Patient variables impact lumbar spine dual energy X-ray absorptiometry precision.

INTRODUCTION: Changes in bone mineral density are used to monitor osteoporosis therapy. To determine whether a change in bone mass is clinically significant, the precision of bone mineral density measurements must be known. METHODS: We therefore measured the impact of vertebral body exclusion on dual energy X-ray absorptiometry (DXA) precision. At one university and one Veterans Affairs DXA center, three radiology technologists each scanned 30 participants twice, with repositioning between scans, to estimate DXA precision. Three International Society for Clinical Densitometry-certified physicians reviewed all lumbar spinal scans to note the presence of focal structural defects. We calculated precision for subsets of vertebrae, and for virtual samples of patients with and without physician-identified vertebral focal structural defects. We graphed the reciprocal of least significant change versus bone area to determine the dependence of precision on interpreted scan area. RESULTS: Within each sample, greater interpretable bone area improved precision. The contribution of interpreted bone area to precision differed among the samples, ranging from 57 to 94%. Greater population bone mineral density heterogeneity and presence of focal structural defects each decreased precision. CONCLUSION: All bone densitometry centers must determine precision using a sample representative of their served populations. Failure to do so may lead to incorrect determination of least significant change. Population heterogeneity, vertebral body exclusion and presence of focal structural defects each decreases precision.

Effects of exercise on knee joints with osteoarthritis: a pilot study of biologic markers.

OBJECTIVE: To determine the effects of low intensity weight-bearing exercise on osteoarthritis (OA) of the knee. METHODS: Synovial fluid keratan sulfate (KS) and hydroxyproline were measured as markers of cartilage degradation. The Arthritis Impact Measurement Scales (AIMS) were used to measure health status, and a visual analog scale for pain assessment was used before and after intervention. An exercise (EX) group (n = 15) received a thrice-weekly 12-week low intensity exercise program and a weekly educational program, and a minimal treatment (Min RX) group (n = 15) received only the education program. RESULTS: Pain levels declined in the EX group, and the Min RX group showed improvement on the AIMS. Synovial fluid was obtained in 11 subjects before and after the intervention. Levels of KS and hydroxyproline did not change. CONCLUSION: Further study of exercise effects should include both clinical and biologic parameters to examine the outcome of exercise as a therapeutic intervention in OA of the knee.

Mast cell activation in human synovium explants by calcium ionophore A23187, compound 48/80, and rabbit IgG anti-human IgE, but not morphine sulfate.

To investigate human synovial mast cell physiology, we developed a model in which mast cells in human synovial explant cultures were activated by immunologic or non-immunologic mechanisms. Small (3 mm) cubes of synovial membrane were incubated with or without secretagogue for 30, 45 or 60 min, and supernatant histamine concentrations were quantified. We measured significant histamine release with compound 48/80 at concentrations > or = 1 mg/ml, and with calcium ionophore A23187 at > or = 5 micrograms/ml. Rabbit IgG anti-human IgE induced significant histamine release at all concentrations tested, maximum at 78 micrograms/ml. Morphine sulfate produced no histamine release from synovial explants, in contrast to its significant stimulation of histamine release from neonatal foreskin explants in our explant system. We confirmed synovial mast cell degranulation by electron microscopy, and showed that it corresponded with measurable histamine release. Furthermore, histamine release was not due to secretagogue-induced cytotoxicity, as assessed by supernatant lactate dehydrogenase levels and by ultrastructural analysis. Since morphine sulfate induces mast cell degranulation and histamine release in adult and neonatal human skin, our data show that although synovial and dermal mast cells have a similar granule enzyme profile and electron microscopic morphology, they differ in functional responses. These observations support recent data that among similar human mast cell subtypes there are physiologic differences. Finally, our explant model will be useful in studies of mast cell involvement in arthritis.

Surgical "touch artefacts" of the cerebral cortex. An experimental study with light and electron microscopic analysis.

