The early onset breast cancer patients (age ≤ 40) often display higher incidence of axillary lymph node metastasis, and poorer five-year survival than the late-onset patients. To identify the genes and molecules associated with poor prognosis of early onset breast cancer, we examined gene expression profiles from paired breast normal/tumor tissues, and coupled with Gene Ontology and public data base analysis. Our data showed that the expression of GAS7b gene was lower in the early onset breast cancer patients as compared to the elder patients. We found that GAS7 was associated with CYFIP1 and WAVE2 complex to suppress breast cancer metastasis via blocking CYFIP1 and Rac1 protein interaction, actin polymerization, and β1-integrin/FAK/Src signaling. We further demonstrated that p53 directly regulated GAS7 gene expression, which was inversely correlated with p53 mutations in breast cancer specimens. Our study uncover a novel regulatory mechanism of p53 in early onset breast cancer progression through GAS7–CYFIP1-mediated signaling pathways.

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New potential sources of stem cells for clinical application include bone marrow mesenchymal stem cells (BMMSCs), human embryonic stem cells (hESCs), and induced pluripotent stem cells (iPS). However, each source is not without its own concerns. While research continues in an effort to overcome these problems, the generation of mesenchymal progenitors from existing hESC lines may circumvent many of these issues. We report here a simple and efficient method of generating hESC-derived mesenchymal progenitors (EMPs) and transcriptome profiling using a concise, custom-designed, oligomnucleotide gene expression microarray. Characterization of EMPs shows that these cells are similar to BMMSCs in terms of differentiation capacity as well as cell surface marker expression. In addition, EMPs express several ESC markers and HLA-G, a nonclassical MHC class I molecule with immunomodulatory properties. Morevoer, EMPs possess significantly enhanced proliferative ability over BMMSCs during which karyotypic stability was maintained. Although derived from hESCs, EMPs do not form any tumors in immunocompromised mice. To efficiently profile gene expression in multiple samples, we designed an oligoarray to probe just over 11,000 genes highly expressed in stem cells. We found that the transcriptome of EMPs is more similar to BMMSCs than hESCs. Both cell types highly express genes involved in processes related to the cytoskeleton, extracellular matrix, and cell adhesion, but EMPs show higher expression of genes involved in cell proliferation whereas BMMSCs showed higher expression of immune-related genes. Based on our data, EMPs may be an accessible source of mesenchymal progenitor for therapeutic use.

Areca nut is the most widely used psychoactive substance and an important environmental risk factor for development of oral premalignant lesions and cancer. Arecoline, the major alkaloid of areca nut, has been known to cause cytotoxicity and genotoxicity in mammalian cells in vivo and in vitro and even contributes to carcinogenicity. However, the susceptible genes accounting for arecoline-induced damage in normal human oral cells are still lacking, which possibly involves in initial molecular damage via alternation of gene expression level on biological pathways. The present study was undertaken to characterize the toxic effects of arecoline in gene expression profiling on normal human gingival fibroblasts (HGF) using cDNA microarray and quantitative real-time reverse transcription PCR. The cytotoxicity of arecoline on HGF-1 cell line was elevated in a dose-dependent manner (p < 0.05) accompanied with distinct morphological change and formation of intracellular vacuoles were observed. At optimum concentration of arecoline determined from dose-response curve of the cytotoxicity, a large number of genes were significantly repressed than induced by arecoline in global gene expression profiling. Five induced- and seven repressed genes including glutathione synthetase were further validated, and their gene expression changes were increased in a dose-dependent manner in a concentration range of 50-150 microg/ml. In conclusion, we proposed a tentative model to explain arecoline-induced effects on contribution of oral pathogenesis. The findings identified that 12 susceptible genes can potentially serve as biomarkers of arecoline-induced damage in betel chewers.

Multilineage tissue-source mesenchymal stem cells (MSCs) possess strong immunomodulatory properties and are excellent therapeutic agents, but require constant isolation from donors to combat replicative senescence. The differentiation of human induced pluripotent stem cells (iPSCs) into MSCs offers a renewable source of MSCs; however, reports on their immunomodulatory capacity have been discrepant. Using MSCs differentiated from iPSCs reprogrammed using diverse cell types and protocols, and in comparison to human embryonic stem cell (ESC)-MSCs and bone marrow (BM)-MSCs, we performed transcriptome analyses and assessed for functional immunomodulatory properties. Differentiation of MSCs from iPSCs results in decreased c-Myc expression and its downstream pathway along with a concomitant downregulation in the DNA replication pathway. All four lines of iPSC-MSCs can significantly suppress in vitro activated human peripheral blood mononuclear cell (PBMC) proliferation to a similar degree as ESC-MSCs and BM-MSCs, and modulate CD4 T lymphocyte fate from a type 1 helper T cell (Th1) and IL-17A-expressing (Th17) cell fate to a regulatory T cell (Treg) phenotype. Moreover, iPSC-MSCs significantly suppress cytotoxic CD8 T proliferation, activation, and differentiation into type 1 cytotoxic T (Tc1) and IL-17-expressing CD8 T (Tc17) cells. Coculture of activated PBMCs with human iPSC-MSCs results in an overall shift of secreted cytokine profile from a pro-inflammatory environment to a more immunotolerant milieu. iPSC-MSC immunomodulation was also validated in vivo in a mouse model of induced inflammation. These findings support that iPSC-MSCs possess low oncogenicity and strong immunomodulatory properties regardless of cell-of-origin or reprogramming method and are good potential candidates for therapeutic use. Stem Cells 2018;36:903-914.

