Splenectomy enhances the therapeutic effect of adipose tissue-derived mesenchymal stem cell infusion on cirrhosis rats.

BACKGROUND & AIMS: Clinical studies suggest that splenectomy improves liver function in cirrhotic patients, but the influence of splenectomy on stem cell transplantation is poorly understood. This study investigated the effect of splenectomy on stem cell infusion and elucidated its mechanism. METHODS: Rat adipose tissue-derived mesenchymal stem cells were infused into cirrhosis rats with or without splenectomy, followed by the assessment of the in vivo distribution of stem cells and pathological changes. Stromal cell-derived factor-1 and hepatocyte growth factor expression were also investigated in splenectomized cirrhosis patients and rats. RESULTS: Splenectomy, prior to cell infusion, improved liver function and suppressed fibrosis progression more efficiently than cell infusion alone in the experimental cirrhosis model. Stromal cell-derived factor-1 and hepatocyte growth factor levels after splenectomy were increased in patients and rats. These upregulated cytokines significantly facilitated stem cell motility, migration and proliferation in vitro. C-X-C chemokine receptor type 4 neutralization weakened the promotion of cell migration by these cytokines. The infused cells integrated into liver fibrosis septa and participated in regeneration more efficiently in splenectomized rats. Direct coculture with stem cells led to inhibition of hepatic stellate cell proliferation. In addition, hepatocyte growth factor induced hepatic stellate cell apoptosis via the c-jun N-terminal kinase-p53 pathway. CONCLUSIONS: Splenectomy prior to cell infusion enhanced the therapeutic effect of stem cells on cirrhosis, which involved upregulation of stromal cell-derived factor-1 and hepatocyte growth factor after splenectomy.

The microenvironment in hepatocyte regeneration and function in rats with advanced cirrhosis.

UNLABELLED: In advanced cirrhosis, impaired function is caused by intrinsic damage to the native liver cells and from the abnormal microenvironment in which the cells reside. The extent to which each plays a role in liver failure and regeneration is unknown. To examine this issue, hepatocytes from cirrhotic and age-matched control rats were isolated, characterized, and transplanted into the livers of noncirrhotic hosts whose livers permit extensive repopulation with donor cells. Primary hepatocytes derived from livers with advanced cirrhosis and compensated function maintained metabolic activity and the ability to secrete liver-specific proteins, whereas hepatocytes derived from cirrhotic livers with decompensated function failed to maintain metabolic or secretory activity. Telomere studies and transcriptomic analysis of hepatocytes recovered from progressively worsening cirrhotic livers suggest that hepatocytes from irreversibly failing livers show signs of replicative senescence and express genes that simultaneously drive both proliferation and apoptosis, with a later effect on metabolism, all under the control of a central cluster of regulatory genes, including nuclear factor κB and hepatocyte nuclear factor 4α. Cells from cirrhotic and control livers engrafted equally well, but those from animals with cirrhosis and failing livers showed little initial evidence of proliferative capacity or function. Both, however, recovered more than 2 months after transplantation, indicating that either mature hepatocytes or a subpopulation of adult stem cells are capable of full recovery in severe cirrhosis. CONCLUSION: Transplantation studies indicate that the state of the host microenvironment is critical to the regenerative potential of hepatocytes, and that a change in the extracellular matrix can lead to regeneration and restoration of function by cells derived from livers with end-stage organ failure.

Autologous bone marrow cell infusions suppress tumor initiation in hepatocarcinogenic mice with liver cirrhosis.

