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Biomedical subjects

C J Walsh

Publications and source records attributed to C J Walsh.

At least 19 recordsLinked to original sources
PubMed2002

Dexamethasone-induced apoptosis in immune cells from peripheral circulation and lymphomyeloid tissues of juvenile clearnose skates, Raja eglanteria.

Juvenile clearnose skates (Raja eglanteria) were injected intramuscularly with dexamethasone-21-phosphate at 50, 75, and 100mg/kg body weight. After 24h, skates were sacrificed and lymphomyeloid tissues (thymus, spleen, Leydig organ, and epigonal organ) were removed and whole blood was sampled. Tissues were used fresh for imprints or prepared for histology by solvent fixation or freezing in liquid nitrogen. Apoptosis in fixed tissues was assessed by transmission electron microscopy. Frozen sections and cytospin preparations of peripheral blood leukocytes (PBL) were evaluated by the TUNEL reaction to detect DNA strand breaks. Dexamethasone treatment increased apoptotic activity in all lymphomyeloid tissues as well as in PBL. These studies demonstrate that immune cells of elasmobranchs have the capacity for glucocorticoid-driven apoptosis, and that programmed cell death as a mechanism to regulate immune cell production appears to have been conserved during vertebrate evolution.

Animals↗
PubMed2001

Complex expression patterns of lymphocyte-specific genes during the development of cartilaginous fish implicate unique lymphoid tissues in generating an immune repertoire.

Cartilaginous fish express canonical B and T cell recognition genes, but their lymphoid organs and lymphocyte development have been poorly defined. Here, the expression of Ig, TCR, recombination-activating gene (Rag)-1 and terminal deoxynucleosidase (TdT) genes has been used to identify roles of various lymphoid tissues throughout development in the cartilaginous fish, Raja eglanteria (clearnose skate). In embryogenesis, Ig and TCR genes are sharply up-regulated at 8 weeks of development. At this stage TCR and TdT expression is limited to the thymus; later, TCR gene expression appears in peripheral sites in hatchlings and adults, suggesting that the thymus is a source of T cells as in mammals. B cell gene expression indicates more complex roles for the spleen and two special organs of cartilaginous fish-the Leydig and epigonal (gonad-associated) organs. In the adult, the Leydig organ is the site of the highest IgM and IgX expression. However, the spleen is the first site of IgM expression, while IgX is expressed first in gonad, liver, Leydig and even thymus. Distinctive spatiotemporal patterns of Ig light chain gene expression also are seen. A subset of Ig genes is pre-rearranged in the germline of the cartilaginous fish, making expression possible without rearrangement. To assess whether this allows differential developmental regulation, IgM and IgX heavy chain cDNA sequences from specific tissues and developmental stages have been compared with known germline-joined genomic sequences. Both non-productively rearranged genes and germline-joined genes are transcribed in the embryo and hatchling, but not in the adult.

Animals↗
PubMed1999

Meniscus regeneration in a rabbit partial meniscectomy model.

Meniscectomy is known to be associated with osteoarthrosis of the knee. The purpose of this study was to compare the natural and augmented repair of menisci in the knees of New Zealand White rabbits. To create a partial defect in the medial meniscus, we used an experimental model that has been well characterized and extensively used in the study of osteoarthrosis and articular cartilage repair. The defect was left untreated or treated with one of the following: a periosteal autograft, a type I collagen sponge, or the same sponge loaded with autologous, bone marrow-derived, cultured mesenchymal stem cells. The natural repair was always incomplete and degenerative changes within these joints were progressive. The periosteal autograft underwent differentiation into a bone and hyaline cartilage composite that was ineffectual as a meniscus and accelerated the degenerative changes in those joints when compared to natural repair controls. There was evidence of a consistent sequence of events in the transformation of the periosteal grafts to a core of cartilage that underwent endochondral ossification. In the last two groups, the collagen sponge functioned as a scaffold that resulted in more abundant repair tissue. The collagen sponge alone supported a largely fibrous repair process. The cultured mesenchymal stem cells were observed to augment the repair process in some specimens to include fibrocartilage histologically similar to normal meniscus. Degenerative changes were present in both of these groups, which indicates that the biomechanical function of the meniscus was not restored, or an irreversible osteoarthrosis cascade was initiated during the repair period. Based on these preliminary studies, further investigation of cell-based meniscus regeneration appears to be warranted.

