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Biomedical subjects

L Philipson

Publications and source records attributed to L Philipson.

At least 55 records · Page 3Linked to original sources
PubMed1991

Cloning of a growth arrest-specific and transforming growth factor beta-regulated gene, TI 1, from an epithelial cell line.

By cDNA cloning and differential screening, five genes that are regulated by transforming growth factor beta (TGF beta) in mink lung epithelial cells were identified. A novel membrane protein gene, TI 1, was identified which was downregulated by TGF beta and serum in quiescent cells. In actively growing cells, the TI 1 gene is rapidly and transiently induced by TGF beta, and it is overexpressed in the presence of protein synthesis inhibitors. It appears to be related to a family of transmembrane glycoproteins that are expressed on lymphocytes and tumor cells. The four other genes were all induced by TGF beta and correspond to the genes of collagen alpha type I, fibronectin, plasminogen activator inhibitor 1, and the monocyte chemotactic cell-activating factor (JE gene) previously shown to be TGF beta regulated.

Amino Acid Sequence↗
PubMed1991

Epidural anesthesia with 0.5% bupivacaine: influence of age on sensory and motor blockade.

To determine the influence of age on epidural blockade, 10 elderly male patients (mean age, 73 yr) and 11 young male volunteers (mean age, 25 yr) received epidural anesthesia with 20 mL of 0.5% bupivacaine with epinephrine at the L2-3 interspace. Median cephalad analgesia level (pinprick) was T-4 in the elderly and T-10 in the young subjects (P less than 0.01). The duration of analgesia at the L-1 to S-3 segments was about 60 min shorter in the older patients than in the young volunteers (P less than 0.05). Motor blockade was evaluated quantitatively by isometric muscle force measurements in the lower extremities and by electromyographic recordings from the abdominal and quadriceps muscles. Motor blockade in the lower extremities was also assessed qualitatively by the Bromage scale. The intensity of motor blockade was not influenced by age. The mean duration of maximal motor blockade of the lower abdominal muscles was 69 min in elderly patients and 140 min in young subjects (P less than 0.05). The corresponding mean duration in the lower extremities was approximately 1.5 h in the older patients and 3 h in the young subjects (P less than 0.01). The elderly patients could be mobilized 2 h earlier than the young volunteers. Duration of Bromage grade 1 was about 1.5 h shorter in the elderly than in the young subjects (P less than 0.01). Not until 1.5-2 h after attainment of Bromage grade 0 was the muscle force in the lower extremities restored to 90% of baseline value. The authors conclude that cephalad spread was more extensive and duration of epidural blockade was shorter in elderly patients versus younger volunteers.

Adult↗
PubMed1991

Single cell assay with an automated capillary microinjection system.

An automated capillary microinjection system with computer controlled positioning of the cells and of the capillary, and its applications and advantages are described. About 1500 injections are possible in one hour, with high reproducibility. In cytoplasmic and nuclear injections more than 90% and 85% of the cells are successfully injected. Using FITC-Dextran at a concentration of 0.5% as a fluorescently labelled coinjection marker, 99% of the cells can be retrieved in culture medium even 48 hours after injection. The coordinates of the cells are stored in the computer and accuracy in statistical evaluation of experiments is improved in comparison to the manual techniques. Methods for preparation and handling of glass capillaries were developed resulting in reproducible form and significantly reduced clogging rate. The improved characteristics offered by this system are demonstrated in studies leading to the confirmation of existence of mRNA(s) inhibiting cell proliferation. Functional screening by cell injections of cDNA libraries and of size fractionated mRNA molecules can be performed efficiently with the automated microinjection system.

Animals↗
PubMed1990

Cell proliferation inhibited by MyoD1 independently of myogenic differentiation.

Cell growth and differentiation are usually mutually exclusive. Transformation of myoblasts by retroviruses containing the myc oncogene inhibits differentiation, preventing cells from withdrawing from the cell cycle. If cell-cycle withdrawal is a prerequisite for myoblast differentiation, it is probably an early event in terminal cell differentiation, but this has not yet been established. MyoD1 regulates myogenesis. It is expressed only in skeletal muscle, but can convert other cells to muscle cells. The MyoD1 protein, a nuclear phosphoprotein in part similar to the myc family of proteins, is a DNA-binding protein binding to the enhancer sequences of the muscle-specific creatine phosphokinase gene. Thus, introduction of MyoD1 into cells provides a simple approach to study the effect of induction of differentiation on cell growth. In cultured NIH 3T3 cells, inhibition of cell proliferation occurs within 18 hours, and expression of myosin starts after 72 hours. Furthermore, injection of MyoD1 into quiescent NIH 3T3 cells inhibit cell proliferation independently of induction of differentiation. Deletion of the myc-like domain in the MyoD1 gene eliminates the inhibition of DNA synthesis, but substitution of the basic domain with the analogous domain from the E12 transcription factor inhibits growth yet fails to induce differentiation. Inhibition of DNA synthesis, therefore, seems to be controlled separately from myogenic differentiation.

Animals↗
PubMed1990

Influence of intravenous local anaesthetics upon quadriceps muscle function.

