Biomedical subjects

M Crowley

Publications and source records attributed to M Crowley.

At least 37 records · Page 2Linked to original sources

PubMed1994

The importance of fixation procedures on DNA template and its suitability for solution-phase polymerase chain reaction and PCR in situ hybridization.

Conventional solution-phase polymerase chain reaction (PCR) and in situ PCR/PCR in situ hybridization are powerful tools for retrospective analysis of fixed paraffin wax-embedded material. Amplification failure using these techniques is now encountered in some centres using archival fixed tissues. Such 'failures' may not only be due to absent target DNA sequences in the tissues, but may be a direct effect of the type of fixative, fixation time and/or fixation temperature used. The type of nucleic acid extraction procedure applied will also influence amplification results. This is particularly true with in situ PCR/PCR in situ hybridization. To examine these effects in solution-phase PCR, beta-globin gene was amplified in 100 mg pieces of tonsillar tissue fixed in Formal saline, 10% formalin, neutral buffered formaldehyde, Carnoy's Bouin's, buffered formaldehyde sublimate, Zenker's, Helly's and glutaraldehyde at 0 to 4 degrees C, room temperature and 37 degrees C fixation temperatures and for fixation periods of 6, 24, 48 and 72 hours and 1 week. DNA extraction procedures used were simple boiling and 5 days' proteinase K digestion at 37 degrees C. Amplified product was visible primarily yet variably from tissue fixed in neutral buffered formaldehyde and Carnoy's, whereas fixation in mercuric chloride-based fixatives produced consistently negative results. Room temperature and 37 degrees C fixation temperature appeared most conducive to yielding amplifiable DNA template. Fixation times of 24 and 48 hours in neutral buffered formaldehyde and Carnoy's again favoured amplification.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗

PubMed1994

PCR in situ hybridisation detection of HPV 16 in fixed CaSki and fixed SiHa cell lines.

AIMS: To investigate the feasibility of using fixed cells with the polymerase chain reaction (PCR) in situ hybridisation and to investigate possible reasons for reaction failure. METHODS: Fixed SiHa and CaSki cells were used in an experimental model of PCR in situ hybridisation for the detection of low and intermediate copy number viral infection in fixed cells. RESULTS: PCR in situ hybridisation was able to detect one to two copies of human papillomavirus (HPV) 16 in SiHa cells, using small fragment amplicons (120 base pairs), confirming the high detection sensitivity and flexibility of the technique. Problems were encountered with localisation of PCR amplified product in CaSki cells (200-300 copies of HPV 16 per cell) owing to diffusion of product post amplification. Overall, 40% of reactions were successful, which confirms the current unreliability of the technique. Within cell preparations, about 50% of cells contained amplified product. CONCLUSION: PCR in situ hybridisation represents the marriage of two revolutionary molecular pathological techniques. However, it is currently unreliable, with reaction failure common. Standardised, dedicated equipment is urgently required if the technique is to achieve universal acceptance. In the future, the technique may be used to detect chromosomal translocations in human tumours and to study cellular gene expression.

Base Sequence↗

PubMed1993

Oral cancer in Ireland, 1984-1988.

Age standardised oral cancer incidence rates were calculated for the Republic of Ireland from data collected by the Southern Tumour Registry for the years 1984-1988. The annual incidence rates (per 100,000) for men were lip 3.8, tongue 1.2 and mouth 1.8 while the corresponding rates for women were lip 0.2, tongue 0.7 and mouth 0.5. The incidence of lip cancer is shown to be much lower than indicated by previously published Irish rates. In general, oral cancer incidence rates in Ireland are comparable to those reported for Denmark. The vast majority (94%) of cases were histologically verified. The results, along with recent findings on mortality, contradict the view that Ireland has a very high rate of oral cancer by west European standards.

Adenocarcinoma↗

PubMed1993

Epstein-Barr virus in normal, pre-malignant, and malignant lesions of the uterine cervix.

