Biodegradation of DEHP, DBP, and DINP: poorly water soluble and widely used phthalate plasticizers.
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Biodegradation, Environmental↗
Biomedical subjects
D Linnemann
Publications and source records attributed to D Linnemann.
Two types of amyloid precursor protein (APP) mRNA in rat glioma cell lines: upregulation via a cyclic AMP-dependent pathway.
APP is a multifunctional transmembrane glycoprotein and the only known natural source of beta A4 peptide-the major constituent of senile plaques in Alzheimer's disease (AD). The expression and cAMP-dependent regulation of the APP gene were investigated in primary cultures of rat astrocytes and two related glioma cell lines, BT4C and BT4Cn, which exhibit distinct invasive phenotypes. Besides the well-characterized 3.5 kb APP mRNA class, a robust expression of an unusual 2.8 kb APP mRNA class was revealed by Northern blotting in both glioma cell lines, but not in the astrocytes. Low amounts of the 2.8 kb APP mRNA species were also observed in rat liver and occasionally in aged rat brain. The 2.8 kb APP mRNA contained exons 1-18 and may thus be generated by truncation of the 3' untranslated region. For the first time, regulation of the APP gene via a cAMP-dependent mechanism was shown. Exposure to dBcAMP dramatically upregulated the 3.5 and 2.8 kb transcripts in BT4C cells, and, to a lesser extent, in BT4Cn cells where the constitutive expression of the APP gene was much higher. Elucidation of the factors involved in cAMP-dependent induction of APP mRNA in these cells may shed more light on the molecular mechanisms of APP overexpression.
Amyloid beta-Protein Precursor↗
NCAM-antibodies modulate induction of long-term potentiation in rat hippocampal CA1.
The neural cell adhesion molecule (NCAM) probably plays a role in neural plasticity in the adult vertebrate brain. We here present evidence that NCAM may be involved in long-term potentiation (LTP) in the CA1-region of rat hippocampal slices. It is shown that local application of antibodies against NCAM inhibits subsequent LTP-induction. Thus NCAM may be directly involved in the initial phase of LTP-induction. These results have important implications for the possible involvement of NCAM in learning and memory.
2-Amino-5-phosphonovalerate↗
The ability to re-express polysialylated NCAM in soleus muscle after denervation is reduced in aged rats compared to young adult rats.
The neural cell-adhesion molecule, NCAM, contains an unusual homopolymer of sialic acid units, polysialic acid. This carbohydrate seems to be involved in neurite outgrowth, bundling and branching, processes which are important during reinnervation. In aged rats, reinnervation of denervated muscle fibres is incomplete. In this study, age-related changes in the degree of polysialylation of NCAM re-expressed after denervation were examined using a monoclonal antibody recognizing polysialic acid and a polyclonal antibody recognizing NCAM. The results show that, after denervation, the degree of polysialylation on NCAM was clearly reduced in rat soleus muscle of aged, compared to young, adult rats. This age-related change in expression of polysialic acid probably influences the reinnervation process in aged muscle.
Animals↗
Characterization of N-cadherin messenger RNA and polypeptide expression in rat.
The cell adhesion molecule N-cadherin is a member of the cadherin gene superfamily. The protein is involved in morphogenetic processes, including neurite extension. In this study, N-cadherin mRNA and polypeptide expression were investigated in rat brain, liver, muscle, heart, kidney and lung during postnatal development and aging. Six synthetic oligonucleotide probes covering different parts of mouse N-cadherin cDNA all hybridized to 5.2, 4.3-4.4 and 3.5 kb mRNAs in rat tissues. The mRNA pattern differed between tissues and, furthermore, the amount of N-cadherin mRNA and polypeptides in brain, liver and heart was higher than in muscle, kidney and lung. N-cadherin expression decreased slightly during early postnatal development in all tissues, whereas no changes in N-cadherin expression were observed during aging. Antibodies against a fusion protein containing the transmembrane and cytoplasmic sequence of chick N-cadherin were produced. These antibodies, termed anti-N-cad-cyt, were compared to the R-156 antibodies which recognize the 24 C-terminal amino acids of N-cadherin and which have been shown to react with a broad spectrum of cadherins. Using these two antibodies, it was shown that the 130 kDa N-cadherin polypeptide was subject to calcium-dependent cleavage of the cytoplasmic domain. Conversely, in the absence of calcium the polypeptide was cleaved extracellularly, producing two C-terminal fragments of 85 and 95 kDa. A 122 kDa polypeptide was recognized by both antibodies and may be either an alternatively spliced form of N-cadherin or a closely related cadherin.
