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Objective To investigate the effect of knock⁃down 3⁃phosphoglycerin dehydrogenase (PHGDH) targeting energy metabolism on malignant biological behavior and osteogenic differentiation of human osteosarcoma 143B cells. Methods Real⁃time PCR and Western blotting were used to detect the expression of PHGDH in osteoblasts hFOB1. 19 and osteosarcoma cells TE85, MG63 and 143B with different malignant degrees. The short hairpin RNA (shRNA)⁃PHGDH recombinant plasmid was transfected into 143 B cells by liposome transfection method. The expression of PHGDH was detected by Real⁃time PCR and Western blotting. Crystal violet staining, cell counting and CCK⁃8 assay were used to detect cell proliferation; wound healing assay was used to detect cell parallel migration ability, and Transwell assay was used to detect cell vertical migration and invasion ability. Annexin V⁃FITC/ PI double staining and DAPI staining were used to detect apoptosis; Alkaline phosphatase(ALP) staining and alizarin red S staining were used to detect osteogenic differentiation. Western blotting was used to detect the expression of Runt related transcription factor 2 (Runx2) and osteocalcin (OC) . The expression of genes related to energy metabolism, glucose transporter⁃1 (GLUT1), 6⁃ phosphofructokinase⁃1(PFK1), pyruvate kinae subtype M2 (PKM2), lactate dehydrogenase A (LDHA) was detected by Real⁃time PCR. Lactic acid secretion was detected by lactic acid detection kit. Adenosine triphosphate(ATP) production was detected by ATP detection kit. Results The expression of PHGDH in 143B cells was significantly higher than that in hFOB1. 19, MG63 and TE85 cells (P < 0. 01) . After the transfection of shRNA⁃PHGDH recombinant plasmid, the expression of PHGDH in 143 B cells decreased (P<0. 01), proliferation ability decreased (P<0. 01), cell migration and invasion ability decreased (P < 0. 01), apoptosis rate increased (P < 0. 01), ALP staining positive rate increased (P < 0. 01), alizarin red staining positive rate increased (P < 0. 05), Runx2 (P < 0. 05) and OC expression increased (P < 0. 01), expression of genes related to energy metabolism (GLUT1, PFK1, PKM2, LDHA) decreased (P < 0. 01), lactic acid decreased (P < 0. 01), ATP increased (P < 0. 05) . Conclusion Knocking down of PHGDH can inhibit the proliferation, migration and invasion of human osteosarcoma 143B cells through energy metabolism, promote their apoptosis and promote their osteogenic differentiation.
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[Abstract] Objective To investigate the effect of wingless-type MMTV integration site family member 5b(Wnt5b) gene overexpression mediated by recombinant adenovirus on the differentiation of mouse embryonic liver stem cells and repair of chronic liver injury in mice. Methods Recombinant adenoviruses expressing Wnt5b and green fluorescent protein (GFP) were applied respectively to infect mouse fetal liver stem cells HP14-19, and induced its differentiation and verified the expression of Wnt5b through Real-time PCR and Western blotting. It also applied indocyanine grean(ICG) uptake experiment and periodic acid-schiff(PAS) staining to detect the differentiation ability of HP14-19 into hepatocyte-like cells. The Real-time PCR was chosen to detect hepatocyte markers albumin (Alb) and cytokeratin 18 (CK18) expression. Forty-eight experimental male BALB/ c mice were randomly divided into control group, model group, stem cell treatment group and Wnt5b modified stem cell treatment group. The carbon tetrachloride(CCl
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With the development and maturity of liver transplant techniques, liver transplantation has become one of the vital treatment options for hepatocellular carcinoma (HCC). Postoperative recurrence and metastasis of HCC after liver transplantation is one of critical factors that affect the long-term survival of recipients. Exploring the prevention and therapeutic strategies for HCC recurrence and metastasis after liver transplantation plays a pivotal role in improving the clinical efficacy of liver transplantation for HCC recipients. Intimate monitoring, active prevention, early diagnosis, comprehensive surgical treatment and local treatment, especially targeted immunotherapy, and individualized prevention and therapeutic strategies are of significance for the prevention and treatment of HCC recurrence and metastasis after liver transplantation. In this article, the monitoring, diagnosis, prevention and treatment of tumor recurrence and metastasis after liver transplantation for HCC were reviewed, aiming to provide reference for the prevention and treatment of tumor recurrence and metastasis after liver transplantation, enhancing clinical efficacy of liver transplantation and prolonging the survival of recipients.
