WO2001010428A2 - Utilisation de bases n-methyl d'ethanolamine pour prevenir la mort cellulaire induite par le stress oxydant - Google Patents

Utilisation de bases n-methyl d'ethanolamine pour prevenir la mort cellulaire induite par le stress oxydant Download PDF

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WO2001010428A2
WO2001010428A2 PCT/IL2000/000464 IL0000464W WO0110428A2 WO 2001010428 A2 WO2001010428 A2 WO 2001010428A2 IL 0000464 W IL0000464 W IL 0000464W WO 0110428 A2 WO0110428 A2 WO 0110428A2
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cells
dea
dha
oxidative stress
cell
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WO2001010428A3 (fr
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Ephraim Yavin
Annette Brand
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Yeda Research and Development Co Ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/13—Amines
    • A61K31/131—Amines acyclic

Definitions

  • the present invention relates to prevention of cell death induced by oxidative stress and, more particularly, to the use of ethanolamine analogues for this purpose.
  • CDP cytidine diphosphate
  • Cho choline
  • CPG choline phosphoglyceride
  • dEa N.N'-dimethylethanolamine
  • dEPG N,N'-dimethylethanol- amine phosphoglyceride
  • DHA docosahexaenoic acid
  • DMEM Dulbecco's modified Eagle's medium
  • Ea ethanolamine
  • EPG ethanolamine phosphoglyceride
  • FA fatty acid
  • FACS fluorescence-activated cell sorter
  • FCS fetal calf serum
  • G6PD glucose-6-phosphate dehydrogenase
  • IPG inositol phosphoglyceride
  • LPO lipid peroxide
  • mEa N-monomethylethanolamine
  • mEPG N-monomethylethanolamine phosphoglyceride
  • MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-
  • Oxygen free radicals are continuously formed as intermediates of enzymatic reactions during normal cellular function, e.g. mitochondrial respiratory activity, and some may be involved in growth regulation and intercellular signaling
  • ROS reactive oxygen species
  • the nervous system is particularly vulnerable to oxidative damage (Halliwell, 1992).
  • One major reason is the high content of polyunsaturated fatty acids (PUP As), which are especially sensitive to free radical damage due to the multiplicity of double bonds (Halliwell and Chirico, 1993; Alexander-North et al., 1994).
  • PUP As polyunsaturated fatty acids
  • Free radicals of oxygen generated during oxidative stress can attack PUFAs in the membrane bilayer and generate lipid hydroperoxides (Halliwell, 1992). The latter can initiate complex chain reactions, leading to more radicals, which damage cells and enhance cell death (Halliwell, 1992.
  • Docosahexaenoic acid is a major PUFA and an essential constituent of neuronal and glia cell plasma membranes and may participate in growth regulation (Yavin et al., 1975) or subserve as second messenger in cellular signal transduction processes, although less evidence is available for the latter effect.
  • DHA has the potential to increase the susceptibility of the membranes to form lipid peroxides (LPOs)
  • LPOs lipid peroxides
  • OLN 93 modified cell membrane lipid composition of actively proliferating clonal cells of oligodendroglia origin, OLN 93, by adding DHA and different N-bases, i.e. ethanolamine (Ea), N-monomethylethanolamine (mEa), N,N'-dimethylethanolamine (dEa), and choline (Cho), to the growth medium.
  • DHA ethanolamine
  • mEa N-monomethylethanolamine
  • dEa N,N'-dimethylethanolamine
  • Cho choline
  • the present invention thus relates to pharmaceutical compositions comprising N-monomethylethanolamine and /or N,N'-dimethylethanolamine for preventing cell death induced by oxidative stress.
  • These compositions are useful for protecting brain tissue from ischemic stroke and for the alleviation of injuries or disorders such as ischemic stroke, thromboembolic or hemorrhagic stoke, cardiac arrest, oxidative distressed pregnancy and neurodegenerative disorders such as aging and Alzheimer's disease.
  • the active ingredient of the composition is
  • composition is formulated for oral administration.
