Isoamylamine Induces B16-F1 Melanoma Cell Autophagy by Upregulating the 5' Adenosine Monophosphate-Activated Protein Pathway.

Isoamylamine (IA) is an aliphatic monoamine molecule present in cheese, eggs, and wine. It belongs to the family of polyamines and also can be synthesized endogenously. It has been known that regulation of polyamines in cells is related to cell cycle and tumor formation. Malignant melanoma is difficult to treat and easily resistant to chemotherapy/radiotherapy through autophagy. In this study, we aim to clarify whether IA has a growth control effect on melanoma tumor cells and the regulatory mechanism. We treated B16-F1 melanoma cells with IA at concentrations of 0, 200, 400, and 600 ppm for 24 h. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was checked for cell viability and results showed that IA has an inhibitory effect on B16-F1 melanoma cells. The signaling molecules, which included Raf/MEK/ERK, were activated, while MSK1 and protein kinase B (AKT) were suppressed. Autophagy was also confirmed to be induced by IA. The acridine orange stain-positive cells were increased and BECN-1/LC3 upregulated. The data also showed that the autophagy regulatory molecule, 5'-adenosine monophosphate-activated protein kinase (AMPK), was induced after IA treatment, so we used dorsomorphin to inhibit AMPK and found that it could suppress autophagy. In conclusion, IA has an effect of inducing autophagy in B16-F1 cells and it is regulated through AMPK.

The minor C-allele of the rs2014355 variant in ACADS gene is associated with exercise-induced increase in HDL cholesterol levels in Taiwanese adults.

ABSTRACT: We investigated the association between high-density lipoprotein cholesterol (HDL-C) and rs2014355 variant in the gene, short-chain acyl-coenzyme A dehydrogenase (ACADS) based on exercise habits.Data collected between 2008 and 2015 for individuals aged 30 to 70 years were available in the Taiwan Biobank (TWB) database. Backward stepwise linear regression was used to evaluate the associations of rs2014355 and exercise with HDL-C levels.We analyzed data of 5515 physically active and 4169 inactive biobank participants. The HDL-C concentrations were higher in the exercise compared to no exercise group (beta value, ß = 1.79856; P < .0001). We observed that the test for interaction was significant for the ACADS rs2014355 variant and exercise (P for interaction =.0412). Multivariate analyses showed significant association between TC+CC genotype and HDL-C in the exercise (ß = 1.09785; P value = .0146) compared to the no-exercise group (ß = -0.03754, P = .9154).In summary, the association between HDL-C and exercise differed significantly with respect to ACADS rs2014355 genotypes. Compared to the TT genotype, the TC+CC genotype together with exercise was associated with higher levels of HDL-C.

Interaction of Osteoarthritis and BMI on Leptin Promoter Methylation in Taiwanese Adults.

Leptin (LEP) regulates glucose metabolism and energy storage in the body. Osteoarthritis (OA) is associated with the upregulation of serum LEP. LEP promoter methylation is associated with obesity. So far, few studies have explored the association of BMI and OA with LEP methylation. We assessed the interaction between body mass index (BMI) and OA on LEP promoter methylation. Data of 1114 participants comprising 583 men and 558 women, aged 30-70 years were retrieved from the Taiwan Biobank Database (2008-2015). Osteoarthritis was self-reported and cases were those who reported having ever been clinically diagnosed with osteoarthritis. BMI was categorized into underweight, normal weight, overweight, and obesity. The mean LEP promoter methylation level in individuals with osteoarthritis was 0.5509 ± 0.00437 and 0.5375 ± 0.00101 in those without osteoarthritis. The interaction between osteoarthritis and BMI on LEP promoter methylation was significant (p-value = 0.0180). With normal BMI as the reference, the mean LEP promoter methylation level was significantly higher in obese osteoarthritic individuals (ß = 0.03696, p-value = 0.0187). However, there was no significant association between BMI and LEP promoter methylation in individuals without osteoarthritis, regardless of BMI. In conclusion, only obesity was significantly associated with LEP promoter methylation (higher levels) specifically in osteoarthritic patients.

Juniperus communis Suppresses Melanoma Tumorigenesis by Inhibiting Tumor Growth and Inducing Apoptosis.

Melanoma, which has a high metastatic capacity and death rate, is a common skin cancer in Western countries. The purpose of this study was to address whether Juniperus communis (JCo) extract is effective in the suppression of melanoma and to elucidate the anticancer mechanisms involved in vitro and in vivo. The antitumor capacities of JCo extract on tumor suppression and toxicity were evaluated and the results demonstrated that the tumor burden was reduced via mediation of cell cycle, reduction of autocrine signaling, and induction of apoptosis. Moreover, JCo extract significantly prolonged the survival rate of the test subjects with only low pathological and physiological toxicity. Additionally, JCo extract also reduced cancer stem cell-related angiogenic and metastatic proteins in the process of tumor elimination. Based on these results, this study suggests that JCo extract suppresses tumor growth and induces apoptosis, and JCo extract may be useful for the prevention of melanoma tumorigenesis.