We have previously reported the efficacy and safety of autologous bone marrow cell infusion (ABMi) therapy for liver cirrhosis patients without hepatocellular carcinoma in a multicenter clinical trial. However, since liver cirrhosis is highly oncogenic, evaluation of the effects of ABMi on the mechanisms of hepatocarcinogenesis is of great importance. Therefore, frequent ABMi was performed in hepatocarcinogenic mice, and its effects on hepatocarcinogenesis were analyzed. The N-nitrosodiethylamine (DEN)/green fluorescent protein (GFP)-carbon tetrachloride (CCl(4) ) model was developed by administering DEN once, followed by repeated administration of CCl(4) intraperitoneally as for the control group. In the administration (ABMi) group, GFP-positive bone marrow cells were infused through a tail vein. The kinetics of hepatocarcinogenesis were evaluated histologically 4.5 months after DEN treatment. At 4.5 months, there was significantly lower incidence of foci and tumors in the ABMi group, and they were smaller in number, while their size was almost equal. No GFP-positive tumors were found in ABMi livers. Moreover, ABMi livers showed significantly reduced liver fibrosis, consistent with significantly lower 8-hydroxy-2'-deoxyguanosine levels, higher superoxide dismutase activity, and increased nuclear translocation of nuclear factor-erythroid 2 p45-related factor 2. These results demonstrate that frequent ABMi might contribute to suppressed tumor initiation during stages of hepatocarcinogenesis, consistent with improvements in liver fibrosis and stabilization of redox homeostasis.

Role of angiotensin-1 receptor blockade in cirrhotic liver resection.

BACKGROUND: The regeneration capacity of cirrhotic livers might be affected by angiotensin-1 (AT1) receptors located on hepatic stellate cells (HSC). The effect of AT1 receptor blockade on microcirculation, fibrosis and liver regeneration was investigated. MATERIALS AND METHODS: In 112 Lewis rats, cirrhosis was induced by repetitive intraperitoneal injections of CCl(4) . Six hours, 3, 7 and 14 days after partial hepatectomy or sham operation, rats were sacrificed for analysis. Animals were treated with either vehicle or 5 mg/kg body weight losartan pre-operatively and once daily after surgery by gavage. Microcirculation and portal vein flow were investigated at 6 h. The degree of cirrhosis was assessed by Azan Heidenhein staining, activation of HSC by desmin staining, apoptosis by ssDNA detection and liver regeneration by Ki-67 staining. Changes in expression of various genes important for liver regeneration and fibrosis were analysed at 6 h and 3 days. Haemodynamic parameters and liver enzymes were monitored. RESULTS: Losartan treatment increased sinusoidal diameter, sinusoidal blood flow and portal vein flow after partial hepatectomy (P<0.05), but not after sham operation. AT1 receptor blockade resulted in increased apoptosis early after resection. HSC activation was reduced and after 7 days, a significantly lower degree of cirrhosis in resected animals was observed. Losartan increased the proliferation of hepatocytes at late time-points and of non-parenchymal cells early after partial hepatectomy (P<0.05). Tumour necrosis factor (TNF)-α was significantly upregulated at 6 h and stem cell growth factor (SCF) was downregulated at 3 days (P<0.05). CONCLUSION: Losartan increased hepatic blood flow, reduced HSC activation and liver fibrosis, but interfered with hepatocyte proliferation after partial hepatectomy in cirrhotic livers.

Macrophage-mediated phagocytosis of apoptotic cholangiocytes contributes to reversal of experimental biliary fibrosis.

Studies have suggested the reversibility of liver fibrosis, but the mechanisms of fibrosis reversal are poorly understood. We investigated the possible functional link between apoptosis, macrophages, and matrix turnover in rat liver during reversal of fibrosis secondary to bile duct ligation (BDL). Biliary fibrosis was induced by BDL for 4 wk. After Roux-en-Y (RY)-bilio-jejunal-anastomosis, resolution of fibrosis was monitored for up to 12 wk by hepatic collagen content, matrix metalloproteinase (MMP) expression and activities, and fibrosis-related gene expression. MMP expression and activities were studied in macrophages after engulfment of apoptotic cholangiocytes in vitro. Hepatic collagen decreased to near normal at 12 wk after RY-anastomosis. During reversal, profibrogenic mRNA declined, whereas expression of several profibrolytic MMPs increased. Fibrotic septa showed fragmentation at week 4 and disappeared at week 12. Peak histological remodeling at week 4 was characterized by massive apoptosis of cytokeratin 19+ cholangiocytes, >90% in colocalization with CD68+ macrophages, and a 2- to 7.5-fold increase in matrix-degrading activities. In vitro, phagocytosis of apoptotic cholangiocytes induced matrix-degrading activities and MMP-3, -8, and -9 in rat peritoneal macrophages. We concluded that reconstruction of bile flow after BDL leads to an orchestrated fibrolytic program that results in near complete reversal of advanced fibrosis. The peak of connective tissue remodeling and fibrolytic activity is associated with massive apoptosis of cholangiocytes and their phagocytic clearance by macrophages in vivo. Macrophages upregulate MMPs and become fibrolytic effector cells upon apoptotic cholangiocyte engulfment in vitro, suggesting that phagocytosis-associated MMP induction in macrophages significantly contributes to biliary fibrosis reversal.