  • the invention also relates to a method for preventing cell death induced by oxidative stress comprising administering to a subject in need thereof an effective amount of N-monomethylethanolamine and /or N,N'-dimethylethanolamine.
  • Fig. 1 P incorporation into phospholipids of OLN 93 cells following N-methyl base administration.
  • OLN 93 cells were grown in 60-mm-diameter poly-L-lysine-coated dishes for 2 days with 1 mCi/ml carrier-free ortho[ o2 P]phosphate in either the absence or presence of 1 mM mEa or dEa.
  • the radioactive medium was discarded and cells were rinsed with PBS before lipid extraction as indicated in Materials and Methods. Radioactively labeled phospholipids were separated by TLC and after exposure to an X-ray film, spots were scraped off and counted by the Cerenkov method. The experiment was repeated on several occasions with nearly identical results (p ⁇ 0.05).
  • IPG Inositol phosphoglyceride
  • SPG serine phosphoglyceride
  • SPG sphingomyelin
  • CPG choline phosphoglyceride
  • EPG ethanolamine phosphoglyceride
  • mEPG monomethylethanolamine phosphoglyceride
  • dEPG dimethylethanolamine phosphoglyceride
  • mEa dimethylethanolamine
  • Fig. 2 Oxidative stress-induced lipid peroxidation in OLN 93 cells after lipid modification. Cells were grown for 3 days in the presence of 0.1 mM DHA and/or 1 mM mEa or dEa. Oxidative stress was induced by a 30-min incubation with 0.1 mM H 2 0 2 and the release of TBARS into the culture medium was determined as detailed in Materials and Methods. DHA, docosahexaenoic acid.
  • Fig. 3 Oxidative stress-induced TBARS release after lipid modification with various concentrations of N-methyl bases.
  • OLN 93 cells were grown for 3 days in the presence of either 0.02, 0.1 or 1 mM concentrations of dEa or Cho. Oxidative stress was induced by a 30-min incubation with 0.1 mM H 2 O 2 , and cellular lipid composition and the release of TBARS into the culture medium were determined as detailed in Materials and Methods. Cho, choline.
  • Fig. 4 Oxidative stress-induced TBARS release after time dependent lipid modification.
  • OLN 93 cells were incubated with 1 mM dEa or Cho for 4, 8, 24 or 48 h (a and c).
  • OLN 93 cells were grown for 3 days in the presence of 1 mM dEa, the culture medium was changed to DMEM not containing any dEa and cultures were incubated for another 1-3 days (b and d).
  • Oxidative stress was induced by a 30-min incubation with 0.1 mM H 2 0 2 , and cellular lipid composition (c and d) and the release of TBARS into the culture medium (a and b) were determined as detailed in Materials and Methods.
  • Fig. 5 H 2 0 2 clearance from the medium of modified OLN 93 cells.
  • Cells were grown for 3 days in the presence of 0.1 m DHA and/or 1 m mEa, or Cho.
  • H 2 0 2 was added at 0.1 m to the cultures, and for 30 min the disappearance of the H 2 0 2 from the culture medium was determined as detailed in Materials and Methods.
  • Fig. 6 Cell viability of modified OLN 93 cells after H 2 0 2 treatment.
  • Cells were grown for 3 days in the presence of 0.1 mM DHA and/or 1 mM dEa.
  • Oxidative stress was induced by a 30-min incubation with 0.1 mM H 2 0 2 .
  • Medium was discarded and fresh medium containing 5 % FCS was added.
  • mitochondrial activity was determined by using the MTT reduction assay as detailed in Materials and Methods. Con, control; OD, optical density.
  • Fig. 7 Cell-cycle analysis of modified OLN 93 cells after H 2 0 2 treatment.
  • OLN 93 cells were grown for 2 days in 35-mm-diameter culture dishes in DMEM containing 10 % FCS. Medium was changed to serum-free DMEM containing either 0.1 mM DHA or a combination of 0.1 mM DHA and 1 mM dEa. Oxidative stress was induced by a 30-min incubation with 0.5 mM H 2 0 2 (right panels). Medium was discarded and fresh medium containing 5 % FCS was added for 24 h. FACS analysis was performed as detailed in Materials and Methods, au, arbitrary units. DETAILED DESCRIPTION OF THE INVENTION
