The extracellular domain of epithelial cell adhesion molecule (EpCAM) enhances multipotency of mesenchymal stem cells through EGFR-LIN28-LET7 signaling.

Mesenchymal stem cells (MSCs) are widely considered to be an attractive cell source for regenerative therapies, but maintaining multipotency and self-renewal in cultured MSCs is especially challenging. Hence, the development and mechanistic description of strategies that help promote multipotency in MSCs will be vital to future clinical use. Here, using an array of techniques and approaches, including cell biology, RT-quantitative PCR, immunoblotting, immunofluorescence, flow cytometry, and ChIP assays, we show that the extracellular domain of epithelial cell adhesion molecule (EpCAM) (EpEX) significantly increases the levels of pluripotency factors through a signaling cascade that includes epidermal growth factor receptor (EGFR), signal transducer and activator of transcription 3 (STAT3), and Lin-28 homolog A (LIN28) and enhances the proliferation of human bone marrow MSCs. Moreover, we found that EpEX-induced LIN28 expression reduces the expression of the microRNA LET7 and up-regulates that of the transcription factor high-mobility group AT-hook 2 (HMGA2), which activates the transcription of pluripotency factors. Surprisingly, we found that EpEX treatment also enhances osteogenesis of MSCs under differentiation conditions, as evidenced by increases in osteogenic markers, including Runt-related transcription factor 2 (RUNX2). Taken together, our results indicate that EpEX stimulates EGFR signaling and thereby context-dependently controls MSC states and activities, promoting cell proliferation and multipotency under maintenance conditions and osteogenesis under differentiation conditions.

The TRAX, DISC1, and GSK3 complex in mental disorders and therapeutic interventions.

Psychiatric disorders (such as bipolar disorder, depression, and schizophrenia) affect the lives of millions of individuals worldwide. Despite the tremendous efforts devoted to various types of psychiatric studies and rapidly accumulating genetic information, the molecular mechanisms underlying psychiatric disorder development remain elusive. Among the genes that have been implicated in schizophrenia and other mental disorders, disrupted in schizophrenia 1 (DISC1) and glycogen synthase kinase 3 (GSK3) have been intensively investigated. DISC1 binds directly to GSK3 and modulates many cellular functions by negatively inhibiting GSK3 activity. The human DISC1 gene is located on chromosome 1 and is highly associated with schizophrenia and other mental disorders. A recent study demonstrated that a neighboring gene of DISC1, translin-associated factor X (TRAX), binds to the DISC1/GSK3ß complex and at least partly mediates the actions of the DISC1/GSK3ß complex. Previous studies also demonstrate that TRAX and most of its interacting proteins that have been identified so far are risk genes and/or markers of mental disorders. In the present review, we will focus on the emerging roles of TRAX and its interacting proteins (including DISC1 and GSK3ß) in psychiatric disorders and the potential implications for developing therapeutic interventions.

SARS-CoV-2 N protein mediates intercellular nucleic acid dispersion, a feature reduced in Omicron.

The coronavirus nucleocapsid (N) protein is known to bind to nucleic acids and facilitate viral genome encapsulation. Here we report that the N protein can mediate RNA or DNA entering neighboring cells through ACE2-independent, receptor (STEAP2)-mediated endocytosis, and achieve gene expression. The effect is more pronounced for the N protein of wild-type SARS-CoV-2 than that of the Omicron variant and other human coronaviruses. This effect is enhanced by RANTES (CCL5), a chemokine induced by N protein, and lactate, a metabolite produced in hypoxia, to cause more damage. These findings might explain the clinical observations in SARS-CoV-2-infected cases. Moreover, the N protein-mediated function can be inhibited by N protein-specific monoclonal antibodies or p38 mitogen-activated protein kinase inhibitors. Since the N-protein-mediated nucleic acid endocytosis involves a receptor commonly expressed in many types of cells, our findings suggest that N protein may have an additional role in SARS-CoV-2 pathogenesis.

OxLDL upregulates caveolin-1 expression in macrophages: Role for caveolin-1 in the adhesion of oxLDL-treated macrophages to endothelium.

