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Biomedical subjects

L Philipson

Publications and source records attributed to L Philipson.

At least 37 records · Page 2Linked to original sources
PubMed1994

Regulation of G1 progression by E2A and Id helix-loop-helix proteins.

In NIH3T3 fibroblasts, the ubiquitous helix-loop-helix (HLH) protein E2A (E12/E47) and the myogenic HLH proteins MyoD, MRF4 and myogenin are growth-inhibitory, while two ubiquitous Id proteins lacking the basic region are not. The dimerization domain mediates inhibition. However, in addition to the HLH region, E2A contains two inhibitory regions over-lapping with the main transcriptional activation domains. The growth-suppressive activity of the intact E47 as well as MyoD was counteracted by the Id proteins. When E47 lacking the HLH domain was overexpressed, Id could no longer reverse growth inhibition. By increasing the amount of E47 with an inducible system or neutralizing the endogenous Id with microinjected anti-Id antibodies, withdrawal from the cell cycle occurred within hours before the G1-S transition point. The combined results suggest that the Id proteins are required for G1 progression. The antagonism between the E2A and Id proteins further suggests that both are involved in regulatory events prior to or near the restriction point in the G1 phase of the cell cycle.

3T3 Cells↗
PubMed1994

Id proteins control growth induction in mammalian cells.

Id1, Id2, and Id3 (HLH462) dimerize with members of the basic helix-loop-helix protein family, but due to the absence of the basic region, the resulting heterodimers cannot bind DNA. Therefore Id-type proteins negatively regulate DNA binding of the basic helix-loop-helix proteins. Here we report that Id1, Id2, and Id3 are induced shortly after serum stimulation in arrested NIH 3T3. Antisense oligonucleotides against the Id mRNAs delay the reentry of arrested cells into the cell cycle elicited by stimulation with serum or growth factors. Antisense oligonucleotides against all three Id mRNAs are more effective than individual ones. Combined, these results indicate that Id proteins are involved in the control of growth induction.

3T3 Cells↗
PubMed1994

CHOP (GADD153) and its oncogenic variant, TLS-CHOP, have opposing effects on the induction of G1/S arrest.

The growth arrest and DNA damage-inducible gene CHOP (GADD153) encodes a small nuclear protein from the C/EBP family, originally isolated from adipocytes in culture. Although inactive in cells under normal conditions, the CHOP gene is markedly induced by a variety of cellular stresses, including nutrient deprivation and metabolic perturbations. These lead to accumulation of CHOP protein in the nucleus. Because cellular stress normally leads to growth arrest, we examined the implication of CHOP in this process. Microinjection of CHOP expression plasmids into NIH-3T3 cells blocked the cells from progressing through the cell cycle, measured by an attenuation in the fraction of cells incorporating BrdU, an S-phase marker. The precise point in the cell cycle at which CHOP acts was mapped by microinjection of bacterially expressed CHOP protein into synchronized cells--this blocked the cells from progressing from G1 to S phase. This effect of CHOP was observed only when the protein was introduced early after serum stimulation suggesting that CHOP works at or around the so-called G1/S checkpoint. CHOP dimerizes with other C/EBP proteins and the CHOP-C/EBP dimers are directed away from "classical" C/EBP sites recognizing instead unique "nonclassical" sites. Mutant forms of the CHOP protein that lack the leucine zipper dimerization domain or the unusually structured basic region, potentially involved in DNA binding, fail to induce growth arrest. A tumor-specific form of CHOP, TLS-CHOP, that has been found so far exclusively in the human adipose tissue tumor myxoid liposarcoma, fails to cause growth arrest and furthermore interferes with the ability of normal CHOP to induce growth arrest. CHOP has been shown recently to be markedly inducible by nutritional deprivation of cells. This suggests that CHOP may play a role in an inducible growth arrest pathway that is triggered by metabolic cues and is of particular importance in adipose tissue--an organ that undergoes marked changes in its metabolic activity. Blocking of this pathway by TLS-CHOP may play a mechanistic role in the establishment of myxoid liposarcoma.

3T3 Cells↗
PubMed1994

Expression of growth arrest-specific (gas) genes in senescent murine cells.

