Biomedical subjects

C A Ryan

Publications and source records attributed to C A Ryan.

At least 55 records · Page 3Linked to original sources

PubMed2000

The systemin signaling pathway: differential activation of plant defensive genes.

Systemin, an 18-amino-acid polypeptide released from wound sites on tomato leaves caused by insects or other mechanical damage, systemically regulates the activation of over 20 defensive genes in tomato plants in response to herbivore and pathogen attacks. Systemin is processed from a larger prohormone protein, called prosystemin, by proteolytic cleavages. However, prosystemin lacks a signal sequence and glycosylation sites and is apparently not synthesized through the secretory pathway, but in the cytoplasm. The polypeptide activates a lipid-based signal transduction pathway in which the 18:3 fatty acid, linolenic acid, is released from plant membranes and converted to the oxylipin signaling molecule jasmonic acid. A wound-inducible systemin cell surface receptor with an M(r) of 160,000 has recently been identified. The receptor regulates an intracellular cascade including, depolarization of the plasma membrane, the opening of ion channels, an increase in intracellular Ca(2+), activation of a MAP kinase activity and a phospholipase A(2) activity. These rapid changes appear to play important roles leading to the intracellular release of linolenic acid from membranes and its subsequent conversion to jasmonic acid, a potent activator of defense gene transcription. Although the mechanisms for systemin processing, release, and transport are still unclear, studies of the timing of the synthesis and of the intracellular localization of wound- and systemin-inducible mRNAs and proteins indicates that differential syntheses of signal pathway genes and defensive genes are occurring in different cell types. This signaling cascade in plants exhibits extraordinary analogies with the signaling cascade for the inflammatory response in animals.

Amino Acid Sequence↗

PubMed2000

Acute animal studies of the STARFlex system: a new self-centering cardioSEAL septal occluder.

The STARFlex system is a modified CardioSEAL device with a flexible self-centering mechanism comprised of nitinol springs strung between opposing arms, a connecting ball (sleeve joint that allows the device to pivot prerelease), and a front-loading delivery system. It was designed to allow a smaller device/defect sizing ratio and delivery profile, provide centering capability, and improve closure rates. To test this system, 13 devices (23, 28, and 33 mm) were deployed in six sheep within created atrial septal defects (12- to 22-mm diameter; n = 10), in the left atrium (n = 2), and in inferior vena cava (n = 1). All implantations in atrial septal defects were successful, with device/defect ratio ranging from 1.3 to 1.9 (median, 1.3), with no residual leak by angiography or echocardiography in seven (3/10 had </= small immediate leaks). The STARFlex system was effective in closing created atrial septal defects using a 10 Fr delivery sheath and low device/defect sizing ratios, comparing favorably with the standard CardioSEAL. Cathet. Cardiovasc. Intervent. 49:225-233, 2000.

Animals↗

PubMed2000

Sexually transmitted diseases in Morocco: gender influences on prevention and health care seeking behavior.

Increased awareness of the medical and social costs of sexually transmitted diseases (STD) has resulted in greater attention to the control of these illnesses. STDs are responsible for a significant amount of morbidity in Morocco and have become a key target of the HIV control program. In 1996, the Ministry of Health conducted a qualitative study in order to enhance information, education and communication strategies in the national STD/HIV program. Data on the conceptualization and knowledge of STD, information sources and health-care-seeking behavior were gathered through 70 semidirected, in-depth interviews conducted with men and women in the general population and health care providers (HCPs). Two commonly applied health behavior theories in STD/HIV prevention, the Health Belief Model (HBM) and the Theory of Reasoned Action (TRA) served as a framework for data analysis. The most common name for STD is berd, which means "the cold" in Moroccan Arabic. Berd is caused either by cold striking the genital area or sexual intercourse and most often designates a syndrome of genital discharge. However, the term was also often used to indicate STD in general. The dual causality of berd maintains social stability by providing an honorable excuse for individuals who become infected, while warning against unsanctioned sexual behavior. Clear gender differences in understanding STDs and health-care-seeking behavior emerged through these interviews. STDs in Morocco are viewed as women's illnesses and men with STD often reported feeling victimized by women. Men appear to have more extensive informal information sources for STD than women. Consequences of STD, both physical and psychosocial, were viewed as more severe for women than men, and men had greater access to treatment, for both social and economic reasons.

