Overexpression of CEACAM6 activates Src-FAK signaling and inhibits anoikis, through homophilic interactions in lung adenocarcinomas.

Among carcinoembryonic antigen-related cell adhesion molecule (CEACAM) family proteins, CEACAM6 has received less attention than CEACAM5 and its presence and role in lung cancer are largely unknown. The application of CellphoneDB on the single cell RNA sequencing dataset showed that the homophilic interactions among CEACAM6 molecules, which are overexpressed in lung cancer cells were highly significant. CEACAM6 was overexpressed in 80.1% of lung adenocarcinomas and its overexpression had a significant relationship with non-smoking history and activating EGFR mutations. The effect of CEACAM6 overexpression on patient prognosis was evaluated using TCGA-LUAD dataset; the CEACAM6 overexpression group showed a shorter overall survival than that of the control group when matched for stage, age, sex, and pack-years. Immunoblotting of cell culture soup and ELISA of human derived material suggested that the majority of CEACAM6 was present on the cancer cell surface and interacted with other cancer cells in the crowded tumor microenvironment. Treatment with CEACAM6 showed CEACAM6 homophilic interactions in the cell membrane and anoikis inhibition through the activation of the Src-FAK pathway. Inhibition of CEACAM6 or its homophilic interactions in the cancer cell membrane may provide another therapeutic strategy for lung cancer.

Early lung carcinogenesis and tumor microenvironment observed by single-cell transcriptome analysis.

With the increasing interest in health screening with chest CT Ground-glass nodule (GGN) has become one of the common lung lesions encountered in daily medical practice. Because lung adenocarcinoma in the form of GGN is an ideal model for studying early lung carcinogenesis, 11 GGN and normal lung specimens from 6 never smoker patients were studied by single-cell RNA sequencing. Lung cancer cells showed enrichment of gene sets related to small vesicle processing and surfactant homeostasis compared to non-malignant lung epithelial cells, suggesting the dysregulation of surfactant pathway may be involved in early lung carcinogenesis. Along with cancer-associated fibroblasts showing enrichment of gene sets involved in negative regulation of protein kinase activity and negative regulation of endothelial cell proliferation, tumor microenvironment (TME) was dominated by infiltration of TNFRSF4+/TNFRSF18+/CTLA4+ regulatory T cells (Treg) and depletion of CD8+ cytotoxic T cells (TC) and γδTC. Majority of mucosa-associated lymphoid tissue B cells (BCs) and follicular BCs were detected within tumor tissue, which was associated with CXCL13 overexpressed in intratumoral Tregs and CD4+ memory TCs. Coordination of components of the TME towards immune evasion is governed by Tregs from the onset of lung cancer, requiring unremitting efforts to target and overcome them. This provision of information on changes in cancer cell-specific biomarkers and TME using early lung cancer from never smokers will provide new insight into early lung carcinogenesis and useful targets for treatment.

Three-Dimensional Multistack-Printed, Self-Powered Flexible Pressure Sensor Arrays: Piezoelectric Composites with Chemically Anchored Heterogeneous Interfaces.

Recently, the development of pressure sensor devices composed of mechanically flexible materials has gained a tremendous attention for emerging wearable electronics applications. Compared with various sensing materials, piezoelectric composite materials provide a characteristic advantage of enabling energy unit-free integration of sensor compartments. In this study, we develop a new chemical method of synthesizing highly functioning piezoelectric composite materials with electrostatically reinforced heterogeneous interfaces to improve the voltage output signal in all-printed sensor arrays. The surfaces of piezoelectric oxide nanoparticles are decorated subsequently with a cationic polyelectrolyte, polyethyleneimine, and a tri-block copolymer, styrene-ethylene/butylene-styrene grafted with maleic anhydride. To elucidate the factors determining the performance of pressure sensor devices, both the electrical properties and piezoelectric characteristics are investigated comprehensively for various compositional composite materials prepared from chemical and physical rubbers. The resulting device exhibits a sensitivity of 0.28 V·kPa-1 with a linear increment of output voltage in a pressure range up to 30 kPa. It is also demonstrated that the all-printed sensor array is fabricated successfully by a multistack-printing process of conductive, insulating, and piezoelectric composite materials in an additive manufacturing fashion.

Development of organoid-based drug metabolism model.

Cytochrome P450 (CYP) gene superfamily catalyzes oxidative metabolism of a wide variety of drugs, carcinogens, and endogenous biomolecules in the liver and intestinal organs. In vitro assay platforms such as primary hepatocyte and immortalized liver-derived cell lines have been developed to evaluate drug effects. However, several limitations have been suggested regarding discrepancies between in vitro and in vivo assays. In this study, we aimed to investigate drug metabolism and toxicity based on mouse small intestinal and liver organoids derived from resident stem cells. At first, expressions and activities of CYP subfamilies (CYPs) in intestinal and liver organoids were investigated. Organoids treated with three CYPs-inducers dexamethasone (Dex), ß-naphthoflavone (BNF), and 1,4-bis-2-(3, 5-dichloropyridyloxy)-benzene (TCPOBOP) were evaluated for CYPs activities. The CYPs-induced intestinal and liver organoids were confirmed to digest more docetaxel, as colon cancer cell-line survived more in CYPs-induced organoid's medium than in non-induced organoid's medium. Then, the activity of docetaxel in a co-culture platform of mouse liver organoids and human pancreatic tumoroids was measured. We obtained significant statistical values on CYPs-induced metabolic activities: cell survival rates of pancreatic tumoroids co-cultured with docetaxel-treated undifferentiated, differentiated, and CYPs-induced differentiated organoids were 66.05 ±â€¯2.14%, 89.20 ±â€¯2.67%, and 101.90 ±â€¯0.94%, respectively. To sum up, gene expression modification and drug metabolism evaluation were able to be done with organoids as done with tissues. In vivo-like in vitro investigation on drug toxicity may potentially be done with organoids as a stepping bridge to the clinical trial.

