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At least 127 records · Page 7Linked to original sources
PubMed1999

Regeneration of a germinal layer in the adult mammalian brain.

Neuronal precursors reside in the subventricular zone (SVZ) of adult mammals. This region is composed of a network of chains of migrating neuroblasts ensheathed by astrocytes and juxtaposed by clusters of immature precursors (type C cells). Here we show that after antimitotic treatment with cytosine-beta-D-arabinofuranoside, neuroblasts and type C cells are eliminated but some astrocytes remain. Remarkably, the SVZ network rapidly regenerates. Soon after cytosine-beta-D-arabinofuranoside treatment astrocytes divide. Two days later, type C cells reappear, followed at 4.5 days by migrating neuroblasts. By 10 days the SVZ network is fully regenerated, and the orientation and organization of chains of migrating neuroblasts resemble that of normal mice. This regeneration reveals an unexpected plasticity in the adult central nervous system and should provide a model system to study the early stages of neurogenesis in the adult brain.

Animals↗
PubMed1997

Short-chain fatty acids promote the migration of colonic epithelial cells in vitro.

BACKGROUND & AIMS: Repair of colonic epithelial erosions requires cell migration. This study aimed to examine the effects of physiologically relevant short-chain fatty acids on migration in colonic epithelial cell lines. METHODS: Butyrate, propionate, and acetate were added to confluent monolayers of LIM1215 colon cancer cells after wounding. Migration in circular wounds was assessed after 24 hours. RESULTS: The migration of LIM1215 cells was stimulated in a concentration-dependent manner by all short-chain fatty acids. In four experiments, 2 mmol/L butyrate, 8 mmol/L propionate, and 16 mmol/L acetate induced 112.6% +/- 6.7%, 98.5% +/- 5.4%, and 63.4% +/- 7.2% (mean +/- SEM) stimulation above control migration, respectively. Their effects were additive at submaximal concentrations and reversible. Butyrate also stimulated migration in two other colon cancer cell lines, Caco-2 and LIM2405. However, butyrate failed to stimulate the migration of nongastrointestinal and nonepithelial cell lines. The stimulatory effect of butyrate required protein and RNA synthesis but was independent of cell proliferation, presence of serum, beta-oxidation, transforming growth factor beta, intracellular acidification, and substratum composition. CONCLUSIONS: In wounded in vitro models of colonic epithelium, short-chain fatty acids promote cell migration. If such an effect occurs in vivo, it would have ramifications for the biology and pathobiology of the colonic mucosa.

Animals↗
PubMed1988

Problems in prenatal diagnosis of beta-thalassaemia by fetal blood analysis: beta-chain variant comigrating with gamma chains.

This report describes a couple at risk for beta-thalassaemia in which one spouse was heterozygous for classical high Hb A2 beta-thalassaemia while the other had the compound heterozygous state for beta+-thalassaemia and a beta-chain variant. This variant comigrates on carboxymethyl-cellulose columns (CMC) with gamma-chains, indicating that globin separation on CMC columns could not have been used for fetal diagnosis. The beta-chain variant migrates separately from the other globin chains on HPLC and the respective abnormal haemoglobin can be separated by isoelectrofocusing. Oligonucleotide hybridization showed that both parents were carriers of the beta+ IVS-1, nt 6 mutation. Prenatal diagnosis was successfully accomplished by oligonucleotide analysis on trophoblast DNA. This case indicates that an Antenatal Service should have alternative methods to CMC columns so as to carry out prenatal diagnosis of beta-thalassaemia in uncommon cases.

Blood Protein Electrophoresis↗
PubMed1997

Oncogenic Ki-ras but not oncogenic Ha-ras blocks integrin beta1-chain maturation in colon epithelial cells.

