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Biomedical subjects

M Chou

Publications and source records attributed to M Chou.

At least 19 recordsLinked to original sources
PubMed2005

Exercise primes a molecular memory for brain-derived neurotrophic factor protein induction in the rat hippocampus.

Exercise is an important facet of behavior that enhances brain health and function. Increased expression of the plasticity molecule brain-derived neurotrophic factor (BDNF) as a response to exercise may be a central factor in exercise-derived benefits to brain function. In rodents, daily wheel-running exercise increases BDNF gene and protein levels in the hippocampus. However, in humans, exercise patterns are generally less rigorous, and rarely follow a daily consistency. The benefit to the brain of intermittent exercise is unknown, and the duration that exercise benefits endure after exercise has ended is unexplored. In this study, BDNF protein expression was used as an index of the hippocampal response to exercise. Both daily exercise and alternating days of exercise increased BDNF protein, and levels progressively increased with longer running duration, even after 3 months of daily exercise. Exercise on alternating days was as effective as daily exercise, even though exercise took place only on half as many days as in the daily regimen. In addition, BDNF protein remained elevated for several days after exercise ceased. Further, after prior exercise experience, a brief second exercise re-exposure insufficient to cause a BDNF change in naïve animals, rapidly reinduced BDNF protein to levels normally requiring several weeks of exercise for induction. The protein reinduction occurred with an intervening "rest" period as long as 2 weeks. The rapid reinduction of BDNF by an exercise stimulation protocol that is normally subthreshold in naïve animals suggests that exercise primes a molecular memory for BDNF induction. These findings are clinically important because they provide guidelines for optimizing the design of exercise and rehabilitation programs, in order to promote hippocampal function.

Animals↗
PubMed2000

Potent and selective non-peptidic inhibitors of endothelin-converting enzyme-1 with sustained duration of action.

Potent and selective non-peptidic inhibitors of human endothelin-converting enzyme-1 (ECE-1) have been designed as potential modulators of endothelin (ET-1) production in vivo. Because of its unique structural characteristics and long duration of action in vivo, the dual ECE-1 and neutral endopeptidase 24.11 (NEP) inhibitor, CGS 26303, was selected as an attractive lead for further optimization of potency and selectivity. Replacement of the P(1)' biphenyl substituent of CGS 26303 by a conformationally restricted 3-dibenzofuranyl group led to more potent and more selective ECE-1 inhibitors, such as the tetrazole 27. The remarkable effect of this P(1)' modification allowed for the first time phosphonomethylcarboxylic acids, such as 29, to display both potent (IC(50) = 22 nM) and selective (104-fold vs NEP) ECE-1 inhibition. Chemoenzymatic syntheses of the new alpha-amino acid (S)-3-dibenzofuran-3-ylalanine intermediate were developed, and improved procedures to generate substituted alpha-aminoalkylphosphonic acids were devised to support the production of various analogues. Although additional gains in intrinsic ECE-1 inhibitory potency could occasionally be achieved by addition of a P(1) side chain, these compounds (e.g. 43a) showed poor functional activity in vivo in the big ET-1 pressor test. Phosphonoalkyl dipeptides featuring 3-dibenzofuranyl groups in both the P(1)' and P(2)' positions were also very potent ECE-1 inhibitors, albeit lacking the desired selectivity against NEP. Functionally, 27and 29 were the two most efficacious compounds from this study, producing sustained inhibition of ECE-1 activity in rats, as measured by their ability to block the hypertensive effects induced by big ET-1. This profile was similar to that of a potent ET(A)/ET(B) dual receptor antagonist, SB 209670. Due to their favorable in vitro and in vivo profiles, 27 (CGS 34043) and 29 (CGS 35066) constitute new pharmacological tools useful in assessing the role of ECE-1 in pathological conditions.

Animals↗
PubMed2000

CGS 34043: a non-peptidic, potent and long-acting dual inhibitor of endothelin converting enzyme-1 and neutral endopeptidase 24.11.

