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Biomedical subjects

R A Robbins

Publications and source records attributed to R A Robbins.

At least 73 records · Page 4Linked to original sources
PubMed1995

Glucocorticoid inhibition of RANTES expression in human lung epithelial cells.

An influx of eosinophils into the lungs occurs in several pulmonary disorders. However, the mechanisms involved remain unknown. Lung epithelial cell release of eosinophil chemotactic factors such as RANTES or macrophage inflammatory protein-1 alpha (MIP-1 alpha) could account for the influx of eosinophils into the lungs. In order to demonstrate the potential role for lung epithelial cells to release RANTES and/or MIP-1 alpha, we investigated the mRNA expression and protein release in cultured A549 cells. Tumor necrosis factor-alpha (TNF alpha) and interleukin-1 beta (IL-1 beta) induced a time- and dose-dependent increase in RANTES mRNA expression and protein release. In contrast, MIP-alpha protein release was not detectable in these cells. As corticosteroids decrease the influx of eosinophils into the lungs in vivo, we also investigated the capacity of dexamethasone to decrease the TNF alpha-induced RANTES release and mRNA expression; both were decreased in a time- and concentration-dependent manner. Dexamethasone did not affect the TNF alpha-induced RANTES mRNA half-life and did not require protein synthesis to manifest an inhibitory effect. Supernatant from cells stimulated with TNF alpha and IL-1 beta increased eosinophil chemotaxis and this was also inhibited by dexamethasone. These findings suggest a role for RANTES release by lung epithelial cells in the recruitment of eosinophils into the lungs in pulmonary disorders such as interstitial lung diseases, idiopathic pulmonary fibrosis, or asthma and suggest that one beneficial effect of corticosteroids may be inhibition of lung epithelial cell RANTES mRNA expression and protein release.

Cell Line↗
PubMed1995

Calcium and protein kinase C dependency of lipoxygenase-derived neutrophil chemotactic activity release from bronchial epithelial cells.

In the present investigation, we evaluated the roles of calcium, calcium-calmodulin, inositol turnover, and protein kinase C in the release of lipoxygenase-derived metabolites with neutrophil chemotactic activity (NCA) from bronchial epithelial cells (BECs) in response to endotoxin (ETX) and opsonized zymosan (OZ). Both ETX and OZ stimulated BECs to release NCA [56.9 +/- 5.1 (ETX), 65.2 +/- 5.1 (OZ) versus 15.2 +/- 3.0 (controls) cells/high power field, P < 0.001]. The lipoxygenase inhibitors, nordihydroguaiaretic acid and diethylcarbamazine, and phospholipase A2 inhibitors, mepacrine and dibucaine, blocked the release of NCA in response to ETX, OZ, calcium ionophore A23187 (A23187), and phorbol myristate acetate (PMA). The calcium channel blockers, nifedipine and verapamil, and two putative inhibitors of calcium release from intracellular storage sites, 8-(N,N-diethylamine)-octyl-3,4,5-trimethoxybenzoate hydrochloride and ruthenium red, inhibited the release of NCA induced by ETX but had little effect on the release of NCA induced by OZ. However, depletion of extracellular calcium inhibited the release of NCA in response to both ETX and OZ. Calmodulin inhibitors, compound 48/80 and N-(6-aminohexyl)-1 naphthalenesulfonamide (W-7), inhibited the release of NCA in response to a variety of endotoxin concentrations. Lithium chloride, an inositol turnover inhibitor, inhibited the release of NCA in response to both ETX and OZ, but less attenuation was observed in the response to OZ. A protein kinase C inhibitor, dihydrosphingosine, inhibited the release of NCA in response to both ETX and OZ. Finally, A23187 and PMA stimulated the release of NCA and [3H]arachidonic acid independently and additively. These data suggest that the release of NCA from BECs in response to OZ may be predominantly mediated by inositol turnover and protein kinase C and that the release of NCA in response to ETX may be regulated by calcium influx and calcium release from intracellular storage sites, calcium-calmodulin activation, inositol turnover, and protein kinase C activation. Protein kinase C augmented the release of NCA and [3H]arachidonic acid independent of and in combination with calcium. These results may suggest the calcium and protein kinase C dependency of the release of NCA from BECs.

Animals↗
PubMed1995

Immunological functions of the pulmonary epithelium.

The mature pulmonary epithelium forms a continuous lining to the airspace. Recent data suggest that this specialized epithelium may also contribute to host defence via interactions with inflammatory cells. Pulmonary epithelial cells can serve as part of the local immune system, providing structures and functions crucial for the maintenance of normal pulmonary function. This article will briefly review the morphology and development of the pulmonary epithelial cells, their function with regard to host defence, alterations of the pulmonary epithelium associated with airway diseases, and potential therapeutic implications for the treatment of respiratory diseases.