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@#To study the effects of sarmentosin(SA)on the intervention and regulation of juvenile intrahepatic cholestasis in rats, 48 young SD rats were randomly divided into the control group, α-naphthylisothiocyanate(ANIT)model group, ursodeoxycholic acid(UDCA)positive control group and low-, medium- and high- dosage groups of SA, with 8 rats in each group. Except for the control group, rats in each group were given corresponding drugs by gavage once a day for a week, and 80 mg/kg ANIT model was established on the 5th day. Bile excretion was measured 48 hours after the establishment of the model; the activities of alanine aminotransferase(ALT), aspartate aminotransferase(AST)and alkaline phosphatase(ALP)and the contents of total bilirubin(TBIL), direct bilirubin(DBIL)and total bile acid(TBA)in serum were measured. The pathological changes of liver tissue and the contents of malondialdehyde(MDA), superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)in tissue homogenate were detected, and the expressions of tumor necrosis factor-α(TNF-α), γ-interferon(IFN-γ)and interleukin-1β(IL-1β)in serum were determined. Bile acid transporters and synthetic proteins were analyzed by Western blot. Compared with the control group, bile excretion was significantly inhibited in the model group; liver tissue showed obvious pathological damage; serum levels of ALT, AST, ALP, TBIL, DBIL and TBA were significantly increased; MDA content in tissue homogenate was significantly increased, SOD and GSH-Px contents were significantly decreased; inflammatory factors TNF-α, IFN-γ and IL-1β were significantly decreased in the model group. The expression of FXR, SHP-1, SHP-2, MREP2, BSEP and NTCP decreased, while the expression of CYP7A1 and CYP27A1 increased. Compared with the model group, the bile excretion of rats in each dose of SA group increased in varying degrees; the pathological damage of liver tissue was improved; the levels of ALT, AST, ALP, TBIL, DBIL and TBA in serum were decreased; the contents of MDA in liver homogenate were decreased; and the contents of SOD and GSH-Px were increased; the expression of TNF-α, IFN-γ and IL-1β decreased. The results showed that sarmentosin had a certain therapeutic effect on cholestasis. The effect of high dose of SA was similar to that of UDCA group, while SA could up-regulate the expression of FXR, SHP-1, SHP-2, MREP2, BSEP and NTCP, down-regulate the expression of CYP7A1 and CYP27A1, suggesting that the drug plays a role by regulating related proteins. SA has obvious intervention and regulation effect on ANIT-induced intrahe patic cholestasis in young SD rats, with possible therapeutic function by participating in the transport and synthesis of bile acids.
