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David G Taylor

Publications and source records attributed to David G Taylor.

13 recordsLinked to original sources
PubMed2006

Catecholamines and development of cardiac pacemaking: an intrinsically intimate relationship.

A generation ago, a melding of imagination and experimental evidence led to the hypothesis that catecholamines were essential in establishing basal cardiac pacemaking rhythm. Subsequent discoveries of depolarizing "pacemaker" currents and viable adult catecholamine-deficient animals raised serious doubts about the necessity of catecholamines in pacemaking. However, the findings that catecholamines are produced in pacemaking regions prior to innervation, and that they are required for embryonic survival during a defined "critical period" of embryonic development have revitalized the original hypothesis. Recent results have further suggested that intrinsic cardiac adrenergic cells can differentiate into pacemaking myocytes, and that protein kinase A, a prominent downstream mediator of beta-adrenergic signaling, is required for pacemaking activity. Here, we discuss how catecholamines and the intrinsic cardiac adrenergic cells that produce them may influence ontological development of cardiac pacemaking.

Animals↗
PubMed2006

Role of chromatin accessibility in the occupancy and transcription of the insulin gene by the pancreatic and duodenal homeobox factor 1.

The pancreatic and duodenal homeobox factor 1 (Pdx-1) is a Hox-like transcription factor that is responsible for the activation of the insulin gene. Previous studies have demonstrated the interaction in vitro of Pdx-1 with short (20-40 nucleotide) DNA fragments corresponding to A boxes of the insulin promoter. Precisely how Pdx-1 binds to DNA in the complex milieu of chromatin, however, has never been studied. In this study, we explored how Pdx-1-DNA interactions might be influenced by chromatin accessibility at the insulin gene in beta-cells (betaTC3) vs. pancreatic ductal cells (mPAC). We demonstrate that Pdx-1 occupies the endogenous insulin promoter in betaTC3 cells but not in mPAC cells, a finding that is independent of the intracellular Pdx-1 protein concentration. Based on micrococcal nuclease protection assays, the difference in promoter binding between the two cell types appears to be secondary to chromatin accessibility at predicted Pdx-1 binding sites between bp -126 to -296 (relative to the transcriptional start site) of the insulin promoter. Binding studies using purified Pdx-1 and reconstituted chromatin in vitro suggest that the positioning of a nucleosome(s) within this crucial region of the promoter might account for differences in chromatin accessibility. Consistent with these observations, fluorescence colocalization studies show that Pdx-1 does not occupy regions of compacted, nucleosome-rich chromatin within the nucleus. Our findings suggest a model whereby insulin transcription in the beta-cell is at least partially facilitated by enhanced chromatin accessibility within a crucial regulatory region between bp -126 to -296, thereby permitting occupancy by transactivators such as Pdx-1.

Acetylation↗
PubMed2006

Surface-directed boundary flow in microfluidic channels.

Channel geometry combined with surface chemistry enables a stable liquid boundary flow to be attained along the surfaces of a 12 microm diameter hydrophilic glass fiber in a closed semi-elliptical channel. Surface free energies and triangular corners formed by PDMS/glass fiber or OTS/glass fiber surfaces are shown to be responsible for the experimentally observed wetting phenomena and formation of liquid boundary layers that are 20-50 microm wide and 12 microm high. Viewing this stream through a 20 microm slit results in a virtual optical window with a 5 pL liquid volume suitable for cell counting and pathogen detection. The geometry that leads to the boundary layer is a closed channel that forms triangular corners where glass fiber and the OTS coated glass slide or PDMS touch. The contact angles and surfaces direct positioning of the fluid next to the fiber. Preferential wetting of corner regions initiates the boundary flow, while the elliptical cross-section of the channel stabilizes the microfluidic flow. The Young-Laplace equation, solved using fluid dynamic simulation software, shows contact angles that exceed 105 degrees will direct the aqueous fluid to a boundary layer next to a hydrophilic fiber with a contact angle of 5 degrees. We believe this is the first time that an explanation has been offered for the case of a boundary layer formation in a closed channel directed by a triangular geometry with two hydrophobic wetting edges adjacent to a hydrophilic surface.

Dimethylpolysiloxanes↗
PubMed2005

The C-terminal domain of the beta cell homeodomain factor Nkx6.1 enhances sequence-selective DNA binding at the insulin promoter.

