Lung Microenvironment Research Articles

Lung-derived soluble factors support stemness/plasticity and metastatic behaviour of breast cancer cells via the FGF2-DACH1 axis

Patients with triple-negative breast cancer (TNBC) have an increased propensity to develop lung metastasis. Our previous studies demonstrated that stem-like ALDHhiCD44+ breast cancer cells interact with lung-derived soluble factors, resulting in enhanced migration and lung metastasis particularly in TNBC models. We have also observed that the presence of a primary TNBC tumor can ‘prime’ the lung microenvironment in preparation for metastasis. In this study, we hypothesized that soluble lung-derived factors secreted in the presence of a primary TNBC tumor can influence stemness/plasticity of breast cancer cells. Using an ex vivo pulmonary metastasis assay (PuMA), we observed that the lung microenvironment supports colonization and growth of ALDHhiCD44+ TNBC cells, potentially via interactions with lung-derived FGF2. Exposure of TNBC cells to lung-conditioned media (LCM) generated from mice bearing TNBC primary tumors (tbLCM) significantly enhanced the proportion of ALDHhiCD44+ cells compared to control or LCM from tumor-naïve mice (tnLCM). Further analysis using a human cancer stem cell qPCR array revealed that, relative to tnLCM or control, exposure of TNBC cells to tbLCM leads to downregulation of the transcription factor and putative tumor suppressor Dachshund homolog 1 (DACH1), a downstream regulator of FGF2. In addition, inhibition of DACH1 using siRNA or treatment with recombinant FGF2 enhanced the ALDHhiCD44+ phenotype. Taken together, our findings suggest that the FGF2-DACH1 signaling axis supports stemness/plasticity of TNBC cells in the lung microenvironment and lays the foundation for future evaluation of FGF2 as a potential novel therapeutic target for treatment or prevention of breast cancer metastasis to the lung.

Abstract SY38-02: Clinical investigations of obesity in cancer: BMI and other confounders

Abstract SY38-02: Clinical investigations of obesity in cancer: BMI and other confounders

Schwann cells in the normal and pathological lung microenvironment.

The lungs are a key organ in the respiratory system. They are regulated by a complex network of nerves that control their development, structure, function, and response to various pathological stimuli. Accumulating evidence suggests the involvement of a neural mechanism in different pathophysiological conditions in the lungs and the development and progression of common respiratory diseases. Lung diseases are the chief source of death globally. For instance, lung cancer is the second most commonly diagnosed malignancy, after prostate cancer in men and breast cancer in women, and is the most lethal cancer worldwide. However, although airway nerves are accepted as a mechanistically and therapeutically important feature that demands appropriate emphasizing in the context of many respiratory diseases, significantly less is known about the role of the neuroglial cells in lung physiology and pathophysiology, including lung cancer. New data have uncovered some cellular and molecular mechanisms of how Schwann cells, as fundamental components of the peripheral nervous system, may regulate lung cancer cells' survival, spreading, and invasiveness in vitro and in vivo. Schwann cells control the formation and maintenance of the lung cancer microenvironment and support metastasis formation. It was also reported that the number of lung cancer-associated Schwann cells correlates with patients' survival. Different factors secreted by Schwann cells, including microRNA, are known to sharpen the lung cancer environment by regulating the tumor-neuro-immune axis. Further clinical and experimental studies are required to elucidate the detailed role of Schwann cells in creating and maintaining pulmonary tumor-neuro-immune axis, which will advance our understanding of the pathogenesis of lung cancer and may inform therapeutic hypotheses aiming neoplasms and metastases in the lung.

