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Biomedical subjects

C A Schmitt

Publications and source records attributed to C A Schmitt.

At least 19 recordsLinked to original sources
PubMed2000

Genetic analysis of chemoresistance in primary murine lymphomas.

Understanding the basis of chemoresistance is a principal goal of molecular oncology. We have exploited a murine lymphoma model and retroviral gene transfer to rapidly generate a series of spontaneous tumors differing only in a gene of interest, and subsequently studied the impact of the test gene on the treatment sensitivity of tumors at their natural site. We demonstrate that the Bcl-2 oncoprotein produces multi-drug resistance when assessed in primary lymphomas in vivo. In contrast, this effect was dramatically reduced when the primary lymphomas were subjected to long-term culture, and completely missed in the standard clonogenic survival assay. This model highlights the importance of physiological test systems to address the complexity of clinical drug resistance and provides a novel strategy to evaluate compounds targeting specific genetic lesions.

Adaptation, Biological↗
PubMed1999

INK4a/ARF mutations accelerate lymphomagenesis and promote chemoresistance by disabling p53.

The INK4a/ARF locus encodes upstream regulators of the retinoblastoma and p53 tumor suppressor gene products. To compare the impact of these loci on tumor development and treatment response, the Emu-myc transgenic lymphoma model was used to generate genetically defined tumors with mutations in the INK4a/ARF, Rb, or p53 genes. Like p53 null lymphomas, INK4a/ARF null lymphomas formed rapidly, were highly invasive, displayed apoptotic defects, and were markedly resistant to chemotherapy in vitro and in vivo. Furthermore, INK4a/ARF(-/-) lymphomas displayed reduced p53 activity despite the presence of wild-type p53 genes. Consequently, INK4a/ARF and p53 mutations lead to aggressive tumors by disrupting overlapping tumor suppressor functions. These data have important implications for understanding the clinical behavior of human tumors.

Animals↗
PubMed1999

Accumulation of 3,4-dihydroxyphenylglycolaldehyde, the neurotoxic monoamine oxidase A metabolite of norepinephrine, in locus ceruleus cell bodies in Alzheimer's disease: mechanism of neuron death.

3,4-Dihydroxyphenylglycolaldehyde (DOPEGAL) is the neurotoxic monoamine oxidase A (MAO-A) metabolite of norepinephrine (NE). NE neurons in the locus ceruleus (LC) die in Alzheimer's disease (AD). To determine if DOPEGAL could contribute to NE neuron death in AD we measured levels of DOPEGAL, NE and their synthesizing enzymes in LC from AD and matched controls. We found 2.8- and 3.6-fold increases in DOPEGAL and MAO-A in AD LC neuronal cell bodies compared to controls. NE and dopamine beta-hydroxylase were increased by 3.8- and 10.7-fold, respectively. Implications for the mechanism of neuron death in AD are discussed.

Aged↗
PubMed1999

Apoptosis and therapy.

The dogma that antineoplastic treatments kill tumour cells by damaging essential biological functions has been countered by the notion that treatment itself initiates a programmed cellular response. This response often produces the morphological features of apoptosis and is determined by a network of proliferation and survival genes, some of which are differentially expressed in normal and malignant cells. Correspondingly, mutations that interfere with the initiation or execution of apoptosis may produce tumour-cell drug resistance. Remarkably, many of the genes that modulate apoptosis in response to cytotoxic drugs also affect apoptosis during tumour development; hence, the process of apoptosis provides a conceptual framework for understanding how cancer genes can influence the outcome of cancer therapy. Although the relative contribution of apoptosis to radiation and drug-induced cell death remains controversial, clinical studies have associated anti-apoptotic mutations with treatment failure. While careful preclinical and clinical studies will be necessary to resolve this point, our current understanding of apoptosis should facilitate the design of rational new therapies.

Apoptosis↗
PubMed1999

Expression and regulation by interferon-gamma of the membrane-bound complement regulators CD46 (MCP), CD55 (DAF) and CD59 in gastrointestinal tumours.