Human colorectal tumors commonly contain mutations in Ki-ras but rarely, if ever, in Ha-ras. The selectivity for Ki-ras mutations in this tumor was explored using the HD6-4 colon epithelial cell line which contains no ras mutations. After adhesion to an extracellular matrix, HD6-4 cells polarize into columnar goblet cells with distinct apical and basal regions. Stable HD6-4 transfectants were made with mini-gene constructs of the oncogenic cellular Ki-ras4BG12V gene, the oncogenic Ha-rasG12V gene, or mini-gene constructs of wild-type Ki-ras4B as a control. Ki-ras mutations, but not Ha-ras mutations, disrupted colon epithelial cell apicobasal polarity and adhesion to collagen I and laminin. Three Ha-ras transfectants and three Ki-ras transfectants exhibited Ras proteins expressing the Val-12 mutation by Western blotting with pan-rasG12V antibody. Only wild-type Ki-ras transfectant cells and oncogenic Ha-ras transfectant cells synthesized the mature, fully glycosylated forms of beta1 integrin. Instead of the mature integrin beta1-chain, a faster migrating beta1-chain intermediate was detected on the cell surface and in the cytoplasm of the oncogenic Ki-ras transfectants. Expression of the oncogenic Ki-ras gene caused the altered beta1 integrin maturation because phosphorothiolated antisense oligonucleotides to Ki-ras reduced expression of both the mutant Ki-Ras protein and the aberrant integrin beta1-chain and increased expression of the mature integrin beta1-chain. Altered glycosylation generated the new beta1 integrin form since integrin core beta1-chain proteins of the same molecular weight were yielded in Ki-ras, Ha-ras, and control transfectants after removal of sugar residues with endoglycosidase F or following tunicamycin treatment to inhibit glycosylation. The selective effect of oncogenic Ki-ras on beta1 integrin glycosylation was not due to selective activation of mitogen-activated protein kinases because both mutated Ki- and Ha-ras genes activated this pathway and increased cell proliferation. Since blocking the glycosylation of integrin beta1-chain inhibited the adherence, polarization, and subsequent differentiation of colon epithelial cells, the selective effects of the oncogenic cellular Ki-ras gene on integrin beta1-chain glycosylation may account, at least in part, for the selection of Ki-ras mutations in human colon tumors.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed2003

Matrix metalloproteinases process the laminin-5 gamma 2-chain and regulate epithelial cell migration.

Matrix metalloproteinase (MMP)-2 and membrane type 1-MMP can process the laminin-5 (Ln-5) gamma2-chain, revealing a cryptic site inducing epithelial cell migration. We investigated whether other MMPs process the Ln-5 gamma2-chain and related their ability to induce epithelial cell migration. The N-terminal sequences of the MMP-3, -12, -13, and -20 processed 80kDa Ln-5 gamma2x-chains were identical whereas the N-terminus of the 80kDa(MMP-8) Ln-5 gamma2x-chain was not. MMP-3, -13, -14, and -20 induced MCF-7 cell migration over Ln-5 while MMP-8 was a poor inducer of MCF-7 cell migration. In conclusion, several MMPs can process the Ln-5 gamma2-chain and induce epithelial cell migration.

Amino Acid Sequence↗
PubMed2001

Distinguishing migration from isolation: a Markov chain Monte Carlo approach.

A Markov chain Monte Carlo method for estimating the relative effects of migration and isolation on genetic diversity in a pair of populations from DNA sequence data is developed and tested using simulations. The two populations are assumed to be descended from a panmictic ancestral population at some time in the past and may (or may not) after that be connected by migration. The use of a Markov chain Monte Carlo method allows the joint estimation of multiple demographic parameters in either a Bayesian or a likelihood framework. The parameters estimated include the migration rate for each population, the time since the two populations diverged from a common ancestral population, and the relative size of each of the two current populations and of the common ancestral population. The results show that even a single nonrecombining genetic locus can provide substantial power to test the hypothesis of no ongoing migration and/or to test models of symmetric migration between the two populations. The use of the method is illustrated in an application to mitochondrial DNA sequence data from a fish species: the threespine stickleback (Gasterosteus aculeatus).