Condoms↗

PubMed2000

The importance of exposure estimation in the assessment of skin sensitization risk.

The development of new ingredients and products for the consumer market requires a thorough assessment of their potential for skin sensitization and the possible clinical manifestation of allergic contact dermatitis. The process by which low molecular weight chemicals induce and elicit skin sensitization reactions is complex and dependent on many factors relevant to the ability of the chemical to penetrate the skin, react with protein, and trigger the cell-mediated immune response. These major factors include inherent potency, chemical dose, duration and frequency of exposure, vehicle or product matrix, and occlusion. The fact that a chemical is a contact allergen does not mean that it cannot be formulated into a consumer product at levels well tolerated by most individuals. Many common ingredients (e.g., fragrances, preservatives) are known skin allergens. However, all allergens show dose-response and threshold characteristics. Therefore, one should be able to incorporate these chemicals into products at levels that produce acceptably low incidences of skin sensitization under foreseeable conditions of exposure. The critical exposure determinant for evaluating skin sensitization risk is dose per unit area of skin exposed. Use of this parameter allows for comparative assessments from different types of skin sensitization tests (including cross-species comparisons), and, at least for known potent allergens, there is remarkable similarity in threshold dose/unit area determinations across species. The dose/unit area calculation enables a judgment of the sensitization risk for different product types. This is illustrated using the chemical preservative methylchloroisothiazolinone/methylisothiazolinone (MCI/MI) as a case study.

Animals↗

PubMed2000

Use of the local lymph node assay for the estimation of relative contact allergenic potency.

The effective toxicological evaluation of skin sensitization demands that potential contact allergens are identified and that the likely risks of sensitization among exposed populations assessed. By definition, chemicals which possess the toxicological property of skin sensitization potentially are capable of causing allergic contact dermatitis (ACD) in humans. However, this hazard is not an all-or-none phenomenon; clear dose-response relationships can be discerned and thresholds identified for both the induction of sensitization and the elicitation of contact dermatitis. Commonly, these parameters are grouped under the heading of potency, determination of which is vital for risk assessment. In the present investigation, the local lymph node assay (LLNA) has been employed to determine the relative potency of a range of 20 chemicals. The parameter used is the estimated concentration required to produce a 3-fold increase in draining lymph-node cell proliferative activity, the EC3 value. These measurements have been compared with an assessment of the human sensitizing potency of the 20 selected chemicals, each being assigned to 1 of 5 classes based on their human sensitizing potency. The EC3 value, derived from LLNA work carried out in acetone/ olive oil vehicle, correlated well with the human classification, with the strongest sensitizers having low EC3 values (<O.1%), weaker sensitizers having EC3 values generally in the 1-10% range, and non-sensitizing chemicals having EC3 values in excess of 100%. In conclusion, the derivation of the EC3 for a chemical provides an objective and quantitative estimate of potency that is of considerable utility for skin sensitization risk assessment.

Allergens↗

PubMed2000

Activity of human contact allergens in the murine local lymph node assay.

The murine local lymph node assay (LLNA) is a predictive test for the identification of chemicals that have the potential to cause skin sensitization. Since its original development, the assay has been the subject of national and international evaluation studies and extensive comparisons with guinea pig tests and human data. On the basis of these investigations, the LLNA has recently been endorsed by ICCVAM (Interagency Coordinating Committee on the Validation of Alternative Methods) as a stand-alone method for skin sensitization hazard identification. At the same time, ICCVAM confirmed that, although the LLNA is not an in vitro method, it does represent a refinement in the way animals are used and can provide a means for reducing the number of animals used in sensitization hazard assessment. The investigations described here were designed to explore further the ability of the LLNA to identify accurately those chemicals that cause allergic contact dermatitis in humans. To that end we have measured, amongst 3 independent laboratories, LLNA responses induced by a total of 18 test chemicals, 11 of which are known to cause skin sensitization and 7 of which are believed not to be associated with any significant evidence of allergic contact dermatitis in humans. The LLNA correctly classified 16 of the 18 materials. The 11 chemicals tested which are associated with allergic contact dermatitis in humans were found to be positive in the LLNA. Of the 7 materials believed to be non-sensitizers, 5 were negative in the LLNA and 2 produced positive results. Collectively, these data provide additional evidence that the LLNA is able to discriminate skin sensitizers from those chemicals which do not possess a significant skin sensitization potential and thus provides a method for hazard identification that offers important animal welfare benefits.

