Faculty Bibliography

Rationale: Exposure to World Trade Center (WTC) dust and fumes is associated with onset of asthma-like respiratory symptoms in exposed community members including local workers, residents and clean-up workers. Although abnormal spirometry measurements are often not detected in these patients, impulse oscillometry (IOS) suggests abnormalities localized to the smaller airways. Peripheral C-reactive protein (CRP) is a marker of systemic inflammation. Since an association between CRP and asthma has been reported, we hypothesized that levels of CRP would be associated with lung function abnormalities as assessed by spirometry and IOS measurements in community members exposed to WTC dust/fumes and gasses. Methods: The WTC Environmental Health Center (EHC) is a treatment program for community members, with presumed WTC-related symptoms. Patients undergo a standardized evaluation including questionnaires and routine blood work, spirometry and IOS measurements. Between 10/1/2009 and 9/29/2011, a measurement of CRP was included for patients undergoing standardized visits. Measurements of lung function were compared utilizing the Wilcoxon test between subjects with normal vs. elevated CRP and further analyzed using linear regression models with log(CRP) as a continuous predictor. Regression analyses were adjusted for potential confounding factors including Body mass index (BMI), exposure category, and smoking history. Results: 208 WTC-exposed individuals met inclusion criteria. Valid spirometry and IOS data were available in 204 and 189 patients, respectively. Mean age was 49 years, 53% were female. Exposure categories (local workers, clean-up workers, residents) were associated with normal/elevated CRP levels (P=0.01). Smokers had a larger portion of elevated CRP (P=0.048). BMI was higher among the high CRP group (Wilcoxon test, P<0.001). FEV₁ and FVC were lower for the high CRP group (P=0.016, P=0.033). However, CRP level was not associated with the ratio FEV₁/FVC (P=0.58). The IOS measurements (R₅, R_5-20, AX ) were higher (P=0.01, P=0.003, P=.001, respectively) among the high CRP group. Multiple regression analysis confirmed that log(CRP) values were inversely correlated with % of predicted FEV 1 (P=0.009) and positively correlated with log(R₅) (P=0.02) and log(AX) (P=0.0045) after adjustment for log(BMI). Conclusions: Peripheral CRP was negatively correlated with levels of FEV₁ and positively correlated with IOS measurements in community members with WTC dust/gas/fume exposure. These data suggest a relationship between systemic inflammation, as reflected by CRP, and both large and small airway abnormalities in a WTC- exposed population

INTRODUCTION: We have previously shown improvement in spirometry parameters in symptomatic WTC dust exposed community members enrolled in the WTC Environmental Health Center treatment program. Additionally, impulse oscillometry (IOS) has demonstrated evidence for distal lung injury not apparent on spirometry. We hypothesize that longitudinal change of spirometry will differ based on presence or absence of distal airway injury and its response to bronchodilator at baseline. METHODS: 810 patients were identified with more than one spirometry and IOS assessment. IOS parameters included resistance at 5 and 20Hz (R₅ and R₂₀) and frequency dependence of resistance assessed as the difference between these parameters (R_5-20). Linear mixed effects modeling evaluated longitudinal changes in IOS parameters, FVC and FEV₁ for the entire population. Separate models were fit for subgroups categorized based on normal vs. abnormal baseline spirometry and normal vs. abnormal baseline IOS (R₅>3.96 cmH2O/L/s). Analyses were adjusted for confounding factors (age, gender, BMI, race/ethnicity, smoking, exposure category and dust cloud exposure). RESULTS: Mean age was 50yr. Patients were mostly female (52%) and had diverse race/ethnicity. At baseline, mean FVC was 91+/-17% predicted and FEV₁ was 88+/-18% predicted. A normal spirometry pattern was noted in the majority (67%; n=542). Despite normal spirometry, IOS revealed abnormalities in 67% (n=364). Longitudinal analysis of IOS parameters (R₅, R₂₀, R_5-20) over time revealed no significant trends for the entire population and for subgroups categorized by baseline spirometry pattern. In contrast, the longitudinal change in spirometry variables differed based on presence of IOS abnormality. In patients with normal spirometry, FEV₁ increased more rapidly in patients with abnormal baseline IOS compared to those with normal IOS (0.76 vs. 0.52 % predicted/yr; Table 1). For patients with abnormal baseline spirometry, FVC increased more rapidly in the abnormal vs. normal IOS patients (1.73 vs. 1.02 % predicted/yr). Patients with IOS response to bronchodilator (highest quartile for improvement of R₅ post bronchodilator) demonstrated a more rapid longitudinal increase in FEV₁ compared with patients without bronchodilator response (lowest quartile)(0.88 vs. 0.53 % predicted/yr,; Table 2). CONCLUSIONS: Spirometry parameters demonstrated improvement over time, while improvement in IOS parameters was not evident, suggesting potential irreversible injury in the distal lung. However, assessment of baseline distal airway function and its acute response to bronchodilator predicted longitudinal response of spirometry in patients enrolled in a treatment program. (Table Presented)

