Novel substituted methylenedioxy lignan suppresses proliferation of cancer cells by inhibiting telomerase and activation of c-myc and caspases leading to apoptosis

Conventional solvent fractionation and bioactivity based target assays were used to identify a new anti-cancer molecule from Phyllanthus urinaria, a herbal medicinal plant used in South India. At each step of the purification process the different fractions that were isolated were tested for specific anti-proliferative activity by assays measuring the inhibition of [(3)H]thymidine incorporation, and trypan blue drug exclusion. The ethyl acetate fraction that contained the bioactivity was further purified and resolved by HPLC on a preparative column. The purity of each of the fractions and their bioactivity were checked. Fraction 3 demonstrated a single spot on TLC and showed maximum anti-proliferative activity. This fraction was further purified and the structure was defined as 7'-hydroxy-3',4',5,9,9'-pentamethoxy-3,4-methylene dioxy lignan using NMR and mass spectrometry analysis. The pure compound and the crude ethyl acetate fraction which showed anti-proliferative activities were examined for ability to target specific markers of apoptosis like bcl2, c-myc and caspases and for effects on telomerase. Four specific cancer cell lines HEp2, EL-1 monocytes, HeLa and MCP7 were used in this study. The results indicate that 7'-hydroxy-3',4',5,9,9'-pentamethoxy-3,4-methylene dioxy lignan was capable of inhibiting telomerase activity and also could inhibit bcl2 and activate caspase 3 and caspase 8 whose significance in the induction of apoptosis is well known. We believe that this compound could serve as a valuable chemotherapeutic drug after further evaluations.     

Circulating levels of insulin-like growth factor binding protein-1 in relation to insulin resistance, type 2 diabetes mellitus, and metabolic syndrome (Chennai Urban Rural Epidemiology Study 118)

The objective was to assess the association of insulin-like growth factor binding protein-1 (IGFBP-1) with insulin resistance (IR), type 2 diabetes mellitus (T2DM), and metabolic syndrome (MS) in Asian Indians. Fifty subjects with normal glucose tolerance (NGT) and 50 with T2DM were randomly selected from the Chennai Urban Rural Epidemiology Study. Insulin-like growth factor binding protein-1 was measured by sandwich enzyme-linked immunosorbent assay. Serum insulin was estimated using Dako (Glostrup, Denmark) kits. Insulin resistance was calculated using the homeostasis model assessment. Subjects with T2DM had significantly decreased levels of IGFBP-1 (21.7 ± 3.5 ng/mL) compared with NGT subjects (34.4 ± 7.6 ng/mL, P < .001). The IGFBP-1 was significantly lower in NGT subjects with IR as measured by the homeostasis model assessment (25.5 ± 6.5 ng/mL) compared with NGT subjects without IR (40.7 ± 9.5 ng/mL, P < .001). On regression analysis, IR showed a significant association with IGFBP-1 even after adjusting for age, sex, body mass index, and glycated hemoglobin (β = -3.714, P < .001). Type 2 diabetes mellitus was significantly associated with IGFBP-1 even after adjusting for age, sex, and body mass index (β = -12.798, P < .001). The IGFBP-1 levels decreased with increasing number of metabolic abnormalities (P for trend < .001). Logistic regression analysis showed that IGFBP-1 had a strong negative association with MS even after adjusting for age and sex (odds ratio, 0.942; 95% confidence interval, 0.914-0.971; P < .001). Among Asian Indians, lower levels of circulating IGFBP-1 are seen in subjects with IR, T2DM, and MS.     

Cyclic Fatigue Resistance of RaCe and Mtwo Rotary Files in Continuous Rotation and Reciprocating Motion

Introduction

The purpose of this study was to evaluate and compare the cyclic fatigue resistance of RaCe (FKG Dentaire, La Chaux-de-Fonds, Switzerland) and Mtwo (VDW, Munich, Germany) rotary files in continuous rotation and reciprocating motion.

Methods

A total of 60 new rotary Mtwo and RaCe files (ISO size = 25, taper = 0.06, length = 25 mm) were selected and randomly divided into 4 groups (n = 15 each): Mt_c (Mtwo NiTi files in continuous rotation), R_c (RaCe NiTi files in continuous rotation), Mt_r (Mtwo NiTi files in reciprocating motion), and R_r (RaCe NiTi files in reciprocating motion). A cyclic fatigue testing device was fabricated with a 60° angle of curvature and a 5-mm radius. All instruments were rotated or reciprocated until fracture occurred. The time taken for each instrument to fracture and the length of the broken fragments were recorded. All the fractured files were analyzed under a scanning electron microscope to detect the mode of fracture. The Kolmogorov-Smirnov test was used to assess the normality of samples distribution, and statistical analysis was performed using the independent sample t test.

Results

The time taken for the instruments of the Mt_r and R_r groups to fail under cyclic loading was significantly longer compared with the Mt_c and R_c groups (P < .001). Scanning electron microscopic observations showed that the instruments of all groups had undergone a ductile mode of fracture. The length of the fractured segments was between 5 and 6 mm, which was not statistically significant among the experimental groups.

Conclusions

Mtwo and RaCe rotary instruments showed a significantly higher cyclic fatigue resistance in reciprocating motion compared with continuous rotation motion.     

Estrogen and ERα enhanced β‐catenin degradation and suppressed its downstream target genes to block the metastatic function of HA22T hepatocellular carcinoma cells via modulating GSK‐3β and β‐TrCP expression

In our previous experiments, we found β‐catenin was highly expressed in the tumor area with high invasive ability and poor prognosis. In this study, we have examined the mechanism by which ERα regulates β‐catenin expression as well as the metastasis ability of hepatocellular cancer HA22T cells. To identify whether the anticancer effect of estrogen and ERα is mediated through suppression of β‐catenin expression, we co‐transfected pCMV‐β‐catenin and ERα into HA22T cells, and determined the cell motility by wound healing, invasion, and migration assays. Results showed that estrogen and/or ERα inhibited β‐catenin gene expression and repressed HA22T cell motility demonstrated that similar data was observed in cells expressing the ERα stable clone. Moreover, we examined the protein‐protein interaction between ERα and β‐catenin by immunostain, co‐immunoprecipitation, and Western blotting. E2 enhanced the binding of ERα with β‐catenin and then triggered β‐catenin to bind with E3 ligase (βTrCP) to promote β‐catenin degradation. Finally by employing systematic ChIP studies, we showed ERα can interact directly with the β‐catenin promoter region following E2 treatment. All our results reveal that estrogen and ERα blocked metastatic function of HA22T cells by modulating GSK3β and βTrCP expression and further enhanced β‐catenin degradation and suppressed its downstream target genes. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 519–529, 2017.