The roles of Oct4 and Nanog in maintaining self-renewal and undifferentiated status of adult stem cells are unclear. Here, increase in Oct4 and Nanog expression along with increased proliferation and differentiation potential but decreased spontaneous differentiation were observed in early-passage (E), hypoxic culture (H), and p21 knockdown (p21KD) mesenchymal stem cells (MSCs) compared to late-passage (L), normoxic culture (N), and scrambled shRNA-overexpressed (Scr) MSCs. Knockdown of Oct4 and Nanog in E, H, and p21KD MSCs decreased proliferation and differentiation potential and enhanced spontaneous differentiation, whereas overexpression of Oct4 and Nanog in L, N, and Scr MSCs increased proliferation and differentiation potential and suppressed spontaneous differentiation. Oct4 and Nanog upregulate Dnmt1 through direct binding to its promoter, thereby leading to the repressed expression of p16 and p21 and genes associated with development and lineage differentiation. These data demonstrate the important roles of Oct4 and Nanog in maintaining MSC properties.

Gene inactivation through DNA hypermethylation plays a pivotal role in carcinogenesis. This study aimed to profile aberrant DNA methylation in different stages of liver disease, namely noncirrhosis, cirrhosis and hepatocellular carcinoma (HCC), and also to clarify the influence of hepatitis B virus (HBV) infection on the aberrant DNA methylation in HCCs. Promoter methylation in p14 ARF , p16 INK4a , O 6 -methylguanine-DNA methyltransferase (MGMT), glutathione S-transferase pi (GSTP1) and E-cadherin (E-Cad) genes of 58 HCCs paired with adjacent nontumorous tissues was assayed by methylation-specific PCR. HBV infection was determined using a hepatitis B virus surface antigen (HBsAg) serological assay. The frequency of p16 INK4a promoter methylation increased from noncirrhotic, cirrhotic, to HCC tissues (noncirrhotic vs. HCC, p < 0.001), while that of GSTP1 promoter methylation increased in cirrhotic tissues compared to noncirrhotic ones (p 5 0.029). The frequency of GSTP1 promoter hypermethylation is significantly higher in HCC than in nontumorous tissues (p 5 0.022) from HBsAg-positive patients, but not the HBsAg-negative controls (p 5 0.289). While the frequency of E-Cad promoter hypermethylation remained high in both nontumorous tissues and HCCs from HBsAg-positive patients (p 5 0.438), it was lower in HCCs than in nontumorous tissues from HBsAg-negative patients (p 5 0.002). In contrast, the frequency of p16 INK4a , MGMT and p14 ARF promoter hypermethylation in HCCs was unrelated to HBsAg status. In conclusion, aberrant DNA methylation may begin at different stages of liver disease in a gene-dependent manner. Moreover, HBV infection may enhance or maintain GSTP1 and E-Cad promoter methylation and thereby affect hepatocarcinogenesis. ' 2007 Wiley-Liss, Inc.

No abstract

To investigate the immunomodulatory activities of phytocompounds for potential therapeutics, we devised an in vivo, transgenic, human cytokine gene promoter assay using defined epidermal skin cells as test tissue. Test compounds were topically applied to mouse skin before or after gene gun transfection, using a cytokine gene promoter-driven luciferase reporter. Croton oil, an inflammation inducer, induced transgenic GM-CSF and TNF-alpha promoter activities in skin epidermis 6-fold and 3.4-fold, respectively; however, it produced a less than 1.5-fold and 1.7-fold change in IL-1beta and IL-18 promoter activity, respectively. The phytocompound shikonin drastically inhibited inducible GM-CSF promoter activity. However, a fraction of Dioscorea batatas extract significantly increased the GM-CSF promoter activity in normal and inflamed skin. Shikonin suppressed the transcriptional activity of GM-CSF promoter by inhibiting the binding of TFIID protein complex (TBP) to TATA box. Our results demonstrate that this in vivo transgenic promoter activity assay system is cytokine gene-specific, and highly responsive to pro-inflammatory or anti-inflammatory stimuli. Currently it is difficult to profile the expression and cross-talk of various types of cytokines in vivo. This investigation has established a bona fide in vivo, in situ, immune tissue system for research into cytokine response to inflammation.

Inorganic arsenic is an environmental carcinogen. The generation of toxic trivalent methylated metabolites complicates the study of arsenic-mediated carcinogenesis. This study systematically evaluated the effect of chronic treatment with sodium arsenite (iAsIII), monomethylarsonous acid (MMAIII), and dimethylarsinous acid (DMAIII) on immortalized human uroepithelial cells (SV-HUC-1 cells) using cDNA microarray. After exposure for 25 passages to iAsIII (0.5 μM), MMAIII (0.05, 0.1, or 0.2 μM), or DMAIII (0.2 or 0.5 μM), significant compound-specific morphologic changes were observed. A set of 114 genes (5.7% of the examined genes) was differentially expressed in one or more sets of arsenical-treated cells compared with untreated controls. Expression analysis showed that exposure of cells to DMAIII resulted in a gene profile different from that in cells exposed to iAsIII or MMAIII, and that the iAsIII-induced gene profile was closest to that in the tumorigenic HUC-1–derived 3-methylcholanthrene–induced tumorigenic cell line MC-SV-HUC T2, which was derived from SV-HUC-1 cells by methylcholanthrene treatment. Of the genes affected by all three arsenicals, only one, that coding for interleukin-1 receptor, type II, showed enhanced expression, a finding confirmed by the reduced increase in NF-κB (nuclear factor kappa B) activity seen in response to interleukin-1β in iAsIII-exposed cells. The expression of 11 genes was suppressed by all three arsenicals. 5-Aza-deoxycytidine partially restored the transcription of several suppressed genes, showing that epigenetic DNA methylation was probably involved in arsenical-induced gene repression. Our data demonstrate that chronic exposure to iAsIII, MMAIII, or DMAIII has different epigenetic effects on urothelial cells and represses NF-κB activity.