Camellia oil has become an important plant oil in China in recent years, but its effects on non-alcoholic fatty liver disease (NAFLD) have not been documented. In this study, the effects of camellia oil, soybean oil, and olive oil on NAFLD were evaluated by analyzing the fatty acid profiles of the plant oils, the serum lipids and lipoproteins of rats fed different oils, and by cytological and ultrastructural observation of the rats' hepatocytes. Analysis of fatty acid profiles showed that the polyunsaturated fatty acid (PUFA) n-6/n-3 ratio was 33.33 in camellia oil, 12.50 in olive oil, and 7.69 in soybean oil. Analyses of serum lipids and lipoproteins of rats showed that the levels of total cholesterol and low-density lipoprotein cholesterol in a camellia oil-fed group (COFG) were lower than those in an olive oil-fed group (OOFG) and higher than those in a soybean oil-fed group (SOFG). However, only the difference in total cholesterol between the COFG and SOFG was statistically significant. Cytological observation showed that the degree of lipid droplet (LD) accumulation in the hepatocytes in the COFG was lower than that in the OOFG, but higher than that in the SOFG. Ultrastructural analysis revealed that the size and number of the LDs in the hepatocytes of rats fed each of the three types of oil were related to the degree of damage to organelles, including the positions of nuclei and the integrity of mitochondria and endoplasmic reticulum. The results revealed that the effect of camellia oil on NAFLD in rats was greater than that of soybean oil, but less than that of olive oil. Although the overall trend was that among the three oil diets, those with a lower n-6/n-3 ratio were associated with a lower risk of NAFLD, and the effect of camellia oil on NAFLD was not entirely related to the n-6/n-3 ratio and may have involved other factors. This provides new insights into the effect of oil diets on NAFLD.
Animals ; Camellia/chemistry* ; Fatty Acids/analysis* ; Hepatocytes/ultrastructure* ; Lipid Droplets/physiology* ; Lipids/blood* ; Male ; Non-alcoholic Fatty Liver Disease/pathology* ; Plant Oils/administration & dosage* ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the anti-cancer effects of crude extract from Melia toosendan Sieb. et Zucc and its possible molecular mechanisms in vitro and in vivo.

METHODSTransonic alcohol-chloroform extraction method was used to extract toosendanin from the bark of Melia toosendan Sieb. et Zucc, and the content of toosendanin in the crude extract was measured by high performance liquid chromatography (HPLC). Anti-cancer effects of crude extract from Melia toosendan Sieb. et Zucc were investigated in in vivo and in vitro studies. In the in vitro experiment, human hepatocellular carcinoma cell lines SMMC-7721 and Hep3B were co-incubated with toosendanin crude extract of different concentrations, respectively. In the in vivo experiment, BALB/c mice were subcutaneously inoculated with mouse hepatocellular carcinoma H22 cells and treated with crude extract.

RESULTSHPLC revealed the content of toosendanin was about 15%. Crude extract from Melia toosendan Sieb. et Zucc inhibited cancer cells growth in a dose- and time-dependent manner. The 50% inhibitory concentration (IC50, 72 h) was 0.6 mg/L for SMMC-7721 cells and 0.8 mg/L for Hep3B cells. Both high-dose [0.69 mg/(kg d)] and low-dose [0.138 mg/(kg d)] crude extract could markedly suppress cancer growth, and the inhibition rate was greater than 50%. Hematoxylin and eosin staining showed necrotic area in cancers and transmission electron microscopy displayed necrotic and apoptotic cancer cells with apoptotic bodies. Immunohistochemistry showed that the expression of Bax and Fas increased and the expression of Bcl-2 reduced.

CONCLUSIONSToosendanin extract has potent anti-cancer effects via suppressing proliferation and inducing apoptosis of cancer cells in vivo and in vitro. The mechanism of apoptosis involves in mitochondrial pathway and death receptor pathway.

Animals ; Antineoplastic Agents ; pharmacology ; therapeutic use ; Apoptosis ; drug effects ; Carcinoma, Hepatocellular ; drug therapy ; pathology ; ultrastructure ; Cell Proliferation ; drug effects ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; therapeutic use ; Female ; Immunohistochemistry ; Liver Neoplasms ; drug therapy ; pathology ; ultrastructure ; Male ; Melia ; chemistry ; Mice, Inbred BALB C ; Mitochondria ; drug effects ; metabolism ; Neoplasm Transplantation ; Plant Extracts ; therapeutic use ; Reference Standards ; bcl-2-Associated X Protein ; metabolism ; fas Receptor ; metabolism

Animals ; Arsenic Poisoning ; pathology ; Imaging, Three-Dimensional ; Liver ; blood supply ; ultrastructure ; Mice

