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S Szuchet

Publications and source records attributed to S Szuchet.

At least 19 recordsLinked to original sources
PubMed1992

Oligodendrocyte-substratum adhesion activates the synthesis of specific lipid species involved in cell signaling.

Ovine oligodendrocytes (OLGs) undergo biochemical and morphological changes following attachment to polylysine. Autoradiographs of two-dimensional thin-layer chromatograms of [14C]Gal-labeled OLG cultures revealed that attachment of OLGs to a polylysine substratum and their subsequent morphological differentiation is accompanied by an increased synthesis of multiple forms of galactosylceramide, sulfogalactosylceramide, and both sulfogalactosyl- and galactosyl-diglycerides, together with an array of complex sialoglycosphingolipids, predominantly GM2 ganglioside. As previously reported, overall lipid synthesis measured by [14C]acetate incorporation into glycerophosphatides, sphingomyelin, and neutral lipids also increased dramatically for up to 60 days (last time point examined) following OLG-substratum adhesion, reflecting membrane growth. Attachment was associated with a rapid augmentation in the synthesis of ethanolamine plasmalogen from 12 to 27% within 24 hr to reach a 35% plateau at 30 days and remain constant thereafter. In contrast, the plasmalogen content of phosphatidylcholine remained constant at 3-5%. This rapid increase in lipid synthesis (especially in the ethanolamine plasmalogen content following attachment) closely paralleled increased diacylglycerol (DAG) production and protein kinase C-dependent phosphorylation of both myelin basic protein and 2',3'-cyclic nucleotide phosphohydrolase. Labeling studies indicated that the major source of [3H]arachidonate-labeled DAG following attachment was from phosphatidylinositol turnover (and to a lesser extent phosphatidylcholine) rather than polyphosphoinositides or plasmalogens. Enhanced lipid synthesis is not only required for the production of membranes in these myelin-producing cells but is also a source of second messengers required in the posttranslational modification of key myelin and cellular proteins.

Animals↗
PubMed1992

Myelin palingenesis. 1. Electron microscopical localization of myelin/oligodendrocyte proteins in multilamellar structures by the immunogold method.

The issue of the capacity of mature oligodendrocytes to remyelinate naked axons has not been totally resolved. The impression is that for this to happen, oligodendrocytes have to undergo cell division. We are interested in providing an answer to the question: can oligodendrocytes myelinate more than once? To address this question, we are using a model system consisting of pure cultures of postmyelination oligodendrocytes. We have previously shown that these cells have plasticity and are able to regenerate, but for this to occur they need a signal. In vitro, this signal is provided by interaction with a positively charged substratum and a serum factor(s). Over time in culture, without cell division, oligodendrocytes assemble multilamellar structures that ultrastructurally resemble myelin. We have named this process myelin palingenesis--rebirth--to distinguish it from remyelination. In order to ascertain the significance of the regenerative process observed in vitro and its predictive value as to the capabilities of oligodendrocytes to ensheathe axons, we undertook an ultrastructural and immunocytochemical characterization of the multilamellar membranes to assess whether they contain all the myelin-characteristic proteins, i.e., myelin basic protein, proteolipid protein, 2',3'-cyclic nucleotide 3'-phosphodiesterase and myelin-associated glycoprotein. We have used antibodies against these proteins as immunocytochemical probes in conjunction with the immunogold method at the electron microscopic level. Oligodendrocyte cultures were processed for electron microscopy. Blocks were serially sectioned parallel to the culture plate at 0.09-micron spacings. Staining of cells was done prior to embedding. Analysis of these micrographs brought to light the existence of membrane-membrane interaction between multilamellar structures; this is reminiscent of the interactions observed in situ between myelinated axons. We have found that the membranous structures possess all the myelin-characteristic proteins. Furthermore, based on the accessibility of these proteins to the antibodies, e.g., whether or not permeabilization was required, we can surmise that these proteins are incorporated into the membranous structures with the same orientation as in myelin. These results, in conjunction with our earlier work, show that mature oligodendrocytes stripped of their myelin are able to regenerate and reassemble multilamellar membranes that have all the myelin proteins. It remains to be proven that when provided with axons, these oligodendrocytes will generate typical myelin.

