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Biomedical subjects

M D Walker

Publications and source records attributed to M D Walker.

At least 73 records · Page 4Linked to original sources
PubMed1985

Homologous recombination between a defective virus and a chromosomal sequence in mammalian cells.

Replacement of the early region of simian virus 40 results in virus that cannot replicate in a normal host, CV-1 cells, but can replicate in COS cells, a derivative of CV-1 cells that constitutively express simian virus 40 tumor antigen (T antigen). However, passage of such an early replacement simian virus 40 mutant in COS cells results in the emergence of virus that can propagate in CV-1 cells. Analysis of this virus revealed that the mutant rescued the integrated T-antigen gene from the COS cell genome. Comparison of the sequence of the recovered virus with that of the viral DNA resident in COS cells (strain 776) and the mutant used in our studies (derived from strain 777) proves that the mutant virus acquired the T-antigen gene from the COS cell chromosome via homologous recombination. Most probably this process was mediated by a direct genetic exchange.

Amino Acid Sequence↗
PubMed1983

Expressing a human proinsulin cDNA in a mouse ACTH-secreting cell. Intracellular storage, proteolytic processing, and secretion on stimulation.

The AtT-20 cell line, derived from the mouse anterior pituitary, synthesizes an adrenocorticotropic hormone (ACTH) precursor, proteolytically processes it to mature ACTH, stores it in secretory granules, and releases mature ACTH on stimulation with a secretagogue. A cDNA for human proinsulin inserted downstream from the SV40 early promoter in an SV40-pBR322 recombinant vector was introduced into AtT-20 cells. The stably transformed cell line, AtT-20ins4b/1, stores immunoreactive insulin, proteolytically processes proinsulin to smaller fragments, and on stimulation with secretagogues releases insulin-like material, not proinsulin, into the medium. Similarly transformed fibroblast L-cells secrete only proinsulin; they do not store it, and their secretion rate is unaffected by secretagogues. The transport mechanism for precursor ACTH thus appears to recognize other prohormones.

Adrenocorticotropic Hormone↗
PubMed1983

Effects of inhibitors of cyclic nucleotide phosphodiesterase on actions of cholecystokinin, bombesin, and carbachol on pancreatic acini.

In dispersed acini from guinea pig pancreas two inhibitors of cyclic nucleotide phosphodiesterase, Ro 20-1724 and 3-isobutyl-1-methylxanthine (IBMX), augmented the increase in amylase secretion caused by supramaximal concentrations of cholecystokinin but did not alter the stimulation of enzyme secretion caused by bombesin. The augmentations of the action of cholecystokinin caused by Ro 20-1724 or IBMX could be reproduced by 8-bromo-cAMP. When tested alone or with theophylline, cholecystokinin did not alter cAMP in pancreatic acini; however, with Ro 20-1724 or IBMX, concentrations of cholecystokinin that were supramaximal for stimulating amylase secretion caused a significant increase in cellular cAMP. These findings indicate that Ro 20-1724 and IBMX augment the action of cholecystokinin on enzyme secretion by inhibiting cyclic nucleotide phosphodiesterase and allowing a significant cholecystokinin-induced increase in cellular cAMP. IBMX but not Ro 20-1724 caused a parallel rightward shift in the dose-response curve for the stimulation of amylase secretion caused by carbachol. IBMX also caused a parallel rightward shift in the dose-response curve for the stimulation of outflux of 45Ca caused by carbachol. These results indicate that IBMX, but not Ro 20-1724, can function as a muscarinic cholinergic antagonist.

1-Methyl-3-isobutylxanthine↗
PubMed1983

Comparisons of carmustine, procarbazine, and high-dose methylprednisolone as additions to surgery and radiotherapy for the treatment of malignant glioma.

Within 3 weeks of definitive surgery, 609 patients with histologically demonstrated, supratentorial malignant glioma were randomized to receive, in addition to 6000 rads of radiotherapy, one of four treatment regimens: carmustine (BCNU), high-dose methylprednisolone, procarbazine, or BCNU plus high-dose methylprednisolone. We analyzed the data for the total randomized population and for the 527 patients (87% with glioblastoma multiforme) in whom the initial protocol specifications were met (the valid study group). Significantly longer survival was experienced by patients receiving procarbazine or BCNU alone compared to those receiving only high-dose methylprednisolone. No other pairwise comparisons demonstrated differences significant at the 0.05 level. However, the combination of BCNU plus high-dose methylprednisolone tended to be less effective than BCNU alone in patients with poor prognosis. This study indicates that BCNU and procarbazine are moderately useful agents in conjunction with radiotherapy for patients with malignant glioma. In addition, future protocols may allow use of corticosteroids in conventional dosages for treating cerebral edema and controlling symptoms; conclusions based on survival as the endpoint are unlikely to be affected by administering steroids at somewhat greater than the usual dose. More effective regimens for the treatment of malignant glioma should be sought.