Endothelin-1 (ET- 1) is a potent vasoconstrictor. Its biosynthesis is catalyzed by endothelin converting enzyme (ECE). In contrast, atrial natriuretic peptide (ANP) is a potent vasorelaxant and diuretic, and it is mainly degraded by neutral endopeptidase 24.11 (NEP). Therefore, compounds that can suppress the production of ET-1 by inhibiting ECE while simultaneously potentiating the levels of ANP by inhibiting NEP may be novel agents for the treatment of cardiovascular and renal dysfunction. CGS 34043 is one such compound, which inhibited the activities of ECE-1a and NEP with IC50 values of 5.8 and 110 nM, respectively. In vivo, it inhibited the pressor response induced by big ET-1, the precursor of ET-1, dose-dependently in rats, and the inhibition was sustained for at least 2 hr. In addition, CGS 34043 increased plasma ANP by 150% up to 4 hr after an intravenous dose of 10 mg/kg in conscious rats infused with ANP. However, this compound had no effect on the angiotensin I-induced pressor response. These results demonstrate that CGS 34043 is a potent and long-lasting dual inhibitor of ECE-1 and NEP. Consequently, it may be beneficial for the treatment of diseases in which an overproduction of ET-1 and/or enhanced degradation of ANP plays a pathogenic role. The activity of CGS 34753, an orally active prodrug of CGS 34043, is also described.

Animals↗
PubMed2000

Design and synthesis of a potent and selective endothelin-converting enzyme inhibitor, CGS 35066.

CGS 26303 has previously been shown to inhibit human endothelin converting enzyme-1 (ECE-1) with an IC50 of 410 nM and to be efficacious in several animal disease models. However, it is a more potent inhibitor of neutral endopeptidase 24.11 (NEP) with an IC50 of 1 nM. The aim of this study was to optimize CGS 26303 for greater potency and selectivity towards ECE-1 inhibition. The in vivo activity of the compounds was assessed by inhibition of the big endothelin-1 (ET-1)-induced pressor response in anesthetized rats at 90 min after treatment with a dose of 10 mg/kg, i.v. Under these conditions, CGS 26303 inhibited the pressor response to big ET-1 by 50%. Replacement of the biphenyl and tetrazol groups in CGS 26303 with a dibenzofuran and carboxylic acid, respectively, yielded CGS 35066, a potent ECE-1 inhibitor having an IC50 of 22 nM. In contrast, these substitutions markedly weakened the NEP inhibitory activity of the compound to an IC50 of 2.3 microM. CGS 35066 also exhibited a potent and sustained ECE-1 inhibitory activity in vivo, blocking the pressor response to big ET-1 by 84%. Its orally active prodrug, CGS 35339, was obtained by introducing two phenyl groups at the phosphonic acid substituent in CGS 35066. Therefore, CGS 35066 and CGS 35339 represent novel compounds for assessing the pathogenic role of ET-1 overproduction in various disease states.

Animals↗
PubMed2000

Enhanced expression of renal endothelin-converting enzyme-1 and endothelin-A-receptor mRNA in rats with interstitial fibrosis following ureter ligation.

Endothelin-1 (ET-1) has been implicated in many chronic renal glomerular diseases. The purpose of this study was to investigate whether the levels of mRNA expression of endothelin-converting enzyme-1 (ECE-1) and endothelin-A- and -B- (ET(A) and ET(B)) receptors are altered during the progression of interstitial fibrosis following ureter ligation. Rats were subjected to left ureter ligation or a sham operation and euthanized 5 days afterward. Kidneys were fixed in Carnoy's fixative, embedded in paraffin, and sectioned for assessment of interstitial fibrosis by staining for collagen III using immunofluorescence techniques. The area occupied by collagen staining was quantified by image analysis. Kidneys from obstructed rats showed a 54% increase in the area occupied by collagen III staining compared to the contralateral kidney, and an 89% increase compared to sham-operated kidneys. The mRNA levels of ECE-1, as well as ET(A)- and ET(B)-receptors in the kidney were analyzed by Northern blots. It was found that the ECE-1 and ET(A)-receptor mRNA levels in kidneys subjected to ureter ligation increased by 92% and 71%, respectively, when compared with those obtained from the contralateral kidneys. In contrast, mRNA levels of ET(B)-receptors were not significantly different between the two groups. These results suggest that ET-1, through interaction with the ET(A)-receptors, may play a role in the progression of interstitial fibrosis following ureter ligation.

Animals↗
PubMed2000

Attenuation of puromycin aminonucleoside-induced glomerular lesions in rats by CGS 26303, a dual neutral endopeptidase/endothelin-converting enzyme inhibitor.

