Bioactive Vitamin D Attenuates MED28-Mediated Cell Growth and Epithelial-Mesenchymal Transition in Human Colorectal Cancer Cells.

Inadequate vitamin D status may increase the risk of developing multiple types of cancer. Epidemiological studies suggest an inverse association between 25-hydroxyvitamin D3 (25(OH)D3) and malignancy, including colorectal cancer. Previous studies have suggested that MED28, a Mediator subunit involved in transcriptional regulation, is associated with the growth of colorectal cancer cells; however, its role in the progression of metastasis such as epithelial-mesenchymal transition (EMT) and cell migration of colorectal cancer is unclear at present. The aim of this study was to investigate a potentially suppressive effect of calcitriol, 1,25-dihydroxyvitamin D3 (1,25(OH)2D3), a bioactive form of vitamin D, and the role of MED28 in the progression of EMT in human colorectal cancer cells. Suppression of MED28 increased the expression of E-cadherin and reduced the expression of several mesenchymal and migration biomarkers and Wnt/ß-catenin signaling molecules, whereas overexpression of MED28 enhanced the EMT features. Calcitriol suppressed the expression of MED28, and the effect of calcitriol mirrored that of MED28 silencing. Our data indicate that calcitriol attenuated MED28-mediated cell growth and EMT in human colorectal cancer cells, underlining the significance of MED28 in the progression of colorectal cancer and supporting the potential translational application of calcitriol.

Calcitriol Suppresses Warburg Effect and Cell Growth in Human Colorectal Cancer Cells.

Increasing lines of evidence indicate that the biologically active form of vitamin D, calcitriol (1,25-dihydroxyvitamin D3), prevents cancer progression by reducing cell proliferation, increasing cell differentiation, and inhibiting angiogenesis, among other potential roles. Cancer cells in solid tumors preferably undergo the "Warburg effect" to support cell growth by upregulating glycolysis, and the glycolytic intermediates further serve as building blocks to generate biomass. The objective of the current study is to investigate whether calcitriol affects glucose metabolism and cell growth in human colorectal cancer cells. Calcitriol reduced the expression of cyclin D1 and c-Myc. In addition, calcitriol reduced the expression of glucose transporter 1 (GLUT1) and key glycolytic enzymes and decreased extracellular acidification rate but increased oxygen consumption rate in human colorectal cancer cells. In a subcutaneous HT29 xenograft NOD/SCID mouse model, the volume and weight of the tumors were smaller in the calcitriol groups as compared with the control group, and the expression levels of GLUT1 and glycolytic enzymes, hexokinase 2 and lactate dehydrogenase A, were also lower in the calcitriol groups in a dose-responsive manner. Our data indicate that calcitriol suppresses glycolysis and cell growth in human colorectal cancer cells, suggesting an inhibitory role of the biologically active form of vitamin D in colorectal cancer progression.

The Enhancing Effect of Fungal Immunomodulatory Protein-Volvariella Volvacea (FIP-vvo) on Maturation and Function of Mouse Dendritic Cells.

Volvariella volvacea, also known as straw mushroom, is a common edible mushroom in Chinese cuisine. It contains many nutrients for human health. A fungal immunomodulatory protein (FIP) has been isolated from V. volvacea and named FIP-vvo. Although the regulatory effects of many FIPs on immunity have been identified, the impact of FIP-vvo in modulating dendritic cells (DCs), which play a key role to connect the innate and the adaptive immunity, is not known. In this study, we aim to study the effect of FIP-vvo on the DC maturation and function. We found that FIP-vvo slightly increased the generation of CD11c+ bone marrow-derived DC (BMDC). In addition, the surface expression of MHCII was promoted in BMDCs after the treatment of FIP-vvo, suggesting that FIP-vvo induces DC maturation. Furthermore, FIP-vvo enhanced the ability of BMDCs to activate antigen-specific T cell responses in vitro. In the in vivo study, the FIP-vvo treatment facilitated T cell response in lymph nodes. Therefore, for the first time, our data demonstrated that FIP-vvo promoted DC maturation and function and suggested that FIP-vvo could have benefits for human health by enhancing immunity.

MED28 and forkhead box M1 (FOXM1) mediate matrix metalloproteinase 2 (MMP2)-dependent cellular migration in human nonsmall cell lung cancer (NSCLC) cells.

Non-small-cell lung cancer (NSCLC) accounts for the majority of the lung cancer cases that have become a leading cause of cancer deaths worldwide. Overexpression of transcription factor forkhead box M1 (FOXM1) is involved in the inauspicious development of several types of cancer, including lung tumor aggressiveness. Our laboratory has previously found that MED28, a Mediator subunit for transcriptional activation, modulates cell growth, epithelial-mesenchymal transition, migration, and invasion in human breast cancer cells. The objective of the current study is to investigate the potential role of MED28 and FOXM1 in NSCLC. In addition to A549 and PC9 cells, we also used a doxycycline-inducible system to generate FOXM1-overexpressed A549-DN cells, and we explored the connection of MED28 with FOXM1 and their effect on migration. Herein, we report that the increased expression levels of both MED28 and FOXM1 elevated the expression of matrix metalloproteinase 2 (MMP2), a metastasis marker, which enhanced cell migration and matrigel invasion of NSCLC cells. Furthermore, MED28 interacted with FOXM1, and both exhibited a mutual effect on the expression and subcellular localization. Moreover, MED28 small interfering RNA-mediated MMP2 gene suppression could be attenuated by inducible expression of a constitutively active form of FOXM1, which consequently restored the migration and invasion ability of NSCLC cells. Our data indicate that MED28 interacts with FOXM1, and each affects the expression and localization of the other, and, more importantly, both regulate MMP2-dependent migration and invasion in human lung cancer cells.