Allergens↗

PubMed2000

Effects of magnesium sulfate in a newborn piglet meconium aspiration model.

OBJECTIVE: To examine the hemodynamic effects of magnesium sulfate (MgSO4) in a neonatal model of meconium aspiration syndrome. DESIGN: A prospective animal study. PATIENTS AND PARTICIPANTS: Eight anesthetized neonatal piglets. METHODS: Animals were instrumented under fentanyl anesthesia for the determination of pulmonary and systemic blood pressures and cardiac index. A model of meconium aspiration was produced by instilling 6 to 7 ml of 20% fresh human meconium in normal saline down the endotracheal tube, resulting in hypoxemia (mean arterial O2 saturation 73 SD, 10%), respiratory acidosis (mean pH 7.08, SD 0.04; PaCO2 89, SD 2 torr), and pulmonary arterial hypertension (mean pulmonary artery pressure 52 SD, 5 mm Hg). A total of four aliquots of MgSO4 (25 mg/kg equivalent to 0.1 mM/kg of magnesium) were administered intravenously to five animals (total of 100 mg/kg MgSO4). Three animals served as controls and did not receive MgSO4. RESULTS: Each dose of MgSO4 was associated with a transient decrease in heart rate, cardiac index, pulmonary and systemic arterial pressures, and SaO2, followed by a partial gradual recovery of all these variables within 2 to 3 minutes. According to measurements taken after MgSO4 therapy, there was a significant decrease in arterial saturations (from 73 SD (10%) to 63 SD (13%); p < 0.05), mean systemic blood pressure (from 77 SD (16 mm Hg) to 74 SD (15 mm Hg); p < 0.05) and pulmonary artery pressure (to 52 SD (5 mm Hg); p < 0.05). The ratio between pulmonary and systemic vascular resistance was not significantly affected by MgSO4 therapy. There were no changes in the control animals. CONCLUSION: The administration of magnesium sulfate is associated with increased hypoxemia in this model of meconium aspiration syndrome, probably because of worsening ventilation perfusion mismatch. These data do not provide any physiological support for the use of MgSO4 in the neonate with meconium aspiration and pulmonary hypertension and suggest that this agent should be used cautiously, if at all, in such situations.

Animals↗

PubMed2000

Local lymph node assay: validation assessment for regulatory purposes.

For the prediction of skin sensitization potential of substances, the murine local lymph node assay (LLNA) is an alternative to the widely used guinea pig tests. For more than 10 years, this method has undergone extensive development, evaluation, and validation. In this review, the validation status of the LLNA is considered, specifically with regard to its use for regulatory identification of skin sensitization hazards. The LLNA is a method for the predictive identification of chemicals that have a potential to cause skin sensitization. Activity is measured as a function of lymph node cell proliferative responses stimulated by topical application of test chemicals. The LLNA has successfully passed all reasonable validation stages. It provides a reliable and relevant source of predictive skin sensitization data, which unlike results from guinea pig tests, are reproducible from laboratory to laboratory. In summary, the LLNA is now ready for acceptance as a viable and complete alternative to traditional methods, offering a substantial reduction in animal numbers and refinement opportunities without compromising the standards for the identification of important skin sensitizers.

Allergens↗

PubMed2000

UVB/UVA radiation activates a 48 kDa myelin basic protein kinase and potentiates wound signaling in tomato leaves.

