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Biomedical subjects

David A Brown

Publications and source records attributed to David A Brown.

At least 19 recordsLinked to original sources
PubMed2003

Exercise training preserves coronary flow and reduces infarct size after ischemia-reperfusion in rat heart.

The effect of endurance training on the resistance of the heart to left ventricular (LV) functional deficit and infarction after a transient regional ischemia and subsequent reperfusion was examined. Female Sprague-Dawley rats were randomly assigned to an endurance exercise training (Tr) group or a sedentary (Sed) control group. After 20 wk of training, hearts were excised, perfused, and instrumented for assessment of LV mechanical function, and the left anterior descending coronary artery was occluded to induce a transient regional ischemia (1 h) that was followed by 2 h of reperfusion. Throughout much of the regional ischemia-reperfusion protocol, coronary flow rates, diastolic function, and LV developed pressure were better preserved in hearts from Tr animals. During the regional ischemia, coronary flow to myocardium outside the ischemic zone at risk (ZAR) was maintained in Tr hearts, whereas it progressively fell in Sed hearts. On release of the coronary artery ligature, flow to the ZAR was greater in Tr than in Sed hearts. Infarct size, expressed as a percentage of the ischemic ZAR, was significantly smaller in hearts from Tr rats (24 +/- 3 vs. 32 +/- 2% of ZAR, P < 0.05). Mn- and CuZn-SOD protein expression were higher in the LV myocardium of Tr animals (P < 0.05 for both isoforms). Our data indicate that long-term exercise training leads to infarct sparing and better maintenance of coronary flow and mechanical function after ischemia-reperfusion.

Animals↗
PubMed2003

Macrophage inhibitory cytokine 1 reduces cell adhesion and induces apoptosis in prostate cancer cells.

Macrophage inhibitory cytokine 1 (MIC-1), a divergent member of the transforming growth factor-beta superfamily, is linked to the pathogenesis of cancer. To delineate possible roles for MIC-1 in prostate cancer, a number of prostate epithelial cell lines have been studied, including PZ-HPV-7, DU-145, PC-3, and LNCaP cells. Factors regulating the production of MIC-1 protein by these cells and some of the effects of MIC-1 on them were investigated. Although PZ-HPV-7 and DU-145 produced no MIC-1 protein, PC-3 and LNCaP cells secreted MIC-1 protein at high levels. The secretion of MIC-1 in LNCaP cells was modulated by both androgen and estrogen. Although neither MIC-1 nor anti-MIC-1 antibody had any effect on the proliferation of epithelial cells, MIC-1 induced changes in DU-145 cells. These cells became flattened and more spread out, and this was accompanied by reduced intercellular actin filaments and intercellular junctions. The DU-145 cells then detached from their substrate and underwent caspase-dependent apoptosis. To define some of the genes responsible for these changes, cDNA microarrays, followed by confirmatory reverse transcription-PCR, was used to analyze differential gene expression induced by MIC-1. The antiapoptotic gene metallothionein 1E and cell adhesion genes RhoE and catenin delta 1 were down-regulated by more than 2-fold by MIC-1, suggesting that they were, at least in part, responsible for the observed changes in the behavior of DU-145 cells. These findings suggest that although MIC-1 has no effect on cell proliferation, it reduces cell adhesion and consequently induces cell detachment. It is likely that caspase-dependent apoptosis is secondary to loss of cell adhesion and may suggest a role for MIC-1 in tumor dissemination in vivo.

Apoptosis↗
PubMed2003

KCNQ/M currents in sensory neurons: significance for pain therapy.

Neuronal hyperexcitability is a feature of epilepsy and both inflammatory and neuropathic pain. M currents [IK(M)] play a key role in regulating neuronal excitability, and mutations in neuronal KCNQ2/3 subunits, the molecular correlates of IK(M), have previously been linked to benign familial neonatal epilepsy. Here, we demonstrate that KCNQ/M channels are also present in nociceptive sensory systems. IK(M) was identified, on the basis of biophysical and pharmacological properties, in cultured neurons isolated from dorsal root ganglia (DRGs) from 17-d-old rats. Currents were inhibited by the M-channel blockers linopirdine (IC50, 2.1 microm) and XE991 (IC50, 0.26 microm) and enhanced by retigabine (10 microm). The expression of neuronal KCNQ subunits in DRG neurons was confirmed using reverse transcription-PCR and single-cell PCR analysis and by immunofluorescence. Retigabine, applied to the dorsal spinal cord, inhibited C and Adelta fiber-mediated responses of dorsal horn neurons evoked by natural or electrical afferent stimulation and the progressive "windup" discharge with repetitive stimulation in normal rats and in rats subjected to spinal nerve ligation. Retigabine also inhibited responses to intrapaw application of carrageenan in a rat model of chronic pain; this was reversed by XE991. It is suggested that IK(M) plays a key role in controlling the excitability of nociceptors and may represent a novel analgesic target.

