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M D Brennan

Publications and source records attributed to M D Brennan.

At least 37 records · Page 2Linked to original sources
PubMed1990

Tissue-specific expression phenotypes of Hawaiian Drosophila Adh genes in Drosophila melanogaster transformants.

Interspecific differences in the tissue-specific patterns of expression displayed by the alcohol dehydrogenase (Adh) genes within the Hawaiian picture-winged Drosophila represent a rich source of evolutionary variation in gene regulation. Study of the cis-acting elements responsible for regulatory differences between Adh genes from various species is greatly facilitated by analyzing the behavior of the different Adh genes in a homogeneous background. Accordingly, the Adh gene from Drosophila grimshawi was introduced into the germ line of Drosophila melanogaster by means of P element-mediated transformation, and transformants carrying this gene were compared to transformants carrying the Adh genes from Drosophila affinidisjuncta and Drosophila hawaiiensis. The results indicate that the D. affinidisjuncta and D. grimshawi genes have relatively higher levels of expression and broader tissue distribution of expression than the D. hawaiiensis gene in larvae. All three genes are expressed at similar overall levels in adults, with differences in tissue distribution of enzyme activity corresponding to the pattern in the donor species. However, certain systematic differences between Adh gene expression in transformants and in the Hawaiian Drosophila are noted along with tissue-specific position effects in some cases. The implications of these findings for the understanding of evolved regulatory variation are discussed.

Alcohol Dehydrogenase↗
PubMed1989

Isolation of a cDNA encoding functional Drosophila alcohol dehydrogenase in Escherichia coli and purification of the bacterially produced enzyme.

Because of the severe limitations on growing large quantities of Drosophila affinidisjuncta in the laboratory, direct purification of alcohol dehydrogenase (ADH) from this species has proven impossible. As an alternative source of this enzyme, a cDNA encoding functional ADH was isolated from a newly constructed cDNA library made from larval poly(A)-containing RNA. The cDNA was recovered by virtue of its hybridization to a previously isolated genomic ADH gene. Nucleotide sequence analysis confirmed the identity of the newly isolated cDNA. When the cDNA was inserted in the proper orientation downstream of the lac promoter on the vector pUC8, the cDNA directed the synthesis of functional ADH by the bacterial host. The bacterially produced enzyme was purified to homogeneity and used to elicit polyclonal antibodies in rabbits. The purified ADH has identical apparent subunit molecular weight to that of authentic ADH in larval fly extracts as determined by immunoblotting. Further, comparisons of the kinetic parameters of the bacterially produced enzyme and ADH activity in larval fly extracts indicate similar substrate preferences, pH dependencies, and Km values for 2-propanol and NAD. These results show that expression of a cDNA in Escherichia coli is a valid strategy for isolation of an ADH that would otherwise be difficult or impossible to purify.

Alcohol Dehydrogenase↗
PubMed1988

Expression of tumor-associated aldehyde dehydrogenase gene in rat hepatoma cell lines.

Significant changes in aldehyde dehydrogenase (ALDH) activity occur during rat hepatocarcinogenesis in vivo. To compare the structure and expression of the tumor aldehyde dehydrogenase gene in rat hepatoma cell lines and normal rat liver, several rat hepatoma cell lines, including HTC, H4-II-EC3, JM2, McA-RH7777, and four lines established in this laboratory have been examined for T-ALDH gene expression using a tumor ALDH complementary DNA. Northern blot analysis of polyadenylate-containing RNA from log-phase cells and normal rat liver with T-ALDH complementary DNA indicates production of a single major 1.7-kilobase transcript in the high activity lines HTC, JM2, RLT-2M, RLT-3C, RLT-9F, and intermediate activity line RLT-5G. There is a direct correlation between expression of T-ALDH enzyme activity and the amount of 1.7-kilobase transcript. S1 nuclease protection experiments confirm that there is only one major T-ALDH transcript in the high activity lines. Thus, cell line differences in T-ALDH activity are reflected in the level of a single T-ALDH transcript. Southern analysis was used to identify the T-ALDH gene in genomic DNA. The results indicate that no significant amplification or rearrangement of the T-ALDH gene has occurred in these hepatoma cells. DNA methylation has been proposed to play an important role in gene expression. Genomic DNA from HTC, JM2, McA-RH7777, H4-II-EC3, RLT-2M, RLT-9F, RLT-3C, RLT-5G, rat embryo and normal rat liver were digested with MspI and HpaII to examine methylation patterns. A digestion pattern consistent with hypomethylation was detected only in DNA from the high T-ALDH activity cell lines HTC, JM2, RLT-2M, and RLT-9F. This suggests that constitutive expression of T-ALDH in the hepatoma cells is related to changes in DNA methylation patterns.

