Endoplasmic reticulum stress and autophagy dysregulation in alcoholic and non-alcoholic liver diseases

Alcoholic and non-alcoholic liver diseases begin from an imbalance in lipid metabolism in hepatocytes as the earliest response. Both liver diseases share common disease features and stages (i.e., steatosis, hepatitis, cirrhosis, and hepatocellular carcinoma). However, the two diseases have differential pathogenesis and clinical symptoms. Studies have elucidated the molecular basis underlying similarities and differences in the pathogenesis of the diseases; the factors contributing to the progression of liver diseases include depletion of sulfhydryl pools, enhanced levels of reactive oxygen and nitrogen intermediates, increased sensitivity of hepatocytes to toxic cytokines, mitochondrial dysfunction, and insulin resistance. Endoplasmic reticulum (ER) stress, which is caused by the accumulation of misfolded proteins and calcium depletion, contributes to the pathogenesis, often causing catastrophic cell death. Several studies have demonstrated a mechanism by which ER stress triggers liver disease progression. Autophagy is an evolutionarily conserved process that regulates organelle turnover and cellular energy balance through decomposing damaged organelles including mitochondria, misfolded proteins, and lipid droplets. Autophagy dysregulation also exacerbates liver diseases. Thus, autophagy-related molecules can be potential therapeutic targets for liver diseases. Since ER stress and autophagy are closely linked to each other, an understanding of the molecules, gene clusters, and networks engaged in these processes would be of help to find new remedies for alcoholic and non-alcoholic liver diseases. In this review, we summarize the recent findings and perspectives in the context of the molecular pathogenesis of the liver diseases.

Sex-Biased Molecular Signature for Overall Survival of Liver Cancer Patients

Sex/gender disparity has been shown in the incidence and prognosis of many types of diseases, probably due to differences in genes, physiological conditions such as hormones, and lifestyle between the sexes. The mortality and survival rates of many cancers, especially liver cancer, differ between men and women. Due to the pronounced sex/gender disparity, considering sex/ gender may be necessary for the diagnosis and treatment of liver cancer. By analyzing research articles through a PubMed literature search, the present review identified 12 genes which showed practical relevance to cancer and sex disparities. Among the 12 sex-specific genes, 7 genes (BAP1, CTNNB1, FOXA1, GSTO1, GSTP1, IL6, and SRPK1) showed sex-biased function in liver cancer. Here we summarized previous findings of cancer molecular signature including our own analysis, and showed that sexbiased molecular signature CTNNB1High , IL6High , RHOAHigh and GLIPR1Low may serve as a female-specific index for prediction and evaluation of OS in liver cancer patients. This review suggests a potential implication of sex-biased molecular signature in liver cancer, providing a useful information on diagnosis and prediction of disease progression based on gender.

Clinical Outcomes of Occupational Exposure to N,N-Dimethylformamide: Perspectives from Experimental Toxicology

N,N-Dimethylformamide (DMF) is globally used as an organic solvent in the production of synthetic leather and resins because of its low volatility, making it an attractive industrial material. Despite its excellent property as a chemical solvent, utilization of DMF is somewhat controversial nowadays due to its hazardous effects on exposed workers in work places. Many toxification cases are being reported globally and the number of cases of liver damage is still increasing in developing countries. On account of this, a series of epidemiologic surveys are being conducted to understand the degrees of liver damage caused by DMF exposure. Furthermore, many investigations have been performed to clarify the mechanism of DMF-induced liver toxicity using both human and experimental animal models. This review summarizes the current occupational cases reported on liver damage from workers exposed to DMF in industrial work places and the research results that account for DMF-induced liver failure and possible carcinogenesis. The findings reviewed here show the synergistic toxicity of DMF exposure with other toxicants, which might occur through complicated but distinct mechanisms, which may extend our knowledge for establishing risk assessments of DMF exposure in industrial work places.

