Carl R. Fuhrman

Experimental Mycobacterium tuberculosis Infection of Cynomolgus Macaques Closely Resembles the Various Manifestations of Human M. tuberculosis Infection

ABSTRACT Nonhuman primates were used to develop an animal model that closely mimics human Mycobacterium tuberculosis infection. Cynomolgus macaques were infected with low doses of virulent M. tuberculosis via bronchoscopic instillation into the lung. All monkeys were successfully infected, based on tuberculin skin test conversion and peripheral immune responses to M. tuberculosis antigens. Progression of infection in the 17 monkeys studied was variable. Active-chronic infection, observed in 50 to 60% of monkeys, was characterized by clear signs of infection or disease on serial thoracic radiographs and in other tests and was typified by eventual progression to advanced disease. Approximately 40% of monkeys did not progress to disease in the 15 to 20 months of study, although they were clearly infected initially. These monkeys had clinical characteristics of latent tuberculosis in humans. Low-dose infection of cynomolgus macaques appears to represent the full spectrum of human M. tuberculosis infection and will be an excellent model for the study of pathogenesis and immunology of this infection. In addition, this model will provide an opportunity to study the latent M. tuberculosis infection observed in ∼90% of all infected humans.

Association of Radiographic Emphysema and Airflow Obstruction with Lung Cancer

RATIONALE To study the relationship between emphysema and/or airflow obstruction and lung cancer in a high-risk population. OBJECTIVE We studied lung cancer related to radiographic emphysema and spirometric airflow obstruction in tobacco-exposed persons who were screened for lung cancer using chest computed tomography (CT). METHODS Subjects completed questionnaires, spirometry, and low-dose helical chest CT. CT scans were scored for emphysema based on National Emphysema Treatment Trial criteria. Multiple logistic regressions estimated the independent associations between various factors, including radiographic emphysema and airflow obstruction, and subsequent lung cancer diagnosis. MEASUREMENTS AND MAIN RESULTS Among 3,638 subjects, 57.5, 18.8, 14.6, and 9.1% had no, trace, mild, and moderate-severe emphysema, and 57.3, 13.6, 22.8, and 6.4% had no, mild (Global Initiative for Chronic Obstructive Lung Disease [GOLD] I), moderate (GOLD II), and severe (GOLD III-IV) airflow obstruction. Of 3,638 subjects, 99 (2.7%) received a lung cancer diagnosis. Adjusting for sex, age, years of cigarette smoking, and number of cigarettes smoked daily, logistic regression showed the expected lung cancer association with the presence of airflow obstruction (GOLD I-IV, odds ratio [OR], 2.09; 95% confidence interval [CI], 1.33-3.27). A second logistic regression showed lung cancer related to emphysema (OR, 3.56; 95% CI, 2.21-5.73). After additional adjustments for GOLD class, emphysema remained a strong and statistically significant factor related to lung cancer (OR, 3.14; 95% CI, 1.91-5.15). CONCLUSIONS Emphysema on CT scan and airflow obstruction on spirometry are related to lung cancer in a high-risk population. Emphysema is independently related to lung cancer. Both radiographic emphysema and airflow obstruction should be considered when assessing lung cancer risk.

Quantitative Comparison of Active and Latent Tuberculosis in the Cynomolgus Macaque Model

ABSTRACT We previously described that low-dose Mycobacterium tuberculosis infection in cynomolgus macaques results in a spectrum of disease similar to that of human infection: primary disease, latent infection, and reactivation tuberculosis (S. V. Capuano III, D. A. Croix, S. Pawar, A. Zinovik, A. Myers, P. L. Lin, S. Bissel, C. Fuhrman, E. Klein, and J. L. Flynn, Infect. Immun. 71:5831-5844, 2003). This is the only established model of latent infection, and it provides a unique opportunity to understand host and pathogen differences across of range of disease states. Here, we provide a more extensive and detailed characterization of the gross pathology, microscopic histopathology, and immunologic characteristics of monkeys in each clinical disease category. The data underscore the similarities between human and nonhuman primate M. tuberculosis infection. Furthermore, we describe novel methods of quantifying gross pathology and bacterial burden that distinguish between active disease and latent infection, and we extend the usefulness of this model for comparative studies. Early in infection, an abnormal chest X ray, M. tuberculosis growth by gastric aspirate, and increased mycobacterium-specific gamma interferon (IFN-γ) in peripheral blood mononuclear cells (PBMCs) and bronchoalveolar lavage (BAL) cells were associated with the development of active disease. At necropsy, disease was quantified with respect to pathology and bacterial numbers. Microscopically, a spectrum of granuloma types are seen and differ with disease type. At necropsy, monkeys with active disease had more lung T cells and more IFN-γ from PBMC, BAL, and mediastinal lymph nodes than monkeys with latent infection. Finally, we have observed a spectrum of disease not only in monkeys with active disease but also in those with latent infection that provides insight into human latent tuberculosis.

