Biomedical subjects

A Chow

Publications and source records attributed to A Chow.

At least 37 records · Page 2Linked to original sources

PubMed1999

Molecular diversity of K+ channels.

K+ channel principal subunits are by far the largest and most diverse of the ion channels. This diversity originates partly from the large number of genes coding for K+ channel principal subunits, but also from other processes such as alternative splicing, generating multiple mRNA transcripts from a single gene, heteromeric assembly of different principal subunits, as well as possible RNA editing and posttranslational modifications. In this chapter, we attempt to give an overview (mostly in tabular format) of the different genes coding for K+ channel principal and accessory subunits and their genealogical relationships. We discuss the possible correlation of different principal subunits with native K+ channels, the biophysical and pharmacological properties of channels formed when principal subunits are expressed in heterologous expression systems, and their patterns of tissue expression. In addition, we devote a section to describing how diversity of K+ channels can be conferred by heteromultimer formation, accessory subunits, alternative splicing, RNA editing and posttranslational modifications. We trust that this collection of facts will be of use to those attempting to compare the properties of new subunits to the properties of others already known or to those interested in a comparison between native channels and cloned candidates.

Alternative Splicing↗

PubMed1999

Contributions of Kv3 channels to neuronal excitability.

Four mammalian Kv3 genes have been identified, each of which generates, by alternative splicing, multiple protein products differing in their C-terminal sequence. Products of the Kv3.1 and Kv3.2 genes express similar delayed-rectifier type currents in heterologous expression systems, while Kv3.3 and Kv3.4 proteins express A-type currents. All Kv3 currents activate relatively fast at voltages more positive than -10 mV, and deactivate very fast. The distribution of Kv3 mRNAs in the rodent CNS was studied by in situ hybridization, and the localization of Kv3.1 and Kv3.2 proteins has been studied by immunohistochemistry. Most Kv3.2 mRNAs (approximately 90%) are present in thalamic-relay neurons throughout the dorsal thalamus. The protein is expressed mainly in the axons and terminals of these neurons. Kv3.2 channels are thought to be important for thalamocortical signal transmission. Kv3.1 and Kv3.2 proteins are coexpressed in some neuronal populations such as in fast-spiking interneurons of the cortex and hippocampus, and neurons in the globus pallidus. Coprecipitation studies suggest that in these cells the two types of protein form heteromeric channels. Kv3 proteins appear to mediate, in native neurons, similar currents to those seen in heterologous expression systems. The activation voltage and fast deactivation rates are believed to allow these channels to help repolarize action potentials fast without affecting the threshold for action potential generation. The fast deactivating current generates a quickly recovering after hyperpolarization, thus maximizing the rate of recovery of Na+ channel inactivation without contributing to an increase in the duration of the refractory period. These properties are believed to contribute to the ability of neurons to fire at high frequencies and to help regulate the fidelity of synaptic transmission. Experimental evidence has now become available showing that Kv3.1-Kv3.2 channels play critical roles in the generation of fast-spiking properties in cortical GABAergic interneurons.

Alternative Splicing↗

PubMed1999

Investigating the role played by protein-lipid and protein-protein interactions in the membrane association of the p120GAP CaLB domain.

The GTPase activating protein, p120GAP, contains an amino acid sequence motif called the Ca2+-dependent lipid binding domain (CaLB) which mediates a protein-protein interaction between p120GAP and annexin VI and also binds to negatively charged phospholipids. Because membrane association of p120GAP is important for the regulation of p21 Ras activity, we have studied the roles played by Ca2+, phospholipids and annexin VI in the membrane association of p120GAP. Here we demonstrate that a truncated CaLB domain GST fusion protein (GSTGAP618-632), lacking the ability to bind to phospholipids, is able to bind to rat fibroblast membranes in a Ca2+- and concentration-dependent manner. In addition, this fusion protein also binds to annexin VI in an amino acid sequence specific but Ca2+ independent manner. Also, when bound to annexin VI in the presence of Ca2+, this fusion protein has the ability to co-bind to phosphatidylserine vesicles. Thus, annexin VI may simultaneously mediate an interaction with p120GAP and also an interaction with membrane phospholipids. This may in part explain the mechanism by which p120GAP associates with membranes in response to Ca2+ elevation and suggests the potential importance of annexin VI in the regulation of p21 Ras and the role CaLB domains may play in the specific recognition of cellular membranes.