We detected a significant elevation of serum HSP90α levels in pancreatitis patients and even more in pancreatic ductal adenocarcinoma (PDAC) patients. However, there was no significant difference in the serum HSP90α levels between patients with early-stage and late-stage PDAC. To study whether elevation of serum HSP90α levels occurred early during PDAC development, we used LSL-KrasG12D/Pdx1-Cre transgenic mice as a studying model. Elevated serum HSP90α levels were detected before PDAC formation and an extracellular HSP90α (eHSP90α) inhibitor effectively prevented PDAC development. Both serum HSP90α level and pancreatic lesion were suppressed when the mice were administered a CD11b-antagonizing antibody, suggesting that CD11b+-myeloid cells were associated with eHSP90α levels and pancreatic carcinogenesis. Consistently, in CD11b-DTR-EGFP transgenic mouse model with CD11b+-myeloid cells depletion, serum HSP90α levels were suppressed and Panc-02 cell grafts failed to develop tumors. Macrophages and granulocytes are two common tissue-infiltrating CD11b+-myeloid cells. Duplex in situ hybridization assays suggested that macrophages were predominant HSP90α-expressing CD11b+-myeloid cells during PDAC development. Immunohistochemical and immunohistofluorescent staining results revealed that HSP90α-expressing cells included not only macrophages but also pancreatic ductal epithelial (PDE) cells. Cell culture studies also indicated that eHSP90α could be produced by macrophages and macrophage-stimulated PDE cells. Macrophages not only secreted significant amount of HSP90α, but also secreted interleukin-6 and interleukin-8 to induce a JAK2−STAT3 signaling axis in PDE cells, stimulating them to express and secrete HSP90α. eHSP90α further promoted cellular epithelial-mesenchymal transition, migration, and invasion in PDE cells. Besides myeloid cells, eHSP90α can be potentially taken as a target to suppress PDAC pathogenesis.

Oral squamous cell carcinoma (OSCC) carcinogenesis involves heterogeneous tumor cells, and the tumor microenvironment (TME) is highly complex with many different cell types. Cancer cell–TME interactions are crucial in OSCC progression. Candida albicans (C. albicans)—frequently pre-sent in the oral potentially malignant disorder (OPMD) lesions and OSCC tissues—promotes malignant transformation. The aim of the study is to verify the mechanisms underlying OSCC car-cinogenesis with C. albicans infection and identify the biomarker for the early detection of OSCC and as the treatment target. The single-cell RNA sequencing analysis (scRNA-seq) was performed to explore the cell subtypes in normal oral mucosa, OPMD, and OSCC tissues. The cell composi-tion changes and oncogenic mechanisms underlying OSCC carcinogenesis with C. albicans infec-tion were investigated. Gene Set Variation Analysis (GSVA) was used to survey the mechanisms underlying OSCC carcinogenesis with and without C. albicans infection. The results revealed spe-cific cell clusters contributing to OSCC carcinogenesis with and without C. albicans infection. The major mechanisms involved in OSCC carcinogenesis without C. albicans infection are the IL2/STAT5, TNFα/NFκB, and TGFβ signaling pathways, whereas those involved in OSCC carcinogenesis with C. albicans infection are the KRAS signaling pathway and E2F target down-stream genes. Finally, stratifin (SFN) was validated to be a specific biomarker of OSCC with C. albicans infection. Thus, the detailed mechanism underlying OSCC carcinogenesis with C. albicans infection was determined and identified the treatment biomarker with potential precision medicine applications.

Lymph-node metastasis is a prognosis factor for poor clinical outcome of breast cancer patients. Currently, how breast cancer cells establish pre-metastatic niche in the tumor-draining lymph nodes (TDLNs) is still unclear. To address this question, we isolated heterogeneous cells including immune and stromal cells from naive lymph nodes (LNs) of the FVB/NJ mice and TDLNs of the MMTV-PyMT mice. Single-cell RNA sequencing was performed to investigate the transcriptome of the cells and various bioinformatics analyses were used to identify the altered pathways. Our results revealed several significant changes between naïve LNs and TDLNs. First, according to immunologic signature and pathway analysis, CD4+ and CD8 + T cells showed upregulated angiogenesis pathway genes and higher regulatory T (Treg)-associated genes while they demonstrated downregulation of interferon response and inflammatory response gene signatures, concurrently suggesting an immunosuppressive microenvironment in the TDLNs. Second, profiling of B cells showed down-regulation of marginal zone B lymphocytes in the TDLNs, which was validated by flow cytometric analysis. Third, we found the enhancement of oxidative phosphorylation pathway in the fibroblastic reticular cells (FRCs) of the MMTV-PyMT mice and the elevation of related genes including Prdx3, Ndufa4 and Uqcrb, suggesting massive ATP consumption and TCA cycle metabolism in the FRCs. Collectively, our results reveal the reprogramming of TDLNs during breast cancer progression at single-cell level in a spontaneous breast cancer model and suggest the changes in immune modulation and metabolic switch are key alterations in the preparation of pre-metastatic niche by breast cancer cells.