Animals↗
PubMed2003

Influence of age and run training on cardiac Na+/Ca2+ exchange.

Effects of age and training on myocardial Na+/Ca2+ exchange were examined in young sedentary (YS; 14-15 mo), aged sedentary (AS; 27-31 mo), and aged trained (AT; 8- to 11-wk treadmill run training) male Fischer Brown Norway rats. Whole heart performance and isolated cardiocyte Na+/Ca2+ exchange characteristics were measured. At the whole heart level, a small but significant slowing of late isovolumic left ventricular (LV) relaxation, which may be indicative of altered Na+/Ca2+ exchange activity, was seen in hearts from AS rats. This subtle impairment in relaxation was not observed in hearts from AT rats. At the single-cardiocyte level, late action potential duration was prolonged, resting membrane potential was more positive, and overshoot potential was greater in cardiocytes from AS rats than from YS rats (P < 0.05). Training did not influence any of these age-related action potential characteristics. In electrically paced cardiocytes, neither shortening nor intracellular Ca2+ concentration ([Ca2+]i) dynamics was influenced by age or training. Similarly, neither age nor training influenced the rate of [Ca2+]i clearance via forward (Nain+ /Caout2+) Na+/Ca2+ exchange after caffeine-induced Ca2+ release from the sarcoplasmic reticulum or cardiac Na+/Ca2+ exchanger protein (NCX1) expression. However, when whole cell patch-clamp techniques combined with fluorescence microscopy were used to evaluate the ability of Na+/Ca2+ exchange to alter cytosolic [Ca2+] ([Ca2+]c) under conditions where membrane potential (Vm) and internal and external [Na+] and [Ca2+] could be controlled, we observed age-associated increases in forward Na+/Ca2+ exchange-mediated [Ca2+]c clearance (P < 0.05) that were not influenced by training. The age-related increase in forward Na+/Ca2+ exchange activity provides a hypothetical explanation for the late action potential prolongation observed in this study.

Action Potentials↗
PubMed2003

Critical role for NHE1 in intracellular pH regulation in pancreatic acinar cells.

The primary function of pancreatic acinar cells is to secrete digestive enzymes together with a NaCl-rich primary fluid which is later greatly supplemented and modified by the pancreatic duct. A Na+/H+ exchanger(s) [NHE(s)] is proposed to be integral in the process of fluid secretion both in terms of the transcellular flux of Na+ and intracellular pH (pHi) regulation. Multiple NHE isoforms have been identified in pancreatic tissue, but little is known about their individual functions in acinar cells. The Na+/H+ exchange inhibitor 5-(N-ethyl-N-isopropyl) amiloride completely blocked pHi recovery after an NH4Cl-induced acid challenge, confirming a general role for NHE in pHi regulation. The targeted disruption of the Nhe1 gene also completely abolished pHi recovery from an acid load in pancreatic acini in both HCO3--containing and HCO3--free solutions. In contrast, the disruption of either Nhe2 or Nhe3 had no effect on pHi recovery. In addition, NHE1 activity was upregulated in response to muscarinic stimulation in wild-type mice but not in NHE1-deficient mice. Fluctuations in pHi could potentially have major effects on Ca2+ signaling following secretagogue stimulation; however, the targeted disruption of Nhe1 was found to have no significant effect on intracellular Ca2+ homeostasis. These data demonstrate that NHE1 is the major regulator of pHi in both resting and muscarinic agonist-stimulated pancreatic acinar cells.

Acids↗
PubMed2003

Stoichiometry of expressed KCNQ2/KCNQ3 potassium channels and subunit composition of native ganglionic M channels deduced from block by tetraethylammonium.