Amino Acid Sequence↗

PubMed1999

The effect of site-specific monoclonal antibodies directed to toxic shock syndrome toxin-1 in experimental Staphylococcus aureus arthritis.

Staphylococcus aureus produces a large number of potential virulence factors, among these the superantigen toxic shock syndrome toxin-1 (TSST-1). We have recently demonstrated that TSST-1 is involved in the pathogenesis of septic arthritis. Recent data show that the TSST-1 molecule is composed of two distinct domains, one proposed to interact with T cell receptor (TCR) and one with the MHC class II. The aim of this study was to assess if interaction between TSST-1-specific MoAbs directed to sites on the MHC and/or TCR Vbeta affects the development of experimental S. aureus-induced arthritis. For that purpose we used a panel of seven MoAbs, which were injected intraperitoneally before and after inoculation with a TSST-1-producing S. aureus strain. Administration of antibodies did not affect the development of arthritis, suggesting inefficacy of such a procedure in neutralization of exotoxin-mediated disease manifestations.

Animals↗

PubMed1999

Kv3.1-Kv3.2 channels underlie a high-voltage-activating component of the delayed rectifier K+ current in projecting neurons from the globus pallidus.

The globus pallidus plays central roles in the basal ganglia circuitry involved in movement control as well as in cognitive and emotional functions. There is therefore great interest in the anatomic and electrophysiological characterization of this nucleus. Most pallidal neurons are GABAergic projecting cells, a large fraction of which express the calcium binding protein parvalbumin (PV). Here we show that PV-containing pallidal neurons coexpress Kv3. 1 and Kv3.2 K+ channel proteins and that both Kv3.1 and Kv3.2 antibodies coprecipitate both channel proteins from pallidal membrane extracts solubilized with nondenaturing detergents, suggesting that the two channel subunits are forming heteromeric channels. Kv3.1 and Kv3.2 channels have several unusual electrophysiological properties when expressed in heterologous expression systems and are thought to play special roles in neuronal excitability including facilitating sustained high-frequency firing in fast-spiking neurons such as interneurons in the cortex and the hippocampus. Electrophysiological analysis of freshly dissociated pallidal neurons demonstrates that these cells have a current that is nearly identical to the currents expressed by Kv3.1 and Kv3.2 proteins in heterologous expression systems, including activation at very depolarized membrane potentials (more positive than -10 mV) and very fast deactivation rates. These results suggest that the electrophysiological properties of native channels containing Kv3.1 and Kv3.2 proteins in pallidal neurons are not significantly affected by factors such as associated subunits or postranslational modifications that result in channels having different properties in heterologous expression systems and native neurons. Most neurons in the globus pallidus have been reported to fire sustained trains of action potentials at high-frequency. Kv3.1-Kv3.2 voltage-gated K+ channels may play a role in helping maintain sustained high-frequency repetitive firing as they probably do in other neurons.

Animals↗

PubMed1998

Characterization of hemoglobin variants by MALDI-TOF MS using a polyurethane membrane as the sample support.

A new method for the sampling and off-site analysis of hemoglobin variants by mass spectrometry is reported. This technique uses a nonporous polyurethane membrane as the collection device and transportation medium of a blood sample for analysis. The same membrane is then used as the MALDI-TOF MS sample support for mass spectrometric analysis. Minimal invasive sample collection is afforded by collecting less than 1 microL of blood using a common lancet device. MALDI-TOF MS is performed directly on the membrane, after washing off the interfering plasma components, followed by the addition of matrix. This reduces the time of analysis and prevents sample loss. Enzymatic digestion can be performed directly on the membrane, using in this case trypsin, allowing for further characterization of the sample. The method is much less invasive compared to drawing blood with a syringe. The sample may be transported to the laboratory by regular mail, and thus the method can serve remote locations. We demonstrate the procedure by characterizing the Hb Shepherds Bush hemoglobin variant, b74-(E18)Gly-->Asp.

