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Biomedical subjects

J M Carroll

Publications and source records attributed to J M Carroll.

11 recordsLinked to original sources
PubMed1992

Cytosolic free calcium and gene expression during chemical hypoxia.

Understanding the cellular response to hypoxia may help elucidate the role of altered oxidation in neuronal death or abnormal cell function. In PC12 cells, 30 min of chemical hypoxia (i.e., KCN) reduced ATP concentrations by 92%, but diminished viability by only 10%. Ten minutes of hypoxia increased cytosolic free calcium ([Ca2+]i) 2.5-fold above control, but after 30 min of hypoxia, [Ca2+]i was slightly below that of nonhypoxic cells. Short periods of hypoxia also exaggerated the K(+)-induced elevation of [Ca2+]i, but by 30 min these ATP-depleted cells reestablished a calcium gradient that was equal to nonhypoxic, K(+)-depolarized cells. Thus, 30 min of severe ATP depletion left [Ca2+]i and viability relatively unaffected. Nerve growth factor caused slight, but significant, improvements in ATP and viability of hypoxic cells, but had no effect on [Ca2+]i. Although [Ca2+]i was equivalent in control and hypoxic cells after 30 or 60 min, hypoxia abolished the K(+)-stimulated elevation of [Ca2+]i. The nerve growth factor induction of c-fos, an indicator of the genomic response, was diminished by approximately 80%. Thus, hypoxic PC12 cells with greatly reduced ATP stores maintained normal [Ca2+]i, but their ability to respond to external stimulation was impaired. Further, the reduced oxidation that occurs in the brain in a variety of pathological conditions may interfere with the cellular response to stimulation and growth factors.

Adenosine Triphosphate↗
PubMed1992

Characterization of the human involucrin promoter using a transient beta-galactosidase assay.

Involucrin, a component of the cornified cell envelope, is expressed specifically in differentiating keratinocytes of stratified squamous epithelia. To explore the regulation of involucrin expression, 3.7 kb of upstream sequences of the human involucrin gene was cloned into a plasmid containing a beta-galactosidase reporter gene and transfected into early passage keratinocytes and a variety of human cell types. The full-length construct gave maximal and tissue-specific expression. Deletion analysis showed that sequences between 900 and 2500 bp upstream of the transcriptional start site and the intron located between the transcriptional and translational start sites were required for maximal expression. Further analysis of the intron indicated that its effects on expression were independent of it being present in nascent RNA and suggested that sequences within the intron have regulatory activity. These results suggest that the involucrin intron operates in vivo to regulate expression in the epidermis.

Breast Neoplasms↗
PubMed1991

Effects of second messenger system activation on functional expression of tyrosine hydroxylase fusion gene constructs in neuronal and nonneuronal cells.

A genomic clone for rat tyrosine hydroxylase (TH) was isolated and a fragment containing 503 bp upstream of the transcription start site was sequenced. The BamHI/AluI fragment was inserted into a plasmid carrying the coding sequence for bacterial chloramphenicol acetyltransferase (CAT). Another construct with the 5' sequence truncated to -151 bp also was prepared. When these were introduced into several mammalian cell lines, including C6 glioma, BE(2) neuroblastoma, CV-1 or Ltk- fibroblasts, different basal levels of CAT expression were observed. In the fibroblast lines, THCAT constructs were not expressed unless the cells were treated with forskolin or TPA. However, the low basal expression was not correlated to endogenous expression as THCAT constructs expressed comparably in BE(2)C, HeLa, and C6 glioma. Treatment of any of the cell lines with forskolin, TPA, or a combination of the two agents stimulated the expression by at least two-fold in all cell lines and the maximally induced levels were at least 10-fold over promoterless controls. These data indicate that the essential promoter elements as well as those conferring responsivity to cyclic AMP reside within 151 bp of the transcription start site. However, the array of elements regulating cell-type expression lie, at least in part, beyond the 500-bp region examined. Further, a role for phosphorylation in the regulation of basal and induced transcription of TH is suggested.

Animals↗
PubMed1991

Differential and coordinate regulation of TH and PNMT mRNAs in chromaffin cell cultures by second messenger system activation and steroid treatment.