Epithelium↗
PubMed1995

Antiproteases modulate bronchial epithelial cell responses to endotoxin.

Escherichia coli endotoxin (0.1 to 1000 micrograms/ml) stimulated the release of neutrophil chemotactic activity (P < 0.001) and induced bronchial epithelial cell (BEC) cytotoxicity assessed by lactate dehydrogenase release (P < 0.001). Endotoxin (100 micrograms/ml) inhibited BEC accumulation (P < 0.001). In the present study, we investigated the role of proteolytic activity of BECs per se in response to endotoxin. Several structurally and functionally different antiproteases, alpha 1 protease inhibitor, soybean trypsin inhibitor, two chloromethyl ketone derivatives (N-tosyl-L-lysine chloromethyl ketone and methoxysuccinyl-Ala-Ala-Pro-Val chloromethyl ketone), and L-658,758, a neutrophil elastase inhibitor, attenuated the release of neutrophil chemotactic activity and lactate dehydrogenase (P < 0.01). alpha 1-Protease inhibitor and N-tosyl-L-lysine chloromethyl ketone attenuated the inhibition of BEC accumulation by endotoxin (P < 0.001). The proteolytic enzyme activity measured by synthetic substrates revealed that endotoxin significantly augmented the serine proteolytic activity in the cell layers. Culture supernatant fluids and cell lysates of BECs in the presence of endotoxin solubilized 14C-labeled casein. These data suggest that responses of BECs to endotoxin may involve activation of cellular proteolytic activity.

Bronchi↗
PubMed1994

Expression of inducible nitric oxide in human lung epithelial cells.

Nitric oxide (NO) is increased in the exhaled air of subjects with several airway disorders. To determine if cytokines could stimulate epithelial cells accounting for the increased NO, the capacity of the proinflammatory cytokines (cytomix: tumor necrosis factor-alpha, interleukin-1 beta, and interferon-gamma) to increase inducible nitric oxide synthase (iNOS) was investigated in A549 and primary cultures of human bronchial epithelial cells. Cytomix induced a time-dependent increase in nitrite levels in culture supernatant fluids (p < 0.05). Increased numbers of cells stained for iNOS and increased iNOS mRNA was detected in the cytokine-stimulated cells compared to control (p < 0.05). Dexamethasone diminished the cytokine-induced increase in nitrite, iNOS by immunocytochemistry, and iNOS mRNA. These data demonstrate that cytokines, such as those released by mononuclear cells, can induce lung epithelial iNOS expression and NO release, and that this is attenuated by dexamethasone.

Amino Acid Oxidoreductases↗
PubMed1994

Inducible nitric oxide synthase is increased in murine lung epithelial cells by cytokine stimulation.

Nitric oxide (NO) is detectable in exhaled air. To elucidate whether airway epithelial cells could be a source of NO, we investigated the expression of inducible nitric oxide synthase (iNOS) by the murine lung epithelial cell line, LA-4, in response to cytokine stimulation and the ability of corticosteroids to modulate this effect. Stimulation with cytomix, a combination of interleukin-1 beta, tumor necrosis factor-alpha, and interferon-gamma, elevated nitrite levels by 873% in the culture supernatants and enhanced the conversion of arginine to citrulline by 273% at 24 h. An increased number of cells stained for iNOS and an increase in iNOS mRNA was also observed. Dexamethasone decreased the cytokine-induced increase in nitrite levels, NOS activity, iNOS immunoreactivity, and mRNA but did not change the half life of iNOS mRNA. These results show that lung epithelial cells can release NO, a process which can be inhibited by dexamethasone.

Amino Acid Oxidoreductases↗
PubMed1994

Substance P increases neutrophil adhesion to bronchial epithelial cells.

