Calcium ; metabolism ; Hepatocytes ; metabolism ; Homeostasis ; Humans

Animals ; Hepatocytes ; metabolism ; Humans ; Lipoproteins ; metabolism ; Protein Binding ; Proteins ; metabolism

Hepatocytes ; metabolism ; pathology ; Liver Diseases ; genetics ; metabolism ; pathology

Carcinoma, Hepatocellular ; metabolism ; pathology ; Hepatocytes ; metabolism ; pathology ; Humans ; Liver Neoplasms ; metabolism ; pathology ; NF-kappa B ; metabolism

Aquaporin-9 (AQP9) is a membrane-span transport protein expressed in the liver. It is located in the cytoplasm membrane of hepatic cells. In addition to water, it is also permeable to glycerol, urea, and other small solutes. Several evidences have revealed that AQP9 is involved in multiple physiological and pathological process of the liver. This paper summarized the expression of AQP9 in the liver and the effect on the physiological and pathological processes of the liver. AQP9 may be defined as a novel target for diagnosis and treatment of hepatic diseases.
Aquaporins ; metabolism ; Cell Membrane ; metabolism ; Glycerol ; Hepatocytes ; metabolism ; Humans ; Liver ; metabolism ; Urea

Primary human hepatocytes (PHH) are the gold standard of in vitro human liver model for drug screening. However, a problem of culturing PHH in vitro is the rapid decline of cytochrome P450 (CYP450) activity, which plays an important role in drug metabolism. In this study, thermo-responsive culture dishes were used to explore the conditions for murine embryonic 3T3-J2 fibroblasts to form cell sheet. Based on the cell sheet engineering technology, a three-dimensional (3D) "sandwich" co-culture system of 3T3-J2 cell sheet/PHH/collagen gel was constructed. The tissue structure and protein expression of the model section were observed by hematoxylin eosin staining and immunofluorescence staining respectively. Phenacetin and bupropion were used as substrates to determine the activity of CYP450. The contents of albumin and urea in the system were determined by enzyme linked immunosorbent assay (ELISA). The results showed that the complete 3T3-J2 cell sheet could be obtained when the cell seeding density was 1.5×10⁶ /dish (35 mm dish) and the incubation time at low temperature was 60 min. Through cell sheet stacking, a 3D in vitro liver model was developed. Compared with the two-dimensional (2D) model, in the 3D model, the cell-cell and cell-matrix connections were tighter, the activities of cytochrome P450 CYP1A2 and cytochrome P450 CYP2B6 were significantly increased, and the secretion levels of albumin and urea were increased. These indexes could be maintained stably for 21 d. Therefore, cell sheet stacking is helpful to improve the level of liver function of 3D liver model. This model is expected to be used to predict the metabolism of low-clearance drugs in preclinical, which is of great significance for drug evaluation and other studies.
Albumins/metabolism* ; Animals ; Cytochrome P-450 Enzyme System/metabolism* ; Hepatocytes/metabolism* ; Humans ; Liver ; Mice ; Urea/metabolism*

OBJECTIVETo investigate the effect of N,N-dimethylformamide (DMF) on calcium homeostasis and calpain I and II gene expression in human hepatocytes (HL-7702).

METHODSHL-7702 cells were exposed to different concentrations of DMF (10, 25, 50, 100, or 200 mmol/L); other HL-7702 cells, which were used as a control group, were exposed to the equal volume of DMEM; the intracellular Ca(2+) concentration was monitored using the calcium fluorescent probe (fluo-3/AM). After 24-h exposure to DMF (10, 25, 50, 100, 150, or 200 mmol/L), the morphology of hepatocytes was observed under an inverted phase contrast microscope, and the cell viability was measured by MTT assay. After 24-h exposure to DMF (10, 25, 50, 100, or 150 mmol/L), the mRNA expression levels of calpain I and II in hepatocytes were measured by real-time quantitative PCR.

RESULTSThere were significant differences in cell viability among different exposure groups (P < 0.01); the 50, 100, 150, and 200 mmol/L DMF exposure groups had a significantly lower cell viability than the control group (P < 0.05). Under the inverted phase contrast microscope, HL-7702 cells gradually lost the original shape, with swelling and shrinking, as the dose of DMF increased, and those treated with 150 mmol/L DMF even became round and floated. The fluorescence density of fluo-3 in hepatocytes increased as the dose of DMF rose, demonstrating a dose-response relationship, and there were significant differences among these exposure groups (P < 0.05). There were significant differences in mRNA expression levels of calpain I and II among these exposure groups (P < 0.01), and the expression increased as the dose of DMF rose; but DMF did not promote the mRNA expression of calpain I at a concentration of 150 mmol/L.

CONCLUSIONDMF can cause damage to hepatocytes, which is related to intracellular calcium increase and calpain mRNA increase.