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<p><b>OBJECTIVE</b>To investigate the changes of tumor necrosis factor-α (TNF-α) and transforming growth factor-β (TGF-β) in mice with cholestatic cirrhosis and their role in regulating the balance of liver stem cell differentiation.</p><p><b>METHODS</b>Balb/c mice were subjected to bile duct ligation (BDL), and serum biochemical parameters were measured and hepatic histopathology was observed using HE staining to evaluate the modeling of cholestatic cirrhosis. Immunohistochemistry and Western blotting were used to detect the changes of TNF-α and TGF-β in the mice after modeling. Mouse embryonic hepatic stem cells (HP14-19) were treated with different concentrations of TNF-α and TGF-β, and the cell differentiation was assessed using Western blotting, real-time PCR, and PAS staining.</p><p><b>RESULTS</b>The mice receiving BDL showed significantly increased blood biochemical parameters (P<0.05), and HE staining revealed obviously increased collagen fibers in the liver with significantly increased expressions of TNF-α and TGF-β (P<0.05). In HP14-19 cells, induction with TNF-α and TGF-β for 3 days did not cause significant changes in cell differentiation, but induction for 5 days resulted in significantly increases intensity of PAS staining in the cells. The cells induced with 20, 40, and 80 ng/mL TNF-α for 5 days exhibited a significantly stronger expression of cytokeratin 18 than cytokeratin 19 (P<0.05), while induction with 20, 40, and 80 ng/mL TGF-β produced opposite changes in cytokeratin 18 and cytokeratin 19 expressions. Further induction of the cells with TNF-α and TGF-β for 10 days, did not alter the expression patterns of cytokeratin 18 and cytokeratin 19 observed on day 5, but their protein expression levels and PAS staining intensity of the cells were enhanced and their mRNA expressions became lowered.</p><p><b>CONCLUSION</b>Common bile duct ligation can induce conditions simulating cholestatic cirrhosis in mice. TNF-α and TGF-β are elevated in cholestatic cirrhosis and play opposite roles in regulating the differentiation balance of liver stem cells: the former promotes the differentiation of liver stem cells into hepatocytes, while the latter promotes the cell differentiation into colangiocytes.</p>
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Objective To study the clinical characteristics of immune tolerance after liver transplantation in children and to identify possible predictors.Methods The clinical data of 37 pediatric patients who underwent liver transplantation between April 2006 and April 2014 at the Children's Hospital of Chongqing Medical University were retrospectively analyzed.The patients were divided into the no-drug (n =4),single-drug (n =16) and multi-drug (n =17) groups according to the status of their current immunosuppressant medications.The possible predictive factors were screened based on their clinical data,and statistical analysis was performed.Results The 37 liver transplantation recipients included 16 males (43.2％) and 21 females (56.8％).The factors that differed among the groups included age at transplantation and the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) of the transplant recipients.Age,ALT level,and AST level of the transplant recipients were significantly different between the single-drug group and the multi-drug group (all P ＜ 0.05).However,only the ALT Ievel was significantly different (P ＜ 0.05) between the no-drug group and the multi-drug group.No significant differences were found in the various other factors between the no-drug and single-drug groups.Conclusion The age of the recipient at transplantation was a predictive factor affecting clinical immune tolerance in pediatric liver transplantation,while ALT and AST levels were potential predictors of postoperative immune tolerance.
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<p><b>OBJECTIVE</b>To construct a recombinant adenovirus co-expressing bone morphogenic protein (BMP) 9 and BMP6 and observe its effect on the osteogenesis in C3H10 cells.</p><p><b>METHOD</b>The full-length sequences of BMP9 and BMP6 were amplified from AdEasy vector by PCR and cloned into the shuttle plasmid pASG2 vector to construct the co-expression shuttle plasmid pASG2-BMP9, 6 followed by homologous recombination with plasmid pAdeasy-1 in BJ5183. After confirmation by restriction endonuclease digestion, the recombinant vector was transfected into HEK293 cells, and high-titer recombinant adenovirus (Ad-BMP9, 6) was collected after amplification. Ad-BMP9, 6 was then transduced into C3H10 cells in vitro, and the mRNA expression of BMP9 and BMP6 was detected by RT-PCR. The osteogenic capability of the transfected cells was observed by alkaline phosphatase staining and calcium-alizarin red staining.</p><p><b>RESULTS</b>AdBMP9,6 was constructed successfully and effectively infected in C3H10 cells, in which high expressions of BMP6 and BMP9 were detected. C3H10 cells infected with Ad-BMP9,6 showed stronger alkaline phosphatase and calcium-alizarin red staining than the cells transfected by either BMP9 or BMP6 alone.</p><p><b>CONCLUSION</b>The recombinant adenovirus co-expressing BMP9 and BMP6 we constructed shows a more potent effect than the adenoviruses expressing either BMP9 or BMP6 alone in inducing the osteogenic differentiation of C3H10 cells into osteoblasts.</p>