In surgically excised brain tissue a very common artefact, unrelated to the presence or absence of a genuine pathological process, involves the water content of cortical neurons. Nerve cells may show massive watery swelling of both their cytoplasm and nuclei or conversely may become shrunken and dark-staining. The most important aspect of this alteration is that it may lead to mistaken histopathological interpretation, but the question also arises whether such changes, presumably caused by exposure, touch, pressure or the combination of all 3, in the living patient, would persist after surgery and would eventually result in irreversible damage to the involved neurons? Thirty rats were operated in this experiment: craniotomy and opening of the dura mater was done over one convexity and slight pressure (uniformly calibrated for all animals) was applied to the exposed cortex. The wound was closed and the animals were sacrificed at various intervals, ranging from immediately after the operation to 6 weeks. The areas that suffered compression were examined by light and electron microscopy. Initial changes included marked watery swelling of neuronal perikarya and nuclei which predominated over pyknotic changes. The changes thus produced were identical with those seen in portions of the cortex excised immediately after pressure was applied to the area in 10 additional rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Cholestatic hepatitis after ingestion of chaparral leaf: confirmation by endoscopic retrograde cholangiopancreatography and liver biopsy.

The use of herbal and other "natural" health products by healthy and ill people is more common than is appreciated by many health care providers. Since most of these substances are not categorized as medicines, they are exempt from U.S. Government approval processes, and are essentially uncontrolled. In this article we describe a patient who developed painless jaundice, fatigue, and pruritus after taking chaparral tablets, 160 mg/day, for approximately 2 months. Serial liver biopsies and serum chemistries documented severe cholestasis and hepatocellular injury, i.e., a severe cholangiolitic hepatitis. Serum enzyme levels were markedly elevated: alk. phos. to four-fold, alanine aminotransferase and aspartate aminotransferase to 25-fold, total bilirubin to 30-fold, and gamma-glutamyl transpeptidase to 35-fold. Endoscopic retrograde cholangiopancreatography showed smooth, but severely narrowed biliary ducts without sclerosing cholangitis, distal obstruction, tumor, or stenosis. The diagnosis remained in doubt until the publication of two cases of chaparral hepatotoxicity. Because of the similarity of our patient's illness to those cases we concluded that chaparral was almost certainly the cause. Chaparral, also known as creosote or greasewood, is used by some practitioners to treat a diverse group of ailments including ethanol withdrawal. This report should heighten the awareness by primary care physicians and gastroenterologists that any chaparral herbal preparation is a potential hepatotoxin that can lead to serious illness.(ABSTRACT TRUNCATED AT 250 WORDS)

Interferon gamma induced production of indoleamine 2,3 dioxygenase in cultured human synovial cells.

OBJECTIVE: Synovial membrane cells from inflamed joints share morphological and functional properties with malignant mesenchymal cells. Interferon gamma (IFN-gamma) has antitumor cell activity related to stimulation of 2,3 indoleamine dioxygenase (IDO), a widely distributed tryptophan catabolizing enzyme. Our objective was to measure synoviocyte IDO production to determine if the varied clinical and in vitro effects of IFN-gamma on nonmalignant immunocompetent cells might involve a similar mechanism. METHODS: Using an established radioenzymatic assay, we measured IDO activity in suspensions of freshly isolated cells obtained by enzymatic dispersion of human synovial membrane, and in fresh and longterm (> or = 2 months) cultures of these cells in response to varying concentrations of recombinant human interferons alpha 2a, beta ser, or gamma. RESULTS: In fresh and in > or = 2 month-old cultures, IFN-gamma strongly stimulated IDO activity, a corresponding fall in supernatant tryptophan levels, and an elevation in the supernatant concentration of kynurenine, tryptophan's principal metabolite, mRNA for IDO was likewise markedly increased in cells after 4 days' incubation with IFN-gamma. Staining studies indicated that the IDO producing cells in synovium were not typical macrophages. Interferon beta ser had weak IDO stimulatory activity that was in a few cases additive to that of IFN-gamma. In no case did interferon beta ser abrogate IFN-gamma induced IDO activity increases. Interferon alpha 2a also had weak stimulatory activity. CONCLUSIONS: IFN-gamma stimulates IDO production and tryptophan metabolism in cultured human synovial cells, and therefore may contribute to this cytokine's in vitro and clinical effects in arthritis and inflammation.

Osteomyelitis of the skull base.