2',3'-Cyclic Nucleotide 3'-Phosphodiesterase↗
PubMed1992

Myelin palingenesis. 2. Immunocytochemical localization of myelin/oligodendrocyte glycolipids in multilamellar structures.

Myelin is a membrane with unique characteristics that set it apart from any other membrane. It has a very high lipid:protein ratio (approximately 75:25) not found in other multilamellar membranes; it has a high content of two glycolipids--galactocerebrosides and sulfatides--which account for 26.5% of its lipids. The physiological role of these glycolipids in myelin and/or oligodendrocytes is unknown, but evidence that Abs directed against them interfere with myelination has been presented. Moreover, one of the early events in the process of oligodendrocyte differentiation prior to myelination is the acquisition of galactocerebroside on their surface. In earlier work, we have demonstrated that adhesion of oligodendrocytes to a positively charged substratum signals the commencement of myelinogenesis. Among the events that take place following adhesion are the rapid synthesis of glycolipids. Since over time in culture, oligodendrocytes elaborate multilamellar membranes that contain all the myelin characteristic proteins, we undertook to investigate whether these structures also accumulate myelin characteristic glycolipids. We have used three monoclonal antibodies--two are directed against galactocerebroside, the third one is an antisulfatide--in conjunction with the immunogold method, at the electron microscopic level, to examine the distribution of these glycolipids in the multilammelar structures that accumulate in long-term oligodendrocyte cultures. Our data show that galactocerebrosides and sulfatides are present in the multilamellar structures. Staining is easily demonstrated on the outermost membrane. However, as was the situation with myelin proteins, detection of these glycolipids on the inner lamellae is only achieved at the expense of destroying the ultrastructure. This is so, independent of whether the structures are compact or not.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Tumor necrosis factor inhibits K+ current expression in cultured oligodendrocytes.

The effects of tumor necrosis factor-alpha (TNF-alpha), a cytokine secreted by activated macrophages, on the electrical membrane properties of cultured adult ovine oligodendrocytes (OLGs) were investigated using the whole-cell voltage-clamp technique. Treatment with recombinant human TNF-alpha (rhTNF) for 24 to 72 hr produces (i) process retraction in some but not all OLGs, (ii) a reduction in the resting membrane potential with no significant change in membrane capacitance or input resistance over control cells and (iii) a decrease in the expression of both the inwardly rectifying and outward K+ current. The magnitude of the membrane potential change as well as K+ current inhibition was larger in cells with retracted processes. The electrophysiological effects of rhTNF were attenuated when rhTNF was neutralized with a polyclonal anti-rhTNF antibody. The binding of rhTNF to its receptor has been reported to increase GTP binding, to increase GTPase activity of a pertussis-sensitive G protein, and to produce an elevation in intracellular cAMP in other cell types. However, pretreatment of OLGs with activated pertussis toxin failed to attenuate or mimic the effects of rhTNF. Chronic exposure of OLGs to the membrane permeant analogue of cAMP, 8-bromo-cAMP, resulted primarily in an inhibition of the inwardly rectifying K+ current, an effect which was less than that produced by rhTNF alone and without any of the associated rhTNF-induced morphological changes. This indicates that the effects of rhTNF cannot be entirely accounted for by an elevation in intracellular cAMP. Cycloheximide (CHX), an inhibitor of protein synthesis, mimicked the effects of rhTNF; however, the effects of rhTNF and CHX were not additive. The finding that both ionic current expression and membrane potential were reduced in cells treated with rhTNF that appeared morphologically normal suggests that abnormal ion channel expression in OLGs precedes and may contribute to eventual myelin swelling and damage.