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Humans , Adenoviridae , Genetics , Bone Morphogenetic Protein 6 , Genetics , Genetic Vectors , Growth Differentiation Factors , Genetics , HEK293 Cells , Osteoblasts , Cell Biology , Osteogenesis , Plasmids , Recombinant Fusion Proteins , Genetics , TransfectionABSTRACT
Objective To summarize the clinical experience of successful liver transplantation from infant donation after cardiac death (DCD) for infant with biliary astresia (BA).Methods The donor was a 16-months-old girl with a body weight of 10 kg,who died of irreversible anoxic cerebral damage after sudden asphyxiation.The recipient was a 24-months-old girl with a body weight of 12 kg,who suffered from icteric concurrent late biliary cirrhosis after the Porta-jejunum anastomosis because of congenital BA.The DCD liver was classically orthotopically transplanted into the infants recipient.The warm ischemia time was 7 min,the cold ischemia time was 360 min,and the graft volume to the standard liver volume (GV/SLV) was 1.02.After operation,the vital signs and transplanted liver function of the recipient were monitored,and the recipient was given treatments of anti-infection,anticoagulation,and improving the microcirculation.The recipient was treated with the triple immunosuppression protocol of tacrolimus,mycophenolate and prednisone to prevent rejection.Results The operating time of the recipient was 480 min,the non-liver stage was 65 min,and the blood loss was 230 mL.The endotracheal intubation was removed from the recipient at 12 h,and the recipient started to eat at 48 h aftcr operation.The recipient had a hepatic artery thrombus on the 3rd and 15th day after operation,and the hepatic artery had re-blood-supply after the hepatic artery catheterization and continuous perfusion with urokinase.The recipient was discharged on the 42nd day,and the recipient was in satisfactory condition to present.Conclusion The infant DCD liver is a better graft for infant liver transplantation for BA.The surgical complications can be reduced with matched volume of donor-recipient liver; and it can guarantee a successful operation with perfect operative technique and careful perioperative management.
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<p><b>OBJECTIVE</b>To explore the expression of connective tissue growth factor (CTGF) in pancreatic cancer and its influence on the proliferation and migration of cancer cells.</p><p><b>METHODS</b>The expression of CTGF in pancreatic cell line PANC-1 cells was analyzed by real-time PCR and in pancreatic carcinoma (50 cases) tissues by immunohistochemistry. The ability of proliferation and migration in vitro of PANC-1 cells was tested by MTT assay, scratch test and Boyden chamber test after the CTGF gene was overexpressed by Ad5-CTGF or silenced with Ad5-siCTGF transfection.</p><p><b>RESULTS</b>CTGF was overexpressed in both pancreatic cancer cells and tissues. Overxpression of CTGF leads to increased proliferation and migration of PANC-1 cells. The CTGF-transfected PANC-1 cells showed apparent stronger proliferation ability and scratch-repair ability than that of empty vector controls. The results of Boyden chamber test showed that there were 34 cells/field (200× magnificantion) of the CTGF-transfected overexpressing cells, much more than the 11 cells/field of the empty vector control cells; and 6 cells/microscopic field of the Ad5-siCTGF-transfected silenced cells, much less than the 15 cells/field of the control cells.</p><p><b>CONCLUSIONS</b>CTGF is overexpressed in both pancreatic cancer cells in vitro and in vivo, indicating that it may play an important role in the cell proliferation and migration in pancreatic cancer.</p>
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Humans , Adenoviridae , Genetics , Cell Line, Tumor , Cell Movement , Cell Proliferation , Connective Tissue Growth Factor , Genetics , Metabolism , Pancreatic Neoplasms , Metabolism , Pathology , Recombinant Proteins , Genetics , Metabolism , TransfectionABSTRACT
Objective To investigate the cause of jejunum perforation after infantile livingrelated liver transplantation (ILRLT) and summarize the experience of treatment. Methods The clinical data of 28 infants with biliary atresia who underwent ILRLT were analyzed and 4 of 28 infantile recipients (14. 3%) developed jejunum perforation after ILDLT. Results Four patients had 7 episodes of jejunum perforation after transplantation among 28 infantile recipients who underwent ILRLT because of biliary atresia. The median time between transplantation and perforation was 11 days.Perforation occurred at the point of silk in jejunum stoma (n = 3) and the Roux-en-Y limb (n = 1 ).None had a history of prior operation including Kasai in 4 patients. Clinical manifestation included fever, increased heart rate, abdominal distention, leukocytosis, and no free air on abdominal roentgenograrns. A simple repair was performed in three infants with silk: two developed recurrent perforation (67%) and underwent a re-exploration,and another had a third perforation and underwent a third repair because of re-perforation. Another child underwent a simple repair with prolene, and there was no recurrence. None died from the perforation in our study. Conclusion The occurrence and location of jejunum perforation after ILDLT suggests that the cause of the perforation is related to the jejunal anastomosis with silk, and the jejunum perforation may be avoided in the jejunal anastomosis with prolene. Early diagnosis and exploration may ensure better survival.