The membrane-bound complement inhibitors CD46 (membrane cofactor protein), CD55 (decay-accelerating factor) and CD59 (protectin) protect tumour cells against lysis by activated complement. In this study, a total of 14 (3 gastric, 3 colonic and 8 pancreatic) gastrointestinal tumour cell lines were examined for the expression of CD46, CD55 and CD59 with respect to the regulatory efficacy of interferon-gamma (IFN-gamma). The effects of IFN-gamma on mRNA and protein expression levels of CD46, CD55 and CD59 were evaluated by Northern blot hybridisation, RT-PCR, flow cytometry and immunostaining. In unstimulated cell lines, CD46 and CD59 transcripts were expressed at comparable levels, whereas the basal expression of CD55 mRNA was heterogeneous. The complement inhibitor proteins were detected in all cell lines using specific antibodies. Additional immunohistochemical stainings of gastrointestinal tissue specimens supported these findings. IFN-gamma evoked a weak induction of certain transcripts in a subset of the cell lines. Upregulation of protein expression was only observed in HT29 cells for CD55 and CD59 and was accompanied by a marked increase of the corresponding transcripts. We conclude that membrane-bound complement inhibitors are broadly expressed in gastrointestinal tumour cells and vary in their susceptibility to IFN-gamma. Thus, they may be involved in tumour escape mechanisms in gastric, pancreatic and colorectal cancer.

Antigens, CD↗
PubMed1998

Detection of the DCC gene product in normal and malignant colorectal tissues and its relation to a codon 201 mutation.

Protein expression of the putative tumour-suppressor gene DCC on chromosome 18q was evaluated in a panel of 16 matched colorectal cancer and normal colonic tissue samples together with DCC mRNA expression and allelic deletions (loss of heterozygosity, LOH). Determined by a polymerase chain reaction (PCR)-LOH assay, 12 of the 16 (75%) cases were informative with LOH occurring in 2 of the 12 cases. For DCC mRNA, transcripts could be detected in all analysed normal tissues (eight out of eight) by RT-PCR, whereas 6 of the 15 tumours were negative. DCC protein expression, investigated by immunohistochemistry using the monoclonal antibody 15041 A directed against the intracellular domain, was homogeneously positive in all normal tissue samples. In tumour tissues, no DCC protein was seen in 11 out of 16 samples (69%). For the DCC codon 201, we found a loss of a wild-type codon sequence caused by mutation or LOH in at least 8 out of 15 cases (53%) compared with the corresponding normal tissue. DCC protein expression was undetectable in eight of the nine tumours missing both wild-type codons. Only one of the five tumours with retained DCC protein expression had no detectable wild-type codon 201. In addition, 9 out of 15 normal tissue specimens were mutated in codon 201. In two out of three cases with homozygous wild-type codons in peripheral blood lymphocyte (PBL) DNA, mutations were already observed in the tumour adjacent normal colonic mucosa. We conclude that DCC immunostaining should be introduced in the clinicopathological routine because of its strong correlation with the known prognostic markers 18q LOH and mutation of codon 201.

Blotting, Southern↗
PubMed1997

Norepinephrine transmitter metabolite induces apoptosis in differentiated rat pheochromocytoma cells.

3,4-Dihydroxyphenylglycolaldehyde (DOPEGAL) is the monoamine oxidase A metabolite of norepinephrine and epinephrine. DOPEGAL, but not other metabolites, kills differentiated PC-12 cells. However, the type of DOPEGAL induced cell death, whether necrosis or apoptosis, is not known. To determine the type of cell death triggered by DOPEGAL, PC-12 cells cultured in the presence or absence of 30 microM DOPEGAL were examined by electron microscopy and DNA agarose gel electrophoresis for characteristic features of apoptosis. Results indicate that DOPEGAL induces apoptosis in these cells. Implications for degenerative diseases are discussed.

Adrenergic alpha-Agonists↗
PubMed1996

Norepinephrine transmitter metabolite is a selective cell death messenger in differentiated rat pheochromocytoma cells.

3,4-Dihydroxyphenylglycolaldehyde (DOPEGAL) is the monoamine oxidase A (MAO-A) metabolite of norepinephrine (NE) and epinephrine (Epi). Oxidative metabolites of amines are predicted toxins. In this study we determine the toxicity of DOPEGAL, its tautomer 2',3,4-trihydroxyacetophenone (THAP) as well as NE, Epi and their oxidative and methylated metabolites in cultures of differentiated PC-12 cells. At 59.5 microM DOPEGAL, THAP and Epi, but not NE or other NE or Epi metabolites decreased PC-12 cells by 43.8%, 26.7% and 16.8% respectively. DOPEGAL toxicity was concentration and time dependent. Possible implications for degenerative diseases are discussed.