Genetic Variation↗

PubMed1998

Metal-catalyzed oxidation of immunoglobulin G impairs Fc receptor-mediated binding to macrophages.

Enhanced oxidative stress is a feature of inflammatory and infectious conditions. Proteins may be important targets of oxidation and this may alter their function. We evaluated whether metal-catalyzed oxidation of IgG could alter its ability to bind to Fc receptors on macrophages. Human IgG incubated with an FeCl3/EDTA/ascorbate metal-catalyzed oxidation system resulted in a significant increase in carbonyl content, a measure of protein oxidation, compared to IgG treated with EDTA alone (control). Western blot analysis using an antibody to oxidized protein revealed an increase in antibody binding to both the heavy (Fc portion-containing) and light chains of IgG treated with the oxidizing system. Western blot analysis of papain-digested IgG confirmed oxidative modification of the Fc portion. Binding studies carried out with J774.16 macrophages demonstrated significantly diminished ability of the oxidized IgG to bind to macrophage Fc receptors compared to control IgG. These data demonstrate that IgG is susceptible to metal-catalyzed oxidation and that this impairs its ability to bind to macrophage Fc receptors. Oxidation of IgG might play a role in modulating immune function in infection and disorders associated with immune complex formation by diminishing IgG binding to phagocytic cells.

Ascorbic Acid↗

PubMed1998

Metal-catalyzed oxidation of extracellular matrix increases macrophage nitric oxide generation.

BACKGROUND: Oxygen radicals are believed to play a significant role in glomerular disease. In part this may be due to oxidation of lipids, but protein oxidation may play a contributory role as well. We have demonstrated that the mesangial extracellular matrix is susceptible to metal-catalyzed oxidation and that this increases scavenger receptor-mediated adhesion of macrophages, cells which appear to be important participants in glomerular injury via their secretory products. As other scavenger receptor ligands can increase macrophage nitric oxide generation, we examined whether oxidation of matrix could increase the activity of macrophage inducible nitric oxide synthase (iNOS). METHODS: Extracellular matrix was oxidized using a metal-catalyzed oxidation system. Matrix oxidation was measured using carbonyl analysis, and iNOS activity in macrophages seeded onto the matrix was measured by nitrite determination and Western and Northern analyses for iNOS. RESULTS: Macrophages exposed to oxidized matrix demonstrated a significant enhancement of iNOS activity. This enhancement could be antagonized by cotreatment of matrix with the radical spin trap N-tert-butyl-a-phenylnitrone, resulting in a corresponding decrease in protein carbonyl content, a measure of protein oxidation. Seeding macrophages onto oxidized matrix and adding the scavenger receptor ligand polyinosinic acid further augmented iNOS activity, suggesting that additional scavenger receptors were available to bind ligand and that further augmentation of iNOS activity did not require an additional change in cell shape. Western blot analysis revealed an increase in iNOS protein expression as a consequence of interaction with the oxidized matrix, but there was no difference in iNOS mRNA expression by Northern analysis suggesting a post-transcriptional mechanism for enhanced iNOS activity. CONCLUSION: These data demonstrate that oxidation of extracellular matrix enhances macrophage nitric oxide generation, and suggest a previously undescribed role for extracellular matrix modification in the regulation of cellular function and possibly the mediation of glomerular injury.

Animals↗

PubMed1998

Natural killer cell activity in patients with Graves' disease and Hashimoto's thyroiditis.

