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Biomedical subjects

M E O'Donnell

Publications and source records attributed to M E O'Donnell.

At least 19 recordsLinked to original sources
PubMed2006

Surgical treatment of malignant carcinoid tumours of the appendix.

Since their first description in 1882, malignant neoplasms of the appendix still remain rare. Malignant carcinoid tumours are the most common accounting for 85% of all appendiceal neoplasms. Preoperative diagnosis is invariably difficult, and precise treatment protocols for these neoplasms remain unclear. We reviewed our experience and searched published evidence to produce management guidelines. A retrospective review of all malignant carcinoid tumours diagnosed in our hospital between April 1994 and December 2003 was performed. Patient demographics, operative details, histological types and clinical outcomes were retrieved from case notes supplemented by a questionnaire to the patient's General Practitioner. A literature search was then performed. Nine patients were identified with classical carcinoid tumours (CCT); (M = 3 and F = 6, mean age: 43, range 14-81) and two patients with goblet-cell morphology (F = 2, age 46 and 76). Mean follow-up was 63 months (range 1-125 months). Nine patients were alive at the end of follow-up. Appendicectomy was performed for acute appendicitis. Other appendiceal pathologies were identified following hemicolectomy and oophorectomy. CCT are the most common tumours and have the better prognosis. From our experience and subsequent review of the literature, we recommend right hemicolectomy as the treatment of choice for malignant carcinoid tumours. However, small CCTs less than 2 cm in diameter at the tip of the appendix, with a low proliferative index, without angiolymphatic or mesoappendiceal extension can be treated by appendicectomy. Following oncological assessment, further adjuvant therapy should be considered for patients with advanced disease.

Adolescent↗
PubMed2006

Physiology and pathophysiology of Na+/H+ exchange and Na+ -K+ -2Cl- cotransport in the heart, brain, and blood.

Maintenance of a stable cell volume and intracellular pH is critical for normal cell function. Arguably, two of the most important ion transporters involved in these processes are the Na+/H+ exchanger isoform 1 (NHE1) and Na+ -K+ -2Cl- cotransporter isoform 1 (NKCC1). Both NHE1 and NKCC1 are stimulated by cell shrinkage and by numerous other stimuli, including a wide range of hormones and growth factors, and for NHE1, intracellular acidification. Both transporters can be important regulators of cell volume, yet their activity also, directly or indirectly, affects the intracellular concentrations of Na+, Ca2+, Cl-, K+, and H+. Conversely, when either transporter responds to a stimulus other than cell shrinkage and when the driving force is directed to promote Na+ entry, one consequence may be cell swelling. Thus stimulation of NHE1 and/or NKCC1 by a deviation from homeostasis of a given parameter may regulate that parameter at the expense of compromising others, a coupling that may contribute to irreversible cell damage in a number of pathophysiological conditions. This review addresses the roles of NHE1 and NKCC1 in the cellular responses to physiological and pathophysiological stress. The aim is to provide a comprehensive overview of the mechanisms and consequences of stress-induced stimulation of these transporters with focus on the heart, brain, and blood. The physiological stressors reviewed are metabolic/exercise stress, osmotic stress, and mechanical stress, conditions in which NHE1 and NKCC1 play important physiological roles. With respect to pathophysiology, the focus is on ischemia and severe hypoxia where the roles of NHE1 and NKCC1 have been widely studied yet remain controversial and incompletely elucidated.

Animals↗
PubMed2001

Quantitation of Na+-K+-2Cl- cotransport splice variants in human tissues using kinetic polymerase chain reaction.

A kinetic reverse transcription-polymerase chain reaction (RT-PCR)-based assay is described that can discriminate and quantitate differentially spliced mRNAs. This assay should be generally applicable for high-throughput quantitation of differentially spliced transcripts. The utility of this method was assessed for spliced transcripts encoded by the human Na+-K+-2Cl- cotransporter gene hNKCC1. Evidence is presented that the NKCC1 isoform of the human Na+-K+-2Cl- cotransporter is differentially spliced analogous to that recently described for the mouse Na+-K+-2Cl- cotransporter gene BSC2. The nucleotide sequences of the two human splice variants predict Na+-K+-2Cl- cotransporter proteins differing only in length. Stable transfectants expressing these human splice variants, designated NKCC1a or NKCC1b, were constructed. Both splice variants produce functional Na+-K+-2Cl- cotransporters in vivo. The abundance of NKCC1 mRNA and patterns of differential splicing in 10 different tissue types and three cell lines were quantitated using the kRT-PCR assay. The results showed that the total amount of NKCC1 mRNA varied by more than 30-fold in the human tissues and cell lines examined. The ratio of NKCC1a/NKCC1b varied nearly 70-fold among these same tissues and cell lines suggesting that differential splicing of the NKCC1 transcript may play a regulatory role in human tissues.