Anticancer Effect of AG60 (Acriflavine-Guanosine Compound) on the Ehrlich Cancer Cells Light Microscopic, Autoradiographic and Electron Microscopic Study / 대한해부학회지

To evaluate the effect and working mechanism of a newly developed anti-cancer drug, AG60 (acriflavine-guanosine compound, Taerim Pharm. Co. Seoul, Korea), histotologic, autoradiographic and electron microscopic studies were carried out. For the histologic study, each Ehrlich carcinoma cells (10(7) cells)-inoculated mouse was subcutaneously injected with saline (0.2 ml), 10 mg/kg of AG60, or 30 mg/kg of AG60, every other day, respectively. Animals were sacrified on the 14th day from the first injection, and tumor masses were fixed in 10% formalin solution. Tissue sections of the tumor were stained with hematoxylin and eosin. For the electron microscopic study, Ehrlich carcinoma (10(7) cells)-inoculated mice were subcutaneously injected every other day with saline (0.2 ml) or 30 mg/kg of AG60, respectively. The day after 7th injection (14th day), animals were sacrified, small piece of tumor masses were fixed in 2.5% glutaraldehyde-1.5% paraformaldehyde solution followed by fixation in 2% osmium tetroxide solution. Ultrathin sections were counter stained with uranyl acetate-lead citrate solutions, and observed with JEM 100CX electron microscope. For the autoradiographic study, each Ehrlich carcinoma (10(7) cells)-inoculated mouse was injected every day with 0.2 ml of saline, 5 mg/kg of AG60, or 30 mg/kg of AG60, respectively. The day following the last injection, each animal was given a single dose of 0.7 micricurie/g of methyl-3H-thymidine (Amersham Lab., England) through the tail vein. Seventy minutes after the thymidine injection, animals were sacrified, tumor masses were collected and fixed in 10% neutral formalin. Tissue blocks were washed, dehydrated, embedded and cut in 6 micrometer-thick sections. Deparaffinzied sections were dipped in the autradiographic emulsion E1 (Amersham Lab., England) and dried and stocked in the dark room. Filmed sections were exposured five weeks in the dark room, and were developed in the developer. Labeled indices (mean number of labeled cells per 100 cancer cells) and labeled grain indices (mean number of labeled silver grains per one cancer cell, and total granule numbers per every 100 cancer cell) were observed and calculated. The results were as follows : 1. On histological study, massive apoptosis were occured following the injection of AG60. Only small number of live cancer cells were observed. 2. On electron microscopic study, massive apoptotic figures including fragmentation of nuclei and cytoplasms, multiple nucleoli, condensation of nucleus and cytoplasm, deep invaginations and microcleft formations of nuclei, margination of heterochromatin along the inner nuclear membrane and microcleft , etc. were noticed. Giant cells represent the "tumor cell-tumor cell emperipolesis", and many of them seem to be in process of "cytolytic emperipolesis". 3. On autoradiographic study, labeled grains of 3H-thymidine were suppressed to only 11%~5% of control cancer cells following AG60 administrations. Discussed on the above experiments, it is suggested that severe suppression of DNA, RNA and protein syntheses by AG60 induce massive apoptosis of cancer cells. AG60 is expected as one of most effective anticancer drugs for the cytostatic therapy, the disease stabilization, the improved quality of life, the prolongation of life, and possibly the chemoprevention.

Autoradiographic Study on the Effect of AG60 to the DNA Synthesis of Gastric Epithelium of Mouse Inoculated with Ehrlich Carcinoma / 대한해부학회지

To study the tumor-suppression effect of a newly developed anti-tumor agent AG60 [acriflavine (1) : guanosine (1) composition, Taerim Pharm. Co., Seoul, Korea], each Ehrlich carcinoma (10(7) cells)-inoculated mouse received the subcutaneous injection of 0.2 ml of saline, 5 mg/kg of AG60, and 30 mg/kg of AG60 per day for a week. The day following the last injection, each mouse was injected with a single dose of 0.7 microcurie/g of methyl-3H-thymidine (25Ci/mmol, Amersham Lab., England) through tail vein. Seventy minutes after the thymidine injection, animals were sacrificed, and gastric tissues were collected and fixed in 10% neutral formalin. Tissue blocks were washed, dehydrated, embedded and cut in 6 micrometer-thick sections. Deparaffinized sections were coated with autoradiographic emulsion EM 1 (Amersham Lab. England) in a dark room and dried and were placed in a light-tight box. The sections were exposured for 5 weeks in the dark room, and were then developed in D-19 developer. Labeled indices (mean number of labeled cells per 100 epithelial cells in the isthmus) were observed and calculated. The results are as follows; 1. On histological study, gastric mucosa had no morphological changes following the injection of AG60. 2. On autoradiographic study, labeled grains of 3H-thymidine were restricted on the isthmus portion of the gatric gland. 3. On autoradiographic study, labeling indicies of gastric epithelial cells of normal control, experimental control, AG60 (5 mg/kg)-treated and AG60 (30 mg/kg)-treated groups were 21.9+/-0.28%, 18.8+/-0.03%, 21.6+/-1.61% and 6.3+/-0.93%, respectively. These result suggest that AG60 is expected as one of most effective anticancer drugs, and the dosage under 5 mg/kg of AG60 does not result any defect on the DNA synthesis in gastric epithelial cells.