Animals↗
PubMed1990

Channel-mediated and carrier-mediated uptake of K+ into cultured ovine oligodendrocytes.

Uptake of radioactive K+ by mature ovine oligodendrocytes (OLGs) maintained in primary culture was measured under steady-state conditions, i.e., in cells maintained in a normal tissue culture medium (5.4 mM K+), and in cells after depletion of intracellular K+ to less than 15% of its normal value by pre-incubation in K(+)-free medium. The latter value is dominated by an active, carrier-mediated uptake (although it may include some diffusional uptake), whereas the former, in addition to active uptake, also reflects passive K+ diffusion through ion selective channels and possible self-exchange between extracellular and intracellular K+, which may be carrier-mediated. The total uptake rate was 144 +/- 10 nmol/min/mg protein, and the uptake after K+ depletion was 60 +/- 2 nmol/min/mg protein, much lower rates than previously observed in astrocytes. The uptake into K(+)-depleted cells was inhibited by about 80% in the presence of ouabain (1 mM) and about 30% in the presence of furosemide (2 mM). Activators of protein kinase C (phorbol esters) and cAMP-dependent protein kinase (forskolin) have been shown to alter the myelinogenic metabolism as well as outward K+ current in cultured OLGs. The present study demonstrates that K+ homeostasis in OLGs is modulated through similar second messenger pathways. Active uptake was inhibited by about 60% in the presence of active phorbol esters (100 nM) but was not affected by forskolin (100 nM). Forskolin likewise had no effect on total uptake, whereas phorbol esters caused a much larger inhibition than expected from their effect on carrier-mediated uptake alone, suggesting that channel-mediated uptake was also reduced.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Phosphorylation of myelin basic protein in intact oligodendrocytes: inhibition by galactosylsphingosine and cyclic AMP.

We have previously shown that cyclic AMP (cAMP) inhibits the protein kinase C (PKC)-mediated phosphorylation of myelin basic protein (MBP) in cultured oligodendrocytes (OLGs). Recently, it has been demonstrated that the long chain base sphingosine inhibits PKC by competing PKC effectors (diacylglycerol and phorbol esters) for a binding site on the kinase (Hannun and Bell: Science 235: 670-674, 1987). In this report we define further the mechanism by which cAMP inhibits MBP phosphorylation by comparing the effects of cAMP with that of galactosylsphingosine (psychosine), a potential catabolite of galactocerebroside, the major OLG glycosphingolipid. We identify the consequences of psychosine treatment and PKC down-regulation on OLG morphology and electrophysiology and discuss their relevance. Our results in intact ovine oligodendrocytes are consistent with a mechanism in which cAMP inhibits MBP phosphorylation by interfering with the release of diacylglycerol (DAG) from phosphatidylinositol. First, the effects of cAMP on MBP phosphorylation are reversed with exogenous TPA; and second, cAMP inhibits the incorporation of 1-[14C]arachidonate into DAG and specifically inhibits the turnover (as judged by 32PO4 3-incorporation) of phosphatidylinositol. Psychosine inhibits MBP phosphorylation, and its action can be reversed by TPA suggesting a mechanism of inhibition similar to that described for other systems. In addition, psychosine has profound effects on OLG morphology; it disintegrates OLG processes while leaving the cell soma intact. Stable hyperpolarized resting potentials were obtained following psychosine treatment, but there was a 66% decrease in membrane capacitance indicating a significant decrement in membrane surface area. The morphological changes induced by psychosine are reversible and can be eliminated by removing the drug but not by the addition of TPA. Whether inhibition of PKC by psychosine plays any role in process dissolution remains an unanswered question. However, current evidence suggests that a PKC-independent mechanism may be at play. This investigation in conjunction with our previous work emphasizes a role for the interregulation of protein kinase A (PKA) and PKC in the control of OLG somal vs. myelin components. This may have significant implications for central nervous system myelin assembly.