Acetophenones↗
PubMed1994

Blood pressure regulation in Alzheimer's disease.

Brain neurons which regulate blood pressure (BP), including the C-1 tonic vasomotor neurons, degenerate in Alzheimer's disease (AD). This study determines whether BP is decreased in AD. We reviewed records of three autopsy proven AD patients. Medical causes for decreased BP were investigated. Yearly averages for systolic (SBP), diastolic (DBP), mean arterial (AP) blood pressure and pulse pressure (PP) were calculated. BP in the year of diagnosis was compared to the sum of all BP in subsequent years. In addition, each yearly measurement through the course of AD was compared to its counterpart in the year of diagnosis. Three BP measurements were significantly decreased by from 6.9% to 15.9% in all patients when BP in the year of diagnosis was compared to the sum of each pressure in subsequent years. Sustained BP declines started in the third to fourth year after diagnosis of AD and continued for up to 9 years. The PP was decreased by 19.9% in one patient. There was a strong correlation between the number of C-1 neurons in these cases and their AP and SBP in the years after diagnosis. Hypothalamic phenylethanolamine N-methyltransferase activity was decreased by 63% in AD compared to control cases. Neurofibrillary tangles were found in the paraventricular nucleus of the hypothalamus in an AD case. We postulate that BP is altered in AD as neurons which regulate it degenerate.

Aged↗
PubMed1989

Clue cells in bacterial vaginosis: immunofluorescent identification of the adherent gram-negative bacteria as Gardnerella vaginalis.

Clue cells are epithelial cells covered by adherent gram-negative rods, observed in vaginal smears from women with bacterial vaginosis. Immunofluorescence studies were used to identify the gram-negative bacteria adhering to clue cells. Specific antisera to four common gram-negative vaginal bacteria (Gardnerella, Bacteroides, Fusobacterium, and Mobiluncus) were prepared by long-term, multiple, small-inoculum immunization of rabbits. Cross-reactivity with heterologous common vaginal bacteria was removed by absorption against whole cells of heterologous bacteria and by serial dilution. Gardnerella vaginalis was most often observed adhering to the surface of clue cells and was detected on the surface of exfoliated vaginal epithelial cells significantly more frequently and in higher numbers than were Mobiluncus, Bacteroides, and Fusobacterium, suggesting that this species of gram-negative bacteria is responsible for clue cell formation.

Bacteroides↗
PubMed1988

Thyroid hormone regulates alpha and alpha + isoforms of Na,K-ATPase during development in neonatal rat brain.

The brain contains two molecular forms of Na,K-ATPase designated alpha found in non-neuronal cells and neuronal soma and alpha + found in axolemma. Previously we have shown that the abundance of both forms (determined by immunoblots) as well as Na,K-ATPase activity increases 10-fold between 4 days before and 20 days after birth (Schmitt, C. A., and McDonough, A. A. (1986) J. Biol. Chem. 261, 10439-10444). Hypothyroidism in neonates blunts these increases. Neonatal, but not adult brain Na,K-ATPase is thyroid hormone (triiodothyronine, T3) responsive. This study defines the period during which brain Na,K-ATPase responds to T3. The start of the critical period was defined by comparing Na,K-ATPase activity and alpha and alpha + abundance in hypothyroid and euthyroid neonates (birth to 30 days of age). For all parameters, euthyroid was significantly higher by 15 days of age. The end of the critical period was defined by dosing hypothyroid neonates with T3 daily (0.1 micrograms/g body weight) beginning at increasing days of age, and sacrificing all at 30 days then assaying enzyme activity and abundance. Those starting T3 treatment on or before day 19 were restored to euthyroid levels of Na,K-ATPase activity and abundance, while those starting T3 treatment on or after day 22 remained at hypothyroid levels of enzyme activity and abundance. We conclude that brain Na,K-ATPase alpha and alpha + isoforms are sensitive to T3 by as late as 15 days of age and that the period of thyroid hormone responsiveness is over by 22 days.

