Faculty Bibliography

Community-acquired pneumonia affects approximately 4 million people in the United States, with 40,000 deaths per year. The incidence is increased about 35-fold in HIV-infected individuals, and this rate has decreased since the antiretroviral era has begun. Bacterial pneumonia has decreased from 5 to 20 cases per 100 person-years to less than 1 to 5 cases per 100 person-years in the era of antiretroviral therapy. HIV-1 infection impairs the function of neutrophils in the lung and infects CD4(+) cells and alveolar macrophages. Opportunistic infections dramatically increase local HIV replication in the lung cells, especially alveolar macrophages and CD4(+) cells. This enhanced replication increases viral mutations and provides opportunities for viral escape from latent reservoirs. Mortality is increased with more comorbidities in this highly susceptible population. Immunization with vaccines is recommended, especially pneumococcal vaccines, although the vaccine itself may stimulate viral replication. Recent studies show that the lower respiratory tract is a microbial reservoir in HIV-infected individuals rather than being a sterile environment, as originally thought. This may provide new opportunities for preventing opportunistic infections in HIV-infected subjects. Bacterial pneumonia presents an ongoing challenge in these high-risk individuals, particularly in studying the functions of the innate and acquired immune response

Background: In the 7 years after the WTC disaster, the annual number of FDNY retirements attributable to pulmonary disability increased nearly 4-fold. FDNY rescue workers evaluated for pulmonary disability at our subspecialty center were previously found to have an obstructed phenotype based on FEV₁/FVC, Methacholine Challenge Test (MCT) reactivity, or bronchodilator (BD) reactivity (59% of N=1720). In this pilot study, we examined the long-term efficacy of treatment and its effect on disease progression in a subgroup (N=450) of individuals with subspecialty pulmonary testing who were referred for disability evaluation before and after a course of treatment. Methods: Our study cohort includes individuals that were symptomatic, had subspecialty pulmonary testing (pre-treatment), and were referred for disability evaluation after a full course of treatment (post-treatment). Disability evaluations were performed 30 days after weaning from inhaled medication. Individuals were excluded if their post-treatment PFT was less than three months after starting treatment. Change in lung function over time and post treatment were analyzed (N=415). Values are presented as medians with IQR. We defined statistical significance as P<0.05 using the paired t-test. Analyses were performed using GraphPad and SPSS software. Results: Individuals evaluated by MCT presented for disability evaluation earlier, with a shorter interval of treatment (7.5 months (5-10)) compared to those that received spirometry with BD, (14.5 months (8-25) p<0.0001). FEV₁ %Pred and FEV₁/FVC declined significantly; p<0.0001. FEV₁ observed declined at a slower rate in the treatment group than in the entire WTC-exposed FDNY cohort, including those not referred for subspecialty evaluation: 15cc/year vs. 26cc/year. After treatment course, BD response was unchanged, but methacholine reactivity improved significantly; Fig. 1. (Figure presented) Airway Hyperreactivity Post-Treatment. (A): BD response did not improve from pre- to post-treatment, (N=135). Shading represents a positive BD response. (B): Methacholine reactivity significantly improved, from failing at a cumulative dose of 1.4 mg/mL to failing at a cumulative dose of 64 mg/mL (N=35). There was no difference between individuals receiving bronchodilator response and methacholine challenge in FEV1 or FEV1/FVC both pre- and post treatment, the decline FEV1 from pre- to post-Treatment, time to entry into treatment cohort, or demographics (years of service, exposure, age at 9/11, or BMI). Conclusion: Treatment with removal from fire fighting, inhaled steroids and beta agonists did not improve spirometric measures, or BD response, but slowed the rate of decline and decreased hyper-reactivity to the MCT. Improvement in reactivity defined by MCT but not defined by BD response suggests that the pathways leading to BD response and MCT reactivity may be different and may respond differently to treatment