Early Events in Mycobacterium tuberculosis Infection in Cynomolgus Macaques

ABSTRACT Little is known regarding the early events of infection of humans with Mycobacterium tuberculosis. The cynomolgus macaque is a useful model of tuberculosis, with strong similarities to human tuberculosis. In this study, eight cynomolgus macaques were infected bronchoscopically with low-dose M. tuberculosis; clinical, immunologic, microbiologic, and pathologic events were assessed 3 to 6 weeks postinfection. Gross pathological abnormalities were observed as early as 3 weeks, including Ghon complex formation by 5 weeks postinfection. Caseous granulomas were observed in the lung as early as 4 weeks postinfection. Only caseous granulomas were observed in the lungs at these early time points, reflecting a rigorous initial response. T-cell activation (CD29 and CD69) and chemokine receptor (CXCR3 and CCR5) expression appeared localized to different anatomic sites. Activation markers were increased on cells from airways and only at modest levels on cells in peripheral blood. The priming of mycobacterium-specific T cells, characterized by the production of gamma interferon occurred slowly, with responses seen only after 4 weeks of infection. These responses were observed from T lymphocytes in blood, airways, and hilar lymph node, with responses predominantly localized to the site of infection. From these studies, we conclude that immune responses to M. tuberculosis are relatively slow in the local and peripheral compartments and that necrosis occurs surprisingly quickly during granuloma formation.

Tumor necrosis factor neutralization results in disseminated disease in acute and latent Mycobacterium tuberculosis infection with normal granuloma structure in a cynomolgus macaque model

OBJECTIVE An increased risk of tuberculosis has been documented in humans treated with tumor necrosis factor alpha (TNFalpha)-neutralizing agents. In murine models, impaired signaling by TNF causes exacerbation of both acute and chronic infection associated with aberrant granuloma formation and maintenance. This study was undertaken to investigate immune modulation in the setting of TNF neutralization in primary and latent tuberculosis in a non-human primate model. METHODS Cynomolgus macaques 4 years of age or older were infected with Mycobacterium tuberculosis and subjected to clinical, microbiologic, immunologic, and radiographic examinations. Monkeys were classified as having active or latent disease 6-8 months after infection, based on clinical criteria. Monkeys used in acute infection studies were randomized to receive either adalimumab (prior to and during infection) or no treatment. Monkeys with latent infection that were randomized to receive TNF-neutralizing agent were given either an inhibitor of soluble TNF, recombinant methionyl human soluble TNF receptor I (p55-TNFRI), or adalimumab. Control monkeys with latent infection were given no treatment or saline. Data from previously studied monkeys with active or latent disease were also used for comparison. RESULTS Administration of TNF-neutralizing agents prior to M tuberculosis infection resulted in fulminant and disseminated disease by 8 weeks after infection. Neutralization of TNF in latently infected cynomolgus macaques caused reactivation in a majority of animals as determined by gross pathologic examination and bacterial burden. A spectrum of dissemination was noted, including extrapulmonary disease. Surprisingly, monkeys that developed primary and reactivation tuberculosis after TNF neutralization had similar granuloma structure and composition to that of control monkeys with active disease. TNF neutralization was associated with increased levels of interleukin-12, decreased levels of CCL4, increased chemokine receptor expression, and reduced mycobacteria-induced interferon-gamma production in blood but not in the affected mediastinal lymph nodes. Finally, the first signs of reactivation often occurred in thoracic lymph nodes. CONCLUSION These findings have important clinical implications for determining the mechanism of TNF neutralization-related tuberculosis.

