ABSTRACT
Rats have long been an ideal model for disease research in the field of biomedicine, but the bottleneck of in vitro culture of rat embryonic stem (ES) cells hindered the wide application as genetic disease models. Here, we optimized a special medium which we named 5N-medium for rat embryonic stem cells, which improved the in vitro cells with better morphology and higher pluripotency. We then established a drug selection schedule harboring a prior selection of 12 h that achieved a higher positive selection ratio. These treatments induced at least 50% increase of homologous recombination efficiency compared with conventional 2i culture condition. Moreover, the ratio of euploid ES clones also increased by 50% with a higher germline transmission rate. Finally, we successfully knocked in a 175 kb human Bacterial Artificial Chromosome (BAC) fragment to rat ES genome through recombinase mediated cassette exchange (RMCE). Hence, we provide a promising system for generating sophisticated rat models which could be benefit for biomedical researches.
Subject(s)
Embryonic Stem Cells/cytology , Animals , Cell Proliferation , Cells, Cultured , Models, Animal , Rats , Rats, Inbred F344 , Rats, Sprague-DawleyABSTRACT
Post-hepatectomy liver dysfunction is a life-threatening morbidity that lacks efficient therapy. Bioactive lipids involved in macrophage polarization crucially regulate tissue injury and regeneration. Herein, we investigate the key bioactive lipids that mediate the cytotherapeutic potential of polarized-macrophage for post-hepatectomy liver dysfunction. Untargeted lipidomics identified elevation of ceramide (CER) metabolites as signature lipid species relevant to M1/M2 polarization in mouse bone-marrow-derived-macrophages (BMDMs). M1 BMDMs expressed a CER-generation-metabolic pattern, leading to elevation of CER; M2 BMDMs expressed a CER-breakdown-metabolic pattern, resulting in upregulation of sphingosine-1-phosphate (S1P). After infusing M1- or M2-polarized BMDMs into the mouse liver after hepatectomy, we found that M1-BMDM infusion increased M1 polarization and CER accumulation, resulting in exaggeration of hepatocyte apoptosis and liver dysfunction. Conversely, M2-BMDM infusion enhanced M2 polarization and S1P generation, leading to alleviation of liver dysfunction with improved hepatocyte proliferation. Treatment of exogenous CER and S1P or inhibition CER and S1P synthesis by siRNA targeting relevant enzymes further revealed that CER induced apoptosis while S1P promoted proliferation in post-hepatectomy primary hepatocytes. In conclusion, CER and S1P are uncovered as critical lipid mediators for M1- and M2-polarized BMDMs to promote injury and regeneration in the liver after hepatectomy, respectively. Notably, the upregulation of hepatic S1P induced by M2-BMDM infusion may have therapeutic potential for post-hepatectomy liver dysfunction.
Subject(s)
Ceramides/metabolism , Hepatectomy/methods , Liver/pathology , Lysophospholipids/metabolism , Metabolomics/methods , Sphingosine/analogs & derivatives , Animals , Disease Models, Animal , Humans , Liver/surgery , Mice , Sphingosine/metabolism , TransfectionABSTRACT
BACKGROUND: Liver hepatocellular carcinoma (HCC) is the third most common cause of death by cancer and has a high mortality world-widely. Approximately 75-85% of primary liver cancers are caused by HCC. Uncovering novel genes with prognostic significance would shed light on improving the HCC patient's outcome. OBJECTIVE: In this research, we aim to identify novel prognostic biomarkers in hepatocellular carcinoma. METHODS: Integrated proteomics and bioinformatics analysis were performed to investigate the expression landscape of prognostic biomarkers in 24 paired HCC patients. RESULTS: As a result, eight key genes related to prognosis, including ACADS, HSD17B13, PON3, AMDHD1, CYP2C8, CYP4A11, SLC27A5, CYP2E1, were identified by comparing the weighted gene co-expression network analysis (WGCNA), proteomic differentially expressed genes (DEGs), proteomic turquoise module, The Cancer Genome Atlas (TCGA) cohort DEGs of HCC. Furthermore, we trained and validated eight pivotal genes integrating these independent clinical variables into a nomogram with superior accuracy in predicting progression events, and their lower expression was associated with a higher stage/risk score. The Gene Set Enrichment Analysis (GSEA) further revealed that these key genes showed enrichment in the HCC regulatory pathway. CONCLUSION: All in all, we found that these eight genes might be the novel potential prognostic biomarkers for HCC and also provide promising insights into the pathogenesis of HCC at the molecular level.