458 Influence of a Gastrointestinal Infection on Lung Immunity

OBJECTIVES/GOALS: We aim to characterize how Heligmosomoides polygyrus bakeri (H. poly) alleviates murine allergic asthma which shares many characteristics of human asthma. This approach of has already identified helminth-produced human immune cell ligand “mimics” that hold great potential for next-generation clinical biologics METHODS/STUDY POPULATION: We examined the lung tissue of C57BL/6 mice infected with H. poly for changes in the pulmonary microenvironment. At ten days post infection, four infected mice and two co-housed uninfected mice were sacrificed, and their lung tissue harvested for examination of RNA via RT-qPCR. This design allows for the comparison between the lung microenvironments of infected and naīve mice. In future experiments, we intend to characterize what small molecules produced by the helminth drive changes in the lung using germ-free models of H. poly infection. RESULTS/ANTICIPATED RESULTS: We found key differences in lung chemokines between mice infected with H. poly and naīve mice. Using a student t-test with naīve correction for variance, we were able to show significant differences in the expression of E cadherin (p = 0.0355), CXCL10 (p = 0.0025), CX3CL1 (p = 0.0029), CCR2 (p = 0.017), and IDO1 (0.0078). We also found that differences in the expression of CCL5 bordered on significant with a p-value of 0.066. The expression of most of these markers (CXCL10, CCR2, CCL5, and IDO1) was elevated in the lungs of infected mice compared to naīve controls. In contrast, E cadherin and CX3CL1 showed the opposite trend with naīve mice showing greater expression. These clear differences in lung tissue gene expression underscore the connection between the gastrointestinal and pulmonary mucosal immune compartments. DISCUSSION/SIGNIFICANCE: The changes are unexpected for an infection that has been shown to attenuate allergic inflammation in the lung with increases in the IFN-Y responsive genes IDO1 and CXCL10 and inflammatory lung markers, CCL5 and CCR2. In contrast, there were decreases in inflammatory lung marker CX3CL1 and the tight junction protein E cadherin in infected mice.

Osteosarcoma cells exhibit functional interactions with stromal cells, fostering a lung microenvironment conducive to the establishment of metastatic tumor cells.

Osteosarcoma (OS) stands out as the most common bone tumor, with approximately 20% of the patients receiving a diagnosis of metastatic OS at their initial assessment. A significant challenge lies in the frequent existence of undetected metastases during the initial diagnosis. Mesenchymal stem cells (MSCs) possess unique abilities that facilitate tumor growth, and their interaction with OS cells is crucial for metastatic spread. We demonstrated that, in vitro, MSCs exhibited a heightened migration response toward the secretome of non-metastatic OS cells. When challenged to a secretome derived from lungs preloaded with OS cells, MSCs exhibited greater migration toward lungs colonized with metastatic OS cells. Moreover, in vivo, MSCs displayed preferential migratory and homing behavior toward lungs colonized by metastatic OS cells. Metastatic OS cells, in turn, demonstrated an increased migratory response to the MSCs' secretome. This behavior was associated with heightened cathepsin D (CTSD) expression and the release of active metalloproteinase 2 (MMP2) by metastatic OS cells. Our assessment focused on two complementary tumor capabilities crucial to metastatic spread, emphasizing the significance of inherent cell features. The findings underscore the pivotal role of signaling integration within the niche, with a complex interplay of migratory responses among established OS cells in the lungs, prometastatic OS cells in the primary tumor, and circulating MSCs. Pulmonary metastases continue to be a significant factor contributing to OS mortality. Understanding these mechanisms and identifying differentially expressed genes is essential for pinpointing markers and targets to manage metastatic spread and improve outcomes for patients with OS.

A multi-omics investigation of the lung injury induced by PM2.5 at environmental levels via the lung-gut axis

Long-term exposure to fine particulate matter (PM2.5) posed injury for gastrointestinal and respiratory systems, ascribing with the lung-gut axis. However, the cross-talk mechanisms remain unclear. Here, we attempted to establish the response networks of lung-gut axis in mice exposed to PM2.5 at environmental levels. Male Balb/c mice were exposed to PM2.5 (dose of 0.1, 0.5, and 1.0 mg/kg) collected from Chengdu, China for 10 weeks, through intratracheally instillation, and examined the effect of PM2.5 on lung functions of mice. The changes of lung and gut microbiota and metabolic profiles of mice in different groups were determined. Furthermore, the results of multi-omics were conjointly analyzed to elucidate the primary microbes and the associated metabolites in lung and gut responsible for PM2.5 exposure. Accordingly, the cross-talk network and key pathways between lung-gut axis were established. The results indicated that exposed to PM2.5 0.1 mg/kg induced obvious inflammations in mice lung, while emphysema was observed at 1.0 mg/kg. The levels of metabolites guanosine, hypoxanthine, and hepoxilin B3 increased in the lung might contribute to lung inflammations in exposure groups. For microbiotas in lung, PM2.5 exposure significantly declined the proportions of Halomonas and Lactobacillus. Meanwhile, the metabolites in gut including L-tryptophan, serotonin, and spermidine were up-regulated in exposure groups, which were linked to the decreasing of Oscillospira and Helicobacter in gut. Via lung-gut axis, the activations of pathways including Tryptophan metabolism, ABC transporters, Serotonergic synapse, and Linoleic acid metabolism contributed to the cross-talk between lung and gut tissues of mice mediated by PM2.5. In summary, the microbes including Lactobacillus, Oscillospira, and Parabacteroides, and metabolites including hepoxilin B3, guanosine, hypoxanthine, L-tryptophan, and spermidine were the main drivers. In this lung-gut axis study, we elucidated some pro- and pre-biotics in lung and gut microenvironments contributed to the adverse effects on lung functions induced by PM2.5 exposure.