Animals↗
PubMed1989

Expression and modulation of K+ currents in oligodendrocytes: possible role in myelinogenesis.

We have used whole-cell and single-channel recording techniques to investigate the electrophysiological properties of cultured ovine oligodendrocytes (OLGs). Our studies have led to the following conclusions. (1) Cultured mature OLGs express a variety of voltage-dependent K+ conductances including an outward current that consists of a transient component and a steady-state component, as well as an inwardly rectifying K+ current. (2) These conductances are expressed sequentially as a function of development in culture. The inwardly rectifying K+ current appears later than the outward current. (3) Although process extension may influence the expression of the ion channels, the majority of the K+ channels are located in the soma of OLGs, probably concentrated in the basal plasma membrane. (4) Finally, the activation of K+ channels in OLGs can be inhibited by two distinct second messengers, cAMP acting through protein kinase A and diacylglycerol acting through protein kinase C, the effects of which perhaps converge at the level of a common phosphorylated enzyme or regulatory protein. Both cAMP and diacylglycerol have been implicated as factors important in controlling the induction of a myelinogenic metabolism associated with OLG substratum attachment. Thus, membrane ion channels may provide an important intermediate step linking cellular substratum attachment to the eventual induction of myelinogenesis.

Animals↗
PubMed1988

Plasma membrane of cultured oligodendrocytes: I. Isolation, purification, and initial characterization.

Oligodendrocytes generate myelin as extensions of the plasma membrane. Myelin has been well characterized, yet little is known concerning oligodendrocyte plasma membrane. We have developed a reproducible method for the isolation of an oligodendrocyte plasma membrane-rich fraction (F2.2). F2.2 has a 25-fold enrichment in K+-dependent p-nitrophenyl phosphatase, a plasma membrane marker. Impurities are composed of Golgi elements (8-12%), microsomes (4-6%), and lysosomal membranes (1-5%). Our starting material was oligodendrocytes kept in culture in a nonattached state for 3 to 5 d. After disrupting the cells and removing nuclei (P1), the supernatant (SP1) was fractionated on a self-generating 20% Percoll gradient into three bands: F1, F2, and F3. F1 had only 3% of the applied protein and was not characterized. F2, with 11% of the protein, was fivefold enriched in plasma membrane. F3 had 27% of initial protein; it consisted of a crude mitochondrial and lysosomal fraction. F2 was further purified by first washing it hypotonically, treating it with Mg2+, and then fractionating it on a Ficoll step gradient that yielded F2.2 at the interphase. Morphologically F2.2 comprises 1) membranous sheets, often with more than one membrane in close apposition; 2) membrane vesicles of various sizes and shapes frequently filled with amorphous material; 3) Golgi elements; and 4) unrecognizable profiles. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profile of F2.2 reveals CNPase as a major component in agreement with the high CNPase specific activity (3,860 mumol/mgP/h) found in F2.2. Other significant polypeptides have Mr = 170,000, 135,000, 108,000, 80,000, 53,000, 38,500, 32,000, and 22,200.

Animals↗
PubMed1988

Plasma membrane of cultured oligodendrocytes: II. Possible structural and functional domains.

An oligodendrocyte plasma membrane-rich fraction, F2.2, was resolved by equilibrium density centrifugation on a linear sucrose gradient from 0.5 M to 1.3 M into three fractions, F2.2a, F.2.2b, F2.2c, and a pellet F2.2p. F2.2a and F.2.2b were enriched 1.5-fold relative to F2.2 in plasma membrane markers at the expense of F2.2c and F2.2p, which became correspondingly impoverished. This gave F2.2a and F2.2b a 42-fold and 37-fold enrichment, respectively, in plasma membrane markers relative to the initial cell homogenate. F2.2c had a sevenfold enrichment in a Golgi marker; together with F2.2p, they contained all the Golgi marker initially present in F2.2. Preliminary data indicated that the F2.2-subfractions differed from one another in their molar ratios of cholesterol to phospholipids and protein to lipids but had similar protein profiles when examined by sodium dodecylsulfate-polyacrylamide gel electrophoresis. Their content of fucosylated glycoproteins appeared also to be different. Morphologically, F2.2a and F2.2b were very similar: they contained large membrane vesicles, membrane sheets, and vesicles entrapped within other vesicles. Membrane-membrane interaction was apparent in these fractions. F2.2c had many of the same elements, but most of the membrane structures contained amorphous material. F2.2p differed morphologically from the other fractions in that it had principally electron-dense structures. It is postulated that F2.2a, F2.2b, and perhaps F2.2c represent different domains of oligodendrocyte plasma membrane. Alternatively, these fractions might correspond to the plasma membrane of oligodendrocyte subtypes.