Aldehyde Dehydrogenase↗
PubMed1988

Characterization of a functional recombinant rat liver aldehyde dehydrogenase: expression as a non-fusion protein in E. coli.

A cDNA encoding a rat liver inducible aldehyde dehydrogenase carried in a pUC8 plasmid is expressed in E. coli as a dimeric enzyme molecule functionally and physically identical to the authentic rat enzyme. The cDNA appears to be transcribed using the lac promoter, but is translated from an initiator codon 174 base pairs from the 5' end of the cDNA. The aldehyde dehydrogenase polypeptide is not produced as a fusion protein. This is the first example of the production by E. coli of a catalytically active, multimeric eukaryotic protein which is not a fusion protein.

Aldehyde Dehydrogenase↗
PubMed1988

Molecular analysis of alcohol dehydrogenase electromorphs in wild type and transformed Drosophila melanogaster.

The protein expressed by the alcohol dehydrogenase locus (Adh) in D. melanogaster comprises a small group of electromorphs. We are able to study the expression of these electromorphs by electrophoretic separation and subsequent probing of blots of the separated polypeptides with antiserum for alcohol dehydrogenase (ADH). In the present study we have utilized this technique to study and compare the ADH electromorphs in wild type D. melanogaster with D. melanogaster transformants which carry an Adh gene from D. grimshawi, D. hawaiiensis or D. affinidisjuncta and produced functional ADH (10, 19). We have determined that polypeptides are produced by the donor loci in the transformed flies and further show that although the molecular weight of the expressed polypeptides is similar to D. melanogaster electromorphs, the isoelectric points are not similar. Thus this methodology offers the potential to study naturally occurring ADH electromorphs and null alleles independent of enzymatic activity assays.

Alcohol Dehydrogenase↗
PubMed1988

Complex developmental regulation of the Drosophila affinidisjuncta alcohol dehydrogenase gene in Drosophila melanogaster.

During development, the alcohol dehydrogenase genes of Drosophila melanogaster and D. affinidisjuncta are expressed in similar, yet distinct, tissue- and stage-specific patterns. Transcripts from both of these genes arise from two promoters (distal and proximal) that also display tissue and stage specificity. We used P-element-mediated transformation to introduce the D. affinidisjuncta Adh gene into the germ line of D. melanogaster. We show that the D. affinidisjuncta Adh gene is expressed at comparable overall levels in both species and that the tissue- and stage-specific expression for this gene (including promoter utilization) is similar in the donor and the host species. However, in some details, the expression of the D. affinidisjuncta gene in D. melanogaster resembles the host pattern, and one novel tissue-specific expression phenotype is displayed by transformants. In general, our results suggest that there has been strong conservation of cis- and trans-acting regulatory factors since the divergence of the two species but that this conservation has not been perfect.

Alcohol Dehydrogenase↗
PubMed1988

Tissue-specific regulatory differences for the alcohol dehydrogenase genes of Hawaiian Drosophila are conserved in Drosophila melanogaster transformants.

Naturally occurring regulatory variation is a source of genetic variability that is well documented but poorly understood. Two members of the Hawaiian picture-winged Drosophila, D. affinidisjuncta and D. hawaiiensis, display markedly different levels of alcohol dehydrogenase (alcohol: NAD+ oxidoreductase, EC 1.1.1.1) in the larval midgut and Malpighian tubules. To analyze the regulation of the alcohol dehydrogenase genes from these two species, their homologous alcohol dehydrogenase genes were cloned and introduced, via P element-mediated transformation, into the germ line of Drosophila melanogaster. Histochemical and electrophoretic analyses of larval transformants demonstrate that major differences in the tissue-specific levels of alcohol dehydrogenase production are characteristic of the alcohol dehydrogenase genes themselves. While these results do not directly address possible species-specific differences in the tissue distribution of trans-acting regulatory components, they indicate that demonstrable differences in cis-dominant regulatory information are sufficient to account for the observed regulatory variation.