Primary cultures of chromaffin cells were prepared from bovine adrenal medullae and the levels of mRNA for tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT) determined. The cells expressed moderate levels of TH mRNA and low levels of PNMT mRNA. The latter appeared to be more sensitive than TH mRNA to variations in the culture medium. The treatment of cultures with agents that activate signal transduction pathways, forskolin or phorbol esters, dramatically enhanced the expression of both mRNAs. The forskolin-induced increases in the steady-state levels of TH and PNMT mRNAs occurred rapidly and were apparent within 5 hours. These data suggest that the TH and PNMT genes can be regulated by second messengers. In contrast, dexamethasone treatment dramatically increased PNMT mRNA with no change in TH mRNA. The increase in PNMT mRNA was apparent within 6 hours of addition of the drug to the culture medium.

1-Methyl-3-isobutylxanthine↗
PubMed1990

Identification of a functional glucocorticoid response element in the phenylethanolamine N-methyltransferase promoter using fusion genes introduced into chromaffin cells in primary culture.

The rat gene encoding phenylethanolamine N-methyltransferase (PNMT) was cloned and a consensus sequence for a glucocorticoid response element (GRE) was found at -513 bp, 5' to the transcriptional start site. In order to define the function of this element, fusion genes containing the PNMT promoter and a chloramphenicol acetyltransferase (CAT) reporter gene were constructed. These constructs did not express after transfection into any of 7 continuous cell lines, none of which endogenously produce PNMT. A system for transfecting chromaffin cells in primary culture was therefore devised using constructs containing 200 bp of the proenkephalin (ENK) promoter, whose expression characteristics are well known. pENK beta GAL-1, containing the ENK promoter with a lac Z reporter, was introduced into these cells and beta-galactosidase activity was visualized in situ. Approximately 90% of cells transfected were chromaffin; transfection efficiency was 5%. High levels of CAT activity were measured in chromaffin cells transfected with pENKAT12, possessing a CAT reporter. In contrast to tumor cell lines, pENKAT12 induction in these cells by forskolin and phorbol esters did not require a phosphodiesterase inhibitor. In this chromaffin system, both basal and regulated expression of the PNMT fusion genes were detected. Dexamethasone (dex) induced expression of pPNMT3000 and pPNMT900, containing the putative GRE and 3000 bp or 863 bp of PNMT promoter sequence, 4- to 10-fold. Expression of pPNMT300 and pPNMT100, which lack the GRE and contain 273 bp or 99 bp of PNMT promoter sequence, was unaffected by dex. Addition of the PNMT region spanning -490 to -863 bp conferred full dex responsiveness to a thymidine kinase promoter. Deletion of the putative GRE sequence by site-directed mutagenesis abolished the dex response. These data identify the sequence at -513 bp in the rat PNMT gene as a functional, positively acting GRE. Primary cultures of bovine chromaffin cells provide a biologically relevant expression system for transcriptional studies of catecholamine genes and their related neuropeptides.

Animals↗
PubMed1979

Multiple choroidal metastasis from bronchial carcinoid treated with photocoagulation and proton beam irradiation.

A 23-year-old man had bilateral multiple choroidal lesions eight years after removal of a bronchial carcinoid tumor. Choroidal biopsy confirmed that the clinical diagnosis and conventional radiotherapy to one eye resulted in rubeosis iridis, neovascular glaucoma, cataract, and vitreous hemorrhage. We were able to destroy the tumors in the other eye by photocoagulation and proton beam irradiation without any adverse effects. The patient is in good health 1 1/2 years after therapy and he has 6/6 (20/20) vision in the eye that was treated.

Adult↗
PubMed1978

Functional clauses and sentence segmentation.

In two experiments subjects listened to a sentence containing a brief tone, then wrote out the sentence and marked the location of the tone. The experimental sentences were biclausal with the tone placed before or after the clause break. The initial clause was either functionally complete or functionally incomplete. Functionally complete clauses contain a complete set of fully specified grammatical relations, while functionally incomplete clauses do not. In Experiment 1 tones were mislocated toward the clause break and the final word of the first clause significantly more often for functionally complete clauses. Experiment 2 replicated this finding holding deep- and surface-structure variables constant. The results indicate that functionally complete clauses are better segmentation units during sentence perception than functionally incomplete clauses. Purely structural theories of the units of sentence perception cannot account for this finding.

Adult↗
PubMed1976

Segmentation in cinema perception.

Viewers perceptually segment moving picture sequences into their cinematically defined units: excerpts that follow short film sequences are recognized faster when the excerpt originally came after a structural cinematic break (a cut or change in the action) than when it originally came before the break.

Humans↗