  • the present invention is directed to a novel and important use of N-monomethylethanolamine, N,N' -dimethylethanolamine or a mixture thereof, namely, the prevention of mammalian cell death induced by oxidative stress.
  • a major reason for brain tissue vulnerability to oxidative damage is the high content of PUFAs which are highly sensitive to free radicals attack.
  • Docosahexaenoic acid (DHA, 22:6 n3) is a PUFA exclusively acylated at the 2 position of certain phospholipid species and confers asymmetry to the membrane which may be of importance in cell-cell interaction, cell recognition and cell death.
  • Oligodendroglia-like OLN 93 cells lack PUFAs and are relatively insensitive tp oxidative stress.
  • compositions of the invention may be useful both as a preventive measure in the treatment of persons known to be prone to stroke or heart attack and to the treatment/alleviation of the patient once the stroke or heart attack occurs,
  • compositions are also useful for alleviating the effects of aging and of oxidative distressed pregnancy.
  • compositions of the invention are preferably formulated for oral administration by standard procedures, e.g. in the form of dosage units, such as capsules, cachets, tablets or lozenges; in the form of a powder or granules for reconstitution; in the form of a solution or a suspension in an aqueous liquid or nonaqueous liquid.
  • dosage will depend on the disease, the age and the condition of the patient, as determined by a skilled physician.
  • the dosage to be administered to a patient will be lower in cases in which the composition is administered frequently as a preventive measure, and will be higher in cases of emergency, e.g. after stroke or heart attack.
  • oligodendroglia-like OLN 93 clonal cell line (Richter-Landsberg and Heinrich, 1996) at less than 30 passages.
  • DMEM Dulbecco's modified Eagle's medium
  • FCS 10% fetal calf serum
  • OLN 93 cells were seeded on plastic culture dishes (35 mm in diameter) pre-coated with 50 ⁇ g/ml poly-L-lysine (PLL, MW 78,000, Sigma, St. Louis, MO, USA) in DMEM containing 10% FCS. If not stated otherwise, the culture medium was changed after 1-2 days to DMEM containing different hormones and growth factors, i.e. 10 ng/ml glycyl-L-histidyl-L-lysine (Calbiochem, La Jolla, CA, USA), 40 ⁇ g/ml hydrocortisone (Sigma, St.
  • lipid-modified and control OLN 93 cells were pre-incubated with DMEM containing 25 ⁇ M Fe 2 S0 4 . After 30 min, Fe 2 S0 4 was added to cells in a final concentration of 50 ⁇ M, and H 2 0 2 was added in a final concentration of 0.1 or 0.5 mM. After another 30-min incubation, the medium was collected for thiobarbituric acid (TBA) reactive substances (TBARS) measurement and the cells were immediately subjected to lipid extraction or returned to the incubator in DMEM containing 5% FCS, until the time of harvesting for fluorescence-activated cell sorter (FACS) analysis.
  • TAA thiobarbituric acid
  • TBARS thiobarbituric acid
  • FACS fluorescence-activated cell sorter
  • Lipid extraction After removal of the culture medium, the cell monolayer was washed once with cold PBS and dried as much as possible. An organic solvent mixture consisting of hexane/isopropanol, 3:2 vol/vol containing 0.01% (wt/vol) butylated hydroxytoluene was added to the culture dish and left for 10 min at room temperature. The extract was then collected into an Eppendorf tube and each plate was washed with an additional volume of hexane/isopropanol. The extracts were pooled and the solvent was evaporated under a nitrogen gas flow. The dried lipid extract was dissolved in a minimal volume of hexane/isopropanol and subjected to TLC or gas chromatography.
  • TLC and gas chromatography analysis The major phospholipids, choline phosphoglyceride (CPG), ethanolamine phosphoglyceride (EPG), serine phosphoglyceride (SPG) and inositol phosphoglyceride (IPG), were separated by thin-layer chromatography (TLC) on heat-activated silica gel G plates (Merck, Darmstadt, Germany) with a solvent mixture of chloroform/methanol/methylamine (40%) (130:70:30 by vol). All the solvents were of HPLC grade. Lipid bands resolved on TLC were visualized under UN light after spraying with 0.2% dichlorofluorescein in ethanol. Individual spots were identified and scraped into glass tubes. Transmethylation of FAs and gas-chromatographic separation of the FA methyl esters were performed by standard methods. Lipid phosphorus content was determined according to Bartlett, 1959.