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<p><b>OBJECTIVE</b>To establish a novel Myc gene transgenic mouse model for spontaneously forming B-lymphoma and assessing its tumorigenesis potential.</p><p><b>METHODS</b>Freshly isolated hematopoietic progenitor cells served as the target for Myc gene transfer mediated by a retrovirus vector. These cells were engrafted into C57BL/6 mice with (60)Co-gamma ray radiation in advance. Tumor latency was measured and the tumor loaded mice were followed for survival time. Tumor was identified with histology and immunostaining. The exogenous Myc gene was detected by Western blot (in liver, spleen, tumor tissue) and flow cytometry (FCM) \[in bone marrow (BM)\].</p><p><b>RESULTS</b>Mice BM-infected with mutant Myc gene more readily gave rise to B-cell lymphomas than those infected with wild type Myc gene did Myc gene was expressed highly in BM and tumor tissues but not in liver and spleen.</p><p><b>CONCLUSION</b>Our model will be a tool in assessing the transforming potential of Myc mutants and in studying cooperation between Myc and other oncogenes. Mutant Myc is more effective than wild-type Myc in promoting B cell lymphomagenesis in mice.</p>
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Animals , Mice , B-Lymphocytes , Cell Transformation, Neoplastic , Flow Cytometry , Lymphoma , Lymphoma, B-Cell , Mice, Inbred C57BL , Mice, Transgenic , Retroviridae InfectionsABSTRACT
Objective:To design and construct the replication-deficient recombinant adenovirus Ad-siCTGF which can silence the expression of connective tissue growth factor (CTGF) by RNA interference and verified its function. Methods:A specific sequence, which was verified to be able to silence CTGF gene with high efficiency, was cloned into pSES-HUS vector to produce the shuttle plasmid pSES-siCTGF. The plasmid after Pme Ⅰ linearization was cotransduced with pAdEasy into BJ5183 E.coli strains to construct recombinant vector Ad-siCTGF. After linearization treatment with Pac Ⅰ enzyme digestion Ad-siCTGF was transfected into HEK293 cells via liposome mediation. The recombinant adenovirus was packaged. The titer of the Ad-siCTGF was increased after three times of cross-infection. 4T1 cells were infected with the adenovirus. The silencing efficiency was tested by real-time fluorescence quantitative (RFQ)-PCR and Western blotting.Results:Pac Ⅰ enzyme digestion electrophoresis indentified that recombinant adenovirus was successfully constructed. The titer of the recombinant adenovirus Ad-siCTGF was 2.6×10~(10) pfu/mL after amplification and purification. The CTGF mRNA and protein expression in 4T1 cells were decreased by 36.27% and 31.56%, respectively, compared with the control groups.Conclusion:The recombinant adenovirus which can silence the expression of CTGF was successfully constructed. It laid a good foundation for further investigation of the action mechanism of CTGF in tumor cells.