Animals↗
PubMed1988

Plasma membrane of cultured oligodendrocytes: III. Relatedness to myelin.

We have compared highly purified fractions of oligodendrocyte plasma membrane to myelin by one- and two dimensional gel electrophoresis and found them to be distinct. The major myelin proteins--proteolipid protein (PLP), DM-20, and myelin basic protein (MBP), which dominate the sodium dodecyl sulfate polyacrylamide gel electrophoresis pattern of myelin--were minor components of the plasmalemma. However, 2',3', cyclic nucleotide phosphodiesterase (CNPase) and myelin-associated glycoprotein (MAG) were represented equally in both membranes. Labeling the cells with various precursors followed by isolation of plasmalemma revealed that newly synthesized PLP, DM-20, CNPase, and MAG were incorporated into the plasma membrane of "floating" oligodendrocytes (i.e., nonattached to substratum). This was not so with MBP. Nevertheless, scattered patches of MBP were localized on the plasma membrane of intact cells using the immunogold method at the electron microscopic level. The data are consistent with the notion that MBP is not a constituent of the plasma membrane of mature oligodendrocytes (the MBP patches on intact cells are likely remnants from past association with myelin) but is rapidly associated with the plasmalemma of myelinating oligodendrocytes (i.e., attached cells). It is suggested that phosphorylation of MBP provides the triggering signal for plasma membrane association. In order to analyze the minor proteins in myelin and compare them to the plasma membrane by two-dimensional gel electrophoresis, myelin was extracted with chloroform:methanol to remove PLP, DM-20, and MBP. Even in the absence of PLP, DM-20, and MBP the pattern of extracted myelin still differed from that of plasmalemma indicating that their minor protein compositions were not the same. Myelin was characterized by a group of proteins that clustered at pI 5.5-6.5 and Mr 40,000-60,000 of which alpha-tubulins, beta-tubulins, and actin are part: the plasmalemma had tubulins and actin but in different proportions. Our findings indicate that in addition to PLP, DM-20, and MPB, myelin is also enriched relative to the plasmalemma in another group of proteins.

Animals↗
PubMed1988

Forskolin and phorbol esters decrease the same K+ conductance in cultured oligodendrocytes.

Cultured ovine oligodendrocytes (OLGs) express a number of voltage-dependent potassium currents after they attach to a substratum and as they begin to develop processes. At 24-48 hours following plating, an outward potassium current can be identified that represents a composite response of a rapidly inactivating component and a steady-state or noninactivating component. After 4-7 days in culture, OLGs also develop an inward rectifier current. We studied the effects of forskolin and phorbol 12-myristate 13-acetate (PMA) on OLG outward currents. These compounds are known to alter the myelinogenic metabolism of OLGs. PMA, an activator of protein kinase C (PK-C), has been shown to enhance myelin basic protein phosphorylation while forskolin acting on adenylate cyclase, and thereby increasing cAMP, inhibits it. Both forskolin and PMA increase the phosphorylation of 2'3'-cyclic nucleotide phosphodiesterase, an OLG/myelin protein. We found that forskolin decreased the steady-state outward current at 120 mV by 10% at 100 nM, and by 72% at 25 microM from a holding potential of -80 mV. The time course of inactivation of the peak currents was decreased, affecting both the fast and slow time constants. There was no significant change in the steady-state parameters of current activation and inactivation. The effect of forskolin was attenuated when the adenylate cyclase inhibitor adenosine (2 mM) was present in the intracellular/pipette filling solution. The results of PMA experiments were similar to those obtained with forskolin. Whereas the amplitude of the currents in the presence of PMA was reduced by 28% at 1.5 nM and 60% and 600 nM, the decay phase of the peak currents was less affected. The PMA effect could still be seen when the intracellular Ca2+ was reduced to less than or equal to 10 nM with 5 mM BAPTA, but was inhibited when the cells were pre-exposed to 50 microM psychosine, a PK-C inhibitor. It is postulated that the potassium currents in OLG can be physiologically modulated by two distinct second-messenger systems, perhaps converging at the level of a common phosphorylated enzyme or regulatory protein.