Alcohol Dehydrogenase↗
PubMed1988

Cloning and complete nucleotide sequence of a full-length cDNA encoding a catalytically functional tumor-associated aldehyde dehydrogenase.

To study the mechanism(s) controlling expression of the tumor-associated aldehyde dehydrogenase (tumor ALDH), which appears during rat hepatocarcinogenesis, cDNAs encoding this isozyme were cloned and identified with an antibody probe. Poly(A)-containing RNA from HTC rat hepatoma cells, which have been shown to possess high levels of tumor ALDH, was used as template to synthesize double-stranded cDNA. The cDNA was methylated to protect internal sites. Two different synthetic DNA linkers were added sequentially to the cDNA to insure correct orientation for expression from the lac promoter of pUC8. A library of 100,000 independent members carrying inserts greater than 1 kilobase was obtained. From this library, two apparently identical tumor ALDH clones, differing only in size, were identified with an indirect immunological probe. The larger of the cDNA clones identified, pTALDH, was chosen for further study. Interestingly, since tumor ALDH is a dimeric enzyme, pTALDH directs synthesis of a functional tumor ALDH in the bacterial cell. The cDNA sequence has been confirmed by comparison to the amino acid sequence of tumor ALDH purified from HTC cells.

Aldehyde Dehydrogenase↗
PubMed1987

Heterophilic serum antibodies: a cause for falsely elevated serum thyrotropin levels.

We report two cases of euthyroid patients with inappropriately elevated serum levels of thyrotropin (TSH) in whom investigation led to the detection of heterophilic antibodies. In addition, we compare the performance of two TSH immunometric assay systems in this setting. Even when monoclonal assay systems are used, TSH results must be critically interpreted when they are at variance with clinical and other biochemical findings.

Adult↗
PubMed1987

Pathology of the thyroid in amiodarone-associated thyrotoxicosis.

Among 83 consecutive patients operated on for thyrotoxicosis at the Mayo Clinic between January 1, 1980 and May 1, 1986, four had a combination of pathologic findings that were similar and unexpected: involutional changes; degenerative and destructive follicular lesions; and zones of fibrosis. These four, but none of the remaining 79, were being treated with amiodarone for cardiac tachyarrhythmias; this drug is known to be taken up by the thyroid gland. Characteristically, small groups of involuted follicles exhibited varying degrees of damage ranging from degenerative changes in a few lining cells to total follicular destruction. Damaged follicular cells were swollen and featured granular or vacuolated cytoplasm. This type of cytoplasmic alteration has been reported to occur in pneumocytes and hepatocytes damaged by amiodarone. Therefore, the drug probably is the cause of the thyroid cell damage, and follicular disruption (with consequent release of iodothyronines into the circulation) is likely to be an important contributor to the associated thyrotoxicosis.

Adult↗
PubMed1987

Amiodarone-associated thyrotoxicosis (AAT): experience with surgical management.

Amiodarone (Cordarone), a benzofuran derivative containing two atoms of iodine per molecule (37% of molecular weight), has recently been released in the United States for the management of refractory cardiac arrhythmias. While it was still under investigation, 529 patients were managed with this drug at the Mayo Clinic between 1981 and 1986; eight of them (1.5%) had thyrotoxicosis 12 to 37 months (median, 28 months) after initiation of treatment. Low 24-hour iodine-131 uptake (less than 4%) and recognized refractoriness to thionamide treatment precluded the use of radioablation or antithyroid drugs. Cessation of the amiodarone treatment was considered undesirable from a cardiac standpoint. Near-total thyroidectomies were undertaken in six patients, and rapid correction of hyperthyroidism followed. There were no intraoperative or postoperative arrhythmias. The median postoperative hospital stay was 4.3 days (range, 2 to 13 days). Subsequently, all patients required thyroid hormone replacement. Near-total thyroidectomy has proved a safe and effective treatment for amiodarone-associated thyrotoxicosis in patients who required drug continuation.

Adult↗
PubMed1986

Developmentally regulated RNA transcripts coding for alcohol dehydrogenase in Drosophila affinidisjuncta.