Calcium ; metabolism ; Calpain ; metabolism ; Cell Line ; Dimethylformamide ; pharmacology ; Hepatocytes ; drug effects ; metabolism ; Humans

Acetaminophen ; metabolism ; Animals ; Cells, Cultured ; Drug Interactions ; Flavonoids ; metabolism ; Hepatocytes ; metabolism ; Humans

OBJECTIVETo determine the effect of intermittent hypoxia on lipid metabolism in liver cells and to explore the possible molecular pathways involved in this process.

METHODSAn intermittent hypoxia cell model system was established by incubating the human hepatic cell lines L02 and HepG2 in an atmosphere of 2% O₂, 5% CO₂ and 93% N₂ for 8 hours per day over a period of 1, 2, 3, 4 or 5 days. Cells cultured in normoxia conditions (21% O₂) served as controls. Changes in intracellular lipid droplets and triglyceride (TG) levels were assessed by biochemical assays and oil red staining. Changes in intracellular reactive oxygen species (ROS) were assessed by inverted fluorescence microscopy and flow cytometry. Changes in expression of hypoxia-inducible factor (HIF)-1a and HIF-2a proteins, and the downstream ADFP, SREBP-1c and FAS proteins, were detected by western blotting.

RESULTSFor both L02 and HepG2 cell lines, the cells grown under hypoxic conditions showed significantly higher lipid droplet accumulation and TG content than the cells grown under normoxic conditions (F(L02) =61.83, FHepG2 =104.19, P less than 0.01). Both oxygen concentration and time appeared to be correlated with these lipid-related changes (F(L02) =39.60, FHepG2 =76.39, P less than 0.01). The ROS fluorescence index was significantly increased after 2 days of intermittent hypoxia L02: 0.703 ± 0.129 vs. 3.310 ± 0.198, t =22.0637 and HepG2:0.617 ± 0.156 vs. 2.33 ± 0.42, t =7.2003, P < 0.05); in addition, increasing trends were observed in the ROS content and intensity of green fluorescence in conjunction with increased time of exposure to intermittent hypoxia (F(L02) =1021.84, FHepG2 =49.89, P less than 0.01). Compared with their respective control groups, the L02 and HepG2 cells both showed significantly upregulated expression of HIF-1a ADFP, SREBP-1c and FAS (L02:FHIF-1a =371.19, FsREBP-1c =204.49, FFAS =38.20, FADFP =154.31, P less than 0.05 and HepG2:FHF-1a =150.84, FSRERBP-1c =107.35, FFAs =279.71, FADFP =352.06, P less than 0.01). In contrast, the HIF-2a level was markedly decreased in the cells after 4 and 5 days of exposure to intermittent hypoxia (F(L02) =125.29, FHcpG2 =10.68, P less than 0.05).

CONCLUSIONUnder intermittent hypoxic conditions, ROS may regulate the expression of hypoxia-inducible factors and the adipose differentiation-related protein,as well as influence fatty acid metabolism via a HIF-1 a-SREBP-1 c-FAS signal and upregulation of the ADFP protein, in liver cells.

Cell Hypoxia ; Cell Line ; Hep G2 Cells ; Hepatocytes ; metabolism ; Humans ; Lipid Metabolism ; Reactive Oxygen Species ; metabolism

OBJECTIVETo investigate the characteristics of the intra- and extra-hepatocyte sodium ions distribution in scalded rats during early postburn stage,with the aim of improving burn shock resuscitation regime and the resuscitation effects.

METHODSAdult Sprague-Dawley rats were randomly divided into sham scalding (C, n = 12) and scalding (S, n = 7) groups. The rats in S group were subjected to 40% TBSA III degree scalding on the back and were catheterized via jugular vein for fluid resuscitation. The rats in C group were catheterized via jugular vein without fluid infusion and were sham scalded by warm water in temperature of 37 degrees. The changes in the intra- and extra-hepatocyte sodium ion contents were determined in vivo by (23)Na-magnetic resonance spectrum technology, while the existing state of the intra- and extra-hepatocyte sodium ion was determined by detecting (23)Na-magnetic resonance horizontal delaying time (T(2)).

RESULTSThe extra-hepatocyte sodium content in S group at 24 postburn hours (PBHs) was 17% less than that in C group. In addition, the T(2f) (fast T(2)) in S group remained stable but maintained a higher ratio during the observation time. This suggested that the sodium binding sites in extra-hepatocyte matrix increased relatively and that intra-hepatocyte sodium content increased by 57%. But the T(2) and the fast and slow parts of the T(2) kept stable, which implied that intra-hepatocyte catabolizing products were increased. This led to an increase in the sodium ion binding sites within intra-hepatocyte matrix in proportion to the sodium ion content.

CONCLUSIONDuring early postburn stage, the extra-hepatocyte sodium in a remote organ such as the liver exhibited relative deficiency due to its ingress into hepatocyte cytoplasm and to the increase of sodium combining sites.

Animals ; Binding Sites ; Burns ; metabolism ; Hepatocytes ; metabolism ; Magnetic Resonance Spectroscopy ; Rats ; Rats, Sprague-Dawley ; Sodium ; metabolism