Mulberry leaf polyphenol extract induced apoptosis involving regulation of adenosine monophosphate-activated protein kinase/fatty acid synthase in a p53-negative hepatocellular carcinoma cell.

The polyphenols in mulberry leaf possess the ability to inhibit cell proliferation, invasion, and metastasis of tumors. It was reported that the p53 status plays an important role in switching apoptosis and the cell cycle following adenosine monophosphate-activated protein kinase (AMPK) activation. In this study, we aimed to detect the effect of the mulberry leaf polyphenol extract (MLPE) on inducing cell death in p53-negative (Hep3B) and p53-positive (Hep3B with transfected p53) hepatocellular carcinoma cells and also to clarify the role of p53 in MLPE-treated cells. After treatment of the Hep3B cells with MLPE, apoptosis was induced via the AMPK/PI3K/Akt and Bcl-2 family pathways. Transient transfection of p53 into Hep3B cells led to switching autophagy instead of apoptosis by MLPE treatment. We demonstrated that acridine orange staining and protein expressions of LC-3 and beclin-1 were increased in p53-transfected cells. These results implied induction of apoptosis or autophagy in MLPE-treated hepatocellular carcinoma cells can be due to the p53 status. We also found MLPE can not only activate AMPK but also diminish fatty acid synthase, a molecular target for cancer inhibition. At present, our results indicate MLPE can play an active role in mediating the cell death of hepatocellular carcinoma cells and the p53 might play an important role in regulating the death mechanisms.

Mithramycin inhibits human epithelial carcinoma cell proliferation and migration involving downregulation of Eps8 expression.

Mithramycin is an inhibitor of the binding of the Sp-family transcription factor to the GC box. Many studies show that mithramycin may reduce the expression of many oncogenes by inhibiting the mRNA and protein synthesis and it has been used as an antibiotic chemotherapy drug for a long time. Recently, Eps8 (EGFR pathway substrate 8) has been revealed to be a novel proto-oncogene related to cellular transformation, Rac activation and actin barbed-end-capping activity. Therefore, the aim of this study was to verify whether Eps8 might be regulated by mithramycin. Results showed that mithramycin could reduce the mRNA and protein levels of Eps8 in dose- and time-dependent manners in several cancer cell lines. Furthermore, cell growth and migration ability were also reduced significantly by mithramycin treatment. Since Src is a well-known Eps8 activity enhancer, a v-Src transfected IV5 cell line was subjected to mithramycin treatment and then analyzed to show that Src expression was unable to restore the mithramycin-induced decrease in Eps8 expression, cell growth, and migration ability. To further confirm the above mentioned results, the expression of Eps8 was eliminated by a transient transfection with siRNA and subsequent analysis showed that silencing of Eps8 might also lead to a reduced growth and migration ability of cancer cells. These findings suggested that Eps8 was involved in the regulation of growth and motility of cancer cells and mithramycin might exert its anticancer ability via a pathway involving the downregulation of Eps8.

Elevated circulatory MMP-2 and MMP-9 levels and activities in patients with rheumatoid arthritis and systemic lupus erythematosus.

OBJECTIVES: Matrix metalloproteinases (MMPs) are suggested to play important roles in rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). This study is to examine the MMPs expressions and activities in Taiwanese RA and SLE patients. DESIGN AND METHODS: Levels and activities of plasma MMP-2 and MMP-9 were investigated by enzyme-linked immunosorbent assay and zymography, respectively. RESULTS: MMP-2 levels in control subjects, RA and SLE patients were 146.1+/-34.2, 194.0+/-24.2 and 208.9+/-75.9 ng/mL respectively, and for MMP-9 were 51.4+/-57.1, 567.7+/-313.1 and 208.7+/-105.5 ng/mL respectively. Both MMP-2 and MMP-9 levels and activities from all patients were significantly higher than that from control subjects. CONCLUSIONS: MMP-2 levels in both patients groups were approximately 1.3-1.4 folds higher than that in control subjects, notably, MMP-9 levels were 11- and 4-folds significantly higher, respectively, in RA and SLE patients. The results which MMP-2 and MMP-9 levels and activities are significantly elevated support the involvement of MMPs proteins in these autoimmune disorders.

Induction of antiphospholipid antibodies and antiphospholipid syndrome-like autoimmunity in naive mice with antibody against human parvovirus B19 VP1 unique region protein.