RATIONALE: Analysis of local in vivo inflammation is relevant to the understanding of pathogenesis and disease progression in emphysema. Bronchial reactivity is an early marker of disease in asthma but the relevance of reactivity to the natural history of emphysema is not understood. We hypothesize that bronchial reactivity is a phenotype of early emphysema that might be related to the degree of inflammation in the lung. METHODS: Normal subjects were enrolled as part of a normal volunteer protocol. Emphysema subjects were identified from the NYU Lung Cancer Biomarker Center CT-scan screening cohort. All patients underwent spirometry, plethysmography, diffusion, and oscillometry, as well as bronchoscopy with bronchoalveolar lavage (BAL). Bronchial reactivity was assessed by changes in FEV1, V50 and R5 . From the BAL fluid, cell count differential was obtained, as well as measurement of 39 cytokines in concentrated BAL fluid with Luminex using Human Cytokine Panel I (Millipore). Results amongst the groups were compared with ANOVA and post-hoc LSD comparison. RESULTS: Twenty patients were available for analysis: Six subjects in the control group, 6 emphysema subjects without bronchial reactivity (BR-), and 8 emphysema subjects with bronchial reactivity (BR+). Baseline demographics and pertinent spirometry/oscillometry are listed in Table 1. Emphysema subjects were all GOLD stage 0 or 1. Post-bronchodilator spirometric and oscillometric parameters were not significantly different between BR- and BR+ emphysema groups. There were 28/39 cytokines with reliably measurable levels. Both emphysema groups had elevated neutrophils and higher degree of inflammation as compared to controls (significant data shown Table 1). However, the BR+ emphysema group evidenced higher degree of neutrophils, IL-6, IL-8, G-CSF, Eotaxin, GRO and Fractalkine as compared with the BR- emphysema group. CONCLUSION: These data suggest that in early emphysema a phenotype of proximal and/or distal bronchial reactivity is associated with an increased degree of inflammation as assessed by neutrophils and in vivo inflammatory cytokines. In contrast with early asthma, the phenotype of bronchial reactivity in early emphysema may be characterized by neutrophilic inflammation produced by increased IL-8 in the lung. The role of IL-6, G-CSF, Eotaxin, GRO and Fractalkine in producing emphysema related bronchial reactivity requires further investigation. (Table Presented)

OBJECTIVE: Patterned photo-stimulation offers a promising path towards the effective control of distributed neuronal circuits. Here, we demonstrate the feasibility and governing principles of spatiotemporally patterned microscopic photo-absorber induced neural-thermal stimulation (PAINTS) based on light absorption by exogenous extracellular photo-absorbers. APPROACH: We projected holographic light patterns from a green continuous-wave (CW) or an IR femtosecond laser onto exogenous photo-absorbing particles dispersed in the vicinity of cultured rat cortical cells. Experimental results are compared to predictions of a temperature-rate model (where membrane currents follow I proportional, variant dT/dt). MAIN RESULTS: The induced microscopic photo-thermal transients have sub-millisecond thermal relaxation times and stimulate adjacent cells. PAINTS activation thresholds for different laser pulse durations (0.02 to 1 ms) follow the Lapicque strength-duration formula, but with different chronaxies and minimal threshold energy levels for the two excitation lasers (an order of magnitude lower for the IR system <50 nJ). Moreover, the empirical thresholds for the CW system are found to be in good agreement with detailed simulations of the temperature-rate model, but are generally lower for the IR system, suggesting an auxiliary excitation mechanism. SIGNIFICANCE: Holographically patterned PAINTS could potentially provide a means for minimally intrusive control over neuronal dynamics with a high level of spatial and temporal selectivity.