Three cases of osteomyelitis of the skull base with associated problems in diagnosis and therapy are discussed. Patients with atypical skull base osteomyelitis are difficult to diagnose as they have no ear abnormalities, but they often develop multiple cranial nerve deficits mimicking symptoms of a posterior fossa mass. We conclude that computed tomographic scans, magnetic resonance imaging studies, bone scans indium-labeled white blood cell scans, and gallium scans are useful in making the diagnosis. A biopsy of the bony lesion often is needed to identify the causative organism and to rule out a tumor. Intravenously administered antibiotics are the mainstay of therapy and should be continued until 1 week after the gallium scan shows no abnormalities. Follow-up gallium scans then are done at 1 week and 3 months after the cessation of antibiotic therapy to search for a recurrence.

A lavage method for dynamic intraarticular monitoring of animal joints in situ: quantification and release kinetics of histamine after selective synovial mast cell activation by diverse secretagogues.

Evidence implicating synovial mast cells in the initiation and perpetuation of arthritis has increased. We have developed a method of joint lavage to monitor dynamic intraarticular events in the intact animal. Lavage was done before and after immunologic (passive sensitization followed by intravenous specific antigen or intraarticular anti-rat immunoglobulin E [IgE] heteroantiserum) and nonimmunologic (intraarticular calcium ionophore A-23187; phorbol 12-myristate, 13-acetate; and compound 48/80) synovial mast cell activation. To quantify and analyze synovial mast cell mediator release kinetics in situ, we measured lavage fluid histamine. With all activation protocols except A-23187, histamine release was evident within 5 minutes after introduction of the stimulus. The quantitative and chronological similarities between immunologically induced and compound 48/80-induced synovial mast cell histamine release kinetics suggested that connective tissue type mast cells are an important source of inflammatory mediators in rat joints. We also measured joint lavage fluid histamine levels in rats immunized with an active sensitization protocol. Histamine levels were determined by the autoanalyzer method and were confirmed by using a commercially available radioimmunoassay that uses a monoclonal antibody against acetylated histamine. We found that in many of these animals, at the peak of the serum IgE response, joint lavage fluid histamine levels were very high even before challenge with specific antigen, and that this increase was not due to diffusion into the joint of abnormally elevated plasma histamine. These data suggested that synovial mast cells are preferentially activated in states of high serum IgE immune responses. We have used a simple, inexpensive, rapid lavage technique to generate the first data on histamine release kinetics after selective synovial mast cell activation in the intact animal. The technique can be adapted for investigation of release kinetics of a variety of other substances from activated synovial cells and can be used in other arthritis models.

Human synovial mast cell involvement in rheumatoid arthritis and osteoarthritis. Relationship to disease type, clinical activity, and antirheumatic therapy.

Mast cells were isolated by enzymatic digestion of synovium obtained from 48 patients with rheumatoid arthritis (RA) and 42 patients with osteoarthritis (OA). A significantly lower percentage of stainable synovial mast cells was obtained by tissue digestion from patients with clinically active RA compared with those with less active disease. The 54 patients treated with nonsteroidal antiinflammatory drugs had a significantly lower percentage of stainable synovial mast cells in cell suspension than did the other 36 patients. When anti-IgE antibody was used as a secretagogue in vitro, significantly greater histamine release was observed from synovial mast cells of RA patients compared with OA patients. Greater histamine release in response to anti-IgE was observed in the RA patients with more clinically active disease and those who were treated with prednisone, compared with RA patients without these features. Synovial mast cells of RA patients treated with a disease-modifying antirheumatic drug had a significantly lower mean histamine content than did cells from patients not receiving such treatment. Our data suggest that there are differences between synovial mast cells from tissues of patients with RA and OA and suggest that synovial mast cells may be activated in clinically active RA. In addition, the data indicate an effect of systemic antirheumatic therapy on mast cells isolated from synovium of patients with arthritis.

In vivo effects of nonsteroidal antiinflammatory drugs on rat skin and synovial mast cell-induced vasopermeability.