We investigated the effect of UV radiation on early signaling events in the response of young tomato plants (Lycopersicon esculentum) to wounding. Ultraviolet-C (< 280 nm) and UVB/UVA (280-390 nm) radiation both induced 48 kDa myelin basic protein kinase activity in leaves. The activation was associated with phosphorylation of tyrosine residues on the kinase, which is indicative of protein kinases of the mitogen-activated protein kinase family. Ultraviolet-C irradiation resulted in a strong proteinase inhibitor synthesis, as reported previously (Conconi et al., Nature 383, 826-829, 1996). Under the conditions used, UVB/UVA radiation did not induce proteinase inhibitor synthesis but resulted in a strong potentiation of systemic proteinase inhibitor synthesis in response to wounding. The UVB/UVA-irradiated plants that were subsequently wounded accumulated 2.5-4-fold higher levels of proteinase inhibitor I when compared to wounded non-irradiated plants. The potentiating effect was most prominent in the systemic unwounded leaf of a wounded plant. Levels of 12-oxo-phytodienoic acid and jasmonic acid that have been well documented to increase in response to wounding were not detected in response to UVB/UVA irradiation alone. The effect of UVB/UVA radiation in potentiating plant defense signaling should be further considered as a factor that may influence the ecological balance between plants and their predators.

Calcium-Calmodulin-Dependent Protein Kinases↗

PubMed2000

Trend analysis and socio-economic differentials in infant mortality in the Southern Health Board, Ireland (1988-1997).

The objective of this paper is to examine trends in infant mortality (IMR) and low-birth weight (LBW) in the Southern Health Board (SHB) area and to investigate the relationship between IMR and social and economic factors. IMR data were obtained from the Central Statistics Office (CSO) over a 10-year period (1988-1997). Social deprivation was measured using the Small Area Health Research Unit (SAHRU) Deprivation Index, which consists of five census-based indicators; unemployment, low social class, car ownership, rented accommodation and overcrowding. During the 10 years, 556 infants died, 380 deaths (68%) occurring in the neonatal period and 176 (32%) in the post-neonatal period. There was a downward trend in IMR throughout the study period, from 11.3 in 1988 to 4.0 in 1997 (p < 0.001). The IMR in Cork City was higher than the rate in the SHB area as a whole (p = 0.0001). Congenital anomalies accounted for 34% of neonatal deaths. Sudden infant death syndrome (SIDS) and congenital anomalies accounted for 45% (79/176) and 22% (39/176) of postnatal deaths, respectively. Only 7% of all district electoral division (DED's) in the SHB were classed in the most deprived SAHRU Deprivation Index category (level 5). Yet, almost half (43%) of the DED's in Cork City were level 5 compared to only 1% in Cork County and 2% in Kerry. Despite significant overall improvements in infant mortality in the SHB, infants born into the lower socio-economic area (Cork City) continue to experience higher relative risks of mortality in comparison with those born in the higher socio-economic areas. Further investigation into the apparently divergent socio-economic patterns of infant mortality within the SHB is necessary.

Cause of Death↗

PubMed1999

A method for distinguishing 1-acyl from 2-acyl lysophosphatidylcholines generated in biological systems.

Phospholipases A(1) and A(2) frequently coexist in biological systems. Generation of lysophosphatidylcholine (LPC) in such systems cannot be assigned to any of these types of enzymes unless the position of the fatty acid in the lysocompound can be unambiguously determined. We here present a simple method to achieve this purpose. It is based on the initial chemical acylation of the isolated LPC with a labeled fatty acid, followed by the enzymatic analysis of the resulting phosphatidylcholine (PC), using snake or bee venom phospholipase A(2). Thus, if treatment of the PC with this enzyme releases a labeled free fatty acid, it is demonstrated that the initial LPC was acylated at position sn-1, whereas if the product of hydrolysis yields labeled LPC, then the initial LPC was acylated at position sn-2. This is the first method devised to determine the source of LPC in the presence of mixtures of phospholipases A(1) and A(2) in complex biological systems.

Acylation↗

PubMed1999

Proteinase inhibitor-inducing activity of the prohormone prosystemin resides exclusively in the C-terminal systemin domain.