Decreased collagen types I and IV, laminin, CK-19 and α-SMA expression after bone marrow cell transplantation in rats with liver fibrosis.

Bone marrow cells have frequently been tested in animal models of liver fibrosis to assess their role in hepatic regeneration. The mononuclear fraction of bone marrow cells is of particular interest, as many studies show that these cells may be beneficial to treat hepatic fibrosis. In this study, we used the bile duct ligation model to induce hepatic fibrosis in an irreversible manner, and rats were treated with bone marrow mononuclear (BMMN) cells after fibrosis was established. Analysis of collagen types I and IV, laminin and α-SMA showed a decreased expression of these proteins in fibrotic livers after 7 days of BMMN cell injection. Moreover, cytokeratin-19 analysis showed a reduction in bile ducts in the BMMN cell-treated group. These results were accompanied by ameliorated levels of hepatic enzymes GPT (Glutamic-pyruvic transaminase), GOT (glutamic-oxaloacetic transaminase) and alkaline phosphatase (AP). Therefore, we showed that BMMN cells decrease hepatic fibrosis by significantly reducing myofibroblast numbers and through reduction of the collagen and laminin-rich extracellular matrix of fibrotic septa and hepatic sinusoids.

[Cryosurgery in diffuse hepatic diseases].

Comparative studying, using histological and biomicroscopic methods, of the dosed cryohepatodestruction (CHD), periarterial cryodenervation of hepatic artery (CDHA) and their concomitant application influence on the dynamics of hepatic restoration processes in experimental cirrhosis was performed. The investigations were done on 215 male rats owing body mass 200-280 g in a not changed and pathologically changed liver. There was shown, that CDHA promotes changes in hepatic tissue microhemocirculation, as well as the enhancement of the sinusoidal vessels diameter and relative square of vascular bed. CHD stimulates the reparative processes course in a pathologically changed organ. There was established, that while simultaneous application of two cryosurgical methods, the velocity and grade of restoration processes in cirrhotically-changed liver are enhanced in comparison with such indices changes while separate usage of these two methods.

Transplanted endothelial progenitor cells ameliorate carbon tetrachloride-induced liver cirrhosis in rats.

Cirrhosis is the most common end stage of liver diseases, and there are no effective treatment methods. Here we evaluated the effect of endothelial progenitor cell (EPC) transplantation from rat bone marrow (BM) on the development of cirrhosis induced by carbon tetrachloride (CCl(4)). Ex vivo generated, characterized, and cultivated rat BM-derived EPCs were identified by their vasculogenic properties in vitro. EPCs from male rats were transplanted into female rats via the intraportal vein 12 weeks after they had been challenged with CCl(4), and the rats were killed 16 weeks later. The control rats received only a saline infusion. The fibrosis index and donor cell engraftment were assessed after EPC transplantation. After transplantation via the portal vein, PKH26 labeling, polymerase chain reaction, and in situ hybridization analysis revealed that the donor EPCs had adhered to the vasolateral surfaces of blood vessels and established in the liver. EPCs reduced the expression of alpha-smooth muscle actin, collagen III, and transforming growth factor beta (P < 0.05) as well as levels of aspartate aminotransferase, alanine aminotransferase, and total bilirubin in the serum (P < 0.05), but at the same time they increased the levels of albumin and Ki67. CCl(4) treatment increased the international prothrombin ratio (P < 0.05) and reduced albumin levels, whereas EPCs restored these parameters to normal levels. These results suggest that EPC transplantation could play a role in regulating hepatocyte regeneration and ameliorating established liver cirrhosis.