Alternative Splicing↗
PubMed2001

Vesicle-mediated trafficking of parasite proteins to the host cell cytosol and erythrocyte surface membrane in Plasmodium falciparum infected erythrocytes.

During the development of the asexual stage of the malaria parasite, Plasmodium falciparum, the composition, structure and function of the host cell membrane is dramatically altered, including the ability to adhere to vascular endothelium. Crucial to these changes is the transport of parasite proteins, which become associated with or inserted into the erythrocyte membrane. Protein and membrane targeting beyond the parasite plasma membrane must require unique pathways, given the parasites intracellular location within a parasitophorous vacuolar membrane and the lack of organelles and biosynthetic machinery in the host cell necessary to support a secretory system. It is not clear how these proteins cross the parasitophorous vacuolar membrane or how they traverse the erythrocyte cytosol to reach their final destinations. The identification of: (1) a P. falciparum homologue of the protein Sar1p, which is an essential component of the COPII-based secretory system in mammalian cells and yeast and (2) electron-dense, possibly coated, secretory vesicles bearing P. falciparum erythrocyte membrane protein 1 and P. falciparum erythrocyte membrane protein 3 in the host cell cytosol of P. falciparum infected erythrocytes recently provided the first direct evidence of a vesicle-mediated pathway for the trafficking of some parasite proteins to the erythrocyte membrane. The major advance in uncovering the parasite-induced secretory pathway was made by incubating infected erythrocytes with aluminium tetrafluoride, an activator of guanidine triphosphate-binding proteins, which resulted in the accumulation of the vesicles into multiple vesicle strings. These vesicle complexes were often associated with and closely abutted the erythrocyte membrane, but were apparently prevented from fusing by the aluminium fluoride treatment, making their capture by electron microscopy possible. It appears that malaria parasites export proteins into the host cell cytosol to support a vesicle-mediated protein trafficking pathway.

Animals↗
PubMed2001

Flow-induced expression of endothelial Na-K-Cl cotransport: dependence on K(+) and Cl(-) channels.

Steady laminar shear stress has been shown previously to markedly increase Na-K-Cl cotransporter mRNA and protein in human umbilical vein endothelial cells and also to rapidly increase endothelial K(+) and Cl(-) channel conductances. The present study was done to evaluate the effects of shear stress on Na-K-Cl cotransporter activity and protein expression in bovine aortic endothelial cells (BAEC) and to determine whether changes in cotransporter expression may be dependent on early changes in K(+) and Cl(-) channel conductances. Confluent BAEC monolayers were exposed in a parallel-plate flow chamber to either steady shear stress (19 dyn/cm(2)) or purely oscillatory shear stress (0 +/- 19 dyn/cm(2)) for 6-48 h. After shearing, BAEC monolayers were assessed for Na-K-Cl cotransporter activity or were subjected to Western blot analysis of cotransporter protein. Steady shear stress led to a 2- to 4-fold increase in BAEC cotransporter protein levels and a 1.5- to 1.8-fold increase in cotransporter activity, increases that were sustained over the longest time periods studied. Oscillatory flow, in contrast, had no effect on cotransporter protein levels. In the presence of flow-sensitive K(+) and Cl(-) channel pharmacological blockers, the steady shear stress-induced increase in cotransporter protein was virtually abolished. These results suggest that shear stress modulates the expression of the BAEC Na-K-Cl cotransporter by mechanisms that are dependent on flow-activated ion channels.

Aorta↗
PubMed2001

Expression of aquaporin-1 in human trabecular meshwork cells: role in resting cell volume.