Abstract 5507: High-resolution spatial atlas reveals insight into spatial landscape of lung cancer and chronic lung diseases

Abstract Background: Patients with lung cancer often have a high prevalence of severe comorbidities, primarily due to the significant correlation with cigarette smoking and the aging process. Dysregulations of the immune response are the cause of chronic inflammatory diseases such as chronic obstructive pulmonary disease (COPD), pulmonary arterial hypertension (PAH), pulmonary fibrosis (PF) and lung cancer. In addition, the lung microenvironment in these chronic inflammatory diseases has not yet been sufficiently investigated. Therefore, we aimed to investigate the spatio-temporal localization and cellular distribution in the lung microenvironment and to compare lung cancer with the concomitant diseases of lung cancer. Methods: FFPE lung sections, each 3µm in thickness, were collected from patients with diagnoses of lung cancer, COPD, PAH, PF, and from healthy donors, with three samples taken for each category. These sections were subsequently imaged using the PhenoCycler®-Fusion system, utilizing a comprehensive panel of 45 antibodies. including immune, epithelial, vascular, proliferation and apoptosis markers. The first step of the analysis workflow consists of cell segmentation using a fine-tuned deep learning model leading to a total of 5.2 million cells across the whole dataset. Protein expressions were then calculated from the segmented cells, and unsupervised clustering was performed based on the normalized expression values. The resulting clusters were manually annotated into 16 cell phenotypes based on their protein expression patterns as displayed on a hierarchical clustering heatmap. The percentages of cell phenotypes were then calculated for each sample and compared between disease groups. Furthermore, the spatial proximity and cellular neighborhood analyses were performed to study the spatial organization of the different cell phenotypes in the tissues. Results: Quantitative assessment revealed unique characteristics specific to each disease, identified by the varying abundance of different immune cell subtypes. Additionally, when comparing lungs from healthy donors with those affected by lung diseases, notable differences were observed in the spatial arrangement of immune cells and their proximity to lung blood vessels. Conclusions: This study presents the first architectural map of lung cancer, COPD, PAH, and PF linking various immune cell types to their spatial location within the tissue. Such in-depth spatial analyses will aid in comprehending the spatial and temporal interplay within the lung's microenvironment in diverse lung conditions, as well as in developing targeted therapies for specific cell types. Citation Format: Rajender Nandigama Nandigama, Bassem Ben Cheikh Cheikh, Jamal Nabhanizadeh, Friedrich Grimminger, Werner Seeger, Niyati Jhaveri, Soni Savai Pullamsetti, Rajkumar Savai. High-resolution spatial atlas reveals insight into spatial landscape of lung cancer and chronic lung diseases [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5507.

Abstract 5537: How aging promotes immune mediated reactivation from metastatic melanoma dormancy

