Overexpression of secretagogin promotes cell apoptosis and inhibits migration and invasion of human SW480 human colorectal cancer cells.

OBJECTIVE: In order to investigate the effect of secretagogin (SCGN) on colorectal cancer (CRC) cells apoptosis, invasion and migration in vitro. METHODS: Expression of SCGN in CRC tissues and the paired adjacent non-tumorous tissues (n = 36) and four human CRC cell lines (HT29, HCT116, SW480 and SW620) were detected. SW480 cells were transfected with the SCGN overexpression plasmid (eGFP-SCGN), si-SCGN-773, and the corresponding negative controls (NCs). Then, cell-cycle distribution, cell apoptosis, migration, invasion and expression of apoptosis- and metastasis-related proteins were detected. RESULTS: SCGN was significantly downregulated in CRC tissues as compared with the adjacent non-tumorous tissues. The expression of SCGN in HT29 and SW480 cells were lower than those in HT116 and SW620 cells. We transfected SW480 cells with SCGN overexpression plasmid eGFP-SCGN and found the increased cell apoptosis, with cell arresting at G0/G1 phase. SW480 cells with SCGN overexpression showed wider wound width and fewer invaded cells than control and blank cells, with upregulated Bax, cleaved Caspase 3 and E-cadherin, and downregulated Bcl-2 and Vimentin. We also transfected SW480 cells with si-SCGN-773 and found si-SCGN increased cell migration and invasion, but did not affect cell apoptosis and expression of related proteins. CONCLUSION: We concluded that the overexpression of SCGN in SW480 cells promoted cell apoptosis and inhibited cell migration and invasion.

Itraconazole attenuates hepatic gluconeogenesis and promotes glucose uptake by regulating AMPK pathway.

The primarily metabolic abnormality in type 2 diabetes mellitus (T2DM) is the defect in gluconeogenesis and glucose uptake. Itraconazole (ITCZ) is a traditional azole drug with anti-fungal and anticancer properties. However, limited attention has been directed towards the contribution of ITCZ to hepatic gluconeogenesis and glucose uptake in T2DM. The present study aimed to investigate the potential effects of ITCZ on hepatic gluconeogenesis and glucose uptake as well as the underlying mechanisms. No obvious change in cell viability was detected by MTT assay in HepG2 cells with ITCZ treatment at gradually increasing concentrations. Western blot analysis demonstrated that the phosphorylation level of 5' adenosine monophosphate-activated protein kinase (AMPK) was significantly elevated by ITCZ treatment at ≥5 µg/ml (P<0.05). Moreover, ITCZ repressed the gluconeogenesis of HepG2 cells, as evidenced by the dose-dependently increased glycogen synthase kinase 3ß phosphorylation level and a notably decreased glucose production rate (P<0.05). Simultaneously, the expression of peroxisome proliferator-activated receptor γ co-activator 1α, phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) in HepG2 cells was reduced by ITCZ in a dose-dependent manner (P<0.001). Furthermore, a 2-deoxyglucose uptake assay revealed that the glucose uptake of HepG2 cells was notably enhanced, accompanied by the ITCZ dose-dependent upregulation of glucose transporter-4 (GLUT-4) (P<0.05). Conversely, silencing of AMPK by small interfering RNA resulted in an increase of ITCZ-reduced gluconeogenesis and inhibition of ITCZ-induced glucose uptake with relative upregulation of PEPCK and G6Pase and downregulation of GLUT4 in the presence of 50 µg/ml ITCZ (P<0.05). Overall, the results indicated that AMPK has an important role in regulating ITCZ-induced glucose uptake by stimulating GLUT4 in HepG2 cells. Therefore, ITCZ may become a promising candidate for T2DM therapy.

Clock1a affects mesoderm development and primitive hematopoiesis by regulating Nodal-Smad3 signaling in the zebrafish embryo.

Circadian clock and Smad2/3/4-mediated Nodal signaling regulate multiple physiological and pathological processes. However, it remains unknown whether Clock directly cross-talks with Nodal signaling and how this would regulate embryonic development. Here we show that Clock1a coordinated mesoderm development and primitive hematopoiesis in zebrafish embryos by directly up-regulating Nodal-Smad3 signaling. We found that Clock1a is expressed both maternally and zygotically throughout early zebrafish development. We also noted that Clock1a alterations produce embryonic defects with shortened body length, lack of the ventral tail fin, or partial defect of the eyes. Clock1a regulates the expression of the mesodermal markers ntl, gsc, and eve1 and of the hematopoietic markers scl, lmo2, and fli1a Biochemical analyses revealed that Clock1a stimulates Nodal signaling by increasing expression of Smad2/3/4. Mechanistically, Clock1a activates the smad3a promoter via its E-box1 element (CAGATG). Taken together, these findings provide mechanistic insight into the role of Clock1a in the regulation of mesoderm development and primitive hematopoiesis via modulation of Nodal-Smad3 signaling and indicate that Smad3a is directly controlled by the circadian clock in zebrafish.