PURPOSE: Drainage of aqueous humor from the human eye appears dependent on intracellular volume of trabecular meshwork (TM) cells, the predominant cell type of the human outflow pathway. Thus, the modulation of water and solute flux across the plasma membrane of TM cells is predicted to be an important factor in regulating outflow facility. Aquaporin (AQP)-1 is a hexahelical integral membrane protein that functions as a regulated channel for water and cations in fluid-secreting and -absorbing tissues. AQP1 is present in many tissues of the human eye, including the TM; however, its role in outflow facility is unknown. The purpose of the present study was twofold: to evaluate the prospect of manipulating AQP1 protein levels in TM cells using sense and antisense mRNA and to investigate the functional role of AQP1 in TM cells. METHODS: An adenovirus (AV) expression system was used to alter AQP1 protein levels. AQP1 protein expression was monitored using immunoblot analysis, and resting cell volume was measured by forward light scatter, electronic cell sizing, and [(14)C]-sucrose/urea equilibration. Permeability of TM monolayers to [(14)C]-sucrose was also assessed as an indirect evaluation of cell volume. RESULTS: AV-mediated gene transfer of AQP1 cDNA to TM cells resulted in a titer-dependent increase in recombinant AQP1, whereas transfer of antisense cDNA decreased native AQP1 protein by 71.7% +/- 5.5% (P < 0.01) after 5 days. A novel finding of this study is that mean resting volumes of AQP1(s) AV-infected TM cells in suspension were 8.7% +/- 3.0% greater (P < 0.05) than control cells. Conversely, AQP1 antisense (as) AV-infected cells had resting volumes 7.8% +/- 2.9% less than control cells (P < 0.05). Similar effects of AQP1 expression on resting cell volume were observed in TM monolayers. Consistent with this finding, paracellular permeability of AQP1(s) AV-infected TM monolayers to [(14)C]-sucrose decreased by 8.0% +/- 1.4% (P < 0.001). CONCLUSIONS: In addition to influencing the osmotic permeability of TM plasma membranes, the level of AQP1 protein expression influences resting intracellular volume and thus paracellular permeability of TM cell monolayers in vitro. These data suggest that AQP1 expression may affect outflow facility in vivo.

Adenoviruses, Human↗
PubMed1999

Biobehavioral responses to acute pain in adolescents with a significant neurologic impairment.

OBJECTIVE: To further understand acute pain response in children with a significant neurologic impairment (SNI), we undertook a descriptive hypothesis-generating study of the response to a routine vaccine among adolescents with SNI. DESIGN: Within-subject crossover design. SETTING: Tertiary care facility for children and adolescents with SNI. PATIENTS: Eight adolescents (mean age = 15 years). INTERVENTIONS: Mock and real vaccine injections. OUTCOME MEASURES: Quantitative measures of heart rate, videotaped facial action, Child Facial Coding System (CFCS), and Facial Action Coding System (FACS); observer ratings visual analog scale (VAS) were obtained before, during, and after a mock injection and routine annual influenza vaccine injection presented in a counterbalanced order. RESULTS: VAS scores were significantly higher during the injection phase than during the other time periods; however, there were no significant differences across study time periods when using the other outcome measures. CONCLUSIONS: Although the dampened behavioral and physiologic reactions to an acute noxious stimulus were similar to those of previous work with developmentally delayed children and frail elderly, it remains unclear what underlies the apparent reduced pain response in this setting. These findings have potentially important implications for the daily care of individuals with significant neurologic impairment and illustrate the compelling need tor further study of the unique character of the pain experience in this setting.

Acute Disease↗
PubMed1999

Pain in children with significant neurological impairment.

The pain experience in the child with a significant neurological impairment is complex and confusing, and it raises many questions about the very nature of pain itself. Early work in this field suggests that the pain experience may be blunted. The neurological impairment associated with conditions such as cerebral palsy may alter the neurological system and hence the ability to comprehend and communicate pain; there is no evidence to date that this reflects true pain insensitivity or indifference. From recent work, the emerging body of evidence supports a relationship between the pain system and the motor, sensory, and autonomic systems and demonstrates how alterations to these systems may have a profound and unique impact on the pain experience. Beyond the altered neurological substrate, communication disabilities and social/environmental factors also seem to alter the pain experience. Establishing a clear pain history, including baseline information of child-specific patterns of behaviors and ongoing comparative use of this information over time, can provide clinically meaningful measures. Pain management should be directed at the underlying sources of pain and should include the analgesic ladder for everyday pain, opioids for acute/procedural pain (+/- benzodiazepine adjuvants), and antispasticity medications for high tone. With appropriate monitoring, demand and regional analgesic techniques can provide effective and safe postoperative pain control. The lack of basic and clinical knowledge in this field, however, adds another challenge to the clinician.