ABSTRACT
Long noncoding RNAs have been implicated in many biological processes, but their roles in liver regeneration still need to be illustrated. Therefore, we aimed to investigate the role of LINC00265 as a pivotal regulator of hepatocyte proliferation during liver regeneration. It was found that LINC00265 is significantly upregulated in rat liver tissues at various time points after 2/3 liver resection. LINC00265 knockdown inhibited hepatocyte proliferation, induced cell apoptosis and led to G2/M phase cell cycle arrestment. In rats subjected to surgery, LINC00265 knockdown decreased liver/body weight ratio, attenuated improvement from liver damage and reduced Ki67 and PCNA expression. Luciferase reporter assays confirmed that miR-28-5p was a direct target of LINC00265, and inhibition of miR-28-5p abolished the effect of LINC00265 knockdown. In summary, LINC00265 might maintain hepatocyte proliferation by targeting miR-28-5p during liver regeneration and should be considered as a promising therapeutic option for hepatocyte regeneration after liver resection.
Subject(s)
Cell Proliferation/physiology , Hepatocytes/cytology , Liver Regeneration , MicroRNAs/physiology , RNA, Long Noncoding/metabolism , Apoptosis/genetics , Cell Line , Gene Knockdown Techniques , Hepatocytes/metabolism , Humans , Ki-67 Antigen/metabolism , Proliferating Cell Nuclear Antigen/metabolism , RNA, Long Noncoding/geneticsABSTRACT
Hepatocellular carcinoma (HCC) has been extensively studied as one of the most aggressive tumors worldwide. However, its mortality rate remains high due to ideal diagnosis and treatment strategies. Uncovering novel genes with prognostic significance would shed light on improving the HCC patient's outcome. In our study, we applied data-independent acquisition (DIA) quantitative proteomics to investigate the expression landscape of 24 paired HCC patients. A total of 1029 differentially expressed proteins (DEPs) were screened. Then, we compared DEPs in our cohort with the differentially expressed genes (DEGs) in The Cancer Genome Atlas, and investigated their prognostic significance, and found 183 prognosis-related genes (PRGs). By conducting protein-protein interaction topological analysis, we identified four subnetworks with prognostic significance. Acyl-CoA oxidase 2 (ACOX2) is a novel gene in subnetwork1, encodes a peroxisomal enzyme, and its function in HCC was investigated in vivo and in vitro. The lower expression of ACOX2 was validated by real-time quantitative PCR, immunohistochemistry, and Western blot. Cell Counting Kit-8 assay, wound healing, and transwell migration assay were applied to evaluate the impact of ACOX2 overexpression on the proliferation and migration abilities in two liver cancer cell lines. ACOX2 overexpression, using a subcutaneous xenograft tumor model, indicated a tumor suppressor role in HCC. To uncover the underlying mechanism, gene set enrichment analysis was conducted, and peroxisome proliferator-activated receptor-α (PPARα) was proposed to be a potential target. In conclusion, we demonstrated a PRG ACOX2, and its overexpression reduced the proliferation and metastasis of liver cancer in vitro and in vivo through PPARα pathway.