Endothelin-1 (ET-1) has been implicated in the pathogenesis of various renal diseases. The purpose of this study was to investigate the effect of CGS 26303, an endothelin-converting enzyme (ECE) inhibitor, on puromycin aminonucleoside (PA)-induced nephrosis in rats. The animals (three groups; n = 8 per group) received 50 mg/kg PA or NaCl, intravenously. CGS 26303 (5 mg/kg/day, s.c. via osmotic minipumps) or vehicle was administered to the PA-treated animals for 4 weeks, starting within 5 min after PA injection. Uninephrectomy was performed 2 weeks after PA to accelerate the renal damage. Rats received no treatment between 4 and 8 weeks. At 8 weeks rats were euthanized and kidneys removed for histology and analysis for mRNA levels of endothelin-A- and -B- (ET(A) and ET(B)) receptors and ECE-1. Glomeruli (100 glomeruli/section; 800/group) were graded as normal (N), mild lesion (ML = few periodic acid-Schiff positive [PAS+] droplets and small adhesions to Bowman's capsule), and moderate to severe lesion (SL = many PAS+ droplets, adhesions to and thickening of Bowman's capsule, mesangial expansion, and cystic dilations of glomerular capillaries). In the PA + vehicle group N, ML and SL were 39.5%, 11.9% and 48.6%, respectively, while the respective values were 68.3%, 9.4%, and 22.3% in PA + CGS 26303-treated rats. However, the renal mRNA levels of ECE-1 and ET(A)- and ET(B)-receptors were not significantly different among the three groups. These results confirm the efficacy of ECE inhibition in this disease model. On the other hand, the mRNA data suggest that either there was no change in the expression of the genes examined or their levels had already returned to normal by the end of the experiment.

Animals↗
PubMed1999

Feasibility of bioremediation of trichloroethylene contaminated sites by nitrifying bacteria through cometabolism with ammonia.

The autotrophic ammonia-oxidizing bacteria (Nitrosomonas sp.) are able to dechlorinate trichloroethylene (TCE) through cometabolism using ammonia (NH(3)) as a growth substrate. Cometabolic kinetics models suggest that TCE is a potent competitive inhibitor of NH(3) oxidation because it competes with NH(3) for oxidation by the enzyme of ammonia monooxygenase (AMO). In this study, an enriched culture of nitrifying bacteria was used to investigate the efficiencies of cometabolism of TCE by AMO. In addition, the relationships among specific growth substrate (NH(3)) utilization rate (qNH(3)), specific nongrowth substrate (TCE) cometabolic rate (qTCE), NH(3) and TCE concentrations, and NH(3)/TCE and TCE/NH(3) ratios were also analyzed. We found that the relationships between qNH(3) and NH(3) for the systems with and without TCE followed the Alvarez-Cohen competitive inhibition model and Monod model, respectively. Our results demonstrate that TCE could be cometabolized in a nitrification system when sufficient oxygen and NH(3)200 microg/l) were also found to show inhibitory effects towards NH(3) oxidation in enriched nitrifying culture. We also found that the NH(3)/TCE ratio rather than TCE concentrations alone exhibited strong correlation with qNH(3), much the same as the Ely activity recovery model presented. Our results suggest that the relationship between qTCE and TCE concentrations followed the Oldenhuis enzyme inactivation model for systems without NH(3).

Ammonia↗
PubMed1999

Effect of matrix metalloproteinase inhibition on progression of atherosclerosis and aneurysm in LDL receptor-deficient mice overexpressing MMP-3, MMP-12, and MMP-13 and on restenosis in rats after balloon injury.