KCNQ2 and KCNQ3 potassium-channel subunits can form both homomeric and heteromeric channels; the latter are thought to constitute native ganglionic M channels. We have tried to deduce the stoichiometric contributions of KCNQ2 and KCNQ3 subunits to currents generated by the coexpression of KCNQ2 and KCNQ3 cDNA plasmids in Chinese hamster ovary (CHO) cells, and to native M currents in dissociated rat superior cervical ganglion (SCG) neurons, by comparing the block of these currents produced by tetraethylammonium (TEA) with the block of currents generated by a tandem KCNQ3/2 construct. TEA concentration-inhibition curves against coexpressed KCNQ2 plus KCNQ3 currents, and against native M currents in SCG neurons from 6-week-old [postnatal day 45 (P45)] rats, were indistinguishable from those for the expressed tandem construct, and fully accorded with a 1:1 stoichiometry. Inhibition curves in neurons from younger (P17) rats could be better fitted assuming an additional small proportion of current carried by KCNQ2 homomultimers. Single-cell PCR yielded signals for KCNQ2, KCNQ3, and KCNQ5 mRNAs in all SCG neurons tested from both P17 and P45 rats. Quantitative PCR of whole-ganglion mRNA revealed stable levels of KCNQ2 and KCNQ5 mRNA between P7 and P45, but excess and incrementing levels of KCNQ3 mRNA. Increasing levels of KCNQ3 protein between P17 and P45 were confirmed by immunocytochemistry. We conclude that coexpressed KCNQ2 plus KCNQ3 cDNAs generate channels with 1:1 (KCNQ2:KCNQ3) stoichiometry in CHO cells and that native M channels in SCG neurons adopt the same conformation during development, assisted by the increased expression of KCNQ3 mRNA and protein.

Animals↗
PubMed2003

Large-scale delineation of secreted protein biomarkers overexpressed in cancer tissue and serum.

Genetic alterations in tumor cells often lead to the emergence of growth-stimulatory autocrine and paracrine signals, involving overexpression of secreted peptide growth factors, cytokines, and hormones. Increased levels of these soluble proteins may be exploited for cancer diagnosis and management or as points of therapeutic intervention. Here, we combined the use of controlled vocabulary terms and sequence-based algorithms to predict genes encoding secreted proteins from among approximately 12,500 sequences represented on oligonucleotide microarrays. Expression of these genes was queried in 150 carcinomas from 10 anatomic sites of origin and compared with 46 normal tissues derived from the corresponding sites of tumor origin and other body tissues and organs. Of 74 different genes identified as overexpressed in cancer tissues, several encode proteins with demonstrated clinical diagnostic application, such as alpha-fetoprotein in liver carcinoma, and kallikreins 6 and 10 in ovarian cancer, or therapeutic utility, such as gastrin-releasing peptide/bombesin in lung carcinomas. We show that several of the other candidate genes encode proteins with high levels of tumor-associated expression by immunohistochemistry on tissue microarrays and further demonstrate significantly elevated levels of another novel candidate protein, macrophage inhibitory cytokine 1, a distant member of the transforming growth factor-beta superfamily, in the serum of patients with metastatic prostate, breast, and colorectal carcinomas. Our results suggest that the combination of annotation/protein sequence analysis, transcript profiling, immunohistochemistry, and immunoassay is a powerful approach for delineating candidate biomarkers with potential clinical significance and may be broadly applicable to other human diseases.

Biomarkers, Tumor↗
PubMed2003

Off-pump coronary artery bypass grafting does not decrease the incidence of atrial fibrillation.

BACKGROUND: Postoperative atrial fibrillation remains a significant source of morbidity after coronary artery bypass grafting. We reviewed the data on 2,569 patients to determine if the absence of cardiopulmonary bypass resulted in a lower incidence of atrial fibrillation. METHODS: All patients undergoing coronary artery bypass grafting without cardiopulmonary bypass from January 1, 1997 through June 30, 2001 were evaluated for postoperative atrial fibrillation. The data of 252 patients with no cardiopulmonary bypass (group 1) were reviewed and compared with three other patient groups. Group 2 consisted of 1,470 patients using cardiopulmonary bypass during the same study period. Group 3 consisted of 841 patients with a similar number of grafts as the study group but using cardiopulmonary bypass. Group 4 consisted of historical data for 847 patients operated on using cardiopulmonary bypass collected from January 1995 through December 1996. Prophylactic beta-blockade was instituted in January 1997. Groups 1 to 3 received this treatment, but group 4 did not. RESULTS: Group 1 had an incidence of atrial fibrillation of 8.8%. Groups 2, 3, and 4 had incidences of atrial fibrillation of 11.6%, 9.4%, and 28.0%, respectively. When compared with group 1, the incidence of atrial fibrillation in group 4 was statistically different (p <. 0001). CONCLUSIONS: Avoiding cardiopulmonary bypass did not aid the reduction of atrial fibrillation at our institution.