RATIONALE: In the first year post 9/11/2001, there was a decline of 439 mL in FEV1 in FDNY rescue workers, stabilizing to a 25 ml/year decline in the subsequent seven years. Airflow obstruction predominated in 1720 individuals who sought a subspecialty pulmonary evaluation for treatment. We noted that the most severely obstructed patients had significant weight gain. This well phenotyped cohort had serum drawn within the five months post 9/11/2001, allowing us to investigate relationships of metabolic syndrome biomarkers and decline in lung function. METHODS: The treatment cohort (N=1720) was stratified by FEV1 into either obstructed, FEV1<76% predicted (Lower limit of normal for this cohort), or normal airflow, FEV1>76% (Figure 1A). A pilot analysis assayed 41 patients' serum drawn 2.88+/-0.99 months following 9/11 for 15 biomarkers of metabolic syndrome by Luminex; 10 cases were obstructed and 31 were normal. All patients had normal pre-9/11 lung function. Serum cholesterol and triglycerides were available on 157 patients, 20/157 were obstructed. The subspecialty PFT was used to stratify patients, months post-9/11. Data represented as means +/-SD and significance was assessed based on p<=0.05 by t-test. RESULTS: BMIs at the time of serum sampling were no different between individuals with obstruction and normal FEV1. However, at subspecialty PFT, the obstructed patients had higher BMIs, (Figure 1B). The cohort with the greatest lung function decline had significantly greater cholesterol and Cholesterol/HDL ratios, with a trend towards elevated triglyceride levels, (Figure 1C-E). The subgroup with available Luminex data showed higher levels of sE-Selectin, tPAI-1, and s-ICAM in the obstructed patients, (Figure 2A-C). The same cohort had a trend towards elevated levels of C-peptide (Figure 2D). Individuals with obstruction at treatment entry had accelerated decline in lung function post 9/11, increased airway reactivity, and evidence of air trapping based on elevated RV when compared to those who maintained a normal FEV1%. (Figure presented) CONCLUSIONS: Blood drawn within 5 months of WTC exposure identified a subgroup of patients with markers of metabolic syndrome. This subgroup had more weight gain and greater decline in lung function during the next 7 years. The finding of metabolic syndrome biomarkers prior to decline in lung function raises the possibility that the combination of irritant exposure and mediators of metabolic syndrome interact and promote lung injury

BACKGROUND: Pulmonary function declined among 13,234 WTC exposed FDNY personnel in the first year after 9/11/2001. In subsequent years, the rate of decline in FEV1 returned to a group's mean about 25ml/yr (Aldrich et. al 2010). The subgroup who presented for treatment frequently had airway obstruction (Weiden et al. 2010) with some patients having accelerated decline of lung function compared to the total cohort. IgA and IgG4 mediate mucosal immunity while IgG3 drives lung inflammation. Serum from 2001-2002 was stored and tested for immunoglobulin expression to identify biomarkers that predict accelerated decline in lung function in WTC exposed individuals. METHODS: We conducted a pilot investigation of serum antibody expression in 40/1720 firefighters who received treatment for pulmonary complaints. All patients have at least 3 pulmonary function tests available for the study: (1) pre 9/11 (2) 3 months post 9/11 and (3) upon entry into treatment (32 months post 9/11/2001). All of these patients had normal pre-9/11 lung function (Fig. 1). At treatment entry, 9/40 patients had abnormal FEV1 (Mean 58 +/-13% predicted) and 31/40 had normal FEV1 (92 +/- 11% predicted). The serum samples, drawn 3 months post 9/11 were analyzed for IgM, IgA, IgE and IgG1 through IgG4 profiles by using Luminex multiplex technology. Immunoglobulins expression was then correlated with changes in FEV1 from the PFT done soon after 9/11/2001 to the PFT done at the time of presentation for treatment. RESULTS: In the serum collected prior to development lung function abnormalities, patients who maintained normal lung function have different Ig levels than those who developed airflow obstruction. Patients who went on to develop abnormal lung function post 9/11 had reduced IgA (p=0.06), IgG4 (p<0.0005) levels and increased IgG3 (p<0.01) level compared to those who maintained normal lung function (Fig. 2). There were no differences in IgM, IgG1, IgG2, or IgE levels. There were no significant differences between two groups in FEV1 prior to 9/11, age, BMI, exposure intensity status. 9 patients with abnormal lung function had accelerated decline in lung function post 9/11, increased airway reactivity and evidence of air trapping when compared to those who maintained normal lung function. (Figure presented) CONCLUSIONS: Reduced levels of IgG4 and IgA are biomarkers for future decline in lung function after an irritant exposure. Since IgG4 and IgA mediate mucosal immunity deficiency in mucosal immune response may predispose to non-resolving inflammation and progressive lung injury after an irritant exposure