The broad-spectrum MMP inhibitor CGS 27023A was tested to determine its potential as a therapy for atherosclerosis, aneurysm, and restenosis. LDL receptor-deficient (LDLr -/-) mice fed a high-fat, cholic acid-enriched diet for 16 weeks developed advanced aortic atherosclerosis with destruction of elastic lamina and ectasia in the media underlying complex plaques. Lesion formation correlated with a 4.6- to 21.7-fold increase in MMP-3, -12, and -13 expression. Treatment with CGS 27023A (p.o., b.i.d. at 50 mg/kg) had no effect on the extent of aortic atherosclerosis (36 +/- 4% versus 30 +/- 2% in controls), but both aortic medial elastin destruction and ectasia grade were significantly reduced (38% and 36%, respectively, p < 0.05). In the rat ballooned-carotid-artery model, CGS 27023A (12.5 mg/kg/day via osmotic minipump) reduced smooth muscle cell migration at 4 days by 83% (p < 0.001). Intimal lesions were reduced by 85% at 7 days (p < 0.001), but intimal smooth muscle proliferation was unaffected, and inhibitory efficacy was lost with time. At 12 days, intimal lesion reduction was less potent (52%, p < 0.01). At 3 and 6 weeks, reductions of 11% and 4%, respectively, were not significant. This demonstrates that it is essential to include late time points when the ballooned-carotid-artery model is employed to ensure that lesion size does not "catch up" when a compound solely inhibits smooth muscle cell migration. In summary, MMP inhibitor therapy delayed but did not prevent intimal lesions, thereby demonstrating little promise to prevent restenosis. In contrast, MMP inhibitor therapy may prove useful to retard progression of aneurysm.

Animals↗
PubMed1999

NH4+ transport system of a psychrophilic marine bacterium, Vibrio sp. strain ABE-1.

NH4(+) transport system of a psychrophilic marine bacterium Vibrio sp. strain ABE-1 (Vibrio ABE-1) was examined by measuring the uptake of [14C]methylammonium ion (14CH3NH3+) into the intact cells. 14CH3NH3+ uptake was detected in cells grown in medium containing glutamate as the sole nitrogen source, but not in those grown in medium containing NH4Cl instead of glutamate. Vibrio ABE-1 did not utilize CH3NH3+ as a carbon or nitrogen source. NH4Cl and nonradiolabeled CH3NH3+ completely inhibited 14CH3NH3+ uptake. These results indicate that 14CH3NH3+ uptake in this bacterium is mediated via an NH4+ transport system and not by a specific carrier for CH3NH3+. The respiratory substrate succinate was required to drive 14CH3NH3+ uptake and the uptake was completely inhibited by KCN, indicating that the uptake was energy dependent. The electrochemical potentials of H+ and/or Na+ across membranes were suggested to be the driving forces for the transport system because the ionophores carbonylcyanide m-chlorophenylhydrazone and monensin strongly inhibited uptake activities at pH 6.5 and 8.5, respectively. Furthermore, KCl activated 14CH3NH3+ uptake. The 14CH3NH3+ uptake activity of Vibrio ABE-1 was markedly high at temperatures between 0 degrees and 15 degrees C, and the apparent Km value for CH3NH3+ of the uptake did not change significantly over the temperature range from 0 degrees to 25 degrees C. Thus, the NH4+ transport system of this bacterium was highly active at low temperatures.

Ammonium Chloride↗
PubMed1999

MEC-12, an alpha-tubulin required for touch sensitivity in C. elegans.

mec-12 is one of a dozen genes required for touch receptor neuron function in Caenorhabditis elegans. Some mec-12 mutants (mechanosensory-defective) lack the large-diameter microtubules that are characteristic of these neurons (15 protofilaments, as opposed to 11). Mutants of mec-7, a alpha-tubulin encoding gene, have a similar phenotype. We have identified the nature of mec-12 by germline transformation rescue and characterization of a point mutation. Sequence analysis of the mec-12 encoded product (MEC-12) indicates that it corresponds to a novel C. elegans alpha-tubulin. MEC-12 is the only identified C. elegans alpha-tubulin that contains a lysine at position 40, a known site of post-translational acetylation. Some mec-12 mutations eliminate microtubule acetylation as assayed immunocyto-chemically; phenotypic rescue using a MEC-12 variant lacking the lysine-40 showed that acetylation is not required for MEC-12 activity. Although functionally needed only in the touch neurons, mec-12 is expressed in several other neuron types. These results support the notion that tubulin isotype diversity contributes to the formation of distinct classes of microtubules; 15-protofilament microtubule assembly requires MEC-12 alpha-tubulin and MEC-7 beta-tubulin, which are both highly expressed in the touch receptor neurons. MEC-12 is the first reported alpha-tubulin isotype that appears to be required in a single class of neuronal microtubules.

