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Biomedical subjects

D Morrissey

Publications and source records attributed to D Morrissey.

12 recordsLinked to original sources
PubMed2001

In vitro selection criteria for probiotic bacteria of human origin: correlation with in vivo findings.

The enteric flora comprises approximately 95% of the total number of cells in the human body and can elicit immune responses while protecting against microbial pathogens. However, the resident bacterial flora of the gastrointestinal tract may also be implicated in the pathogenesis of diseases such as inflammatory bowel disease (ulcerative colitis and Crohn disease). The objectives of the Probiotic Research Group based at University College Cork were to isolate and identify lactic acid bacteria exhibiting beneficial probiotic traits, such as bile tolerance in the absence of deconjugation activity, acid resistance, adherence to host epithelial tissue, and in vitro antagonism of pathogenic microorganisms or those suspected of promoting inflammation. To isolate potentially effective probiotic bacteria, we screened the microbial population adhering to surgically resected segments of the gastrointestinal tract (the environment in which they may subsequently be reintroduced and required to function). In total, 1500 bacterial strains from resected human terminal ilea were assessed. From among these organisms, Lactobacillus salivarius subsp. salivarius strain UCC118 was selected for further study. In mouse feeding trials, milk-borne L. salivarius strain UCC118 could successfully colonize the murine gastrointestinal tract. A human feeding study conducted in 80 healthy volunteers showed that yogurt can be used as a vehicle for delivery of strain UCC118 to the human gastrointestinal tract with considerable efficacy in influencing gut flora and colonization. In summary, we developed criteria for in vitro selection of probiotic bacteria that may reflect certain in vivo effects on the host such as modulation of gastrointestinal tract microflora.

Animals↗
PubMed1999

Invasion by esophageal cancer cells: functional contribution of the urokinase plasminogen activation system, and inhibition by antisense oligonucleotides to urokinase or urokinase receptor.

Early metastasis contributes to the very poor prognosis of esophageal carcinoma. The recent immunohistochemical finding that invasive esophageal carcinomas express elevated levels of urokinase (uPA) and urokinase receptor (uPA-R) in vivo suggest that the plasminogen activation system may contribute to metastasis in esophageal cancer. The aim of our study was to functionally investigate, at the molecular level, the relative contribution of uPA and uPA-R to the invasiveness of esophageal cancer cells in vitro. The three esophageal cancer cell lines, OC1-3, generated in our laboratory, were analyzed for uPA and uPA-R expression by RT-PCR, immunoenzymatic staining, and quantitative ELISA. Invasiveness of all cell lines was quantified as percentage cellular invasiveness in a standardized Matrigel in vitro assay. OC1 and OC3, which were found to coexpress both uPA and uPA-R, displayed stronger invasiveness (44% and 32.5% respectively) relative to OC2 (19%) which expressed uPA-R but was negative for uPA. Transfection of OC2 cells with the uPA cDNA resulted in two variants, OC2.uPA1 and OC2.uPA2, stably expressing functional uPA. Both transfectants exhibited enhanced invasiveness (60% and 50% respectively) relative to the parent uPA-negative OC2 cells (19%). Antisense oligonucleotide inhibition of either uPA or uPA-R expression resulted in a similar, marked reduction in invasiveness of esophageal tumor cells which normally coexpress both molecules (OC1, OC3 and the uPA-expressing OC2-transfectant clones). Neither antisense treatment altered the basal invasiveness of OC2, which expresses uPA-R but not uPA. In conclusion, coexpression of uPA with its receptor, uPA-R, is required for functional involvement of the urokinase system in invasion by esophageal carcinoma cells. Our results suggest that these synergistic mediators of invasiveness are quantitatively major contributors to the invasiveness of esophageal carcinoma.

DNA, Complementary↗
PubMed1998

Plasticity of Drosophila paired function.

The Drosophila Paired (Prd) transcription factor has homeodomain (HD) and paired domain (PD) DNA-binding activities required for in vivo function. Correspondingly, Prd activation of late even-skipped (eve) expression occurs through a conserved target sequence (PTE) with HD and PD half sites, both of which are required for activation. To investigate the relationship between the HD and PD, and their roles in conferring specificity to Prd function, we tested altered versions of the Prd protein and of the PTE target site using in vivo assays in embryos. We found that function through PTE was constrained by the targeting specifications of both the HD and PD as well as the spatial relationship between these two domains. PTE function was also constrained by the spacing between the target half sites for the PD and HD, although surprisingly, late eve activation was retained when PTE was replaced by in vitro optimized binding sites for either the PD alone or for an HD dimer. In contrast to late eve regulation, other Prd targets tolerated more changes in the Prd protein, suggesting that their target sequences may be qualitatively different from PTE.

Animals↗
PubMed1997

Identification of CDK4 sequences involved in cyclin D1 and p16 binding.

