The elucidation of the molecular mechanisms that govern the differentiation and proliferation of human adipose tissue-derived mesenchymal stem cells (hASCs) could improve hASC-based cell therapy. In this study, we examined the roles of microRNA (miRNA)-196a on hASC proliferation and osteogenic differentiation. Lentiviral overexpression of miR-196a decreased hASC proliferation and enhanced osteogenic differentiation, without affecting adipogenic differentiation. Overexpression of miR-196a decreased the protein and mRNA levels of HOXC8, a predicted target of miR-196a. HOXC8 expression was decreased during osteogenic differentiation of hASCs, and this decrease in HOXC8 expression was concomitant with an increase in the level of miR-196a. In contrast, inhibition of miR-196a with 29-O-methyl-antisense RNA increased the protein levels of HOXC8 in treated hASCs and was accompanied by increased proliferation and decreased osteogenic differentiation. The activity of a luciferase construct containing the miR-196a target site from the HOXC8 39UTR was lower in LV-miR196a-infected hASCs than in LV-miLacZ-infected cells. RNA interference-mediated downregulation of HOXC8 in hASCs increased their proliferation and decreased their differentiation into osteogenic cells, without affecting their adipogenic differentiation. Our data indicate that miR-196a plays a role in hASC osteogenic differentiation and proliferation, which may be mediated through its predicted target, HOXC8. This study provides us with a better knowledge of the molecular mechanisms that govern hASC differentiation and proliferation.

Accumulation of ␤-catenin and subsequent stimulation of ␤-catenin-T cell-factor (Tcf)/lymphoid-enhancerfactor (Lef) transcriptional activity causes dedifferentiation of articular chondrocytes, which is characterized by decreased type II collagen expression and initiation of type I collagen expression. This study examined the mechanisms of ␣-catenin degradation, the role of ␣-catenin in ␤-catenin signaling, and the physiological significance of ␣-catenin regulation of ␤-catenin signaling in articular chondrocytes. We found that both ␣-and ␤-catenin accumulated during dedifferentiation of chondrocytes by escaping from proteasomal degradation. ␤-Catenin degradation was ubiquitination-dependent, whereas ␣-catenin was proteasomally degraded in a ubiquitination-independent fashion. The accumulated ␣-and ␤-catenin existed as complexes in the cytosol and nucleus. The complex formation between ␣-and ␤-catenin blocked proteasomal degradation of ␣-catenin and also inhibited ␤-catenin-Tcf/Lef transcriptional activity and the suppression of type II collagen expression associated with ectopic expression of ␤-catenin, the inhibition of proteasome, or Wnt signaling. Collectively, our results indicate that ubiquitin-independent degradation of ␣-catenin regulates ␤-catenin signaling and maintenance of the differentiated phenotype of articular chondrocytes.␤-Catenin interacts with cadherin to participate in cell-cell adhesion and regulates gene expression by acting as a transcriptional co-activator (1, 2). In the absence of extracellular stimuli, cytosolic ␤-catenin is phosphorylated by glycogen synthase kinase-3␤, leading to its ubiquitination and subsequent degradation by the 26 S proteasome. However, extracellular stimuli, such as Wnt signaling, lead to the inhibition of glycogen synthase kinase-3␤, escape of ␤-catenin from ubiquitin-dependent proteolytic degradation, and subsequent cytosolic accumulation of ␤-catenin (3). The accumulated ␤-catenin translocates into the nucleus in association with members of the T cell-factor (Tcf) 1 /lymphoid-enhancer-factor (Lef) family of transcription factors, leading to stimulation or suppression of target gene transcription (2). As a transcriptional co-activator, ␤-catenin is involved in the regulation of several biological functions. Our group previously has shown that ␤-catenin regulates maintenance of differentiated phenotypes (4, 5) and expression of cyclooxygenase-2 (6) in chondrocytes by acting as a transcriptional co-activator. We have also shown that Wnt-7a causes dedifferentiation of chondrocytes characterized by suppression of type II collagen expression and the onset of type I collagen expression. Accumulation and stimulation of ␤-catenin transcriptional activity by Wnt-7a signaling is sufficient to cause chondrocyte dedifferentiation (4, 5).␤-Catenin signaling can be regulated by a variety of proteins. Cadherins regulate the transcriptional activity of ␤-catenin in a cell adhesion-independent manner (7, 8) via sequestration of ␤-catenin in the cytoplasm (9), whereas ␥-catenin ...

b-Catenin/Tcf and NF-jB pathways play an important role in biological functions. We determined the underlying mechanisms of differential interaction between two pathways in various human cancer cell lines. NF-jB positively regulated b-catenin/Tcf pathways in human glioblastoma, whereas it has an opposite effect on b-catenin/Tcf pathways in colon, liver, and breast cancer cells. Expression of lucine zipper tumor suppressor 2 (lzts2) was positively regulated by NF-jB activity in colon, liver, and breast cancer cells, whereas negatively regulated in glioma cells. Downregulation of lzts2 increased the b-catenin/ Tcf promoter activity and inhibited NF-jB-induced modulation of the nuclear translocation of b-catenin. These data indicate that the differential crosstalk between b-catenin/Tcf and NF-jB pathway in various cancer cells is resulted from the differences in the regulation of NF-jB-induced lzts2 expression.

Injection of the Helicobacter pylori cytotoxin-associated gene A (CagA) is closely associated with the development of chronic gastritis and gastric cancer. Individuals infected with H. pylori possessing the CagA protein produce more reactive oxygen species (ROS) and show an increased risk of developing gastric cancer. Sirtuins (SIRTs) are nicotinamide adenine dinucleotide (NAD+)-dependent deacetylases and mitochondrial SIRT3 is known to be a tumor suppressor via its ability to suppress ROS and hypoxia inducible factor 1α (HIF-1α). However, it is unclear whether increased ROS production by H. pylori is regulated by SIRT3 followed by HIF-1α regulation and whether intracellular CagA acts as a regulator thereof. In this study, we investigated correlations among SIRT3, ROS, and HIF-1α in H. pylori-infected gastric epithelial cells. We observed that SIRT3-deficient AGS cells induce HIF-1α protein stabilization and augmented transcriptional activity under hypoxic conditions. In CagA+ H. pylori infected cells, CagA protein localized to mitochondria where it subsequently suppressed SIRT3 proteins. CagA+ H. pylori infection also increased HIF-1α activity through the ROS production induced by the downregulated SIRT3 activity, which is similar to the hypoxic condition in gastric epithelial cells. In contrast, overexpression of SIRT3 inhibited the HIF-1α protein stabilization and attenuated the increase in HIF-1α transcriptional activity under hypoxic conditions. Moreover, CagA+ H. pylori attenuated HIF-1α stability and decreased transcriptional activity in SIRT3-overexpressing gastric epithelial cells. Taken together, these findings provide valuable insights into the potential role of SIRT3 in CagA+ H. pylori-mediated gastric carcinogenesis and a possible target for cancer prevention via inhibition of HIF-1α.