Aged↗
PubMed2003

Two cases of anomalous origin of LAD from right coronary artery requiring coronary artery bypass.

We experienced two cases with anomalous origin of the left anterior descending artery (LAD) from the proximal right coronary artery requiring coronary artery bypass grafting. A 66-yr old female with a long history of angina and a positive stress test had the anomalous artery coursing anterior to the right ventricular outflow tract. A 42-yr old male with worsening angina after an anteroseptal myocardial infarction had the anomalous artery running between the great vessels. Both patients underwent left internal mammary artery-to-LAD bypass on the beating heart with complete resolution of ischemic symptoms. Isolated coronary artery anomaly is an uncommon disease (0.6-1.2%) in patients undergoing cardiac catheterization. An anomalous origin of the LAD from the proximal right coronary artery (RCA) or the right sinus of Valsalva (RSV) is very rare, found in 1.2-6.1% of all coronary anomalies. This coronary anomaly has been considered potentially serious but functionally unimportant. We report two cases of anomalous LAD from the proximal RCA resulting in anterior wall ischemia which was effectively treated by coronary artery bypass surgery.

Adult↗
PubMed2003

AKAP150 signaling complex promotes suppression of the M-current by muscarinic agonists.

M-type (KCNQ2/3) potassium channels are suppressed by activation of G(q/11)-coupled receptors, thereby increasing neuronal excitability. We show here that rat KCNQ2 can bind directly to the multivalent A-kinase-anchoring protein AKAP150. Peptides that block AKAP150 binding to the KCNQ2 channel complex antagonize the muscarinic inhibition of the currents. A mutant form of AKAP150, AKAP(DeltaA), which is unable to bind protein kinase C (PKC), also attenuates the agonist-induced current suppression. Analysis of recombinant KCNQ2 channels suggests that targeting of PKC through association with AKAP150 is important for the inhibition. Phosphorylation of KCNQ2 channels was increased by muscarinic stimulation; this was prevented either by coexpression with AKAP(DeltaA) or pretreatment with PKC inhibitors that compete with diacylglycerol. These inhibitors also reduced muscarinic inhibition of M-current. Our data indicate that AKAP150-bound PKC participates in receptor-induced inhibition of the M-current.

A Kinase Anchor Proteins↗
PubMed2003

Coupling of the nucleotide P2Y4 receptor to neuronal ion channels.

1. G protein-linked P2Y nucleotide receptors are known commonly to stimulate the phosphoinositide signalling pathway. However, we have previously demonstrated that the cloned P2Y(2), P2Y(6) and P2Y(1) receptors couple to neuronal N-type Ca(2+) channels and to M-type K(+) channels. Here we investigate the coupling of recombinant, neuronally expressed rat- and human P2Y(4) receptors (rP2Y(4), hP2Y(4)) to those channels. 2. Rat sympathetic neurones were nuclear-injected with a P2Y(4) cDNA plasmid. A subsequent activation of rP2Y(4) or hP2Y(4) by UTP (100 micro M) in whole-cell (ruptured-patch) mode produced only about 12% inhibition of the N-type Ca(2+) current (I(Ca(N))). Surprisingly, in perforated patch mode, UTP produced much more inhibition of I(Ca(N)) (maximally 51%), with an IC(50) value of 273 nM. This inhibition was voltage-dependent and was blocked by co-expression of the betagamma-binding transducin Galpha-subunit. Pertussis toxin (PTX) pretreatment also suppressed I(Ca(N)) inhibition. 3. UTP inhibited the M-current, recorded in perforated patch mode, by (maximally) 52%, with IC(50) values of 21 nM for rP2Y(4) and 28 nM for hP2Y(4). This inhibition was not affected by PTX pretreatment. 4. With rP2Y(4), ATP inhibited the M-current (IC(50) 524 nM, 26 times weaker than UTP), whereas ATP had no agonist activity at hP2Y(4). This suggests a difference in agonist binding site between rP2Y(4) and hP2Y(4). 5. We conclude that, in contrast to other nucleotide receptors studied, the P2Y(4) receptor couples much more effectively to M-type K(+) channels than to Ca(2+) channels. Coupling to the Ca(2+) channels involves the betagamma-subunits of G(i/o)-proteins and requires a diffusible intracellular component that is lost in ruptured-patch recording.