Using our animal model of synovial mast cell-mediated arthritis in rats, we tested the effects of 3 nonsteroidal antiinflammatory drugs (NSAIDs) (aspirin, indomethacin, and ketoprofen) and an H1 and an H2 histamine receptor antagonist (diphenhydramine and cimetidine, respectively) on synovial and dermal mast cell-induced vasopermeability. Drug effects were assessed by quantifying the leakage of radiolabeled albumin into tissues following specific antigen-initiated activation of passively sensitized dermal and synovial mast cells. The 3 NSAIDs tested had different effects on synovial and dermal mast cell-induced vasopermeability. Aspirin and indomethacin significantly increased dermal and synovial plasma exudation (P less than or equal to 0.008). Ketoprofen decreased dermal (P = 0.015), but had no effect on synovial, vascular exudation. Complete histamine H1 and H2 receptor blockade with diphenhydramine and cimetidine, respectively, substantially decreased (P less than or equal to 0.0008), but did not completely inhibit, dermal and synovial mast cell-induced vasopermeability. However, the addition of indomethacin to the combined antihistamine regimen resulted in an increase in the leakage of the radiolabel into skin and synovium (back to control levels), despite the complete blockade of H1 and H2 receptors. Results of experiments with antihistamines and indomethacin suggest that mediators other than histamine are involved in synovial mast cell-induced inflammation. Furthermore, the differential response to ketoprofen indicates that the specific antigen-stimulated mediator release profiles of dermal and synovial mast cells are different. Our finding of enhanced synovial vascular leakage in animals treated with some NSAIDs, and no such effect by other NSAIDs, perhaps explains in part the diverse effects of these agents in humans with arthritis.

Implications of interferon-induced tryptophan catabolism in cancer, auto-immune diseases and AIDS.

Tryptophan (Trp) is an indispensable amino acid required for biosynthesis of proteins, serotonin and niacin. Indoleamine 2,3-dioxygenase (IDO) is induced by infections, viruses, lipopolysaccharides, or interferons (IFNs) and this results in significant catabolism of Trp along the kynurenine (Kyn) pathway. Intracellular growth of Toxoplasma gondii and Chlamydia psittaci in human fibroblasts in vitro is inhibited by IFN-gamma and this inhibition is negated by extra Trp in the medium. Similarly, growth of a number of human cell lines in vitro is inhibited by IFN-gamma and addition of extra Trp restores growth. Thus, in some in vitro systems, antiproliferative effects of IFN-gamma are mediated by induced depletion of Trp. We find that cancer patients given Type I or Type II IFNs can induce IDO which results in decreased serum Trp levels (20-50% of pretreatment) and increased urinary metabolites of the Kyn pathway (5 to 500 fold of pretreatment). We speculate that in vivo antineoplastic effects of IFNs and clinical side effects are mediated, at least in part, by a general or localized depletion of Trp. In view of reported increases of IFNs in autoimmune diseases and our earlier findings of elevated urinary Trp metabolites in autoimmune diseases, it seems likely that systemic or local depletion of Trp occurs in autoimmune diseases and may relate to degeneration, wasting and other symptoms in such diseases. We find high levels of IDO in cells isolated from synovia of arthritic joints. IFNs are also elevated in human immunodeficiency virus (HIV) patients and increasing IFN levels are associated with a worsening prognosis. We propose that IDO is induced chronically by HIV infection, is further increased by opportunistic infections, and that this chronic loss of Trp initiates mechanisms responsible for the cachexia, dementia, diarrhea and possibly immunosuppression of AIDS patients. In these symptoms, AIDS resembles classical pellagra due to dietary deficiency of Trp and niacin. In preliminary studies, others report low levels of Trp and serotonin, and elevated levels of Kyn and quinolinic acid in AIDS patients. The implications of these data in cancer, autoimmune diseases and AIDS are discussed.

Droperidol-fentanyl inhibits mast cell histamine release in rat synovium, but not skin, after immunologic activation: evidence for mast cell heterogeneity.