  • TBA Thiobarbituric acid assay.
  • the TBA reagent was prepared by dissolving 0.67% TBA (wt/vol) in 50% acetic acid and adding 0.01% (wt/vol) of butylated hydroxytoluene.
  • 0.5 ml of culture medium collected after stress 0.5 ml of TBA reagent was added and heated in a boiling water bath for 15 min. The samples were cooled with tap water and the developed color was read in a Shimadzu RF-540 spectrofluorophotometer at 535 nm excitation and 553 nm emission.
  • a standard curve was prepared by using a 100 ⁇ M 1, 1,3,3-tetraethoxypropane stock solution diluted in ethanol. Medium without cells was assayed as the blank, exactly under the same conditions.
  • H 0 2 The removal of H 0 2 from the incubation buffer of modified OLN 93 cells was determined using a method described by Dringen et al. (1998). Briefly, to cells grown on 35-mm-diameter poly-L-lysine-precoated plates, supplements were added for 3 days. Cultures were washed with HEPES buffer solution (20 mM HEPES, 145 mM NaCl, 1.8 mM CaCl 2 , 5.4 mM KC1, 1 mM MgCl 2 , 0.8 mM Na 2 HP0 4 , and 5 mM glucose, pH 7.4) and incubated for 30 min with buffer containing 0.1 mM H 2 0 .
  • HEPES buffer solution (20 mM HEPES, 145 mM NaCl, 1.8 mM CaCl 2 , 5.4 mM KC1, 1 mM MgCl 2 , 0.8 mM Na 2 HP0 4 , and 5 mM glucose
  • the culture plates were kept at 37°C on a shaking water bath and 10 ⁇ l aliquots of the incubation buffer were collected at designated times. Each aliquot was combined with 180 ⁇ l of 25 mM H S0 4 on a 96-microtiter plate, and 190 ⁇ l of reaction mixture [0.5 mM (NH 4 ) 2 Fe(S0 4 ) 2 , 200 mM sorbitol, and 200 ⁇ M xylenol orange in 25 mM H 2 S0 4 ] were added. After a 45-min incubation at room temperature the absorbance was read in an ELISA reader (ELX 808, BIO-TEK Instruments, Inc., USA) at 540 nm.
  • ELISA reader ELX 808, BIO-TEK Instruments, Inc., USA
  • G6PD glucose-6-phosphate dehydrogenase
  • the assay of Langdon (1996) was used with minor modifications.
  • the reduction of NADP + toNADPH was detected as the rate of change of absorbance at 340 nm.
  • the medium for measurements consisted of 40 mM Tris-HCl (pH 7.5), 11.4 mMMgCl 2 , 0.17 mM ⁇ -NADP sodium salt, and 1.14 mM D-glucose-6-phosphate disodium salt dihydrate, and the reaction was started by addition of the cell homogenate (1-20 ⁇ l). Cellular fractionation was prepared in the presence of protease inhibitors.
  • OLN cells were scraped off in homogenization buffer, consisting of 20 mM Tris-HCl (pH 7.5), 2 mM EDTA, 0.5 mM EGTA, 2 mM dithiothreitol, 2 mM phenylmethylsulfonyl fluoride, 10 ⁇ g/ml leupeptin, 10 ⁇ g/ml pepstatin, and 25 ⁇ g/ml aprotinin.
  • Cells were homogenized with a Polytron homogenizer (Kinematica, Switzerland) and centrifuged for 10 min at 1,000 rpm and 4°C.
  • the supernatant was centrifuged for 30 min at 45,000 g and 4°C to yield the cytosolic fraction.
  • the pellet was reextracted with the same buffer containing 1% Triton X-100 and centrifuged for 10 min at 11,000 g and 4°C, and the Triton X-100 extract was used for enzymatic activity determination. Protein content was measured according to the method of Bradford (1986). Cell viability assay and DNA content.
  • the samples were vigourosly vortexed until the pellets appeared colorless and centrifuged for 2 min (10,000 g). Color reading of the blue supernatant was done in an ELISA reader at 540 nm and 630 nm.
  • the cell pellet was dissolved in 0.2 M NaOH and subjected to DNA content determination according to the method described by Burton (1968).
