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Biomedical subjects

R Lim

Publications and source records attributed to R Lim.

At least 127 records · Page 7Linked to original sources
PubMed1987

Rat astrocytes and Schwann cells in culture synthesize nerve growth factor-like neurite-promoting factors.

Neurite-promoting activity in feeding medium conditioned by rat astrocytes and Schwann cells in culture was examined. The conditioned medium (CM) from both types of glial cultures stimulated extensive neurite outgrowth from embryonic chick dorsal root ganglia (DRG) as well as pheochromocytoma (PC12) cells. Both the DRG and PC12 cells also produce neurite outgrowth in the presence of nerve growth factor (NGF). With the DRG, the neurite growth rates observed with the glial cell CM were identical to growth rates seen with NGF. Although anti-NGF antibody did not inhibit the neurite outgrowth produced by either of the glial CM, a nerve growth factor radioreceptor assay did detect an NGF-like molecule in both CM. Since the extensive neurite outgrowth stimulated by the glial CM was not mimicked by pure laminin alone, we conclude that the glial neurite promoting factors are distinct from laminin.

Adrenal Gland Neoplasms↗
PubMed1987

Endogenous immunoreactive glia maturation factor-like molecule in astrocytes and glioma cells.

Using the monoclonal antibody G2-09 raised against bovine glia maturation factor (GMF), we conducted a survey of GMF-like immunoreactivity in various cell types. Of all the normal and neoplastic cells tested, only extracts from astroblasts, gliomas, Schwann cells and schwannomas, but not their conditioned media, possessed endogenous GMF-like immunoreactivity. The presence of immunoreactive GMF correlated well with GMF bioactivity. Using the same monoclonal antibody, the GMF-like factor in astroblasts and C6 glioma cells was characterized by immunofluorescence, immunoadsorption and immunoblotting. Immunofluorescence confirmed the intracellular location of GMF. Immunoadsorption completely eliminated the GMF-like bioactivity from the cell extracts. Immunoblotting identified a protein band having a mol. wt. of 14,000 Da. Thus, the evidence strongly supports the argument that the GMF-like factor in astroblasts and C6 cells is identical with GMF from the bovine brain. In order to explain the fact that astroblasts and C6 cells are both the source and targets of GMF, we propose the hypothesis that GMF functions as an injury signal, being released from the injured glia and serving as a stimulant for gliosis in the neighboring intact glia.

Animals↗
PubMed1987

Distribution of immunoreactive glia maturation factor-like molecule in organs and tissues.

Using the monoclonal antibody G2-09 raised against bovine glia maturation factor (GMF), we screened various rat organs and tissues for GMF-like immunoreactivity. In the adult animal, with the exception of the heart, GMF was found exclusively in the nervous system, with the cerebellum exhibiting higher specific activity than other brain regions. The nature of the immunoactivity in the heart is presently unclear. None of the body fluids collected from humans, including serum and cerebrospinal fluid, possessed detectable GMF immunoactivity. A phylogenetic comparison revealed the presence of GMF in the brain of al vertebrates studied, from fish to primates. GMF was absent from bacteria and yeast. An ontogenetic study on rats showed the highest GMF level in the fetal brain, with a gradual but steady decrease after birth. However, a substantial amount of GMF persisted even in older animals. GMF was localized in astrocytes and Bergmann glia in the rat brain, using immunostaining at the light microscopic level.

Age Factors↗
PubMed1987

Induction of cytoskeletal alterations in C6 glioma by glia maturation factor.

C6 rat glioma cells respond to glia maturation factor (GMF) with characteristic morphological alterations. Observed under phase-contrast microscopy, the cells changed from a rounded morphology in random formation to a spindle-shaped appearance in parallel arrays. Observed under scanning electron microscopy, GMF led to a decrease in the number of microvilli and cell surface knobs. Transmission electron microscopy demonstrated the appearance of numerous microtubules aligned with the long axis of the cells after GMF stimulation. The change in cell shape and histotypic pattern was inhibited by vinblastin, further implicating the involvement of microtubules. Immunofluorescence using anti-alpha-tubulin revealed a well-defined cytoskeletal system in GMF-stimulated cells but not in the control cells. Finally, an increase in tubulin was confirmed with enzyme-linked immunosorbent assay (ELISA) on extracts from these cultures. The findings indicate that morphological alterations induced by GMF are associated with changes in the quantity and arrangement of microtubules.

Animals↗
PubMed1987

Acute cholecystitis in pregnancy.

Thirty cases of acute cholecystitis in pregnancy were identified during a 12-year period. Twenty-one patients were successfully managed with medical therapy alone. Nine underwent surgical intervention, four after failing medical therapy and five as the initial approach to therapy. Surgical therapy was complicated by preterm labor and delivery in two patients undergoing surgery during the early third trimester. One patient underwent cholecystectomy during the first trimester and subsequently aborted. There were no serious complications in patients undergoing surgery during the second trimester. These data suggest that conservative medical management can be successfully used in most patients with acute cholecystitis in pregnancy.