Caveolin-1, a principle component of caveolae, is present in several cell types known to play an important role in the development of atherosclerosis. In this study, its distribution and expression were studied in the arterial walls of hypercholesterolemic rabbits and apo-E-deficient mice and in oxidized low-density lipoprotein (oxLDL)-treated RAW264.7 macrophages. Immunohistochemical studies showed that staining for caveolin-1 expression was stronger in atherosclerotic lesions in hypercholesterolemic rabbits and apo-E-deficient mice compared to normal rabbits and mice and was closely associated with macrophages. OxLDL treatment increased caveolin-1 protein expression in RAW264.7 macrophages in a time- and dose-dependent manner. The increase in caveolin-1 expression was dependent on phosphorylation of the mitogen-activated protein kinases (MAPKs) extracellular signal-regulated kinase1/2 (ERK1/2), p38, and Jun N-terminal kinase (JNK) and the transcriptional activation and translocation of nuclear factor-?B (NF-kappaB). OxLDL also induced caveolin-1 mRNA expression and this effect was not seen in the presence of inhibitors for transcription or de novo protein synthesis. OxLDL increased the adhesion of RAW264.7 macrophages to endothelial cells via an increase in caveolin-1 expression, and the adhesion was reduced by the use of anti-caveolin-1 antibody or caveolin-1-specific shRNA. These results show that oxLDL increases caveolin-1 expression in macrophages through the MAPKs/NF-kappaB pathway. The caveolin-1 levels are closely associated with the adherence of monocytes/macrophages to endothelial cells and their accumulation within the arterial intima after hypercholesterolemia insult, resulting in the progression of atherosclerosis.

Extracellular domain of EpCAM enhances tumor progression through EGFR signaling in colon cancer cells.

Epithelial cell adhesion molecule (EpCAM) is highly expressed in colon cancers, but its role in cancer progression remains to be elucidated. In this work, we found that the extracellular domain of EpCAM (EpEX) activated EGFR and downstream ERK1/2 signaling to promote colon cancer cell migration and proliferation, as well as tumor growth. Mechanistically, we discovered that EpEX-EGFR-ERK1/2 signaling positively regulated intramembrane proteolysis (RIP) of EpCAM and shedding of the intracellular domain (EpICD). Treatment with an EGFR inhibitor ablated the EpEX-induced phosphorylation of ERK1/2 and AKT. Additionally, treatment with inhibitors of either EGFR or MEK decreased EpEX-induced EpICD shedding and further revealed that EpICD is necessary for nuclear accumulation of ß-catenin and the induction of HIF1α target gene expression in vitro and in vivo. Moreover, an anti-EpCAM neutralizing monoclonal antibody, EpAb2-6, inhibited the nuclear translocation of EpICD and ß-catenin and induced apoptosis in colon cancer cells. Importantly, analysis of colorectal cancer tissues showed that nuclear accumulation of EpICD was highly correlated with metastasis and poor prognosis, suggesting that it may play an important functional role in cancer progression. Thus, we provide novel insights into the mechanisms and functions of EpEX-mediated signaling, which may be considered as a promising target for the treatment of colon cancer.

EpEX/EpCAM and Oct4 or Klf4 alone are sufficient to generate induced pluripotent stem cells through STAT3 and HIF2α.

Epithelial cell adhesion molecule (EpCAM) was reported to be cleaved into extracellular domain of EpCAM (EpEX) and intracellular domain of EpCAM (EpICD). We previously reported that EpCAM serves as a potent stem cell marker which is highly and selectively expressed by undifferentiated rather than differentiated hESC. However, the functional role of EpCAM remains elusive. Here, we found that EpEX and EpCAM enhance the efficiency of OSKM reprogramming. Interestingly, Oct4 or Klf4 alone, but not Sox2, can successfully reprogram fibroblasts into iPSCs with EpEX and EpCAM. Moreover, EpEX and EpCAM trigger reprogramming via activation of STAT3, which leads to the nuclear-translocation of HIF2α. This study reveals the importance of a novel EpEX/EpCAM-STAT3-HIF2α signal in the reprogramming process, and uncovers a new means of triggering reprogramming by delivery of soluble and transmembrane proteins.

Ganoderma lucidum polysaccharides attenuate endotoxin-induced intercellular cell adhesion molecule-1 expression in cultured smooth muscle cells and in the neointima in mice.