Animals↗
PubMed2003

Macrophage inhibitory cytokine 1 in fetal membranes and amniotic fluid from pregnancies with and without preterm labour and premature rupture of membranes.

The placenta and fetal membranes are the site of expression of macrophage inhibitory cytokine (MIC-1), a member of the transforming growth factor (TGF)-beta superfamily. We hypothesized that MIC-1 may act as an immune regulator in pregnancy complications associated with intrauterine inflammation. Decidual cells, chorionic trophoblasts and amnion epithelial cells were identified by immunohistochemistry as the predominant MIC-1-containing cell type in term membranes. Amnion and choriodecidual explants all produced MIC-1 in culture, the latter having the greatest production rate (206 +/- 74.5 pg/mg tissue/24 h, n=6; mean +/- SEM). Production was not responsive to stimulation by pro-inflammatory cytokines. MIC-1 was detectable in 217 transabdominal amniotic fluid (AF) samples taken from 15 to 41 weeks gestation, concentrations ranging from 0.9-51.1 ng/ml. AF MIC-1 concentrations in pregnancies with premature rupture of membranes (PROM) or preterm labour, either with or without microbial invasion of the amniotic cavity, were not significantly different from those delivered at term either with or without labour. Treatment with MIC-1 (0.25-25 ng/ml) did not alter production of interleukin-6 or -8 by amnion or choriodecidual cells in vitro. We conclude that AF MIC-1 is derived from the fetal membranes and decidua, but that MIC-1 is unlikely to be involved in the pathophysiology of preterm birth or PROM.

Amniotic Fluid↗
PubMed2003

Mortality among hourly motor vehicle manufacturing workers.

We evaluated mortality among 198,245 motor vehicle industry workers during the period of 1973 to 1995. Workers' mortality rates were lower than expected overall (40,131 observed/43,859 expected deaths, standardized mortality ratio [SMR] = 92, CI = 91-92) and for all major cause of death categories except cancer (SMR = 100, CI = 98-102). Mortality rates were higher than expected for lung cancer overall (SMR = 110, CI = 107-113) and among employees in transmission/gear manufacturing (SMR = 121, CI = 112-130), casting operations (SMR = 122, CI = 110-135), engine manufacturing (SMR = 111, CI = 101-123), and vehicle assembly (SMR = 111, CI = 105-117); for stomach cancer in engine manufacturing (SMR = 147, CI = 110-192); and for prostate cancer in casting operations (SMR = 128, CI = 102-158). Excesses of lung cancer in transmission, vehicle assembly, and casting operations and of stomach cancer in engine manufacturing have been observed in other investigations. Further information on employees' occupational exposures and personal attributes is required to clarify the interpretation of these results.

Accidents, Occupational↗
PubMed2003

MIC-1 serum level and genotype: associations with progress and prognosis of colorectal carcinoma.

PURPOSE: Macrophage inhibitory cytokine-1 (MIC-1) is a divergent member of the tumor growth factor beta (TGF-beta) superfamily. Several observations suggest that it plays a role in colorectal carcinoma (CRC). In particular, MIC-1 is markedly up-regulated in colorectal cancers as well as in premalignant adenomas. This study examines the relationship of serum MIC-1 levels and genotypes to clinical and pathologic features of colonic neoplasia. EXPERIMENTAL DESIGN: We confirmed the presence of MIC-1 in CRC tissue and the cell line CaCo-2. The normal range for serum MIC-1 levels was defined in 260 healthy blood donors, and the differences between normal subjects and 193 patients having adenomatous polyps or CRC were then determined. In a separate cohort of 224 patients, we evaluated the relationship of MIC-1 serum level and genotype to standard tumor parameters and outcome measures. RESULTS: MIC-1 was expressed in CRC tissue and the cancer cell line CaCo-2. There was a progressive increase in serum MIC-1 levels between normal individuals [mean (M) = 495 pg/ml, SD = 210), those with adenomatous polyps (M = 681 pg/ml, SD = 410), and those with CRC (M = 783 pg/ml, SD = 491)]. Serum MIC-1 level was correlated with the extent of disease so that the levels were higher in patients with higher Tumor-Node-Metastasis stage. There were significant differences in time to relapse and overall survival between subjects with different MIC-1 levels and genotypes. CONCLUSIONS: This study identifies a strong association between MIC-1 serum levels and neoplastic progression within the large bowel. We suggest that the measurement of serum MIC-1 levels and determination of MIC-1 genotype may have clinical use in the management of patients with CRC.