  • OLN-93 cells were grown in serum-free, growth factor-supplemented medium in the presence of DHA either alone or in combination with either mEa or dEa. After a 48-h incubation, marked changes in polar head groups and FA composition, including formation of dEa and mEa lipid analogues were detected by TLC. As illustrated in Fig. 1, in the presence of millimolar concentrations of mEa and dEa, 43% and 56% of the total lipid-soluble P radioactivity were taken up into the newly synthesized N-monomethyl- and N,N'-dimethyl-ethanolamine phosphoglyceride (EPG) (mEPG and dEPG) lipid analogues, respectively.
  • EPG N-monomethyl- and N,N'-dimethyl-ethanolamine phosphoglyceride
  • FIG. 3 depicts the relationship between the steady state levels of dEPG and CPG after 3 days of incubation in the presence of dEa and Cho, respectively, and the corresponding TBARS levels after H 2 0 2 stress.
  • increasing the concentration of Cho up to ImM did not cause a significant rise in the levels of CPG, which regularly accounts for about 35 % of the total phospholipids.
  • Apart from an initial decrease at 0.02 mM Cho no changes were noticed in TBARS levels up to 1 mM Cho.
  • dEPG After 3 days of incubation with 0.1 mM dEa, formation of dEPG reached a steady state level of 24 % of total cellular phospholipids. Under these conditions, TBARS formation after H 2 0 2 stress was decreased by 40 % compared with the control value. At 1 mM dEa, dEPG accounted for 50 % of the total cellular phospholipids; at this concentration the H 2 0 2 -induced TBARS levels were down to 30% of the control levels. Thus, it would appear that inhibition of TBARS production and accumulation of dEPG are mutually related events.
  • the inducible glucose-6-phosphate dehydrogenase (G6PD; EC 1.1.1.49) activity was examined in cytosolic and paniculate fractions prepared from control and dEa supplemented cultures. As shown in Table I, a 25% increase in the specific activity was found in cytosolic preparations from dEa-supplemented cultures.
  • H 2 0 2 -induced stress in OLN 93 cells is death by apoptosis as recently reported (Kameshwar-Rao et al, 1999). Therefore, the ability of dEa supplments to rescue cells from death was deemed worthy of investigation.
  • Mitochondrial activity measured by the MTT reduction assay was a helpful indicator to link lipid peroxidation to impaired cellular metabolism after H 2 0 2 stress. As illustrated in Fig. 6, treatment with 0.1 mM H 2 0 2 followed by a 24-h recovery period that included addition of serum, caused more than 50% decrease in the mitochondrial activity in control cells.
  • Fig. 7 shows the results of cell cycle distribution under the various experimental conditions. Addition of dEa, DHA, or a combination of both (Fig. 7, left panels) showed few if any (3-5%) cells in the sub- Gi phase, a fraction that is considered to contain the apoptotic cells (Rodriguez-Tarduchi et al., 1990). Most of the cells were found in the Gj phase whereas a high proportion of cells resumed the cycle (13-18%, G 2 /M phase) most likely because of withdrawal from serum starvation.
  • OLN 93 cells derived from a spontaneously growing oligodendroglia primary culture contain between 4-6% of both n-3 and n-6 long chain PUFAs of which DHA represents less than 0.55 ⁇ 0.02%.
  • This deficiency was instrumental in our hands in manipulating the cellular DHA content by exogenous supplements.
  • 0.1 mM DHA After 3 days following administration of 0.1 mM DHA, a 6 to 8-fold increase compared with control levels was noticed. Most of the DHA was esterified to enrich the SPG and EPG species as determined by gas chromatography (data not shown).
  • H 2 0 is a useful genotoxic agent to induce oxidative stress in cell culture systems. It easily penetrates cellular membranes, and in the presence of divalent iron, hydroxyl radicals are generated (Halliwell, 1992). Subsequent attacks by hydroxyl radicals on single and multiple double bonds of fatty acids causes production of aldehydic lipid peroxidation products, which can be detected as TBARS.
  • Malondialdehyde is the most abundant aldehyde resulting from lipid peroxidation of PUFAa (mainly arachidonic acid and DHA) with 3 or more methylene-interrupted double bonds that is detected by the TBA assay. The fact that more than 90% of the TBARS were released in the incubation medium, was advantageous in establishing an index for lipid peroxidation while cells were permitted to grow or die, depending on the stress.