Introduction: Macrolides are associated with a significant reduction in exacerbations compared with placebo in both chronic obstructive lung disease and cystic fibrosis. However, this may be due anti-bacterial and/or anti-inflammatory effects. The mechanism of the anti-inflammatory effects of macrolides in the lung are poorly understood. Alveolar macrophages (AM) have a central role on innate immune homeostasis of the lung; AM have developed many mechanisms to prevent autonomous induction and excess production of inflammatory cytokines through an increased expression of immune inhibitors such as C/EBPbeta and MafB. Interferon induces inhibitory C/EBPbeta, is regulated by MafB and has anti-inflammatory effect in the lung. Here, we test the hypothesis that azithromycin suppresses innate immune response by induction of inhibitory transcription factors. Methods: Alveolar macrophages were obtained from five subjects and incubated with lipopolysaccharide (LPS), interferon (IFN)-alpha, azithromycin (AZM) and AZM+LPS for 24hrs. Thirty nine supernatant cytokines were measured using Luminex. Protein extracts were obtained from AM and used for immunoblots for C/EBPbeta and MafB. Results: LPS strongly induced TNF-alpha production by AM, while the combination AZM and LPS inhibited LPS induced TNF-alpha production (Figure 1, p=0.02). AZM by itself had minimal effect. AZM also inhibited LPS induced CCL22, MDC, IL-1b, IL-6, IL10, G-GSF and GM-CSF. The inhibitory C/EBPbeta transcription factor and MafB expression was induced when AZM was added with LPS (Figure 2). Conclusions: AZM inhibits inflammatory cytokine production after LPS stimulation and induces inhibitory C/EBPbeta and MafB in AM. We will investigate if induction of innate immune inhibitors that blocks TLR signaling pathways underlies the broad anti-inflammatory effect achieved by macrolides. (Figure presented)

Introduction:Regulatory T cells (Treg) play important roles in immunological self-tolerance, and are functionally immunosuppressive subsets of T cells. Th17 cells are critical in the defense against microbes, particularly at mucosal surfaces. It has been shown that the balance between Treg and Th17 cells is a key factor that regulates helper T-cell (CD4⁺ or Th) function. However, there is limited information on the balance between Treg and Th17 cells in bronchoalveolar lavage (BAL). We investigated the distribution of Th17 cells in relation to Treg in PBMC and BAL of healthy volunteers, emphysema patients and HIV-1 infected patients. Methods:BAL lymphocytes were obtained by plating BAL cells for >=1 hour to remove adherent alveolar macrophages. PBMC were isolated by Ficoll gradient fractionation. BAL and PB lymphocytes were stimulated with PMA and ionomycin, treated with monensin, permeabilized, then labeled with PerCP-anti-CD4, PE-anti-IL-17 and Alexa Fluor 647 anti-Foxp3, and analyzed on a FACSCalibur to determine the percentage of CD4⁺ lymphocytes expressing IL-17 and Foxp3 (mean+/-SD). Results:Ten subjects were available for analysis (2 healthy volunteers, 5 emphysema and 3 HIV-1 infected subjects). Both PB and BAL Th cells presented a wide range of FoxP3⁺ cells (19.2+/-18.6 and 6.3+/-5.9 respectively) and IL-17⁺ cells (9.5+/-8.1 and 8.8+/-4.6 respectively). There was a greater percentage of FoxP3⁺ in the HIV-infected group compared to the healthy volunteer and emphysema groups (11.7+/-6.4 vs. 1.7+/-1.1 and 4.9+/-4.7 respectively). Similarly, there was a trend towards higher percentage of IL-17⁺ cells in the BAL of HIV-infected subjects compared to the healthy volunteer and emphysema subjects (10.8+/-3.6 vs. 5.6+/-0.9 and 8.9+/-5.8 respectively). When the expression of FoxP3⁺ and IL-17⁺ cells in BAL was compared for each subject, a direct correlation with an r2 of 0.36 and a p-value of 0.06 was found Conclusions:We observed a relative preservation of FoxP3⁺ and IL-17⁺ cells in BAL CD4⁺ lymphocytes, despite prior evidence suggesting a preferential loss of both Treg and Th17 subsets in PBMC of HIV-1 infected subjects. The co-ordinated expression of these subsets in BAL Th cells warrants further investigation to evaluate its immune significance in the alveolar compartment

Alveolar macrophages (AMs) are exposed to respirable microbial particles. Similar to phagocytes in the gastrointestinal tract, AMs can suppress inflammation after exposure to nonpathogenic organisms. IL-1R-associated kinase-M (IRAK-M) is one inhibitor of innate immunity, normally suppressing pulmonary inflammation. During pneumonia, polymorphonuclear neutrophils (PMNs) are recruited by chemotactic factors released by AMs to produce an intense inflammation. We report that intact IRAK-M is strongly expressed in resting human AMs but is cleaved in patients with pneumonia via PMN-mediated induction of caspase-6 (CASP-6) activity. PMN contact is necessary and PMN membranes are sufficient for CASP-6 induction in macrophages. PMNs fail to induce TNF-alpha fully in macrophages expressing CASP-6 cleavage-resistant IRAK-M. Without CASP-6 expression, PMN stimulation fails to cleave IRAK-M, degrade IkappaBalpha, or induce TNF-alpha. CASP-6(-/-) mice subjected to cecal ligation and puncture have impaired TNF-alpha production in the lung and decreased mortality. LPS did not induce or require CASP-6 activity demonstrating that TLR2/4 signaling is independent from the CASP-6 regulated pathway. These data define a central role for CASP-6 in PMN-driven macrophage activation and identify IRAK-M as an important target for CASP-6. PMNs de-repress AMs via CASP-6-mediated IRAK-M cleavage. This regulatory system will blunt lung inflammation unless PMNs infiltrate the alveolar spaces

