Recent Publications by CFE Educators

1. This study addressed the contribution of primary afferents, mast cells, and sympathetic efferents to the control of vascular permeability in synovial joints. Extravasation of Evans blue dye into the synovial space was measured by perfusion of the knee joint in the adult rat. Plasma extravasation (PE) was evoked by pharmacologic activation of either unmyelinated primary afferents, mast cells, or sympathetic postganglionic nerve (SPGN) terminals with acute injection of either capsaicin, compound 48/80, or 6-hydroxydopamine (6-OHDA), respectively. In otherwise untreated control rats, acute infusion of capsaicin or compound 48/80 produced a brief increase in vascular permeability; infusion of 6-OHDA produced a larger and more prolonged increase. 2. To evaluate the contribution of an interaction of different cellular elements in the joint to PE, we repeated these experiments in rats pretreated with capsaicin, compound 48/80, or 6-OHDA; administered quercetin; or surgically sympathectomized by excision of the lumbar sympathetic chain. Eliminating unmyelinated afferent nerve terminals by neonatal treatment with capsaicin only reduced the increase in PE produced by acute infusion of capsaicin. Degranulating mast cells by pretreatment with compound 48/80, or preventing the degranulation of mast cells by treatment with quercetin, reduced the increase in PE evoked by infusion of either capsaicin or compound 48/80. Finally, sympathectomy, produced by excision of the lumbar sympathetic chain or by pretreatment with 6-OHDA, significantly reduced PE elicited by acute infusion of capsaicin, compound 48/80, or 6-OHDA. 3. Neither infusing substances normally localized to sympathetic efferents nor inducing changes in blood pressure could mimic the profound increase in PE evoked by activation of sympathetic postganglionic neurons with acute infusion of 6-OHDA. Thus norepinephrine produced a significant decrease in PE, adenosine triphosphate produced only a brief increase, neuropeptide Y had no effect, and manipulating blood pressure (either up or down) had no effect on either base-line or 6-OHDA-induced PE. 4. Indomethacin treatment significantly reduced the increase in PE produced by 6-OHDA. This effect of indomethacin was reversed by the addition of prostaglandin E2 (PGE2) to the 6-OHDA in the perfusion fluid. This finding implicates prostaglandins (i.e., cyclooxygenase products of arachidonic acid metabolism) in SPGN-dependent generation of PE.(ABSTRACT TRUNCATED AT 400 WORDS)

Beryllium lung disease is a chronic granulomatous disorder in which a beryllium-specific immune response plays a central role. By using a measure of cellular immune response to beryllium salts, bronchoalveolar lavage (BAL), and lung biopsy, we have identified 12 new cases of beryllium disease. Each of these individuals had pathologic changes on biopsy, lymphocytic alveolitis on BAL, and positive BAL lymphocyte transformation tests (LTT) in response to beryllium sulfate. This group of patients was remarkable for its paucity of clinical findings. At initial evaluation, seven had no respiratory symptoms, and four had normal physical examinations. Five had no increase in interstitial markings on chest radiograph. In eight cases, flow rates and lung volumes were normal. Diffusing capacity for carbon monoxide was low in only one case, and oxygen exchange during exercise was normal in six of nine subjects studied. In addition to the 12 cases, we evaluated eight beryllium-exposed workers who had other (nonberyllium) lung diseases; two of these eight demonstrated beryllium sensitization based on BAL LTT. We conclude that use of fiberoptic bronchoscopy with transbronchial biopsy and BAL facilitates diagnosis of beryllium workers who have histopathologic and immunologic alterations consistent with chronic beryllium disease. These findings may precede frank clinical illness and physiologic impairment, having important implications for our understanding of the natural history of beryllium disease.

In previous studies we reported that the rat spinal cord contains relatively high levels of uric acid and that the levels in a rat model of bilateral chronic pain, experimental adjuvant arthritis. In this report we evaluate the changes in UA in the unilaterally deafferented rat, a preparation which has also been used to study chronic pain. Uric acid was measured by high-pressure liquid chromatography with electrochemical detection in the spinal cord of rats that underwent unilateral, multiple cervical dorsal rhizotomy. Compared to control and sham-operated rats, there was a significant increase in the level of uric acid in the dorsal quadrant of the spinal cord ipsilateral to the dorsal rhizotomy. This increase was present at 1 and 4 weeks after surgery. At 1 week, we also observed a small but statistically insignificant increase in uric acid levels in the dorsal quadrant contralateral to the deafferentation and in sham-operated rats. Four weeks after surgery the levels of UA in all regions except for the deafferented dorsal quadrant returned to normal. The possibility was raised that the changes in uric acid reflect an increase in purinergic metabolism in the spinal cord secondary to the increased activity of the dorsal horn neurons that occurs with deafferentation.