Abortion, Spontaneous↗
PubMed1986

Suppression of glioma growth in vitro and in vivo by glia maturation factor.

Glia maturation factor (GMF), a 14,000 Mr acidic protein of the brain, is capable of promoting differentiation of cultured astroblasts. In this study we report the effect of GMF on two glioma cell lines: the C6 line, of rodent origin, and the HG-1 line, of human origin. When tested in culture, GMF promotes the initial growth of the two cell lines when the cells are sparse but limits proliferation by restoring contact inhibition when the cells are confluent. Cell cycle analysis confirms the arrest of the cells at the G0/G1 phase when the tumor cells are contact inhibited by GMF. When C6 cells are inoculated into the athymic (nude) mice at a s.c. site, a single solid tumor grows out with a 100% take. Intraperitoneal injection of GMF leads to the slowing down of tumor growth. That the in vivo effect of GMF is not due to cytotoxicity is evidenced by the lack of necrosis and by the appearance of more mature astrocytic cells in the tumors. The results lend support to the concept of GMF as a cellular regulator and suggest the therapeutic potential of GMF for brain tumors.

Animals↗
PubMed1985

Purification of bovine glia maturation factor and characterization with monoclonal antibody.

Glia maturation factor (GMF) is purified 100 000-fold to apparent homogeneity from bovine brains by a procedure consisting of ammonium sulfate precipitation, column chromatography with diethylaminoethyl-Sephacel, Sephadex G-75, and hydroxylapatite, and a final step using C4 reverse-phase high-performance liquid chromatography. The product shows a single protein band in sodium dodecyl sulfate-polyacrylamide gel. It has a molecular weight of 14 000 and an isoelectric point of pH 5.2. Purified GMF stimulates cultured astroblasts to proliferate and to grow out cell processes with half-maximal activity at 8 ng/mL. A monoclonal antibody raised against partially purified GMF adsorbs the activity of pure GMF and immunologically binds the putative GMF protein band.

Amino Acids↗
PubMed1985

Glia maturation factor influences recovery from injury in neonatal rat brains.

Newborn rats were injured with a puncture wound in one cerebral hemisphere. Experimental animals were treated with three i.p. injections of Glia Maturation Factor (GMF) at daily intervals starting from the time of injury, whereas control littermates were treated with equivalent amounts of bovine serum albumin. At 25 days old the size of the cerebral cortex at the plane of injury was measured on representative brain sections. In control rats the injured side was 18% smaller than the normal side whereas in GMF-treated animals the difference was only 1%. The results suggest a possible regulatory role of GMF in promoting tissue recovery from brain damage.

Animals↗
PubMed1985

Mitogenic activity of glia maturation factor. Interaction with insulin and insulin-like growth factor-II.

The mitogenic activity of glia maturation factor (GMF) was tested on sparse-cultured cells. GMF stimulates the growth rate of normal astroblasts and fibroblasts grown in the presence of fetal calf serum (FCS), and raises the saturation density of the cells over what is imposed by the corresponding serum concentrations. GMF has no mitogenic effect in the complete absence of serum. The mitogenicity of GMF is also demonstrable in defined media where certain serum components are present. In particular, GMF in combination with the defined medium N2 partially mimics the proliferative effect of serum alone. Insulin, an ingredient of N2, can substitute for the complete N2 formula. Insulin-like growth factor-II (IGF-II), in turn, can substitute for insulin. The interaction of GMF with insulin or IGF-II can be demonstrated in a sequential manner, suggesting that GMF is a competence factor. Since insulin is required at a concentration well above the physiologic serum level, and must be used at a dose 1000 times higher than IGF-II, we suspected that insulin acts on IGF-II receptors. This was substantiated by the demonstration of IGF-II receptors and the absence of detectable insulin receptors on the astroblasts. The combined effect of IGF-II and GMF mimics the combined effect of 10% FCS and GMF, in both growth rate and saturation density.

Animals↗
PubMed1985

Protein phosphorylation in a tetradecanoyl phorbol acetate-nonproliferative variant of 3T3 cells.

The 3T3-TNR9 cell line is a variant of Swiss 3T3 cells which does not respond mitogenically to tumor promoters, but does respond mitogenically to epidermal growth factor, fibroblast growth factor, and serum. To elucidate differences between tumor promoters and polypeptide mitogens in the pathway(s) of mitogenesis which might be responsible for the nonresponsiveness of the 3T3-TNR9 cells, we have examined in these cells the early protein phosphorylation events known to be associated with mitogenesis in the parental 3T3 cells. We find that the 3T3-TNR9 cells display levels of tetradecanoyl phorbol acetate binding and of a calcium- and phospholipid-dependent protein kinase activity which are at least the equal of those seen in the parental 3T3 cells, implicating some postreceptor event in the nonmitogenic phenotype. In addition, we find that phosphorylation of the epidermal growth factor receptor and of 80-kDa and 22-kDa proteins, as well as the tyrosine phosphorylation of a 42-kDa protein, all proceed normally in the nonmitogenic variant, even though these phosphorylations must depend on the activation of different kinases. Thus, all these early phosphorylation reactions are intact in the 3T3-TNR9 cells. Although these phosphorylations may be necessary, they clearly are insufficient to trigger mitogenesis.

