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Biomedical subjects

L F Johnson

Publications and source records attributed to L F Johnson.

At least 127 records · Page 7Linked to original sources
PubMed1985

Palliative dilation of esophageal carcinoma.

The authors' experience with palliative dilation of 46 consecutive patients evaluated for squamous cell carcinoma of the esophagus was retrospectively reviewed. Thirty-nine of 46 patients (85%) underwent dilation in order to palliate symptoms, enable endoscopy and biopsy, or prepare for placement of an esophageal prosthesis. Thirty-two of the 46 patients (70%) were treated with radiation therapy and seven (15%) underwent placement of an esophageal prosthesis. Thirty-five of the 39 patients dilated (90%) noted improvement in swallowing, allowing resumption of a soft or regular diet. Complications were noted in three of the 39 patients dilated (8%). The authors conclude that peroral dilation is a safe, effective, and probably underutilized method of palliation in patients with squamous cell esophageal carcinoma.

Adult↗
PubMed1985

Control of thymidylate synthase mRNA content and gene transcription in an overproducing mouse cell line.

We studied the content and metabolism of thymidylate synthase mRNA in cultured mouse fibroblasts that were undergoing a serum-induced transition from the resting to growing state. The studies were performed with a 5-fluorodeoxyuridine-resistant 3T6 cell line (LU3-7) that over produces the enzyme and its mRNA about 50-fold and that regulates the expression of the thymidylate synthase gene in the same manner as the parental cell line. We have previously shown that the rate of synthesis of thymidylate synthase increases at least ninefold when the serum-stimulated cells traverse the S phase. Here we show, by Northern blot analysis, that thymidylate synthase mRNA increased 20- to 40-fold as cells progressed from resting to late S phase. About 85% of poly(A)+ thymidylate synthase mRNA was associated with polysomes at all times. The increase in thymidylate synthase poly(A)+ mRNA content was the result of an eightfold increase in the rate of production of this species, as shown by pulse-labeling studies. Pulse-chase analysis revealed that the half-life of thymidylate synthase poly(A)+ mRNA was similar in resting (9 h) and growing (7 h) cells. The rate of transcription of the thymidylate synthase gene, as determined in isolated nuclei, increased only by a factor of three to four during the S phase. Since the content of the message increased to a much greater extent than the rate of transcription of the gene, posttranscriptional controls must also play a role in regulating the content of thymidylate synthase mRNA under these conditions. Our results suggest that the cell may regulate the distribution of thymidylate synthase mRNA between a relatively stable poly(A)+ RNA species and a labile poly(A)- RNA species.

Animals↗
PubMed1985

Cimetidine for prevention and treatment of gastroduodenal mucosal lesions in patients in an intensive care unit.

The efficacy of cimetidine in the prevention and treatment of stress-induced gastroduodenal lesions was evaluated in a randomized, double-blind, placebo-controlled study in which serial endoscopy was used to examine patients without clinical evidence of bleeding who were admitted to a medical intensive care unit. Endoscopy showed that 14 of 21 patients treated with cimetidine, compared with 5 of 18 patients treated with placebo, had normal or improved gastroduodenal mucosa (p less than 0.05). Endoscopic signs of bleeding cleared or did not develop in 20 patients treated with cimetidine and in 11 patients treated with placebo (p less than 0.01). Significantly fewer blood transfusions were given to patients with endoscopic signs of bleeding in the cimetidine-treated group (0.5 +/- 0.3 [SE] units) than in placebo-treated patients (4.5 +/- 1.5 units; p less than 0.05). The mortality rate was not statistically different between treatment groups. By preventing established gastroduodenal stress lesions from progressing in severity, cimetidine diminished both bleeding and the need for transfusions.

Adult↗
PubMed1985

Thymidylate synthase overproduction and gene amplification in fluorodeoxyuridine-resistant human cells.

Cytotoxicity to 5-fluoro-2'-deoxyuridine (FdUrd) derives from its conversion to 5-fluorodeoxyuridine-5'-monophosphate, which binds to and inhibits thymidylate synthase (TS) in the presence of the cofactor, 5,10-methylenetetrahydrofolate. We have selected FdUrd-resistant variants of the human cell line HEp-2 following adaptation to stepwise increases in drug concentration. In the initial selection, maximal drug resistance was associated with a 26-fold increase in the cellular level of TS. Greater TS overproduction (80-fold) was obtained by selection for FdUrd resistance in the presence of 10 microM folinic acid and 100 microM deoxyinosine. The latter agents were included to expand the folate pool to ensure adequate levels of cofactor during the selection process. Using cDNA plasmid pMTS-4, which is complementary to mouse TS mRNA, we show that TS overproduction in the HEp-2 variants is accompanied by a 100-fold increase in TS mRNA and a 100-fold amplification of the TS structural gene. Thus, TS overproduction and gene amplification is a mechanism of resistance to FdUrd in human cells.