Abstract Disseminated cancer cells that escape the primary tumor can seed distal tissues but may take several years to grow into clinically detectable metastases, a phenomenon termed tumor dormancy. Despite its importance in metastasis and residual disease, few studies have been able to successfully model melanoma dormancy. We have previously shown that while tumors grow more slowly in aged mouse skin, they form more lung metastases in aged mice compared to young mice. We queried whether the lung microenvironment undergoes age-related changes that establish a permissive niche for metastatic outgrowth. We identified that Wnt5A and AXL are not only required for efficient dissemination away from the primary tumor but also activate dormancy of melanoma cells within the lung. The young lung microenvironment maintains melanoma cells in a dormant state; however, age-associated reprograming of lung fibroblasts increases secretion of the soluble Wnt antagonist sFRP1, which inhibits both Wnt5A and AXL to enable reactivation from dormancy and the formation of larger metastatic colonies in the aged lung. We found that downregulation of AXL in melanoma cells leads to an increase in the related TAM family receptor MERTK, which promotes a proliferative melanoma phenotype and enables reactivation from dormancy. We have now found that both the aged-pre and post metastatic niche regulate immune cells in the lung to create a permissive niche for reactivation of metastatic melanoma. NK cells are significantly decreased in the healthy lungs of aged mice, are significantly less active, and have increased exhaustion markers. NK cell infiltration remains low in early and late-stage metastasis in aged mice. Depletion of NK cells in young mice is sufficient to induce reactivation from dormancy. Further, MERTK induced reactivation of melanoma cells results in paracrine secretion of its own ligand PROS1 to facilitate a growth permissive lung microenvironment. Melanoma-secreted PROS1 increases infiltration of PMN-MDSCs and decreases macrophage infiltration to maintain an immunosuppressive lung niche for efficient outgrowth. Selective inhibition of PMN-MDSCs using a TRIAL-R2 agonistic antibody significantly decreases metastatic outgrowth and promotes smaller, non-proliferative dormant colonies. Our findings indicate a significant role that aging plays in altering NK-cell infiltration and function to create a growth permissive pre-metastatic niche and how reactivation from dormancy can further contribute to create an immunosuppressive microenvironment for efficient outgrowth via PROS1 induced infiltration of PMN-MDSCs. Importantly, the clear differences found in our young and aged mouse modelling systems for outgrowth in metastatic tissues necessitates the need for age-appropriate pre-clinical modelling for better patient and disease representation. Citation Format: Pulkit Datt, Jhon Passamonte, Christopher Price, Daniel Zabransky, Yash Chhabra, Ashani Weeraratna, Mitchell E. Fane. How aging promotes immune mediated reactivation from metastatic melanoma dormancy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5537.

Abstract 5527: Spatiotemporal analysis of growth factor regulation of osteosarcoma AKT and ERK signaling dynamics in an ex vivo lung metastasis model

Abstract In the present study, we investigated the relationships between spatiotemporal distribution of growth factors in the lung microenvironment, metastatic osteosarcoma cell signaling, and intratumoral gene expression heterogeneity. Osteosarcoma is an aggressive bone malignancy that commonly metastasizes to the lungs, resulting in significant patient mortality. During metastasis, disseminated cancer cells are exposed to microenvironment-derived growth factors that result in signaling pathway activation. Two such pathways are MAPK/ERK and PI3K/AKT, which regulate complex transcriptional programs underlying cell proliferation and survival decisions. In this study, we utilized precision cut lung slices (PCLS) obtained from mice with established osteosarcoma metastases to ex vivo model associations between the spatiotemporal distributions of growth factors in the lung, osteosarcoma signaling dynamics, and intratumoral gene expression heterogeneity, with single cell precision. To accomplish this, we engineered osteosarcoma cells that co-expressed genetically-encoded ERK and AKT sensors, engrafted them into PCLS tissues, then recorded sensor dynamics via live-cell microscopy. These tissues were subsequently fixed and immuno-stained for measurement of growth factor expression in resident lung cells and spatial correlation with observed tumor cell ERK-AKT signaling dynamics. Using this approach, we found that growth factor expression was diffusely increased within areas bordering osteosarcoma metastases. FGF2 and IGF1, which we have previously demonstrated to potently induce ERK signaling in osteosarcoma cells, were both significantly upregulated in these regions. However, despite relatively uniform overexpression of growth factors adjacent to tumors, individual osteosarcoma cells displayed stochastics patterns of ERK and AKT signaling, and downstream gene targets. These findings revealed that upregulation of lung growth factors, and the inherent systems-level properties of osteosarcoma signaling networks, can converge to promote intratumoral signaling and gene expression heterogeneity. Together, our results, using a novel PCLS and live-cell biosensor imaging approach, provide new insight into how spatial and temporal changes in microenvironmental signaling composition are integrated by metastatic osteosarcoma cells to promote heterogeneity. Citation Format: Rawan Makkawi, Ryan D. Roberts, Jeremy Copperman, Alexander E. Davies. Spatiotemporal analysis of growth factor regulation of osteosarcoma AKT and ERK signaling dynamics in an ex vivo lung metastasis model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5527.