The growth arrest-specific (gas) genes were initially identified on the basis of their preferential expression in mouse fibroblasts during quiescence, followed by down-regulation upon reentry into the cell cycle. We here report studies on the expression of these genes in murine fibroblasts undergoing replicative senescence in vitro. Our results indicate a different behavior between senescent and GO-arrested quiescent fibroblasts. Expression of the gas1 and 6 genes was dramatically reduced in senescent cells. Only basal levels of gas2, 3, and 5 genes were detected in senescent fibroblasts, and they were independent of the growing conditions of the cultures. Down-regulation of the gas1 gene expression in senescent cells was apparently due to reduced transcription of the gas1 gene. This correlates with an altered pattern of factors that bind to the promoter region of the gas1 gene, as measured by band shift assay with nuclear extracts of senescent fibroblasts.

Animals↗
PubMed1994

F response and H reflex for monitoring nerve block during epidural analgesia with ropivacaine.

The effect of a single epidural injection of ropivacaine on the motor and sensory function controlled from the L5/S1 level was investigated in 28 male volunteers. Concentrations of 1%, 0.75%, or 0.5% ropivacaine, 20 mL, administered at the L2/3 level were studied. Motor function was assessed quantitatively (measurement of muscle force by mechano-transducers), and sensory function by the pinprick method. In addition, F response and H reflex, tests which measure the conduction velocity in the central parts of peripheral nerves, were used. Epidural ropivacaine caused dose-dependent prolongation of the latencies of both these variables. F response latency recovered significantly later than motor function measured by mechano-transducers in the two lower concentration groups. H reflex latency recovered significantly later than sensory function assessed by the pinprick method in all three concentration groups. The time needed for recovery of F and H latencies was not significantly longer than the time from epidural injection to mobilization. At the time when the subjects could go through the mobilization procedure, 12 of 28 subjects were not completely recovered. In 5 of these 12 subjects, the H reflex latency was persistently prolonged at the end of the investigation, long after the subjects felt "normal" again. On follow-up recordings 5 mo later, the baseline latency had been regained in all five subjects. We conclude that F response and H reflex latencies are good indicators of the inhibition of nerve impulse conduction induced by epidural analgesia.

Adult↗
PubMed1993

Structure and regulation of the growth arrest-specific (gas-1) promoter.

We have isolated recombinant genomic clones encompassing several kilobase pairs of the 5'-flanking regions of both the human and murine gas-1 gene (growth arrest-specific gene 1). Both species share a highly conserved region of approximately 550 base pairs upstream of the gas-1 transcription start site. Deletion analysis of the murine gas-1 promoter demonstrated that a fragment containing the first 665 base pairs is sufficient to drive the serum-regulated expression of a luciferase reporter gene in NIH3T3 cells, in a manner qualitatively reflecting the activity of the endogenous gene. Gel retardation assays indicated the presence of a number of DNA-binding proteins specific for sequences contained within the gas-1 transcription regulatory region. Comparative studies with extracts prepared from growing and resting cells revealed several growth state-specific binding activities. One promoter fragment that bound prominent growth- and arrest-specific complexes was further analyzed by copper-phenanthroline footprinting. It was found that the same DNA element is a target both for growth- and for arrest-specific activities. The factors characterized in this study are the first candidates for transcriptional regulators mediating cell growth-specific repression and/or growth arrest-specific activation of gene expression.

3T3 Cells↗
PubMed1993

Blockade of the abdominal muscles measured by EMG during lumbar epidural analgesia with ropivacaine--a double-blind study.

A single shot of 20 ml of 1%, 0.75% or 0.5% ropivacaine was administered epidurally (at L2/3 level) to 30 volunteers, in a double-blind manner. The blockade of the rectus abdominis muscle was measured quantitatively by registration of the average rectified electromyographic signal (AREMG) at the T7, T9 and T11 motor segmental levels and with a qualitative test for blockade of the rectus abdominis muscle (the so-called RAM test). The maximal cranial spread of analgesia, evaluated by the pin-prick method, was not significantly different for the three concentrations (T8-T10 dermatome; median value). The intensity of motor blockade, measured by the AREMG method, increased progressively from the T7 segment and caudally with all three concentrations. The blockade was partial (i.e. 85-25% of baseline AREMG activity was present at its maximum) in all subjects. When the effect of the three concentrations of ropivacaine was compared at the same segmental level, the intensity and duration of maximal motor blockade seemed to be dose-dependent, but the difference was not statistically significant. The total duration of motor blockade was shorter with the 0.5% solution than with the higher concentrations. The AREMG method gave a more exact and graded picture of blockade of the rectus abdominis muscle than the RAM test. The duration of sensory blockade did not outlast motor blockade at any level. In half of our subjects the maximal spread of sensory blockade was either equal to or higher than the spread of partial motor block. In the other half, this relationship was reversed--the maximal cranial level of partial motor block was 1-4 segments higher than the maximal level of analgesia.