Multipotent cells from the human third molar: feasibility of cell-based therapy for liver disease.

Adult stem cells have been reported to exist in various tissues. The isolation of high-quality human stem cells that can be used for regeneration of fatal deseases from accessible resources is an important advance in stem cell research. In the present study, we identified a novel stem cell, which we named tooth germ progenitor cells (TGPCs), from discarded third molar, commonly called as wisdom teeth. We demonstrated the characterization and distinctiveness of the TGPCs, and found that TGPCs showed high proliferation activity and capability to differentiate in vitro into cells of three germ layers including osteoblasts, neural cells, and hepatocytes. TGPCs were examined by the transplantation into a carbon tetrachloride (CCl4)-treated liver injured rat to determine whether this novel cell source might be useful for cell-based therapy to treat liver diseases. The successful engraftment of the TGPCs was demonstrated by PKH26 fluorescence in the recipient's rat as to liver at 4 weeks after transplantation. The TGPCs prevented the progression of liver fibrosis in the liver of CCl4-treated rats and contributed to the restoration of liver function, as assessed by the measurement of hepatic serum markers aspartate aminotransferase and alanine aminotransferase. Furthermore, the liver functions, observed by the levels of serum bilirubin and albumin, appeared to be improved following transplantation of TGPCs. These findings suggest that multipotent TGPCs are one of the candidates for cell-based therapy to treat liver diseases and offer unprecedented opportunities for developing therapies in treating tissue repair and regeneration.

Observation of the effect of bone marrow mesenchymal stem cell transplantation by different interventions on cirrhotic rats.

Bone marrow mesenchymal stem cells (BMSCs) transplantation has attracted attention for the treatment of liver cirrhosis and end-stage liver diseases. Therefore, in this study, we evaluated the effect of different methods of BMSCs transplantation in the treatment of liver cirrhosis in rats. Seventy-two male Sprague-Dawley rats were divided into 7 groups: 10 were used to extract BMSCs, 10 were used as normal group, and the remaining 52 rats were randomly divided into five groups for testing: control group, BMSCs group, BMSCs+granulocyte colony-stimulating factor (G-CSF) group, and BMSCs+Jisheng Shenqi decoction (JSSQ) group. After the end of the intervention course, liver tissue sections of rats were subjected to hematoxylin and eosin (H&E) and Masson staining, and pathological grades were scored. Liver function [aminotransferase (ALT), aspartate aminotransferase (AST), albumin (ALB)] and hepatic fibrosis markers [hyaluronidase (HA), laminin (LN), type III procollagen (PCIII), type IV collagen (CIV)] were measured. BMSCs+JSSQ group had the best effect of reducing ALT and increasing ALB after intervention therapy (P<0.05). The reducing pathological scores and LN, PCIII, CIV of BMSCs+G-CSF group and BMSCs+JSSQ group after intervention therapy were significant, but there was no significant difference between the two groups (P>0.05). The effect of JSSQ on improving stem cell transplantation in rats with liver cirrhosis was confirmed. JSSQ combined with BMSCs could significantly improve liver function and liver pathology scores of rats with liver cirrhosis.

Potentials of Differentiated Human Cord Blood-Derived Unrestricted Somatic Stem Cells in Treatment of Liver Cirrhosis.