Adolescent↗
PubMed1982

Secretion of intrinsic factor from dispersed mucosal cells isolated from guinea pig and rabbit stomach.

In dispersed mucosal cells prepared from rabbit and guinea pig stomach, the secretion of intrinsic factor was constant (0.3-0.4%/min) for at least 30 min incubation at 37 degrees C. Histamine or isobutyl methylxanthine increased cyclic AMP and intrinsic factor secretion in both cells preparations. Isobutyl methylxanthine potentiated and cimetidine competitively inhibited (ki = 5.10-7 M) both effects of histamine. Dibutyryl cyclic AMP (1.0 mM), also caused a 3-fold increase in intrinsic factor secretion. These results suggest that in rabbit and guinea pig histamine interacts with H2-receptors to increase cyclic AMP which mediates the rise in the rate of intrinsic factor secretion.

1-Methyl-3-isobutylxanthine↗
PubMed1982

Synthesis and some pharmacological properties of Z-Tyr(SO3H)-Met-Gly-Trp-Met-Asp(Phe-NH2)-OH, a 32-beta-aspartyl analogue of cholecystokinin (pancreozymin) 27-33.

The heptapeptide carbobenzoxy-L-tyrosyl(O-sulfate)-L-methionylglycyl-L-tryptophyl-L-methionyl-L- aspartyl-beta-L-phenylalanine amine (Z-32-beta-Asp-CCK-27-33) was synthesized and tested for its ability to stimulate secretion from dispersed pancreatic acini in vitro, to increase protein secretion from cat pancreas in vivo, and to cause contraction of guinea pig gallbladder in situ. In increasing amylase secretion in vitro, the Z-32-beta-Asp-CCK-27-33 was equal in efficacy with but approximatively one-third as potent as the Boc-CCK-27-33, and when tested in vivo its activity is approximately 10 Ivy dog units (Idu)/microgram. In stimulation of the contraction of the gallbladder, it showed an activity lower than 1 Idu/microgram. This analogue has more pancreozyminic activity than cholecystokin-like activity. This seems to indicate different affinities for the two receptors.

Amylases↗
PubMed1982

Effects of inhibitors of cyclic nucleotide phosphodiesterase on the actions of vasoactive intestinal peptide and secretin on pancreatic acini.

Theophylline, 3-isobutyl-1-methylxanthine (IBMX), and Ro 20-1724 each augmented the increase in cAMP and the stimulation of amylase secretion caused by vasoactive intestinal peptide (VIP) or secretin. With IBMX the dose-response curve for the stimulation of amylase secretion caused by VIP or secretin spanned a range of lower concentrations than did that obtained with Ro 20-1724, which in turn spanned a range of lower concentrations than did that obtained with theophylline. The configuration of the dose-response curve for the action of VIP on cAMP differed with each phosphodiesterase inhibitor tested. With Ro 20-1724 the dose-response curve was monophasic, whereas with the two methylxanthines the dose-response curve was biphasic. With theophylline the magnitude of the second component of the dose-response curve was larger than the first; with IBMX the magnitude of the first component was larger than the second. The configuration of the dose-response curve for the action of secretin on cAMP also differed with each phosphodiesterase inhibitor tested. With theophylline the dose-response curve was monophasic, whereas with Ro 20-1724 and IBMX the dose-response curve was biphasic. With Ro-20-1724 the magnitude of the second component of the dose-response curve was larger than the first; with IBMX the magnitude of the first component was larger than the second. These results indicate that cAMP is compartmentalized in pancreatic acinar cells and that the different compartments of cAMP are affected differently by various inhibitors of cyclic nucleotide phosphodiesterase. These findings also suggest that the different compartments of cAMP are acted on by phosphodiesterases with different sensitivities to various inhibitors.

1-Methyl-3-isobutylxanthine↗
PubMed1981

mRNA for the rat uterine estrogen-induced protein. Translation in vitro and regulation by estrogen.

Poly(A)-rich RNA was isolated from uterus and brain of immature rats and translated in the rabbit reticulocyte lysate cell-free protein-synthesizing system. The translation products were analyzed by 1) specific immunoprecipitation using antibodies directed against purified estrogen-induced protein (IP), 2) two-dimensional gel electrophoresis, and 3) partial protease digestion of selected spots from two-dimensional gels. These analyses showed that IP mRNA from both uterus and brain is faithfully translated in the cell-free system. Comparison of translation products derived from uterine mRNA of untreated and 1-h estrogen-treated rats revealed a specific increase in the ability of mRNA from treated animals to direct the synthesis of IP. Thus, estrogen stimulation of IP synthesis results, at least partly, from increased uterine concentrations of IP mRNA.