  • the first question we addressed in this invention concerned whether lipid peroxidation in DHA-enriched cells is linked or not to apoptotic death.
  • H 2 0 2 used in the examples herein was 10-fold lower than in a previous study (Kameshwar-Rao et al., 1999)
  • DHA-enriched cultures produced more TBARS and showed a greater proportion of dying cells.
  • a direct correlation between TBARS formation (Fig 3) and apoptotic cell death (Fig 7) after DHA supplements was apparent, suggesting that the nature of the fatty acid composition may play an important role in cell death.
  • DHA supplements affected energy metabolism as evident by a 35% decrease in MTT reduction even without H 2 0 2 stress.
  • the second subject addressed in this invention was aimed at modifying the polar head group composition to study its relation to stress-induced cell damage and cell death.
  • the asymmetric distribution of polar head groups between the inner and outside part of the membrane bilayer is well known and is believed to fulfill an essential role in normal cell physiology.
  • some phospholipids such as SPG, a phospholipid normally present in the inner membrane layer, appears to be externalized during apoptosis presumably to serve as a recognition target for phagocytic clearance by macrophages (Shiratsuchi et al., 1998).
  • SPG a phospholipid normally present in the inner membrane layer
  • EPG appears to be redistributed at the cell surface during early stages of apoptosis, resulting in a total loss of asymmetric distribution of aminophospholipids in the plasma membrane bilayer.
  • CDP-Cho and CDP-Ea may have the potential to diminish the damage following global ischemia (Dorman et al., 1983). This has been attributed to the CDP-nucleotide negating the harmful release of free FAs observed in ischemic brains by stimulating de novo phospholipid synthesis. In our study, neither Cho nor Ea showed a protective effect, ruling out this possibility. Furthermore, as shown by the incorporation of ortho[ 32 P]phosphate into phospholipids (Fig.l), the turnover rate is high for both control and N-base analogues suggesting a similar amount of newly synthesized phospholipids. Therefore, it is likely that the concentration of the corresponding CDP-amines is about the same, nevertheless only dEa-modif ⁇ ed cells were ultimately protected against H 2 0 2 stress.
  • phosphorylated Ea analogues in particular dEa-phosphate
  • dEa-phosphate may function to extend longer-term cell survival in cultures subjected to serum deprivation-induced apoptotic cell death.
  • the results obtained herein after a chase using high Cho concentrations to deplete the former modified cells of dEa-phospholipid intermediates indicate that the protective potential of dEa treatment is not exclusively related to the concentration of newly synthesized dEa phospholipid derivatives or its intermediates.
  • H 2 0 2 and lipid hydroperoxides by glial cells depends on the presence of highly effective enzymes, i.e. catalase and glutathione peroxidase (Dringen and Hamprecht, 1997).
  • catalase and glutathione peroxidase Dringen and Hamprecht, 1997.
  • dEa-supplemented cultures were able to detoxify H 2 0 2 by 15% more rapidly than control cells, measuring the clearance of H 2 0 2 from the incubation medium as described by Dringen et al., (1998).
  • Cellular antioxidant defense mechanisms are dependent on a high reduction potential in the form of high NADPH concentrations.
  • the pentose phosphate pathway By replenishing the intracellular NADPH pool, the pentose phosphate pathway, a shunt to glycolysis, is enzymatically coupled to glutathione peroxidase.
  • an enhanced antioxidant capacity may play a role in the protective mechanisms based on dEa supplements, the changes observed in H 2 0 2 detoxification rate and G6PD activity seem to be too small to be the only reason for the reduced lipid peroxidation and, moreover, prevention from cell death.

