Biomedical subjects

Cecilio J Vidal

Publications and source records attributed to Cecilio J Vidal.

15 recordsLinked to original sources

PubMed2006

Butyrylcholinesterase activity and molecular components in thymus of healthy and merosin-deficient Lama2dy mice.

The laminin-alpha2 chain, referred to as merosin, forms part of the laminin-2 heterotrimer (alpha2beta1gamma1), which is principally expressed in the basement membrane of muscle. Nearly half of patients suffering from congenital muscular dystrophy (CMD) have abnormalities in the laminin-alpha2 chain (LAMA2) gene, and the merosin-deficient Lama2dy mouse shows CMD. The expression of merosin in thymus, the abnormalities in the gland of Lama2dy mice, and the presence of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in thymus prompted us to study the possible effects of the deficiency of merosin on thymus BuChE. We found that, while AChE activity decreased by approximately 50% in merosin-deficient thymus, the deficiency had little effect on BuChE activity. About 65% of thymus BuChE activity was extracted with a saline buffer and 30% with 1% Triton X-100. Sedimentation analyses and phenyl-agarose chromatography showed that thymus contained amphiphilic BuChE monomers (G(1)(A),44%) and dimers (G(2)(A),33%), and hydrophilic tetramers (G(4)(H),23%). Binding assays with various plant lectins revealed differences between the oligoglycans linked to BuChE tetramers and lighter components. The deficiency of merosin had no effect on the biosynthesis of thymus BuChE as judged by the lack of major changes between control and Lama2dy mice thymuses in the distribution of BuChE molecules and the level of lectin binding. The detoxifying action of BuChE, its role as a backup to AChE, and the relevance of the cholinergic dialogue between T cells and stromal cells for T lymphocyte proliferation, maturation and survival support a physiological function for BuChE in thymus.

Animals↗

PubMed2006

Thymus acetylcholinesterase activity is reduced in mice with congenital muscular dystrophy.

Lama2dy mice constitute an animal model for congenital muscular dystrophy (CMD) by merosin (laminin alpha2-chain) deficiency. This pathology affects the properties of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) of mouse skeletal muscle and nerves (Moral-Naranjo et al., 1999, 2002). AChE and BChE are involved through catalytic and noncatalytic actions in multiple processes, such as hydrolysis of acetylcholine (ACh), morphogenesis, hematopoiesis, and tumorigenesis (Soreq and Seidman, 2001). AChE and BChE molecules can be globular (G1, G2, and G4) or asymmetric forms (A4, A8, and A12) (Massoulié, 2002), and G molecules can show amphiphilic (detergent-interacting, GA) or hydrophilic (GH) behavior. AChE catalytic subunits are encoded by three mRNAs (T, H, or R) generated by alternative splicing. The presence of AChE in lymphoid tissues (Rossi et al., 1991; Nieto-Cerón et al., 2004), the role of immune responses in muscular dystrophy (Spencer and Tidball, 2001), the abnormalities of Lama2dy thymus (Magner et al., 2000), and the role of ACh in thymocyte function (Kawashima and Fujii, 2000) prompted us to investigate thymus AChE and the possible effect of merosin deficiency on it.

Acetylcholinesterase↗

PubMed2006

Acetyl- and butyrylcholinesterase activities decrease in human colon adenocarcinoma.

Apart from the hydrolysis of acetylcholine (ACh), acetyl- (AChE) and butyrylcholinesterase (BChE), through noncatalytic mechanisms, intervene in hematopoiesis, morphogenesis, and neurogenesis (Layer and Willbold, 1995; Soreq and Seidman, 2001). Cholinesterase (ChE) molecules occur as globular (G1, G2, and G4) and asymmetric (A4, A8, and A12) forms (Legay, 2000; Massoulié, 2002). The G species might display amphiphilic (GA) or hydrophilic (GH) properties (Perrier et al., 2002). The involvement of ChEs in tumorigenesis is supported by the measurement of ChE activity in tumors (García-Ayllón et al., 2001; Ruiz-Espejo et al., 2003), the amplification of ChE genes in leukemias and ovarian tumors, and the relationship between the expression of AChE and the aggressiveness of astrocytomas(Perry et al., 2002). This research was undertaken to determine whether ChE activity is altered in gut carcinomas.

Acetylcholinesterase↗

PubMed2005

Acetylcholinesterase biogenesis is impaired in lung cancer tissues.

