Yu-Dong Qiu

Identification and Clinical Significance of Mobilized Endothelial Progenitor Cells in Tumor Vasculogenesis of Hepatocellular Carcinoma

Purpose: To investigate the distribution, frequency, and clinical significance of mobilized endothelial progenitor cells (EPC) in hepatocellular carcinoma (HCC). Experimental Design: In healthy controls and patients with HCC, the frequency of circulating EPCs was determined by colony-forming assays, fluorescence-activated cell sorting, and real-time PCR. One hundred sixty-five–amino acid form of vascular endothelial growth factor and platelet-derived growth factor-BB in plasma and tissue were quantified by ELISA. The distribution and frequency of EPCs were evaluated by immunofluorescence, immunohistochemistry, and real-time PCR in normal liver (n = 8), and tumor tissue (TT), adjacent nonmalignant liver tissue (AT), and tumor-free tissue 5 cm from the tumor edge (TF) from 64 patients with HCC. Clinicopathologic data for these patients were evaluated. Results: Compared with values for healthy controls, colony-forming unit scores were higher in the peripheral blood of patients with HCC. Plasma 165-amino acid form of vascular endothelial growth factor and platelet-derived growth factor-BB correlated with the expression level of the AC133 gene, which was also higher in the peripheral blood of patients with HCC. Immunohistochemical analysis showed that EPCs were incorporated into the microvessels in cirrhotic and tumor tissue. Compared with normal liver (9.00), increased AC133+ microvessel density (microvessels/0.74 mm2) was found in TT (53.56), AT (84.76), and TF (48.33). The levels of AC133 gene expression and AC133-microvessel density in AT, which were the highest among four groups, correlated with clinicopathologic variables (the absence of tumor capsule, venous invasion, proliferating cell nuclear antigen intensity, and early recurrence). Conclusions: Mobilized EPCs participate in tumor vasculogenesis of HCC. AC133 gene or antigen in peripheral blood and liver tissue could be used as a biomarker for predicting the progression of HCC.

Differentiation of embryoid-body cells derived from embryonic stem cells into hepatocytes in alginate microbeads in vitro

AbstractAim:Embryonic stem (ES) cells are being widely investigated as a promising source of hepatocytes with their proliferative, renewable, and pluripotent capacities. However, controlled and scalable ES cell differentiation culture into functional hepatocytes is challenging. In this study, we examined the differentiating potential of embryoid-body cells derived from ES cells into hepatocytes in alginate microbeads containing exogenous growth factors in vitro.Methods:Embryoid bodies were formed from ES cells by suspension methods. Embryoid bodies cultured for 5 d were treated with trypsin-EDTA. The disaggregated cells were encapsulated in alginate microbeads and stimulated with exogenous growth factors to induce hepatic differentiation. In the course of cell differentiation, cell morphology and viability were observed, and the expression patterns of some genes of the hepatocyte were confirmed by RT-PCR. An immunofluorescence analysis revealed the expression of albumin (ALB) and cytokeratin-18 (CK18). Hepatocyte functional assays were confirmed by the secretion of ALB and urea.Results:We showed that embryoid-body cells could maintain cell viability in alginate microbeads in vitro. We also found that directed differentiated cells expressed several hepatocyte genes including α-fetoprotein (AFP), ALB, Cyp7a1, CK18, transthyretin (TTR) and tyrosine aminotransferase (TAT) and produced ALB and urea in alginate microbeads. The directed differentiated cells expressed ALB and CK18 proteins on d 14. However, embryoid-body cells could not form hepatocytes without exogenous growth factors in alginate microbeads.Conclusion:The differentiation of embryoid-body cells into hepatocytes containing exogenous growth factors in alginate microbeads gives rise to functional hepatocytes and may develop scalable stem cell differentiation strategies for bioartificial livers and hepatocyte transplantation.

Effects of ω-3 Fish Oil Lipid Emulsion Combined With Parenteral Nutrition on Patients Undergoing Liver Transplantation