This study addressed the opiate receptor subclass which underlies the paradoxical analgesic action of intrathecal administration of low doses of the stereospecific opiate receptor antagonist, naloxone. The analgesic effect of low dose naloxone was abolished in rats that had been pretreated 24 hr earlier with a large intrathecal dose of naloxone or 20 min previously with the delta receptor specific antagonist, ICI-174,864. A large intrathecal dose of naloxone administered 24 hr previously also abolished the analgesic effects of the delta-specific ligands [D-Pen2,5]-enkephalin and [D-Ser2]-Leu enkephalin-Thr but not those produced by mu-ligands Tyr-D-Ala-Gly-NMe-Phe-Gly-ol and beta-Casomorphin(1-4) amide or by the kappa-specific ligand, U50,488H. Furthermore, the analgesic action of low dose naloxone (on thermal and mechanical nociceptive threshold tests) persisted in rats made tolerant to mu-opioid receptor specific ligands. We conclude that the analgesic action of a low dose of naloxone is mediated via interaction with a stereospecific binding site with the characteristics of the delta-opioid receptor.

Light microscopic studies have demonstrated important differences in the distribution of enkephalin and dynorphin cells and terminals in the dorsal horn. Most importantly, dynorphin neurons are located in regions almost exclusively associated with the transmission and/or control of nociceptive messages (laminae I, IIo, and V); enkephalin neurons, although located in the same regions, are also found in areas involved in the transmission of nonnociceptive messages, e.g., laminae IIi and III. To determine whether there are also differences in the synaptic organization of the two opioid peptides, we have examined the distribution of dynorphin B immunoreactivity at the ultrastructural level. The studies were performed in colchicine-treated rats that underwent dorsal rhizotomy so that the relationship of dynorphin terminals and cells to primary afferent terminals could be established. Dynorphin B-immunoreactive cell bodies and dendrites in laminae I and IIo receive convergent primary and nonprimary afferent input, which suggests that dynorphin neurons receive a small-diameter, nociceptive input. Dynorphin terminals predominantly contain round, agranular vesicles; some terminals also contain a few dense core vesicles. Most dynorphin terminals are presynaptic to unlabelled dendrites; both asymmetric and symmetrical axonal contacts were noted. Dynorphin-immunoreactive boutons are also presynaptic to unlabelled cell bodies and spines. Twenty-nine percent of dynorphin terminals were associated with axonal profiles, including degenerating primary afferent terminals; only rarely could a synaptic density be detected. Although some degenerating primary afferent terminals were clearly presynaptic to dynorphin-immunoreactive terminals, in most cases, the polarity of the relationship between primary afferents and dynorphin terminals could not be established. These data indicate that synaptic interactions made by and with dynorphin-immunoreactive cells and terminals in the superficial dorsal horn are not very different from those that were previously reported for enkephalin cells and terminals. Thus, it is unlikely that dynorphin terminals provide a significant presynaptic input to primary afferent fibers. On the other hand, the presence of a primary afferent input to dynorphin cell bodies and dendrites in the superficial dorsal horn suggests that dynorphin cells receive a direct input from small-diameter, nociceptive primary afferents. That connection might contribute to the increased levels of dynorphin message and peptide that have been reported in rats experiencing a chronic inflammatory condition.(ABSTRACT TRUNCATED AT 400 WORDS)

