Role of Oxidative Stress-Dependent C/EBPß Expression on CAF Transformation Inducing HCT116 Colorectal Cancer Cell Progression; Migration and Invasion.

OBJECTIVE: To investigate oxidative stress-related CAF transformation through C/EBPß, which affects CRC progression and may have a potential implication for CRC treatment. METHODS: The conditioned media (CM) from HCT116, CRC cells, was used to activate CCD-18Co, colon fibroblasts, then the ability of activated FBs to induce HCT116 growth and progression was assessed using MTT assay, transwell migration, and matrix invasion assay. Alteration of the cytokine profile and oxidative stress of the activated FBs were studied with cytokine arrays and DCFH-DA assay, respectively. The protein expressions of the CAF markers (α-SMA and FAP) and C/EBPß were investigated with immunofluorescence and western blotting. RESULT: It was found that CM from HCT116 cells induced oxidative stress, change of cytokine profile, CAF markers, and the C/EBPß expression of activated FBs. Furthermore, when the oxidative stress of the activated FBs was suppressed, FAP and C/EBPß expression were downregulated, correlating with the disabling of their capability to support the cancer progression. The C/EBPß and prognosis for CRC patients were accessed using the GEPIA dataset, in which high C/EBPß expression was associated with a poor prognosis. CONCLUSION: These findings suggest that C/EBPß expression has a role in CAF transformation in an oxidative stress-related manner and might be used as a target to improve aggressive CRC treatment outcomes.

Recent Advances in Cell Sheet Engineering: From Fabrication to Clinical Translation.

Cell sheet engineering, a scaffold-free tissue fabrication technique, has proven to be an important breakthrough technology in regenerative medicine. Over the past two decades, the field has developed rapidly in terms of investigating fabrication techniques and multipurpose applications in regenerative medicine and biological research. This review highlights the most important achievements in cell sheet engineering to date. We first discuss cell sheet harvesting systems, which have been introduced in temperature-responsive surfaces and other systems to overcome the limitations of conventional cell harvesting methods. In addition, we describe several techniques of cell sheet transfer for preclinical (in vitro and in vivo) and clinical trials. This review also covers cell sheet cryopreservation, which allows short- and long-term storage of cells. Subsequently, we discuss the cell sheet properties of angiogenic cytokines and vasculogenesis. Finally, we discuss updates to various applications, from biological research to clinical translation. We believe that the present review, which shows and compares fundamental technologies and recent advances in cell engineering, can potentially be helpful for new and experienced researchers to promote the further development of tissue engineering in different applications.

YAP, a novel target regulates F-actin rearrangement-associated CAFs transformation and promotes colorectal cancer cell progression.

Colorectal cancer (CRC) progression is strongly influenced by the tumor microenvironment (TME) in which cancer-associated fibroblasts (CAFs) are the major components influencing CRC growth and progression. The present study aimed to investigate the effect of YAP on F-actin arrangement in CAF transformation and the possibility of using YAP as a target for inhibiting CRC growth and progression. Conditioned media were collected from direct interaction between CRC cells and fibroblasts. CAF markers were investigated by flow cytometry, western blot analysis, and immunofluorescence assay in CM-treated fibroblasts. Promoting the CRC progression of conditioned media was determined in CRC cells by using MTT assay, fluorescence assay, wound healing assay, transwell migration assay, and tubulogenesis. The results showed that the conditioned media induced the expression of CAF markers associated with the central rearrangement of F-actin in colon fibroblasts, upregulating and promoting the nuclear translocation of YAP. The conditioned media also significantly promoted the proliferation, migration, invasion, and angiogenesis of CRC cells. Interestingly, Verteporfin, a YAP inhibitor during cocultivation, abolished the conversion of CAFs and inhibited proliferation, migration, invasion, and angiogenesis in CRC cells. Moreover, bioinformatics analysis was employed to determine the potential role of YAP as a prognostic marker in CRC patients from databases. The results suggested that YAP has higher expression in CRC patients and is associated with a poor prognosis. In conclusion, these findings demonstrate that YAP-related F-actin rearrangement may be a potential new target of combination therapy with a focus on targeting TME.

Thioredoxin Reductase-1 as a Potential Biomarker in Fibroblast-Associated HCT116 Cancer Cell Progression and Dissemination in a Zebrafish Model.

The tumor microenvironment, especially that of fibroblasts, strongly promotes colorectal cancer (CRC) progression. Progressive cancers usually accumulate high reactive oxygen species (ROS), leading to oxidative stress. The stress relates to the expression of thioredoxin reductase-1 (TrxR-1), which is an oxidative stress sensitivity molecule. This study aimed to investigate TrxR-1 expression as an indication of colon-fibroblast-inducing colorectal cancer progression and metastasis. We found that the high proliferative fibroblast-cultured media (FCM) contained pro-inflammatory cytokines that have a high ability to influence HCT116 and CRC cell progression, when compared with complete media (CM) as a control in terms of growth (CM = 100.00%, FCM = 165.96%), migration (CM = 32.22%, FCM = 83.07%), invasion (CM = 130 cells/field, FCM = 449 cells/field), and EMT transformation while decreasing E-cadherin expression (CM = 1.00, FCM = 0.69) and shape factor (CM = 0.94, FCM = 0.61). In addition, the overexpression of TrxR-1 is associated with cellular oxidant enchantment in FCM-treated cells. A dot plot analysis showed a strong relation between the EMT process and the overexpression of TrxR-1 in FCM-treated cells (CM = 13/100 cells, FCM = 45/100 cells). The cancer transplantation of the adult zebrafish model illustrated a significantly higher number of microtumors in FCM-treated cells (CM = 4.33 ± 1.51/HPF, FCM = 25.00 ± 13.18/HPF) disseminated in the intraperitoneal cavity with TrxR-1 positive cells. The overexpression of TrxR-1 indicated fibroblast-associated CRC progression in HCT116 cells and the zebrafish model. Therefore, TrxR-1 could be applied as a novel biomarker for colorectal cancer progression and prognostic evaluation.