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Abstract

L'invention concerne un groupe N-monométhyléthanolamine et N,N'-diméthyléthanolamine et des mélanges de ceux-ci utilisés pour prévenir la mort cellulaire induite par le stress oxydant, en particulier pour protéger les tissus cérébraux des accidents ischémiques cérébraux et pour alléger les souffrances induites par des accidents ischémiques cérébraux, des attaques d'apoplexie thromboemboliques ou hémorragiques, des arrêts cardiaques, des grossesses à trouble oxydatif et des troubles neurodégénératifs tels que le vieillissement et la maladie d'Alzheimer.
PCT/IL2000/000464 1999-08-10 2000-08-03 Utilisation de bases n-methyl d'ethanolamine pour prevenir la mort cellulaire induite par le stress oxydant Ceased WO2001010428A2 (fr)

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AU64652/00A AU6465200A (en) 1999-08-10 2000-08-03 Use of n-methyl bases of ethanolamine for prevention of cell death induced by oxidative stress
CA002378482A CA2378482A1 (fr) 1999-08-10 2000-08-03 Utilisation de bases n-methyl d'ethanolamine pour prevenir la mort cellulaire induite par le stress oxydant

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IL13133499A IL131334A0 (en) 1999-08-10 1999-08-10 Pharmaceutical compositions comprising hydroxyamines

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2013188417A3 (fr) * 2012-06-11 2014-03-20 The Cleveland Clinic Foundation Traitement et prévention d'une maladie cardiovasculaire et de la thrombose
EP2672965A4 (fr) * 2011-02-10 2014-07-16 Cleveland Clinic Foundation Traitement et prévention d'une maladie cardiovasculaire et d'une thrombose

Family Cites Families (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3088871A (en) * 1959-05-19 1963-05-07 Riker Laboratories Inc Process of stimulating the central nervous system
WO1988009171A1 (fr) * 1987-05-19 1988-12-01 Baylor College Of Medicine Phosphoethanolamine permettant le traitement de la maladie d'alzheimer
JPH03227921A (ja) * 1990-02-01 1991-10-08 Kanebo Ltd ケロイド治療剤
US6008188A (en) * 1994-05-06 1999-12-28 Kanebo Limited Cytokine potentiator and pharmaceutical formulation for cytokine administration
CA2221075C (fr) * 1997-11-14 2006-02-07 Meiji Milk Products Co., Ltd. Utilisation des derives d'amino-alcool dans le traitement de maladie hepatique
AU2186699A (en) * 1998-02-09 1999-08-23 Meiji Milk Products Co., Ltd. Preventives and remedies for intestinal mucosal disorder

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2672965A4 (fr) * 2011-02-10 2014-07-16 Cleveland Clinic Foundation Traitement et prévention d'une maladie cardiovasculaire et d'une thrombose
US9265736B2 (en) 2011-02-10 2016-02-23 The Cleveland Clinic Foundation Treatment of cardiovascular disease and thrombosis
WO2013188417A3 (fr) * 2012-06-11 2014-03-20 The Cleveland Clinic Foundation Traitement et prévention d'une maladie cardiovasculaire et de la thrombose
US9168233B2 (en) 2012-06-11 2015-10-27 The Cleveland Clinic Foundation Treatment and prevention of cardiovascular disease and thrombosis
US10064830B2 (en) 2012-06-11 2018-09-04 The Cleveland Clinic Foundation Treatment and prevention of cardiovascular disease and thrombosis
EP3524235A1 (fr) * 2012-06-11 2019-08-14 The Cleveland Clinic Foundation Traitement et prévention des maladies cardiovasculaires et de la thrombose

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AU6465200A (en) 2001-03-05
CA2378482A1 (fr) 2001-02-15
IL131334A0 (en) 2001-01-28
WO2001010428A3 (fr) 2002-06-13

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