Animals↗
PubMed1988

Oligodendrocyte substratum adhesion modulates expression of adenylate cyclase-linked receptors.

The molecular mechanisms of myelin formation/reformation in the central nervous system are unknown. In previous work we have demonstrated that mature oligodendrocytes (OLG) respond to a signal(s), elicited by their adhesion to a substratum, by turning on a myelinogenic metabolism. Events occurring within 24 hr of adhesion include generation of diacylglycerol, activation of protein kinase C, phosphorylation of myelin basic protein, and enhanced synthesis of myelin lipids and proteins. To elucidate the mechanism(s) of signal transduction, we have investigated whether OLG-substratum interaction influences the level of basal cAMP and the expression of receptors coupled to adenylate cyclase. By using ovine brain OLG we have found that adhesion to a polylysine-coated surface for 24 hr increased the basal level of cAMP 2-fold and altered the expression (assessed by cAMP production) of receptors coupled to adenylate cyclase. Isoproterenol (beta-adrenergic agonist) augmented cAMP from 4 to 26 pmol/mg of protein in adhering OLG but had no such effect in nonattached OLG. Adhesion of OLG was accompanied by rapid synthesis of ethanolamine plasmalogen, a class of lipids believed to be associated with beta-adrenergic receptors. Nonattached OLG responded to prostaglandin E1 with only a 3-fold stimulation in their cAMP content; in attached OLG, 6-fold stimulation was observed. In contrast, vasoactive intestinal polypeptide elicited a 3-fold increase in cAMP in nonattached OLG but, following 24 hr of attachment, OLG did not respond to vasoactive intestinal polypeptide. The increase of cellular cAMP levels was accompanied by a 2.5-fold gain in protein kinase A. OLG-substratum adhesion resulted also in phosphorylation of the OLG/myelin protein, 2',3'-cyclic nucleotide 2'-phosphodiesterase, which proved to be a substrate for cAMP and phospholipid-, Ca2+-dependent protein kinases. These findings, in conjunction with our earlier work, implicate cAMP and diacylglycerol in signaling myelinogenesis; they suggest that phosphorylation/dephosphorylation of myelin basic protein and 2',3'-cyclic nucleotide 2'-phosphodiesterase may be key processes in the cascade of events that are initiated by adhesion of OLG to a polylysine surface (possibly acting as a surrogate for axons) and culminate in the reformation of myelin.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1988

A peanut agglutinin binding glycoprotein in CNS myelin and oligodendrocytes.

We isolated and characterized a 120-kd PNA-binding polypeptide from the human CNS. This polypeptide is linked to membranes through a PI linkage. After release from membranes by PLC it measures 105 kd, 30 kd of which appear to be contributed by N-linked carbohydrates. We isolated the polypeptide by the use of PLC and PNA affinity chromatography and used it to raise polyclonal antibodies and to determine the N-terminal sequence. Immunohistochemical and immunochemical studies using these antibodies showed that 120 kdpp is present in both myelin and oligodendrocytes.