OBJECTIVES: Liver transplantation is the well-known treatment for chronic liver diseases; however, postoperative complications and lack of donors continue to be limitations with this treatment. Investigating new modalities for treatment of chronic liver illness is a must. In the present study, we aimed to clarify the effects of an in vitro hepatocyte-differentiated human unrestricted somatic stem cell transplant as a new cell-based therapy in an experimental model of chronic liver failure. MATERIALS AND METHODS: Human umbilical cord blood-derived unrestricted somatic stem cells were isolated, cultured, propagated, and characterized. Cells were directed to differentiate into hepatocyte-like cells. An animal model of carbon tetrachloride cirrhotic liver failure was prepared, and the human in vitro differentiated unrestricted somatic stem cells were transplanted into the experimental model. Animals that did not receive transplant served as the pathologic control group. Animals were euthanized 12 weeks after transplant, and liver functions and histopathology were assessed. RESULTS: Compared with the pathologic control group, the transplant group showed improvements in levels of alanine aminotransferase, aspartate aminotransferase, albumin, and bilirubin. Histopathologic examination of the transplant group also showed improvements in hydropic degeneration and fibrosis. CONCLUSIONS: The use of unrestricted somatic stem cells, isolated and propagated from cord blood and then differentiated into hepatocyte-like cells, improved both fibrosis and normal function of cirrhotic livers. These cells could be considered as a line of cell-based therapy in cases of chronic liver disease.

IL-6 Promotes compensatory liver regeneration in cirrhotic rat after partial hepatectomy.

Major hepatic resection in cirrhotic patients is associated with impaired liver regeneration and failure, leading to high peri-operative mortality. In this work, the causes of defective regeneration in cirrhotic liver and the utility of IL-6 treatment were investigated in an experimental model combining cirrhosis and partial hepatectomy in the rat. Relative to normal controls, decompensated cirrhotic animals showed decreased survival, while compensated cirrhotic animals showed similar survival but reduced hepatic DNA synthesis and newly regenerated liver mass amount. Defective liver regeneration was associated with a decrease in STAT3 and NF-kB activation, consistent with an increased accumulation of their respective inhibitors PIAS3 and IkBalpha, and with a decreased induction of Bcl-xL. Treatment with recombinant IL-6 enhanced survival of decompensated cirrhotic animals, while it did not affect survival of compensated cirrhotic animals but sustained liver regeneration, by restoring STAT3 and NF-kB activation and Bcl-xL induction to the levels found in normal controls. The pro-growth effects exerted by IL-6 treatment in cirrhotic liver were attained also at low, pharmacologically acceptable doses. In conclusion, our results suggest that IL-6 treatment may be therapeutic in major resection of cirrhotic liver.

Bone marrow cell transplant does not prevent or reverse murine liver cirrhosis.

We tested the effect of bone marrow cell (BMC) transplantation in either preventing or reversing cirrhosis on an experimental model of chronic liver disease. Female Wistar rats were fed a liquid alcohol diet and received intraperitoneal injections of carbon tetrachloride (CCl4) over 15 weeks. Ten animals (cell-treated group) received five injections of BMCs during the cirrhosis induction protocol (on the 4th, 6th, 8th, 10th, and 12th weeks) and four animals received the cells after liver injury was established through tail vein. Nine animals (nontreated group) were submitted to the previously described protocols; however, they received vehicle injections. Analyses were performed to verify whether the infusion of cells was effective in preventing the development of cirrhosis in our model of induction, and if the cells could reverse cirrhosis once it was established. Hepatic architecture and fibrotic septa were analyzed in liver slices stained with hematoxilin & eosin and Sirius red, respectively. Fibrosis quantification was measured by Sirius red histomorphometry. Indirect immunofluorescence was performed to detect the amount of tissue transglutaminase 2. Blood analyses were performed to assess liver injury and function by the assessment of alanine aminotransferase and albumin. Ultrasound was performed to analyze the portal vein caliber and presence of ascitis. Cirrhosis features (regenerative nodules and fibrous septa) were observed in histopathology after 15 weeks of continuous hepatic injury in nontreated and cell-treated groups. Collagen content, immunofluorescence analysis, and biochemical and ultrasound parameters were similar in nontreated and cell-treated groups; however, both groups showed significant differences compared to a normal control group. Cell infusions with bone marrow-derived cells seem to be ineffective in improving morphofunctional parameters of the liver when applied to chronic cases either during or after establishment of the hepatic lesion.