Animals↗
PubMed1999

Cloning, sequencing, and nucleolar targeting of the basal-body-binding nucleolar protein BN46/51.

BN46/51 is an acidic protein found in the granular component of the nucleolus of the amebo-flagellate Naegleria gruberi. When Naegleria amebae differentiate into swimming flagellates, BN46/51 is found associated with the basal body complex at the base of the flagella. In order to determine the factors responsible for targeting BN46/51 to a specific subnucleolar region, cDNAs coding for both subunits were isolated and sequenced. Two clones, JG4.1 and JG12.1 representing the 46 kDa and 51 kDa subunits, respectively, were investigated in detail. JG12.1 encoded a polypeptide of 263 amino acids with a predicted size of 30.1 kDa that co-migrated with the 51 kDa subunit of BN46/51 when expressed in yeast. JG4.1 encoded a polypeptide of 249 amino acids with a predicted size of 28.8 kDa that co-migrated with the 46 kDa subunit of BN46/51. JG4.1 was identical to JG12.1 except for the addition of an aspartic acid between positions 94 and 95 of the JG12.1 sequence and the absence of 45 amino acids beginning at position 113. The predicted amino acid sequences were not closely related to any previously reported. However, the sequences did have 26-31% identity to a group of FKPBs (FK506 binding proteins) but lacked the peptidyl-prolyl cis-trans isomerase domain of the FKBPs. Both subunits contained two KKE and three KKX repeats found in other nucleolar proteins and in some microtubule binding proteins. Using 'Far Western' blots of nucleolar proteins, BN46/51 bound to polypeptides of 44 kDa and 74 kDa. The 44 kDa component was identified as the Naegleria homologue of fibrillarin. BN46/51 bound specifically to the nucleoli of fixed mammalian cells, cells which lack a BN46/51 related polypeptide. When the JG4.1 and JG12.1 cDNAs were expressed in yeast, each subunit was independently targeted to the yeast nucleolus. We conclude that BN46/51 represents a unique nucleolar protein that can form specific complexes with fibrillarin and other nucleolar proteins. We suggest that the association of BN46/51 with the MTOC of basal bodies may reflect its role in connecting the nucleolus with the MTOC activity for the mitotic spindle. This would provide a mechanism for nucleolar segregation during the closed mitosis of Naegleria amebae.

3T3 Cells↗
PubMed1996

Laparoscopic common bile duct exploration: evolution of a new technique.

Laparoscopic exploration of the common bile duct (CBD) with a choledochoscope or a stone basket during laparoscopic cholecystectomy was attempted in 60 patients and was successful in 56. The cystic duct was used for entry to the CBD in 46 patients and in 14 a choledochotomy was performed. Of 51 patients with confirmed common duct stones, 38 had complete laparoscopic clearance (75 per cent). In 13 patients the duct was not cleared or was only partially cleared, of whom four went on to have clearance by postoperative percutaneous choledochoscopy down a cystic duct or T-tube track. Two patients with cystic duct tubes passed their remaining stones spontaneously. One patient had open exploration and six required endoscopic retrograde cholangiopancreatography. Of nine patients without stones, choledochoscopy was impossible in three patients whose cholangiogram was later considered to be normal. In five patients stones were excluded by choledochoscopy and in one patient laparoscopic choledochoscopy was undertaken to better define abnormal biliary anatomy; this helped to avoid major bile duct injury. Choledochoscopy was easier with the smaller 3.6-mm ureteroscope but stone removal was more difficult when the basket was too small for the stones, the cystic duct too small relative to stone size or the number of stones was too great. Successful stone clearance was proportional to the level of effort expended, and was limited by operating time and equipment.

Cholangiography↗
PubMed1996

The effects of olfactory and somatosensory desensitization on Fos-like immunoreactivity in the brains of pup-exposed postpartum rats.