OBJECTIVE: To observe the micromorphological changes of ultrastructure, apoptosis-related proteins expression and tumor cell apoptosis after ablation with the high-intensity focused ultrasound (HIFU), and to explore the mechanisms responsible for the thermal and non-thermal effect.
 METHODS: Forty rabbits with hepatic VX2 tumors were randomly divided into a thermal group (n=20) and a non-thermal group (n=20), and were subjected to HIFU ablation with thermal or non-thermal condition, respectively. Five animals in each group were sacrificed on the 1st, 3rd, 7th or 14th day after the ablation. The changes of ultrastructure, apoptosis-related proteins expression and tumor cell apoptosis were detected.
 RESULTS: The results of transmission electron microscope (TEM) revealed more severe injury on tissue and cells in the non-thermal group than that in the thermal group. The changes of apoptosis-related proteins expression and tumor cell apoptosis in transient zone were significantly different in comparison with that in the ablated area or peripheral area between the two groups. The expression of vascular endothelial growth factor (VEGF) was at low level on the 1st and 3rd day and elevated gradually on the 7th and 14th day, with no significant difference (all P>0.05). The expression of caspase-3 reached peak on the 3rd day and decreased on the 7th and 14th day. It was significantly higher in the non-thermal group than that in the thermal group on the 3rd and 7th day (all P<0.05). The expression of NF-κB was elevated from the 3rd day and reached peak on the 7th day while decreased on the 14th day. There was no significant difference at every time point between the 2 groups (all P>0.05). The apoptosis index in the non-thermal group and the thermal group on the 3rd and 7th day were (28.60±1.14)% vs (21.80±1.92)% and (21.00±1.58)% vs (14.80±1.48)%, respectively. It was higher in the non-thermal group than that in the thermal group (both P<0.01).
 CONCLUSION Both the thermal and the non-thermal effect of HIFU can induce apoptosis in transient zone, but the latter have a stronger effect.
Animals ; Apoptosis ; Caspase 3 ; metabolism ; High-Intensity Focused Ultrasound Ablation ; Liver Neoplasms ; pathology ; ultrastructure ; NF-kappa B ; metabolism ; Neoplasms, Experimental ; pathology ; ultrastructure ; Rabbits ; Vascular Endothelial Growth Factor A ; metabolism

Donation after brain death followed by circulatory death (DBCD) is a unique practice in China. The aim of this study was to define the pathologic characteristics of DBCD liver allografts in a porcine model. Fifteen male pigs (25-30 kg) were allocated randomly into donation after brain death (DBD), donation after circulatory death (DCD) and DBCD groups. Brain death was induced by augmenting intracranial pressure. Circulatory death was induced by withdrawal of life support in DBCD group and by venous injection of 40 mL 10% potassium chloride in DCD group. The donor livers were perfused in situ and kept in cold storage for 4 h. Liver tissue and common bile duct samples were collected for hematoxylin and eosin staining, TUNEL testing and electron microscopic examination. Spot necrosis was found in hepatic parenchyma of DBD and DBCD groups, while a large area of necrosis was shown in DCD group. The apoptosis rate of hepatocytes in DBD [(0.56±0.30)%] and DBCD [(0.50 ± 0.11)%] groups was much lower than that in DCD group [(3.78±0.33)%] (P<0.05). And there was no significant difference between DBD group and DBCD group (P>0.05)). The structures of bile duct were intact in both DBD and DBCD groups, while the biliary epithelium was totally damaged in DCD group. Under electron microscope, the DBD hepatocytes were characterized by intact cell membrane, well-organized endoplasmic reticulum, mild mitochondria edema and abundant glycogens. Broken cell membrane, mild inflammatory cell infiltration and sinusoidal epithelium edema, as well as reduced glycogen volume, were found in the DBCD hepatocytes. The DCD hepatocytes had more profound cell organelle injury and much less glycogen storage. In conclusion, the preservation injury of DBCD liver allografts is much less severe than that of un-controlled DCD, but more severe than that of DBD liver allografts under electron microscope, which might reflect post-transplant liver function to some extent.
Allografts ; Animals ; Apoptosis ; Brain Death ; China ; Death ; Heart Arrest ; Hepatocytes ; pathology ; ultrastructure ; Humans ; In Situ Nick-End Labeling ; Liver ; pathology ; ultrastructure ; Liver Transplantation ; methods ; Microscopy, Electron ; Organ Preservation ; methods ; Swine ; Tissue Donors ; Tissue and Organ Procurement ; methods

OBJECTIVETo determine the roles of mitochondrial apoptosis and energy metabolism in hepatocytes during the pathogenic process of acute renal failure (ALF) by assessing disease-related differential activities of several key mitochondrial enzymes, including citrate synthase (CS), carnitine palmitoyltransferase-1 (CPT-1) and cytochrome c oxidase (COX).