The Pittsburgh lung screening study (PLuSS): Outcomes within 3 years of a first computed tomography scan

RATIONALE The role of computed tomography (CT) screening for lung cancer is controversial, currently under study, and not yet fully elucidated. OBJECTIVES To report findings from initial and 1-year repeat screening low-radiation-dose CT of the chest and 3-year outcomes for 50- to 79-year-old current and ex-smokers in the Pittsburgh Lung Screening Study (PLuSS). METHODS Notified of findings on screening CT, subjects received diagnostic advice from both study and personal physicians. Tracking subjects for up to three years since initial screening, we obtained medical records to document diagnostic procedures, lung cancer diagnoses, and deaths. MEASUREMENTS AND MAIN RESULTS 3,642 and 3,423 subjects had initial and repeat screening. A total of 1,477 (40.6% of 3,624) were told about noncalcified lung nodules on the initial screening and, before repeat screening, 821 (55.6% of 1,477, 22.5% of 3,642) obtained one or more subsequent diagnostic imaging studies (CT, positron emission tomography [PET], or PET-CT). Tracking identified 80 subjects with lung cancer, including 53 subjects with tumor seen at initial screening. In all, 36 subjects (1.0% of the 3,642 screened), referred for abnormalities on either the initial or repeat screening, had a major thoracic surgical procedure (thoracotomy, video-assisted thoracoscopic surgery [VATS], median sternotomy, or mediastinoscopy) leading to a noncancer final diagnosis. Out of 82 subjects with thoracotomy or VATS to exclude malignancy in a lung nodule, 28 (34.1%) received a noncancer final diagnosis. Forty of 69 (58%) subjects with non-small cell lung cancer had stage I disease at diagnosis. CONCLUSIONS Though leading to the discovery of early stage lung cancer, CT screening also led to many diagnostic follow-up procedures, including major thoracic surgical procedures with noncancer outcomes.

Automated lung segmentation in X-ray computed tomography

RATIONALE AND OBJECTIVES To develop and evaluate a reliable, fully-automated lung segmentation scheme for application in X-ray computed tomography. MATERIALS AND METHODS The automated scheme was heuristically developed using a slice-based, pixel-value threshold and two sets of classification rules. Features used in the rules include size, circularity, and location. The segmentation scheme operates slice-by-slice and performs three key operations: (1) image preprocessing to remove background pixels, (2) computation and application of a pixel-value threshold to identify lung tissue, and (3) refinement of the initial segmented regions to prune incorrectly detected airways and separate fused right and left lungs. RESULTS The performance of the automated segmentation scheme was evaluated using 101 computed tomography cases (91 thick slice, 10 thin slice scans). The 91 thick cases were pre- and post-surgery from 50 patients and were not independent. The automated scheme successfully segmented 94.0% of the 2,969 thick slice images and 97.6% of the 1,161 thin slice images. The mean difference of the total lung volumes calculated by the automated scheme and functional residual capacity plus 60% inspiratory capacity was -24.7 +/- 508.1 mL. The mean differences of the total lung volumes calculated by the automated scheme and an established, commonly used semi-automated scheme were 95.2 +/- 52.5 mL and -27.7 +/- 66.9 mL for the thick and thin slice cases, respectively. CONCLUSION This simple, fully-automated lung segmentation scheme provides an objective tool to facilitate lung segmentation from computed tomography scans.

Effect of Cardiopulmonary Bypass on Early Graft Dysfunction in Clinical Lung Transplantation