Fos-like immunoreactivity (fos-lir) was examined in sites within the "maternal circuit" in postpartum female rats that received various sensory desensitizations and were exposed to pups for 1 or 2 hr. Neither olfactory bulbectomy nor thelectomy (nipple removal) significantly reduced the fos-lir in the anterior medial preoptic area (MPOA), although reductions following bulbectomy in medial amygdala did occur. Peripherally induced hyposmia by ZnSo4 reduced fos-lir in the olfactory structures (olfactory bulbs, piriform cortex, and olfactory tubercle), in medial and cortical nuclei of the amygdala, but not in anterior MPOA. Application of the topical anesthetic Emla to the ventrum only reduced fos-lir in the somatosensory cortex. Combined olfactory and ventral desensitizations produced marginal reductions in posterior MPOA. It is suggested that the MPOA is primarily involved as part of the effector system in the expression of the behavior. In contrast, the amygdala is involved in processing sensory cues received from pups during dam-litter interactions.

Afferent Pathways↗
PubMed1996

Incidence of third-degree perineal tears in labour and outcome after primary repair.

The incidence and outcome of third-degree tears following 16,583 vaginal deliveries were prospectively assessed over a 5.5-year period. Ninety-three deliveries (0.56 per cent) were complicated by a third-degree tear and the patients underwent primary repair. Eighty-one patients were reviewed 3 months postpartum at a colorectal clinic. Third-degree tears were significantly more common in primigravidae and mothers with higher birth-weight babies. They were significantly associated with the use of forceps and were not prevented by episiotomy. Of the 81 patients reviewed, 30 had an abnormal anorectal examination. Six patients (7 per cent) were incontinent of faeces. A further ten (12 per cent) were incontinent of flatus only. The overall incidence of faecal incontinence was 0.04 per cent. An important group of women with significant subclinical sphincter injury was identified. Obstetric trauma causes significant anorectal dysfunction and patients with third-degree tears require assessment by a colorectal specialist.

Adult↗
PubMed1996

Evaluation of p53 protein expression in renal cell carcinoma: comparison of two antibodies.

The aims of this study were to establish a profile for the expression of p53 in primary renal cell carcinoma using the polyclonal antibody NCL-CB1 and the monoclonal antibody D07, and to compare the results of staining with both antibodies. Ninety-six cases were studied using formalin-fixed paraffin-embedded tissue. Positive nuclear staining ranged from 5% (D07) to 12% (NCL-CM1). Positive cytoplasmic staining ranged from 7% (D07) to 25% (NCL-CM1) of cases. Interobserver agreement was excellent. The findings suggest that such a low level of immunohistochemical positivity reduces any potential prognostic value for p53 in this tumour type.

Antibodies↗
PubMed1995

Tumor necrosis factor-alpha and interleukin-6 selectively regulate neutrophil function in vitro.

The neutrophil is an important effector cell of the host response to sepsis. Tumor necrosis factor-alpha (TNF-alpha), a cytokine mediator of the septic response, is rapidly released following endotoxemia or gram-negative bacteremia. Interleukin-6 (IL-6) is another cytokine mediator of the host response to sepsis whose role is less well understood than that of TNF-alpha. It is known to be elevated in gram-negative sepsis, where peak levels have been correlated with mortality. This study examined the effect of IL-6 alone and in combination with TNF-alpha on three neutrophil functions--CD18 adhesion receptor expression, phagocytosis, and superoxide anion generation. Neutrophils from human volunteers were incubated with amounts of IL-6 ranging from 10 to 1000 ng/ml. At a concentration of 1000 ng/ml, IL-6 increased neutrophil phagocytosis of opsonized bacteria (826 +/- 255 x 10(3) MESF vs 552 +/- 103 MESF, P < 0.05) and also increased neutrophil superoxide anion generation (18.41 +/- 1.86 vs 12.6 nmol O2-/10(6) PMN/10 min, P < 0.05). Lesser amounts of IL-6 had no effect on phagocytosis or superoxide generation. IL-6 did not increase neutrophil CD18 adhesion receptor expression. Combining IL-6 with TNF-alpha at doses of 100 ng/ml and 100 U/ml, respectively, neutrophil phagocytosis (221 +/- 455 MESF vs 552 +/- 103 MESF) and superoxide generation (23.18 +/- 1.86 vs 12.6 nmol O2-/10(6) PMN/10 min) were significantly (P < 0.05) increased above control by an amount similar to that seen with 1000 U/ml TNF-alpha alone.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

Delayed tumor necrosis factor alpha blockade attenuates pulmonary dysfunction and metabolic acidosis associated with experimental gram-negative sepsis.