Abstract 5516: Aberrant activation and perpetuation of lung wound response facilitates osteosarcoma metastasis

Abstract Introduction: The mechanisms that allow tumor cells to integrate with unfamiliar parenchymal cells within hostile distant tissues during metastatic colonization remain poorly defined. Nowhere are these mechanisms more important than in osteosarcoma-a pediatric bone tumor where metastases cause nearly all osteosarcoma related deaths and occur almost exclusively in the lung. The purpose of this study was to define the cellular and molecular events that facilitate osteosarcoma lung colonization to devise novel therapeutic strategies that prevent and treat metastasis. Experimental Approach: We generated experimental lung metastases using a panel of immunocompetent murine and human xenograft osteosarcoma models. A combination of immunohistochemistry, single cell RNA sequencing, and spatial transcriptomics were used to define the metastatic niche. Relevant findings were validated in human primary tumor, lung metastasis, and normal lung tissue specimens. Results: Disseminated osteosarcoma cells physically associate with alveolar epithelial cells and trigger changes in surrounding epithelial cells that are characteristic of the aberrant, non-resolving wound response seen in non-malignant lung diseases such as idiopathic pulmonary fibrosis (IPF). This process drives the accumulation of fibrotic epithelial cells and scar-associated macrophages confined within a dense fibronectin-based matrix. Anti-fibrotic tyrosine kinase inhibitors used to treat patients with IPF (nintedanib) prevent the observed fibrogenic changes and inhibit metastatic colonization. Epithelial-osteosarcoma paracrine signaling is dominated by an inflammatory cytokine milieu that facilitates this transition towards fibrosis. Blocking one of the dominant paracrine signals with a recombinant IL1 receptor antagonist (anakinra) has similar effects on both fibrosis and metastatic colonization. Conclusions: Our study demonstrates that tumor interactions with host stromal cells are critical for metastasis. Metastatic colonization requires transformation of the local lung microenvironment into a fibrotic, non-resolving, cytokine-rich, wound-like state that is both provoked and sustained by tumor-epithelial interactions. These insights have identified several potential therapeutic approaches to prevent metastatic colonization and treat established metastases. Citation Format: James Brandon Reinecke, Amy C. Gross, Maren Cam, Leyre Jimenez Garcia, Matthew V. Cannon, Ryan D. Roberts. Aberrant activation and perpetuation of lung wound response facilitates osteosarcoma metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5516.

Abstract 3934: Smoking's influence on divergent paths to lung cancer

Abstract Lung cancer is the leading cause of cancer mortality in the world. Many studies have highlighted how mutagens from cigarette smoke promote specific mutations such as G12 mutations in KRAS, contributing to clonal evolution that leads to lung cancer. Despite the indisputable correlation between smoking and lung cancer, people who have never smoked account for the 7th leading cause of cancer mortality. Given the differences in predominant exposures, we hypothesize that the clonal evolution between people with and without smoking histories are markedly different. Therefore, we analyzed lung samples from people with different cigarette smoking exposure to elucidate smoking’s effect on the somatic mutational landscape in the progression to lung cancer. To observe somatic mutations subjected to different exposures of smoking, we used a rare-mutation detection technique called DuplexSeq to analyze somatic variants from 200+ lung samples. We acquired DNA from cells from bronchoscopy brushings and lung punches of histologically normal tissue from people with and without lung cancer and who currently, formerly, or never smoked cigarettes. Through collaborations, we characterized mutations using bioinformatic pipelines, various databases, and statistical methods. In samples from people who smoke, we observe an enhancement of mutations with higher variant allele frequencies and pervasive positive selection acting on mutations in many cancer-associated genes. From our bronchoscopy brushings, NOTCH1 mutations show the most definitive increase of protein-altering variants, while NF1 and KRAS show a predominance of driver mutation in lung punches. Unexpectedly, people without lung cancer who never smoked had a notable presence of pathogenic mutations in PTEN and PIK3CA. Furthermore, multiple punches from the same individual highlight how expansions of mutations have stronger similarity within an individual than across individuals regardless of smoking status. This association points to the importance of the inherent lung microenvironment in influencing the mutational landscapes in lung cancer progression. Altogether, these results show how smoking increases the oncogenicity of observable mutations in the lung but does not preclude the presence of oncogenic clones, emphasizing the value of precision diagnoses and prevention. Ongoing research will be centered around functional assays to determine the impact of observed mutations on tissue function and longitudinal studies using mutational landscapes as a proxy for lung cancer risk. Citation Format: Edward James Evans, Fabio Marongiu, Emilia Lim, Oriol Pich, Yonghua Zhuang, Dexiang Gao, William Hill, Cian Murphy, Ryan Turner, Moumita Ghosh, York Miller, Mariam Jamal-Hanjani, Charles Swanton, James DeGregori. Smoking's influence on divergent paths to lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3934.