Natural killer (NK) cell activity of peripheral blood lymphocytes (PBL) against k562 human tumor cell targets was studied in patients with Graves' disease and Hashimoto's thyroiditis. NK activity was measured in a standard 4-hour 51chromium (Cr) release assay. Cytotoxicity was expressed as lytic units (LU)/10(6) PBL. Significantly decreased NK cell activity was demonstrated in both groups of patients, with mean (+/- SE) lytic units of 10.3 (+/- 9.1) and 13.3 (+/- 10.3) for patients with Graves' disease and Hashimoto's thyroiditis, respectively, compared with 36.0 (+/- 26.3) for age- and sex-matched normal subjects. When patients with Graves' disease were analyzed according to their thyroid status; NK activity was significantly depressed in (1) hyperthyroid patients before treatment; (2) hyperthyroid patients receiving antithyroid therapy; and (3) euthyroid patients receiving antithyroid therapy, compared with normal subjects. Graves' disease patients who were hypothyroid after radioactive iodine therapy or thyroidectomy had normal NK activity. No significant differences between hyperthyroid and euthyroid patients or between hypothyroid patients and normal subjects were demonstrated. NK activity in patients with Graves' disease did not correlate with serum levels of thyroxine, the presence or severity of ophthalmopathy, or titers of serum thyroid antibodies. In patients with Hashimoto's thyroiditis there was no correlation between NK activity and goiter size, titers of antithyroid antibodies, or thyroid status. These findings suggest that depression of NK activity in both disorders is secondary to abnormalities of thyroid hormone secretion, although an effect of the underlying autoimmune reactions has not been excluded.

Adolescent↗

PubMed1998

Glutamine reduces heat shock-induced cell death in rat intestinal epithelial cells.

Glutamine supplementation is beneficial for preventing intestinal atrophy and maintaining mucosal functions in metabolically stressed patients. The mechanisms by which glutamine prevents mucosal atrophy remain unclear. In particular, the role of glutamine in the survival of cells under stress is unknown. Intestinal epithelial cells (IEC-6) were cultured in media with or without supplementation of L-glutamine. A low concentration of L-glutamine (1.0 mmol/L) was sufficient to minimize the percentage of floating cells under basal conditions. Heat shock at 43 degrees C for 90 min decreased (P < 0. 001) the number of attached cells, while increasing (P < 0.001) the number of floating cells, which is a measurement of the extent of cell death in these cultures. Glutamine enhanced attached cell count and diminished heat shock-induced cell death in a dose-dependent manner. Of note, 2 mmol/L was suboptimal in both respects, thus indicating that heat-shocked cells require higher concentrations of glutamine for optimal cell survival. Maximal effect was achieved with 8 mmol/L glutamine, which increased (P < 0.001) cell growth (indicated by the number of attached cells) and diminished (P < 0. 001) cell death (indicated by the number of floating cells). Further increase of L-glutamine concentration to 12 or 20 mmol/L did not provide additional benefit in minimizing cell death. Heat shock protein 70 (hsp 70) mRNA was induced by heat shock only in cultures supplemented with L-glutamine, and the induction was more consistent and greater in cultures containing higher concentrations of glutamine. Thus, glutamine supplementation reduced heat shock-induced cell death. This effect, together with the maintenance of cell growth, may play a key role in the prevention of intestinal mucosal atrophy.

Animals↗

PubMed1997

Use of a non-porous polyurethane membrane as a sample support for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of peptides and proteins.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) of proteins and peptides was performed on samples deposited onto non-porous ether-type polyurethane (PU) membranes. Spectra obtained using PU membranes showed that mass resolution and accuracy were equivalent to values observed using a metal target, and superior to those obtained using poly(vinylidene difluoride) (PVDF) membranes. A small apparent increase in the mass of proteins and also loss of resolution were observed at very high laser irradiance due to charging, but were not observed under normal conditions. Analysis of NaCl-doped standards demonstrated that PU membranes yielded better results than a metallic target for salt-containing solutions. Relatively strong hydrophobic interactions between the proteins and peptides and the PU membrane allowed the incorporation of a washing step. This step allowed for the removal of salts and buffer components and thus provided an increase in resolution and mass accuracy. Digestion of citrate synthase (a protein of molecular weight 47,886) with trypsin was performed directly on the surface of the membrane for variable periods of time, and characteristic peptide fragments were observed by MALDI-TOFMS. Delayed extraction was used to increase the resolution and to permit more accurate mass assignments for those fragments. The use of PU membranes for MALDI-TOFMS analysis of proteins with higher molecular weights is also demonstrated.

