Biomedical subjects

Judy M Hickman-Davis

Publications and source records attributed to Judy M Hickman-Davis.

11 recordsLinked to original sources

PubMed2006

Surfactant dysfunction in SP-A-/- and iNOS-/- mice with mycoplasma infection.

Surfactant dysfunction was studied in C57BL/6 (B6), B6.SP-A(-/-), and B6.iNOS(-/-) mice with pulmonary mycoplasma infection (10(7) colony-forming units). Cell-free bronchoalveolar lavage (BAL) from uninfected B6.SP-A(-/-) versus B6 mice had a reduced content of very large aggregates (VLA) and an increase in intermediate large aggregates (ILA), with no difference in total large aggregates (LA = VLA + ILA). However, LA from uninfected B6.SP-A(-/-) versus B6 mice contained less protein and were more sensitive to inhibition by serum albumin and lysophosphatidylcholine in pulsating bubble studies in vitro. Infection with Mycoplasma pulmonis caused significant lung injury and surfactant abnormalities in B6.SP-A(-/-), B6.iNOS(-/-), and B6 mice at 24, 48, 72 h after infection compared with uninfected mice of the same strain. Analyses of time-pooled data indicated that mycoplasma-infected B6.SP-A(-/-) and B6.iNOS(-/-) mice had significantly lower levels of LA and higher protein/phospholipid ratios in BAL compared with infected B6 mice. Infected B6.iNOS(-/-) versus B6 mice also had increased minimum surface tensions on the pulsating bubble and decreased levels of surfactant protein (SP)-B in BAL. These results indicate that pulmonary mycoplasma infection in vivo causes lung injury and surfactant abnormalities that are dependent in part on iNOS and SP-A. In addition, SP-A deficiency modifies surfactant aggregate content and lowers the inhibition resistance of LA surfactant in vitro compared with congenic normal mice.

Animals↗

PubMed2006

Bactericidal function of alveolar macrophages in mechanically ventilated rabbits.

Protective ventilation strategies have been universally embraced because of reduced mortality. We tested the hypothesis that tidal volume (VT) in an in vivo model of mechanical ventilation would modulate bactericidal function of alveolar macrophages (AMs). Adult New Zealand White rabbits were mechanically ventilated for 4 h with a VT of 6 ml/kg (low) or a VT of 12 ml/kg (traditional), with each group receiving 3 cm H2O positive end-expiratory pressure with and without intratracheal lipopolysaccharide (LPS) instillation (20 mg/kg). AMs were isolated from bronchoalveolar lavage fluid taken from the whole left lung and used for bacterial killing assays. There were no significant differences in steady-state levels of nitrite or AM phagocytosis and killing of Klebsiella pneumoniae, although these values trended to be slightly higher in the traditional VT group. However, bronchoalveolar lavage fluid protein concentrations were significantly increased in traditional VT groups receiving LPS compared with animals ventilated with a low VT (1,407.8 +/- 121.4 versus 934.7 +/- 118.2; P < 0.001). Lung wet:dry weight ratio in the traditional VT group was increased when compared with the low VT group without LPS (7.3 +/- 0.4 versus 6.1 +/- 0.3, respectively; P < 0.05). Additionally, IL-8 expression was significantly greater under conditions of LPS treatment and mechanical ventilation at VT of 12 ml/kg. These results suggest that the traditional ventilator approach (12 ml/kg VT) in a model of in vivo mechanical ventilation results in lung pathology without affecting AM antibacterial function.

Animals↗

PubMed2005

Leflunomide prevents alveolar fluid clearance inhibition by respiratory syncytial virus.