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<p><b>OBJECTIVE</b>To summarize our experience in adult-to-infant living donor liver transplantation (A-ILDLT) and to analyze the efficacy and complications of A-ILDLT.</p><p><b>METHODS</b>The clinical data, surgical strategies and complications of 28 adult donors and infantile recipients who underwent A-ILDLT from April 2006 to December 2009 were retrospectively analyzed. These 28 patients (14 boys and 14 girls) aged from 80 days to 11.5 months with body weights of 3.08 to 10.3 kg at the time of operation . They suffered from biliary atresia with decompensated cirrhosis. The living donors were 15 mothers, 9 fathers, 3 grandma and 1 elder brother with ABO compatible with the infantile recipients. 27 Donor organs were the left lateral lobe grafts (segment II, III) and 1 graft was segment II. All patients were followed up for 5 to 24 months.</p><p><b>RESULTS</b>These grafts were orthotopically transplanted into the infantile recipients. The average length of stay was 9.3 days for the donor group without any complications. Postoperative immunosuppression included prednisone, Cyclosporin and mycophenolate mofetil (MMF). A total of 24 postoperative complications occurred in 20 recipients, including 5 vascular complications, 4 bleeding, 7 pneumonia, 2 bowel obstruction, 4 intestinal perforation and 3 rejection. Three recipients died of hepatic arterial thrombosis (HAT). The perioperative mortality rate of recipients was 10.7% (3/28) and the survival rate was 89.3% in peroperative period. One died of stricture of hepatic vein and 1 of accidental asphyxia during follow-up term. At present, 23 cases are still alive.</p><p><b>CONCLUSION</b>A-ILDLT has become an effective method to infants with end-stage liver disease. The postoperative vascular complication is the predominant cause of death.</p>
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Female , Humans , Infant , Male , Liver Diseases , General Surgery , Liver Transplantation , Methods , Living Donors , Retrospective Studies , Treatment OutcomeABSTRACT
Objective To summarize the clinical experience of segmental living related liver transplantation for very small infant with biliary atresia. Methods The recipient was a 145-day-old male with congenital biliary atresia. The infant was 66 cm in height and weighed 3.08 kg. The donor was his 36-year-old mother. Her segment Ⅱ of the liver was excised and orthotopically transplanted into the infant's body as the graft. The portal vein of the graft was end-to-end anastomosed to the portal vein of the recipient, the hepatic artery of the graft was end-to-end anastomosed to the proper hepatic artery of the recipient with lateral superficial vein of left great saphenous vein from donor as a bridge, and the hepatic vein was end-to-end anastomosed to the hepatic vein of the recipient whose hepatic vein was conformed from right, middle and left hepatic vein. Biliary tract was reconstructed via Roux-en-Y operation. Results Segment Ⅱ (160 g) of liver from donor was resected, and there was no blood infusion. The donor retained her liver function within 5 days and was discharged on the eighth day. The operating time of graft implantation was 451 min. The blood loss was 250 ml. Non-liver stage was 71 min. The cold ischemic time was 132 min. Cyclosporine, mycophenolate mofetil (MMF) and prednisone were used for postoperative immunosuppression. The bilirubin level of the infant was decreased to the normal level one week after operation, and the liver function became normal in 9 days. Jejuno-leakage on the 7th day after the transplantation was recovered by mend and drainage and discharged on the 35th day. The donor and recipient were in satisfactory condition to present. Conclusion The segmental living related liver transplantation is advisable for very small infant with biliary atresia. Perfect operative technique and postoperative intensive care are the keys to ensure the success of the procedure.