Autologous bone marrow cell infusion therapy for liver cirrhosis.

The plasticity of bone marrow cells (BMC) has been confirmed by autopsy results of female recipients of BMC from male donors. To establish new clinical therapies for patients with liver cirrhosis using autologous BMC, we developed a new in vivo murine model using green fluorescent protein (GFP) and repeated carbon tetrachloride (CCl(4)) injection. We found that BMC infused through the tail vein, efficiently repopulated cirrhotic liver tissue and, under the influence of persistent liver damage induced by carbon tetrachloride, differentiated into albumin-producing hepatocytes. Moreover, such BMC infusions into mice with cirrhosis improved liver function and reduced mortality. The latter observation correlated with the strong expression of matrix metalloproteinases (MMP), particularly MMP-9, and reduced hepatic fibrosis. The results from the 'GFP/CCl(4) model' showed that cell therapy using autologous BMC has the potential to become an effective treatment for patients with liver failure due to advanced liver cirrhosis. This review summarizes previous findings plus these recent experimental results, as well as recent clinical trials of BMC transfusion into patients with end-stage chronic liver disease.

Functional hepatic recovery after xenotransplantation of cryopreserved fetal liver cells or soluble cell-factor administration in a cirrhotic rat model: are viable cells necessary?

BACKGROUND AND AIM: Chronic liver failure results in the decrease of the number of functioning hepatocytes. It dictates the necessity of using exogenous viable cells or/and agents that can stimulate hepatic regenerative processes. Fetal liver contains both hepatic and hematopoietic stem cells with high proliferative potential, which may replace damaged cells. Also, immature cells produce fetal-specific factors which may support the injured liver. Our aim was to test the ability of human fetal liver cells and cell-free fetal-specific factors of non-hepatic origin to stimulate recovery processes in an experimental model of carbon tetrachloride-induced cirrhosis in rats. METHODS: Cirrhotic rats were intrasplenically injected with fetal liver cells (1 x 10(7) cells/0.3 mL medium) or cell-free fetal-specific factors (0.3 mL/1 mg protein). Control groups received medium alone. Serum indexes, hepatic functions, and morphology were evaluated for 15 days. RESULT: Human fetal liver cell transplantation was shown to abrogate the mortality of cirrhotic animals, to improve serum markers, and to restore liver mitochondrial function and detoxification. Morphological patterns of liver recovery were observed by histology. In comparison, an injection of fetal-specific factors produced similar functional recovery, whilst a more limited liver regeneration was observed by histology. CONCLUSIONS: The positive effects of fetal liver cell and cell-free fetal-specific factors in experimental cirrhosis may result from the presence of stage-specific factors activating hepatocellular repair.

Restrictive model of compensated carbon tetrachloride-induced cirrhosis in rats.

AIM: To develop a simplified and quick protocol to induce cirrhosis and standardize models of partial liver resection in rats. METHODS: In Fischer F344 rats two modified protocols of phenobarbital-carbon tetrachloride (CCl4) (dilution 50%) gavage to induce cirrhosis (frequency adjusted according to weight, but each subsequent dose was systematically administered) were tested, i.e. the rapid and slow protocols. Prothrombin time (PT) and total bilirubin (TB) were also evaluated. Animals from the rapid group underwent 15% hepatectomy and animals from the slow group underwent 70% hepatectomy. RESULTS: Rapid protocol: This corresponded to 1 gavage/4 d over 6 wk (mortality 30%). Mean PT was 35.2+/-2.8 s (normal: 14.5 s), and mean TB was 1.8+/-0.2 mg/dL (normal: 0.1 mg/dL). Slow protocol: This corresponded to 1 gavage/6 d over 9 wk (mortality 10%). Mean PT was 11.8+/-0.2 s (normal: 14.5 s), and mean TB was 0.4+/-0.04 mg/dL (normal: 0.1 mg/dL). Pathological analyses were performed in both protocols which showed persistent cirrhosis at 3 mo. Rat mortality in the rapid gavage group who underwent 15% hepatectomy and in the slow gavage group who underwent 70% hepatectomy was 50% and 70%, respectively. CONCLUSION: Our modified model is a simplified method to induce cirrhosis which is rapid (6 to 9 wk), efficient and stable up to 3 mo. Using this method, "Child Pugh A" or "Child Pugh BC" cirrhotic rats were obtained. Our models of cirrhosis and hepatectomy can be used in various situations focusing on postoperative survival.