RATIONALE: Previously, we demonstrated that intranasal infection of BALB/c mice with respiratory syncytial virus (RSV) resulted in an early 40% reduction in alveolar fluid clearance (AFC), an effect mediated via P2Y purinergic receptors. OBJECTIVES: To confirm that RSV-induced inhibition of AFC is mediated by uridine triphosphate (UTP), and to demonstrate that inhibition of de novo pyrimidine synthesis with leflunomide prevents increased UTP release after RSV infection, and thereby also prevents inhibition of AFC by RSV. METHODS: BALB/c mice were infected intranasally with RSV strain A2. AFC was measured in anesthetized, ventilated mice by instillation of 5% bovine serum albumin into the dependent lung. Some mice were pretreated with leflunomide or 6-mercaptopurine. MEASUREMENTS AND MAIN RESULTS: RSV-mediated inhibition of AFC is associated temporally with a 20-nM increase in UTP and ATP content of bronchoalveolar lavage fluid, hypoxemia, and altered nasal potential difference. RSV-mediated nucleotide release, AFC inhibition, and physiologic sequelae thereof can be prevented by pretreatment of mice with the de novo pyrimidine synthesis inhibitor leflunomide, which is not toxic to the mice, and which does not affect RSV replication in the lungs. In contrast, pretreatment of mice with 6-mercaptopurine, an inhibitor of de novo purine synthesis, has no beneficial effect on AFC or other indicators of disease progression. Finally, RSV-mediated inhibition of AFC is prevented by volume-regulated anion channel inhibitors. CONCLUSION: Pyrimidine synthesis or release pathways may provide novel therapeutic targets to counter the pathophysiologic sequelae of impaired AFC in RSV disease.

Animals↗

PubMed2005

Transgenic mice.

For the past 20 years researchers have used transgenic mice to help understand the basic mechanisms associated with inherited human and animal diseases. The ability to integrate exogenous genetic information into the mouse genome has revolutionised the analysis of gene function. Both gene addition and gene replacement can be performed and the capability exists to create 'conditional' mutations and to study gene dosage effects. The aim of the present review is to provide a framework of information on transgenic mouse methodologies that can be applied to any area of research. A basic understanding of transgenic technology, recognising its advantages and disadvantages, is essential knowledge for the scientist in the 21st century.

Animals↗

PubMed2005

Reactive species mediate inhibition of alveolar type II sodium transport during mycoplasma infection.

RATIONALE: Mycoplasma pneumoniae is a significant cause of pneumonia in humans. OBJECTIVES: To determine the impact of mycoplasma infection and the host inflammatory response on alveolar type II (ATII) cell ion transport in vivo and in vitro. METHODS: Mice were infected with M. pulmonis for measurements of alveolar fluid clearance (AFC) in vivo and isolation of ATII cells. ATII cells were infected in vivo for determination of epithelial Na+ channel (ENaC) total and cell surface protein levels by biotinylation and Western blot and in vitro for whole cell patch clamp recording and measurement of nitric oxide (NO) production by chemiluminescence. RESULTS: Mycoplasma infection significantly inhibited AFC at 24 h and total and amiloride-sensitive AFC by 48 h postinfection (pi). In contrast, infected myeloperoxidase-deficient mice had similar basal and amiloride-sensitive AFC values to uninfected control mice at 48 h pi. Addition of forskolin restored total and amiloride-sensitive AFC to control values at 48 h pi. ATII cells isolated from infected mice demonstrated normal alpha, beta, and gamma ENaC total protein levels; however, infected whole-lung cell-surface levels of gamma ENaC were significantly decreased. Patch-clamp recordings demonstrated a significant decrease in total and amiloride-sensitive Na+ currents at 24 h pi. ATII cells demonstrated a significant increase in the production of NO at 24 h pi and inhibition of NO by ATII cells before infection reversed the decrease in total Na+ currents. CONCLUSIONS: These data indicate that mycoplasma infection results in decreased AFC and functional ENaC via the production of reactive oxygen nitrogen intermediates.

Animals↗

PubMed2003

Role of surfactant protein-A in nitric oxide production and mycoplasma killing in congenic C57BL/6 mice.