Prosystemin is the 200-amino acid precursor of the 18-amino acid polypeptide defense hormone, systemin. Herein, we report that prosystemin was found to be as biologically active as systemin when assayed for proteinase inhibitor induction in young tomato plants and nearly as active in the alkalinization response in Lycopersicon esculentum suspension-cultured cells. Similar to many animal prohormones that harbor multiple signals, the systemin precursor contains five imperfect repetitive domains N-terminal to a single systemin domain. Whether the five repetitive domains contain defense signals has not been established. N-terminal deletions of prosystemin had little effect on its activity in tomato plants or suspension-cultured cells. Deletion of the C-terminal region of prosystemin containing the 18-amino acid systemin domain completely abolished its proteinase inhibitor induction and alkalinization activities. The apoplastic fluid from tomato leaves and the medium of cultured cells were analyzed for proteolytic activity that could process prosystemin to systemin. These experiments showed that proteolytic enzymes present in the apoplasm and medium could cleave prosystemin into large fragments, but the enzymes did not produce detectable levels of systemin. Additionally, inhibitors of these proteolytic enzymes did not affect the biological activity of prosystemin. The cumulative data indicated that prosystemin and/or large fragments of prosystemin can be active inducers of defense responses in both tomato leaves and suspension-cultured cells and that the only region of prosystemin that is responsible for activating the defense response resides in the systemin domain.

Journal Article↗

PubMed1999

Hydrogen peroxide is generated systemically in plant leaves by wounding and systemin via the octadecanoid pathway.

Hydrogen peroxide (H2O2) generated in response to wounding can be detected at wound sites and in distal leaf veins within 1 hr after wounding. The response is systemic and maximizes at about 4-6 hr in both wounded and unwounded leaves, and then declines. The timing of the response corresponds with an increase in wound-inducible polygalacturonase (PG) mRNA and enzyme activity previously reported, suggesting that oligogalacturonic acid (OGA) fragments produced by PG are triggering the H2O2 response. Systemin, OGA, chitosan, and methyl jasmonate (MJ) all induce the accumulation of H2O2 in leaves. Tomato plants transformed with an antisense prosystemin gene produce neither PG activity or H2O2 in leaves in response to wounding, implicating systemin as a primary wound signal. The antisense plants do produce both PG activity and H2O2 when supplied with systemin, OGA, chitosan, or MJ. A mutant tomato line compromised in the octadecanoid pathway does not exhibit PG activity or H2O2 in response to wounding, systemin, OGA, or chitosan, but does respond to MJ, indicating that the generation of H2O2 requires a functional octadecanoid signaling pathway. Among 18 plant species from six families that were assayed for wound-inducible PG activity and H2O2 generation, 14 species exhibited both wound-inducible PG activity and the generation of H2O2. Four species, all from the Fabaceae family, exhibited little or no wound-inducible PG activity and did not generate H2O2. The time course of wound-inducible PG activity and H2O2 in Arabidopsis thaliana leaves was similar to that found in tomato. The cumulative data suggest that systemic wound signals that induce PG activity and H2O2 are widespread in the plant kingdom and that the response may be associated with the defense of plants against both herbivores and pathogens.

Hydrogen Peroxide↗

PubMed1999

A wound- and systemin-inducible polygalacturonase in tomato leaves.

Oligogalacturonide fragments that activate defensive genes in plant leaves heretofore have been thought to be generated only by pathogen-derived pectin-degrading enzymes, because polygalacturonase (PG) activity has not been reported in leaves. Here, we report that mRNAs encoding a PG catalytic subunit protein and its regulatory (beta-subunit) protein are expressed in tomato leaves in response to wounding, systemin, and oligosaccharide elicitors. Synthesis of the two subunits in response to wounding is systemic and is accompanied by an increase in PG activity in extracts from both wounded and unwounded leaves. The finding that PG subunit mRNAs and PG enzyme activity are induced by wounding indicates that herbivore attacks can produce endogenous oligogalacturonide elicitors that may be involved in the local and systemic activation of defense responses against both herbivores and pathogens.

Amino Acid Sequence↗

PubMed1999

Local lymph node assay: differentiating allergic and irritant responses using flow cytometry.