Subject(s)
Carcinoma, Hepatocellular/metabolism , Liver Neoplasms/metabolism , Oxidoreductases/physiology , PPAR alpha/metabolism , Animals , Biomarkers, Tumor/physiology , Cell Line, Tumor , Cell Proliferation , Disease Progression , Female , Gene Expression Regulation, Neoplastic , Humans , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Middle AgedABSTRACT
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
ABSTRACT
Overload of palmitic acids is linked to the dysregulation of ceramide metabolism in nonalcoholic steatohepatitis (NASH), and ceramides are important bioactive lipids mediating the lipotoxicity of palmitic acid in NASH. However, much remains unclear about the role of ceramidases that catalyze the hydrolysis of ceramides in NASH. By analyzing the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database, we found that alkaline ceramidase 3 (ACER3) is upregulated in livers of patients with NASH. Consistently, we found that Acer3 mRNA levels and its enzymatic activity were also upregulated in mouse livers with NASH induced by a palmitate-enriched Western diet (PEWD). Moreover, we demonstrated that palmitate treatment also elevated Acer3 mRNA levels and its enzymatic activity in mouse primary hepatocytes. In order to investigate the function of Acer3 in NASH, Acer3 null mice and their wild-type littermates were fed a PEWD to induce NASH. Knocking out Acer3 was found to augment PEWD-induced elevation of C18:1-ceramide and alleviate early inflammation and fibrosis but not steatosis in mouse livers with NASH. In addition, Acer3 deficiency attenuated hepatocyte apoptosis in livers with NASH. These protective effects of Acer3 deficiency were found to be associated with suppression of hepatocellular oxidative stress in NASH liver. In vitro studies further revealed that loss of ACER3/Acer3 increased C18:1-ceramide and inhibited apoptosis and oxidative stress in mouse primary hepatocytes and immortalized human hepatocytes induced by palmitic-acid treatment. These results suggest that ACER3 plays an important pathological role in NASH by mediating palmitic-acid-induced oxidative stress.
Subject(s)
Alkaline Ceramidase/metabolism , Apoptosis/genetics , Non-alcoholic Fatty Liver Disease/enzymology , Oxidative Stress/genetics , Alkaline Ceramidase/deficiency , Alkaline Ceramidase/genetics , Animals , Cell Survival/genetics , Chromatography, Liquid , Diet, Western , Hepatocytes/enzymology , Hepatocytes/metabolism , Humans , Inflammation/diet therapy , Liver/metabolism , Liver/pathology , Liver Cirrhosis/enzymology , Liver Cirrhosis/genetics , Liver Cirrhosis/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Non-alcoholic Fatty Liver Disease/genetics , Non-alcoholic Fatty Liver Disease/metabolism , Palmitic Acid/pharmacology , Tandem Mass Spectrometry , Up-RegulationABSTRACT
BACKGROUND Liver failure after resection for liver cancer is associated with increased patient mortality. This study aimed to investigate the mechanism of the protective effects of resveratrol, a natural plant-derived compound, on liver injury in a rat model of partial hepatectomy. MATERIAL AND METHODS Adult male Sprague-Dawley (SD) rats (n=60) were divided into the sham group (n=20), the liver resection group (n=20), and the liver resection plus resveratrol-treated group (n=20). Liver resection removed 2/3 of the liver resection; resveratrol was given at a dose of 30 mg/kg/day from one week before surgery until death. Liver injury was assessed by serum liver function tests, including alanine aminotransferase (ALT), aspartate aminotransferase (AST), γ-glutamyl-transferase (γ-GT) and total bilirubin, histological examination of the rat liver, and liver cell apoptosis using the TUNEL assay. High mobility group box 1 (HMGB1) expression was measured by enzyme-linked immunoassay (ELISA). Sirtuin 1 (SIRT1) and acetylated HMGB1 (Ac-HMGB1) expression were detected by Western blot. Normal human liver cells and HepG2 liver cancer cells were incubated with acetylated HMGB1, and albumin production and ammonia elimination assays were performed. RESULTS Resveratrol reduced postoperative liver injury as shown by reduced ALT, AST, γ-GT, and total bilirubin levels, maintained liver structure, and reduced cell apoptosis. Resveratrol treatment reduced the expression and acetylation levels of HMGB1 via the SIRT1 signaling pathway. Resveratrol reversed Ac-HMGB1 induced dysfunction in liver cells cultured in vitro. CONCLUSIONS Resveratrol reduced liver damage after liver resection in a rat model by upregulating SIRT1 and reducing the acetylation of HMGB1.