Animals↗
PubMed1999

Rho-kinase in human neutrophils: a role in signalling for myosin light chain phosphorylation and cell migration.

The role of a Rho-associated coiled-coil forming kinase in migration of neutrophils has been investigated. Rho-associated coiled-coil forming kinase I was expressed in human neutrophils. Chemotactic peptide led to a Rho-associated coiled-coil forming kinase-dependent increase in phosphorylation of myosin light chain. This was determined with the help of an antibody directed against serine 19-phosphorylated myosin light chain and an inhibitor of Rho-associated coiled-coil forming kinase (Y-27632). Y-27632 suppressed myosin light chain phosphorylation and chemotactic peptide-induced development of cell polarity and locomotion with similar potency (ED50 0.5-1.1 microM). The data strongly suggest that a Rho-associated coiled-coil forming kinase isoform, activated in human neutrophils exposed to chemotactic peptide, is important for motile functions of these cells.

Amides↗
PubMed2001

Transplantation of mammalian olfactory progenitors into chick hosts reveals migration and differentiation potentials dependent on cell commitment.

In vertebrates, interneurons of the olfactory bulb are continuously generated postnatally and throughout life at the subventricular zone of the forebrain. From there, the neuronal progenitors migrate tangentially in a typical chain-like structure to the olfactory bulb in which they differentiate as interneurons. We have used a mouse/chick xenograft strategy to explore the migration and differentiation potential of the mouse olfactory progenitors in a heterochronic and heterotypic environment. We compared the migration of primary cells derived from the subventricular zone of adult or newborn lateral ventricule with the behavior of in vitro amplified cells derived from the same structures. We show that in the chick environment, olfactory bulb progenitors from newborn brain tissue perform chain migration along the neural crest cell routes, whereas grafted neurosphere-derived-cells migrate as isolated cells. These results, together with in vitro observations, allow us to propose that neuronal chain migration is a community effect independent of environmental cues but which is closely regulated by the differentiation program of the cells. We established that the progenitor cells performing chain migration are already committed, while neurosphere-derived-cells are able to integrate and differentiate as components of the peripheral nervous system.

Age Factors↗
PubMed1986

Osteogenesis imperfecta type IV: evidence of abnormal triple helical structure of type I collagen.

Skin fibroblasts from a patient with mild osteogenesis imperfecta (OI) type IV synthesize two populations of type I procollagen molecules. One population contains pro alpha 1(I) and pro alpha 2(I) chains that migrate normally in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and a second population contains only slower migrating pro alpha 1(I) and pro alpha 2(I) chains. The total amount of type I procollagen made by OI cells and the ratio of pro alpha 1(I):pro alpha 2(I) is normal. When labeled under conditions that inhibit post-translational modification of pro alpha chains, the OI cells produce only single populations of pro alpha 1(I) and pro alpha 2(I) chains indicating that the apparent increased molecular weight of some OI pro alpha chains is due to excessive post-translational modification rather than peptidyl insertions. Peptide maps indicate that excessive post-translational modification occurs along the entire triple helical segment of some alpha 1(I) and alpha 2(I) chains produced by OI cells. The effect of the mutation is to lower the melting temperature of the molecules containing slow migrating alpha 1(I) and alpha 2(I) chains to 39.5 degrees C (compared to 41.5 degrees C for control), and to delay secretion of the over-modified type I procollagen from OI cells. These data are consistent with a mutation near the carboxyl-terminal end of the triple helical domain which delays triple helical formation and renders all chains available for further post-translational modification amino-terminal to the mutation. Such alterations in triple helical structure, thermal stability, and secretion previously associated only with the lethal OI type II phenotype are thus also seen in the mild OI type IV phenotype.

Cells, Cultured↗
PubMed1992

Effect of medium- and long-chain triglycerides on human neutrophil migration.