Abstract 5517: Chronic stress causes metastasis via neutrophil-mediated changes to the microenvironment

Abstract Chronic stress is associated with increased risk of metastasis and poor survival in cancer patients, yet the reasons are unclear. We show that chronic stress increased lung metastasis from disseminated cancer cells 2- to 4-fold in mice. Chronic stress significantly altered the lung microenvironment, with fibronectin accumulation, reduced T cell infiltration, and increased neutrophil infiltration. Depleting neutrophils abolished stress-induced metastasis, indicating their critical role. Chronic stress shifted neutrophils’ normal circadian rhythm, and, via glucocorticoid release, caused increased neutrophil extracellular trap (NET) formation. Importantly, in mice with neutrophil-specific glucocorticoid receptor deletion, chronic stress failed to increase NETs and metastasis. Furthermore, digesting NETs with deoxyribonuclease (DNase) I prevented chronic stress-induced metastasis. Together, our data show that glucocorticoids released during chronic stress cause NET formation and establish a metastasis-promoting microenvironment. Therefore, NETs could be targets for preventing metastatic recurrence in cancer patients, many of whom will experience chronic stress due to their disease. Citation Format: Xue-Yan He, Yuan Gao, David Ng, Evdokia Michalopoulou, Shanu George, Jose M. Adrover, Lijuan Sun, Jean Albrengues, Juliane Daßler-Plenker, Xiao Han, Ledong Wan, Xiaoli S. Wu, Longling S. Shui, Yu-Han Huang, Bodu Liu, Chang Su, David L. Spector, Christopher R. Vakoc, Linda VanAelst, Mikala Egeblad. Chronic stress causes metastasis via neutrophil-mediated changes to the microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5517.

Regulatory T cells: Supporting lung homeostasis and promoting resolution and repair after lung injury

Regulatory T cells, characterized by their expression of the transcription factor Forkhead box P3, are indispensable in maintaining immune homeostasis. The respiratory system is constantly exposed to many environmental challenges, making it susceptible to various insults and infections. Regulatory T cells play essential roles in maintaining homeostasis in the lung and promoting repair after injury. Regulatory T cell function dysregulation can lead to inflammation, tissue damage, or aberrant repair. Research on regulatory T cell mechanisms in the lung has unveiled their influence on lung inflammation and repair mechanisms. In this review, our goal is to highlight the advances in regulatory T cell biology with respect to lung injury and resolution. We further provide a perspective that a deeper understanding of regulatory T cell interactions in the lung microenvironment in health and disease states offers opportunities for therapeutic interventions as treatments to promote lung health.

The activation of the AIM2 inflammasome after cigarette smoke exposure leads to an immunosuppressive lung microenvironment

Cigarette smoke is widely known as contributing to chronic inflammation underlying several airway diseases, such as chronic obstructive pulmonary disease (COPD) and lung cancer. In our previous studies we found that the lung of both COPD and cancer patients were characterized by the presence and activation of the AIM2 inflammasome. Here, we wanted to investigate the upstream step during the establishment of chronic lung inflammation after cigarette smoke exposure. We took advantage of a mouse model of smoking exposure and public scRNAseq data. We found that AIM2 mRNA was expressed in both alveolar type II, B cells, T regulatory (Treg) and macrophages detected in the lung of non-smokers (n = 4) and smokers (n = 3). The activation of AIM2 in smoking mice by using PolydA:dT did not alter cigarette-smoke-induced alveoli enlargement and mucus production, rather it induced higher recruitment of immunosuppressive cells, such as non-active dendritic cells (DCs), Arginase I+ macrophages, myeloid-derived suppressor cells (MDSC) and Tregs. In addition, the inflammatory environment after AIM2 activation in smoking mice was characterized by higher levels of IL-1α, IL-1β, IL-33, TNFα, LDH, IL-10 and TGFβ. This scenario was not altered after the pharmacological inhibition of both caspase-1 and STING pathway.In conclusion, these data suggest that chronic inflammation after cigarette smoke exposure is associated with AIM2 activation, which could lead towards cigarette smoke-associated lung diseases.

Galectin-3 levels in children with cystic fibrosis.