Aging↗
Expression of amyloid precursor protein (APP) in rat brain and cultured neural cells.
Cell type-specific and developmental patterns of APP expression were investigated in rat brain and cultured neural cells. Nearly all astrocytes were APP-positive, whereas only selected population of neurons appeared to express APP. In these neurons, APP immunoreactivity was preferentially restricted to single processes. mRNAs encoding the major APP isoforms, APP695 and APP770, were co-expressed as 3.4-3.6 kb transcripts both in astrocytes and neurons. In addition, an unusual 2.8 kb mRNA size class was revealed in cultured cerebellar granule neurons by means of the probe recognizing APP770 mRNA. Also, for the first time, APP714 mRNA was detected in rat brain by northern blotting. The steady-state levels of these transcripts were increased from birth up to postnatal day 20, whereas no apparent changes were observed after reaching adulthood. These data hint at the involvement of APP in the major morphogenetic events taking place in rat brain during the first three postnatal weeks.
Age Factors↗
Reexpression of the neural cell adhesion molecule (NCAM) on cardiac myocytes in aging rat heart.
The neural cell adhesion molecules, NCAM, is present in several non-neuronal tissues including heart. Using biochemical methods it has been recently shown that NCAM is expressed on cardiac myocytes in early development. During postnatal development NCAM expression is down-regulated and is restricted to neural components of rat heart. However, in the aged rat heart NCAM expression is increased compared to young adult rats. In the present immunohistochemical study the localization of NCAM in different regions of aging rat heart was investigated. Cardiac myocytes expressed NCAM in newborn rat heart whereas NCAM was absent from myocytes in young adult heart. In aged rat heart, NCAM was reexpressed on cardiac myocytes in the ventricles. Thus, NCAM reexpression may be an element in regenerative processes or alternatively a marker of degenerating or dying myocytes.
Aging↗
Age-related changes in expression of the neural cell adhesion molecule in skeletal muscle: a comparative study of newborn, adult and aged rats.
Neural cell adhesion molecule (NCAM) is expressed by muscle and involved in muscle-neuron and muscle-muscle cell interactions. The expression in muscle is regulated during myogenesis and by the state of innervation. In aged muscle, both neurogenic and myogenic degenerative processes occur. We here report quantitative and qualitative changes in NCAM protein and mRNA forms during aging in normal rat skeletal muscle. Determination of the amount of NCAM by e.l.i.s.a. showed that the level decreased from perinatal to adult age, followed by a considerable increase in 24-month-old rat muscle. Thus NCAM concentration in aged muscle was sixfold higher than in young adult muscle. In contrast with previous reports, NCAM polypeptides of 200, 145, 125 and 120 kDa were observed by immunoblotting throughout postnatal development and aging, the relative proportions of the individual NCAM polypeptides remaining virtually unchanged at all ages examined. However, changes in the extent of sialylation of NCAM were demonstrated. Even though the relative amounts of the various NCAM polypeptides were unchanged during aging, distinct changes in NCAM mRNA classes were observed. Three NCAM mRNA classes of 6.7, 5.2 and 2.9 kb were present in perinatal and young adult skeletal muscle, whereas only the 5.2 and 2.9 kb mRNA classes could be demonstrated in aged muscle. This indicates that metabolism of the various NCAM polypeptides is individually regulated during aging. Alternative splicing of NCAM mRNA in skeletal muscle was studied by Northern blotting using DNA oligonucleotide probes specifically hybridizing to selected exons or exon combinations. Exon VASE, which has previously been shown to be present in both brain and heart NCAM mRNA, was virtually absent from skeletal muscle at all ages studied. In contrast, the majority of NCAM mRNA in postnatal skeletal muscle was shown to contain extra exons inserted between exons 12 and 13. Of the various possible exon combinations at this splice site, the combinations 12-a-AAG-13 and 12-a-b seemed to be prevalent in postnatal skeletal muscle. No significant change in the relative proportion of these two exon combinations occurred during aging. The observed upregulation of NCAM protein in aged muscle supports the assumption that an increasing proportion of muscle fibres are denervated in aged muscle. Selective upregulation of the 5.2 and 2.9 kb mRNA forms have previously been demonstrated in muscle cell lines and in primary cultures of muscle cells during formation of myotubes in vitro, and this switch in NCAM mRNA classes has been suggested to correlate with myogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
Aging↗
Expression of NCAM mRNA and polypeptides in aging rat brain.