The characteristics of mast cell heterogeneity include differences in mast cell staining, cellular structure, content of proteoglycans and vasoactive amines, and in vitro responses to secretagogues. We have found that the anesthetic agent, droperidol-fentanyl, significantly decreased rat synovial vascular permeability after selective synovial mast cell degranulation. In contrast, there was no effect on skin mast cell-induced vascular permeability. To address the possibility that this difference in response represents mast cell heterogeneity and uniqueness of synovial mast cells, we investigated the effects of three different anesthetic agents on synovial mast cells. In these studies we examined whether (1) the drugs affect synovial mast cells and (2) whether the drugs affect synovial mast cells differently from mast cells in skin. Our data indicate that, compared to inhaled methoxyflurane and ethyl ether, subcutaneous droperidol-fentanyl had a significant inhibitory effect on synovial, but not skin, mast cell-mediated vascular permeability, which was associated with a significant inhibition of synovial mast cell histamine release. Thus, the permeability differences were not due only to effects on histamine receptors. Our data suggest the possibility that there are significant functional differences between synovial and skin mast cells, a phenomenon that may have important therapeutic implications for treatment of mast cell-mediated synovial inflammation.

Mast cells and arthritis.

There is suggestive evidence, both in vitro and in vivo, that mast cells and their products are involved in the pathogenesis of some forms of arthritis. Because mast cells may be activated by both IgE-mediated and non-IgE-mediated stimuli, they provide a putative link between specific antigenic, neurologic, and inflammatory stimuli and synovial inflammation. The degree to which this occurs and the relative clinical significance remains to be defined.

Demonstration and characterization of a transient arthritis in rats following sensitization of synovial mast cells with antigen-specific IgE and parenteral challenge with specific antigen.

We have developed a model of IgE-dependent, mast cell-mediated arthritis in rats. One knee joint (test joint) of a Sprague-Dawley rat was injected with 1 micrograms of a monoclonal IgE specific for dinitrophenol, and the contralateral (control) joint was injected with the same amount of an irrelevant monoclonal IgE in phosphate buffered saline or with phosphate buffered saline alone. Within 5 minutes of intravenous injection of antigen, an acute, transient arthritis occurred in the test joints only, with swelling and extravasation of intravascular blue dye and 125I-labeled albumin, decreased numbers of stainable mast cells, and decreased histamine content of the joint synovium. Pretreatment of experimental animals with H1 and H2 antihistamines did not completely block the reaction. These data show that IgE-dependent synovial mast cell degranulation causes a transient, nondestructive arthritis, reminiscent of lupus arthritis and intermittent hydrarthrosis.

Production of granulocyte-macrophage colony-stimulating factor by primary cultures of unstimulated rat microvascular endothelial cells.

Small vessel (microvascular) endothelial cells are in close contact with hematopoietic progenitor cells in the bone marrow and therefore may have an important role in hematopoiesis. Although other studies have shown that endothelial cells produce various colony-stimulating factors (CSFs), these studies examined large vessel endothelial cells, which are different in many respects from microvascular endothelial cells and which do not contact cells in the bone marrow. We show in this study that primary cultures of unstimulated rat fat capillary endothelial cells grown in serum-free medium produce a substantial amount of granulocyte-macrophage CSF (GM-CSF). The medium conditioned by these cells stimulated proliferation of two different lines of GM-CSF-responsive cells--PT-18 mast cells and FDC-P1 cells--and supported the growth of cells of the granulocyte and macrophage lines in cultures of rat bone marrow cells. The factor responsible for this activity had physical properties consistent with those of GM-CSF, namely, a similar apparent mol wt by gel filtration, resistance to repeated freeze-thaws, resistance to boiling for ten minutes but not for 30 minutes, and resistance to heating to 56 degrees C for one hour. The factor causing target cell stimulation was not B cell-stimulating factor-1 (BSF-1, or IL 4), since it failed to stimulate a BSF-1-responsive cell line HT2-JH, and target cells (PT-18) did not respond appreciably to recombinant BSF-1. Northern blot analysis of mRNA from rat fat capillary endothelial cells showed high levels of expression of GM-CSF, confirming that this factor is produced by microvascular endothelial cells. This is the first report of CSF production by unstimulated microvascular endothelial cells, demonstrating that these ubiquitous cells are capable of producing sizable amounts of at least one growth factor for hematopoietic progenitor cells.

Mast cell numbers in rheumatoid synovial tissues. Correlations with quantitative measures of lymphocytic infiltration and modulation by antiinflammatory therapy.