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<p><b>OBJECTIVE</b>To explore the risk factors for hepatoblastoma.</p><p><b>METHODS</b>A case-cohort study using Logistic regression multiple variables analysis of medical record data sets was conducted to examine infant and perinatal risk factors for hepatoblastoma.</p><p><b>RESULTS</b>Birth weight less than 1,000 g was associated with a strongly increased risk of hepatoblastoma (odds risk, OR = 26.0, 95% confidence interval, CI: 14.0 to 65.7). After adjustment of birth weight, a moderately increased risk of hepatoblastoma was found for older maternal age ( > 35 years vs. 20 to 34 years: OR = 2.6, 95% CI: 0.9 to 5.9), maternal smoking (OR = 2.9, 95% CI: 1.1 to 4.2) and higher maternal pregnancy body mass index (OR = 3.2, 95% CI: 1.0 to 6.7).</p><p><b>CONCLUSION</b>Very low birth weight and maternal characteristics including overweight, smoking are associated with hepatoblastoma risk.</p>
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Child , Child, Preschool , Female , Humans , Infant , Infant, Newborn , Male , Pregnancy , Case-Control Studies , Confidence Intervals , Follow-Up Studies , Hepatoblastoma , Epidemiology , Infant, Very Low Birth Weight , Liver Neoplasms , Epidemiology , Overweight , Retrospective Studies , Risk Factors , SmokingABSTRACT
<p><b>OBJECTIVE</b>To review the outcomes of living-related liver transplantation (LRLT) in treating 3 cases of cavernous transformation of portal vein (CTPV) with severe portal hypertension.</p><p><b>METHODS</b>Three children (two boys and one girl) were presented to our hospital with recurring esophageal variceal bleeding, decompensating ascites, splenomegaly and refractory anemia. CTPV was confirmed by intravenous computed tomographic portography using a helical computed tomography scanner and 3-dimensional image reconstruction. LRLT were performed in these 3 patients from July 2006 to January 2007. The evaluation of the outcomes was made by referring to their clinical features and laboratory and imaging examination findings.</p><p><b>RESULTS</b>Although one patient died from early graft thrombosis, the other two patients showed excellent prognoses. They lived and stayed well during a follow-up period of 12-14 months. Following the transplantations, there had been no esophageal variceal hemorrhage, the ascites disappeared and the portal hypertension vanished. Their hemoglobin, blood platelets count, and serum albumin reached normal values.</p><p><b>CONCLUSION</b>LRLT is an effective procedure in treating CTPV with severe portal hypertension. The reconstruction of the portal vein is the difficult part of the LRLT procedure.</p>
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Child , Female , Humans , Male , Hypertension, Portal , Pathology , General Surgery , Liver Transplantation , Living Donors , Parents , Portal Vein , Pathology , Treatment OutcomeABSTRACT
<p><b>OBJECTIVE</b>To explore the myelo-protection effect of mdr1 transfected cord blood cells (CBMNCs) graft against high-dose homoharringtonine leukemia-bearing severe combined immunodeficient (SCID) mice model.</p><p><b>METHODS</b>Multidrug resistant (mdr1)gene was transferred into CBMNCs by a retrovirus vector, containing full-length cDNA of human mdr1 gene. CBMNCs and high-titer retrovirus supernatant were cocultured with cytokine combinations for 5 - 6 days. The SCID mouse models bearing human HL-60 cell leukemia were divided into three groups. Group A received tail vein injection of 2 x 10(6) mdr1 gene transduced CBMNCs at day 1 and 3, groups B and C 2 x 10(6) un-transduced CBMNCs and same volume of normal saline, respectively. The 3 groups of the mouse model were treated with weekly escalated doses of homoharringtonine. The peripheral white blood cell (WBC) counts, the human leukemia cells percentage in peripheral blood, the histological findings of main organs were assayed. The CD33 positive HL-60 cells in bone marrow were determined by flow cytometry. The function and expression of mdr1 gene were examined by PCR, immunochemistry (IC) and DNR extrusion test in vivo.