AIM--To detect the presence or absence of Epstein-Barr virus (EBV) in cervical lesions ranging from normality to invasive malignancy. METHODS--Eighteen randomly selected cases of invasive squamous cell carcinomas of the uterine cervix were examined as well as 25 cases each of normal cervices and those showing cervical intra-epithelial neoplasia (CIN) I, II, and III. DNA-DNA in situ hybridisation, using a biotinylated probe to the Bam H1 "W" fragment of EBV, was carried out in addition to the polymerase chain reaction using specific primer sequences that flank a 153 base pair segment of the Bam H1 "W" region of the EBV genome and which do not cross-amplify other DNA herpes viruses. Positive control material included paraffin wax embedded P3 HR1 lymphoblastoid cells (containing high copy numbers of EBV) and two nasopharyngeal carcinomas positive for EBV. RESULTS--Neither normal nor CIN I tissue was positive. Eight per cent of CIN II tissue was positive; 8% of CIN III, and 43% of carcinomas were positive for EBV. CONCLUSION--The study shows that the virus is present in some cases of cervical carcinoma and to a lesser degree in some premalignant lesions of the cervix, but the exact association between it and cervical oncogenesis, be it causative or incidental, remains to be determined.

Base Sequence↗

PubMed1993

Evidence for cigarette smoking as a major risk factor for periodontitis.

The role of smoking as a risk factor for periodontitis was assessed separately in diabetic and nondiabetic study groups. Subject listings stratified for age (19 to 40 years) and sex were obtained for subjects with insulin-dependent diabetes mellitus (IDDM) and nondiabetic subjects. For both the IDDM group (n = 132) and the nondiabetic group (n = 95), age and sex stratified samples were constructed by random selection of subjects from each subject listing. Patients were recruited by phone, examined, and their medical and dental histories obtained. Among nondiabetic subjects, the prevalence of periodontitis was markedly higher among current smokers compared with never smokers (P < or = 0.005) in both the 19 to 30 year-old (46% vs. 12%) and 31 to 40 year-old groups (88% vs. 33%). The subject mean percent of sites with gingival pocket depth > or = 4 mm was higher among current smokers than never smokers (P = 0.001) in the 19 to 30 (8.2% vs. 3.4%) and 31 to 40 (14.3% vs. 4.3%) age groups. The effects of smoking among IDDM subjects were similar to that observed in the nondiabetic population. There were no differences between current and never smokers in the proportion of sites positive for plaque. Attributable risk percents from prevalence data suggest that among nondiabetic subjects, a large proportion, perhaps as much as 51% of the periodontitis in the 19 to 30 year old group and 32% of the periodontitis in the 31 to 40 year old group, is associated with smoking.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1992

Magnetic resonance imaging of the rat brain following acute carbon monoxide poisoning.

Magnetic resonance (MR) may be used for repeatedly and non-invasively imaging the brain. Until now, no studies have used this approach to study the effects of carbon monoxide (CO) poisoning in a defined animal model. Conscious, Levine-prepared female rats (unilateral carotid artery and jugular vein occlusion) were exposed to 2400 ppm CO for 90 min, with or without the infusion of 50% glucose solution; CO-stimulated increases in blood glucose and lactate occurred in both groups, while blood pressure and body temperature fell. One to four hours following termination of CO exposure, increased cortical pixel intensity, cortical surface area and brain midline shift were observed on the operated side of the brain in some rats of both groups (i.e. responders = R), providing evidence of edema. At sacrifice, 5 h following termination of CO exposure, gross water content was increased on the left side in the corresponding cortical slices in R rats, providing another measure of edema. Significant positive correlations were found between left to right pixel intensity difference and water content difference, and between the extent of midline shift and water content difference. The elevations of blood glucose and lactate concentrations, and the magnitudes of CO-induced hypothermia and hypotension were similar to those in past studies, but appeared to exert no effect on the severity of cortical edema in terms of differences in pixel intensity, surface area, midline shift or gross tissue water content. Thus, the observed differences between the R rats is not explained by the available data.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Living longer and better than expected. A wellness-based model keeps CCRC residents active, healthy, and out of nursing homes.

The McCauley, a continuing care retirement community in West Hartford, CT, promotes a wellness philosophy that helps its residents retain their health and independence and extend their lives. The wellness program centers around the individual, not around the services provided. The residents have significant input and initiative in maintaining their own health. The health program is based on a nursing model, with each resident relating to a primary nurse as needs become evident. The nurses' goal is to know each person's holistic needs--to assist in maintaining physical, emotional, and spiritual balance. The program's approach to physical health requires a strong emphasis on prevention, including individualized fitness programs. The McCauley fosters mental and social well-being by allowing patients to maintain independent living for as long as they are able and by maximizing their self-direction. Another contributor to mental well-being is the opportunity for personal growth and intellectual stimulation. The McCauley promotes spiritual well-being by supporting continued participation in churches or synagogues from previous neighborhoods, fostering a supportive network within the community, and recognizing the value of residents' volunteer activities.