Synovial biopsy specimens from 20 patients with rheumatoid arthritis were subjected to quantitative analysis for several parameters of inflammation and for enumeration of synovial tissue mast cells. Strong positive correlations were found between numbers of mast cells per cubic millimeter of synovial tissue and the following synovial tissue parameters: inflammatory index (a quantification of lymphocytic infiltration), Leu-3a grade (T helper/inducer lymphocytes), Leu-1 grade (T lymphocyte), and plasma cell grade. A strong negative correlation was found between the synovial mast cell count and the extent of sublining layer fibrin deposition. Correlations between synovial mast cell count and Leu-2a grade, ratio of Leu-3a grade:Leu-2a grade, OKM1 grade, HLA-DR grade, and lining layer thickness grade did not reach statistical significance. In addition, we obtained synovial specimens from 6 of the patients both before and after long-term therapy with oral methotrexate and from 3 of the patients before, and 1 week after, an intraarticular injection of steroid. The 3 patients who had an intraarticular steroid injection showed a 67-96% decrease in the number of synovial tissue mast cells; there was no significant change in the number of synovial mast cells in the tissues of the 6 patients who received oral methotrexate. These observations are the first documentation of a quantitative relationship between the number of mast cells and the number and phenotypic profile of infiltrating lymphocytes in an inflamed tissue, which in this case, is human synovium. Our findings suggest that mast cells are involved in the pathologic interactions in rheumatoid arthritis and might play a role in the early phases of exacerbations of disease activity.

Mast cell numbers and histamine levels in synovial fluids from patients with diverse arthritides.

Thirty-five synovial fluid (SF) specimens were examined for the presence of mast cells and for their histamine content. Mast cells were seen in SF cells from 27 of 35 fluids, and histamine was measurable in 19 of 34. There was a strong correlation between mast cell number and histamine content. No consistent relationship was found between either the mast cell number or histamine level and the patients' diagnoses, except that the 2 patients with systemic mastocytosis had markedly elevated values for both SF mast cell number and histamine content. SF mast cells from one of the mastocytosis patients were studied for histamine release; significant amounts of histamine were released upon exposure to anti-human IgE, but not compound 48/80. Thus, mast cells similar to those present in connective tissue are frequently present in SF in numbers which correlate with SF histamine levels. These mast cells contain active proteases and are capable of degranulation. Mast cells were consistently present in large numbers in the SF of patients with systemic mastocytosis, but their numbers were highly variable in fluids of patients with other diseases.

Role of the thymus in streptococcal cell wall-induced arthritis and hepatic granuloma formation. Comparative studies of pathology and cell wall distribution in athymic and euthymic rats.

Systemic administration of an aqueous suspension of group A streptococcal cell wall fragments to susceptible rats induces acute and chronic polyarthritis, as well as noncaseating hepatic granulomas. To gain insight into the role of the thymus in the pathogenesis of this experimental model, pathologic responses and cell wall tissue distribution were compared in congenitally athymic rats (rnu/rnu) and their euthymic littermates (NIH/rnu). Within 24 h, both rat strains developed acute arthritis, characterized by polymorphonuclear leukocytic exudate in the synovium and joint spaces. This acute process was maximal at day 3 and gradually subsided. Beginning 2-3 wk after injection, the euthymic, but not the athymic, rats developed the typical exacerbation of arthritis, characterized by synovial cell hyperplasia with villus formation and T helper/inducer lymphocyte-rich mononuclear cell infiltration. This process eventually resulted in marginal erosions and destruction of periarticular bone and cartilage. Parallel development of acute and chronic hepatic lesions was observed. Bacterial cell wall antigen distribution and persistence were similar in the athymic and euthymic rats. Cell wall antigens were demonstrated in the cytoplasm of cells within subchondral bone marrow, synovium, liver, and spleen, coincident with the development of the acute lesions, and persisted in these sites, although in decreasing amounts, for the duration of the experiment. Our findings provide evidence that the acute and chronic phases of the experimental model are mechanistically distinct. The thymus and functional thymus derived-lymphocytes appear not to be required for the development of the acute exudative disease but are essential for the development of chronic proliferative and erosive disease. Induction of disease is dependent upon cell wall dissemination to and persistence in the affected tissues.