Polymorphonuclear neutrophils (PMNs) accumulate within the airways during acute and chronic bronchitis and can adhere to bronchial epithelium. Substance P (SP), a neuropeptide released from the primary afferent nerve endings, has been shown to have a proinflammatory effect on PMNs. To test the hypothesis that SP could modulate PMN bronchial epithelial cell adherence, bovine bronchial epithelial cells (BBECs) were isolated and cultured, and the capacity of SP to modulate PMN-BBEC adherence was evaluated. SP interacted with BBECs to induce an increase in PMN adhesion (14.7 +/- 1.2% vs 5.3 +/- 0.7% adherence, p < 0.01). The effect of SP was both time- and dose-dependent with maximal responses at 6 h and 10(-10) M. The effect was reproduced by the carboxyl-terminal sequence of the molecule (SP 6-11). Importantly, pretreatment of the BBECs with the tachykinin SP receptor (NK1) antagonist, CP-96,345, significantly reduced the increase in adhesion induced by SP (p < 0.01). Furthermore, treatment of the BBECs with antibodies against CD11a (LFA-1), CD11b (MAC-1), or ICAM-1 significantly decreased SP-induced adherence (p < 0.01 all comparisons). Conversely, SP stimulation of PMN induced a dose-dependent, rapid (within 5 min) increase in adherence. This effect was also mediated by the carboxy end of the molecule and was decreased by CP-96,345, again suggesting that this effect was NK1 mediated. These data demonstrate the SP has the capacity for modulating PMN-BBEC interactions and suggest an important role for SP in modulating airway inflammation.

Animals↗
PubMed1994

Increased nitric oxide in exhaled air of asthmatic patients.

Nitric oxide (NO) gas is produced by various cells within the lower respiratory tract, including inflammatory and epithelial cells, and is detectable in the exhaled air of normal human subjects. We have measured exhaled NO in patients with asthma, since several cell types that are activated in asthma can produce NO after induction. NO was measured reproducibly by a slow vital capacity manoeuvre and an adapted chemiluminescence analyser. NO was detectable in exhaled air of 67 control subjects (mean peak concentration 80.2 [SE 4.1] ppb) and was significantly reduced by inhalation of the specific NO synthase inhibitor NG-monomethyl-L-arginine. 61 non-steroid-treated asthmatic subjects had significantly higher peak expired NO concentrations than controls (283 [16] ppb, p < 0.001) but 52 asthmatic patients receiving inhaled corticosteroids had levels similar to controls (101 [7] ppb). High exhaled NO concentrations in asthmatic patients may reflect induction of NO synthase, which is known to be inhibited by steroids. Measurement of exhaled NO concentrations may be clinically useful in detection and management of cytokine-mediated inflammatory lung disorders.

Adrenal Cortex Hormones↗
PubMed1994

Neutrophil chemotaxis to extracts of grain plant components.

Exposure to grain dust has been associated with a neutrophilic bronchitis. In vitro studies suggest several possible mechanisms for recruitment of neutrophils, including direct chemotaxis to aqueous grain dust extracts and release of chemotactic activity by bronchial epithelial cells in response to challenge with grain dust extracts.

Animals↗
PubMed1994

Corticosteroids as adjunctive therapy for diffuse alveolar hemorrhage associated with bone marrow transplantation. University of Nebraska Medical Center Bone Marrow Transplant Group.

BACKGROUND: Diffuse alveolar hemorrhage is a frequent complication of treating malignancies with high-dose chemotherapy and bone marrow transplantation and is associated with very high mortality. This disorder's association with pulmonary inflammation, its coincidence with marrow recovery, and the usefulness of corticosteroids for treating other pulmonary hemorrhage syndromes provided the rationale for this study. METHODS: We retrospectively studied 65 episodes of diffuse alveolar hemorrhage that has occurred in 63 of 603 consecutively treated patients who had undergone high-dose chemotherapy with bone marrow transplantation. Patients were divided into three groups according to the therapy they had received for diffuse alveolar hemorrhage: supportive therapy alone (n = 12); low-dose corticosteroids (30 mg or less of methylprednisolone or its equivalent; n = 10); and high-dose corticosteroids (more than 30 mg methylprednisolone or its equivalent; n = 43). The primary outcome measures were overall survival and survival to hospital discharge, occurrence of respiratory failure requiring intubation, and development of infections subsequent to the diagnosis of diffuse alveolar hemorrhage. RESULTS: Overall survival at the end of the follow-up period was significantly higher for the high-dose corticosteroid group compared with the supportive therapy group (P = 0.005); however, treatment with low-dose steroids did not increase survival over supportive therapy alone (P = 0.198). In addition, survival to discharge was significantly increased for the high-dose group compared with the other two groups combined (33% versus 9.1%, P = 0.038). Respiratory failure after the diagnosis of diffuse alveolar hemorrhage developed in only 12 of the 22 unintubated patients in the high-dose group compared with 9 of the 10 initially unintubated patients in the other two groups (P = 0.056). Although the incidence of infections was high (40%) subsequent to diffuse alveolar hemorrhage, neither high-dose nor low-dose corticosteroid treatment significantly increased the risk of infections (P > 0.4, all comparisons). CONCLUSIONS: In this study, high-dose corticosteroid therapy for diffuse alveolar hemorrhage related to bone marrow transplantation was associated with improved total survival and survival to hospital discharge, and decreased development of respiratory failure in these patients. These results suggest the therapy is beneficial, and further prospective studies are warranted to verify the effectiveness of the treatment.