Acetylation↗
PubMed1999

Use of bioluminescent aequorin for the pharmacological characterization of 5HT receptors.

A convenient functional assay for 5HT2a and 5HT2c receptors is reported utilizing the bioluminescent aequorin to detect intracellular calcium changes. Using this assay, the pharmacological properties of many 5HT ligands can be determined in a 96-well format. The data indicate that the aequorin detection method is superior to the inositol phosphate assay with regard to speed and scope. This system is also appropriate for kinetic studies of receptor desensitization. We showed that the human 5HT2c receptor desensitizes in a biphasic manner, with a fast desensitization of approximately 90% of the total response occurring within 15 minutes while the remaining 10% response remains for at least 3 hours.

Aequorin↗
PubMed1999

Temporal patterns of DNA adduct formation and glutathione S-transferase activity in the testes of rats fed aflatoxin B1: a comparison with patterns in the liver.

Fisher-344 male rats were fed 1.6 ppm of aflatoxin B1 (AFB1) continuously and intermittently for several weeks. At various time periods, DNA was isolated from the testes and livers and analyzed for AFB1-DNA adducts. The ability of the testis to detoxify AFB1 was also investigated by the glutathione S-transferase (GST) activity assay and compared with that of the liver. The levels of testicular AFB1-DNA adducts were 2.4 to 8.1 times lower than those of the liver after 4 to 16 weeks of continuous treatment and 2.2 to 46.2 times lower after 8 to 20 weeks of intermittent treatment. The testicular DNA adducts markedly decreased over time. By 16 weeks of continuous and 20 weeks of intermittent exposure, they had decreased 37 and 91%, respectively. In contrast, hepatic AFB1-DNA adducts increased four-fold from 4 to 16 weeks of continuous treatment but increased at a much slower rate after intermittent exposure. In both the liver and testis, significant levels of AFB1-DNA adducts persisted for at least 1 month after ending the treatment, suggesting that this type of lesion was poorly repaired. In control rats, the testis showed significantly higher GST activity than the liver. In treated rats, these differences were significant during the first 12 weeks of continuous treatment but not at later times. Tissue-specific differences such as germ-cell depletion and increased testicular detoxification may play an important role in the observed differential pattern of DNA adduct formation between the testis and liver.

Aflatoxin B1↗
PubMed1998

Design and synthesis of potent, selective inhibitors of endothelin-converting enzyme.

Endothelin-1 is the most potent peptidic vasoconstrictor discovered to date. The final step of posttranslational processing of this peptide is the conversion of its precursor by endothelin-converting enzyme-1 (ECE-1), a metalloprotease which displays high amino acid sequence identity with neutral endopeptidase 24.11 (NEP) especially at the catalytic center. A series of potent and selective arylacetylene-containing ECE-1 inhibitors have been prepared. (S, S)-3-Cyclohexyl-2-[[5-(2, 4-difluorophenyl)-2-[(phosphonomethyl)amino]pent-4-ynoyl]amino] propio nic acid (47), an arylacetylene amino phosphonate dipeptide, was found to inhibit ECE-1 and NEP with IC50 values of 14 nM and 2 microM, respectively. Similarly, (S)-[[1-[(2-biphenyl-4-ylethyl)carbamoyl]-4-(2-fluorophenyl)but-3- yny l]amino]methyl]phosphonic acid (56), an arylacetylene amino phosphonate amide, had IC50's of 33 nM and 6.5 microM for ECE-1 and NEP, respectively. Slight modification of the aryl moiety was found to have dramatic effects on ECE-1/NEP selectivity. The 2-fluoro dipeptide analogue, (S, S)-2-[[5-(2-fluorophenyl)-2-[(phosphonomethyl)amino]pent-4-ynoyl]+ ++amin o]-4-methylpentanoic acid (40), showed a 72-fold selectivity for ECE-1 over NEP, while the 3-fluoro dipeptide analogue, (S, S)-2-[[5-(3-fluorophenyl)-2-[(phosphonomethyl)amino]pent-4-ynoyl]+ ++amin o]-4-methylpentanoic acid (22), was equipotent for ECE-1 and NEP. Several of these inhibitors were shown to be potent in blocking ET-1 production in vivo as demonstrated by the big ET-1-induced pressor response in rats. These potent inhibitors are the most selective for ECE-1 reported to date and are envisaged to have a variety of therapeutic applications.