Cystic fibrosis (CF) is a multisystemic disease in which airway obstruction, infection, and inflammation play a critical role in the pathogenesis and progression of CF lung disease. The carbohydrate-binding protein Galectin-3 is increased in several inflammatory and fibrotic diseases and has recently been forwarded as a biomarker in these diseases. We aimed to define the role of serum Galectin-3 in children with CF by comparison with healthy subjects. This is a cross-sectional, case-control study. 143 CF and 30 healthy subjects were enrolled in the study. Peripheral blood and sputum concentrations of Galectins-3, interleukin (IL)-17A, IL-8, and neutrophil elastase (NE) were determined with commercial ELISA kits. There was no significant difference between the groups in age and gender (p = 0.592, p = 0.613, respectively). Serum Galectin-3 and NE concentrations were higher in the patient group than in healthy controls (p = 0.002, p < 0.001, respectively). There were no significant differences between groups according to IL-17A and IL-8 concentrations. Serum Galectin-3 was correlated with age (r = 0.289, p < 0.001) and body mass index (BMI) (r = 0.493, p < 0.001) in children with CF. Sputum Galectin-3 levels are negatively correlated with percent predictive forced expiratory volume in 1s (FEV1) (r = - 0.297, p = 0.029), FEV1 z-score, (r = - 0.316, p = 0.020), percent predictive forced vital capacity (FVC) (r = - 0.347, p = 0.010), and FVC z-score (r = - 0.373, p = 0.006). Conclusion: The study shows that serum Galectin-3 levels increased in clinically stable CF patients, and serum Galectin-3 response may depend on age, gender, and BMI. The sputum Galectin-3 was found to be negatively correlated with patients' lung functions. What is known: • Galectin-3 is a key regulator of chronic inflammation in the lung, liver, kidney, and tumor microenvironment. What is new: • Children with cystic fibrosis (CF) have higher serum Galectin-3 concentrations than healthy children. • Serum Galectin-3 expression influenced by age, BMI, and gender in children with CF.

Immunomodulatory effects of BAFF and APRIL cytokines in post-pulmonary infection lung cancer: Implications for drug resistance and progression.

Lung cancer is a complicated and challenging disease and is one of the most common causes of cancer-related mortality worldwide. Within the lung microenvironment, specific cytokines, including the B cell activation factor (BAFF) and the A proliferation-inducing ligand (APRIL), are produce by various cells, notably airway epithelial cells, in response allergic inflammation or pulmonary infection. These cytokines play a critical role in maintaining local immune responses and fostering the survival of immune cells. The BAFF and APRIL system have been connected in a range of malignancies and have shown their potential in inducing drug resistance and promoting cancer progression. This review highlights recent studies on the involvement of BAFF and APRIL in various cancers, focusing mainly on their role in lung cancer, and discusses the possibility of these molecules in contributing to drug resistance and cancer progression following pulmonary infection. We suggest consideration the targeting BAFF and APRIL or their respective receptors as promising novel therapies for effective treatment of lung cancer, especially post pulmonary infection. However, it remains important to conduct further investigations to fully elucidate the precise mechanisms underlying how the BAFF and APRIL systems enhance cancer survival and drug resistance subsequent pulmonary infections.

Chronic stress increases metastasis via neutrophil-mediated changes to the microenvironment

Chronic stress is associated with increased risk of metastasis and poor survival in cancer patients, yet the reasons are unclear. We show that chronic stress increases lung metastasis from disseminated cancer cells 2- to 4-fold in mice. Chronic stress significantly alters the lung microenvironment, with fibronectin accumulation, reduced Tcell infiltration, and increased neutrophil infiltration. Depleting neutrophils abolishes stress-induced metastasis. Chronic stress shifts normal circadian rhythm of neutrophils and causes increased neutrophil extracellular trap (NET) formation via glucocorticoid release. In mice with neutrophil-specific glucocorticoid receptor deletion, chronic stress fails to increase NETs and metastasis. Furthermore, digesting NETs with DNase I prevents chronic stress-induced metastasis. Together, our data show that glucocorticoids released during chronic stress cause NET formation and establish a metastasis-promoting microenvironment. Therefore, NETs could be targets for preventing metastatic recurrence in cancer patients, many of whom will experience chronic stress due to their disease.

Immunological Landscapes in Lung Transplantation: Insights from T Cell Profiling in BAL and PBMC.