BACKGROUND The effect of parenteral nutrition (PN) support supplemented with ω-3 fatty acids was investigated in a randomized, controlled clinical trial at the Affiliated Drum Tower Hospital, Medical School of Nanjing University. MATERIALS AND METHODS Ninety-eight patients with the diagnosis of end-stage liver disease or hepatic cellular carcinoma were admitted for orthotopic liver transplantation at the Affiliated Drum Tower Hospital. The patients were randomly divided into 3 groups: diet group (n = 32), PN group (n = 33), and polyunsaturated fatty acid (PUFA) group (n = 33). Patients in the PN and PUFA groups received isocaloric and isonitrogenous PN for 7 days after surgery. Venous heparin blood samples were obtained for assay on days 2 and 9 after surgery. A pathological test was performed after reperfusion of the donor liver and on day 9. RESULTS Alanine aminotransferase levels were improved significantly by PUFA treatment compared with traditional PN support (P < .05). Compared with the results on day 9 in the PN group, a significant difference was seen in the extent of increase of the prognostic nutrition index and prealbumin in the PUFA group. The pathological results also showed that ω-3 fatty acid supplementation reduced hepatic cell injury. PUFA therapy also decreased the incidence of infectious morbidities and shortened the posttransplant hospital stay significantly. CONCLUSION Posttransplant PN support can greatly improve metabolism of protein and nutrition states of patients. ω-3 fatty acid-supplemented PN significantly reduces injury of the transplanted liver, decreases the incidence of infectious morbidities, and shortens posttransplant hospital stay.

Liver-protecting effects of omega-3 fish oil lipid emulsion in liver transplantation

AIM To investigate the liver-protecting effect of parenteral nutrition (PN) support with omega-3 fatty acids in a randomized controlled clinical trial. METHODS Sixty-six patients with the diagnosis of end-stage liver disease or hepatic cellular carcinoma were admitted to the Affiliated Drum Tower Hospital, Nanjing University, China for orthotopic liver transplantation. The patients were randomly divided into two groups: PN group (n = 33) and polyunsaturated fatty acid (PUFA) group (n = 33). All patients received isocaloric and isonitrogenous PN for seven days after surgery, and in PUFA group omega-3 fish oil lipid emulsion replaced part of the standard lipid emulsion. Liver function was tested on days 2 and 9 after surgery. Pathological examination was performed after reperfusion of the donor liver and on day 9. Clinical outcome was assessed based on the post-transplant investigations, including: (1) post-transplant mechanical ventilation; (2) total hospital stay; (3) infectious morbidities; (4) acute and chronic rejection; and (5) mortality (intensive care unit mortality, hospital mortality, 28-d mortality, and survival at a one-year post-transplant surveillance period). RESULTS On days 2 and 9 after operation, a significant decrease of alanine aminotransferase (299.16 U/L ± 189.17 U/L vs 246.16 U/L ± 175.21 U/L, P = 0.024) and prothrombin time (5.64 s ± 2.06 s vs 2.54 s ± 1.15 s, P = 0.035) was seen in PUFA group compared with PN group. The pathological results showed that omega-3 fatty acid supplement improved the injury of hepatic cells. Compared with PN group, there was a significant decrease of post-transplant hospital stay in PUFA group (18.7 d ± 4.0 d vs 20.6 d ± 4.6 d, P = 0.041). Complications of infection occurred in 6 cases of PN group (2 cases of pneumonia, 3 cases of intra-abdominal abscess and 1 case of urinary tract infection), and in 3 cases of PUFA group (2 cases of pneumonia and 1 case of intra-abdominal abscess). No acute or chronic rejection and hospital mortality were found in both groups. The one-year mortality in PN group was 9.1% (3/33), one died of pulmonary infection, one died of severe intra-hepatic cholangitis and hepatic dysfunction and the other died of hepatic cell carcinoma recurrence. Only one patient in PUFA group (1/33, 3.1%) died of biliary complication and hepatic dysfunction during follow-up. CONCLUSION Post-transplant parenteral nutritional support combined with omega-3 fatty acids can significantly improve the liver injury, reduce the infectious morbidities, and shorten the post-transplant hospital stay.

Effect of early enteral combined with parenteral nutrition in patients undergoing pancreaticoduodenectomy

AIM To investigate the effect of early enteral nutrition (EEN) combined with parenteral nutritional support in patients undergoing pancreaticoduodenectomy (PD). METHODS From January 2006, all patients were given EEN combined with parenteral nutrition (PN) (EEN/PN group, n = 107), while patients prior to this date were given total parenteral nutrition (TPN) (TPN group, n = 67). Venous blood samples were obtained for a nutrition-associated assessment and liver function tests on the day before surgery and 6 d after surgery. The assessment of clinical outcome was based on postoperative complications. Follow-up for infectious and noninfectious complications was carried out for 30 d after hospital discharge. Readmission within 30 d after discharge was also recorded. RESULTS Compared with the TPN group, a significant decrease in prealbumin (PAB) (P = 0.023) was seen in the EEN/PN group. Total bilirubin (TB), direct bilirubin (DB) and lactate dehydrogenase (LDH) were significantly decreased on day 6 in the EEN/PN group (P = 0.006, 0.004 and 0.032, respectively). The rate of grade I complications, grade II complications and the length of postoperative hospital stay in the EEN/PN group were significantly decreased (P = 0.036, 0.028 and 0.021, respectively), and no hospital mortality was observed in our study. Compared with the TPN group (58.2%), the rate of infectious complications in the EEN/PN group (39.3%) was significantly decreased (P = 0.042). Eleven cases of delayed gastric emptying were noted in the TPN group, and 6 cases in the EEN/PN group. The rate of delayed gastric emptying and hyperglycemia was significantly reduced in the EEN/PN group (P = 0.031 and P = 0.040, respectively). CONCLUSION Early enteral combined with PN can greatly improve liver function, reduce infectious complications and delayed gastric emptying, and shorten postoperative hospital stay in patients undergoing PD.