The murine local lymph node assay (LLNA) is a method for assessing the contact sensitization potential of chemicals. Based on events that occur during the induction phase of a contact sensitization response, the LLNA measures the in vivo proliferation of cells in the draining lymph nodes (DLNs) of mice following topical exposure to chemicals. In terms of predictive identification of important skin sensitizers, the LLNA has been shown to be at least as sensitive as, and much more reliable than, current guinea pig tests. However, proliferation has also been observed following treatment with some irritants. In an attempt to distinguish allergic from irritant-induced proliferation, flow cytometric techniques have been used to examine the phenotype of lymphocyte subsets in the DLNs as well as markers of T-lymphocyte activation and memory. Mice were treated on the ears for 3 consecutive days with allergens or irritants. The DLNs were harvested 72 h after the final treatment. Single-cell suspensions were prepared, counted, and stained for analysis of the percentages of T cells and B cells and T-cell expression of two adhesion molecules that have been associated with differentiating naïve and activated/memory T cells, CD62L (L-selectin) and CD44 (H-cam). Increases in lymph node cellularity were observed in both allergen- and irritant-treated mice relative to naïve and vehicle-treated animals. Mice treated with allergens showed a preferential increase in the percentage of B220(+) B cells compared with irritant-treated mice. Treatment with allergens, but not irritants, resulted in a selective increase in the percentages of CD4(+) and CD8(+) cells expressing the T-cell activation/memory phenotype CD62L(lo)CD44(hi). Taken together, flow cytometric analysis of cell phenotype and expression of T-cell activation/memory markers may provide important information for differentiating allergen- and irritant-induced proliferative responses in the DLNs of chemically treated mice.

Allergens↗

PubMed1999

The expression of tomato prosystemin in Escherichia coli: A structural challenge.

Prosystemin is the 200-amino-acid prohormone of the 18-amino-acid polypeptide called systemin, a systemic mobile signal that activates the synthesis of defense genes in solanaceous plants in response to herbivore attacks. The unusual primary structural features of the tomato prosystemin cDNA and protein provided an extraordinary challenge in devising an expression system to obtain the full-length protein. Prosystemin expression inhibited the growth of a eukaryotic and several prokaryotic hosts used. Prosystemin was initially synthesized as a truncated protein of 185 amino acids in length using a T7 RNA polymerase expression system in E. coli strain BL21[DE3]. The truncation was found to be due to two factors: (1) the intramolecular associations of the 5' coding region of the prosystemin sequence with the expression vector's ribosome binding site and (2) the presence of a translation start site just prior to the amino acid methionine at position 15. Mutations that permitted the synthesis of the full-length prosystemin protein were introduced into the amino-terminal 5' coding region of the prosystemin cDNA. A 199-amino-acid recombinant prosystemin lacking the N-terminal methionine was purified from lysates and confirmed by N-terminal amino acid sequence and immunoblot analysis.

Amino Acid Sequence↗

PubMed1999

Preparation/analysis of chromatin replicated in vivo and in isolated nuclei.

This article outlined biochemical methodologies for the labeling, detection, and analysis of newly replicated and newly assembled nucleosomes. The isolation of specific vertebrate factors that may be involved in chromatin assembly in vivo, such as nucleoplasmin, CAF-1, and NAP-1 and their counterparts in Drosophila and yeast add a further dimension to the study of nucleosome assembly in living cells. In particular, the ability to genetically manipulate the yeast system, together with the identification of yeast enzymes that acetylate newly synthesized H4, will certainly provide exciting new avenues for the investigation of chromatin assembly in vivo.

Animals↗

PubMed1999

Wound- and systemin-inducible calmodulin gene expression in tomato leaves.

Using a calmodulin (CaM) cDNA as a probe in northern analyses, transgenic tomato plants that overexpress the prosystemin gene were found to express increased levels of CaM mRNA and protein in leaves compared to wild-type plants. These transgenic plants have been reported previously to express several wound-inducible defense-related genes in the absence of wounding. Calmodulin mRNA and protein levels were found to increase in leaves of young wild-type tomato plants after wounding, or treatment with systemin, methyl jasmonate, or linolenic acid. CaM mRNA appeared within 0.5 h after wounding or supplying young tomato plants with systemin, and peaked at 1 h. The timing of CaM gene expression is similar to the expression of the wound- or systemin-induced lipoxygenase and prosystemin genes, signal pathway genes whose expression have been reported to begin at 0.5-1 h after wounding and 1-2 h earlier than the genes coding for defensive proteinase inhibitor genes. The similarities in timing between the synthesis of CaM mRNA and the mRNAs for signal pathway components suggests that CaM gene expression may be associated with the signaling cascade that activates defensive genes in response to wounding.

3' Untranslated Regions↗