Aging↗
PubMed1988

Thyroid hormone coordinately regulates Na+-K+-ATPase alpha- and beta-subunit mRNA levels in kidney.

Synthesis of the sodium pump, Na+-K+-ATPase, is regulated by thyroid hormone in responsive tissues. The purpose of this study was to determine if triiodothyronine (T3) regulates the concentration of the mRNAs coding for the two enzyme subunits, alpha and beta, and the time course of the response. A single dose of T3 (250 micrograms/100 g body wt) was administered to hypothyroid rats that were killed at various times after injection. In the kidney cortexes of the T3-injected animals, as well as hypothyroid and euthyroid rats, alpha- and beta-mRNA concentrations were measured by dot blot using cDNAs corresponding to the two mRNAs; alpha-subunit abundance was measured by Western blot using antibodies to the enzyme, and Na+-K+-ATPase activity was measured enzymatically. alpha- and beta-mRNAs increased coordinately, after a 6-h time lag to 1.6-fold over hypothyroid levels by 12 h after T3. alpha-Subunit abundance increased significantly by 48 h and to 1.4-fold over hypothyroid by 72 h after T3. Na+-K+-ATPase activity increased with the same time course as the increase in alpha-subunit abundance to 1.3-fold over hypothyroid by 72 h after T3. We conclude that T3 regulates Na+-K+-ATPase synthesis and activity by coordinately increasing the mRNAs of both the alpha- and beta-subunits of the enzyme.

Animals↗
PubMed1987

Isozymes of dog heart Na+-K+-ATPase are immunologically similar to isozymes in brain.

The sodium pump, Na+-K+-ATPase, possesses two populations of cardiac glycoside-binding sites in cardiac tissue. This has been observed in other tissues such as brain where the two sites have been assigned to isozymes of Na+-K+-ATPase termed alpha and alpha +. In a previous study [Am. J. Physiol. 248 (Cell Physiol. 17): C247-C251, 1985], we were unable to demonstrate the presence of an alpha +-form in guinea pig heart sarcolemmal membranes using antibody probes. In the present study, using similar methodology, we show that dog, but not rat or guinea pig, sarcolemmal membranes contain two immunologically distinct alpha-subunits. The antibody-binding characteristics of dog heart alpha and alpha + are similar to the forms found in brain. The relative abundance of the two isozymes, estimated by labeling the sarcolemmal membranes with fluorescein 5'-isothiocyanate, was about equal. We conclude that the two populations of ouabain-binding sites in dog heart may result, at least in part, from the presence of the two isozymes of Na+-K+-ATPase in cardiac tissue of this species.

Animals↗
PubMed1986

Developmental and thyroid hormone regulation of two molecular forms of Na+-K+-ATPase in brain.

In the brain there are two isozymes of Na+-K+-ATPase differing in their catalytic subunits: alpha, indistinguishable from the kidney form of alpha, and alpha +, found in axolemma. The time course of the increase in each alpha during development was described by quantitating the abundance of each form, studied in unpurified membranes resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, with specific antibodies and with fluorescein 5'-isothiocyanate. Both the alpha and alpha + subunits, quantitated with antibodies, increased 10-fold in abundance from 18 days gestation to 20 days of age, with alpha + increasing more rapidly than alpha early in development. A 10-fold increase in enzyme activity was also observed during this period. Using fluorescein 5'-isothiocyanate to quantitate the two alpha subunits, a similar increase in alpha + was observed with less of an increase in alpha. The ratio of alpha + to alpha increased from 0.75 at 18 days gestation to 3 at 3 days of age remaining at this ratio to 20 days of age. The possibility that thyroid hormone, a known regulator of brain Na+-K+-ATPase during development, differentially regulated the two forms was tested using 15-day-old hypothyroid rats. The abundance of both forms of alpha was similarly decreased: alpha + to 69% and alpha to 48% of control values. Na+-K+-ATPase activity was 70% of control. We conclude that both alpha and alpha + abundance increase in the brain during pre-and neonatal development and that the increase in both alpha subunits is regulated, directly or indirectly, by thyroid hormones.

Animals↗