The organization of the gene coding for alcohol dehydrogenase (Adh) in Drosophila affinidisjuncta has been determined by physically mapping Adh RNA transcripts to cloned genomic DNA. Two distinct transcript types accumulate with developmental specificity. Because only a single genomic Adh locus is detected in D. affinidisjuncta, and since all Adh transcripts appear to be identical except at their termini, the two Adh RNA types are products of the same gene. One type of transcript, abundant in adults, contains a small 5' terminal exon that is completely lacking in the other type of transcript, which accumulates in larvae. This 5' end difference suggests that the D. affinidisjuncta Adh gene, like the homologous gene from the distantly related species D. melanogaster, is expressed from two promoters. According to the transcription map, these D. affinidisjuncta promoters are separated by approximately 560 base pairs of genomic DNA sequence. D. affinidisjuncta Adh transcripts also resemble D. melanogaster Adh transcripts in both their overall organization and their developmental distribution. Multiple 3' ends are responsible for the size heterogeneity of both types of D. affinidisjuncta Adh RNA, and some of these also appear with stage specificity.

Alcohol Dehydrogenase↗
PubMed1986

Hashimoto's thyroiditis. An uncommon cause of painful thyroid unresponsive to corticosteroid therapy.

The records of eight patients with thyroid tenderness secondary to Hashimoto's thyroiditis were reviewed. The pathologic characteristics of thyroid tissue sections from these patients were compared with those from patients with nontender Hashimoto's thyroiditis, and no difference was identified. Laboratory features helpful in distinguishing tender Hashimoto's thyroiditis from subacute (de Quervain's) thyroiditis include normal erythrocyte sedimentation rate, significant 131I uptake, and significant antithyroid antibody titer. Diagnosis should be confirmed by fine-needle aspiration biopsy. Corticosteroid therapy was unsuccessful in treating these patients; L-thyroxine and aspirin were successful more often. Two patients required thyroidectomy to control pain.

Adrenal Cortex Hormones↗
PubMed1985

Multidisciplinary management of acromegaly and its deformities.

The skeletal deformities associated with acromegaly are not reversed by correction of abnormal growth hormone dynamic. We describe a patient with acromegaly in whom facial reconstruction was undertaken after successful removal of a pituitary adenoma.

Acromegaly↗
PubMed1985

Volume measurements of normal orbital structures by computed tomographic analysis.

Computed tomographic digital data and special off-line computer graphic analysis were used to measure volumes of normal orbital soft tissue, extraocular muscle, orbital fat, and total bony orbit in vivo in 29 patients (58 orbits). The upper limits of normal for adult bony orbit, soft tissue exclusive of the globe, orbital fat, and muscle are 30.1 cm3, 20.0 cm3, 14.4 cm3, and 6.5 cm3, respectively. There are small differences in men as a group compared with women but minimal difference between right and left orbits in the same person. The accuracy of the techniques was established at 7%-8% for these orbit structural volumes in physical phantoms and in simulated silicone orbit phantoms in dry skulls. Mean values and upper limits of normal for volumes were determined in adult orbital structures for future comparison with changes due to endocrine ophthalmopathy, trauma, and congenital deformity.

Adult↗
PubMed1985

Bilateral subtotal adrenal resection for bilateral pheochromocytomas in multiple endocrine neoplasia, type IIa: a case report.

Our treatment philosophy for the management of the adrenal glands in patients with multiple endocrine neoplasia, type IIa has been bilateral total adrenalectomy. In the patient described, exceptional and pressing reasons necessitated preservation of adrenocortical function. Bilateral pheochromocytomas were resected with preservation of the adrenal cortices. Adrenocortical function was normal 36 months after resection. Although the patient remained symptom free, elevated basal immunoreactive plasma calcitonin levels suggested the presence of residual or metastatic medullary thyroid carcinoma. There was no evidence of recurrent pheochromocytoma.

Adrenal Gland Neoplasms↗
PubMed1984

Introduction of a functional P element into the germ-line of Drosophila hawaiiensis.

When a plasmid carrying a P-transposable element (derived from Drosophila melanogaster) is injected into young embryos of D. hawaiiensis, the P-element sequence from the plasmid transposes into the germ-line chromosomes. The introduction of this P element into D. hawaiiensis provides an opportunity to study the behavior of the transposable element in a novel context. Germ-line transposition and numerical increase of the P elements are readily detected in D. hawaiiensis. Thus these aspects of P-element function do not require chromosomal or cytoplasmic properties that are unique to D. melanogaster. Since D. hawaiiensis is among those Drosophila that are most distantly related to D. melanogaster, these results suggest that P-element-mediated transformation may function in many species.

Animals↗