This study was undertaken to assess the effects of intravenous administration of mepivacaine and etidocaine on muscle function. Seven male volunteers were given mepivacaine (5 mg/kg) and etidocaine (50 mg) intravenously, on separate occasions. A reference group of 11 male volunteers received 0.9% saline solution intravenously. Muscle function was tested by measurements of isometric muscle force of knee extension and by quantitative electromyographic (EMG) recordings from the quadriceps muscle during knee extension at different degrees of isometric muscle force. At the end of the mepivacaine and etidocaine infusions, the mean venous plasma concentrations of the two anaesthetic agents were 2.9 and 1.2 micrograms/ml, respectively. The muscle strength remained unchanged during infusion of the two local anaesthetics. Mepivacaine had a minor effect on the mean rectified EMG amplitudes at the end of the infusion at maximal voluntary muscle contraction, but no such effect was observed at submaximal knee extension force. However, at the plasma concentrations mentioned above, the clinical influence of intravenous infusion of the local anaesthetics on muscle function was negligible.

Adult↗
PubMed1990

Regulation of expression of growth arrest-specific genes in mouse fibroblasts.

The suppression of growth arrest-specific (gas) gene expression by serum appeared to be independent of protein synthesis, but expression in resting cells was sensitive to 2-aminopurine, an inhibitor of intracellular protein kinases. Although accumulation of gas gene mRNA was reduced by serum, nuclear transcription of the gas-2, -3, and -5 genes was observed in serum-stimulated cells, indicating that posttranscriptional events may regulate mRNA levels. Growth induction by serum, on the other hand, led to suppression of transcription of the gas-1 gene. Cell cycle regulation and the serum response of gas-1 were lost in ras-transformed cells.

Animals↗
PubMed1990

A growth arrest-specific (gas) gene codes for a membrane protein.

A set of growth arrest-specific (gas) genes whose expression is negatively regulated by serum has recently been identified. We report on the detailed analysis of one of these genes (gas3). The kinetics of regulation by the presence and absence of serum were investigated, and it was found that this gene is regulated at the post-transcriptional level. The encoded protein deduced from the nucleotide sequence showed some similarity to a mitochondrial oxyreductase, and in vitro translation established that the protein product is a transmembrane glycoprotein.

Amino Acid Sequence↗
PubMed1989

Motor blockade and EMG recordings in epidural anaesthesia. A comparison between mepivacaine 2%, bupivacaine 0.5% and etidocaine 1.5%.

In a double-blind study young volunteers randomly received 20 ml of mepivacaine 2%, bupivacaine 0.5% or etidocaine 1.5% epidurally, all solutions with adrenaline. The mean cephalad spread of pin-prick analgesia was equal (T10) in the groups, but the duration was longest for bupivacaine and etidocaine. The motor blockade of the rectus abdominis muscles was assessed quantitatively by rectified integrated electromyographic recordings (RIEMG) and as number of turns in EMG recordings [changes in the direction (rise/fall) of the EMG; TURNS] from three different segmental levels, T7, T9 and T11. The motor blockade of the quadriceps muscles was estimated by EMG recordings simultaneously with muscle force measurements at maximal isometric knee extension. Motor blockade was also evaluated by the Bromage scale. There was good correlation (correlation coefficient 0.91) between RIEMG values and muscle force in knee extension during epidural anaesthesia. TURNS showed a non-linear relationship to isometric force during epidural anaesthesia and added no further information. At the lower parts of the abdomen (T11), etidocaine gave more profound and longer motor blockade than mepivacaine. For quadriceps muscle function, motor blockade was almost complete with all three local anaesthetics; the duration of maximum motor blockade was short (45-60 min) for mepivacaine, but about 5 h with etidocaine. At the time when the Bromage scale indicated complete regression of motor blockade, the muscle force of knee extension was only 30% and the quadriceps RIEMG 35% of control values and 1-3 h remained until the time of mobilization.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetanilides↗
PubMed1989

Motor and sensory blockade after epidural injection of mepivacaine, bupivacaine, and etidocaine--a double-blind study.

In a double-blind study of epidural anesthesia, 30 young volunteers were given either 2% mepivacaine (400 mg), 0.5% bupivacaine (100 mg), or 1.5% etidocaine (300 mg), all solutions containing epinephrine (1:200,000). The spread of analgesia was equal in the groups, whereas the longest duration was noted in the etidocaine and bupivacaine groups. With use of a method for determining muscle force, motor blockade during anesthesia was recorded quantitatively for hip flexion, knee extension, and plantar flexion of the big toe. Onset of motor blockade was significantly more rapid with etidocaine than with bupivacaine and mepivacaine. All subjects given etidocaine developed complete motor blockade, but with the other local anesthetics 5%-33% of the initial muscle force remained. The least motor blockade was found in the L5-S2 segment (plantar flexion of the big toe). The duration of maximal motor blockade varied between 60 min (mepivacaine) and 360 min (etidocaine). With each of the three local anesthetics, motor function returned simultaneously in the three muscle groups tested. Complete restoration of muscle function occurred significantly later for etidocaine (600 min) than for bupivacaine (360 min) and mepivacaine (180 min). With etidocaine, the motor blockade outlasted the sensory blockade by 150 min. The Bromage scale corresponded to the motor blockade only during the first half of the regression phase. Not until 1-3 h after attainment of Bromage grade 0 was the muscle force of all movements restored (90% of control values).