METHODSThirty-two male Sprague Dawley rats were given D-galactosamine followed by and lipopolysaccharide (LPS) to induce acute liver failure and sacrificed after 4 (4 h group), 8 (8 h group) 12 (12 h group) and 24 hours (24 h group) of treatment. Eight unmodeled rats served as controls. Effects related to apoptosis were evaluated by pathological analysis of hepatic tissues and TUNEL staining. Ultrastructural changes in mitochondria were assessed by electron microscopy. The activity and expression of CS, CPT-1 and COX were measured.

RESULTSHepatocyte apoptosis was present in the 4 h treatment group and was increased obviously in the 8 h treatment group. Hepatocyte necrosis was first observed in the 12 h treatment group and was significantly higher in the 24 h treatment group, with inflammatory cell invasion. Ultrastructural changes in mitochondria were present in the 4 h treatment group, and the 24 h treatment group showed mitochondria with completely destroyed outer membranes, which resulted in mitochondrial collapse. Activity and protein expression of CS, CPT-1 and COX were increased in the 4 h group (vs. controls), were at their peak in the 8 h group (CS:t =1.481, P less than 0.01; CPT-1:t =2.619, P less than 0.05; COX:t =1.014, P less than 0.01) and showed a decreasing trend in the 12 h group. In addition, the activities of CS, CPT-1 and COX were enhanced at the stage of hepatocyte apoptosis, suggesting that these enzymes were involved in the initiation and development of ALF.

CONCLUSIONEnergy metabolism plays an important role in hepatocyte apoptosis during ALF.

Animals ; Apoptosis ; Carnitine O-Palmitoyltransferase ; metabolism ; Citrate (si)-Synthase ; metabolism ; Disease Models, Animal ; Electron Transport Complex IV ; metabolism ; Hepatocytes ; cytology ; enzymology ; Liver Failure, Acute ; metabolism ; pathology ; Male ; Mitochondria ; ultrastructure ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the effects of ancient Chinese medical formula Xiayuxue Decoction ([symbols; see text], XYXD) on activation of hepatic stellate cells (HSCs) and defenestration of sinusoidal endothelial cells (SECs) in CCl4-induced fibrotic liver of mice.

METHODSHigh performance liquid chromatography was used to identify the main components of XYXD and control the quality of extraction. C57BL/6 mice were induced liver fibrosis by CCl4 exposure and administered with XYXD for 6 weeks simultaneously. Liver tissue was investigated by hematoxylin-eosin and Sirius-red staining. Sinusoidal fenestrations were observed by scanning electronic microscopy and fluorescent immunohistochemistry of PECAM-1 (CD31). Whole liver lysates were detected of α-smooth muscle actin (α-SMA) and type-I collagen by Western blot. Primary rat HSCs-T6 cells were analyzed by detecting α-SMA, F-actin, DNA fragmentation through confocal microscopy, Western blot, terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) assay and cellomics arrayscan, respectively.

RESULTSAmygdalin and emodin in XYXD were identified. XYXD (993 mg/kg) inhibited Sirius red positive area up to 70.1% (P<0.01), as well as protein levels of α-SMA and type-I collagen by 42.0% and 18.5% (P<0.05) respectively. In vitro, XYXD (12.5 μg/mL, 50 μg/mL) suppressed the activation of HSCs and reversed the myofibroblastic HSCs into quiescent, demonstrated as inhibition of fluorescent F-actin by 32.3% and 46.6% (P<0.05). Besides, XYXD induced the apoptosis of HSC-T6 cells by 20.0% (P<0.05) and 49.5% (P<0.01), evidenced by enhanced TUNEL positivity. Moreover, ultrastructural observation suggested XYXD inhibited defenestration of SECs, which was confirmed by 31.1% reduction of protein level of CD31 (P<0.05).

CONCLUSIONSXYXD inhibited both HSCs activation and SECs defenestration which accompany chronic liver injuries. These data may help to understand the underlying mechanisms of XYXD for prevetion of chronic liver diseases.

Actins ; metabolism ; Animals ; Carbon Tetrachloride Poisoning ; drug therapy ; Collagen Type I ; metabolism ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Endothelium ; drug effects ; pathology ; Hepatic Stellate Cells ; drug effects ; pathology ; ultrastructure ; Liver Cirrhosis ; chemically induced ; drug therapy ; pathology ; Male ; Mice, Inbred C57BL ; Microscopy, Electron, Scanning ; Myofibroblasts ; drug effects ; pathology ; ultrastructure ; Platelet Endothelial Cell Adhesion Molecule-1 ; metabolism ; Primary Cell Culture ; Rats, Sprague-Dawley

