Deletion of Elovl5 leads to dyslipidemia and atherosclerosis in LDLR-deficient mice.

Atherosclerosis is a chronic inflammatory disease for which hepatic steatosis and atherogenic dyslipidemia are significant risk factors. We investigated the effects of endogenously generated very-long-chain polyunsaturated fatty acids (VL-PUFAs) on dyslipidemia and atherosclerosis development using mice that lack ELOVL5, a PUFA elongase that is required for the synthesis of arachidonic acid, EPA, and DHA from the essential fatty acids linoleic and linolenic acids, and the LDL receptor (LDLR). Elovl5-/-;Ldlr-/- mice manifest increased liver triglyceride and cholesterol concentrations due to the activation of sterol regulatory element binding protein-1, a transcription factor that activates enzymes required for de novo lipogenesis. Plasma levels of triglycerides and cholesterol in VLDL, IDL, and LDL were markedly elevated in Elovl5-/-;Ldlr-/- mice fed a chow and the mice exhibited marked aortic atherosclerotic plaques. Bone marrow-derived monocytes from wild-type (WT) and Elovl5-/- mice were polarized to M1 and M2 macrophages, and the effects of ELOVL5 on inflammatory activity were determined. There were no differences in most of the markers tested for M1 and M2 polarized cells between WT and Elovl5-/- cells, except for a slight increase in PGE2 secretion in Elovl5-/- cells, likely due to elevated Cox-2 expression. These results suggest that the deletion of Elovl5 leads to hepatic steatosis and dyslipidemia, which are the major factors in severe atherosclerosis in Elovl5-/-;Ldlr-/- mice.

Tumor-mediated 4-1BB induces tumor proliferation and metastasis in the colorectal cancer cells.

AIMS: 4-1BB is a member of the tumor necrosis factor receptor superfamily that mainly expressed on activated T-cells and plays important roles in cell proliferation and survival of T-cells and natural killer cells. The roles of 4-1BB in immune cells have been intensively studied, whereas little is known about the expression and roles of 4-1BB in cancer cells. MAIN METHODS: In the present study, we investigated 4-1BB expression in colorectal cancer tissues from human patients and established colorectal cancer cells, using mRNA expression, FACS, and immunostaining. Cancer cell proliferation and metastasis regulated by transfected 4-1BB was evaluated by cell growth rate, colony forming assay, cell migration, and Western blot with antibodies which are involved in epithelial-mesenchymal transition and anti-apoptosis. Expression of 4-1BB was knockdown by 4-1BB shRNA to prove that 4-1BB was involved in the cell proliferation. In vivo, 4-1BB transfected cancer cells were injected into mice, to induce tumor local region or lung. KEY FINDINGS: We found that colorectal cancer tissues from human patients and established colorectal cancer cells expressed 4-1BB at the high level. The higher expression of 4-1BB proliferated faster. In addition, we identified two forms of 4-1BB detected in colorectal cancer cells: full length form that was located on the plasma membrane and a short soluble form in the cytosol. The soluble form was also detected in the plasma from the mice with tumor xenografts expressed 4-1BB. SIGNIFICANCE: Tumor-mediated 4-1BB expression in the colorectal cancer cells showed effects on cancer cell proliferation, invasion, and metastasis.

Arachidonic acid induces ER stress and apoptosis in HT-29 human colon cancer cells.

Polyunsaturated fatty acids (PUFAs) have important functions in biological systems. The beneficial effects of dietary PUFAs against inflammatory diseases, cardiovascular diseases, and metabolic disorders have been shown. Studies using cancer cells have presented the anti-tumorigenic effects of docosahexaenoic acid (DHA), an n-3 PUFA, while arachidonic acid (AA), an n-6 PUFA, has been shown to elicit both pro- and anti-tumorigenic effects. In the current study, the anti-tumorigenic effects of AA were evaluated in HT-29 human colon cancer cells. Upon adding AA in the media, more than 90% of HT-29 cells died, while the MCF7 cells showed good proliferation. AA inhibited the expression of SREBP-1 and its target genes that encode enzymes involved in fatty acid synthesis. As HT-29 cells contained lower basal levels of fatty acid synthase, a target gene of SREBP-1, than that in MCF7 cells, the inhibitory effects of AA on the fatty acid synthase levels in HT-29 cells were much stronger than those in MCF-7 cells. When oleic acid (OA), a monounsaturated fatty acid that can be synthesized endogenously, was added along with AA, the HT-29 cells were able to proliferate. These results suggested that HT-29 cells could not synthesize enough fatty acids for cell division in the presence of AA because of the suppression of lipogenesis. HT-29 cells may incorporate more AA into their membrane phospholipids to proliferate, which resulted in ER stress, thereby inducing apoptosis. AA could be used as an anti-tumorigenic agent against cancer cells in which the basal fatty acid synthase levels are low.