Subject(s)
HMGB1 Protein/metabolism , Liver Diseases/prevention & control , Liver/drug effects , Resveratrol/pharmacology , Sirtuin 1/metabolism , Acetylation/drug effects , Animals , Apoptosis/drug effects , Disease Models, Animal , Hep G2 Cells , Hepatectomy/adverse effects , Hepatectomy/methods , Hepatocytes/drug effects , Hepatocytes/metabolism , Humans , Liver/pathology , Liver/surgery , Liver Diseases/drug therapy , Liver Diseases/etiology , Liver Diseases/metabolism , Male , Rats , Rats, Sprague-Dawley , Up-RegulationABSTRACT
BACKGROUND: Circulating tumors cells (CTCs) may be a promising prognostic marker for patients with malignant tumors. However, there are few reports regarding its value for hepatocellular carcinoma (HCC) patients. AIMS: To investigate CTCs with epithelial and mesenchymal phenotypes as a potential prognostic biomarker for HCC patients. METHODS: Peripheral blood samples were obtained from 165 HCC patients before radical surgery. CTCs were isolated via the CanPatrol CTC enrichment technique and classified using epithelial-mesenchymal transition (EMT) markers. The relationship of CTC phenotype with clinicopathological factors and HCC recurrence in patients was analyzed. RESULTS: CTC-positive status (count ≥ 2/5 mL) was found in 70.9% of the 165 HCC patients. Increased CTC number was more common in patients with higher AFP levels, multiple tumors, advanced TNM and BCLC staging, and presence of embolus or microembolus (P < 0.05). CTCs heterogeneity was noted using EMT markers. Mesenchymal CTCs were significantly correlated with high AFP levels, multiple tumors, advanced TNM and BCLC stage, presence of embolus or microembolus, and earlier recurrence (P < 0.05). The presence of mesenchymal CTCs predicted the shortest relapse-free survival, followed by mixed phenotypic CTCs, and then epithelial CTCs (P < 0.001). CONCLUSION: CTC phenotype may serve as a prognostic indicator for HCC patients. CTCs assessment should include phenotypic identification tailored to characterize cells based on epithelial and mesenchymal markers.
Subject(s)
Carcinoma, Hepatocellular/surgery , Liver Neoplasms/surgery , Neoplasm Recurrence, Local/surgery , Neoplastic Cells, Circulating , Phenotype , Aged , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/mortality , Female , Follow-Up Studies , Humans , Liver Neoplasms/genetics , Liver Neoplasms/mortality , Male , Middle Aged , Neoplasm Recurrence, Local/genetics , Neoplasm Recurrence, Local/mortality , Neoplastic Cells, Circulating/pathology , Prospective Studies , Survival Rate/trendsABSTRACT
Liver regeneration has important clinical importance in the setting of partial hepatectomy (PH). Following PH, quiescent hepatocytes can reenter cell cycle to restore liver mass. Hepatocyte cell cycle progression, as the basic motivations of liver regeneration, can be disrupted by multiple pathological factors such as oxidative stress. This study aimed to evaluate the role of advanced oxidation protein products (AOPP) in S-phase arrest in hepatocytes. Serum AOPP level were measured during the perioperative period of PH in 33 patients with hepatocellular carcinoma (HCC). Normal Sprague Dawley rats, human and murine liver cell line (HL-7702 and AML-12) were challenged with AOPP prepared by incubation of rat serum albumin (RSA) with hypochlorous acid, and the effect of AOPP on hepatocytes cell cycle progression and liver regeneration was studied after PH. AOPP levels were increased following partial hepatectomy (PH) in patients with primary liver cancer. AOPP treatment impaired liver regeneration in rats following 70% partial hepatectomy. S-phase arrest was induced by AOPP administration in hepatocytes derived from the remnant liver at controlled times following partial hepatectomy in rats, and in HL-7702 and AML-12 cells. The effect of AOPP on hepatocyte S phase arrest was mainly mediated by a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-dependent reactive oxygen species (ROS) generation, downregulation of downstream ß-catenin signaling and decreased cyclin-dependent kinase 2 (CDK2) expression, which inhibited S-phase progression in hepatocytes. This study provides preliminary evidence that AOPP can induce S-phase arrest in hepatocytes via the ROS-dependent, ß-catenin-CDK2-mediated pathway. These findings suggest a novel pathogenic role of AOPP contributing to the impaired liver regeneration and may provide the basis for developing new strategies to improve liver regeneration in patients undergoing PH.