When natural photoreception is disrupted, as in outer-retinal degenerative diseases, artificial stimulation of surviving nerve cells offers a potential strategy for bypassing compromised neural circuits. Recently, light-sensitive proteins that photosensitize quiescent neurons have generated unprecedented opportunities for optogenetic neuronal control, inspiring early development of optical retinal prostheses. Selectively exciting large neural populations are essential for eliciting meaningful perceptions in the brain. Here we provide the first demonstration of holographic photo-stimulation strategies for bionic vision restoration. In blind retinas, we demonstrate reliable holographically patterned optogenetic stimulation of retinal ganglion cells with millisecond temporal precision and cellular resolution. Holographic excitation strategies could enable flexible control over distributed neuronal circuits, potentially paving the way towards high-acuity vision restoration devices and additional medical and scientific neuro-photonics applications.

Neuroepithelial cell proliferation must be carefully balanced with the transition to neuroblast (neural stem cell) to control neurogenesis. Here, we show that loss of the Drosophila microRNA mir-8 (the homolog of vertebrate miR-200 family) results in both excess proliferation and ectopic neuroblast transition. Unexpectedly, mir-8 is expressed in a subpopulation of optic-lobe-associated cortex glia that extend processes that ensheath the neuroepithelium, suggesting that glia cells communicate with the neuroepithelium. We provide evidence that miR-8-positive glia express Spitz, a transforming growth factor alpha (TGF-alpha)-like ligand that triggers epidermal growth factor receptor (EGFR) activation to promote neuroepithelial proliferation and neuroblast formation. Further, our experiments suggest that miR-8 ensures both a correct glial architecture and the spatiotemporal control of Spitz protein synthesis via direct binding to Spitz 3' UTR. Together, these results establish glial-derived cues as key regulatory elements in the control of neuroepithelial cell proliferation and the neuroblast transition.

In a recent issue of Neuron, Chen et al. (2013) show that apoptosis is required to ensure the even distribution of a class of retinal ganglion cells (ipRGCs), which sense luminance both intrinsically and through input from rods and cones. Disrupting apoptosis impairs photoentrainment mediated by rods/cones, but not that mediated by ipRGC-expressed melanopsin.

Stochastic mechanisms are sometimes utilized to diversify cell fates, especially in nervous systems. In the Drosophila retina, stochastic expression of the PAS-bHLH transcription factor Spineless (Ss) controls photoreceptor subtype choice. In one randomly distributed subset of R7 photoreceptors, Ss activates Rhodopsin4 (Rh4) and represses Rhodopsin3 (Rh3); counterparts lacking Ss express Rh3 and repress Rh4. In the dorsal third region of the retina, the Iroquois Complex transcription factors induce Rh3 in Rh4-expressing R7s. Here, we show that Ss levels are controlled in a binary on/off manner throughout the retina yet are attenuated in the dorsal third region to allow Rh3 coexpression with Rh4. Whereas the sensitivity of rh3 repression to differences in Ss levels generates stochastic and regionalized patterns, the robustness of rh4 activation ensures its stochastic expression throughout the retina. Our findings show how stochastic and regional inputs are integrated to control photoreceptor subtype specification in the Drosophila retina.

Signaling pathways are often re-used during development in surprisingly different ways. The Hippo tumor suppressor pathway is best understood for its role in the control of growth. The pathway is also used in a very different context, in the Drosophila eye for the robust specification of R8 photoreceptor neuron subtypes, which complete their terminal differentiation by expressing light-sensing Rhodopsin (Rh) proteins. A double negative feedback loop between the Warts kinase of the Hippo pathway and the PH-domain growth regulator Melted regulates the choice between 'pale' R8 (pR8) fate defined by Rh5 expression and 'yellow' R8 (yR8) fate characterized by Rh6 expression. Here, we show that the gene encoding the homolog of human Nuclear respiratory factor 1, erect wing (ewg), is autonomously required to inhibit warts expression and to promote melted expression to specify pR8 subtype fate and induce Rh5. ewg mutants express Rh6 in most R8s due to ectopic warts expression. Further, ewg is continuously required to maintain repression of Rh6 in pR8s in aging flies. Our work shows that Ewg is a critical factor for the stable down-regulation of Hippo pathway activity to determine neuronal subtype fates. Neural-enriched factors, such as Ewg, may generally contribute to the contextual re-use of signaling pathways in post-mitotic neurons.