In aging brain, degenerative as well as compensatory regenerative processes are believed to occur. The neural cell adhesion molecule NCAM is involved in developmental and regenerative processes in the brain. However, the role of NCAM in aging brain has not been characterized. In this study, the expression of NCAM mRNAs and polypeptides was investigated in aging rat brain. The 7.4 and 6.7 kb NCAM mRNAs were selectively downregulated during postnatal development, and the 5.2 and 2.9 kb NCAM mRNAs were upregulated. However, from postnatal day 40 to old age no change in NCAM mRNA classes was observed. The fraction of NCAM mRNA containing the VASE exon increased postnatally but remained stable during adult life. VASE, which is believed to modulate the binding capacity, seemed to be relatively more abundant in the 7.4 and 6.7 kb NCAM mRNAs, encoding transmembrane NCAM forms, than in the 5.2 and 2.9 kb NCAM mRNAs, coding for glycosyl phosphatidylinositol (GPI) linked NCAM. Conversely, insertion of exons a and AAG between exons 12 and 13, a region containing two fibronectin type III repeats, seemed to be more pronounced in 5.2 and 2.9 kb NCAM mRNAs than in the 7.4 and 6.7 kb mRNAs. During postnatal development an increase in the fraction of 6.7 kb NCAM mRNA containing the exons a and AAG was observed. However, during aging the fraction of NCAM mRNAs containing this exon combination seemed constant. At the protein level, NCAM-A was downregulated both during development and aging. No changes were observed during aging in the composition of soluble NCAM forms in the brain, cerebrospinal fluid or blood plasma. The amount of NCAM in rat brain decreased during postnatal development, but remained at a constant level from postnatal day 40 to old age. To conclude, several changes in NCAM expression occur during early postnatal development emphasizing the important role of this molecule in the morphogenetic processes. During aging, a significant selective downregulation of NCAM-A was observed indicating that in general only minor regenerative processes occur in the brain.
Aging↗
Age-related changes in expression of neural cell adhesion molecule (NCAM) in heart: a comparative study of newborn, adult and aged rats.
The neural cell adhesion molecule (NCAM) has been implicated in cellular interactions involved in cardiac morphogenesis and innervation. In this study, expression of NCAM mRNA and protein was characterized in rat heart during postnatal development and aging (postnatal days 1, 10, 40, 270, and 730). Alternative splicing of NCAM mRNA was analyzed by Northern blotting using DNA oligonucleotide probes designed for demonstration of certain exons or exon combinations. Total NCAM mRNA was downregulated during postnatal development followed by upregulation in the aging heart. Three major NCAM mRNA classes of 6.7, 5.2 and 2.9 kb were expressed in newborn heart in approximately equal proportions. At all other ages, the mRNAs of 5.2 and 2.9 kb were more predominant than the 6.7 kb mRNA. During postnatal development and aging, expression of exon VASE was selectively downregulated in the 6.7 kb NCAM mRNA class, whereas it was clearly detectable in NCAM mRNA classes of 5.2 and 2.9 kb. Insertion of exons a and AAG between exons 12 and 13 was more pronounced in the 5.2 and 2.9 kb NCAM mRNAs than in the 6.7 kb mRNA at all ages. Insertions at the 12/13 junctions decreased in the 6.7 kb mRNA as compared to the 5.2 and 2.9 kb mRNAs during postnatal development. Quantification of NCAM protein by enzyme-linked immunosorbent assay showed that NCAM protein amount decreased from a level of 0.93 microgram NCAM/mg total protein at birth to postnatal day 40 where a level of 0.21 microgram NCAM/mg total protein was found. During aging, NCAM increased again to 0.34 micrograms NCAM/mg total protein at postnatal day 730. NCAM isoforms were studied by immunoblotting analysis. At all ages, the major NCAM isoforms of 190, 145 and 120 kDa were observed in heart, the 145 kDa isoform being predominant.(ABSTRACT TRUNCATED AT 250 WORDS)
Aging↗
[Alzheimer's dementia and amyloid precursor-protein].