Subject(s)
Advanced Oxidation Protein Products/metabolism , Cyclin-Dependent Kinase 2/metabolism , Hepatocytes/cytology , Liver/physiology , Regeneration , Signal Transduction , beta Catenin/metabolism , Animals , Cell Line , Hepatocytes/metabolism , Humans , Male , Mice , Rats , Rats, Sprague-Dawley , Reactive Oxygen Species/metabolism , S Phase Cell Cycle CheckpointsABSTRACT
Epithelial-mesenchymal transition (EMT) occurs during the progression of liver fibrosis in response to chronic liver injury. However, the molecular mechanism underlying the regulation of hepatocyte EMT remains unclear. The aim of this study was to determine whether advanced oxidation protein products (AOPP) had an effect on hepatocyte EMT. The human L02 hepatocyte cell line and hepatocytes from normal Sprague-Dawley rats were challenged with AOPP treatment in both in vitro and in vivo studies. The expression of cell and molecular markers of EMT in L02 hepatocytes were studied using Western blotting, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assays. Hepatocyte migratory potential was analyzed using a wound healing assay. Intracellular reactive oxygen species (ROS) were detected using the dichlorofluorescein (DCF) assay. In liver tissue sections, expression of EMT markers was evaluated using immunohistochemistry, and collagen was assessed using histochemical staining with Masson's trichrome. The findings were that AOPP treatment resulted in EMT in hepatocytes, which was associated with reduced expression of E-cadherin, increased expression of vimentin, increased deposition of collagen protein, and enhanced cell migration in vivo and in vitro. AOPP was also found to promote migration in L02 cells, and to promote the production of ROS and the activation of TGF-ßR and Smad signaling. Inhibition of the generation of intracellular ROS and TGF-ß receptor blocking could reverse AOPP-induced EMT in hepatocytes. This study has identified a novel mechanism in the regulation of hepatocyte EMT, and the findings may have implications for the control of liver fibrosis.
Subject(s)
Advanced Oxidation Protein Products/metabolism , Hepatocytes/metabolism , Reactive Oxygen Species/metabolism , Smad Proteins/metabolism , Transforming Growth Factor beta/metabolism , Animals , Cadherins/metabolism , Cell Line , Cell Movement , Collagen/metabolism , Epithelial-Mesenchymal Transition/physiology , Humans , Liver/metabolism , Oxidation-Reduction , Rats , Rats, Sprague-Dawley , Receptors, Transforming Growth Factor beta/metabolism , Signal Transduction , Vimentin/metabolismABSTRACT
BACKGROUND: Suture skills are essential to laparoscopic liver resection. The current suture training models are not ideal enough. The aim of this study is to develop and verify a highly simulated-bleeding continuously perfused training model (CPTM) and to evaluate its training efficacy. METHODS: CPTM was constructed using fresh lamb liver whose portal veins were perfused with red-dyed liquid gelatin. Construct validity of CPTMs was tested in 33 participants with three levels of laparoscopic experience (experts, intermediates, and novices) who were demanded to finish one superficial stitch and one deep stitch for suture hemostasis on CPTMs. The CPTMs were also evaluated by the experts. CPTMs were compared with dry box training models (DBTMs) regarding training efficacy among the novices who were assigned to DBTM and CPTM groups to, respectively, complete a 10-day training on CPTMs or DBTMs. Before and after their assignments, their superficial stitches were assessed by completion time, suture accuracy, and suture knot performance while their deep stitches by completion time and bleeding control. RESULTS: CPTM proved to be construct valid by both superficial and deep stitches. Significant differences were found regarding completion time (763, 271, 174 s), suture accuracy (4.4, 1.8, 0.2 mm), and suturing knot performance (12.1, 21.5, 22.0) for superficial stitches (p < 0.001), as well as regarding completion time (807, 423, 277 s) for deep stitches (p < 0.001). Positive comments were given by all experts. CPTMs helped novices to acquire laparoscopic suture skills. Their training efficacy was significantly better than that of DBTMs (p < 0.05). Learning curves of CPTM group plateaued at the sixth round for superficial stitches and at the seventh round for deep stitches. CONCLUSION: CPTM offers trainees a highly simulated-bleeding means to acquire advanced laparoscopic suture skills. The suture skills learned on CPTMs may improve significantly at the seventh round.