Studies cited by Cowan et al. [J. Appl. Toxicol. 23, 177 (2003)] indicate existence of inflammatory and cholinergic pathways in both nerve agents and sulfur mustard (HD) injury. Increase in AChE synthesis and neurite extension was noted after exposure to HD [K.W. Lanks et al., Exp. Cell Res. 355 (1975)]. Moreover, anti-inflammatory drugs reduce the dermal, respiratory and ocular damage caused by exposure to HD. On the other hand, recent studies have noted the involvement of neuro-inflammatory processes during exposure to the nerve agents sarin or soman [Cowan et al., 2003]. The use of various anti-inflammatory drugs in addition to the classical antidotal drugs (e.g. atropine and oximes) caused decrease in certain toxic symptoms and inflammation-induced brain damage. Our new bifunctional drugs (Scheme 1) are based on CNS-permeable molecular combination of pseudo-reversible AChE inhibitor (pyridostigmine, PYR) coupled via a hydrophobic spacer (octyl or decyl hydrocarbon chain) to a non-steroidal anti-inflammatory drug (NSAID) such as Ibuprofen or Diclofenac (Scheme 1). This study evaluates the efficacy of certain bifunctional compounds against HD and soman poisoning in mice in vivo.

Acetylcholinesterase↗

PubMed2005

Cholinesterase activity and enzyme components in healthy and cancerous human colorectal sections.

The effect of cancer on acetyl- (AChE) and butyrylcholinesterase (BuChE) activities of human gut was investigated. ChE activity was measured in 55 paired samples of healthy and malignant colon, sigmoid colon and rectum. Cancer decreases the mean AChE activity value from 2.17 +/- 1.07 to 1.40 +/- 0.89 mU/mg (p < 0.001), and BuChE activity from 4.16 +/- 2.41 to 1.65 +/- 0.87 mU/mg (p < 0.001). AChE monomers and dimers (light forms), and less asymmetric and tetrameric variants (heavy forms) were identified in gut. The proportions of the heavy species dropped in malignant colon. Since muscarinic stimulation is needed for human colon cancer cell proliferation, the fall of ChE activity in neoplastic colon, with the increased availability of acetylcholine, may increase tumour growth.

Cholinesterases↗

PubMed2005

The increased ecto-5'-nucleotidase activity in muscle, heart and liver of laminin alpha2-deficient mice is not caused by an elevation in the mRNA content.

We have previously shown that mouse muscle and liver contain catalytically active and inactive ecto-5'-nucleotidase (eNT) variants and that eNT activity in these tissues increases in laminin alpha2 (merosin)-deficient Lama2dy mice. These results prompted us to study whether: (1) the increase of eNT activity depends on the change in the content of merosin between healthy and dystrophic organs; (2) the active and inactive eNT variants arise from the same or distinct mRNAs; (3) the enhancement of the activity is caused by an increase in the eNT mRNA content. Compared to healthy organs, the activity in dystrophic organs increased four-fold in muscle, 1.7-fold in liver, 1.4-fold in heart and not at all in kidney and lung. The level of immunolabelled eNT protein per unit of activity suggested a similar ratio of inactive to active eNT in healthy liver, kidney, heart and muscle, which increased greatly in lung. The size of the eNT subunit in liver, kidney, heart and muscle (72 kDa) decreased to 66 kDa in lung. The identification of a single RT-PCR product suggested that active and inactive eNT arise from the same mRNA and are generated by a differential post-translational processing. Compared to the content in muscle, the amount of eNT mRNA was 12-fold higher in liver and kidney, eight-fold in heart and five-fold in lung. The relative content of eNT mRNA was unaffected by the deficiency of merosin.

5'-Nucleotidase↗

PubMed2005

Cholinesterase activity of human lung tumours varies according to their histological classification.