The Hox-like factor Nkx6.1 is required for the formation and secretory function of insulin-producing beta cells, and has the capacity to activate or repress the transcription of relevant target genes in a DNA-context dependent manner. A key determinant of transcriptional activity by Nkx6.1 may be its C terminus, which has been suggested to interfere with DNA binding. To determine how the C terminus modulates homeodomain binding, we assessed the nature of Nkx6.1-DNA interactions at the insulin promoter. By quantitative gel shift analysis, we demonstrate that although the C terminus of Nkx6.1 mitigates the affinity of the homeodomain for DNA slightly (about 2-fold), it enhances the selectivity of the homeodomain for TAAT DNA sequences nearly 10-fold. By reporter gene analysis, this selectivity is also functionally preserved in mammalian cells in vivo. Based on deletional and mutational studies, the sequence selectivity imparted by the C terminus appears to be mediated by a stretch of highly conserved residues between amino acids 318 and 338. Strikingly, these residues impart minimal changes to the secondary structure of the unbound protein as assessed by circular dichroism spectroscopy, suggesting that conformational adjustments of the homeodomain that occur upon binding to DNA may play a more important role in sequence selectivity. The C terminus of Nkx6.1 also functions in a modular fashion, as it can confer similar DNA binding properties when fused to the heterologous homeodomain of Pdx-1. Taken together, our data suggest a model whereby the Nkx6.1 C terminus may function in a regulatory manner by imposing specific functional constraints upon the protein. These constraints may serve to modulate the potential of Nkx6.1 to both recognize target genes and regulate their transcription.

Animals↗
PubMed2005

Microfiber-directed boundary flow in press-fit microdevices fabricated from self-adhesive hydrophobic surfaces.

We report a rapid microfluidic device construction technique which does not employ lithography or stamping methods. Device assembly physically combines a silicon wafer, an elastomer (poly(dimethylsiloxane) (PDMS)), and microfibers to form patterns of hydrophobic channels, wells, elbows, or orifices that direct fluid flow into controlled boundary layers. Tweezers are used to place glass microfibers in a defined pattern onto an elastomeric (PDMS) hydrophobic film. The film is then manually pressed onto a hydrophobic silicon wafer, causing it to adhere to the silicon wafer and form a liquid-tight seal around the fibers. Completed in 15 min, the technique results in an operable microdevice with micrometer-scale features of nanoliter volume. Microfiber-directed boundary flow is achieved by use of the surface wetting properties of the hydrophilic glass fiber and the hydrophobicity of surrounding surfaces. The simplicity of this technique allows quick prototyping of microfluidic components, as well as complete biosensor systems, such as we describe for the detection of pathogenic bacteria.

Air↗
PubMed2005

The Nkx6.1 homeodomain transcription factor suppresses glucagon expression and regulates glucose-stimulated insulin secretion in islet beta cells.

We have previously described rat insulinoma INS-1-derived cell lines with robust or poor glucose-stimulated insulin secretion (GSIS). In the current study, we have further resolved these lines into three classes: class 1, glucose-unresponsive/glucagon-expressing; class 2, glucose-unresponsive/glucagon-negative; and class 3, glucose-responsive/glucagon-negative. The transcription factor Nkx2.2 was expressed with relative abundance of 3.3, 1.0, and 1.0 in class 1, class 2, and class 3 cells, respectively, whereas Nkx6.1 expression had the opposite trend: 1.0, 2.6, and 6.4 in class 1, class 2, and class 3 cells, respectively. In class 1 cells, overexpressed Nkx6.1 suppressed glucagon expression but did not affect the levels of several other prominent beta cell transcription factors. RNA interference (RNAi)-mediated suppression of Nkx6.1 in class 3 cells resulted in a doubling of glucagon mRNA, with no effect on Pdx1 levels, whereas suppression of Pdx1 in class 3 cells caused a 12-fold increase in glucagon transcript levels, demonstrating independent effects of Nkx6.1 and Pdx1 on glucagon expression in beta cell lines. RNAi-mediated suppression of Nkx6.1 expression in class 3 cells also caused a decrease in GSIS from 13.9- to 3.7-fold, whereas suppression of Pdx1 reduced absolute amounts of insulin secretion without affecting fold response. Finally, RNAi-mediated suppression of Nkx6.1 mRNA in primary rat islets was accompanied by a significant decrease in GSIS relative to control cells. In sum, our studies have revealed roles for Nkx6.1 in suppression of glucagon expression and control of GSIS in islet beta cells.