Hepatocyte‐like cells from directed differentiation of mouse bone marrow cells in vitro

AbstractAim:To design the effective directed differentiation medium to differentiate bone marrow cells into hepatocyte-like cells.Methods:Bone marrow cells were cultured in the directed differentiation media including fibroblast growth factor-4 (FGF-4) and oncostatin M (OSM). Hepatocyte-like cells from directed differentiation of bone marrow cells were identified through cell morphology, RNA expressions by reverse transcriptase-polymerase chain reaction (RT-PCR), protein expressions by Western blot, and hepatocellular synthesis and metabolism functions by albumin ELISA, Periodic acid-Shiff staining and urea assay.Results:Some epithelial-like cells or polygonal cells appeared and increased in the course of the cell directed differentiation. Hepatocyte nucleur factor-3β (HNF-3β), albumin (ALB), cytokeratin 18 (CK18), transthyretin (TTR), glucose-6-phosphate (G-6-Pase), and tyrosine aminotransferase (TAT) mRNA were expressed in the course of the directed differentiation. The directed differentiated cells on d 21 expressed HNF-3β, ALB, and CK18 proteins. The directed differentiated cells produced albumin and synthesized urea in a time-dependent manner. They could also synthesize glycogen.Conclusion:Our differentiation media, including FGF-4 and OSM, are effective to differentiate bone marrow cells into hepatocyte-like cells, which could be used for hepatocyte resources for bioartificial liver or hepatocyte transplantation.

Comparative Analysis of the Efficacy and Complications of Nasojejunal and Jejunostomy on Patients Undergoing Pancreaticoduodenectomy

BACKGROUND The efficacy and feeding-related complications of a nasojejunal feeding tube and jejunostomy after pancreaticoduodenectomy (PD) was investigated with a randomized, controlled clinical trial at the Affiliated Drum Tower Hospital. METHODS Sixty-eight patients who underwent PD in the Department of Hepatobiliary Surgery were randomly divided into 2 groups: 34 patients received enteral feeding via a nasojejunal tube (NJT group) and 34 patients received enteral feeding via a jejunostomy tube (JT group). The assessment of clinical outcome was based on postoperative investigation of complications. The second part of the assessment included tube related complications and an index on catheter efficiency. RESULTS There were 15 cases with infectious complications in the JT group and 13 cases in the NJT group, and there was no significant difference in the rate of infectious complications between the 2 groups. The rate of intestinal obstruction and delayed gastric emptying was significantly decreased in the NJT group (P < .05). Catheter-related complications were more common in the JT group as compared with the NJT group (35.3% vs 20.6%, P < .05). The time for removal of the feeding tube and nasogastric tube was significantly decreased in the NJT group. The postoperative hospital stay in the NJT group was significantly decreased (P < .05), and there was no hospital mortality in this study. CONCLUSION Nasojejunal feeding is safer than jejunostomy, and it is associated with only minor complications. Nasojejunal feeding can significantly decrease the incidence of delayed gastric emptying and shorten the postoperative hospital stay.

Efficacy and safety of liver transplantation in patients with cholangiocarcinoma: A systematic review and meta‐analysis

The aim of our study was to evaluate the efficacy and safety of liver transplantation in patients with cholangiocarcinoma. According to the requirements of Cochrane systematic review, a thorough literature search was performed in PubMed/Medline, Embase and Cochrane electronic databases between 1995 and 2009 in terms of the key words “liver transplantation” and “cholangiocarcinoma,” “cholangiocellular carcinoma” or “bile duct cancer,” with restricted articles for the English language. Data were processed for a meta‐analysis by Stata 10 software. Altogether 14 clinical trials containing 605 transplanted patients of bile duct cancers were finally enrolled in our study. The overall 1‐, 3‐ and 5‐year pooled survival rates were 0.73 [95% confidence interval (CI) = 0.65–0.80], 0.42 (95% CI = 0.33–0.51) and 0.39 (95% CI = 0.28–0.51), respectively. Of note, preoperative adjuvant therapies [orthotopic liver transplantation (OLT)‐PAT group] rendered the transplanted individuals with comparably favorable outcomes with 1‐, 3‐ and 5‐year pooled survival rates of 0.83 (95% CI = 0.57–0.98), 0.57 (95% CI = 0.18–0.92) and 0.65 (95% CI = 0.40–0.87). In addition, the overall pooled incidence of complications was 0.62 (95% CI = 0.44–0.78), among which that of OLT‐PAT group (0.58; 95% CI = 0.20–0.92) was relatively acceptable compared to those of liver transplantation alone (0.61; 95% CI = 0.33–0.85) and liver transplantation with extended bile duct resection (0.78; 95% CI = 0.55–0.94). In comparison to curative resection of cholangiocarcinoma with the 5‐year survival rate reported from 20 to 40%, the role of liver transplantation alone is so limited. In the future, attention will be focused on liver transplantation following neoadjuvant radiochemotherapy, which requires a well‐designed, prospective randomized controlled study.