Therapy with bone marrow cells reduces liver alterations in mice chronically infected by Schistosoma mansoni.

AIM: To investigate the potential of bone marrow mononuclear cells (BM-MCs) in the regeneration of hepatic lesions induced by Schistosoma mansoni (S.mansoni) chronic infection. METHODS: Female mice chronically infected with S.mansoni were treated with BM-MCs obtained from male green fluorescent protein (GFP) transgenic mice by intravenous or intralobular injections. Control mice received injections of saline in similar conditions. Enzyme-linked immunosorbent assay (ELISA) assay for transforming growth factor-beta (TGF-beta), polymerase chain reaction (PCR) for GFP DNA, immunofluorescence and morphometric studies were performed. RESULTS: Transplanted GFP(+) cells migrated to granuloma areas and reduced the percentage of liver fibrosis. The presence of donor-derived cells was confirmed by fluorescence in situ hybridization (FISH) analysis for detection of cells bearing Y chromosome and by PCR analysis for detection of GFP DNA. The levels of TGF-beta, a cytokine associated with fibrosis deposition, in liver fragments of mice submitted to therapy were reduced. The number of oval cells in liver sections of S.mansoni-infected mice increased 3-4 fold after transplantation. A partial recovery in albumin expression, which is decreased upon infection with S.mansoni, was found in livers of infected mice after cellular therapy. CONCLUSION: In conclusion, transplanted BMCs migrate to and reduce the damage of chronic fibrotic liver lesions caused by S.mansoni.

[Effects of gene-transfected bone marrow-derived liver stem cell transplantation on accumulation of extracellular matrix in rats with liver fibrosis].

OBJECTIVE: To explore the effects of urokinase-type plasminogen activator (uPA) gene-modified bone marrow-derived stem cell (BDLSC) transplantation on accumulation of extracellular matrix (ECM) in hepatic tissue in liver fibrosis. METHODS: BDLSCs obtained from 10 male Fisher344 rats were transfected by adenovirus-mediated human uPA (AduPA) in vitro. Twenty-seven female rats were randomly divided into 3 equal groups to undergo subcutaneous injection of carbon tetrachloride to establish liver fibrosis models and then randomly divided into 3 equal groups: model group injected with normal saline via caudal vein, BDLSC group injected with 2 x 10(6) BDLSCs via caudal vein, and BDLSC-uPA group injected with 2 x 10(6) AduPA-transfected BDLSCs. Eight weeks later the serum levels of alanine transaminase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), albumin (ALB), and ECM levels, i.e., hyaluronic acid, laminin (LN), and procollagen III (PC III), were detected. Then the rats were killed with their livers taken out. The hydroxyproline (Hyp) content of the liver was detected by alkaline hydrolysis. RT-PCR was used to examine the expression of collagen I and III (COLI and COLIII), matrix metalloproteinases-2, 3, and 9 (MMP-2, 3, and 9), and tissue inhibitor of metalloproteinase-1 and 2 (TIMP-1 and 2). RESULTS: Compared with those of the model group the levels of ALT, AST, and TBIL of the BDLSC-uPA group were all significantly lower, and the ALB level was higher (all P < 0.05). The ECM levels of BDLSC-uPA group were all significantly lower than those of the model group or BDLSC group too (all P < 0.05). Hyp content of the liver decreased dramatically. The mRNA expression levels of COLI and COLIII of the liver of the BDLSC-uPA group were significantly lower (38.9 +/- 2.7, 8.5 +/- 1.6), and the mRNA expression levels of MMP-2, -3, and MMP-9 mRNA (157.5 +/- 32.6, 105.5 +/- 14.6, 187.5 +/- 22.8) were significantly higher than those of the model group or BDLSC group (all P < 0.05), but no significant differences were observed in the mRNA expression of TIMP-1 and 2 mRNA between the 3 groups (all P > 0.05). CONCLUSION: uPA gene-modified BDLSC transplantation improves the liver function and suppresses the hepatic fibrosis in liver cirrhosis through up-regulating the expression of MMPs and promoting the degradation of ECM.