There is some controversy concerning the effect of intravenous long-chain triglyceride (LCT) emulsions on the phagocytic system and little is known about the effect of medium-chain triglyceride (MCT)-containing emulsions. We evaluated the chemotaxis and random migration of human neutrophils from 18 healthy adults after preincubation with the following fat emulsions: LCT, MCT and a mixture of 50% MCT and 50% LCT (MCT/LCT). Leukocyte-rich plasma (4 x 10(6) cells/ml) was diluted 4:1 (v/v) with commercial fat emulsions (LCT, MCT, or MCT/LCT, 1:1) or saline and tumbled at 20 cycles/min for 30 min at 37 degrees C. The final composition of the emulsion was 20 mg/ml fat, 0.24% egg yolk lecithin, and 0.5% glycerol and the dispersion was made isotonic by adding NaCl. In a second set of experiments, the LCT and MCT concentrations were adjusted to be equimolar. Leukocyte viability was > or = 95% after exposure to the treatment with fat emulsions. For emulsions with the same weight of each fat, random migration and chemotaxis of neutrophils were unaffected by the LCT emulsion but there was a significant decrease in both chemotaxis and random migration in MCT-(79 and 74%) or MCT/LCT-treated (60 and 56%) neutrophils. Similar results were obtained when LCT and MCT were equimolar. These results demonstrate an inhibitory effect of MCT on two human neutrophil functions which may be dose dependent.

Adult↗
PubMed1995

Domain-specific antibodies against the B2 chain of laminin inhibit neuronal migration in the neonatal rat cerebellum.

Although the spatial and temporal patterns of neuronal migration have been analyzed in great detail, little direct evidence is available as to what extracellular matrix molecules are involved. Because there is indirect evidence implicating the extracellular matrix protein laminin in neuronal migration, we investigated the effects of antibodies against a synthetic peptide derived from a neurite outgrowth domain of the B2 chain of laminin on neuronal migration in living cerebellar slices. We show by using infrared video microscopy that divalent Fab2 fragments of these antibodies inhibit granule neuronal movement in living slices of (P8) rat cerebellum. This inhibition of neuronal movement manifests itself by cessation of both radial and horizontal translocations of nuclei inside the granule neuronal processes. Fab2 fragments of antibodies against the intact (native) laminin molecule or Fab2 fragments from the preimmune serum do not affect nuclear translocation. Immunocytochemistry shows binding of the divalent Fab2 fragments of the B2 chain-specific antibodies to the Purkinje and Bergmann glial cell areas, and as punctate deposits in between the cells of the external granule cell layer. Native laminin antibodies bind to the basement membranes, and binding of the Fab2 fragments from the preimmune sera cannot be demonstrated. These results indicate that neuronal migration in the postnatal rat cerebellum in vivo involves nuclear translocation that can be inhibited by antibodies against a neurite outgrowth domain of the B2 chain of laminin. Thus, migration of cerebellar granule neurons may depend on the interaction between a neurite outgrowth domain of the B2 chain of laminin and neuronal cytoskeleton involved in nuclear movement.

Animals↗
PubMed2000

[Cp2V] migration along an octatetrayne chain: from the monometallic complex (Cp2V(2-4eta-tBuC triple bond C2-C triple bond CC triple bond CtBu)] to the dimetallic complex.

The oxidative addition of one equivalent of [Cp2V] (4) to the tetrayne ligand tBuC triple bond CC triple bond CC triple bond CC triple bond CtBu (5) gives the monometallic complex [Cp2V(3-4eta-tBuC triple bond C-C2-C triple bond CC triple bond CtBu)] (7). Compound 7 reacts further with a second equivalent of [Cp2V] to give the dimetallic complex [(Cp2V)2(1-2eta:7-8eta-tBuC2-C triple bond CC triple bond C-C2tBu)] (8), which involves a shift of the first coordinated [Cp2V] unit from the internal C3-C4 to the external C1-C2 positions on the alkynyl ligand. Compound 8 is also directly obtained by the addition of two equivalents of [Cp2V] to 5. Reversibly, reaction of 8 with 5 leads to 7. This exchange reaction between 7 and 8 by adding successively 5 and 4 has been monitored by EPR spectroscopy. By contrast, the oxidative addition of one or two equivalents of [Cp2V] to the tetrayne ligand PhC triple bond CC triple bond CC triple bond CC triple bond CPh (6) gives the homodimetallic complex [(Cp2V)2(1-2eta:7-8eta-PhC2-CC triple bond CC triple bond C-C2-Ph)] (9). Both monometallic and dimetallic complexes 7, 8, and 9 have been characterized by X-ray diffraction. Magnetic moment measurements for 8 and 9 from 300 to 4 K indicated a weak antiferromagnetic J exchange coupling of -12.5 and -4.1 cm(-1), respectively.

