Episodic Aspiration with Oral Commensals Induces a MyD88-dependent, Pulmonary T-Helper Cell Type 17 Response that Mitigates Susceptibility to Streptococcus pneumoniae.

Rationale: Cross-sectional human data suggest that enrichment of oral anaerobic bacteria in the lung is associated with an increased T-helper cell type 17 (Th17) inflammatory phenotype.Objectives: In this study, we evaluated the microbial and host immune-response dynamics after aspiration with oral commensals using a preclinical mouse model.Methods: Aspiration with a mixture of human oral commensals (MOC; Prevotella melaninogenica, Veillonella parvula, and Streptococcus mitis) was modeled in mice followed by variable time of killing. The genetic backgrounds of mice included wild-type, MyD88-knockout, and STAT3C backgrounds.Measurements and Main Results: 16S-rRNA gene sequencing characterized changes in microbiota. Flow cytometry, cytokine measurement via Luminex and RNA host-transcriptome sequencing was used to characterize the host immune phenotype. Although MOC aspiration correlated with lower-airway dysbiosis that resolved within 5 days, it induced an extended inflammatory response associated with IL-17-producing T cells lasting at least 14 days. MyD88 expression was required for the IL-17 response to MOC aspiration, but not for T-cell activation or IFN-γ expression. MOC aspiration before a respiratory challenge with S. pneumoniae led to a decrease in hosts' susceptibility to this pathogen.Conclusions: Thus, in otherwise healthy mice, a single aspiration event with oral commensals is rapidly cleared from the lower airways but induces a prolonged Th17 response that secondarily decreases susceptibility to S. pneumoniae. Translationally, these data implicate an immunoprotective role of episodic microaspiration of oral microbes in the regulation of the lung immune phenotype and mitigation of host susceptibility to infection with lower-airway pathogens.

T Stage and Pretreatment Standardized Uptake Values Predict Tumor Recurrence With 5-Fraction SABR in Early-Stage Non-Small Cell Lung Cancer.

Purpose: Five-fraction stereotactic ablative radiotherapy (SABR) regimens are frequently used to treat centrally located early-stage non-small cell lung cancer or disease in the proximity of the chest wall as a means of optimizing tumor control and reducing treatment toxicity. However, increasing these SABR regimens to 5 fractions may reduce tumor control outcomes. We sought to identify the clinical parameters predictive of treatment failures with these 5-fraction courses. Methods: Ninety patients with T1-2 non-small cell lung cancer were treated with 50 or 60 Gy in 5 fractions. Failure over time was modeled using cumulative incidences of local, regional, or distant failure, with death as a competing risk. Cox proportional hazards analysis for incidences of failure was performed to control for patient variables. Results: Of 90 patients, 24 of 53 patients with T1 tumors and 19 of 37 patients with T2 tumors received 50 Gy SABR, and the other 47 patients received 60 Gy. Two-year overall survival and progression-free survival for the whole cohort were 75.8% and 59.3%, respectively. Total SABR dose (50 vs 60 Gy) did not influence survival nor failure rates at 2 and 5 years. Within 2 years of treatment, 7.8% of all patients developed local failure. For all patient and tumor characteristics evaluated, only T stage and pretreatment positron emission tomography standardized uptake values served as predictors of local, regional, and distant failure at 2 and 5 years posttreatment on univariate and multivariable analysis. Conclusions: Five-fraction SABR provides excellent in-field control. T2 and high fluorodeoxyglucose uptake tumors have increased failure rates, suggesting the potential need for adjuvant therapies, which are being assessed in randomized phase 3 trials.

Lower Airway Dysbiosis Affects Lung Cancer Progression.

In lung cancer, enrichment of the lower airway microbiota with oral commensals commonly occurs, and ex vivo models support that some of these bacteria can trigger host transcriptomic signatures associated with carcinogenesis. Here, we show that this lower airway dysbiotic signature was more prevalent in the stage IIIB-IV tumor-node-metastasis lung cancer group and is associated with poor prognosis, as shown by decreased survival among subjects with early-stage disease (I-IIIA) and worse tumor progression as measured by RECIST scores among subjects with stage IIIB-IV disease. In addition, this lower airway microbiota signature was associated with upregulation of the IL17, PI3K, MAPK, and ERK pathways in airway transcriptome, and we identified Veillonella parvula as the most abundant taxon driving this association. In a KP lung cancer model, lower airway dysbiosis with V. parvula led to decreased survival, increased tumor burden, IL17 inflammatory phenotype, and activation of checkpoint inhibitor markers. SIGNIFICANCE: Multiple lines of investigation have shown that the gut microbiota affects host immune response to immunotherapy in cancer. Here, we support that the local airway microbiota modulates the host immune tone in lung cancer, affecting tumor progression and prognosis.See related commentary by Zitvogel and Kroemer, p. 224.This article is highlighted in the In This Issue feature, p. 211.