Aged↗

PubMed1992

Epilepsy and employment, marital, education and social status.

Employment, marital, educational and social status in epileptic patients attending a seizure clinic were assessed. A total of 343 patients were evaluated. The social group, employment and marital status did not compare favourably with the overall population. Forty per cent of patients belonged to social group five and six compared with 14% of the population. Thirty three per cent of male patients aged 20 years or more were married, compared with 65% in the community. The corresponding statistics for females were 46% and 73% respectively. The male unemployment rate of 34% compared poorly with the unemployment rate of 13% for the period during which the study data was collected. Patients with lower educational attainments were significantly more likely to have poor seizure control (P less than 0.001), or required polypharmacy. A similar situation was found between seizure control and social status, with poor seizure control occurring in patients in social group six.

Adolescent↗

PubMed1991

Evidence for a contribution of paramagnetic ions to water proton spin-lattice relaxation in normal and malignant mouse tissues.

Paramagnetic ions complexed to proteins may lose, retain, or enhance solvent paramagnetic relaxation (SPR) relative to free solution. We measured T1 and T2 of three mouse cancers, their normal counterparts, and six additional tissues. Long T1 of cancers was not caused by necrosis or by different contents of water, fat, or blood. Dissociable (TCA-extractable) and nondissociable (ashed) Mn, Cu, and Fe were measured by AA. Cancers had less Mn, Cu, and Fe than did normal counterparts. All 12 tissues had inverse correlations between T1 and dissociable Mn and Cu. For Mn alone to account for reduced T1, the extent to which SPR of the Mn-protein complexes would be enhanced is by factors of 0.6 to 13, below the maximum observed in Mn-enzymes. Different amounts of paramagnetic ion-protein complexes may account for part of the differences in T1 of water protons in different tissues, and the longer T1 of cancer cell water may be caused in part by reduced amounts of such complexes.

Animals↗

PubMed1990

Dendritic cells are the principal cells in mouse spleen bearing immunogenic fragments of foreign proteins.

We monitored the APC function of cells taken from the spleen and peritoneal cavity of mice that had been given protein antigens via the intravenous or intraperitoneal routes. Using the mAb 33D1 and N418 to negatively and positively select dendritic cells, we obtained evidence that dendritic cells are the main cell type in spleen that carries the protein in a form that is immunogenic for antigen-specific T cells. In vivo pulsed macrophages were not immunogenic and did not appear capable of transferring peptide fragments to dendritic cells.

Animals↗

PubMed1990

Comparative behavioral, neurochemical and pharmacological activities of dihydropyridine calcium channel activating drugs.

The behavioral (deficits in motor function in mice), neurochemical (affinity for mouse brain membrane dihydropyridine receptors, effects on neurotransmitter/metabolite levels in mice) and pharmacologic (effect on the contractile activity of guinea pig ileal longitudinal smooth muscle) properties of the calcium channel activators (+/-)-BAY K 8644, (+/-)-202-791 (and their corresponding channel activating and antagonist enantiomers) and CGP-28392 were investigated and compared. The calcium channel activating enantiomers (-)-S-BAY K 8644, (+)-S-202-791 and (+/-)-BAY K 8644, (+/-)-202-791 and CGP-28392 produced a dose-dependent impairment of rotarod ability and decreases in motor activity in mice with the following order of potency: (-)-S-BAY K 8644 greater than (+/-)-BAY K 8644 much greater than (+)-S-202-791 greater than (+/-)-202-791 = CGP-28392. The calcium channel antagonists (+)-R-BAY K 8644 and (-)-R-202-791 were behaviorally inactive but blocked the behavioral effects of (-)-S-BAY K 8644. The binding of dihydropyridine calcium channel activator and antagonist enantiomers to mouse brain membranes was described by both one and two site models. (-)-S-BAY K 8644, (+/-)-BAY K 8644, (+)-S-202-791 and CGP-28392 produced contractions in partially depolarized (15 mM K+) strips of guinea pig ileal longitudinal smooth muscle which differed in the degree of maximum contraction obtained. (+)-R-BAY K 8644 and (-)-R-202-791 inhibited potassium-induced contractions (80 mM K+) in guinea pig ileal longitudinal smooth muscle.(ABSTRACT TRUNCATED AT 250 WORDS)

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1989

A small number of anti-CD3 molecules on dendritic cells stimulate DNA synthesis in mouse T lymphocytes.

