Project description:Our goal and objective of this study is to identify genetic alterations using RNA-seq in a lung cancer organoid (tumoroid) line that we generated in our hospital.
Project description:The biggest problem with lung cancer organoid (LCO) is that most lung cancer organoids are derived from surgical specimens of early-stage lung cancer patients. In fact, patients who initially need chemotherapy are patients with advanced lung cancer who cannot undergo a surgery. So, making lung organoids from biopsy specimens successfully is an urgent task. The success rate of culturing LCO from biopsy tissues is very low because conventional lung biopsies such as transthoracic needle biopsy and forcep biopsy only get a small amount of lung tissue. The possibility of critical complications such as bleeding and pneumothorax makes it difficult to obtain enough specimens. Overgrowth of normal lung cells in late passages is also a critical problem in LCO and optimized culture conditions for LCO remain to be identified. To overcome the hurdles of lung cancer organoid, we made LCOs from cryobiopsy specimens of all stages of lung cancer patients. Transbronchial cryobiopsy can obtain over 10 times greater volume of tissue compared to bronchoscopic forceps biopsy and it can be applied to even peripheral lesions with radial endobronchial ultrasonography. The success rate of LCO culture was also markedly improved and recapitulated the characteristics of primary tumors. LCOs derived from cryobiopsy specimens can overcome the critical limitations of present lung cancer organoids. We expect that cryobiopsy will be a breakthrough strategy of clinical application of LCO in all stages of lung cancer.
Project description:Genes differentially expressed among cells constituting an in vitro human lung carcinogenesis model consisting of normal, immortalized, transformed and tumorigenic bronchial epithelial cells were identified. The differentially expressed genes were then analyzed to determine their relevance to the gene expression patterns of clinical non-small cell lung cancer (NSCLC) samples as well as the clinical outcome of patients with this disease. Experiment Overall Design: Total RNA was extracted from two different replicate culture dishes for each cell line. Differentially expresed genes between the tumorigenic 1170-I lung epithelial cells and the NHBE normal cells that also displayed gradual modulation in expression across the entire in vitro model were determined. The differentially expressed genes were further analyzed by functional pathways analysis and integrated (as gene signatures) with gene expression data of human NSCLC clinical samples obtained from published datasets.
Project description:To more accurately model inhalation toxicity in vitro, we developed a tetra-culture system that combines lung alveolar epithelial cells, endothelial cells, macrophages, and mast cells in a three-dimensional orientation. We characterized the influence of the added complexity, using network perturbation analysis and gene expression data, to gain an insight into the steady-state profile of the assembled complete three-dimensional model using all four cell types and of simpler models of one, two, or three of the cell types. Gene expression data were analyzed using cause-and-effect biological network models, together with a quantitative network-scoring algorithm to determine the biological impact of co-culturing the various cell types. In the assembled tetra-culture, macrophages appeared to be the largest contributors to overall network perturbations, promoting high basal levels of oxidative stress and inflammation. This finding led to further optimization of the model using rested macrophages; the addition of rested macrophages decreased the basal inflammatory and cell stress status of the co-culture. Finally, we compared transcriptional profiles from publicly available datasets of conventional in vitro models representative of the airways and of healthy human lung tissues, to assess similarities between our model and other in vitro models and the human lung. On the transcriptional level, we found an increasing correlation between airway models and normal human lung tissue, particularly as cell types became more physiologically relevant and the complexity of the system increased. This indicates that the combination of multiple lung-relevant cell types in vitro does indeed increase similarity to the physiological counterpart.
Project description:We established a micro-patterned respiratory epithelial cell culture system in vitro. In this culture system, various types of lung epithelial cells were identified.
Project description:In this study we describe some of the challenges of clinical metagenomics through benchtop sequencing of two distinct sample types, designed to highlight current application and opportunities in this field.