Citrate (si)-Synthase↗

PubMed1997

A double-blind, randomized study of three antimicrobial regimens in the prevention of infections after elective colorectal surgery.

The objective of this study was to assess the prophylactic efficacy of cefoxitin, ceftizoxime, and metronidazole-gentamicin in colorectal surgery. A double-blind, randomized prospective clinical trial design was used in a Canadian tertiary care teaching hospital. Patients were randomized to one of three treatment groups and received three doses of a study drug (30 min preoperative and 2 postoperative doses at 12 and 24 h). Cefoxitin and ceftizoxime were given as 1000-mg doses. Metronidazole-gentamicin was given as 500 mg of metronidazole plus 120 mg of gentamicin in a minibag. High-risk patients (bowel ischemia, diabetic, current steroid use, etc.) received 10 postoperative doses. Patients with infections, prior antibiotics, or study drug allergies were excluded. Over 30 months, 153 patients were enrolled. Thirty-one patients were excluded for protocol violations. Of the 122 evaluable patients (38 ceftizoxime, 45 metronidazole-gentamicin, 39 cefoxitin), there was no difference across groups regarding sex, age, weight, preoperative Apache II score, and prior history of bowel surgery. Groups were equivalent regarding surgeon, nursing unit, high-risk status (six ceftizoxime, seven metronidazole-gentamicin, seven cefoxitin), bowel preparation, and procedure (including blood loss, drains, organ injury, intraoperative complications). Clinically significant infection requiring systemic antibiotics (7-day hospital and 30-day follow-up) was identified in 0% of ceftizoxime, 15% of metronidazole-gentamicin, and 26% of cefoxitin receiving patients (p = 0.005). Mean ASEPSIS scores for each group were 2.3 (range 0-15) for ceftizoxime, 9.2 (range 0-45) for metronidazole-gentamicin, and 10.4 (range 0-75) for cefoxitin (p = 0.01). Ceftizoxime patients tended to have a shorter total hospital stay (12.2 days versus 19.7 days for cefoxitin versus 13.9 days for metronidazole-gentamicin; p = 0.04), although the procedure to discharge interval was not significantly different (p = 0.09). There was no difference in clinical outcome according to risk status. Anaerobic bacteria were observed more commonly in the ceftizoxime and cefoxitin groups, whereas enteric Gram-negative aerobes were observed most often in the metronidazole-gentamicin group. The study regimens were generally well tolerated. Drug costs were equivalent between ceftizoxime and cefoxitin and lowest with the metronidazole-gentamicin regimen. Ceftizoxime appears to be more effective for the prevention of infection in colorectal surgery than either cefoxitin or metronidazole-gentamicin in the dosage regimens studied.

Adult↗

PubMed1996

G-protein-gated inward rectifier K+ channel proteins (GIRK1) are present in the soma and dendrites as well as in nerve terminals of specific neurons in the brain.

G-protein-gated inward rectifier potassium (GIRK) channels are coupled to numerous neurotransmitter receptors in the brain and can play important roles in modulating neuronal function, depending on their localization in a given neuron. Site-directed antibodies to the extreme C terminus of GIRK1 (or KGA1), a recently cloned component of GIRK channels, have been used to determine the relative expression levels and distribution of the protein in different regions of the rat brain by immunoblot and immunohistochemical techniques. We report that the GIRK1 protein is expressed prominently in the olfactory bulb, hippocampus, dentate gyrus, neocortex, thalamus, cerebellar cortex, and several brain stem nuclei. In addition to the expected localization in somas and dendrites, where GIRK channels may mediate postsynaptic inhibition, GIRK1 proteins were also found in axons and their terminal fields, suggesting that GIRK channels can also modulate presynaptic events. Furthermore, the distribution of the protein to either somatodendritic or axonal-terminal regions of neurons varied in different brain regions, which would imply distinct functions of these channels in different neuronal populations. Particularly prominent staining of the cortical barrels of layer IV of the neocortex, and the absence of this staining with unilateral kainate lesions of the thalamus, suggest that the GIRK1 protein is expressed in thalamocortical nerve terminals in which GIRK channels may mediate the actions of mu opiate receptors.