Activation of CDK4 is regulated, in part, by its association with a D-type cyclin. Conversely, CDK4 activity is inhibited when it is bound to the cyclin-dependent kinase inhibitor, p16(INK4A). To investigate the molecular basis of the interactions between CDK4 and cyclin D1 or p16(INK4A) we performed site-directed mutagenesis of CDK4. The interaction was examined using in vitro translated wild type and mutant CDK4 proteins and bacterially expressed cyclin D1 and p16 fusion proteins. As mutational analysis of CDC2 suggests that its cyclin binding domain is primarily located near its amino terminus, the majority of the mutations constructed in CDK4 were located near its amino terminus. In addition, CDK4 residues homologous to CDC2 sites involved in Suc1 binding were also mutated. Our analysis indicates that cyclin D1 and p16 binding sites are overlapping and located primarily near the amino terminus. All CDK4 mutations that resulted in decreased p16 binding capability also diminished cyclin D1 binding. In contrast, amino-terminal sequences were identified, including the PSTAIRE region, that are important for cyclin D1 binding but are not involved in p16 binding.

Amino Acid Sequence↗
PubMed1996

Both the paired domain and homeodomain are required for in vivo function of Drosophila Paired.

Drosophila paired, a homolog of mammalian Pax-3, is key to the coordinated regulation of segment-polarity genes during embryogenesis. The paired gene and its homologs are unusual in encoding proteins with two DNA-binding domains, a paired domain and a homeodomain. We are using an in vivo assay to dissect the functions of the domains of this type of molecule. In particular, we are interested in determining whether one or both DNA-binding activities are required for individual in vivo functions of Paired. We constructed point mutants in each domain designed to disrupt DNA binding and tested the mutants with ectopic expression assays in Drosophila embryos. Mutations in either domain abolished the normal regulation of the target genes engrailed, hedgehog, gooseberry and even-skipped, suggesting that these in vivo functions of Paired require DNA binding through both domains rather than either domain alone. However, when the two mutant proteins were placed in the same embryo, Paired function was restored, indicating that the two DNA-binding activities need not be present in the same molecule. Quantitation of this effect shows that the paired domain mutant has a dominant-negative effect consistent with the observations that Paired protein can bind DNA as a dimer.

Animals↗
PubMed1994

Regulation of ciliary neurotrophic factor in cultured rat hippocampal astrocytes.

Ciliary neurotrophic factor (CNTF) is a pleiotropic cytokine which is detectable only at very low levels in the intact adult rat CNS, but following an aspirative lesion to the dorsal hippocampus and overlying cortex, CNTF mRNA levels are dramatically up-regulated in reactive astrocytes. In cultured rat hippocampal astrocytes, CNTF mRNA levels are high, similar to the levels in reactive astrocytes in vivo, but are strongly suppressed after administration of isoproterenol and forskolin, which stimulate the production of intracellular cyclic AMP, induced marked morphological change in the astrocytes and up-regulate glial fibrillary acidic protein mRNA and nerve growth factor mRNA in these cells. Following a single administration of forskolin to cultured astrocytes, suppression of CNTF mRNA was sustained for up to 7 days. A similar down-regulation was observed with the endogenous adrenergic agonists noradrenaline and adrenaline as well as, to a lesser extent, dopamine and adenosine. Down-regulation of CNTF mRNA resulted in a gradual reduction in the level of CNTF protein within the astrocytes. A single addition of forskolin or isoproterenol resulted in a drop in CNTF protein levels to 29 and 52% of control levels respectively after 9 days in vitro, although the rate of turnover of CNTF remained the same. Down-regulation of CNTF mRNA in cultured hippocampal astrocytes by adenylyl cyclase activation was quite specific, as a wide range of growth factors, cytokines and neurotransmitters had little or no effect upon CNTF mRNA levels.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1993

Synergistic activation of transcription is mediated by the N-terminal domain of Drosophila fushi tarazu homeoprotein and can occur without DNA binding by the protein.

Synergistic activation of transcription by Drosophila segmentation genes in tissue culture cells provides a model with which to study combinatorial regulation. We examined the synergistic activation of an engrailed-derived promoter by the pair-rule proteins paired (PRD) and fushi tarazu (FTZ). Synergistic activation by PRD requires regions of the homeodomain or adjacent sequences, and that by FTZ requires the first 171 residues. Surprisingly, deletion of the FTZ homeodomain does not reduce the capacity of the protein for synergistic activation, although this mutation abolishes any detectable DNA-binding activity. This finding suggests that FTZ can function through protein-protein interactions with PRD or other components of the homeoprotein transcription complex, adding a new layer of mechanisms that could underlie the functional specificities and combinatorial regulation of homeoproteins.

Animals↗
PubMed1993

Radiologic case study. Antibiotic modified osteomyelitis.