Tumor-suppressing miR-141 gene complex-loaded tissue-adhesive glue for the locoregional treatment of hepatocellular carcinoma.

microRNAs (miRNAs) regulate gene expression post-transcriptionally and have been extensively tested as therapeutic molecules against several human diseases. In vivo delivery of miRNAs needs to satisfy the following conditions: safety, efficiency, and long-term therapeutic effectiveness. To satisfy these conditions, we developed a tissue-adhesive nucleotide-polymer complex (NPX-glue) for in vivo delivery of miRNAs to treat hepatocellular carcinoma (HCC). Methods: Polyallylamine (PAA), a cationic polymer, was mixed with tumor-suppressing miR-141 to form NPX and then mixed with partially oxidized alginate (OA) to form NPX-glue. Delivery efficiency of miR-141:NPX-glue was determined in cultured HCC cells and in an implanted HCC tumor model. In vivo tumor-suppressive effects of miR-141 on HCC were examined in mice upon intratumoral injection of miR-141:NPX-glue. Result: NPX-glue was generated by mixing of NPX with OA, which eliminated the inherent cytotoxic effect of NPX. NPX-glue led to the efficient delivery of miR-141 and plasmid to cultured cells and solid tumors in mice, where their expression was maintained for up to 30 days. Upon intratumoral injection of miR-141:NPX-glue, the growth of the tumors was dramatically retarded in comparison with the negative control, NCmiR:NPX-glue, (p < 0.05). Molecular examination proved miR-141:NPX-glue efficiently regulated the target genes including MAP4K4, TM4SF1, KEAP1, HDGF, and TIAM1 and finally induced apoptosis of cancer tissues. Conclusion: Here, we show that NPX-glue delivers therapeutic miR-141 to solid tumors in a safe, stable, and long-term manner and prove that locoregional treatment of HCC is possible using the NPX-glue system.

Liver receptor homolog-1 regulates mouse superoxide dismutase 2.

Liver receptor homolog-1 (LRH-1) is a nuclear receptor that plays an important role in the regulation of bile acid biosynthesis, cholesterol reverse transport, steroidogenesis, and exocrine pancreatic enzyme production. In the current study, previously published data from a genome wide analysis of LRH-1 binding in the liver were re-analyzed to identify new LRH-1 targets and propose new roles for LRH-1 in the liver. Superoxide dismutase 2 (Sod2) was identified, which contains putative LRH-1 binding sites in the proximal promoter. When hepatocytes were treated with the LRH-1 agonist RJW101, Sod2 expression was dramatically increased and reactive oxygen species (ROS) production, which was induced by a high concentration of palmitate, was significantly reduced. A LRH-1 binding site was mapped to -288/-283 in the Sod2 promoter, which increased Sod2 promoter activity in response to LRH-1 and its agonist. LRH-1 binding to this site was confirmed using a chromatin immunoprecipitation assay. These results suggest that Sod2 is a target gene of LRH-1, and that LRH-1 agonists can mediate a reduction in ROS production and oxidative stress driven by an excess of fatty acids, as exhibited in nonalcoholic fatty liver disease.

Estrogen-dependent expression of sine oculis homeobox 1 in the mouse uterus during the estrous cycle.

The sine oculis homeobox 1 (SIX1) is a member of the Six gene family. SIX1 is involved in tissue development by regulating proliferation, apoptosis, and differentiation. However, function of SIX1 in the uterus remains unknown. Here, we found that Six1 expression is regulated along the estrous cycle in mouse uterus. Six1 expression was significantly increased at estrus stage and decreased at the rest of stages. SIX1 is detected in the luminal and glandular epithelium of uterine endometrium at the estrus stage. Estrogen injection increased Six1 expression in the ovariectomized mouse uterus, whereas progesterone had no effect on its expression. Estrogen receptor antagonist inhibited estrogen-induced Six1 expression. Our findings imply that SIX1 may play a role as an important regulator to orchestrate the dynamic of uterine endometrium in response to estrogen level during the estrous cycle. These results will give us a better understanding of uterine biology.

Cisplatin Induces Overactivation of the Dormant Primordial Follicle through PTEN/AKT/FOXO3a Pathway which Leads to Loss of Ovarian Reserve in Mice.

Cisplatin is a first-line chemotherapeutic agent for ovarian cancer that acts by promoting DNA cross links and adduct. However drug resistance and considerable side effects including reproductive toxicity remain a significant challenge. PTEN is well known as a tumor suppressor function which plays a fundamental role in the regulation of the cell cycle, apoptosis and development of cancer. At the same time PTEN has been revealed to be critically important for the maintenance of the primordial follicle pool. In this study, we investigated the role of PTEN/Akt/FOXO3 pathway in cisplatin-induced primordial follicle depletion. Cisplatin induced ovarian failure mouse model was used to evaluate how this pathway involves. In vitro maturation was used for oocyte rescue after cisplatin damage. We found that cisplatin treatment decreased PTEN levels, leading to a subsequent increase in the phosphorylation of key molecules in the pathway. The activation of the PTEN/Akt/FOXO3 pathway cascade increased cytoplasmic translocation of FOXO3a in cisplatin-treated follicles, which in turn increased the pool size of growing follicles, and rapidly depleted the number of dormant follicles. Once activated, the follicles were more prone to apoptosis, and their cumulus cells showed a loss of luteinizing hormone (LH) receptor expression, which leads to failure during final maturation and ovulation. In vitro maturation to rescue oocytes in a cisplatin-treated mouse model resulted in successful maturation and fertilization. This study is the first to show the involvement of the PTEN/Akt/FOXO3 pathway in premature ovarian failure after cisplatin treatment and the possibility of rescue through in vitro maturation.