The probable involvement of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) in cancer and the relevance of cholinergic responses for lung cancer growth prompted us to study whether cholinesterase activity of human lung is altered by malignancy. Surgical pieces of non-small lung carcinomas (NSLC) and their adjacent non-cancerous tissues (ANCT) were analysed for AChE and BChE activities. AChE activity in adenocarcinoma (AC) was 7.80 +/- 5.59 nmol of substrate hydrolysed per min and per mg of protein (mU/mg), the same as in their ANCT (8.83 +/- 4.72 mU/mg; P = 0.823); in large cell carcinoma (LCC), 7.52 +/- 3.32 mU/mg, approximately 50% less than in their ANCT (15.39 +/- 5.66 mU/mg; P = 0.043); and in squamous cell carcinoma (SCC), 1.39 +/- 0.58 mU/mg, 80% less than in ANCT (6.08 +/- 2.88 mU/mg; P = 0.003). BChE activity was 5.85 +/- 3.20 mU/mg in AC and 9.56 +/- 3.38 mU/mg in ANCT (P = 0.022); 2.94 +/- 2.01 mU/mg in LCC and 6.50 +/- 6.63 mU/mg in ANCT (P = 0.068); and 4.49 +/- 2.30 mU/mg in SCC and ANCT 6.56 +/- 4.09 mU/mg (P = 0.026). Abundant AChE dimers and fewer monomers were identified in lung and, although their distribution was unaffected by cancer, the binding with concanavalin A revealed changes in AChE glycosylation between SCC and their ANCT. The fall in BChE activity affected all molecules, with a strong decrease of the amphiphilic tetramers. Western blotting revealed protein bands with the expected mass of the principal AChE subunits, and the deeper intensity of the protein signal in SCC than in healthy lung, in lanes loaded with the same units of AChE activity, supported an augment in the amount of AChE protein/unit of AChE activity in SCC. The increased availability of acetylcholine in neoplastic lung, resulting from the fall of cholinesterase activity, may enhance cholinergic signalling and contribute to tumour progression.

Acetylcholine↗

PubMed2005

Expression of cholinesterases in brain and non-brain tumours.

Although the involvement of cholinesterases (ChEs) in the removal of acetylcholine (ACh) at cholinergic synapses is firmly established, there is evidence to suggest that acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) take part in several cellular processes. The early expression of ChE genes during embryonic development and their role in morphogenesis and apoptosis have been explained on the basis of the non-cholinergic actions of ChEs. In addition, the effects of AChE and BuChE, their inhibitors and antisense oligonucleotides in proliferating cellular systems, together with the mitogenic actions of ACh, support a role for ChEs in cell cycle control. The anomalous expression of ChEs may increase cell proliferation and contribute to cancer growth or development. The aim of this report is to compile the available information on ChEs in cancerous tissues in order to stimulating the research to clarify the molecular mechanisms by which ChEs may participate in cancer. Future investigations may throw light into this intriguing issue which will be of benefit to humankind.

Animals↗

PubMed2005

Muscular dystrophy by merosin deficiency decreases acetylcholinesterase activity in thymus of Lama2dy mice.

Half of congenital muscular dystrophy cases arise from laminin alpha2 (merosin) deficiency, and merosin-deficient mice (Lama2dy) exhibit a dystrophic phenotype. The abnormal development of thymus in Lama2dy mice, the occurrence of acetylcholinesterase (AChE) in the gland and the impaired distribution of AChE molecules in skeletal muscle of the mouse mutant prompted us to compare the levels of AChE mRNAs and enzyme species in thymus of control and Lama2dy mice. AChE activity in normal thymus (mean +/- SD 1.42 +/- 0.28 micromol acetylthiocholine/h/mg protein, U/mg) was decreased by approximately 50% in dystrophic thymus (0.77 +/- 0.23 U/mg) (p = 0.007), whereas butyrylcholinesterase activity was little affected. RT-PCR assays revealed variable levels of R, H and T AChE mRNAs in thymus, bone marrow and spinal cord. Control thymus contained amphiphilic AChE dimers (G2A, 64%) and monomers (G1A, 19%), as well as hydrophilic tetramers (G4H, 9%) and monomers (G1H, 8%). The dimers consisted of glycosylphosphatidylinositol-anchored H subunits. Western blot assays with anti-AChE antibodies suggested the occurrence of inactive AChE in mouse thymus. Despite the decrease in AChE activity in Lama2dy thymus, no differences between thymuses from control and dystrophic mice were observed in the distribution of AChE forms, phosphatidylinositol-specific phospholipase C sensitivity, binding to lectins and size of AChE subunits.

Acetylcholinesterase↗

PubMed2004

Molecular properties of acetylcholinesterase in mouse spleen.