The clinical and radiologic presentation of antibiotic-modified osteomyelitis closely resembles primary subacute pyogenic osteomyelitis. The widespread use of antibiotics has increased the incidence of this entity. When a patient is found to have a solitary bone lesion, a careful history should include inquiries regarding recent antibiotic therapy for any condition.

Anti-Bacterial Agents↗
PubMed1991

Functional dissection of the paired segmentation gene in Drosophila embryos.

An ectopic expression assay in Drosophila embryos was used to investigate the roles of pair-rule segmentation genes in the spatial regulation of the segment-polarity gene, engrailed (en). It is hypothesized that the regions of overlap in expression of two genes, paired (prd) and even-skipped (eve), define the odd-numbered en expression stripes. Consistent with this combinatorial model, ectopic expression of prd caused these en stripes to be expanded posteriorly. Surprisingly, however, ectopic expression of a prd gene with a deletion of the conserved paired box resulted in loss of these odd-numbered en stripes. This dominant negative effect is a phenocopy of en expression in prd embryos and suggests that the paired box is necessary for normal prd- function. A similar deletion of odd-numbered en stripes was also observed after ectopic expression of a chimeric fushi tarazu (ftz) gene containing a substituted prd gene homeo box; in addition, in these embryos, the even-numbered en stripes were expanded anteriorly, as observed when the unaltered ftz gene is ectopically expressed. These effects suggest that the chimeric protein may have DNA or protein targets of both the normal Ftz and Prd proteins.

Animals↗
PubMedDate not supplied

Open and closed kinetic chain exercises in the early period after anterior cruciate ligament reconstruction. Improvements in level walking, stair ascent, and stair descent.

Thirty-seven patients who had undergone anterior cruciate ligament reconstruction were tested in a gait analysis laboratory at 2 and 6 weeks after surgery. Between test sessions, patients were randomly assigned to a course of either closed or open kinetic chain resistance exercises (3 sessions per week for 4 weeks). Gait analysis consisted of bilateral calculations of knee joint angle, moment, and power during level walking, stair ascent, and stair descent. An analysis of variance on the effects of training group and test session indicated that the only variable to be significantly affected by the type of exercise program was the amount of knee flexion at the beginning of step-up (P < 0.05). All other measures of knee angle, moment, and power (16 total variables) showed no significant difference between the exercise groups. All variables measured on the injured side showed significant improvement from test 1 to test 2 (P < 0.05), but the injured leg remained functionally deficient when compared with the uninjured leg. These data suggest that there are no clinically significant differences in the functional improvement resulting from the choice of open or closed kinetic chain exercises in the early period after this surgery.

Analysis of Variance↗
PubMedDate not supplied

Probiotics: from myth to reality. Demonstration of functionality in animal models of disease and in human clinical trials.

The enteric flora comprise approximately 95% of the total number of cells in the human body and are capable of eliciting immune responses while also protecting against microbial pathogens. However, the resident bacterial flora of the gastrointestinal tract (GIT) may also be implicated in the pathogenesis of several chronic conditions such as inflammatory bowel disease (IBD). The University College Cork-based Probiotic Research Group has successfully isolated and identified lactic acid bacteria (LAB) which exhibit beneficial probiotic traits. These characteristics include the demonstration of bile tolerance; acid resistance; adherence to host epithelial tissue; and in vitro antagonism of potentially-pathogenic micro-organisms or those which have been implicated in promoting inflammation. The primary objective of this report is to describe the strategy adopted for the selection of potentially effective probiotic bacteria. The study further describes the evaluation of two members of the resulting panel of micro-organisms (Lactobacillus salivarius subsp. salivarius UCC118 and Bifidobacterium longum infantis 35624) under in vitro conditions and throughout in vivo murine and human feeding trials. Specifically, an initial feeding study completed in Balb/c mice focused upon (i) effective delivery of the probiotic micro-organisms to the GIT and evaluation of the ability of the introduced strains to survive transit through, and possibly colonise, the murine GIT; (ii) accepting the complexity of the hostile GIT and faecal environments, development of a method of enumerating the introduced bacterial strains using conventional microbiological techniques; and (iii) assessment of the effects of administered bacterial strains on the numbers of specific recoverable indigenous bacteria in the murine GIT and faeces. Additional research, exploiting the availability of murine models of inflammatory bowel disease, demonstrated the beneficial effects of administering probiotic combinations of Lactobacillus salivarius UCC118 and Bifidobacterium longum infantis 35624 in prevention of illness-related weight loss. A further ethically-approved feeding trial, successfully conducted in 80 healthy volunteers, demonstrated that yoghurt can be used as a vehicle for delivery of Lactobacillus salivarius strain UCC118 to the human GIT with considerable efficacy in influencing gut flora and colonisation.

Animals↗