Lung transplant recipients frequently encounter immune-related complications, including chronic lung allograft dysfunction (CLAD). Monitoring immune cells within the lung microenvironment is pivotal for optimizing post-transplant outcomes. This study examined the proportion of T cell subsets in paired bronchoalveolar lavage (BAL) and peripheral PBMC comparing healthy (n = 4) and lung transplantation patients (n = 6, no CLAD and n = 14 CLAD) using 14-color flow cytometry. CD4+ T cell proportions were reduced in CD3 cells in both PBMC and BAL, and positive correlations were discerned between T cell populations in peripheral PBMC and BAL, suggesting the prospect of employing less invasive PBMC sampling as a means of monitoring lung T cells. Furthermore, regulatory T cells (Tregs) were enriched in BAL when compared to peripheral PBMC for transplant recipients. A parallel positive correlation emerged between Treg proportions in BAL and peripheral PBMC, underscoring potential avenues for monitoring lung Tregs. Finally, the most promising biomarker was the Teff (CD8+Granzyme B+)-Treg ratio, which was higher in both the PBMC and BAL of transplant recipients compared to healthy individuals, and increased in the patients with CLAD compared to no CLAD and healthy patients. Conclusions: Distinct T cell profiles in BAL and peripheral PBMC underscore the significance of localized immune monitoring in lung transplantation. The Teff (CD8+granzyme B+)-Treg ratio, particularly within the context of CLAD, emerges as a promising blood and BAL biomarker reflective of inflammation and transplant-related complications. These findings emphasize the imperative need for personalized immune monitoring strategies that tailored to address the unique immunological milieu in post-transplant lungs.

Human Breast Cancer Cell Lines Differentially Modulate Signaling from Distant Microenvironments, Which Reflects Their Metastatic Potential.

Metastasis is the stage at which the prognosis substantially decreases for many types of cancer. The ability of tumor cells to metastasize is dependent upon the characteristics of the tumor cells, and the conditioning of distant tissues that support colonization by metastatic cells. In this report, we investigated the systemic alterations in distant tissues caused by multiple human breast cancer cell lines and the impact of these alterations on the tumor cell phenotype. We observed that the niche within the lung, a common metastatic site, was significantly altered by MDA-MB-231, MCF7, and T47 tumors, and that the lung microenvironment stimulated, to differing extents, an epithelial-to-mesenchymal transition (EMT), reducing proliferation, increasing transendothelial migration and senescence, with no significant impact on cell death. We also investigated the ability of an implantable scaffold, which supports the formation of a distant tissue, to serve as a surrogate for the lung to identify systemic alterations. The scaffolds are conditioned by the primary tumor similarly to the lung for each tumor type, evidenced by promoting a pro-EMT profile. Collectively, we demonstrate that metastatic and non-metastatic breast cancers condition distant tissues, with distinct effects on tumor cell responses, and that a surrogate tissue can distinguish the metastatic potential of human breast cancer cell lines in an accessible site that avoids biopsy of a vital organ.

Exploring the novel duo of Reticulocalbin, and Sideroflexin as future biomarker candidates for Exacerbated Chronic Obstructive Pulmonary Disease

BackgroundCOPD is a complex respiratory disorder with high morbidity and mortality rates. Even with the current conventional diagnostic methods, including circulating inflammatory biomarkers, underdiagnosis rates in COPD remain as high as 70%. Our study was a comparative cross-sectional study that aimed to address the diagnostic challenges by identifying future biomarker candidates in COPD variants.MethodsThis study used a label-free plasma proteomics approach that combined mass spectrometric data with bioinformatics to shed light on the functional roles of differentially expressed proteins in the COPD lung microenvironment. The predictive capacity of the screened proteins was assessed using Receiver Operating Characteristic (ROC) curves, with Western blot analysis validating protein expression patterns in an independent cohort.ResultsOur study identified three DEPs—reticulocalbin-1, sideroflexin-4, and liprinα-3 that consistently exhibited altered expression in COPD exacerbation. ROC analysis indicated strong predictive potential, with AUC values of 0.908, 0.715, and 0.856 for RCN1, SFXN4, and LIPα-3, respectively. Validation through Western blot analysis confirmed their expression patterns in an independent validation cohort.ConclusionsOur study discovered a novel duo of proteins reticulocalbin-1, and sideroflexin-4 that showed potential as valuable future biomarkers for the diagnosis and clinical management of COPD exacerbations.