Adenoviridae↗
PubMed2005

Caudal anesthesia in pediatric cardiac surgery: does it affect outcome?

OBJECTIVE: The purpose of this study was to examine the influence of caudal anesthesia on outcomes (pediatric intensive care unit [PICU] length of stay, hospital length of stay, ventilatory time, early extubation rate) in pediatric patients undergoing congenital heart disease repair requiring cardiopulmonary bypass (CPB). DESIGN: Retrospective. SETTING: University teaching hospital. PARTICIPANTS: Pediatric patients undergoing surgery to treat congenital heart disease between 1999 and 2002. INTERVENTION: None. MEASUREMENTS AND MAIN RESULTS: Thirty-four patients with atrial septal defect (ASD), 37 with ventricular septal defect, and 46 with tetralogy of Fallot (TOF) were included in the analysis. No differences were found in preoperative and intraoperative data between caudal and noncaudal group for each disorder. There was no difference between caudal and noncaudal groups in PICU and hospital stay. A statistically significant difference was found in the postoperative ventilatory time in patients with ASD and TOF between caudal and noncaudal groups. The early extubation rate was higher in the TOF caudal group compared with the noncaudal group. CONCLUSIONS: This retrospective study demonstrated that postinduction placement of caudal anesthesia does not affect PICU or hospital length of stay. A well-controlled prospective study is needed to confirm these findings.

Analgesics, Opioid↗
PubMed2005

Utility of ultrasound-guided central venous cannulation in pediatric surgical patients: a clinical series.

BACKGROUND: Central venous cannulation can be particularly difficult in pediatric patients. Central line placement is associated with many well-known complications. While ultrasound-guided techniques are well established, the majority of central venous catheters are placed using landmark guidance. This retrospective study compares the safety and efficacy of ultrasound guidance vs landmark guidance in central venous cannulation of pediatric cardiac surgery patients. METHODS: The medical records of 149 pediatric patients undergoing cardiac surgery over 3-year period were reviewed. Patients were classified into two cohorts based on whether central venous cannulation of the internal jugular vein was performed by ultrasound or landmark guidance. Overall success and traumatic complication rates were compared between the two groups. Additionally, comparisons between the groups were made to determine if patient size or age affected the success rate of either approach in different manner. RESULTS: Patients in the ultrasound-guided (n = 47) and the landmark-guided (n = 102) groups were similar with respect to age, weight, and surgical procedure for which central venous access was indicated. The overall success rate for cannulation of the internal jugular vein was 91.5% in the ultrasound-guided group and 72.5% in the landmark-guided group (P = 0.010). But in the subgroup of children under 1 year of age, success rate was 77.8% in ultrasound group and 60.9% in landmark group (P = 0.44); in children under 10 kg in weight, success rate was 80% in ultrasound group and 56.7% in landmark group (P = 0.19). There were no significant differences in the rate of traumatic complications between the two methods. CONCLUSIONS: The overall success of internal jugular vein cannulation for pediatric cardiac surgery is significantly improved with the use of ultrasound guidance, without a significant difference in traumatic complications. However, mostly children above 1 year of age or 10 kg of weight experience advantages of ultrasound technique.

Catheterization, Central Venous↗
PubMed2004

The transcriptional repressor Nkx6.1 also functions as a deoxyribonucleic acid context-dependent transcriptional activator during pancreatic beta-cell differentiation: evidence for feedback activation of the nkx6.1 gene by Nkx6.1.