Adult↗
PubMed1994

Induction of cyclo-oxygenase-2 by cytokines in human pulmonary epithelial cells: regulation by dexamethasone.

1. Cyclo-oxygenase metabolizes arachidonic acid to prostaglandin H2 (PGH2) and exists in at least two isoforms. Cyclo-oxygenase-1 (COX-1) is expressed constitutively whereas COX-2 is induced by lipopolysaccharide (LPS) and some cytokines in vitro and at the site of inflammation in vivo. Epithelial cells may be an important source of prostaglandins in the airways and we have, therefore, investigated the expression of COX-1 or COX-2 isoforms in primary cultures of human airway epithelial cells or in a human pulmonary epithelial cell line (A549). 2. COX-1 or COX-2 protein was measured by western blot analysis using specific antibodies to COX-2 and selective antibodies to COX-1. The activity of COX was assessed by the conversion of either endogenous or exogenous arachidonic acid to four metabolites, PGE2, PGF2 alpha, thromboxane B2 or 6-oxo PGF1 alpha measured by radioimmunoassay. Thus, COX-1 or COX-2 activity was measured under two conditions; initially the accumulation of the COX metabolites formed from endogenous arachidonic acid was measured after 24 h. In other experiments designed to measure COX activity directly, cells were treated with cytokines for 12h before fresh culture medium was added containing exogenous arachidonic acid (30 microM) for 15 min after which COX metabolites were measured. 3. Untreated primary cells or A549 cells contained low amounts of COX-1 or COX-2 protein. Bacterial LPS (1 micro g ml-1 for 24 h) induced COX-2 protein in the primary cells, a process which was enhanced by interferon-gamma, with no further increase in the presence of a mixture of cytokines (interleukin-1 beta, tumour necrosis factor-alpha and interferon-gamma, 10 ng ml-1 for all). In contrast, A549 cells contained only low levels of COX-2 protein after exposure to LPS or LPS plus interferon-y, but contained large amounts of COX-2 protein after exposure to the mixture of cytokines.4. Untreated human pulmonary primary cells or A549 cells released low levels of all COX metabolites measured over a 24 h incubation period. This release was enhanced by treatment of either cell type with the mixture of cytokines (interleukin-1 beta , tumour necrosis factors- and interferon-gamma, 10 ng ml-1 for all).PGE2 was the principal COX metabolite released by cytokine-activated epithelial cells. The release of PGE2 induced by cytokines occurred after a lag period of more than 6 h.5. The glucocorticosteroid, dexamethasone (1 micro M; 30 min prior to cytokines) completely suppressed the cytokine-induced expression of COX-2 protein and activity in both primary cells and A549 cells.6. In experiments where COX-2 activity was supported by endogenous stores of arachidonic acid,treatment of A549 cells with interleukin-l beta but not tumour necrosis factor a or interferon-gamma alone caused a similar release of PGE 2 to that seen when the cytokines were given in combination. However, both interleukin-l beta and necrosis factor- alone produced similar increases in COX-2 activity (measured in the presence of exogenous arachidonic acid) as seen when the mixture of interleukin-l beta, tumour necrosis factor- alpha and interferon-gamma were used to stimulate the cells.7. These findings show that COX-2 expression correlates with the exaggerated release of prostaglandins from cytokine-activated human pulmonary epithelial cells and that the induction of the enzyme is suppressed by a glucocorticosteroid. These findings may be relevant to inflammatory diseases of the lung, such as asthma.

Arachidonic Acid↗
PubMed1994

Peptidases modulate bradykinin-induced arteriolar dilation in the hamster cheek pouch.

The purpose of this study was to investigate whether angiotensin-converting enzyme (ACE; EC 3.4.15.1) and neutral endopeptidase (NEP; EC 3.4.24.11), two membrane-bound metalloenzymes that are widely distributed in the peripheral microcirculation and degrade kinins very effectively, modulate bradykinin-induced arteriolar dilation in vivo. Using intravital microscopy, we measured diameter of second-order arterioles in the hamster cheek pouch during suffusion of bradykinin (0.1-10.0 microM) before and after topical application of captopril (10.0 microM) and phosphoramidon (10.0 nM). We found that each inhibitor significantly potentiated bradykinin-induced increase in arteriolar diameter (P < 0.05). Suffusion of other proteinase inhibitors (excluding ACE and NEP inhibitors) had no significant effect on bradykinin-induced responses. Captopril and phosphoramidon did not potentiate isoproterenol (0.1 microM)-induced arteriolar dilation in the cheek pouch. Collectively, these data indicate that ACE and NEP each plays an important role in regulating bradykinin-induced vasorelaxation in the peripheral microcirculation in vivo.