Analgesics↗
PubMed1999

Intracellular Cl regulates Na-K-Cl cotransport activity in human trabecular meshwork cells.

The trabecular meshwork (TM) of the eye plays a central role in modulating intraocular pressure by regulating aqueous humor outflow, although the mechanisms are largely unknown. We and others have shown previously that aqueous humor outflow facility is modulated by conditions that alter TM cell volume. We have also shown that the Na-K-Cl cotransport system is a primary regulator of TM cell volume and that its activity appears to be coordinated with net efflux pathways to maintain steady-state volume. However, the cellular mechanisms that regulate cotransport activity and cell volume in TM cells have yet to be elucidated. The present study was conducted to investigate the hypothesis that intracellular Cl concentration ([Cl]i) acts to regulate TM cell Na-K-Cl cotransport activity, as has been shown previously for some other cell types. We demonstrate here that the human TM cell Na-K-Cl cotransporter is highly sensitive to changes in [Cl]i. Our findings reveal a marked stimulation of Na-K-Cl cotransport activity, assessed as ouabain-insensitive, bumetanide-sensitive K influx, in TM cells following preincubation of cells with Cl-free medium as a means of reducing [Cl]i. In contrast, preincubation of cells with media containing elevated K concentrations as a means of increasing [Cl]i results in inhibition of Na-K-Cl cotransport activity. The effects of reducing [Cl]i, as well as elevating [Cl]i, on Na-K-Cl cotransport activity are concentration dependent. Furthermore, the stimulatory effect of reduced [Cl]i is additive with cell-shrinkage-induced stimulation of the cotransporter. Our studies also show that TM cell Na-K-Cl cotransport activity is altered by a variety of Cl channel modulators, presumably through changes in [Cl]i. These findings support the hypothesis that regulation of Na-K-Cl cotransport activity, and thus cell volume, by [Cl]i may participate in modulating outflow facility across the TM.

Carrier Proteins↗
PubMed1999

Effects of Na-K-2Cl cotransport regulators on outflow facility in calf and human eyes in vitro.

PURPOSE: Cultured human trabecular meshwork (TM) cells possess substantial Na-K-Cl activity, which is involved in the regulation of TM cell volume. The hypothesis in the present study was that drugs that affect the cotransporter might alter aqueous humor outflow facility (C) in the intact eye. The effects of agents and conditions known to modulate Na-K-CI cotransport activity and/or TM cell volume on C in perfused anterior segments were investigated. METHODS: Human and calf eyes were dissected and perfused, and C was determined according to standard published methods. Perfusates with modified osmolarity were used to cause alterations in TM cell volume. Cl-free perfusate and/or bumetanide (10(-5) M) was used to inhibit Na-K-Cl cotransport activity, and vasopressin (10(-7) M, 10(-8) M) was used to stimulate cotransport activity. RESULTS: In human eyes, hypo-osmotic perfusate decreased C 12%, whereas hyper-osmotic perfusate increased C 44%. These changes lasted approximately 30 minutes, after which C began to normalize. Inhibition of Na-K-Cl cotransport using Cl-free medium or bumetanide resulted in facility increases of 27% and 22%, respectively. There was an additive increase in C with bumetanide plus Cl-free media. Stimulating Na-K-Cl cotransport with 10(-8)M and 10(-7)M vasopressin resulted in 28% and 35% decreases in C, respectively. The results were similar in calf eyes: Cl-free medium or bumetanide resulted in 41% and 52% increases in C, whereas 10(-8) M and 10(-7) M vasopressin resulted in 14% and 19% decreases in C, respectively. CONCLUSIONS: Modulation of Na-K-Cl cotransport results in changes in C that may be mediated in part by cell volume changes.

Aged↗
PubMed1999

How does the trabecular meshwork regulate outflow? Clues from the vascular endothelium.