BACKGROUND: Previous studies have postulated a connection between human parvovirus B19 (B19) infection and anti-phospholipid antibodies (APhL). B19 infection and anti-phospholipid syndrome (APS) exhibit congruent symptoms. Recently, phospholipase A2 (PLA2)-like activity has been linked to the VP1 unique region (VP1u) of B19. However, the precise role of B19-VP1u in pathogenesis of autoimmunity is still obscure. METHODS: To elucidate the roles of VP1u in B19 infection and autoimmunity, the reactivity of B19-VP1u proteins with various autoantibodies were evaluated by ELISA and immunoblotting. Rabbits were immunized with purified recombinant B19-VP1u protein to generate anti-sera. Absorption experiments were conducted to determine the binding specificity of rabbit anti-sera against B19-VP1u, cardiolipin (CL) and beta-2-glycoprotein I (beta2GPI). Moreover, the effects of passive transfer of polyclonal rabbit anti-B19-VP1u IgG antibodies on platelets, activated partial thromboplastin time (aPTT), and autoantibodies were assessed. RESULTS: Autoantibodies against CL, beta2GPI, and phospholipid (PhL) in sera from patients with B19 infection, were cross-reactive with B19-VP1u. Consistently, sera from rabbits immunized with recombinant B19-VP1u protein displayed raised detectable immunoglobulins against B19-VP1u, CL, beta2GPI and PhL. Additionally, the mice immunized with anti-B19-VP1u IgG developed thrombocytopenia, prolongation of aPTT, and autoantibody against beta2GPI and PhL. CONCLUSIONS: These experimental results suggested the association between B19-VP1u and production of anti-beta2GPI antibodies, APhL, and APS-like autoimmunity. Altogether, it may provide a clue in understanding the role of B19-VP1u in inducing autoantibodies and B19-associated APS manifestations.

Cytokine promoter polymorphisms in Taiwanese patients with Graves' disease.

OBJECTIVES: This study aimed to examine the association between promoter polymorphisms of Th1 and Th2 cytokine genes [interleukin-4 (IL-4 T-34C, A-81G, C-285T and T-589C), IL-6 (G-174C), IL-10 (A-592C and T-819C) and tumour necrosis factor-alpha (TNF-alpha G-238A and G-308A)] and Graves' disease (GD) in Taiwanese population. DESIGN AND METHODS: Genomic DNA was extracted from peripheral blood cells of 137 GD patients and 189 control subjects. Cytokine gene polymorphisms were analyzed by polymerase chain reaction and restriction fragment length polymorphism. RESULTS: Genotype frequencies of TNF-alpha G-238A or G-308A between control and GD subjects were significantly different. Frequencies of the high TNF-alpha secreting alleles (-238*A and -308*A) and IL-10 -819*C allele were significantly increased in GD patients. No significant differences regarding IL-4 or IL-6 gene polymorphisms between GD patients and control subjects were found. CONCLUSIONS: Our data demonstrated that TNF-alpha G-238A and G-308A genotypes were strongly associated with GD incidence.

A novel function of BCL-2 overexpression in regulatory volume decrease. Enhancing swelling-activated Ca(2+) entry and Cl(-) channel activity.

The cellular function of the oncogene bcl-2, a key regulator of apoptosis, is still debated. The goal of this study was to explore the relationship between BCL-2 overexpression and cell volume regulation by using two independent models, Madin-Darby canine kidney (MDCK) cells stably transfected with BCL-2 and MDCK clones with inducible BCL-2 expression by the lac operator/repressor. BCL-2 overexpression enhanced the capability of regulatory volume decrease (RVD), a cellular defensive process against hypotonic stress. In various clones of MDCK cells, hypotonic stress induced an outwardly rectified Cl(-) current that was significantly up-regulated by BCL-2 overexpression. Other fundamental characteristics of this channel were similar among different MDCK clones, such as sensitivity to Cl(-) channel inhibitor, anion permeability, and time-dependent inactivation at more positive potential. Most importantly, BCL-2 overexpression up-regulates the swelling-activated Ca(2+) transient that is a critical signaling for normal RVD and the activation of swelling-activated Cl(-) channel in MDCK cells. BCL-2 overexpression also enhances the capacitative Ca(2+) entry that can be differentiated from the swelling-activated Ca(2+) transient by kinetic analysis and sensitivity to Gd(3+). Moreover, neutralization of endogenous BCL-2 by antibody blocks the normal RVD response and the activation of swelling-activated Cl(-) channel in human cervical cancer HT-3 cells. These results provide a new insight into the novel function of BCL-2 overexpression in the regulation of cell volume and ion flux.