Development of Collagen-Based 3D Matrix for Gastrointestinal Tract-Derived Organoid Culture.

Organoid is a cell organization grown in a three-dimensional (3D) culture system which represents all characteristics of its origin. However, this organ-like structure requires supporting matrix to maintain its characteristics and functions. Matrigel, derived from mouse sarcoma, has often been used as the supporting matrix for organoids, but the result may not be desirable for clinical applications because of the unidentified components from the mouse sarcoma. On the other hand, natural characteristics of collagen emphasize toxic-free friendly niche to both organoid and normal tissue. Hence, this study attempts to develop a new, collagen-based matrix that may substitute Matrigel in organoid culture. Collagen-based matrix was made, using type 1 collagen, Ham's F12 nutrient mixture, and bicarbonate. Then, characteristics of mouse colon organoids were analyzed by morphology and quantitative messenger RNA (mRNA) expression, revealing that the mouse colon organoids grown in the collagen-based matrix and in Matrigel had quite similar morphology, specific markers, and proliferative rates. Mouse small intestine-derived organoids, stomach-derived organoids, and human colon-derived organoids were also cultured, all of which were successfully grown in the collagen-based matrix and had similar properties compared to those cultured in Matrigel. Furthermore, possibility of organoid transplantation was observed. When mouse colon organoids were transplanted with collagen matrix into the EDTA-colitis mouse model, colon organoids were successfully engrafted in damaged tissue. For that reason, the use of collagen-based matrix in organoid culture will render organoid cultivation less expensive and clinically applicable.

In vivo evaluation of scaffolds compatible for colonoid engraftments onto injured mouse colon epithelium.

Colon organoids (colonoids) are known to be similar to colon tissue in structure and function, which makes them useful in the treatment of intestinal de-epithelialized disease. Matrigel, which is used as a transplantation scaffold for colonoids, cannot be used in clinical applications because of its undefined composition and tumorigenicity. This study identifies clinically available scaffolds that are effective for colonoid transplantation in damaged intestinal mucosa. The colon crypt was isolated and cultured from C57BL/6-Tg[CAG enhanced green fluorescent protein (EGFP)131Osb/LeySopJ mice into EGFP + colonoids and subsequently transplanted into the EDTA colitis mouse model using gelatin, collagen, or fibrin glue scaffolds. To identify scaffolds suitable for colonoid engraftment in injured colon mucosa, the success rates of transplantation and secondary EGFP colonoid formation were measured, and the scaffolds' mediated toxicity in vitro and in vivo was observed in recipient mice. When colonoids were transplanted with gelatin, collagen, and fibrin glue into the EDTA colitis mouse model, all groups were found to be successfully engrafted. Fibrin glue, especially, showed significant increase in the engrafted area compared with Matrigel after 4 wk. The scaffolds used in the study did not induce colonic toxicity after transplantation into the recipients' colons and were thus deemed safe when locally administrated. This study suggests new methods for and provides evidence of the safety and utility of the clinical application of colonoid-based therapeutics. Furthermore, the methods introduced in this study will be helpful in developing cell treatment using the esophagus or a stomach organoid for various digestive-system diseases.-Jee, J., Jeong, S. Y., Kim, H. K., Choi, S. Y., Jeong, S., Lee, J., Ko, J. S., Kim, M. S., Kwon, M.-S., Yoo, J. In vivo evaluation of scaffolds compatible for colonoid engraftments onto injured mouse colon epithelium.

Leucine-rich repeat-containing G-protein coupled receptor 5 enriched organoids under chemically-defined growth conditions.

An organoid is a complex, multi-cell three-dimensional (3D) structure that contains tissue-specific cells. Epithelial stem cells, which are marked by leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5), have the potential for self-renewal and expansion as organoids. However, in the case of intestinal organoids from Lgr5-EGFP-IRES-CreERT2 transgenic mice, in vitro expansion of the Lgr5 expression is limited in a culture condition supplemented with essential proteins, such as epidermal growth factor (E), noggin (N), and R-spondin 1 (R). In this study, we hypothesized that self-renewal of Lgr5+ stem cells in a 3D culture system can be stimulated by defined compounds (CHIR99021, Valproic acid, Y-27632, and A83-01). Our results demonstrated that dissociated single cells from organoids were organized into a 3D structure in the four compounds containing the ENR culture medium in a 3D and two-dimensional (2D) culture system. Moreover, the Lgr5 expression level of organoids from the ENR- and compound-containing media increased. Furthermore, the conversion of cultured Lgr5+ stem cells from 2D to 3D was confirmed. Therefore, defined compounds promote the expansion of Lgr5+ stem cells in organoids.