Caveolin-1 is important for nitric oxide-mediated angiogenesis in fibrin gels with human umbilical vein endothelial cells

AbstractAim:The role of caveolin-1 (Cav-1) in angiogenesis remains poorly understood. The endothelial nitric oxide (NO) synthase (eNOS), a caveolin-interacting protein, was demonstrated to play a predominant role in vascular endothelial growth factor (VEGF) -induced angiogenesis. The purpose of our study was to examine the role of Cav-1 and the eNOS complex in NO-mediated angiogenesis.Methods:Human umbilical vein endothelial cells (HUVEC) were isolated and cultured in 3-D fibrin gels to form capillary-like tubules by VEGF stimulation. The expression of Cav-1 and eNOS was detected by semiquantitative RT-PCR. The HUVEC were treated with antisense oligonucleotides to downregulate Cav-1 expression. Both transduced and non-infected HUVEC were cultured in fibrin gels in the presence or absence of VEGF (20 ng/mL) and NG-nitro-L-arginine methyl ester (L-NAME; 5 mmol/L). NO was measured using a NO assay kit and capillary-like tubules were quantified by tubule formation index using the Image J program.Results:RT-PCR analysis revealed that Cav-1 levels steadily increased in a time-dependent manner and reached their maximum after 5 d of incubation, but there were no obvious changes in eNOS mRNA expression in response to VEGF in the fibrin gel model. VEGF (20 ng/mL) can promote NO production and the formation of capillary-like tubules, and this promoting effect of VEGF was blocked by the addition of L-NAME (5 mmol/L). When transduced HUVEC with the antisense Cav-1 oligonucleotides were plated in the fibrin gels, the capillary-like tubules were significantly fewer than those of the non-infected cells. The capillary-like tubules formation and NO production of transduced HUVEC with the antisense Cav-1 oligonucleotides cultured in fibrin gels showed no responses to the addition of VEGF (20 ng/mL) and L-NAME (5.0 mmol/L).Conclusion:NO was a critical angiogenic mediator in this model. Cav-1 was essential for NO-mediated angiogenesis and may be an important target of anti-angiogenesis therapy.

Effect of Parenteral Fish Oil Lipid Emulsion in Parenteral Nutrition Supplementation Combined With Enteral Nutrition Support in Patients Undergoing Pancreaticoduodenectomy

BACKGROUND The effect of parenteral fish oil lipid emulsion in parenteral nutrition (PN) supplementation combined with enteral nutrition (EN) support on pancreaticoduodenectomy (PD) was investigated with a randomized controlled clinical trial at the Affiliated Drum Tower Hospital. MATERIALS AND METHODS Seventy-six patients who underwent PD in the Department of Hepatobiliary Surgery were randomly divided into 2 groups: the polyunsaturated fatty acid (PUFA) group (n = 38) and control group (n = 38). Patients in the PUFA group received parenteral fish oil lipid emulsion supplementation combined with early EN support for 5 days after PD. Venous blood samples were obtained for assay on the day before surgery and on day 6 after surgery. RESULTS Compared with the results of the control group, a significant difference was seen in the extent of the decrease in total protein and prealbumin in the PUFA group (P < .05). Alanine aminotransferase (ALT), aspartate aminotransferase (AST), and lactate dehydrogenase (LDH) were significantly decreased on day 6 in the PUFA group (P < .01), and a significant difference was seen in the extent of decrease in ALT, AST, and LDH in the PUFA group (P < .05). The ratio of infectious complications in the PUFA group was significantly decreased, as well as the postoperative hospital stay (P < .05), and there was no hospital mortality in this study. CONCLUSION Parenteral fish oil lipid emulsion in PN supplementation combined with EN support can greatly improve the nutrition state and liver function of patients, decrease the incidence of infectious morbidities, and shorten the postoperative hospital stay.