MED28 Regulates Epithelial-Mesenchymal Transition Through NFκB in Human Breast Cancer Cells.

MED28, a mammalian Mediator subunit, was found highly expressed in several types of malignancy, including breast cancer. Recently, we have identified a role of MED28 in regulating both cell growth and migration in human breast cancer cells. In epithelium-derived solid tumor, migration and invasion are preceded by the progression of epithelial-mesenchymal transition (EMT) which calls for downregulation of epithelial markers as well as upregulation of mesenchymal markers, among other features. The objective of this study was to investigate a putative role of MED28 in the progression of EMT in human breast cancer cells. In fibroblast-like MDA-MB-231 cells, suppression of MED28 attenuated the mesenchymal morphology, concomitantly with a reduction of several mesenchymal biomarkers and Snail, a transcriptional repressor of E-cadherin. The suppression effect was also accompanied by downregulation of p-NFκB/p65. However, overexpression of MED28 exhibited in an opposite manner. In epithelial MCF7 cells, administration of Adriamycin®, an experimental EMT induction system, led to a mesenchyme-like appearance correlated with increased expression of MED28, p-p65, and Snail, and a reciprocal change of epithelial and mesenchymal markers. Furthermore, suppression of MED28 attenuated the experimental EMT effect and restored the original expression status of E-cadherin and MMP9 in MCF7 cells. Our data indicate that MED28 modulates the development of EMT through NFκB in human breast cancer cells, further reinforcing the significance of MED28 in the progression of breast cancer on top of its role in cell growth and migration. J. Cell. Physiol. 232: 1337-1345, 2017. © 2016 Wiley Periodicals, Inc.

All Trans-Retinoic Acid Mediates MED28/HMG Box-Containing Protein 1 (HBP1)/ß-Catenin Signaling in Human Colorectal Cancer Cells.

Vitamin A is required for normal body function, including vision, epithelial integrity, growth, and differentiation. All trans-retinoic acid (ATRA), a family member of vitamin A, has been explored in treating acute promyelocytic leukemia and other types of cancer. Dysregulated Wnt/ß-catenin signaling and disrupted cadherin-catenin complex often contribute to colorectal malignancy. MED28, a mammalian Mediator subunit, is found highly expressed in breast and colorectal cancers. Our laboratory has also reported that MED28 regulates cell growth, migration, and invasion in human breast cancer cells. In the current study we investigated the effect of ATRA on MED28 and Wnt/ß-catenin signaling in colorectal cancer. HCT116, HT29, SW480, and SW620, four human colorectal cancer cell lines representing different stages of carcinogenesis and harboring critical genetic changes, were employed. Our data indicated that regardless of genetic variations among these cells, suppression of MED28 reduced the expression of cyclin D1, c-Myc, and nuclear ß-catenin, but increased the expression of E-cadherin and HMG box-containing protein 1 (HBP1) where HBP1 has been described as a negative regulator of the Wnt/ß-catenin signaling. The reporter activity of an HBP1 promoter increased upon MED28 knockdown, but decreased upon MED28 overexpression. ATRA reduced the expression of MED28 and mimicked the effect of MED28 suppression in down-regulating Wnt/ß-catenin signaling. Taken together, ATRA can reverse the suppressive effect of MED28 on HBP1 and E-cadherin and inactivate the Wnt/ß-catenin pathway in colorectal cancer, suggesting a protective effect of ATRA against colorectal cancer. J. Cell. Physiol. 231: 1796-1803, 2016. © 2015 Wiley Periodicals, Inc.

MED28 regulates MEK1-dependent cellular migration in human breast cancer cells.

MED28, a mammalian Mediator subunit, exhibits several cellular roles, including a merlin, Grb2, and cytoskeleton-associated protein (magicin), a repressor of smooth muscle cell differentiation, and an endothelial-derived gene (EG-1). Overexpression of MED28 may stimulate cell proliferation which presumably results from the transcriptional activation of the Mediator function. Additionally, several tumors, including breast cancer, highly express MED28. We have found recently that MED28 potentiated epidermal growth factor (EGF)-induced migration in human breast cancer cells. Therefore, the objective of this study is to identify the role of MED28 in the aspect of cellular migration and invasion in human breast cancer cells. Suppression of MED28 blocked cellular migration and invasion with concomitant reduced expression levels of matrix metalloproteinase-2 (MMP2) and mitogen-activated protein kinase kinase 1 (MAP2K1; MEK1); overexpression of MED28 enhanced cellular migration and upregulated MMP2 and MEK1 expression. Moreover, suppression of MEK1, by dominant-negative, kinase-dead MEK1 cDNA construct or MEK1-specific small interfering RNA (siRNA) as well as MEK1 inhibitors, blocked MED28-induced MMP2 activation, cellular migration, and invasion in breast cancer cells. Furthermore, ectopic expression of MEK1 rescued the inhibitory effect of MED28 knockdown on invasion, and exogenous MMP2 recombinant protein recovered the suppression on invasion upon MED28 or MEK1 knockdown. Our data indicate that MED28 regulates cellular migration in a MEK1-dependent manner in human breast cancer cells, reinforcing the important cellular roles of MED28.