Opisthorchis viverrini infection causes inflammation and liver injury leading to periductal fibrosis. Little is known about the pathological alterations in bile canaliculi in opisthorchiasis. This study aimed to investigate bile canalicular alterations in O. viverrini-infected hamsters and to examine the chemopreventive effects of curcumin on such changes. Hamsters were infected with O. viverrini and one group of animals was fed with 1% dietary curcumin supplement. Animals were examined during the acute infection phase, days 21 and 30 post-infection (PI) and chronic infection phase (day 90 PI). Scanning electron microscopy revealed that in the infected group fed with a normal diet, bile canaliculi became slightly tortuous by 30 day PI and more tortuous at day 90 PI. Transmission electron microscopy showed a reduction in microvilli density of canaliculi starting at day 30 PI, with a marked loss of microvilli at day 90 PI. These ultrastructral changes were slightly seen at day 21 PI, which was similar to that found in infected animals fed with 1% curcumin-supplemented diet. Notably, curcumin treatment prevented the reduction of microvilli density, reduced the dilation of bile canaliculi, and decreased the tortuosity of the bile canaliculi relative to non-infected animals on a normal diet at days 30 and 90 PI. These results suggest that curcumin reduces alteration of bile canaliculi and may be a promising agent to prevent the onset of bile duct abnormalities induced by O. viverrini infection.
Animals ; Anthelmintics/*administration & dosage ; Bile Canaliculi/*pathology/ultrastructure ; Chemoprevention/methods ; Cricetinae ; Curcumin/*administration & dosage ; Disease Models, Animal ; Electrons ; Liver/pathology/ultrastructure ; Male ; Mesocricetus ; Microscopy, Electron, Scanning ; Microscopy, Electron, Transmission ; Opisthorchiasis/parasitology/*pathology/*prevention & control ; Opisthorchis/*growth & development

OBJECTIVETo compare morphological differences of three drug-resistant hepatocellular carcinoma (HCC) cell subclones (Huh-7/ADM, Huh-7/CBP, Huh-7/MMC) and their parental Huh-7 cell line, to analyze differential microRNA (miRNA) expression profiles in these cells and, finally to screen for the abnormal expressed miRNAs in drug-resistant HCC cells.

METHODSCellular morphology was observed by histology and transmission electron microscopy. MiRNA microarray was used to analyze the differential miRNA expression profiles in these cells (Huh-7, Huh-7/ADM, Huh-7/CBP, Huh-7/MMC) followed by real time quantitative PCR validation.

RESULTSThe drug-resistant cells had more intracytoplasmic organelles and were larger in size along with increased cytological pleomorphism than the parental Huh-7 cells. Compared with the parental Huh-7 cells, 32 simultaneously up-regulated and 22 down-regulated miRNAs were found in three drug-resistant cells. Up-regulation of miR-15a, miR-16, miR-27b, miR-30b, miR-146a, miR-146b-5p, miR-181a, miR-181d and miR-194 was verified by RT-qPCR.

CONCLUSIONDrug-resistant HCC cells have abnormal expressed miRNAs, which may be explored to further investigate the association of miRNA expressions with multidrugs resistance in HCC.

Antineoplastic Agents ; pharmacology ; Carboplatin ; pharmacology ; Carcinoma, Hepatocellular ; genetics ; pathology ; ultrastructure ; Cell Line, Tumor ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Gene Expression Profiling ; Humans ; Liver Neoplasms ; genetics ; pathology ; ultrastructure ; MicroRNAs ; genetics ; metabolism ; Mitomycin ; pharmacology ; Oligonucleotide Array Sequence Analysis

OBJECTIVETo explore the pathological characteristics of inborn hyperbilirubinemia of patients with Gilbert's syndrome (GS).

METHODSPatients with GS (n = 7) and patients with chronic hepatitis B (CHB; n = 8) were enrolled in the study. GS was diagnosed by peripheral blood analysis results showing glucuronyl transferase gene mutation. The histology and ultrastructure of biopsied liver tissues were evaluated by light microscopy and transmission electron microscopy.

RESULTSThe GS group showed normal structure in the hepatic portal area and lobule; however, bile pigment granules with high electron density were noted in the hepatocytes. The CHB group showed abnormal structure of the hepatic lobules, including infiltration of inflammatory cells, necrotic regions, degenerated hepatocytes, bile duct injury, and fibrosis in the portal tracts; a few bile pigment granules were observed. The GS group also showed greater quantity and size of bilirubin deposits than the CHB group.

CONCLUSIONThe histological and ultrastructural features of GS include normal hepatic lobule and deposition of bile pigment granules in hepatocytes.

Adolescent ; Adult ; Child ; Female ; Gilbert Disease ; pathology ; Hepatitis B, Chronic ; pathology ; Hepatocytes ; ultrastructure ; Humans ; Liver ; cytology ; pathology ; Male ; Young Adult