The presence of acetylcholinesterase (AChE) mRNA and activity in the tissues and cells involved in immune responses prompted us to investigate the level and pattern of AChE components in spleen. AChE activity was higher in mouse spleen (0.46 +/- 0.13 micromol of acetylthiocholine split per hour and per mg protein) than in muscle or heart, but lower than in brain. The spleen was essentially free of butyrylcholinesterase (BuChE) activity. About 40% of spleen AChE was extracted with a saline buffer, and a further 40% with 1% Triton X-100. Sedimentation analyses, the splitting of subunits in AChE dimers, phosphatidylinositol-specific phospholipase C (PIPLC) exposure, and phenyl-agarose chromatography showed that hydrophilic (G1H, 43%) and amphiphilic AChE monomers (G1A, 36%), as well as amphiphilic dimers (G2A, 21%), occurred in spleen. All these molecules bound to fasciculin-2-Sepharose, although the extent of binding was higher for G1H (77%) than for G1A (63%) or G2A (48%) forms. Differences in the extent to which wheat germ lectin (WGA) adsorbed with AChE of mouse spleen and of erythrocyte allowed us to discard the blood origin of spleen AChE activity. A 62 kDa protein was labeled in spleen samples using antibodies against human AChE. The protein was attributed to AChE monomers since its size was the same, regardless of whether disulfide bonds were reduced or not. Since cholinergic stimulation modulates proliferation/maturation of lymphoid cells, AChE may be important for regulating the level of acetylcholine (ACh) in the neighborhood of cholinergic receptors (AChR) in spleen and other lymphoid tissues.

Acetylcholinesterase↗

PubMed2004

Active and inactive ecto-5'-nucleotidase variants in liver of control and dystrophic Lama2dy mice.

The ecto-5'-nucleotidase (eNT) activity and the eNT protein content in liver of normal and merosin-deficient dystrophic Lama2dy mice were studied. After the solubilization procedure, the eNT activity in the final extract was 9.2+/-2.5U/mg (nmol of phosphate released from AMP per min and per mg protein) in normal liver, and it rose to 16.1+/-3.9U/mg (P=0.005) in dystrophic liver. The increase of activity was less pronounced in Lama2dy liver (1.7-fold) than the one reported in muscle (four-fold), which probably reflects the lower content of merosin in liver. Similarly to muscle, liver contained active and inactive eNT, as demonstrated by the higher level of immunoreactive protein in normal than in dystrophic liver in Western blots performed with samples containing the same units of eNT activity. PNGase F digestion decreased the size of liver and muscle eNT from 72 and 69kDa, to 63 and 60kDa. Oligoglycan cleavage did not alter eNT activity or the sedimentation coefficient, revealing that oligosaccharides are not required for catalysis or for maintaining the dimeric structure. The eNT protein content in samples of normal liver decreased by 55 or 80% after the trypsinolysis of native or deglycosylated enzyme, but the activity did not change. Such a high proportion of inactive eNT is unlikely to come from aged enzyme, which suggests the involvement of inactive enzyme in non-catalytic actions.

5'-Nucleotidase↗

PubMed2003

Purification and properties of hydrophilic dimers of acetylcholinesterase from mouse erythrocytes.

Differences in the glycosylation of acetylcholinesterase (AChE) subunits which form the dimers of mouse erythrocyte and a suitable procedure to purify the enzyme by affinity chromatography in edrophonium-Sepharose are described. AChE was extracted ( approximately 80%) from erythrocytes with Triton X-100 and sedimentation analyses showed the existence of amphiphilic AChE dimers in the extract. The AChE dimers were converted into monomers by reducing the disulfide bond which links the enzyme subunits. Lectin interaction studies revealed that most of the dimers were bound by concanavalin A (Con A) (90-95%), Lens culinaris agglutinin (LCA) (90-95%), and wheat germ (Triticum vulgaris) agglutinin (WGA) (70-75%), and a small fraction by Ricinus communis agglutinin (RCA(120)) (25-30%). The lower level of binding of the AChE monomers with WGA (55-60%), and especially with RCA (10-15%), with respect to the dimers, reflected heterogeneity in the sugar composition of the glycans linked to each AChE subunit in dimers. Forty per cent of the amphiphilic AChE dimers lost the glycosylphosphatidylinositol (GPI) and, therefore, were converted into hydrophilic forms, by incubation with phosphatidylinositol-specific phospholipase C (PIPLC), which permitted their separation from the amphiphilic variants in octyl-Sepharose. Only the hydrophilic dimers, either isolated or mixed with the amphiphilic forms, were bound by edrophonium-Sepharose, which allowed their purification (4800-fold) with a specific activity of 7700 U/mg protein. The identification of a single protein band of 66 kDa in gel electrophoresis demonstrates that the procedure can be used for the purification of GPI-anchored AChE, providing that the attached glycolipid domain is susceptible to PIPLC.

Acetylcholinesterase↗

PubMed2003

Breast cancer metastasis alters acetylcholinesterase activity and the composition of enzyme forms in axillary lymph nodes.