Base Sequence↗
PubMed1985

Thymidylate synthase gene amplification in fluorodeoxyuridine-resistant mouse cell lines.

We have previously isolated fluorodeoxyuridine-resistant mouse fibroblast (LU3-7) and neuroblastoma (FUdR-R) cell lines that overproduce thymidylate synthase and the mRNA for this enzyme up to 50-fold as compared to the parental cell lines. We have also cloned cDNA corresponding to mouse thymidylate synthase mRNA into pBR322. In the present study, we used this cloned cDNA as a hybridization probe in Southern blot analysis of DNA from the parental and overproducing cell lines. These analyses showed that the thymidylate synthase gene is amplified 50-100 fold in LU3-7 cells and about 30-fold in FUdR-R cells when compared to the respective parental cells. The sizes of the restriction fragments were the same in the parental and overproducing cells of each type, suggesting that extensive rearrangements have not occurred in the vicinity of the thymidylate synthase gene during the amplification process. However, not all of the fragments in the parental cells were amplified in the overproducing cells, suggesting that there may be multiple genes or pseudogenes for the enzyme. Restriction fragment length polymorphisms were detected when analyzing DNA from several different mouse cell lines. When LU3-7 cells were grown in the absence of selective pressure, the level of thymidylate synthase overproduction and the number of copies of the thymidylate synthase gene decreased in parallel.

Animals↗
PubMed1985

Taurodeoxycholate modulates the effects of pepsin and trypsin in experimental esophagitis.

Pepsin and trypsin cause erosive, hemorrhagic lesions in our rabbit model of experimental esophagitis. Since the gastroduodenal contents of patients with reflux esophagitis may also contain bile salts, we used our model to determine the effect that a bile salt, taurodeoxycholate (TDC), would have on the esophageal mucosa when combined with either pepsin in an acid perfusate (pH 2) or trypsin in an alkaline perfusate (pH 7.5). Indexes of esophageal injury included gross appearance of the mucosa, microscopic examination, and mucosal barrier integrity as determined by permeability to hydrogen ion. We found that when 5 mM TDC was combined with pepsin (0.3 mg/ml), the gross and microscopic changes of esophagitis, as well as net hydrogen ion flux, were diminished when compared with those observed with pepsin exposure alone. When increasing concentrations of TDC (2 to 10 mM) were added to pepsin, the morphologic degree of injury as well as hydrogen ion flux decreased in a dose-dependent manner. In contrast, when 5 mM TDC was combined with trypsin (1000 U/ml) in the alkaline perfusate, the gross and microscopic changes of esophagitis and the net of hydrogen ion flux were increased when compared with either bile salt or trypsin alone. These effects were also dose dependent. These data demonstrate that bile salts present in the gastroduodenal contents of patients with reflux esophagitis have the capacity to modulate the effects of pepsin and trypsin on the esophageal mucosa.

Animals↗
PubMed1984

Molecular cloning of DNA sequences complementary to mouse thymidylate synthase messenger RNA.

We describe the isolation of recombinant cDNA clones containing sequences corresponding to mouse thymidylate synthase mRNA. Double-stranded cDNA was made from poly(A+) RNA isolated from the 5-fluoro-2'-deoxyuridine-resistant mouse cell line LU3 -7 that overproduces thymidylate synthase 50- to 100-fold as compared to the parental mouse 3T6 fibroblasts. The cDNA was inserted into pBR322 using the poly(dC)-poly(dG) tailing procedure and transformed into Escherichia coli HB101. A library of 30,000 colonies was screened for thymidylate synthase cDNA sequences by differential colony hybridization. Plasmid DNA was purified from a colony that gave a positive signal. RNA corresponding to this plasmid was isolated by hybridization and translated in vitro to yield a protein that coelectrophoresed with authentic thymidylate synthase. The identity of the translation product was confirmed by immunoprecipitation with thymidylate synthase antiserum and by peptide analysis. A restriction fragment from this plasmid was used to rescreen the library to give a collection of thymidylate synthase cDNA plasmids with overlapping restriction maps. Several plasmids contained cDNA inserts greater than 1 kilobase pair in length. The size and content of thymidylate synthase mRNA in the overproducing and parental cell lines was determined by RNA blot analysis. Multiple thymidylate synthase mRNA species were identified. The predominant species was 1.3 kilobase pairs in length. Thymidylate synthase mRNA was 50 times more abundant in LU3 -7 than in 3T6 cells.

Animals↗
PubMed1984

In vitro and in vivo analysis of the control of dihydrofolate reductase gene transcription in serum-stimulated mouse fibroblasts.