Fluorescent Study on the Tissue-Distribution of Acriflavine-Guanosine Compound (AG60) / 대한해부학회지

The study was carried out to evaluate the tissue-distribution of acriflavine or AG60 (acriflavine-guanosine compound, 1 : 1), the newly developed anticancer remedy. Successful access or distribution of a drug to specific tissue is important to attack the cancer cells in the same area. But it also means that the drug may disturb the activities of labelled tissues or cells. On the other hand, unlabelled elements may survive from massive treatment with the drug. In this study, distribution of acriflavine or AG60 in Yac-1 leukemic cells (0.25~25 microgram/ml) and in the tissues of rats or mice (5~50 mg/kg) were evaluated. Yac-1 cells showed prominent fluorescence on the heterochromatin and more or less prominent fluorescence on the nucleoplasm, cytoplasm and plasma membrane. Cytotoxicity of AG60 led to morphologic changes such as bleb- or baloon-formation on the surface, general swelling of the cell, and lysis of the cell. Following the subcutaneous administration of acriflavine or AG60 (5~50 mg/kg) to the Ehrlich carcinoma-inoculat-ed rats or mice, most tissues including cancer cells showed acriflavine-fluorescence with some exception. The nuclei of cells of tissues were labelled more strongly than those of cytoplasm. Fluorescence was especially strong over biliary tree, renal corpuscle, gastrointestinal mucous coat, and bronchial mucous coat. But parenchymal components of central nervous system did not show any fluorescence. As shown in Yac-1 cells treated with AG60, the drug strongly attached to nucleic acids, and it induced swelling and disintegration of cancer cells. Fast turn-over of AG60 was seen in the secretory passages of bile juice, urine, gastrointestinal mucin, and bronchial mucin. The results show that AG60 could reach most tissues except parenchymes of central nervous system.

Anti-tumor Effect of the Complex of Acriflavine and Guanosine (AG60) / 대한암학회지

PURPOSE: The anti-tumor effect of the complex of acriflavine and guanosine (AG60) was investigated. MATERIALS AND METHODS: In vitro cytotoxicity of AG60 was measured using SRB assay, and in vivo antitumor activity of AG60 was examined in CDF1 mice intraperitoneally inoculated with the P388 leukemic cells and in ICR mice inguinally implanted with S-180 cells. Tumor size and mean survival time were determined. RESULTS: AG60 and acriflavine showed strong anti-tumor effect in vitro on lung cancer (A549), renal cancer (UO-31) and colon cancer (COLO205) cells. However, AG60 did not show the cytotoxicity against normal cell line, 3T3. The range of the IC50 of AG60 to the various tumor cell lines was 0.09 microgram/ml through 1.94 microgram/ml. The treatment of ascitic tumor bearing CDF1 mice with AG60 resulted in over 160% increases in the mean survival time. The most effective dose of AG60 was 30 mg/kg body weight in tumor implanted mice. In solid tumor bearing ICR mice tumor growth and progression were suppressed in response to the different doses at 30 days; 69.8% suppression of tumor size in response to acriflavine, 16.0% to guanosine, 87.7% to AG60 and 78.5% to doxorubicin. In addition, 35% increases were observed in the means survival time of AG60 treated group compared with control group. CONCLUSION: The prominant anti-tumor effects of AG60 shown in this report would represent the possibility of the clinical trials.