Characteristics of Human Nasal Turbinate Stem Cells under Hypoxic Conditions.

This study investigated the influence of hypoxic culture conditions on human nasal inferior turbinate-derived stem cells (hNTSCs), a subtype of mesenchymal stem cells (MSCs). It aimed to discern how hypoxia affected hNTSC characteristics, proliferation, and differentiation potential compared to hNTSCs cultured under normal oxygen levels. After obtaining hNTSCs from five patients, the samples were divided into hypoxic and normoxic groups. The investigation utilized fluorescence-activated cell sorting (FACS) for surface marker analysis, cell counting kit-8 assays for proliferation assessment, and multiplex immunoassays for cytokine secretion study. Differentiation potential-osteogenic, chondrogenic, and adipogenic-was evaluated via histological examination and gene expression analysis. Results indicated that hNTSCs under hypoxic conditions preserved their characteristic MSC phenotype, as confirmed by FACS analysis demonstrating the absence of hematopoietic markers and presence of MSC markers. Proliferation of hNTSCs remained unaffected by hypoxia. Cytokine expression showed similarity between hypoxic and normoxic groups throughout cultivation. Nevertheless, hypoxic conditions reduced the osteogenic and promoted adipogenic differentiation potential, while chondrogenic differentiation was relatively unchanged. These insights contribute to understanding hNTSC behavior in hypoxic environments, advancing the development of protocols for stem cell therapies and tissue engineering.

CD40 Agonists Alter the Pancreatic Cancer Microenvironment by Shifting the Macrophage Phenotype toward M1 and Suppress Human Pancreatic Cancer in Organotypic Slice Cultures.

Background/Aims: CD40 agonists are thought to generate antitumor effects on pancreatic cancer via macrophages and T cells. We aimed to investigate the role of CD40 agonists in the differentiation of macrophages and treatment of human pancreatic adenocarcinoma. Methods: Immunohistochemistry was performed on paraffin-embedded surgical blocks from patients with pancreatic cancers to evaluate macrophage phenotypes and their relationship with survival. The effects of CD40 agonists on macrophage phenotypes and human pancreatic cancer were evaluated utilizing cell cocultures and organotypic slice cultures. Results: CD163+ (predominant in M2 macrophages) and FOXP3+ (predominant in regulatory T cells) expression levels in the tumors were significantly lower in patients with stage IB pancreatic cancer than in those with stage II or III disease (p=0.002 and p=0.003, respectively). Patients with high CD163+ expression had shorter overall survival than those with low CD163+ expression (p=0.002). In vitro treatment of THP-1 macrophages with a CD40 agonist led to an increase in HLA-DR+ (predominant in M1 macrophages) and a decrease in CD163+ expression in THP-1 cells. Cell cocultures showed that CD40 agonists facilitate the suppression of PANC-1 human pancreatic cancer cells by THP-1 macrophages. Organotypic slice cultures showed that CD40 agonists alter the pancreatic cancer microenvironment by shifting the macrophage phenotype toward M1 (increase HLA-DR+ and decrease CD163+ expression), decreasing the abundance of regulatory T cells, and increasing tumor cell apoptosis. Conclusions: CD163 is related to advanced human pancreatic cancer stages and shorter overall survival. CD40 agonists alter macrophage phenotype polarization to favor the M1 phenotype and suppress human pancreatic cancer.

Organotypic slice cultures of pancreatic ductal adenocarcinoma preserve the tumor microenvironment and provide a platform for drug response.

BACKGROUND: /Objective: The conventional models currently used to evaluate various anti-tumor therapeutic agents are not sufficient for representing human pancreatic ductal adenocarcinoma (PDA), which has a unique tumor microenvironment. We aimed to produce an organotypic slice culture model from human PDA that resembles the in vivo situation and to evaluate the responses of PDA slices to established cytotoxic drugs. METHODS: PDA tissues were obtained from 10 patients who underwent pancreatic resection. The tissues were sliced by a vibratome, and the tumor slices were then cultured. The viability of tumor slices during slice culture was evaluated using H&E and immunohistochemical staining, and stromal cells were demonstrated. The effects of cytotoxic drugs on PDA cell lines and slices were analyzed. RESULTS: Tumor slices maintained their surface areas and tissue viability for at least five days during culture. Preserved proliferation and apoptosis in tumor slices were observed by the expression of Ki-67 and cleaved caspase-3. Stromal cells including macrophages (CD68+ and CD163+), T cells (CD3+, CD8+, and FOXP3+), and myeloid cells (CD11b+) were present throughout the culture period. Staurosporine, gemcitabine, and cisplatin treatment of PDA cell lines and tumor slices exerted proportional cytotoxic effects in terms of MTT viability, tumor cell number, and Ki-67 and cleaved caspase-3 expression. CONCLUSIONS: Organotypic human PDA slice cultures preserved their viability and tumor microenvironment for at least five days during slice culture. PDA slice culture appears to be a feasible preclinical test model to assess the response to anti-tumor agents.