Subject(s)
Clinical Competence , Laparoscopy/education , Learning Curve , Liver/surgery , Simulation Training , Suture Techniques/education , Animals , Blood Loss, Surgical , Humans , Male , Models, Anatomic , Reproducibility of Results , SheepABSTRACT
The aim of the current study was to examine the expression of the angiotensin II type 1 receptorassociated protein (ATRAP) in the rat hepatic stellate cell line HSCT6 and to determine its interactions with the local reninangiotensin system (RAS). To achieve this goal, the effect of stimulating HSCT6 cells with angiotensin II (AngII) and angiotensin(17) [Ang(17)], on the expression of ATRAP, the angiotensin II type I receptor (AT1R), the Mas receptor and the angiotensin converting enzyme 2 (ACE2) 2, 6, 12, 18, 24 and 36 h after stimulation was investigated. Changes in expression were quantified at the gene and protein level using RTqPCR and western blotting, respectively. A single dose of AngII (1 µmol/l) significantly increased the gene expression of ATRAP at 12 h, whereas ACE2 gene expression levels were significantly increased at 6 h and then returned to baseline at 12 h, prior to becoming significantly lower. A single dose of Ang(17) at the same concentration as AngII induced ATRAP gene expression, which became statistically significant at the 6 h timepoint, reached a peak at 12 h and remained elevated throughout the experimental timecourse. In addition, ACE2 mRNA expression was significantly suppressed by Ang(17) at 6 h, reaching its lowest expression level at 24 h. The expression of AT1R and the Mas receptor were unaffected by stimulation with AngII and Ang(17). The western blotting results were generally consistent with the mRNA expression data. In conclusion, it was identified that ATRAP is endogenously expressed in HSCT6 cells and therefore, may be critical in regulating the local RAS in these cells.
Subject(s)
Angiotensin II/metabolism , Angiotensin I/metabolism , Peptide Fragments/metabolism , Receptors, Angiotensin/metabolism , Renin-Angiotensin System , Animals , Cell Line , Gene Expression Regulation , RNA, Messenger/genetics , Rats , Receptors, Angiotensin/geneticsABSTRACT
OBJECTIVE: To analyze the long-term therapeutic effects of transcatheter arterial chemoembolization (TACE) before liver transplantation. METHODS: Forty-six patients with hepatocellular carcinoma (HCC) undergoing liver transplantation in our department between November, 2009 and November, 2011 were analyzed. Of these patients, 24 received TACE before the surgery while the other 22 did not. We compared the liver graft function (ALT and AST levels), immune function (CD3, CD4, CD8, CD4/CD8, NK cells, activated T cells), cumulative survival rate and disease-free survival rate between the two groups at 1 year and 2 years after the operation. RESULTS: The patients receiving preoperative TACE showed significantly better liver functions and immune function than those who did not receive TACE (P<0.01 and 0.05). The cumulative disease-free survival rate in the two groups were 72% and 46% at 1 year, and were 57% and 33% at two years, respectively (P<0.05); the cumulative survival rate in the two groups were 84% and 55% at 1 year, and 63% and 34% at two years, respectively (P<0.05). CONCLUSION: TACE prior to liver transplantation can significantly improve the postoperative liver function, immune function, disease-free survival and cumulative survival rates in patients with HCC.
Subject(s)
Carcinoma, Hepatocellular/therapy , Chemoembolization, Therapeutic , Liver Neoplasms/therapy , Liver Transplantation , Disease-Free Survival , Humans , Postoperative Period , Survival RateABSTRACT
BACKGROUND: Metabolomics studies can quantitatively detect the dynamic metabolic response of living systems. OBJECTIVE: To detect urinary metabolomics after hepatic ischemia/reperfusion (I/R) injury induced by the Pringle maneuver using gas chromatography-mass spectrometry (GC-MS). METHODS: Male Sprague-Dawley rats (N = 80) were randomly divided into 4 groups (n = 20/group): sham operation, day 1, day 3, and day 5. Rats in the day 1, day 3, and day 5 groups underwent the Pringle maneuver. Serum alanine transaminase (ALT) and total bilirubin (TBIL) were measured, and hematoxylin and eosin (HE) staining of the liver tissue was performed. GC-MS was used to detect urinary metabolomics. RESULTS: Compared with the sham group, the serum ALT and TBIL levels at day 1 were significantly elevated (P < 0.01) and then decreased and reached close to normal levels at day 5. GC-MS detected 7 metabolites which had similar changes as those of liver tissue revealed by histological examination. Significant differences in lactic acid, pyruvic acid, alanine, serine, and glycerol-3-phosphate were found among the groups (P < 0.001). Principle component analysis showed that 7 metabolites distinguished the day 1 and day 3 groups from the sham group. CONCLUSIONS: Noninvasive urinary metabolomic analysis is a potential means for the early detection and diagnosis of hepatic I/R injury.
