ABSTRACT
Objective To design a new type of hybrid bioartificial liver (HBAL), evaluate its efficacy in vitro, and explore the feasibility in clinical application.Methods CL-1 human hepatocytes were cultured on microcarriers for 5 days, when cell count reached about 4.0 × 109 with cell density of about 4.0 × 107/ml.CL-1 cells cultured on microcarriers in home-made bioreactor constitute the biological part of the HBAL.The abiotic part included blood perfusion and bilirubin adsorption, and blood pump was employed as the circulation driver, which were parts of HBAL.The changes of the concentrations of indirect bilirubin (UBD), chenodeoxycholic acid (CDCD), cholic acid (CA), blood ammonia (AA), AST, ALT and LDH were observed under the condition of in vitro circulation.Meanwhile, the function, morphology and the cell activity of CL-1 cells were also observed.Results After in vitro circulation for 24 h, the concentrations of UBD, CDCD, CA and AA significantly decreased from (335.3 ± 6.0) μmol/L, (395.0 ± 5.6) μmol/L, (155.7 ± 4.5) μmol/L, (39.0 ± 2.6) μmol/L at 0 h to (106.0 ± 10.9) μmol/L, (131.8 ± 28.7) μmol/L, (42.2 ± 7.3) μmol/L, (3.5 ± 1.0) μmol/L, respectively.At 48 h, ALT, AST and LDH significantly increased from (25.9 ± 4.2) IU/L, (22.0 ± 3.6) IU/L, (0.28 ± 0.09) μmol/L to (31.0 ± 2.6) IU/L,(31.6 ± 8.0) IU/L, (0.41 ± 0.12) μmol/L, meanwhile the count and vitality of CL-1 cells were significant declined.Conclusions (1) In the new HBAL system, CL-1 cells can keep its viability and function in vitro;and (2) the HBAL appears to be effective in purifying the serum in liver failure simulation model by clearing out non-conjugated bilirubin, chenodeoxycholic acid, cholic acid and ammonium chloride, which seems to be a promising therapeutic option.
ABSTRACT
<p><b>OBJECTIVE</b>To explore whether the function of the CL-1 hepatocytes, co-cultured with human hepatic stellate cells (HSC) on microcarriers was better than that cultured without HSC.</p><p><b>METHODS</b>CL-1 hepatocytes were divided into 2 groups. The co-culture group was cultured with HSC in DMEM culture medium containing 10% fetal bovine serum, and HSC were not added in single-culture group. The cytomorphology was observed by inverted microscope and scanning electron microscopy. The effects of different culture method on the proliferation in vitro were analyzed by MTT assay. The function of hepatocytes was evaluated through measuring the concentration of ALT and albumin.</p><p><b>RESULTS</b>The inverted microscope and the MTT staining results showed that the quantity and viability of the human hepatocyte (C3A) in bidirectional bioreactor group were much better than the RCMW group. The growth curve results showed that the density of the human hepatocyte (C3A) was firstly increased and then decreased in both groups, and the peak of the curve appeared in day 5. The density of human liver cells in the second generation of bi-directional rotation and perfusion microgravity bioreactor group was significantly higher than the RCMW group from day1 to day 7 (P<0.05). The functional results showed that the albumin and urea concentration, which reached the peak on day 5, also gone up firstly and then gradually gone down in both teams. And the albumin and urea concentration in bidirectional bioreactor group was significantly higher than RCMW group from day 1 to day7 (P<0.01). Besides, the concentration of ALT and AST in bidirectional bioreactor group was significantly lower than RCMW group from day 1 to day 7 (P<0.05).</p><p><b>CONCLUSION</b>This study shows that this new culture method is advantageous in enhancing cell viability and function. It indicates that co-culturing hepatocytes with HSC has a good application prospect in the development of artificial liver technology.</p>
Subject(s)
Humans , Bioreactors , Cells, Cultured , Coculture Techniques , Hepatic Stellate Cells , Cell Biology , Hepatocytes , Cell BiologyABSTRACT
Hepatocellular carcinoma (HCC), among the most common malignancies worldwide, remains a major threat to public health, and there is an urgent need to identify novel biomarkers for diagnosis, prognosis and targets for anti-cancer treatment. In this study, two-dimensional polyacrylamide gel electrophoresis coupled with ESI-Q-TOF MS/MS analysis was used to identify differentially expressed proteins among the HCC tumour centre, tumour margin and nontumourous liver tissues. In total, 52 spots with significant alteration were positively identified byMS/MSanalysis. Altered expression of representative proteins, including CIB1, was validated by Western blotting. Immunostaining suggested an increase tendency of CIB1 expression from nontumourous liver tissue to tumour centre. Knockdown of CIB1 expression by RNA interference led to the significant suppression of the cell growth in hepatoma HepG2 cells. These data suggest that CIB1 may be used as a novel prognostic factor and possibly an attractive therapeutic target for HCC.