OBJECTIVE: To ascertain the effect of delayed tumor necrosis factor alpha (TNF-alpha) on the evolution of systemic and pulmonary injury after the onset of sepsis. DESIGN: Prospective controlled trial. INTERVENTION: Anesthetized swine were made septic with a 1-hour infusion of live Pseudomonas aeruginosa, following which a treatment group received an infusion of anti-TNF-alpha monoclonal antibody (5 mg/kg). Control animals received 0.9% saline. RESULTS: Delayed anti-TNF-alpha treatment had no effect on septic pulmonary hypertension or decline in cardiac output. Late recovery in systemic arterial hypotension was associated with a reversal of arterial acidosis (P < .05 by t test and analysis of variance with Tukey's Studentized Range Test) compared with unprotected septic animals. Septic animals had a significant increase in mean (+/- SEM) plasma lactate levels at 5 hours compared with baseline values (3.8 +/- 0.7 vs 2 +/- 0.4, P < .05), but remained unchanged from baseline following anti-TNF-alpha treatment (1.5 +/- 0.1 vs 1.6 +/- 0.2, not significant). Characteristic septic neutropenia was dramatically reversed by anti-TNF-alpha treatment and was associated with downregulation (P < .05 by t test and analysis of variance) of polymorphonuclear neutrophil (PMN) leukocyte CD18 adhesion receptors and reduction (P < .05 by t test and analysis of variance) in lung PMN sequestration measured by myeloperoxidase activity. The mean (+/- SEM) decrease in bronchoalveolar lavage protein indicated an attenuated permeability injury in anti-TNF-alpha animals (septic animals at 5 hours compared with baseline value, 1044 +/- 270 vs 149 +/- 28 micrograms/mL; control animals at 5 hours compared with baseline value, 217 +/- 83 vs 129 +/- 19 micrograms/mL; P < .05 by t test and analysis of variance). CONCLUSIONS: These data show that delayed anti-TNF-alpha treatment reversed metabolic acidosis associated with sepsis. Furthermore, anti-TNF-alpha treatment reversed septic neutropenia, reduced PMN sequestration, and was associated with attenuated lung injury in a model of fulminant sepsis. This supports evidence of PMN-mediated tissue injury in sepsis and suggests mechanisms for potential therapeutic benefit of anti-TNF-alpha treatment in clinical practice.

Acidosis, Lactic↗
PubMed1994

Multi-agent therapy in the treatment of sepsis-induced microvascular injury.

Cyclo-oxygenase inhibition (with ibuprofen) combined with histamine (H1, H2) receptor antagonism (with diphenhydramine and cimetidine) attenuates microvascular leak injury in sepsis syndromes. Ibuprofen reduces microvascular injury by limiting oxygen radical production by neutrophils. Histamine is known to inhibit this oxygen radical production, an effect antagonized by cimetidine. In the present study neutrophils isolated from pigs made septic with Pseudomonas organisms exhibited a significant (P < 0.05) increase in the production of the oxygen radicals, superoxide anion (O2-, 133 per cent) and hypochlorous acid (HOCl, 38 per cent). Ibuprofen used alone attenuated this sepsis-stimulated overproduction. Addition of the antihistamines cimetidine and diphenhydramine produced a significant increase in oxygen radical production (P < 0.05), by 122 per cent (O2-) and 47 per cent (HOCl), equivalent to that in untreated septic animals. This coincided with a significant deterioration in pulmonary compliance (P < 0.05) compared with that found in control animals and those treated with ibuprofen alone, and a significant accumulation of extravascular lung water (P < 0.05) at 240 and 300 min versus baseline. Histamine receptor antagonism may inadvertently enhance microvascular injury in sepsis.

Animals↗
PubMed1994

Monoclonal antibody to tumor necrosis factor-alpha attenuates plasma interleukin-6 levels in porcine gram-negative sepsis.