Preoperative erythropoietin treatment improves survival following major hepatic resection in a cirrhotic rat model.

AIM: Major hepatic resection of a cirrhotic liver may result in a fatal clinical course. Preoperative erythropoietin (EPO) treatment has been shown to have protective properties and to stimulate liver regeneration. This study aims to investigate the effect of preoperative EPO on survival following major hepatic resection in a cirrhotic rat model. METHODS: Cirrhotic liver was induced by intraperitoneal injection of thioacetamide (200mg/kg/mL) in 72 Lewis rats. Each 36 rats received EPO (1IU/g, every second day, 5 times preoperatively) or saline (control) and major hepatectomy (removal of the left and half of the median lobe) was performed. Biochemical and immunohistochemical parameters, cytokines and overall survival were compared following surgery. RESULTS: Rats that received preoperative EPO had decreased hepatic aspartate aminotransferase, alanine aminotransferase and interleukin (IL)-1ß expression, 48hours following surgery. They had increased hepatocyte growth factor and vascular endothelial growth factor expression at 1hour, increased IL-6 expression at 24, 48 and 120hours and increased Ki-67, 120hours following surgery. Overall, survival was significantly improved among EPO-treated rats (P=0.034). CONCLUSION: Preoperative EPO treatment has a protective effect and stimulates liver regeneration, leading to improved overall survival following major hepatectomy in a cirrhotic rat model.

Transplant of Hepatocytes, Undifferentiated Mesenchymal Stem Cells, and In Vitro Hepatocyte-Differentiated Mesenchymal Stem Cells in a Chronic Liver Failure Experimental Model: A Comparative Study.

OBJECTIVES: Liver transplant is the cornerstone line of treatment for chronic liver diseases; however, the long list of complications and obstacles stand against this operation. Searching for new modalities for treatment of chronic liver illness is a must. In the present research, we aimed to compare the effects of transplant of undifferentiated human mesenchymal stem cells, in vitro differentiated mesenchymal stem cells, and adult hepatocytes in an experimental model of chronic liver failure. MATERIALS AND METHODS: Undifferentiated human cord blood mesenchymal stem cells were isolated, pro-pagated, and characterized by morphology, gene expression analysis, and flow cytometry of surface markers and in vitro differentiated into hepatocyte-like cells. Rat hepatocytes were isolated by double perfusion technique. An animal model of chronic liver failure was developed, and undifferentiated human cord blood mesenchymal stem cells, in vitro hepato-genically differentiated mesenchymal stem cells, or freshly isolated rat hepatocytes were transplanted into a CCL4 cirrhotic experimental model. Animals were killed 3 months after transplant, and liver functions and histopathology were assessed. RESULTS: Compared with the cirrhotic control group, the 3 cell-treated groups showed improved alanine aminotransferase, aspartate aminotransferase, albumin, and bilirubin levels, with best results shown in the hepatocyte-treated group. Histopathologic examination of the treated groups showed improved fibrosis, with best results obtained in the undifferentiated mesenchymal stem cell-treated group. CONCLUSIONS: Both adult hepatocytes and cord blood mesenchymal stem cells proved to be promising candidates for cell-based therapy in liver regeneration on an experimental level. Improved liver function was evident in the hepatocyte-treated group, and fibrosis control was more evident in the undifferentiated mesenchymal stem cell-treated group.