Alzheimer's dementia is a severe form of dementia characterised by three neuropathological changes: amyloid plaques, neurofibrillary tangles and neurodegeneration. Several laboratories are involved in studies of the molecular mechanisms underlying development of Alzheimer's disease. As a result of recent research, the main component of amyloid plaques, the beta A4-peptide, is focused upon in hypotheses of the pathogenesis of the disease. The beta A4-peptide is produced by cleavage of the larger amyloid precursor protein (APP). During normal processing of APP, production of beta A4 is hindered and the peptide is therefore presumably a result of alternative cleavage mechanisms. Several factors may increase alternative processing of APP. One possibility is that conformational changes in APP, e.g. induced by mutations in the APP gene, may facilitate alternative cleavage of APP and thereby production of beta A4. This seems to be the case in certain families with familial Alzheimer's disease. Another possibility for increased beta A4 production is excessive expression of APP. This seems to be the case in persons with trisomy 21 who all have neuropathological signs of Alzheimer's disease after the age of 40 years. The gene for APP is localised to the human chromosome 21 and these persons have 1.5 times the normal expression of APP. Furthermore, excessive APP-expression may be induced by an acute phase response in the brain. In vitro studies and neuropathological examinations indicate that alpha 2-macroglobulin in neurons may be induced by cytokines. The protease inhibitor activity of this protein may prevent normal cleavage of APP, thereby resulting in increased alternative cleavage and deposition of beta A4.(ABSTRACT TRUNCATED AT 250 WORDS)
Alzheimer Disease↗
Characterization of NCAM expression and function in BT4C and BT4Cn glioma cells.
The neural cell adhesion molecule, NCAM, plays an important role in cell-cell adhesion. Therefore, we have studied NCAM expression in the glioma cell lines BT4C and BT4Cn. We demonstrate that the 2 cell lines differ in their metastatic ability; while BT4C cells have a very low capacity for producing experimental metastases, that of BT4Cn cells is high. In BT4C cells NCAM is synthesized as 4 polypeptides with Mr's of 190,000, 140,000, 115,000 and 97,000. The 140,000, 115,000 and 97,000 polypeptides are glycosylated and for the 140,000 and 115,000 polypeptides sulfatation is observed. Conversely, no NCAM protein synthesis is observed in BT4Cn cells, even though NCAM mRNA is expressed. Thus, development of an increased metastatic capacity is accompanied by the disappearance of NCAM protein expression in this model system. The functional importance of NCAM expression was studied by a cell-substratum binding assay in which the binding of BT4C and BT4Cn cells to NCAM immobilized to glass was assessed. We found that BT4C cells adhere specifically to NCAM, and that adhesion is inhibited by anti-NCAM Fab'-fragments, while no specific binding of BT4Cn cells to NCAM was observed. The BT4C and BT4Cn cell lines thus constitute an important new model system for the study of tumor invasion and metastasis and of the role of cell adhesion molecules in these processes.
Animals↗
[Cell adhesion--during morphogenesis and in the immune system].
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Animals↗
Cell adhesion molecules in brain.
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Animals↗
Differential expression of cell adhesion molecules in variants of K1735 melanoma cells differing in metastatic capacity.