This study examined the kinetics of IL-6 release into the systemic circulation in a porcine model of bacterial sepsis induced by infusion of live Pseudomonas aeruginosa. Three groups of animals were studied. Group I (n = 12) animals received a 1 hr infusion of live P. aeruginosa. Group II (n = 6) animals received monoclonal antibody to tumor necrosis factor-alpha (TNF-alpha) (15 mg/kg) prior to induction of sepsis. Group III (n = 7) animals received sterile saline only. TNF-alpha and interleukin-6 (IL-6) levels rose sharply, in group I following pseudomonas infusion. Following a peak at 120 min after the bacterial infusion (4.8 +/- 0.7 U/ml at 120 min vs 0.4 +/- 0.2 U/ml at 0 min), TNF-alpha levels subsequently declined prior to the end of the experiment. In contrast, IL-6 levels rose sharply, subsequent to TNF-alpha, peaked at 180 min, and remained significantly elevated throughout the study period (5.3 +/- 0.9 ng/ml vs 0.05 +/- 0.01 ng/ml, 0 min). In animals pretreated with monoclonal antibody to TNF-alpha, no increase in TNF-alpha activity was detected at any time during the period of study. IL-6 levels in antibody-treated animals, although greatly attenuated, still rose significantly above baseline (2.02 +/- 0.8 ng/ml at 180 min vs 0.05 +/- 0.01 ng/ml at 0 min) and above levels in control animals. We conclude that although TNF-alpha plays an important role in synthesis and release of IL-6, there is a TNF-alpha-independent pathway for release of IL-6 in sepsis.

Animals↗
PubMed1994

Differential activation of alveolar, pulmonary arterial, and systemic arterial neutrophils demonstrates the existence of distinct neutrophil subpopulations in experimental sepsis.

Neutrophils (PMNs) are considered key cellular mediators of sepsis induced acute lung injury. PMN activation is manifest by increased beta 2 integrin expression and enhanced superoxide radical (O2-) generation. What is unclear is at which anatomical sites PMNs are activated and at which sites they release O2- and mediate lung injury. In this study we compared alveolar (ALV), systemic arterial (SA), and pulmonary arterial (PA) PMNs CD18 receptor expression, measured by fluorescent immunophenotyping and, O2- generation, measured by reduction of ferricytochrome C, in septic swine. Swine were anesthetized and ventilated, and given a 1-h infusion of live Pseudomonas aeruginosa. PA, SA, and ALV PMNs were isolated at 0 and 5 h. ALV PMNs O2- was reduced compared to SA blood PMNs O2- at 5 h, (AIV 5 h 23.6 +/- 3 vs. SA 0 h 34.3 +/- 5, p < .05). SA PMNs O2- generation was also significantly reduced compared to PA PMNs at 5 h (PA 5 h 21 +/- 2.5 vs. SA 5 h 16.9 +/- 2.6, p < .05). Alv PMNs expressed significantly greater CD18 receptor levels than SA blood PMNs at 5 h (AIV PMNs 5 h, 76 +/- 6 vs. SA PMNs 5 h 51 +/- 3, p < .05), however, PA PMNs CD18 receptor levels were not significantly different from SA PMNs levels at 5 h. These data corroborate a dissociation between two PMN functions in sepsis. O2- generation was reduced across the lung and following migration. However, alveolar PMNs had significantly upregulated CD18 expression compared to PMNs in PA and SA.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

C-erbB-2 immunostaining: problems with interpretation.

AIMS: To assess the consistency and reproducibility of assessment of c-erbB-2 immunostaining, and to examine some of the problems relating to inter- and intraobserver variability in the documentation of positive staining; to profile the spectrum of cytoplasmic and membranous staining in a wide range of tumour types. METHODS: A total of 283 neoplasms were examined for immunohistochemical expression of the c-erbB-2 oncoprotein. Three independent observers were required to assess intensity both of membrane and cytoplasmic staining on a three point and then a four point scale. Extent of positive staining was also assessed on a two point scale. A minimum of two weeks elapsed between assessments using the differing scales. RESULTS: Positive membrane staining was documented by one or more observers in 16.6% of tumours examined. This positivity was largely restricted to bladder, renal, and breast tumours. The overall level of disagreement as to the presence or absence of membranous staining was 11.3%. Cytoplasmic staining was identified in 55.5% of tumours studied. The level of disagreement as to the presence or absence of cytoplasmic staining was 26.5%. CONCLUSIONS: Intraobserver variability was minimal, indicating that each pathologist was adhering to internal reproducible standards. Interobserver variability was greater, indicating that the interpretation of c-erbB-2 immunostaining may require set guidelines. It is suggested that assessment should be referenced to a standard positive control, that a three tier system for grading of intensity and a two tier system for grading of extent should be adopted, and that the evaluation should be agreed by at least two pathologists. The presence of cytoplasmic staining should continue to be routinely recorded until its biological role and clinical implications are fully understood.

Biomarkers, Tumor↗