In the pancreas, the NK homeodomain transcription factor Nkx6.1 is required for the development of beta-cells and is believed to function as a potent repressor of transcription upon binding to A/T-rich sequences within the promoter region of target genes. Because the nkx6.1 promoter itself contains several such sequences, we considered the possibility that the expression level and restricted pattern of the nkx6.1 gene might be precisely regulated by one or more homeodomain transcription factors, including Nkx6.1 itself. In this report, we identify a novel beta-cell-specific enhancer element in the nkx6.1 gene between -157 and -30 bp (relative to the transcriptional start site) that harbors a conserved A/T-containing sequence flanked by G/C-rich stretches. Although the islet homeodomain-containing activator Pdx-1 was unable to stimulate transcription of a reporter gene through this enhancer element in mammalian cell lines, strikingly, Nkx6.1 robustly activated transcription through direct interaction with the A/T-rich sequence in this element. We demonstrate that this activation is indeed transcriptional in nature (and not secondary to translational effects) and is mediated by a modular acidic sequence within the COOH-terminal domain of Nkx6.1. We show by EMSAs that Nkx6.1 binds to the beta-cell-specific enhancer in vitro and by chromatin immunoprecipitation assays that Nkx6.1 natively occupies this region in vivo in betaTC3 cells. We therefore conclude that Nkx6.1 is a bifunctional transcription factor that serves to maintain the specific expression of its own gene during beta-cell differentiation while simultaneously effecting broader gene repression events.

Animals↗
PubMed2004

Quantification of the rat left ventricle force and Ca2+ -frequency relationships: similarities to dog and human.

OBJECTIVE: To measure and quantify the force-frequency (FFR) and Ca(2+)-frequency (CaFR) relationships in isolated rat left ventricular (LV) muscle at physiological heart rates and compare the obtained FFR to that measured in larger mammalian muscle from dog and human using the same experimental protocol. METHODS: Rat papillary muscle was isolated from the LV of adult male Sprague-Dawley rats, and dog and human muscles were from free-wall LV biopsies, loaded with the Ca(2+) indicator Fura-2, allowed to recover from isolation trauma and then subjected to direct electrical stimulation while measuring force production and intracellular Ca(2+) transients. RESULTS: We obtained a positive FFR between 1 and 4 Hz that is qualitatively similar to that found in isolated LV epicardial muscle strips from dogs and humans with normal LV function. The FFR reflects the cytosolic Ca(2+) transients in amplitude. Isoproterenol yielded an enhancement in force, but flattening of the FFR, whereas cyclopiazonic acid caused depression of FFR amplitude without changing frequency-dependent shape. CONCLUSION: We describe an experimental protocol that consistently yields positive FFRs in rat, dog and human LV muscle at stimulation rates between 1 and 4 Hz, without significant qualitative differences. We attribute previously observed negative FFR in rat muscle to an increase in SERCA activity early after excision and preparation of the muscle strips.

Aged↗
PubMed2004

Prevention of cardiac hypertrophy by angiotensin II type-2 receptor gene transfer.

The role of the angiotensin II type-2 receptor (AT2R) in cardiac hypertrophy remains elusive despite its demonstrated involvement in cardiovascular development. We have previously shown that a lentiviral vector gene delivery system is able to transduce cardiac tissue with high efficiency in vivo. Using such an approach, our objectives in the present study were 2-fold: (1) to overexpress the AT2R in cardiac tissue after completion of natural embryonic development of the heart and (2) to determine the effects of this overexpression on cardiac hypertrophy and basal blood pressure (BP). A lentiviral vector encoding the AT2R (lenti-AT2R) was administered (1.5x10(8) transducing units) into the left ventricular space of 5-day-old spontaneously hypertensive rats (SHRs). AT2R transgene expression increased in these animals and persisted for 30 weeks. In contrast, the expression of the angiotensin II type-1 receptor remained unchanged following lenti-AT2R treatment. At 21 weeks following gene transduction, the lenti-AT2R-treated SHRs exhibited decreased left ventricular wall thickness compared with control animals. In contrast, basal BP did not differ between the two SHR groups. Finally, heart weight to body weight ratios indicated a significant decrease in lenti-AT2R-treated SHRs compared with SHR controls. Our data indicate that AT2R overexpression attenuates cardiac hypertrophy in the SHR. This beneficial outcome was observed despite the existence of elevated BP.

Animals↗
PubMed2002

Biosafety considerations for autopsy.

An autopsy may subject prosectors and others to a wide variety of infectious agents, including bloodborne and aerosolized pathogens such as human immunodeficiency virus, hepatitis B and C viruses, and Mycobacterium tuberculosis. Other hazards include toxic chemicals (e.g., formalin, cyanide, and organophosphates) and radiation from radionuclides used for patient therapy and diagnosis. These risks can be substantially mitigated through proper assessment, personal protective equipment, appropriate autopsy procedures, and facility design.

Autopsy↗