</p><p><b>RESULTS</b>(1) mdr1 gene was transferred into CBMNCs successfully and the transfection frequency was 30%. (2) Leukemia SCID mice were xenotransplanted with mdr1-transfected BMMNCs by a programmed procedure and could be used as a valuable model for in vivo evaluating myelo-protection effects. (3) The transfected mice could tolerate homoharringtonine 5 approximately 6 folds higher than conventional dose and kept peripheral WBC count at a mean of 3 x 10(9)/L, with the peripheral human myeloid leukemia cells percentage decreasing to less than 5%. Histological examination showed that there was no leukemia infiltration in the main organs, the CD33 positive HL-60 cells in bone marrow were less than 5%. (4) The repopulation frequency of the transfected CBMNs in marrow were 9.13%. DNR extrusion test confirmed that the P-gp product maintained its biological function in the marrow.</p><p><b>CONCLUSION</b>mdr1 transferred-human CBMNC can xenotransplanted and repopulated in leukemia-bearing SCID mouse and are protected from chemotherapy-induced myelosuppression.</p>
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Animals , Female , Humans , Male , Mice , ATP Binding Cassette Transporter, Subfamily B, Member 1 , Genetics , Metabolism , Antineoplastic Agents, Phytogenic , Therapeutic Uses , Cord Blood Stem Cell Transplantation , Methods , Fetal Blood , Cell Biology , Genetic Vectors , HL-60 Cells , Harringtonines , Therapeutic Uses , Leukemia, Promyelocytic, Acute , Drug Therapy , Pathology , General Surgery , Leukocytes, Mononuclear , Cell Biology , Metabolism , Transplantation , Mice, SCID , Random Allocation , Retroviridae , Genetics , Transfection , Treatment Outcome , Xenograft Model Antitumor AssaysABSTRACT
Aim To investigate the relationship between the antitumor activity of berberine and the inhibition of Wnt/?-catenin signaling pathway. Methods The antitumor activities of berberine and total alkaloids of Coptis were assessed for their ability to inhibit the proliferation and to induce apopotosis of human colon cancer lines. A well-established Tcf-4 responsive reporter was introduced into human colon cancer lines to test whether berberine and total alkaloids inhibited the ?-catenin/Tcf4 activity in these cancer cells.Results The proliferation of HCT116 and SW480 cells was inhibited substantially within the concentration ranges of 5 to 40 mg?L~(1) of berberine. 72 hours after treament both HCT116 and SW480 cells underwent apparent apopotosis as demonstrated by Hoechst 33258 staining. Total alkaloids of Coptis had similar effects to berberine when comparable concentrations of it with berberine were used.Berberine effectively and specifically inhibited the Tcf4 reporter activity, but not the positive control reporter at the same experimental conditions. Conclusion Our findings suggest that berberine may be mainly responsible for the antitumor activity of total alkaloids of Coptis and this effect may be mediated at least in part by its inhibitory effect on the Wnt/?-catenin signaling pathway, which is aberrantly activated in many types of human cancer.
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<p><b>OBJECTIVE</b>To investigate the expression of mdr1 gene in hematopoietic cells of a murine bone marrow transplantation model and to explore the feasibility of transferring mdr1 gene into hematopoietic cells to pro-vent myelosuppression from chemotherapy.</p><p><b>METHODS</b>mdr1 gene was transferred into hematopoietic cells of murine bone marrow by retrovirus vector. The expression and function of mdr1 gene in vivo was tested in a murine bone marrow transplantation model.</p><p><b>RESULTS</b>(1) mdr1 gene was successfully transferred into murine MNC, the transduction rate was 35%. (2) mdr1 gene transferred murine bone marrow transplantation model was established successfully by scheduled-bone marrow transplantation method. (3) In 1 approximately 5 months period, stable and effective expression of mdr1 gene could be detected in hematopoietic cells of the recipient mouse, the percentage of mdr1 gene expression cells in recipient's hematopoietic cells decreased monthly to 8.0%, 8.0%, 7.5%, 4.0% and 3.0%. (4) mdr1 expression hematopoietic cells had efficient resistance to chemotherapy.</p><p><b>CONCLUSION</b>It is an effective approach to transfer mdr1 gene into hematopoietic cells for preventing myelosuppression in chemotherapy.</p>