Amino Acid Sequence↗

PubMed1996

Double-blind comparison of cefazolin and ceftizoxime for prophylaxis against infections following elective biliary tract surgery.

Antibiotics have been shown to reduce the incidence of wound infections after elective biliary tract procedures. Cefazolin and cefoxitin are among the agents most commonly promoted for this purpose. Cefoxitin has been substituted with ceftizoxime in many institutions; however, the role of ceftizoxime as a prophylactic agent in this setting has not been determined. To assess the comparative prophylactic efficacies of cefazolin and ceftizoxime in biliary tract surgery, we conducted a double-blind, randomized prospective clinical trial in a tertiary-care teaching hospital. Adult patients were randomized to one of two treatment groups and received a 30-min preoperative dose of study drug and as many as two postoperative doses at 12 and 24 h, depending on hospitalization status. Cefazolin and ceftizoxime were given as 1,000-mg doses. Patients with infections, those receiving prior antibiotics, or those with beta-lactam allergies were excluded. Over the 19-month study tenure, 167 patients were enrolled. Seventeen patients were excluded from analysis because of protocol violations. Of the 150 evaluable patients (72 and 78 receiving cefazolin and ceftizoxime doses, respectively), there was no significant difference among groups regarding sex, age, weight, preoperative Apache II score, baseline chemistry, and hematological parameters. Groups were also equivalent regarding the surgeon, type of procedure, characteristics (blood loss, drains, organ injury, and complications), and duration of hospital stay (mean, 5.6 versus 4.3 days [P = 0.31]). No clinical evidence of infection (7-day hospital stay and 30-day follow-up) was identified in 93% of cefazolin and 92% of ceftizoxime patients (P = 1.0). Microbiological confirmation was found in only 18% of primary-site infections. In conclusion, cefazolin and ceftizoxime appear to be equivalent for the prevention of infection in biliary tract surgery with the dosage regimens studied.

Adult↗

PubMed1996

Regulation of AE2 Cl-/HCO3- exchanger during intestinal development.

The Na(+)-independent Cl-/HCO3- exchangers in many tissues are encoded by AE2, which is a member of the anion-exchanger (AE) gene family. Molecular regulation of AE2 remains mostly unknown. In this study, AE2 mRNA levels during enterocyte differentiation and intestinal development were examined in rat small intestines. AE2 mRNA levels did not show obvious segmental variation or vary along the crypt-villus axis. In contrast, AE2 mRNA decreased between days 14 and 21 after birth, the period corresponding to the transition period between the suckling and weanling stages in rats. Effects of L-thyroxine and dexamethasone on AE2 expression were examined because these hormones play important roles in gene regulation during development. L-Thyroxine administration starting on day 9 after birth led to an approximately 60% decrease (P < 0.001) in AE2 mRNA on day 13. Dexamethasone had no effect on AE2 mRNA level regardless of whether the hormone was administered alone or in conjunction with L-thyroxine. Our results indicate that AE2 mRNA in the intestine is regulated during development and that this regulation may in part be related to hormonal changes.

Aging↗

PubMed1996

Biochemical activity, pharmacokinetics and tolerability of tepoxalin, a cyclooxygenase/5-lipoxygenase inhibitor, in man.