Adenoma↗
PubMed2003

A bicyclic monoterpene diol and UVB stimulate BRCA1 phosphorylation in human keratinocytes.

BRCA1 (breast cancer-associated gene 1) is a tumor suppressor gene that plays a role in DNA repair when phosphorylated. Many DNA-damaging agents including UVC and hydrogen peroxide have been shown to induce phosphorylation of BRCA1. Results of this study now show that both UVB and a bicyclic monoterpene diol (BMT diol) result in phosphorylation of BRCA1. This phosphorylation was maximal 2 h after treatment with either agent and declined to basal levels by 24 h. Inhibitor studies revealed that both UVB and the BMT diol phosphorylate BRCA1 through the FK506-binding protein-FKBP rapamycin-associated binding protein pathway, but the BMT diol also led to phosphorylation of BRCA1 through casein kinase II. This suggests that the signaling pathways for UVB and the BMT diol may diverge. Results of this study also show that the BMT diol stimulates the repair of UVB-induced cyclobutane pyrimidine dimers (CPD). Inhibitors of BMT diol-induced BRCA1 phosphorylation blocked the BMT diol-stimulated repair of CPD. This indicates that the BMT diol induces the phosphorylation of BRCA1, which, in turn, leads to an increase in repair of UVB-induced CPD. Therefore, this BMT diol may be useful for ameliorating the damaging effects of UVB.

BRCA1 Protein↗
PubMed2002

Junctional signaling microdomains: bridging the gap between the neuronal cell surface and Ca2+ stores.

Growing evidence suggests that plasma membranes are locally differentiated into microdomains that are important interaction sites for organization of signaling molecules. These signaling microdomains create local conditions that enhance molecular interactions, excluding others, thereby ensuring speed, spatial localization, and specificity of signal transduction. With the special emphasis on InsP(3) and Ca(2+) signaling pathways, we will discuss here the evolving concept of signaling microdomains that provide a key framework for understanding the differential regulation of many cellular target proteins.

Calcium↗
PubMed2002

Concentration in plasma of macrophage inhibitory cytokine-1 and risk of cardiovascular events in women: a nested case-control study.

BACKGROUND: Macrophage inhibitory cytokine-1 (MIC-1) is part of the TGF-beta superfamily. Raised concentrations of MIC-1 in serum arise in several disease states, can be detected in normal individuals, and can partly be genetically determined. We aimed to investigate whether MIC-1 has a role in atherothrombosis. METHODS: We did a prospective, nested, case-control study in 27628 initially healthy women. Of these women, we established baseline concentrations of MIC-1 in 257 who subsequently had myocardial infarction, stroke, or died from a cardiovascular event (cases) and in 257 matched for age and smoking status, who did not report cardiovascular disease during 4-year follow-up (controls). We also assessed polymorphisms in the MIC-1 gene (MIC-1 H and MIC-1 D) in all 514 women. FINDINGS: MIC-1 concentrations were higher at baseline in women who subsequently had cardiovascular events than in those who did not (618 vs 538 pg/mL, p=0.0002). Concentrations above the 90th percentile (>856 pg/mL) were associated with a 2.7-fold increase in risk (95% CI 1.6-4.9, p=0.001). This effect was independent of traditional cardiovascular risk factors and at least additive to that of C-reactive protein. There was no significant association between MIC-1 polymorphism and vascular events. INTERPRETATION: MIC-1 could be a novel target for cardiovascular disease prevention.

Cardiovascular Diseases↗