The expression of adhesion molecules on vessels and subsequent leukocyte recruitment are critical events in vascular diseases and inflammation. The aim of the present study was to examine the effects of an extract of Ganoderma lucidum (Reishi) polysaccharides (EORP), which is effective against cancer and immunological disorders, on adhesion molecule expression by human aortic smooth muscle cells (HASMCs) and the underlying mechanism. EORP significantly suppressed lipopolysaccharide (LPS)-induced intercellular cell adhesion molecule-1 (ICAM-1) mRNA and protein expression and reduced the binding of human monocytes to LPS-stimulated HASMCs. Immunoprecipitation and real-time polymerase chain reaction demonstrated that EORP markedly reduced the interaction of human antigen R protein (HuR) with the 3'-UTR of ICAM-1 mRNA in LPS-stimulated HASMCs. EORP treatment also suppressed extracellular signal-regulated kinase (ERK) phosphorylation and reduced the density of the shifted bands of nuclear factor (NF)-kappaB after LPS-induced activation. In an endothelial-denuded artery model in LPS-treated mice, daily oral administration of EORP for 2 weeks decreased neointimal hyperplasia and ICAM-1 expression in the plasma and neointima. These results provide evidence that EORP attenuates LPS-induced adhesion molecule expression and monocyte adherence and that this protective effect is mediated by decreased ERK phosphorylation and NF-kappaB activation. These findings suggest that EORP has anti-inflammatory properties and could prove useful in the prevention of vascular diseases and inflammatory responses.

Sesamin attenuates intercellular cell adhesion molecule-1 expression in vitro in TNF-alpha-treated human aortic endothelial cells and in vivo in apolipoprotein-E-deficient mice.

Sesame lignans have antioxidative and anti-inflammatory properties. We focused on the effects of the lignans sesamin and sesamol on the expression of endothelial-leukocyte adhesion molecules in tumor necrosis factor-alpha (TNF-alpha)-treated human aortic endothelial cells (HAECs). When HAECs were pretreated with sesamin (10 or 100 microM), the TNF-alpha-induced expression of intercellular cell adhesion molecule-1 (ICAM-1) was significantly reduced (35 or 70% decrease, respectively) by Western blotting. Sesamol was less effective at inhibiting ICAM-1 expression (30% decrease at 100 microM). Sesamin and sesamol reduced the marked TNF-alpha-induced increase in human antigen R (HuR) translocation and the interaction between HuR and the 3'UTR of ICAM-1 mRNA. Both significantly reduced the binding of monocytes to TNF-alpha-stimulated HAECs. Sesamin significantly attenuated TNF-alpha-induced ICAM-1 expression and cell adhesion by downregulation of extracellular signal-regulated kinase 1/2 and p38. Furthermore, in vivo, sesamin attenuated intimal thickening and ICAM-1 expression seen in aortas of apolipoprotein-E-deficient mice. Taken together, these data suggest that sesamin inhibits TNF-alpha-induced extracellular signal-regulated kinase/p38 phosphorylation, nuclear translocation of NF-kappaB p65, cytoplasmic translocalization of HuR and thereby suppresses ICAM-1 expression, resulting in reduced adhesion of leukocytes. These results also suggest that sesamin may prevent the development of atherosclerosis and inflammatory responses.

Atorvastatin induces thrombomodulin expression in the aorta of cholesterol-fed rabbits and in TNFalpha-treated human aortic endothelial cells.

Expression of functionally active thrombomodulin (TM) on endothelial cells is critical for vascular thromboresistance. 3-Hydroxyl-3-methyl coenzyme A reductase inhibitors (statins) can protect the vasculature from inflammation and atherosclerosis caused by cholesterol-dependent and cholesterol-independent mechanisms. In the present study, the effects of atorvastatin on TM expression in the aorta of cholesterol-fed rabbits and in TNFalpha-treated human aortic endothelial cells (HAECs) were investigated. When rabbits were fed a 0.5% cholesterol diet with and without supplementation with atorvastatin for 9 weeks, the neointimal area in the thoracic aorta of the atorvastatin-treated group was significantly reduced and there was significant induction of TM protein expression. In HAECs, TNFalpha treatment decreased the expression of TM in a time- and dose-dependent manner and atorvastatin pretreatment upregulated the expression of TM mRNA and protein in HAECs with or without TNFalpha treatment. Atorvastatin also inhibited monocyte adhesion to control and TNFalpha-treated HAECs via TM expression. ERK1/2 phosphorylation was significantly reduced by 24 h pretreatment with atorvastatin, whereas TNFalpha increased the phosphorylation of the MAPKs, p38, JNK, and ERK1/2. Blocking the transcriptional activation of NF-kappaB and nuclear translocation of NF-kappaB p65 prevented the TNFalpha-induced downregulation of TM. Atorvastatin regulated TM expression in control and TNFalpha-treated HAECs by inhibiting the activation of ERK and NF-kappaB. The increase in endothelial TM activity in response to atorvastatin constitutes an important pleiotropic effect of this commonly used compound and may be of clinical significance in cardiovascular disorders in which deficient endothelial TM plays a pathophysiological role.