Amino Acid Sequence↗
PubMed1988

Voltage-gated potassium currents in cultured ovine oligodendrocytes.

Cultured oligodendrocytes (OLGs) develop processes and form myelin following attachment to a substratum. We applied the whole-cell voltage-clamp technique to identify and characterize the ionic currents of OLGs in culture. Within 2 d after attachment, OLGs extended processes and began to express an outward current that represents a composite response of an inactivating/transient component and a non-inactivating component. The current had a reversal potential of -66 mV and was sensitive to potassium channel blockers. After 4-5 d in culture, the transient component was less prominent, often accompanied by an increase in noninactivating or steady-state outward current. In addition, there was an increase in inward rectifier current. Four of 7 cells that failed to develop processes exhibited only linear high-resistance membranes. We conclude that cultured OLGs express 3 voltage-gated potassium conductances: (1) a transient outward current, (2) a noninactivating outward current, and (3) an inward rectifier current. The sequential appearance of the several currents may relate, at least in part, to process formation.

4-Aminopyridine↗
PubMed1986

Oligodendrocyte adhesion activates protein kinase C-mediated phosphorylation of myelin basic protein.

When isolated adult oligodendrocytes adhere to a substratum myelinogenesis occurs. Investigation of the mechanism by which this happens indicated that the oligodendrocyte-substratum interaction activated protein kinase C-dependent phosphorylation of myelin basic protein and promoted the synthesis of myelin basic protein. In addition, when agents that activate protein kinase C (second messenger diacylglycerol or a tumor-promoting phorbol ester) were added to nonattached oligodendrocytes, they mimicked the influence of the substratum by inducing phosphorylation of myelin basic protein; and reagents that increase cellular adenosine 3', 5'-monophosphate (cyclic AMP) inhibited phosphorylation of myelin basic protein. Thus, at least in vitro, the interaction between oligodendrocytes and the substratum may mediate myelinogenic events, and phosphorylation of myelin basic protein may be an early requirement in the sequence of steps that ultimately results in myelin formation.

1-Methyl-3-isobutylxanthine↗
PubMed1986

Cultured oligodendrocytes. A role for cell-substratum interaction in phenotypic expression.

Oligodendrocytes can be maintained in two states: nonattached; we call these cells B3.f; morphologically they resemble freshly isolated cells; attached; we refer to the latter as B3.fA. Profound morphological, ultrastructural, and biochemical changes take place upon adhesion to a competent surface (Szuchet, S., Yim, S. H., and Monsma, S. (1983) Proc. Natl. Acad. Sci. U. S. A. 80, 7019-7023). Here we present evidence that the transition from B3.f to B3.fA has important consequences for the expression of myelinogenic properties by these cells. We have examined the incorporation of [3H]leucine, [35S]methionine, and [35S]cysteine into polypeptide chains by B3.f and B3.fA cells from 3 days after isolation up to 8 weeks in culture. Specific antisera against myelin and cytoskeletal proteins were used to identify the newly synthesized proteins. Our results indicate that: overall incorporation expressed as cpm/mg of protein remains essentially constant and independent of the state of adhesion or time in culture; B3.f cells keep a low profile in the synthesis of the major myelin proteins but have a high uptake of precursors into 2',3'-cyclic nucleotide phosphodiesterase, actin, and tubulin; adhesion of oligodendrocytes to a polylysine substratum activates the synthesis and phosphorylation of myelin basic protein, and the synthesis and acylation of proteolipid protein and DM-20; over time in culture there is an increased synthesis and accumulation of these proteins and of myelin-associated glycoprotein. We conclude that B3.f cells exhibit a behavior that is distinct from that of B3.fA cells. Our results are consistent with the notion that upon adhesion to a substratum, oligodendrocytes undergo a transition from myelin-maintaining cells (B3.f) to that of myelin-forming cells (B3.fA). This conclusion is substantiated by the finding of myelin membranes in these cultures.

Animals↗