Journal Article↗
PubMed2001

Influence of fluorophor dye labels on the migration behavior of polymerase chain reaction--amplified short tandem repeats during denaturing capillary electrophoresis.

The determination of the length of polymerase chain reaction (PCR)-amplified short tandem repeats (STRs) by denaturing capillary electrophoresis (CE) is a standard procedure for purposes of genotyping. We show that dye-specific mobility anomalies exist for 5'-fluorophor-labelled single-stranded DNA (ssDNA) fragments in CE using the performance-optimized polymer 4 (POP4) buffer sieving matrix, containing the entangled poly(N,N-dimethylacrylamide) polymer, urea, and 2-pyrrolidinone. The dye-specific retardation effects relative to coseparated GeneScan-500 [TAMRA] standard fragments can lead to wrong genotyping, even for allele-specific fragments of pentanucleotide STRs, when comparing the relative calculated sizes of identical fragments, labelled with rhodamine (ROX, TAMRA) or fluorescein dyes (FAM, 6-FAM, HEX, JOE, NED, TET): The size of fluorescein dye-labelled fragments of appr. 100 b in length appears to be smaller by up to 6.5 b. This effect becomes more dramatic with decreasing size: a 6-FAM-labelled 24-mer oligonucleotide appeared to be smaller by 11 b. In contrast, in classical urea/polyacrylamide slab-gel electrophoresis only a small dye-specific retardation of identical fragments is observed. The dye-specific effects are superimposed by weaker size and sequence-dependent anomalies of fragment mobility. Therefore, in denaturing CE the coseparation of a defined allele ladder labelled with the same dye as the unknown sample fragments remains the method of choice for accurate genotyping.

DNA, Single-Stranded↗
PubMed1985

Type IV collagen synthesis by cultured human microvascular endothelial cells and its deposition into the subendothelial basement membrane.

Cultured microvascular endothelial cells isolated from human dermis were examined for the synthesis of basement membrane specific (type IV) collagen and its deposition in subendothelial matrix. Biosynthetically radiolabeled proteins secreted into the culture medium were analyzed by sodium dodecyl sulfate gel electrophoresis after reduction, revealing a single collagenous component with an approximate Mr of 180 000 that could be resolved into two closely migrating polypeptide chains. Prior to reduction, the 180 000 bands migrated as a high molecular weight complex, indicating the presence of intermolecular disulfide bonding. The 180 000 material was identified as type IV procollagen on the basis of its selective degradation by purified bacterial collagenase, moderate sensitivity to pepsin digestion, immunoprecipitation with antibodies to human type IV collagen, and comigration with type IV procollagen purified from human and murine sources. In the basement membrane like matrix elaborated by the microvascular endothelial cells at their basal surface, type IV procollagen was the predominant constituent. This matrix-associated type IV procollagen was present as a highly cross-linked and insoluble complex that was solubilized only after denaturation and reduction of disulfide bonds. In addition, there was evidence of nonreducible dimers and higher molecular weight aggregates of type IV procollagen. These findings support the suggestion that the presence of intermolecular disulfide bonds and other covalent interactions stabilizes the incorporation of the type IV procollagen into the basement membrane matrix. Cultured microvascular endothelial cells therefore appear to deposit a basal lamina-like structure that is biochemically similar to that formed in vivo, providing a unique model system that should be useful for understanding microvascular basement membrane metabolism, especially as it relates to wound healing, tissue remodeling, and disease processes.