Subject(s)
Liver/pathology , Metabolome , Reperfusion Injury/urine , Alanine/metabolism , Alanine/urine , Alanine Transaminase/blood , Animals , Bilirubin/blood , Gas Chromatography-Mass Spectrometry , Glycerophosphates/metabolism , Glycerophosphates/urine , Lactic Acid/metabolism , Lactic Acid/urine , Liver/metabolism , Male , Metabolomics , Pyruvic Acid/metabolism , Pyruvic Acid/urine , Rats , Rats, Sprague-Dawley , Reperfusion Injury/blood , Reperfusion Injury/metabolism , Serine/metabolism , Serine/urineABSTRACT
BACKGROUND: Activation of Kupffer cell (KC) is acknowledged as a key event in the initiation and perpetuation of bile duct warm ischemia/reperfusion injury. The inhibitory effect of gadolinium chloride (GdCl(3)) on KC activation shows potential as a protective intervention in liver injury, but there is less research with regard to bile duct injury. METHODS: Sixty-five male Sprague-Dawley rats (200-250 g) were randomly divided into three experimental groups: a sham group (n = 15), a control group (n = 25), and a GdCl(3) group (n = 25). Specimen was collected at 0.5, 2, 6, 12 and 24 h after operation. Alanine aminotransferase (ALT), alkaline phosphatase (ALP) and total bilirubin (TBIL) of serum were measured. Tumor necrosis factor-α (TNF-α), Capase-3 activity and soluble Fas (sFas) were detected. The pathologic changes of bile duct were observed. Immunochemistry for bile duct Fas was performed. Apoptosis of bile duct cells was evaluated by the terminal UDP nick end labeling assay. RESULTS: GdCl(3) significantly decreased the levels of ALT, ALP and TBIL at 2, 6, 12, and 24 h, and increased serum sFas at 2, 6 and 12 h (P<0.05). TNF-α was lower in the GdCl(3) group than in the control group at 2, 6, 12 and 24 h (P<0.05). Preadministration of GdCl(3) significantly reduced the Caspase-3 activity and bile duct cell apoptosis at 2, 6, 12 and 24 h. After operation for 2, 6 and 12 h, the expression of Fas protein was lower in the GdCl(3) group than in the control group (P<0.05). CONCLUSIONS: GdCl(3) plays an important role in suppressing bile duct cell apoptosis, including decreasing ALT, ALP, TBIL and TNF-α; suppressing Fas-FasL-Caspase signal transduction during transplantation.
Subject(s)
Bile Ducts/blood supply , Bile Ducts/pathology , Gadolinium/therapeutic use , Liver Transplantation , Protective Agents/therapeutic use , Reperfusion Injury/drug therapy , Warm Ischemia , Alanine Transaminase/blood , Animals , Apoptosis/drug effects , Aspartate Aminotransferases/blood , Bile Ducts/drug effects , Bile Ducts/enzymology , Caspase 3/metabolism , Gadolinium/pharmacology , Immunohistochemistry , Male , Protective Agents/pharmacology , Rats , Rats, Sprague-Dawley , Reperfusion Injury/blood , Reperfusion Injury/enzymology , Reperfusion Injury/pathology , Solubility , Time Factors , Tumor Necrosis Factor-alpha/blood , fas Receptor/bloodABSTRACT
OBJECTIVE: To investigate the feasibility and safety of adult-to-adult living-related donor liver transplantation using a right lobe graft. METHODS: The clinical data of 2 cases of living-related donor liver transplantation performed between July, 2010 and November, 2010 were analyzed. RESULTS: Liver transplantation was performed using a right lobe graft including the middle hepatic vein in one case and a right lobe graft without the middle hepatic vein in the other. The ratio of graft volume to standard liver volume was 46.2% and 47.3% in the two cases, with GR/WR of 0.83 and 0.80, and donor residue liver of 42.1% and 39.5%, respectively. The donor operation lasted for 6.5 h and 5 h in the two cases with blood loss of about 200-250 ml without blood transfusion. The donors recovered uneventfully without any surgical complications, whose liver function was normal 7 days after the operation, and were discharged 14 days and 16 days after the surgery, respectively. The recipient operation lasted for 8 h and 7 h with blood loss of about 800-1000 ml. The right hepatic vein, hepatic artery, portal vein and bile duct reconstruction were performed by end-to-end anastomoses in the 2 recipients. Bile duct anastomosis stricture occurred in the first recipient 2 months after transplantation and was treated with percutaneous transhepatic cholangiography and drainage. The second recipient recovered smoothly without any complications. The recipients have so far survived 9 months and 5 months, respectively. CONCLUSION: Adult-to-adult living-related donor liver transplantation is a safe and effective option for treatment of end-stage liver diseases in the context of cadaveric liver graft shortage.