Neutrophils are involved in the development of inflammatory edema in some animal models, but their mechanisms of action are not completely understood. Among injurious agents available for neutrophil-mediated injury are cationic proteins such as cathepsin G. Previous articles have reported that cationic agents decrease the barrier properties of the endothelium both in vivo and in vitro. Using an ex vitro isolated, perfused rabbit lung and a cultured porcine pulmonary artery endothelial cell monolayer (ENDO) as models, we asked whether neutrophil cathepsin G could decrease the barrier properties of the intact vasculature and cultured endothelial monolayers. After the addition of cathepsin G (10 micrograms/ml) to the perfusate of the isolated, perfused lung, there was a fourfold increase in net weight gain after a venous pressure challenge (p less than 0.01). The addition of cathepsin G to a cultured ENDO directly increased the movement of albumin across the monolayers in a dose-dependent fashion (10 micrograms/ml led to a 59% +/- 5% increase, 15 micrograms/ml to a 135% +/- 55% increase, 20 micrograms/ml to a 247% +/- 78% increase, and 30 micrograms/ml to a 381% +/- 89% increase, p less than 0.05 at all concentrations). Heat-inactivation of the enzyme only partially protected the ENDO, but exposure in the presence of either the polyanion heparin or the polyanion dextran sulfate completely protected the ENDO. Normal human serum also protected the ENDO, but serum from two patients with alpha 1-proteinase inhibitor (alpha 1-PI) deficiency was only partially protective. The protection afforded by human serum was time dependent, because addition of the serum 5 or 15 minutes after the addition of cathepsin G offered no protection. Oxidation of alpha 1-PI, as may occur at sites of inflammation, also destroyed its protective effect. Cytotoxic injury of the porcine pulmonary artery endothelial cell monolayer by cathepsin G, which was also prevented by the polyanions, only partially explained these results, because cathepsin G increased albumin transfer across the ENDO at concentrations of 10 to 20 micrograms/ml, which were minimally cytolytic to the ENDO. Additionally, cathepsin G caused cell retraction, with the development of intercellular gaps visible on light microscopy of the ENDO. This effect was also prevented by the polyanions. These results demonstrate that cathepsin G has direct effects on a cultured ENDO that may be caused by the charge or charged site(s) on cathepsin G.

This study provides an anatomical basis for the observation that a unilateral lesion of the spinal dorsolateral funiculus (DLF) can reduce the inhibitory effect of electrical stimulation of the nucleus raphe magnus (NRM) on dorsal horn nociceptive neurons located caudal and contralateral to the lesion. We injected the anterograde tracer Phaseolus vulgaris leucoagglutinin (PHA-L) into the NRM and traced the arborization of single DLF axons in the spinal gray matter. Although the majority of DLF axons arborized in the dorsal horn ipsilaterally, we found some axons which entered the spinal gray matter, traversed the gray matter to the central canal and then abruptly changed direction and coursed to the contralateral superficial dorsal horn. Very few branches were given off en route. These data indicate that some raphe-spinal axons may selectively influence the firing of neurons of the superficial dorsal horn, contralateral to the DLF in which they descend the spinal cord.

The accumulation of inflammatory and immune effector cells in the lungs of patients at early stages of interstitial lung disease has been well documented, but little is known about the functional activity of these cells, particularly alveolar macrophages. The purpose of the experiments described here was to determine whether alveolar macrophages from patients with idiopathic pulmonary fibrosis (IPF) differed from alveolar macrophages of normal subjects using a model system that assesses a component of cell-mediated immunity. Alveolar macrophages were tested for their ability to phagocytose and kill the facultative intracellular bacterium Listeria monocytogenes. Because this system requires the stimulation of macrophages by antigen-specific T-cells, it allows the assessment of effector functions of macrophages found in the lower respiratory tract of patients with IPF. Data showed that alveolar macrophages from normal subjects both phagocytosed and killed those bacteria. However, alveolar macrophages from patients with IPF phagocytosed bacteria normally but expressed little bactericidal or bacteriostatic activity. Further, we found no difference in HLA-DR expression by normal and IPF alveolar macrophages. Therefore, it does not appear that this defect in bactericidal activity in IPF macrophages results from a defect in the antigen-presenting function of these cells. These data suggest that alveolar macrophages from patients with IPF express a functional deficiency in their ability to kill facultative intracellular bacteria.

Bronchiolitis obliterans in the adult patient is a relatively uncommon and vexing clinical entity. This confusion results because this pathologic finding occurs in a variety of diverse clinical settings. Bronchiolitis obliterans is a fibrotic process that primarily affects the small conducting airways. The lesion results from damage to the bronchiolar epithelium and the repair process leads to excessive proliferation of granulation tissue. The alveoli adjacent to the small airway are almost always involved; however, a considerable portion of the interstitium is usually spared. The findings in these patients may physiologically and radiographically mimic chronic obstructive pulmonary disease (COPD). On the other hand, some of the processes associated with bronchiolitis obliterans result in restrictive or mixed restrictive and obstructive ventilatory defects; consequently, they may be confused with other diffuse infiltrative lung disorders. This review will focus principally on bronchiolitis obliterans in adults, which, until recently, was considered rare. There has been heightened interest in this process in adults because of its association with the connective tissue diseases, its development following toxic fume exposure, its occurrence as a result of chronic graft versus host reactions, and the increasing recognition of patients with idiopathic forms of the disease that have an insidious onset often confused with more common problems such as COPD or idiopathic pulmonary fibrosis.