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Tumor necrosis factor alpha (TNF-α)/hypoxia-treated 3T3-L1 adipocytes have been used to model inflamed and insulin-resistant adipose tissue: this study examines gaps in the model. We tested whether modulating TNF-α/hypoxia treatment time could reduce cell death while still inducing inflammation and insulin resistance. Adipocytes were treated with TNF-α (12 h or 24 h) and incubated in a hypoxic chamber for 24 h. To examine maintenance of the phenotype over time, glucose and FBS were added at 24 h post initiation of treatment, and the cells were maintained for an additional 48 h. Untreated adipocytes were used as a control. Viability, insulin resistance, and inflammation were assessed using Live/Dead staining, RT-qPCR, ELISA, and glucose uptake assays. Treatment for 12 h with TNF-α in the presence of hypoxia resulted in an increase in the percentage of live cells compared to 24 h treated cells. Importantly, insulin resistance and inflammation were still induced in the 12 h treated adipocytes: the expression of the insulin sensitive and inflammatory genes was decreased and increased, respectively. In 72 h treated adipocytes, no significant differences were found in the viability, glucose uptake or insulin-sensitive and inflammatory gene expression. This study provides a modified approach to in vitro odeling adipocyte inflammation and insulin resistance.        .

To overcome the limitations of in vitro two-dimensional (2D) cancer models in mimicking the complexities of the native tumor milieu, three-dimensional (3D) engineered cancer models using biomimetic materials have been introduced to more closely recapitulate the key attributes of the tumor microenvironment. Specifically, for colorectal cancer (CRC), a few studies have developed 3D engineered tumor models to investigate cell-cell interactions or efficacy of anti-cancer drugs. However, recapitulation of CRC cell line phenotypic differences within a 3D engineered matrix has not been systematically investigated. Here, we developed an in vitro 3D engineered CRC (3D-eCRC) tissue model using the natural-synthetic hybrid biomaterial PEG-fibrinogen and three CRC cell lines, HCT 116, HT-29, and SW480. To better recapitulate native tumor conditions, our 3D-eCRC model supported higher cell density encapsulation (20 × 10 6  cells/mL) and enabled longer term maintenance (29 days) as compared to previously reported in vitro CRC models. The 3D-eCRCs formed using each cell line demonstrated line-dependent differences in cellular and tissue properties, including cellular growth and morphology, cell subpopulations, cell size, cell granularity, migration patterns, tissue growth, gene expression, and tissue stiffness. Importantly, these differences were found to be most prominent from Day 22 to Day 29, thereby indicating the importance of long-term culture of engineered CRC tissues for recapitulation and investigation of mechanistic differences and drug response. Our 3D-eCRC tissue model showed high potential for supporting future in vitro comparative studies of disease progression, metastatic mechanisms, and anti-cancer drug candidate response in a CRC cell line-dependent manner., (© 2023 Wiley Periodicals LLC.)

Colorectal cancer (CRC) is the third-leading cause of cancer-related deaths in the United States and worldwide. Obesity-a worldwide public health concern-is a known risk factor for cancer including CRC. However, the mechanisms underlying the link between CRC and obesity have yet to be fully elucidated in part because of the molecular heterogeneity of CRC. We hypothesized that obesity modulates CRC in a consensus molecular subtype (CMS)-dependent manner. RNA-seq data and associated tumor and patient characteristics including body weight and height data for 232 patients were obtained from The Cancer Genomic Atlas-Colon Adenocarcinoma (TCGA-COAD) database. Tumor samples were classified into the four CMSs with the CMScaller R package; body mass index (BMI) was calculated and categorized as normal, overweight, and obese. We observed a significant difference in CMS categorization between BMI categories. Differentially expressed genes (DEGs) between obese and overweight samples and normal samples differed across the CMSs, and associated prognostic analyses indicated that the DEGs had differing associations on survival. Using Gene Set Enrichment Analysis, we found differences in Hallmark gene set enrichment between obese and overweight samples and normal samples across the CMSs. We constructed Protein-Protein Interaction networks and observed differences in obesity-regulated hub genes for each CMS. Finally, we analyzed and found differences in predicted drug sensitivity between obese and overweight samples and normal samples across the CMSs. Our findings support that obesity impacts the CRC tumor transcriptome in a CMS-specific manner. The possible associations reported here are preliminary and will require validation using in vitro and animal models to examine the CMS-dependence of the genes and pathways. Once validated the obesity-linked genes and pathways may represent new therapeutic targets to treat colon cancer in a CMS-dependent manner., Competing Interests: The authors have declared that no competing interests exist.

Chitosan-oligosaccharides (COS) are low-molecular weight chitosan derivatives with interesting clinical applications. The optimization of both COS production and purification is an important step in the design of an efficient production system and for the exploration of new COS applications. Trichoderma harzianum is an innocuous biocontrol agent that represents a novel biotechnological tool due to the production of extracellular enzymes, including those that produce a COS mixture. In this work, we propose different systems for the production of COS using the T. harzianum chitinolitic system. A complete qualitative and quantitative analysis of a partially purified COS mixture were performed. Also, an evaluation of the anticancer and antimicrobial effects of the COS mixture was carried out. Three chitosan variants (colloidal, solid and solution) and two fungus stages (spores and mycelia) were tested for COS production. The best system consisted of the interaction of the solid chitosan and the fungal spores, producing a COS mixture containing species from the monomer to the hexamer, in a concentration range of 7-238 mg/mL, according to chromatographic analysis. The proposed purification method isolated the monomer and the dimer from the COS mixture. Moreover, the COS mixture has an inhibitory effect on the growth of bacteria and changes the morphology of yeasts. As anticancer compounds, COS inhibited the growth of cervical cancer cells at concentration of 4 mg/mL and significantly reduced the survival rate of the cells. In conclusion, T. harzianum proved to be an efficient system for COS mixture production., (Copyright © 2019 Elsevier Ltd. All rights reserved.)