Acetanilides↗
PubMed1988

Structure and expression of a Xenopus gene encoding an snRNP protein (U1 70K).

A cDNA and two genes for the Xenopus laevis U snRNP 70K protein have been cloned and partially sequenced. The cDNA encodes a protein whose predicted mol. wt is 57 kd but which migrates as a 70 kd protein in SDS-PAGE when translated in vitro from a cDNA transcript. The predicted protein sequences of the human and Xenopus U1 70K are shown to be very similar. Analysis of several genomic clones suggests that there are at least two, and possibly more, different genes coding for the 70K protein in the Xenopus genome. The two genes analysed in detail cover approximately 16 kb and are divided into 10 exons of which the last exon covers more than half of the protein coding sequence. During Xenopus development several different stage-specific RNAs hybridizing to the U1 70K cDNA are detected. The promoter region of one of the cloned genes is demonstrated to be functionally active, and to show apparent differences from other pol II promoters.

Amino Acid Sequence↗
PubMed1988

Genes specifically expressed at growth arrest of mammalian cells.

A subtraction cDNA library enriched for RNA sequences preferentially expressed in growth-arrested cells was prepared. Six cDNA clones were identified, varying in abundance from 2% to 0.0002% of the library and in size from 0.8 to 10 kb. The corresponding mRNAs are downregulated with different kinetics upon induction of growth by serum. The kinetics of induction after serum starvation and density-dependent inhibition of two of these growth-arrest-specific (gas) genes were investigated in more detail. Two cell lines transformed by viral onc genes did not express the two gas genes. The full-length cDNA for one gene has been sequenced and the protein product preliminarily characterized by in vitro translation.

Amino Acid Sequence↗
PubMed1988

High expression of functional adenovirus DNA polymerase and precursor terminal protein using recombinant vaccinia virus.

Initiation of Adenovirus (Ad) DNA replication occurs by a protein-priming mechanism in which the viral precursor terminal protein (pTP) and DNA polymerase (pol) as well as two nuclear DNA-binding proteins from uninfected HeLa cells are required. Biochemical studies on the pTP and DNA polymerase proteins separately have been hampered due to their low abundance and their presence as a pTP-pol complex in Ad infected cells. We have constructed a genomic sequence containing the large open reading frame from the Ad5 pol gene to which 9 basepairs from a putative exon were ligated. When inserted behind a modified late promoter of vaccinia virus the resulting recombinant virus produced enzymatically active 140 kDa Ad DNA polymerase. The same strategy was applied to express the 80 kDa pTP gene in a functional form. Both proteins were overexpressed at least 30-fold compared to extracts from Adenovirus infected cells and, when combined, were fully active for initiation in an in vitro Adenovirus DNA replication system.

DNA Replication↗
PubMed1988

Single cell assay with an automated capillary microinjection system.

An automated capillary microinjection system with computer-controlled positioning of the cells and of the capillary, and its applications and advantages, is described. The system is easy in handling and manipulation. About 1500 injections are possible in 1 h, with high reproducibility. In cytoplasmic and nuclear injections more than 90 and 85% of the cells are successfully injected. Using FITC-dextran at a concentration of 0.5% as a fluorescently labeled coinjection marker, 99% of the cells can be retrieved in culture medium even 48 h after injection. The coordinates of the cells are stored in the computer and accuracy in statistical evaluation of experiments is improved in comparison to the manual techniques. Methods for preparation and handling of glass capillaries were developed resulting in reproducible form and significantly reduced clogging rate. The improved characteristics offered by this system are demonstrated in studies leading to the confirmation of existence of an mRNA inhibiting DNA synthesis in cells. Functional screening by cell injections of cDNA libraries and of size-fractionated mRNA molecules can be performed efficiently with the automated microinjection system.

Autoradiography↗
PubMed1988

Automatic microinjection system facilitates detection of growth inhibitory mRNA.

Naturally quiescent human lymphocytes, consisting predominantly of T cells, contain mRNA(s) that can inhibit DNA synthesis when injected into either human diploid fibroblasts (IMR-90) or transformed recipient cells (HeLa). By using an automated capillary microinjection system and a fluorescent coinjection marker (fluorescein isothiocyanate-dextran), individually injected cells can be retrieved and analyzed for DNA synthesis. mRNA isolated from resting T cells is able to block the cells from entering the S phase. The block is reversible and leads to a delay in DNA synthesis. The inhibitory effect is not observed if the injected mRNA is isolated from growth-activated T cells. The disappearance of the inhibition coincides with the approach of the G1/S boundary in both the donor T cells and the recipient human fibroblasts. The mRNA of resting T cells was size-fractionated and the peak inhibitory activity was recovered in a fraction approximately equal to 1.5 kilobases long.

Cell Division↗