Animals↗
PubMed1980

Randomized comparisons of radiotherapy and nitrosoureas for the treatment of malignant glioma after surgery.

Within three weeks of definitive surgical intervention, 467 patients with histologically proved malignant glioma were randomized to receive one of four treatment regimens: semustine (MeCCNU), radiotherapy, carmustine (BCNU) plus radiotherapy, or semustine plus radiotherapy. We analyzed the data for the total randomized population and for the 358 patients in whom the initial protocol specifications were met (the valid study group). Observed toxicity included acceptable skin reactions secondary to radiotherapy and reversible leukopenia and thrombocytopenia due to chemotherapy. Radiotherapy used alone or in combination with a nitrosourea significantly improved survival in comparison with semustine alone. The group receiving carmustine plus radiotherapy had the best survival, but the difference in survival between the groups receiving carmustine plus radiotherapy and semustine plus radiotherapy was not statistically significant. The combination of carmustine plus radiotherapy produced a modest benefit in long-term (18-month) survival as compared with radiotherapy alone, although the difference between survival curves was not significiant at the 0.05 level. This study suggests that it is best to use radiotherapy in the post-surgical treatment of malignant glioma and to continue the search for an effective chemotherapeutic regimen to use in addition to radiotherapy.

Aged↗
PubMed1980

Brain tubulin and actin cDNA sequences: isolation of recombinant plasmids.

Rat brain mRNA enriched for tubulin and actin sequences was used to prepare double stranded cDNA. A library of recombinant clones was constructed by inserting the dsDNA into the Pst1 site of pBR322 plasmid and transformation of E. coli chi 1776 host. Clones bearing sequences coding for tubulin and actin were identified and characterized.

Actins↗
PubMed1980

The importance of the amino acid in position 27 of cholecystokinin in determining its biological activity on pancreatic acini.

We tested the synthetic C-terminal heptapeptide of cholecystokinin, which has the same biologic activity as cholecystokinin, and various synthetic analogs of the C-terminal heptapeptide for their abilities to increase amylase secretion from dispersed acini prepared from guinea-pig pancreas. We found that altering the chemical character of the amino acid in position 27 altered the potency with which the peptide stimulated amylase secretion but did not alter the efficacy of the peptide. We also found that, in the amino acid in position 27, the major function of the side-chain seems to be to position to the sulfate ester group at a proper distance from the backbone of the peptide chain, whereas the chemical structure of the side-chain per se seems to be of relatively minor importance.

Amino Acid Sequence↗
PubMed1980

Action of theophylline on secretagogue-stimulated amylase release from dispersed pancreatic acini.

In dispersed acini from guinea pig pancreas, theophylline did not alter basal amylase release, but had three functionally distinct modes of action on the stimulation of amylase release caused by various secretagogues. 1) At relatively low concentrations (0.1-1.0 mM), theophylline augmented the increase in enzyme secretion caused by vasoactive intestinal peptide, secretin, or 8-bromoadenosine 3',5'-monophosphate, but did not alter the increase in amylase release caused by other secretagogues. 2) At intermediate concentrations (1-10 mM), theophylline selectively altered the increase in enzyme secretion caused by carbamylcholine, but did not alter the effects of cholecystokinin or bombesin, secretagogues whose modes of action are similar to that of cabamylcholine. 3) At high concentrations (greater than 10 mM), theophylline inhibited the increase in enzyme secretion caused by all secretagogues tested.

Amylases↗
PubMed1980

Cholecystokinin (pancreozymin). Synthesis and properties of the N alpha-acetyl-derivative of cholecystokinin 27-33.

The acetyl-derivative of the biologically active C-terminal 7-peptide portion of cholecystokinin (CCK), N-acetyl-O-sulfate-L-tyrosyl-L-methionyl-glycyl-L-tryptophyl-L-methionyl-L-aspartyl-L-phenylalanine amide was prepared by the condensation of 2-peptide segments with 1-isobutyloxycarbonyl-2-isobutyloxy-1,2-dihydroquinoline as coupling reagent. The N-terminal residue, tyrosine was incorporated by the active ester method. The same 7-peptide was prepared also by stepwise chain-lengthening, starting with the C-terminal residue. The 9-fluorenylmethyloxycarbonyl group was applied for the protection of alpha-amino functions. In the release of amylase from acinar cells of the pancreas of guinea pigs, the acetyl-7-peptide amide was about 3 times more potent than CCK 27-33 and equal in potency to CCK 26-33. The new derivative strongly stimulated the contraction of the in situ guinea pig gall bladder.

Amino Acid Sequence↗