The records of 100 lung transplant recipients (13 heart-lungs, 45 double-lungs, and 42 single-lungs) from September 1990 through April 1992 were reviewed to determine the role of cardiopulmonary bypass (CPB) in early graft dysfunction. Fifty-five patients requiring CPB (CPB group) for 186 +/- 54 minutes were compared with the 45 patients without CPB (no-CPB group). All of the heart-lung and en-bloc double-lung transplantations were performed under CPB, with pulmonary vascular lung disease the principal diagnosis, resulting in a significantly younger age population in the CPB group. All other donor- and recipient-related factors matched well in both groups. Of 38 bilateral single-lung transplantations, CPB was used in 18. In double-lung and heart-lung recipients gas exchange of the allografts was evaluated by the arterial/alveolar oxygen tension ratios at nine intervals during the first 72 hours. The mean arterial/alveolar oxygen tension ratio in the CPB group was 0.48 +/- 0.19, significantly lower than in the no-CPB group with 0.60 +/- 0.22 (p = 0.025). All patients had radiographic interpretation and scoring of pulmonary infiltrates from chest roentgenograms taken within 12 hours after reperfusion. The CPB group had more severe pulmonary infiltrates than the no-CPB group (p = 0.034). Prolonged intubation defined as 7 days or longer occurred significantly more often (29/55) in the CPB group than in the no-CPB group (8/45) (p = 0.003). Actuarial graft and patient survival at 1 month was better in the no-CPB group than in the CPB group (42/45 versus 44/55 [p = 0.05] and 43/45 versus 45/55 [p = 0.033], respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Non-cardiac findings on coronary electron beam computed tomography scanning.

The objective of this study was to estimate the prevalence and significance of non-cardiac findings on Electron Beam Computed Tomography (EBT) scanning when used in population screening for the quantitative measurement of coronary artery calcium and estimate of coronary risk. Clinic files of 1366 subjects who underwent EBT scanning between September 1996 and December 1998 at the University of Pittsburgh affiliated Comprehensive Heart Care Center were abstracted. The files of 1356 subjects contained the calcium score and non-cardiac findings as reported by board-certified radiologists, who interpreted the scans during the period 1996–1998. A National Death Index (NDI) Plus match was performed to ascertain cause of death. Two hundred seventy-eight of 1356 (20.5%) subjects had 1 or more non-cardiac findings on EBT scanning. Fifty-seven of 1356 (4.2%) received a recommendation for diagnostic CT follow-up. Forty-six of the 57 recommendations were for pulmonary nodules and 11 were for non-nodule, non-cardiac findings. Seven members of the cohort died during a short follow-up period. In 1 case, the non-cardiac finding was the cause of death. Non-cardiac findings in a healthy cohort referred for EBT coronary screening are relatively common. Findings range from clinically insignificant to the cause of death during a short follow-up period. EBT scanning is a frequently used coronary screening procedure. With the relatively high detection of significant, non-cardiac pathology in this increasingly common screening procedure, consideration should be given for radiologists to interpret the scans.

Radiographic Emphysema Predicts Low Bone Mineral Density in a Tobacco-exposed Cohort

RATIONALE Studies demonstrating an association between chronic obstructive pulmonary disease and low bone mineral density (BMD) implicate factors distinct from treatments and severity of lung disease in the pathogenesis of osteoporosis. Whereas emphysema has been independently associated with vascular disease and other comorbidities, its association with BMD has not been well studied. OBJECTIVES We explored the associations of BMD with computed tomography (CT) measures of emphysema and other risk factors in current and former smokers. METHODS One hundred ninety subjects completed a CT scan, pulmonary function testing, questionnaires, and dual x-ray absorptiometry measurements of hip and lumbar spine BMD. Subjects were classified as having normal BMD, osteopenia, or osteoporosis. Demographic, physiologic, and radiographic characteristics were compared and the association of BMD with radiographic emphysema, airflow obstruction, and osteoporosis risk factors was assessed. MEASUREMENTS AND MAIN RESULTS No difference existed in age, tobacco exposure, oral steroid use, or physical activity across BMD categories. Both osteopenia and osteoporosis were associated with the presence of airflow obstruction, inhaled corticosteroid use, and female sex, and demonstrated a significant relationship with the presence of visual emphysema (P = 0.0003). Quantitative emphysema, but not CT-measured indices of airway wall thickness, was inversely associated with BMD. Visual emphysema alone was a significant predictor of osteopenia/osteoporosis (odds ratio = 2.55; 95% confidence interval, 1.24-5.25) in a model including obstruction severity, age, sex, and inhaled and oral steroid use. CONCLUSIONS Radiographic emphysema is a strong, independent predictor of low BMD in current and former smokers. This relationship suggests a common mechanistic link between emphysema and osteopenia/osteoporosis.