Although it is well known that the trabecular meshwork plays a central role in regulating the outflow of aqueous humor from the eye, the cellular mechanisms and events responsible for this function are poorly understood. In contrast, the mechanisms by which vascular endothelial cells modulate the permeability of blood vessels have been more thoroughly investigated. It is hypothesized that the cells of the TM employ mechanisms similar to those observed in the vascular endothelium to modulate aqueous humor permeability. Specifically, it is hypothesized that the cells of the TM employ Na-K-Cl cotransport to modulate their intracellular volume and thus the volume of the paracellular pathways through which aqueous humor may travel. The current knowledge about the role of Na-K-Cl cotransport and volume regulation in the regulation of vascular permeability, and evidence that similar physiologic events occur in the TM are reviewed. In addition, the implications for further study of aqueous humor outflow physiology are discussed.

Aqueous Humor↗
PubMed1999

Na-K-Cl cotransport in normal and glaucomatous human trabecular meshwork cells.

PURPOSE: Previous results from this laboratory showed that intracellular volume of trabecular meshwork (TM) cells is regulated by the Na-K-Cl cotransport system. Other studies suggest that TM cell volume, in turn, is a determinant of permeability across the TM. Given that a decrease in outflow facility across the TM is thought to be the primary cause of elevated intraocular pressure in primary open-angle glaucoma, the present study was conducted to investigate the possibility that Na-K-Cl cotransport function may be altered in glaucomatous TM cells compared with normal TM cells. METHODS: Normal and glaucomatous human TM cells were cultured from donor eyes and trabeculectomy specimens, respectively. Trabecular meshwork cell monolayers were evaluated for Na-K-Cl cotransport activity, assessed as ouabain-insensitive, bumetanide-sensitive K influx using 86Rb as a tracer for K. Cotransporter protein expression was determined by western blot analysis, and intracellular volume was determined radioisotopically using [14C]urea and [14C]sucrose as markers of total and extracellular water space, respectively. RESULTS: Na-K-Cl cotransport activity of glaucomatous TM cells was found to be reduced by 32% +/- 2% compared with that of normal TM cells, whereas western blot analyses showed that cotransporter protein expression in glaucomatous TM cells was reduced by 64% +/- 14% compared with expression in normal TM cells. Also, exposure of normal TM cells to 10 microM norepinephrine or 50 microM 8-bromo-3',5'-cyclic adenosine monophosphate was found to diminish Na-K-Cl cotransport activity, whereas these agents were without effect on glaucomatous TM cell cotransport. Finally, resting cell volume of glaucomatous TM cells was found to be increased compared with that of normal TM cells, whereas intracellular volume of both cell types was reduced after exposure to 10 microM benzmetanide or 10 microM bumetanide. CONCLUSIONS: These findings indicate that Na-K-Cl cotransport function and regulation are altered in glaucomatous TM cells compared with that of normal TM cells. However, the observation that cell volume of glaucomatous TM cells is greater than that of normal TM cells, despite reduced Na-K-Cl cotransport activity, suggests that other volume-regulatory ion flux pathways may be involved in the reduced outflow of glaucoma.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1998

Role of the adenovirus DNA-binding protein in in vitro adeno-associated virus DNA replication.

A basic question in adeno-associated virus (AAV) biology has been whether adenovirus (Ad) infection provided any function which directly promoted replication of AAV DNA. Previously in vitro assays for AAV DNA replication, using linear duplex AAV DNA as the template, uninfected or Ad-infected HeLa cell extracts, and exogenous AAV Rep protein, demonstrated that Ad infection provides a direct helper effect for AAV DNA replication. It was shown that the nature of this helper effect was to increase the processivity of AAV DNA replication. Left unanswered was the question of whether this effect was the result of cellular factors whose activity was enhanced by Ad infection or was the result of direct participation of Ad proteins in AAV DNA replication. In this report, we show that in the in vitro assay, enhancement of processivity occurs with the addition of either the Ad DNA-binding protein (Ad-DBP) or the human single-stranded DNA-binding protein (replication protein A [RPA]). Clearly Ad-DBP is present after Ad infection but not before, whereas the cellular level of RPA is not apparently affected by Ad infection. However, we have not measured possible modifications of RPA which might occur after Ad infection and affect AAV DNA replication. When the substrate for replication was an AAV genome inserted into a plasmid vector, RPA was not an effective substitute for Ad-DBP. Extracts supplemented with Ad-DBP preferentially replicated AAV sequences rather than adjacent vector sequences; in contrast, extracts supplemented with RPA preferentially replicated vector sequences.

Adenoviridae↗