Basement Membrane↗
PubMed1983

Genetic characterization of the folding domains of the catalytic chains in aspartate transcarbamoylase.

In Salmonella typhimurium strains which produce high constitutive levels of aspartate transcarbamoylase due to the pyrH700 mutation, the bulk of the carbamoyl phosphate of the cell is consumed for the biosynthesis of pyrimidines. As a consequence, there is little substrate available for arginine synthesis and the cell growth is impeded. Suppression of arginine auxotrophy by mutations which block aspartate transcarbamoylase activity provides a positive selection technique for mutant strains defective in this enzyme activity. A genetic analysis was performed on 29 mutant strains harboring defects in the structural gene pyrB, encoding the catalytic chains of aspartate transcarbamoylase of Escherichia coli. Extracts from 15 strains contained intact, inactive enzyme-like molecules of the same size as the purified wild type enzyme. These same extracts contained a predominant polypeptide chain which migrated electrophoretically at the same rate as catalytic chains from wild type enzyme. In addition to these 15 different missense mutants, 14 others (presumably chain-terminating mutants) were isolated; no polypeptides corresponding to full length catalytic chains were detected in these strains. Based on their reversion and suppression properties, seven were designated as frameshift and two as amber nonsense. A fine structure recombination map of the pyrB locus was constructed from a series of three-factor transductional crosses. Mutational sites were correlated with regions in the polypeptide sequence by relating their map positions to that of mutation pyrB231 which results in an amino acid replacement at position 128. Moreover, since recent crystallographic studies indicate that residue 128 is located near the junction between the NH2- and COOH-terminal folding domains, the mutational sites can be placed within either of these two regions of tertiary structure. Interallelic complementation experiments showed four units of complementation. Those defining the alpha and beta units were missense mutants with their mutational sites in the NH2- and COOH-terminal domains, respectively. The mutants determining the delta and gamma units involved premature polypeptide chain termination and their mutational sites were correlated with distal regions of the two respective domains. Several mutants of the chain-terminating type failed to complement members of more than one unit. Possible effects of the various mutations and their implications for mechanisms of complementation and enzyme activity are presented.

Aspartate Carbamoyltransferase↗
PubMed1997

Cellular composition and three-dimensional organization of the subventricular germinal zone in the adult mammalian brain.

The adult mammalian subventricular zone (SVZ) contains stem cells that give rise to neurons and glia. In vivo, SVZ progeny migrate 3-8 mm to the olfactory bulb, where they form neurons. We show here that the SVZ of the lateral wall of the lateral ventricles in adult mice is composed of neuroblasts, glial cells, and a novel putative precursor cell. The topographical organization of these cells suggests how neurogenesis and migration are integrated in this region. Type A cells had the ultrastructure of migrating neuronal precursors. These cells were arranged as chains parallel to the walls of the ventricle and were polysialylated neural adhesion cell molecule- (PSA-NCAM), TuJ1- (beta-tubulin), and nestin-positive but GFAP- and vimentin-negative. Chains of Type A cells were ensheathed by two ultrastructurally distinct astrocytes (Type B1 and B2) that were GFAP-, vimentin-, and nestin-positive but PSA-NCAM- and TuJ1-negative. Type A and B2 (but not B1) cells incorporated [3H]thymidine. The most actively dividing cell in the SVZ corresponded to Type C cells, which had immature ultrastructural characteristics and were nestin-positive but negative to the other markers. Type C cells formed focal clusters closely associated with chains of Type A cells. Whereas Type C cells were present throughout the SVZ, they were not found in the rostral migratory stream that links the SVZ with the olfactory bulb. These results suggest that chains of migrating neuroblasts in the SVZ may be derived from Type C cells. Our results provide a topographical model for the adult SVZ and should serve as a basis for the in vivo identification of stem cells in the adult mammalian brain.

Animals↗