Organotypic culture in three dimensions prevents radiation-induced transformation in human lung epithelial cells.

The effects of radiation in two-dimensional (2D) cell culture conditions may not recapitulate tissue responses as modeled in three-dimensional (3D) organotypic culture. In this study, we determined if the frequency of radiation-induced transformation and cancer progression differed in 3D compared to 2D culture. Telomerase immortalized human bronchial epithelial cells (HBECs) with shTP53 and mutant KRas expression were exposed to various types of radiation (gamma, (+)H, (56)Fe) in either 2D or 3D culture. After irradiation, 3D structures were dissociated and passaged as a monolayer followed by measurement of transformation, cell growth and expression analysis. Cells irradiated in 3D produced significantly fewer and smaller colonies in soft agar than their 2D-irradiated counterparts (gamma P = 0.0004; (+)H P = 0.049; (56)Fe P < 0.0001). The cell culture conditions did not affect cell killing, the ability of cells to survive in a colony formation assay, and proliferation rates after radiation-implying there was no selection against cells in or dissociated from 3D conditions. However, DNA damage repair and apoptosis markers were increased in 2D cells compared to 3D cells after radiation. Ideally, expanding the utility of 3D culture will allow for a better understanding of the biological consequences of radiation exposure.

CDDO-Me protects normal lung and breast epithelial cells but not cancer cells from radiation.

Although radiation therapy is commonly used for treatment for many human diseases including cancer, ionizing radiation produces reactive oxygen species that can damage both cancer and healthy cells. Synthetic triterpenoids, including CDDO-Me, act as anti-inflammatory and antioxidant modulators primarily by inducing the transcription factor Nrf2 to activate downstream genes containing antioxidant response elements (AREs). In the present series of experiments, we determined if CDDO-Me can be used as a radioprotector in normal non-cancerous human lung and breast epithelial cells, in comparison to lung and breast cancer cell lines. A panel of normal non-cancerous, partially cancer progressed, and cancer cell lines from both lung and breast tissue was exposed to gamma radiation with and without pre-treatment with CDDO-Me. CDDO-Me was an effective radioprotector when given ∼18 hours before radiation in epithelial cells (average dose modifying factor (DMF) = 1.3), and Nrf2 function was necessary for CDDO-Me to exert these radioprotective effects. CDDO-Me did not protect cancer lines tested from radiation-induced cytotoxicity, nor did it protect experimentally transformed human bronchial epithelial cells (HBECs) with progressive oncogenic manipulations. CDDO-Me also protected human lymphocytes against radiation-induced DNA damage. A therapeutic window exists in which CDDO-Me protects normal cells from radiation by activating the Nrf2 pathway, but does not protect experimentally transformed or cancer cell lines. This suggests that use of this oral available, non-toxic class of drug can protect non-cancerous healthy cells during radiotherapy, resulting in better outcomes and less toxicity for patients.

Chronic stress and a cyclic regimen of estradiol administration separately facilitate spatial memory: relationship with hippocampal CA1 spine density and dendritic complexity.

This study investigated the effects of chronic restraint stress and repeated cyclic estradiol pulses on hippocampal CA3 and CA1 dendritic and/or spine morphology and spatial memory in female rats. Sprague-Dawley adult female rats were ovariectomized and then injected over 2 days with 17ß-estradiol (10 µg, s.c.), which was repeated every 4-5 days. While all rats received similar estradiol injection histories, half of the rats were chronically restrained and/or given a final cyclic pulse of estradiol prior to testing on a hippocampal-dependent object placement (OP) task to assess spatial memory. OP testing was performed 2 days after the last restraint session, as well as when the last 2 estradiol pulses best captured the maximal effect on hippocampal CA1 spine density. The data revealed several novel findings: (a) chronic stress or estradiol separately facilitated spatial memory, but did not have the same effects when coadministered, (b) CA1 spine densities negatively correlated with spatial memory, and (c) repeated estradiol pulses failed to prevent stress-induced CA3 dendritic retraction. We also corroborated previous studies showing increased CA1 spine density following estradiol, chronic stress, and behavioral manipulations. The present study uniquely combined chronic stress, repeated estradiol pulses, hippocampal morphology, and behavior within the same animals, allowing for correlational analyses to be performed between CA1 spine morphology and spatial memory. We demonstrate novel findings that chronic stress or estradiol pulses independently facilitate spatial memory, but not when coadministered, and that these effects may involve a balance of CA1 apical spine expression that is independent of CA3 dendritic complexity.