Because of the probable involvement of cholinesterases (ChEs) in tumorigenesis, this research was addressed to ascertaining whether breast cancer metastasis alters the content of acetylcholinesterase (AChE) and/or butyrylcholinesterase (BuChE) in axillary lymph nodes (LN). ChE activity was assayed in nine normal (NLN) and seven metastasis-bearing nodes (MLN) from women. AChE and BuChE forms were characterised by sedimentation analyses, hydrophobic chromatography and western blotting. The origin of ChEs in LN was studied by lectin interaction. AChE activity dropped from 21.6 mU/mg (nmol of the substrate hydrolysed per minute and per milligram protein) in NLN to 3.8 mU/mg in MLN (p < 0.001), while BuChE activity (3.6 mU/mg) was little affected. NLN contained globular amphiphilic AChE dimers (G2A, 35%), monomers (G1A, 30%), hydrophilic tetramers (G4H, 8%), and asymmetric species (A4, 23%, and A8, 4%); MLN displayed only G2A (65%) and G1A (35%) AChE forms. NLN and MLN contained G4H (79%), G4A (7%), and G1H (14%) BuChE components. Neither the binding of ChE forms with lectins and antibodies nor the subunit size were altered by metastasis. The higher level of AChE in NLN than in brain and the specific pattern of AChE forms in NLN support its role in immunity. The different profile of AChE forms in NLN and MLN may be useful for diagnosis.

Acetylcholinesterase↗

PubMed2002

Muscular dystrophy with laminin deficiency decreases the content of butyrylcholinesterase tetramers in sciatic nerves of Lama2dy mice.

The Lama2dy mouse, a model of congenital muscular dystrophy (CMD) by merosin deficiency (MCMD), shows muscle degeneration and dysmyelination of peripheral nerves. Although it has been reported that MCMD reduces acetylcholinesterase (AChE) activity of mouse sciatic nerve, no information is available regarding its action on butyrylcholinesterase (BuChE). Amphiphilic BuChE monomers (G(1)(A), 39%), dimers (G(2)(A), 18%), and tetramers (G(4)(A), 33%), along with hydrophilic tetramers (G(4)(H), 10%), were identified in mouse sciatic nerve. It also contained abundant G(4)(A) (75%) and less G(1)(A), G(2)(A), G(4)(H) and A(12) AChE components. In dystrophic nerves, the BuChE activity increased 2-fold but the proportion of the G(4)(A) form dropped from 33% to 10%. AChE activity decreased and the composition of enzyme forms was unaffected. Lectin interaction studies showed that, in contrast to skeletal muscle, the defect of merosin did not greatly alter the glycosylation of nerve cholinesterases. The anomalous synthesis of BuChE forms in dystrophic nerve may be related with peripheral neuropathy of MCMD.

Acetylcholinesterase↗

PubMed2002

Cholinesterase activity and acetylcholinesterase glycosylation are altered in human breast cancer.

Increasing evidence supports the involvement of cholinesterases in tumorigenesis. Several tumour cells show ChE activity, while the acetyl- (AChE) and butyrylcholinesterase (BuChE) genes are amplified in leukemias, ovarian carcinoma and other cancers. ChE activity was measured in 31 samples of tumoral breast (TB) and 20 of normal breast (NB). Despite the wide variations observed, BuChE predominated over AChE both in TB and NB. The mean AChE activity in NB was 1.61 nmol of the substrate hydrolysed per minute and per miligram protein (mU/mg), which rose to 3.09 mU/mg in TB (p = 0.041). The BuChE activity dropped from 5.24 mU/mg in NB to 3.39 mU/mg in TB (p = 0.002). Glycolipid-linked AChE dimers and monomers and hydrophilic BuChE tetramers and monomers were identified in NB and TB, and their proportions were unmodified by the neoplasia. The amount of AChE forms reacting with wheat germ agglutinin (WGA) decreased in TB while that of BuChE species was unaffected, demonstrating that the glycosylation of AChE was altered in TB. The binding of AChE and BuChE with antibodies was unaffected by the neoplasia. The difference in lectin reactivity between erythrocyte and breast AChE, the lack of AChE in blood plasma, and the finding of monomeric BuChE in breast but not in plasma suggest that breast epithelial cells produce AChE for membrane attachment and hydrophilic BuChE for secretion. Several reasons are provided to explain the altered expression of ChEs in breast cancer.

Acetylcholinesterase↗