Animals↗
PubMed1984

Glia maturation factor promotes proliferation and morphologic expression of rat Schwann cells.

Glia maturation factor (GMF) is an acidic protein with a molecular weight of about 20,000 daltons, found in the adult brain of many species. Previously GMF was observed to stimulate the proliferation and subsequent maturation of rat astroblasts in culture. We investigated the effects of GMF on Schwann cells. Schwann cells were dissociated from rat sciatic nerve and purified by means of antimitotic agents and by selective immunoadsorption of contaminating fibroblasts. Cultured Schwann cells after 3 passages assumed a flat polygonal shape. Exposure of the cells to GMF converted the cells to the elongated, spindle morphology typical of Schwann cells. GMF also stimulated a 7-fold increase in DNA synthesis when compared with control cultures grown in F10 medium containing 5% fetal calf serum. The mitogenic activity of GMF was still detectable at 5 ng protein/ml medium. The maximal effect on DNA synthesis occurred 72 h after the initial exposure to GMF. Although the cells were positive for the Schwann cell marker Ran-1, GMF failed to induce the production of myelin-associated glycolipids (galactocerebroside) and proteins (Po) nor did it induce the astrocytic marker glial fibrillary acidic protein (GFAP). The effects of GMF on Schwann cells extend its biological role beyond the central nervous system.

Animals↗
PubMed1984

Sequential interaction of glia maturation factor with insulin.

Astroblasts in culture proliferated when exposed to glia maturation factor for at least 2 hours and then to insulin, but not when exposed in the reverse order. The sequential relation suggests that glia maturation factor is a competence factor.

Animals↗
PubMed1984

An improved procedure for the isolation of glia maturation factor.

A procedure for the bulk isolation of glia maturation factor (GMF) in high yield and high purity from bovine brains is outlined. The method involves extraction by homogenization and centrifugation, followed by ammonium sulfate precipitation and column chromatography with diethylaminoethyl (DEAE) Sephacel, Sephadex G-75, and hydroxylapatite. The method results in a 10,000-fold purification, a purity exceeding that of previously published procedures, and enables us to handle as much as 2.8 kg brain tissue or eight brains/week. The ability to mass-produce GMF with this method greatly facilitates its biological studies, further purification, and chemical characterization. The isolated GMF shows a molecular weight of 13,000 on Bio-gel P-30 column and an isoelectric point of about 5.4 on isoelectric focusing. The isolated GMF is heat labile and susceptible to papain and ficin but relatively resistant to trypsin, neuraminidase, and endoglycosidase.

Animals↗
PubMed1984

Dual properties of cultured retinoblastoma cells: immunohistochemical characterization of neuronal and glial markers.

The dual properties of two human retinoblastoma cell lines, WERI-Rb1 and Y79, were investigated with immunohistochemistry. Two neuron-specific markers, dopamine-B-hydroxylase (DBH) and tetanus toxin, and an astrocyte-specific marker, the glial fibrillary acid protein (GFAP), were applied for immunohistochemical reactions. With peroxidase-antiperoxidase (PAP) and immunofluorescence techniques, all of the WERI-Rb1 and Y79 cells showed consistently positive results with both neuronal and glial markers. The findings demonstrate that cultured retinoblastoma cells WERI-Rb1 and Y79 have both neuronal and glial properties.

Cell Line↗
PubMed1983

Purification of rat Schwann cells from cultures of peripheral nerve: an immunoselective method using surfaces coated with anti-immunoglobulin antibodies.

We report a method for deriving purified rat Schwann cells by immunoselective removal of fibroblasts. Contaminating fibroblasts labeled with antibody against specific surface marker Thy 1.1 are bound on plastic surfaces coated with a second antibody. The efficacy of the method is demonstrated by flow cytometry and by specific Schwann cell Ran-1 immunofluorescence.

Animals↗
PubMed1983

Dual effect of glia maturation factor on astrocytes. Differentiation and release of interleukin-1 like factors.

C6 glioma cells, and primary cultures of mouse astrocytes, stimulated with lipopolysaccharide (LPS) release an interleukin-1 like factor (IL-1) which enhances lectin-induced T-lymphocyte proliferation and promotes the release of interleukin-2 (IL-2) by ConA-stimulated thymocytes. In the present study, the glia maturation factor (GMF) was found not only to induce differentiation of glioblasts, but also to elicit the secretion of IL-1 like factors by cultured mouse astrocytes and their precursor cells. GMF was also effective in triggering IL-1 release by macrophages. Contamination of the 23 000 MW GMF preparation with LPS was excluded by the Limulus lysate assay and by using C3H/HeJ LPS-nonresponder mice whose glia and macrophages responded to GMF but not to LPS, by IL-1 release. Through its ability to induce glial differentiation and IL-1 release, GMF may represent an important endogenous signal, triggering both reactive gliosis and the development of an immune response within the central nervous system.

Animals↗