We have investigated the expression of 2 neural-cell adhesion molecules, NCAM and LI, in K1735-C116 and -MI melanoma cells which differ qualitatively in their metastatic potential, i.e., MI cells are metastatic whereas C116 cells are not. We have found that NCAM in C116 cells are expressed as 2 quantitatively major glycosylated polypeptides with Mr of 145,000 and 120,000 and a minor 190,000 Mr polypeptide, whereas MI cells expressed NCAM as 3 glycosylated polypeptides with MR of 200,000, 140,000 and 120,000. The amount of NCAM in MI cells constituted only 60% of the amount observed in C116 cells. In C116 cells, the 145,000 and 120,000 Mr NCAM polypeptides were sulphated whereas NCAM did not appear to be sulphated in MI cells. No phosphorylation of NCAM in the 2 cell lines was observed. LI was expressed as a phosphorylated glycoprotein with Mr of 210,000 in MI cells whereas no LI expression was observed in C116 cells. LI was not sulphated in MI cells.
Animals↗
Soluble neural cell adhesion molecule in brain, cerebrospinal fluid and plasma in the developing rat.
Neural cell adhesion molecule (NCAM) is a glycoprotein found in neurons, glial cells and muscle cells. In this report we describe the polypeptide composition and amount of soluble NCAM in brain, cerebrospinal fluid (CSF) and plasma in rats of various ages. One-two percent of total NCAM in rat brain was shown to be buffer-soluble. Soluble NCAM in brain, extracted at pH 7.2, constituted an increasing relative proportion of total NCAM during development, whereas the concentration of soluble NCAM in both CSF and plasma decreased in the same period. By radioiodination of brain extracts, CSF, and plasma, soluble NCAM was found to be composed of up to five polypeptides with molecular weights (Mr) of 200,000, 170,000, 150,000, 115,000 and 80,000. The relative composition did not vary significantly with age. The major forms in brain were the polypeptides with Mr of 200,000 and 115,000, whereas in CSF all forms seemed to appear in equal amounts. In plasma the 200,000 and the 170,000 Mr polypeptides were not observed and here the major forms were the 150,000 and 115,000 Mr polypeptides.
Aging↗
Heterogeneity of soluble neural cell adhesion molecule.
Soluble neural cell adhesion molecule (NCAM) from rat brain neuronal cell culture media consists predominantly of a polypeptide of Mr approximately 115,000. Minor amounts of a polypeptide of Mr approximately 180,000 and two inconsistently appearing components of Mr 160,000 and 145,000 are also observed. The Mr 115,000 component is derived from the neuronal membrane NCAM components NCAM-A of Mr 190,000, NCAM-B of Mr 140,000, or both. Thus, as a part of the catabolism of membrane NCAM-A plus -B, a minor fraction is posttranslationally cleaved and recovered in the media as discernible soluble NCAM polypeptides. The half-life of membrane NCAM-A plus -B is less than 24 h. Astrocyte culture media contains a predominant soluble NCAM component of Mr 120,000 derived from membrane-associated NCAM-C. A close comparison of deglycosylated soluble NCAM from astrocyte and neuronal cultures showed a small but consistent difference in Mr, a result suggesting that different NCAM polypeptides are released from the membrane of neurons and astrocytes. In contrast to the Mr 115,000-120,000 NCAM polypeptides, the Mr 180,000 polypeptide from neuronal culture media does not seem to be derived from membrane-attached NCAM and may therefore represent a secreted NCAM isoform.
Animals↗
Cell adhesion molecules in neural development.
Adhesive mechanisms have been implicated in several morphogenetic processes during development. In the last decade several molecules mediating specific adhesion between cells and between cells and their environment have been characterized. Possible roles in morphogenesis have been established for some of these molecules. In the brain, four cell-cell adhesion molecules have been characterized: NCAM, L1, the myelin-associated glycoprotein and N-cadherin. Furthermore, a cell-substrate adhesion molecule, cytotactin, and its proteoglycan ligand have been described. We here review the data on structure, localization, developmental regulation and function of these molecules in brain.
Animals↗