Animals↗
PubMed1994

Role of nitric oxide and superoxide anions in interleukin-1 beta-induced airway hyperresponsiveness to bradykinin.

We investigated the role of endogenous nitric oxide (NO) and superoxide anions in recombinant human interleukin-1 beta (rhIL-1 beta)-induced bronchial hyperresponsiveness (BHR) and neutrophilia in Brown-Norway rats. Aminoguanidine (100 mg/kg/d) administered subcutaneously for 3 d, an inhibitor of inducible NO synthase, L-arginine (100 mg/kg/d administered subcutaneously for 3 d, a specific precursor for the synthesis of NO, and apocynin (5 mg/kg/orally), an inhibitor of superoxide anion (O2-)-generating NADPH oxidase in macrophages and neutrophils, were administered prior to administration of rhIL-1 beta (500 U) intratracheally. Aminoguanidine in addition to another inhibitor of NO synthase, NW-nitro-L-arginine methyl ester (L-NAME) 100 mg kg/d administered subcutaneously for 3 d augmented bronchial responsiveness to inhaled bradykinin (BK) but not to acetylcholine (ACh), an effect reversed by L-arginine. rhIL-1 beta-treated rats also demonstrated BHR to BK but not to ACh, associated with neutrophilia in bronchoalveolar lavage fluid (BALF). rhIL-1 beta-induced BHR and neutrophilia were neither further increased by aminoguanidine nor inhibited by L-arginine. Apocynin, however, significantly inhibited rhIL-1 beta-induced BHR but not the BALF neutrophilia. Suppression of NO generation and generation of O2- from macrophages and infiltrating neutrophils may be important in rhIL-1 beta-induced airway hyperresponsiveness to bradykinin.

Acetophenones↗
PubMed1994

Glucocorticoids blunt neutrophil CD11b surface glycoprotein upregulation during cardiopulmonary bypass in humans.

Neutrophil-endothelial adhesion is the initiating event in neutrophil migration to areas of infection or injury. The binding of neutrophils to endothelium depends upon adhesive glycoproteins, of which the CD11/CD18 glycoproteins are the most important. Because of known upregulation of one of these adhesive glycoproteins (CD11b) during cardiopulmonary bypass (CPB) in humans, we evaluated CD11a, CD11b, and CD11c surface expression before, during, and after CPB in humans, with or without pre-CPB administration of a glucocorticoid (methylprednisolone). Fourteen patients were randomized into two groups: Group S received methylprednisolone (1 g intravenously) 5 min prior to CPB; Group N received no steroid. CD11b was significantly upregulated (P < 0.01) during, and 24 h after, CPB in Group N when compared with controls and Group S at similar time intervals, while in Group S no significant changes were found. Since interleukin-1, tumor necrosis factor, and endotoxin are known to upregulate neutrophil CD11b surface expression and are released during CPB in humans, while steroids are known to suppress the release of these cytokines, the authors conclude that the blunting effect by steroids on CD11b surface expression upregulation during and after CPB in humans is attributed to suppressed cytokine release.

Antigens, CD↗
PubMed1994

Impairment of nasal mucociliary clearance during bone marrow transplantation. University of Nebraska Medical Center bone Marrow Transplantation Pulmonary Study Group.

Transient mucositis occurs in almost all patients receiving bone marrow transplantation (BMT) but it is not known whether airway mucociliary transport is impaired during this period. We hypothesized that the preparative regimen associated with BMT impairs mucociliary clearance during the peri-transplant period. To test this hypothesis we determined nasal saccharine transit time (STT) and a mucositis score daily throughout the peri-transplant hospitalization in 13 patients receiving BMT. STT was prolonged in all patients at some time during the peri-transplant period and mucociliary clearance was ineffective in 12 of 13 patients for an average of 22% of the days. STT was most prolonged (about 155% of baseline) in the 2 week period following marrow infusion and correlated with the appearance of mucositis. STT returned to baseline values within 30 days after marrow infusion in all but one patient. These findings demonstrate that nasal mucociliary clearance is significantly impaired in most patients during BMT, if only temporarily. This decrease in local airway host defense may explain, in part, the increased risk of upper respiratory infections that these patients experience.

Adult↗