We generated congenic surfactant protein A (SP-A)-deficient (SP-A[-/-]) mice on the mycoplasma resistant C57BL/6 background (B6.SP-A[-/-]) and characterized their response to mycoplasma infection in comparison to C57BL/6 (B6) mice. B6.SP-A(-/-) mice infected with 10(6) colony-forming units (cfu) of Mycoplasma pulmonis had significantly higher bacterial lung loads than B6 mice at 72 h postinfection (p.i.). At the higher infection dose of 10(7), B6.SP-A(-/-) mice had significantly higher lung cfu at 24 h; however, no difference in mycoplasma cfu was observed between B6 and B6.SP-A(-/-) mice at 48 and 72 h p.i. We found that uninfected B6 mice had lower bronchoalveolar lavage nitrite (NO(2)(-)) and nitrate (NO(3)(-)) levels as compared with B6.SP-A(-/-) mice. On the other hand, infection of B6 mice with mycoplasmas resulted in significantly higher bronchoalveolar lavage NO(2)(-) and NO(3)(-) as compared with B6.SP-A(-/-) mice. These data indicate that SP-A may help regulate NO production in response to a specific stimulus, i.e., suppression of NO in the absence of bacteria and increased NO in the presence of bacteria. These data indicate that the contribution of SP-A to mycoplasma killing may be limited to lower doses of pathogens.

Animals↗

PubMed2003

Nucleotide-mediated inhibition of alveolar fluid clearance in BALB/c mice after respiratory syncytial virus infection.

Respiratory syncytial virus (RSV) is the most common cause of lower respiratory tract disease in infants and children worldwide. Intranasal infection of BALB/c mice with RSV strain A2, but not ultraviolet-inactivated RSV, for 2 or 4 days reduced basal alveolar fluid clearance (AFC), a seminal function of bronchoalveolar epithelium, and caused loss of AFC sensitivity to amiloride inhibition. Reduced AFC was temporally associated with increased lung water content but was not a consequence of increased epithelial permeability or cell death. Reduced AFC was also not due to decreased transcription of epithelial Na+ channel subunit genes in lung tissue. RSV-mediated inhibition of AFC 2 days after infection was rapidly prevented by addition to the instillate of P2Y receptor antagonists (suramin and XAMR-0721) or enzymes that degrade UTP, but not those that degrade ATP. After UTP degradation, AFC returned to control levels but was no longer sensitive to amiloride. UTP at nanomolar concentrations recapitulated the AFC inhibitory effect of RSV in normal mice and mice infected with RSV for 6 days, indicating that normalization of AFC at this time point is a consequence of cessation of UTP release, rather than P2Y receptor desensitization. We conclude that RSV infection of the bronchoalveolar epithelium results in reduced AFC as a consequence of autocrine feedback inhibition mediated by UTP. These studies are the first to demonstrate AFC inhibition by an important pulmonary viral pathogen. Reduced AFC may result in formation of an increased volume of fluid mucus, airway congestion, and rhinorrhea, all features of severe RSV disease.

Animals↗

PubMed2003

Hyperoxia impairs antibacterial function of macrophages through effects on actin.

Oxidative stress may impair alveolar macrophage function in patients with inflammatory lung diseases or those exposed to high concentrations of oxygen. We investigated putative mechanisms of injury to macrophages by oxidative stress, using RAW 264.7 cells exposed to 95% oxygen for 48 h. Hyperoxia-exposed macrophages were less able to phagocytose and kill Klebsiella pneumoniae than normoxic controls, despite increased production of nitric oxide, a free radical important in pathogen killing. Exposure of macrophages to hyperoxia had marked effects on the actin cytoskeleton, including increased actin polymerization, loss of cortical actin, formation of stress fibers, de novo synthesis of actin, and actin oxidation. Hyperoxia induced changes in cell morphology, with increased cell size and pseudopod formation. Exposure of macrophages to jasplakinolide, an agent that increases actin polymerization, also impaired their ability to phagocytose Klebsiella. Alveolar macrophages isolated from mice exposed to 100% oxygen for 84 h also demonstrated impaired phagocytic function, as well as similar effects on the actin cytoskeleton and cell morphology to macrophages exposed to hyperoxia in vitro. We conclude that oxidative stress in vitro and in vivo impairs macrophage antibacterial function through effects on actin.