Tepoxalin, a novel inhibitor of cyclooxygenase (CO) and 5-lipoxygenase (5-LO), was investigated for biochemical activity and pharmacokinetics in two studies. Study I was a 4-period, double-blind, randomized, single rising dose using 2 alternating panels (A, B) with interspersed placebo design (A: 25, 100, 400 mg, B: 50, 200, 800 mg p.o.). Study II was a 3-panel, randomized, placebo-controlled, double-blind, multiple dose study (A: 100 mg, B: 200 mg, C: 400 mg). In both studies, CO inhibition was assessed by generation of serum thromboxane (TxB2), 5-LO activity by LTB4 production ex vivo in Caionophore-stimulated blood. Plasma drug concentrations were assayed by HPLC for tepoxalin and its identified acid metabolite. It was found in both studies that at all dose levels the TxB2 generation was markedly suppressed (> 95% 2 h postdose). In study I, at 2 h postdose, % inhibition of LTB4 biosynthesis was marginal for the 3 lower doses but significant at 200 (14%), 400 (25%) and 800 mg (43%). In study II, the only significant inhibition occurred at the 400 mg dose at 6 h postdose on day 1 (17%) and on day 8 at 4, 6 and 8 h postdose (32, 42 and 32% respectively). In both studies and at all doses, plasma concentrations of tepoxalin varied widely between subjects. Linearity between plasma concentrations and dose could not be ascertained, and correlation between drug plasma levels and effect on LTB4 synthesis was poor. Single doses up to 800 mg and multiple doses up to 400 mg of tepoxalin were generally well tolerated.

Adult↗

PubMed1995

Expression of airway secretory epithelial functions by lung carcinoma cells.

We examined 12 non-small cell lung carcinoma cell lines for expression of airway goblet, serous, and mucous cell characteristics. The cells expressed some ultrastructural traits of secretory epithelial cells but none contained secretory granules typical of the airway secretory cells. Using immunocytochemistry and cell-specific monoclonal antibodies, we identified heterogeneous expression of goblet, mucous, and serous cell markers among the cell lines. After metabolic radiolabeling, cells incorporated isotope into high molecular weight material. Incubation of pulse-radiolabeled cells with a number of known mucus secretogogues revealed that 5 of the 12 cell lines released radiolabeled material in response to the agonists. However, in each cell line only one of the receptor-activated pathways tested was intact. Although we did not identify a single cell line expressing a phenotype similar to normal airway secretory cells, particular functions retained by some of these cell lines may make them useful for specific studies of mucus production or secretion.

Bronchi↗

PubMed1995

Mycetome endosymbionts of tsetse flies constitute a distinct lineage related to Enterobacteriaceae.

Tsetse flies (Diptera: Glossinidae) harbour two morphologically different endosymbionts intracellularly associated with gut tissue: a primary (P) and a secondary (S) organism. The P-endosymbiont is a gram-negative rod, 8-10 microns in size, and resides intracellularly within specialized cells, mycetocytes which are organized into an organelle (mycetome), in the anterior portion of the gut. The S-endosymbiont is a smaller (1-2 microns) gram-negative rod and is harboured in the epithelial sheath cells in midgut. Phylogenetic characterization of S-endosymbionts from taxonomically distant insects including tsetse flies has shown that they are related to the free-living bacterium, Escherichia coli, and are members of the family Enterobacteriaceae within the gamma-3 subdivision of Proteobacteria. In this study, a polymerase chain reaction (PCR) based assay was designed utilizing the conserved sequences of 16S rDNA in order to phylogenetically characterize the mycetome-associated P-endosymbionts directly from tsetse mycetome tissue. Analysis from five species of flies representing the three major subgenera of genus Glossina indicates that P-endosymbionts constitute a distinct lineage within the gamma-3 subdivision of Proteobacteria. Mycetome endosymbiont phylogeny appears to parallel the classic taxonomic assignments independently developed for their insect host species. This suggests an ancient association for this symbiosis, which may have subsequently radiated with time, giving rise to the current species of tsetse flies and their modern-day endosymbionts. Based on endosymbiont phylogeny, the fusca flies constitute the most ancient subgenus, followed by the morsitans and palpalis groups.

Animals↗