Acetylene↗
PubMed1998

Sulfonamide-based hydroxamic acids as potent inhibitors of mouse macrophage metalloelastase.

The structural requirements of sulfonamide-based hydroxamic acid 1 for inhibition of macrophage metalloelastase (MME) were investigated. A short aliphatic group at the R2 position together with an aromatic group at the R3 position significantly improved the inhibitory activity. Compounds 32, 34, and 40 were the most potent inhibitors of MME with IC50 values between 5 and 6 nM.

Animals↗
PubMed1998

Production of leptin in Escherichia coli: a comparison of methods.

A procedure is described for gram-scale refolding of Escherichia coli-derived human leptin inclusion bodies. Refolding was achieved by gradually reducing denaturant using a diafiltration method. Refolded leptin is characterized by in vivo modulation of food intake, reduction in body weight, and lowering of insulin and glucose levels in ob/ob mice. In addition, refolded leptin is characterized by radioimmunoassay (RIA) and activation of the leptin receptor in a cell-based assay. For comparison we also refolded leptin by a simple dilution method and produced periplasmic derived leptin, which did not require ex vivo folding. Leptin produced by these three methods and leptin obtained from commercial sources were compared using the RIA and the cell-based assay and appeared to be of comparable quality and potency.

Animals↗
PubMed1997

Suppression of substance P biosynthesis in sensory neurons of dorsal root ganglion by prodrug esters of potent peptidylglycine alpha-amidating monooxygenase inhibitors.

Substance P as well as many other neuropeptides are synthesized as glycine-extended precursors and converted to the biologically active C-terminal amides by posttranslational modification. The final step of posttranslational processing is catalyzed by peptidylglycine alpha-amidating monooxygenase (PAM). In a previous study, N-substituted homocysteine analogs were found to be potent inhibitors of PAM partially purified from conditioned medium of cultured rat medullary thyroid carcinoma CA-77 cells. These compounds, however, were only modest inhibitors of substance P production in cultured dorsal root ganglion cells, possibly because of poor cell penetration. Several ester derivatives of hydrocinnamoyl-phenylalanyl-homocysteine, one of the most potent PAM inhibitors, were prepared to increase the intracellular accessibility of these compounds. Hydrocinnamoyl-phenylalanyl-(S-benzoyl-homocysteine) benzyl ester was identified as the most potent compound, inhibiting substance P biosynthesis in dorsal root ganglion cells with an IC50 of 2 microM. Inhibition of PAM resulted in a concomitant increase in the glycine-extended substance p (substance P-Gly) precursor peptide. In the presence of 3 microM benzyl ester derivative, the intracellular substance P-Gly level was 2.4-fold higher while the substance P level was 2.1-fold lower than the corresponding peptides in control cells. These results suggest that PAM inhibition represents an effective method for suppression of substance P biosynthesis and, therefore, may have therapeutic utility in conditions associated with elevated substance P levels. Furthermore, PAM inhibition may also prove useful in decreasing other amidated peptides.

Animals↗
PubMed1997

Interferon regulatory factor 1 induces the expression of the interferon-stimulated genes.

The interferon regulatory factor 1 (IRF-1) is a positive transcriptional regulatory protein which acts in the interferon signal transduction pathway to activate the transcription of the type I interferon genes by binding to the PRDI response element. The aim of this study was to explore the role of IRF-1 in regulating the expression of other interferon-stimulated genes in the interferon signal transduction pathway. A transient transfection assay was used to show that IRF-1 induced the expression of interferon-stimulated genes. The induction was a direct result of IRF-1 binding to the promoters of the interferon-stimulated response element (ISRE). The levels of endogenous mRNA of two interferon-stimulated genes, 6-16 and 9-27, were increased in cells containing increased levels of IRF-1. In addition, IRF-1 activates the expression of IRF-2, a negative regulator of the type I interferon genes themselves. Two sequences were found in the IRF-2 promoter which were the binding sites for IRF-1. Mutations in the oligonucleotide sequences of these sites could abolish the binding of the IRF-1. These data suggested that IRF-1 not only plays an important role in the induction of type I interferon genes, but also in the activation of interferon-stimulated genes.

Base Sequence↗