Actins↗

PubMed2002

Role of innate immunity in respiratory mycoplasma infection.

Mycoplasmas are unique among respiratory pathogens. They possess very small genomes, lack cell walls and are strictly dependent on the host for survival. These pathogens have developed the ability to quickly adapt to the host environment through attachment to target cells within the host. Mycoplasmas have been identified as commensal microbial flora of healthy persons yet, infection of the upper and lower respiratory tracts can result in acute cough, fever and headache, and even chronic disease involving multiple organs. The lung contains a complex system of defense mechanisms with which to combat these pathogens, including innate (nonspecific) and acquired (specific) immune responses. Innate defenses include mechanical clearance, cellular responses provided by host phagocytes and molecular protection in the form of antimicrobial peptides. The interaction of mycoplasmas with different components of the innate immune system and mechanisms by which they incite pathology has proved elusive. The mechanisms by which pathogenic mycoplasmas evade the innate immune system are unknown. The purpose of this review is to summarize current knowledge of these interactions in the hope of identifying new avenues for research and therapy.

Animals↗

PubMed2002

Killing of Klebsiella pneumoniae by human alveolar macrophages.

We investigated putative mechanisms by which human surfactant protein A (SP-A) effects killing of Klebsiella pneumoniae by human alveolar macrophages (AMs) isolated from bronchoalveolar lavagates of patients with transplanted lungs. Coincubation of AMs with human SP-A (25 microg/ml) and Klebsiella resulted in a 68% decrease in total colony forming units by 120 min compared with AMs infected with Klebsiella in the absence of SP-A, and this SP-A-mediated effect was abolished by preincubation with N(G)-monomethyl-L-arginine. Incubation of transplant AMs with SP-A increased intracellular Ca(2+) concentration ([Ca(2+)](i)) by 70% and nitrite and nitrate (NO(x)) production by 45% (from 0.24 +/- 0.02 to 1.3 +/- 0.21 nmol small middle dot 10(6) AMs(-1).h(-1)). Preincubation with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester inhibited the increase in [Ca(2+)](i) and abrogated the SP-A-mediated Klebsiella phagocytosis and killing. In contrast, incubation of AMs from normal volunteers with SP-A decreased both [Ca(2+)](i) and NO(x) production and did not result in killing of Klebsiella. Significant killing of Klebsiella was also seen in a cell-free system by sustained production of peroxynitrite (>1 microM/min) at pH 5 but not at pH 7.4. These findings indicate that SP-A mediates pathogen killing by AMs from transplant lungs by stimulating phagocytosis and production of reactive oxygen-nitrogen intermediates.

Bronchoalveolar Lavage Fluid↗

PubMedDate not supplied

The role of nitric oxide in lung innate immunity: modulation by surfactant protein-A.

Surfactant protein A (SP-A) and alveolar macrophages are essential components of lung innate immunity. Alveolar macrophages phagocytose and kill pathogens by the production of reactive oxygen and nitrogen species. In particular, peroxynitrite, the reaction product of superoxide and nitric oxide, appears to have potent antimicrobial effects. SP-A stimulates alveolar macrophages to phagocytose and kill pathogens and is important in host defense. However, SP-A has diverse effects on both innate and adaptive immunity, and may stimulate or inhibit immune function. SP-A appears to mediate toxic or protective effects depending on the immune status of the lung. In contrast to mouse or rat cells, it has been difficult to demonstrate nitric oxide production by human macrophages. We have recently demonstrated that human macrophages produce nitric oxide and use it to kill Klebsiella pneumoniae. SP-A either stimulates or inhibits this process, depending on the activation state of the macrophage. Given its diverse effects on immune function, SP-A may prove to be an effective therapy for both infectious and inflammatory diseases of the lung.

Animals↗