Abdominal Muscles↗
PubMed1992

The growth arrest-specific gene, gas1, is involved in growth suppression.

This report describes the structure of the mRNA, the protein product, and the growth-regulating activity of one of the growth arrest-specific genes, gas1. From the predicted amino acid sequence, in vitro translation of gas1 mRNA, and immunofluorescence of cells in culture, it appears that the gas1 protein is an integral plasma membrane protein whose expression is linked to growth arrest. When gas1 is overexpressed from a constitutive promoter in quiescent cells, the serum-induced transition from the G0 to the S phase of the cell cycle is inhibited without affecting the normal early serum response. Ectopic expression of the gas1 gene by microinjection in normal and transformed NIH 3T3 cell lines with the notable exception of SV40-transformed 3T3 cells leads to inhibition of DNA synthesis. Thus, gas1 appears to be one component of a negative circuit that governs growth suppression. Its effect is, however, abolished in SV40-transformed cells.

3T3 Cells↗
PubMed1992

Consistent policy?

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Base Sequence↗
PubMed1992

Regulation and expression of a growth arrest-specific gene (gas5) during growth, differentiation, and development.

The growth arrest-specific gas5 gene was isolated from mouse genomic DNA and structurally characterized. The transcriptional unit is divided into 12 exons that span around 7 kb. An alternative splicing mechanism gives rise to two mature mRNAs which contain either 11 or 12 exons, and both are found in the cytoplasm of growth-arrested cells. In vivo, the gas5 gene is ubiquitously expressed in mouse tissues during development and adult life. In Friend leukemia and NIH 3T3 cells, the levels of gas5 gene mRNA were high in saturation density-arrested cells and almost undetectable in actively growing cells. Run-on experiments indicated that the gas5 gene is transcribed at the same level in both growing and arrested cells. On the other hand, in dimethyl sulfoxide-induced differentiating cells a sharp decrease in the rate of transcription was observed shortly before the cells reached the postmitotic stage. These results indicate that in density-arrested cells accumulation of gas5 mRNA is controlled at the posttranscriptional level while in differentiating cells expression is regulated transcriptionally.

3T3 Cells↗
PubMed1991

From growth arrest to growth suppression.

Since the introduction of the cell cycle concept two approaches to study growth regulation of cells have been proposed. One claims that cells are naturally quiescent, requiring a stimulatory encouter with growth factors for induction of cell division. The other considers cellular multiplication as the natural steady-state; cessation of multiplication is thus a restriction imposed on the system. In the latter case emphasis is mainly on the signals involved in arrest of multiplication. This Prospect focuses on specific events occurring in mammalian cells at growth arrest, senescence, and terminal differentiation, specifically emphasizing the growth inhibitory factors, tumor suppressor genes, and other signals for growth suppression.

Animals↗
PubMed1991

Behavioral medicine treatment in chronic constipation with paradoxical anal sphincter contraction.

Nine women and five children with severe chronic constipation received behavioral medicine therapy. Before treatment, all patients had a paradoxical contraction of the external anal sphincter at defecation attempts as demonstrated with electromyography and/or anorectal manometry. An electromyographic biofeedback device connected to an anal probe was used for the training that was performed on a regular toilet seat during five 1-hour sessions. Thirteen of the patients improved considerably and could learn to defecate spontaneously, and the use of laxatives ceased or diminished. Simultaneously with improvement, the paradoxical anal contraction disappeared. The results remained after 6 months, although two of the patients had received booster sessions of biofeedback training during follow-up.

Adolescent↗