to have a greater effect on AChE activity than temperature. At a temperature of 12 degrees C, a salinity of 30 per thousand provokes a significant transient increase of AChE 2 days after the beginning of the maintenance period compared with a salinity of 15 per thousand. The effects are short-term stress effects. We noticed only a transient increase of AChE activity between 2 days for individuals maintained in tanks at temperature of 20 degrees C and salinity of 15 and 30 per thousand, respectively, and 8 days for individuals maintained in tanks at salinity of 30 per thousand and at a temperature of 12 degrees C after the beginning of the maintenance period, confirming the more pronounced effect of salinity over temperature.

Activities of acetylcholinesterase, choline acetyltransferase, and catecholamine production in the spinal cord of the axolotl Ambystoma mexicanum during forelimb regeneration.

Amputation of an axolotl limb causes severance of the brachial nerves, followed by their regeneration into a blastema. It is known that these nerves provide a neurotrophic factor to blastemal cells. To approach the problem of the response of spinal cord nerve centers to forelimb amputation, we have studied biosynthetic activities in the nerve centers involved in axonal injury during limb regeneration. We report that the acetylcholinesterase (AChE) activity in the spinal cord is elevated 2 days (+69%) and 7 days (+28%) after limb amputation compared with levels in unamputated control animals, but is not significantly elevated at 3 h or 15 days. The percentages of slow (3.6S and 6.0S) and fast (18S) sedimenting forms of AChE progressively decrease 2 and 7 days after amputation, while those of intermediate sedimenting forms (10.5S and 14.0S) increase. Fifteen days after amputation, lower molecular weight forms return to the control level, but the heavy molecular weight form of AChE is absent as at 7 days; consequently intermediate molecular weight forms are in a greater proportion than the other two forms. Choline acetyltransferase activity was measured only 2 days after amputation (when AChE was at its highest level). It increases by about 34% with regard to the controls. Adrenaline is higher than controls 2 days after amputation, while noradrenaline is not significantly modified. The metabolic changes observed in the spinal cord during limb regeneration probably are the result of a general reaction to the stress of amputation (transection of brachial nerves) and regeneration of nerve fibers, since similar metabolic activities were observed after a simple denervation of the two unamputated forelimbs.

Biochemical and Enzymatic Characterization of an Acetylcholinesterase From Nereis diversicolor (Annelida, Polychaeta): Comparison With the Cholinesterases of Eisenia fetida (Annelida, Oligochaeta).

This study constitutes the first report of a biochemical characterization, involving both substrates and inhibitors and electrophoretic analysis, of a cholinesterase (ChE) from a polychaete annelid (Nereis diversicolor). The ChE of N. diversicolor appears to be an acetylcholinesterase (AChE); i.e., it hydrolyzes acetylthiocholine iodide at a higher rate than other substrates and is inhibited by eserine but not by iso-OMPA. The ChEs of Eisenia fetida are different from that of N. diversicolor and include at least two types of PrChEs. The AChE activity is located principally in the anterior region of the worm (head) in N. diversicolor, whereas the ChE activity of E. fetida is located throughout the body. The electrophoretic characterization of N. diversicolor and E. fetida ChEs showed, respectively, six and two isoforms with disc-PAGE, and three (55,000, 47,000, and 17,000) and five molecular forms (628,000, 301,000, 235,000, 106,000, and 53,800) with PAGGE; substantial activity remained at the top of the PAGGE gel in both species.

Effects of organophosphate and carbamate pesticides on acetylcholinesterase and choline acetyltransferase activities of the polychaete Nereis diversicolor.

A toxicity test for organophosphates (OP) and carbamates (C) was improved with the adult ragworm Nereis diversicolor. Animals were maintained in U-shaped glass tubes of 4-mm inner diameter fixed vertically on a plastic plate and placed in glass aquaria. Each tank was covered with glass in order to reduce evaporation and heat dissipation. Temperature varied between 15 and 16 degrees C and salinity was constant (34 per thousand) during the entire length of the experiment. Experiments were performed with a fixed day length of 12 h and seawater was gently aerated. The maintenance system allowed the administration of OP and C compounds via the seawater. An acclimatization period of 48 h was not sufficient to accomodate worms to their artificial burrows; accordingly, we chose to acclimate worms for a week before beginning the exposure. Choline acetyltransferase (ChAT) activity was very low and was not significantly modified by two OP compounds: malathion and parathion-ethyl. ChAT is not a target for these pesticides and should not be used for future studies about OP and C toxicity. On the other hand, inhibitory effects on acetylcholinesterase (AChE) activity were determined at concentrations of 10(-6) M for three OP compounds-malathion, parathion-ethyl, and phosalone-and a carbamate pesticide-carbaryl. We measured only short-term effects and no cumulative effect was determined, the maximum percentage of AChE activity inhibition being between 2 (carbaryl) and 7 (OP compounds) days after exposure and then remaining stable. Mortality occured only after a period of intoxication of 14 days. N diversicolor, which can be easily maintained at the laboratory, seems to be a good candidate for future laboratory studies to test the toxicity of other pollutants.