We have studied the rate of transcription of the gene for dihydrofolate reductase (DHFR) in mouse 3T6 fibroblasts during serum-induced transitions between the resting (G0) and growing states. As a model system, we have used a methotrexate-resistant 3T6 cell line that overproduces DHFR and its mRNA about 300-fold, yet regulates the expression of the DHFR gene in the same manner as normal 3T6 cells. In previous studies, we showed that the rate of production of cytoplasmic DHFR mRNA relative to total mRNA is about 4 times lower in resting than in exponentially growing cells. The rate increases to the growing value by about 15 hr following serum stimulation of the resting cells. This increase appeared to be controlled by regulating the rate of synthesis of DHFR hnRNA. In this study, we analyze the transcription of the DHFR gene in more detail. We use a variety of labeling times and RNA extraction procedures to measure the rate of synthesis of DHFR hnRNA relative to total hnRNA in pulse-labeled cells or in nuclei isolated from cells at various times following serum stimulation. The amount of labeled DHFR RNA is determined by DNA-excess filter hybridization. In all cases, the relative rate of synthesis of DHFR hnRNA increases at the same time, and to the same extent, as the rate of production of DHFR mRNA, suggesting that the increase in DHFR mRNA production is due to a corresponding increase in the rate of transcription of the DHFR gene. The increase in DHFR gene transcription is not blocked by cytosine arabinoside, showing that the increase does not depend on gene duplication. In isolated nuclei, DHFR RNA synthesis is inhibited by alpha-amanitin (1 microgram/ml), indicating that the DHFR gene is transcribed by RNA polymerase II. Others have shown that when stationary phase cells are stimulated to proliferate, the increase in DHFR mRNA content is controlled primarily at the post-transcriptional level. Therefore, it appears that the rate of production of DHFR mRNA is controlled by different biochemical mechanisms when cells are in different physiological states.

Animals↗
PubMed1984

Lithium chloride: protective and antisecretory properties in rats.

Lithium chloride was evaluated as a potential protective agent against ethanol-induced hemorrhagic gastritis in rats. Rats received lithium chloride intragastrically (30, 60, or 90 mg/kg i.g.) or subcutaneously (3, 5, 10, 15, 20, 30, 60, or 90 mg/kg s.c.), or a placebo (H2O i.g. or 0.9% NaCl s.c.). Ninety minutes later 1 cm3 of 95% ethanol was administered intragastrically. After 30 min, the rats were killed and their stomachs were removed and visually scored for gross hemorrhagic gastritis. Lithium chloride at all doses less than 3 mg/kg significantly improved hemorrhagic gastritis when compared with the placebo. Moreover, rats treated with lithium chloride intragastrically had significantly less hemorrhagic gastritis than those treated subcutaneously even though serum lithium levels were similar. To determine if lithium's protective properties related to acid inhibition, pylorus-ligated and gastric fistula rats were studied. The median effective dose for lithium chloride was 20-30 mg/kg and the nonantisecretory dose was 3 mg/kg. No difference in hemorrhagic gastritis was noted between controls and animals receiving the nonantisecretory dose. However, at higher doses (LiCl 30, 60, and 90 mg/kg), lithium's protective properties persisted in spite of adding 150 mM HCl to the intragastrically administered ethanol (p less than 0.001). To determine further if lithium chloride stimulated endogenous prostaglandins, indomethacin, a prostaglandin inhibitor, was administered. Indomethacin did not alter lithium chloride's protective properties. In fact, when compared with exogenously administered 16,16-dimethyl prostaglandin E2 (5 and 500 micrograms/kg), high-dose LiCl (30, 60, or 90 mg/kg) resulted in significantly more protection against ethanol-induced injury (p less than 0.001). In contrast, when both nonantisecretory doses were compared, 16,16-dimethyl prostaglandin E2 (5 micrograms/kg) gave significantly better protection than LiCl (3 mg/kg). These data indicate that LiCl is a potent gastric antisecretory and protective agent. The protective properties of LiCl appear to be related to acid inhibition and be independent of endogenous prostaglandins, although other protective mechanisms may be present at higher LiCl doses. Additionally, this study indicates that LiCl may have clinical application in protecting the gastric mucosa against hemorrhagic gastritis.

Animals↗
PubMed1984

Effect of sleep on swallowing, esophageal peristalsis, and acid clearance.

Recent studies have shown that impaired esophageal clearance of refluxed material during the sleeping interval plays a major role in the pathogenesis of esophagitis. The present investigation compares esophageal responses to both acid and water infusions during sleep in asymptomatic subjects. Fourteen healthy individuals were studied. The subjects were studied for two nights in the sleep laboratory. Each night involved four to six infusions of 15 ml of either sterile water or 0.1 N HCl. A similar infusion was conducted during the presleep waking interval. Acid clearance time was shown to decrease as a function of increasing wakefulness during the acid clearing interval. Arousal responses were significantly greater with acid infusions during sleep (p less than 0.05). Awakenings from sleep with acid infusions were associated with significantly greater swallowing rates than those associated with water infusions (p less than 0.01). Peristaltic parameters of amplitude, velocity, and duration were not differentially affected by acid versus water infusions or by sleep versus waking.

Adult↗