Resting T cells enter cell cycle when challenged with anti-CD3 mAb and accessory cells that bear required Fc receptors (FcR). Presentation of anti-CD3 is thought to be a model for antigens presented by accessory cells to the TCR complex. We have obtained evidence that the number of anti-CD3 molecules that are associated with the accessory cell can be very small. We first noticed that thymic dendritic cells and cultured, but not freshly isolated, epidermal Langerhans cells (LC) were active accessory cells for responses to anti-CD3 mAb. DNA synthesis was abrogated by a mAb to the FcR but not by mAb to other molecules used in clonally specific antigen recognition, i.e., class I and II MHC products or CD4 and CD8. The requisite FcR could be identified on the LC but in small numbers. Freshly isolated LC had 20,000 FcR per cell, while the more active cultured LC had only 2,000 sites, using 125I-anti-FcR mAb in quantitative binding studies. Individual LC had similar levels of FcR, as evidenced with a sensitive FACS. FcR could not be detected on T cells or within the dendritic cell cytoplasm, at the start of or during the mitogenesis response. When the response was assessed at 30 h with single cell assays, at least 20 T cells became lymphoblasts per added LC, and at least 8 T cells were synthesizing DNA while in contact with the LC in discrete cell clusters. To the extent that anti-CD3 represents a polyclonal model for antigen presentation to specific T cell clones, these results suggest two conclusions. First, only 200-300 molecules of ligand on dendritic cells may be required to trigger a T cell. Second, the maturation of LC in culture entails "sensitizing" functions other than ligand presentation (anti-CD3 on FcR) to clonotypic T cell receptors.

Animals↗

PubMed1989

Presentation of exogenous protein antigens by dendritic cells to T cell clones. Intact protein is presented best by immature, epidermal Langerhans cells.

The capacity of dendritic cells to present protein antigens has been studied with two MHC class II-restricted, myoglobin-specific, T cell clones. Spleen dendritic cells and cultured epidermal Langerhans cells (LC) presented native myoglobin weakly and often not at all. These same populations were powerful stimulators of allogeneic T cells in the primary MLR. Freshly isolated LC were in contrast very active in presenting proteins to T cell clones but were weak stimulators of the MLR. Both fresh and cultured LC could present specific peptide fragments of myoglobin to the clones. These results suggest that dendritic cells in nonlymphoid tissues like skin can act as sentinels for presenting antigens in situ, their accessory function developing in two phases. First antigens are captured and appropriately presented. Further handling of antigen then is downregulated while the cells acquire strong sensitizing activity for the growth and function of resting T lymphocytes. The potent MLR stimulating activity of cultured epidermal LC and lymphoid dendritic cells probably reflects prior handling of antigens leading to the formation of allogeneic MHC-peptide complexes.

Animals↗

PubMed1989

The cell surface of mouse dendritic cells: FACS analyses of dendritic cells from different tissues including thymus.

The surface of dendritic cells from mouse spleen, thymus, and epidermis has been compared with a panel of monoclonal antibodies and the FACS. A method was first developed to isolate populations of large, adherent, thymic dendritic cells that were greater than 90% pure. These were released by collagenase digestion and separated from adherent macrophages after overnight culture. Enrichment was based on the facts that most macrophages remained plastic adherent and rosetted strongly with antibody-coated erythrocytes. As in spleen, thymic dendritic cells were stellate in shape, had abundant class I and II MHC products, lacked many standard macrophage and lymphocyte markers, and actively stimulated the mixed leukocyte reaction. Most spleen and thymic dendritic cells could be lysed by the 7D4 mAb, to the low-affinity IL-2 receptor, and complement but the levels of 7D4 by FACS were low and sometimes not above background. Differences among dendritic cells from different tissues were noted with other mAb. Adherent dendritic cells from thymus all expressed the J11d "B cell" antigen and the NL145 interdigitating cell marker, but lacked the 33D1 spleen dendritic cell antigen. Eighty to ninety percent of spleen dendritic cells were J11d-, NL145-, 33D1+ but the remainder expressed the J11d+, NL145+, 33D1- thymic phenotype. The latter phenotype also was identical to that of epidermal Langerhans' cells. We postulate that the major 33D1+ cell in spleen represents a migratory stage in which dendritic cells are moving from tissues to lymphoid organs.

Animals↗

PubMed1989

Massive hepatic necrosis in a child after administration of phenobarbital.