OBJECTIVES/SPECIFIC AIMS: The first year post 9/11/2001, the FEV1 of FDNY rescue workers declined 439 mL, stabilizing to a 25 ml/yr decline in the subsequent 7 years. Airflow obstruction predominated in firefighters who sought a subspecialty pulmonary evaluation for treatment. We are investigating the relationship between biomarkers of metabolic syndrome (MS) and decline in lung function. METHODS/ STUDY POPULATION: Treatment cohort (N = 1720) was stratified by FEV1 into obstructed, FEV1 < 76% predicted (LLN), or normal airflow, FEV1>76%. A pilot analysis assayed 41 patients' serum drawn 5 months post 9/11 for 15 biomarkers of MS by Luminex, (obstructed N = 10, normal N = 31). All patients had normal pre-9/11 lung function. Serum cholesterol (CHOL) and triglycerides (TG) were available on 157 patients, 20/157 were obstructed. Data presented as means +/- SD; p values -0.05 by t-test considered significant. RESULTS/ANTICIPATED RESULTS: BMIs at time of serum sampling were no different between normal and obstructed individuals. At subspecialty PFT, obstructed patients had higher BMIs with an accelerated decline in lung function post 9/11, increased airway reactivity, and evidence of air trapping based on elevated RV when compared to normals. Obstructed subjects had significantly greater CHOL and CHOL/HDL ratios; higher levels of sE-Selectin, tPAI-1, and s-ICAM; and a trend towards elevated levels of TG and C-peptide. DISCUSSION/SIGNIFICANCE OF IMPACT: Blood drawn post-WTC exposure identified a subgroup of patients with markers of MS. This subgroup had subsequent increased weight gain and decline in lung function. The finding of MS biomarkers prior to lung function decline raises the possibility that the combination of irritant exposure and mediators of MS interact and promote lung injury

BACKGROUND: The World Trade Center (WTC) collapse produced a massive exposure to respirable particulates in New York City Fire Department (FDNY) rescue workers. This group had spirometry examinations pre-September 11, 2001, and post-September 11, 2001, demonstrating declines in lung function with parallel declines in FEV(1) and FVC. To date, the underlying pathophysiologic cause for this has been open to question. METHODS: Of 13,234 participants in the FDNY-WTC Monitoring Program, 1,720 (13%) were referred for pulmonary subspecialty evaluation at a single institution. Evaluation included 919 full pulmonary function tests, 1,219 methacholine challenge tests, and 982 high-resolution chest CT scans. RESULTS: At pulmonary evaluation (median 34 months post-September 11, 2001), median values were FEV(1) 93% predicted (interquartile range [IQR], 83%-101%), FVC 98% predicted (IQR, 89%-106%), and FEV(1)/FVC 0.78 (IQR, 0.72-0.82). The residual volume (RV) was 123% predicted (IQR, 106%-147%) with nearly all participants having normal total lung capacity, functional residual capacity, and diffusing capacity of carbon monoxide. Also, 1,051/1,720 (59%) had obstructive airways disease based on at least one of the following: FEV(1)/FVC, bronchodilator responsiveness, hyperreactivity, or elevated RV. After adjusting for age, gender, race, height and weight, and tobacco use, the decline in FEV(1) post-September 11, 2001, was significantly correlated with increased RV percent predicted (P < .0001), increased bronchodilator responsiveness (P < .0001), and increased hyperreactivity (P = .0056). CT scans demonstrated bronchial wall thickening that was significantly associated with the decline in FEV(1) post-September 11, 2001 (P = .024), increases in hyperreactivity (P < .0001), and increases in RV (P < .0001). Few had evidence for interstitial disease. CONCLUSIONS: Airways obstruction was the predominant physiologic finding underlying the reduction in lung function post-September 11, 2001, in FDNY WTC rescue workers presenting for pulmonary evaluation