Subject(s)
Liver Transplantation/methods , Living Donors , Adult , Female , Hepatectomy , Humans , Liver Cirrhosis/surgery , Liver Neoplasms/surgery , Male , Middle Aged , Retrospective StudiesABSTRACT
OBJECTIVE: To summarize the experience of donor liver procurement and preparation in liver transplantation. METHODS: One hundred and twenty-six cases of donor liver and kidney procurement and 105 cases of donor liver preparation from August, 2004 to December, 2006 were analyzed. The 105 donor liver grafts were all used for orthotopic liver transplantation. RESULTS: The warm ischemia time of the graft ranged from 1 to 8.5 min with a mean of 4 min. The time of graft procurement ranged from 19 to 28 min (mean 22.5 min). Donor liver preparation lasted for 38 to 102 min in the 105 cases, with a mean of 51 min. The cold ischemia time of the donor liver was 5.5 to 13 h (mean 8 h). Anatomical variations were identified in 8 of the donor liver grafts. CONCLUSIONS: Cold perfusion of the donor liver and repair of the hepatic artery are important procedures in donor liver procurement and preparation. Hemorrhage due to the donor graft should be prevented and the procedures should be performed in close cooperation with the recipient operation.
Subject(s)
Liver Transplantation , Organ Preservation/methods , Tissue Donors , Tissue and Organ Procurement/methods , Adult , Female , Humans , Male , Middle Aged , Young AdultABSTRACT
OBJECTIVE: To study the therapeutic effect of glucocorticoid on early postoperative cholangiole cholestasis hyperbilirubinemia after liver transplantation. METHODS: Thirteen liver transplantation recipients with serum total bilirubin above 171 micromol/L at two weeks to one month postoperatively were enrolled in this study. After exclusion of liver blood supply anomalies, bile duct complications, and acute rejection and establishment of a pathological diagnosis of cholangiole cholestasis by hepatic biopsy, hydrocortisone sodium succinate was infused. The liver functions of the patients were tested at 1 day before and 1 day and 1 week after the treatment. Hepatic biopsy was performed before and 1 week after the treatment to observe histopathological changes. RESULTS: The serum levels of total bilirubin decreased significantly after the treatment with glucocorticoid. Pathology of the hepatic biopsy demonstrated the resolution of cholangiole cholestasis 1 week after the treatment. CONCLUSION: Glucocorticoid treatment is effective for early postoperative cholangiole cholestasis hyperbilirubinemia after liver transplantation.
Subject(s)
Cholestasis, Intrahepatic/complications , Hydrocortisone/analogs & derivatives , Hyperbilirubinemia/drug therapy , Liver Transplantation , Postoperative Complications/drug therapy , Adult , Aged , Female , Humans , Hydrocortisone/therapeutic use , Hyperbilirubinemia/etiology , Male , Middle AgedABSTRACT
OBJECTIVE: To summarize the experience with orthotopic liver transplantation (OLT). METHODS: A retrospective analysis of 105 cases of liver transplantation in our hospital was carried out. RESULTS: All the 105 OLT operations were performed successfully and the operation time ranged from 210-350 min (mean 250 min), with anhepatic phase ranging from 35-65 min (mean 53.5 min) and blood transfusion during operation ranging from 0-6600 ml (mean 400 ml). Ninety-five patients recovered smoothly while the rest 10 died, with the success rate of OLT of 90.5%. Postoperative complications included biliary tract complication (12 cases, 11.4%) and abdominal bleeding (6 cases). CONCLUSIONS: Reducing hemorrhage during operation is the most important factor to ensure successful OLT. Thorough hemostasis during operation and sufficient blood supply to the bile duct can significantly reduce postoperative bile duct complications.