A 2-yr-old child developed massive hepatic necrosis caused by an unusual hypersensitivity response to phenobarbital that was prescribed for presumed febrile seizures. Despite discontinuation of the barbiturate, this child experienced fulminant hepatic failure and died. Review of the literature indicates that phenobarbital-induced hepatic injury is uncommon and usually mild. Prompt and permanent cessation of the drug results in resolution of symptoms in most patients.

Chemical and Drug Induced Liver Injury↗

PubMed1988

Macrophages phagocytose thymic lymphocytes with productively rearranged T cell receptor alpha and beta genes.

The thymus gland is important for the formation of competent T lymphocytes. However, there is long-standing evidence that greater than 95% of newly formed thymocytes do not emigrate to peripheral lymphoid tissues but instead die locally. We have identified a rapid and selective pathway for thymocyte turnover in vitro. The mechanism entails binding, uptake, and digestion by macrophages. The susceptible cells are a subpopulation of double-positive thymocytes. These thymocytes can be enriched by virtue of their high buoyant density in Percoll and prove to have low levels of surface CD3 and little or no surface TCR. However TCR-alpha and -beta genes have undergone rearrangement, and full length alpha and beta transcripts are abundant. Therefore many double-positive cells rearrange and express TCR genes but do not have normal levels of TCR on the cell surface. We propose that thymocytes that undergo high turnover in situ are unable to form receptors that can be selected by MHC molecules in the thymus, and that these cells are recognized and cleared by the macrophage.

Gene Rearrangement, T-Lymphocyte↗

PubMed1988

The sensitization phase of T-cell-mediated immunity.

Many cell-mediated immune responses appear to develop in two phases: A sensitization phase in which unprimed or memory T cells interact with dendritic cells to become active lymphoblasts, and an effector phase in which the T lymphoblasts and other presenting cells interact to eliminate the antigen. Antigen presentation is essential to both phases. Here we review several features that are pertinent to the special sensitization role of dendritic cells. First, dendritic cells from lymphoid tissues, blood, and lymph (lymphoid dendritic cells) express very high levels of class I and II MHC products, and these levels cannot be increased by exposure to cytokines such as immune interferon. Second, dendritic cells efficiently cluster antigen-specific T cells during primary responses. Other presenting cells, like macrophages and B lymphocytes, do not form clusters but do bind to sensitized T lymphoblasts. Dendritic-T-cell binding is not inhibited by mAb to CD4 and LFA-1 antigens. It is suggested that a dendritic-cell-specific molecule is required. Third, it is not yet clear if dendritic cells make a "lymphocyte activating factor." However, IL-1 is not produced, even when dendritic cells are in contact with responding T cells. Fourth, dendritic cells have the capacity to migrate from the tissues and move to T-dependent areas. Epidermal Langerhans cells represent a reservoir of tissue dendritic cells but seem to be immunologically immature. The viability and accessory function of the Langerhans cell greatly depend on a single cytokine, granulocyte-macrophage colony stimulating factor (GM-CSF), leading to the proposal that GM-CSF is critical in mobilizing active dendritic cells at the onset of a cell-mediated immune response.

Animals↗

PubMed1987

A comparative study of the relative effects of anticonvulsant drugs and dietary folate on the red cell folate status of patients with epilepsy.

In a survey of the red cell folate status of 200 patients with epilepsy, compared to 72 controls, we found that median red cell folate levels were reduced significantly in patients treated with phenytoin (p less than 0.01) or carbamazepine (p less than 0.001) alone. Patients taking more than one drug had reduced levels also (p less than 0.001), but in patients treated with sodium valproate alone there was no significant decrease in red cell folate levels compared to controls. Twenty-two per cent of patients in the group taking more than one drug had reduced levels of red cell folate compared with 17 per cent of those taking carbamazepine alone, 13 per cent of those taking phenytoin only, and 9 per cent of those taking sodium valproate only. Dietary folate intake was significantly reduced in all the patient groups compared with controls (p less than 0.001 for the carbamazepine and phenytoin groups, p less than 0.01 for the polypharmacy and sodium valproate groups); a significant correlation between red cell folate levels and dietary folate was not established. Significant negative relationships were established between carbamazepine dose (r = -0.35, p less than 0.01) or serum level (r = -0.27, p less than 0.05) and red cell folate level in patients on one drug only. The correlation between dose or serum level of phenytoin and red cell folate level was also negative but did not reach significance. Our findings show that all anticonvulsant drugs interfere with folate metabolism.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