P-450-dependent epoxygenase pathway of arachidonic acid and the products of epoxyeicosatrienoic acids(EETs) have been demonstrated to be involved in angiogenesis and tumor progression.This study examined the expressio...P-450-dependent epoxygenase pathway of arachidonic acid and the products of epoxyeicosatrienoic acids(EETs) have been demonstrated to be involved in angiogenesis and tumor progression.This study examined the expression of EETs and the role of the pathway in the angiogenesis of multiple myeloma(MM).MM cell lines of U266 and RPMI8226 were cultured,and the EETs levels(11,12-EET and 14,15-EET) in the supernatant were determined by ELISA.Human umbilical vein endothelial cells(HUVECs) were cultured and used for analysis of the angiogenesis activity of the two MM cell lines,which was examined both in vitro and in vivo by employing MTT,chemotaxis,tube formation and matrigel plug assays.11,12-EET and 14,15-EET were found in the supernatant of the cultured MM cells.The levels of the two EETs in the supernatant of U266 cells were significantly higher than those in the RPMI8226 cell supernatant(P<0.05),and the levels paralleled the respective angiogenesis activity of the two different MM cell lines.17-octadecynoic acid(17-ODYA),as a specific inhibitor of P450 enzyme,suppressed HUVECs proliferation and tube formation induced by MM cells.Furthermore,17-ODYA decreased the EET levels in the supernatant of MM cells.These results suggest that EETs may play an important role in the angiogenesis of MM,and the inhibitor 17-ODYA suppresses this effect.展开更多
AIM: To assess whether the polymorphisms of NOD2/ CARD15 , autophagy-related 16-like 1 (ATG16L1 ), and interleukin-23 receptor (IL23R ) genes play a more critical role in the susceptibility of childhood-onset than in ...AIM: To assess whether the polymorphisms of NOD2/ CARD15 , autophagy-related 16-like 1 (ATG16L1 ), and interleukin-23 receptor (IL23R ) genes play a more critical role in the susceptibility of childhood-onset than in adult-onset Crohn’s disease (CD). METHODS: Polymorphisms R702W, G908R, and 3020insC of NOD2/CARD15 ; rs2241880 A/G of ATG16L1 , and rs11209026 (R381Q) of IL23R gene were assessed in 110 childhood-onset CD, 364 adult-onset CD, and 539 healthy individuals. Analysis of polymorphisms R702W, G908R, and 3020insC of NOD2/CARD15 genotyping was performed by allele specific polymerase chain reaction (PCR) or by PCR-restriction fragment length polymor-phism analysis. The polymorphisms rs2241880 A/G of the ATG16L1 , and rs11209026 (R381Q) of the IL23R gene in the children’s cohort were genotyped by PCR and melting curve analysis whereas adult group genotyping was performed using the Affymetrix Genome-Wide Human SNP Array 5.0 (500K). RESULTS: The 3020insC allele in NOD2/CARD15 was significantly higher in childhood than in adult-onset CD (P = 0.0067). Association with at least 1 NOD2/CARD15 variant was specific for ileal disease (with or without co- lonic involvement). Even if the frequency of G allele of the rs2241880 ATG16L1 polymorphism was increased in both paediatric and adult CD patients compared to con- trols (P = 0.017 and P = 0.001, respectively), no difference was observed between the childhood and the adult cohort. The rare Q allele of IL23R rs11209026 polymorphism was underrepresented in both paediatric and adult CD cases (P = 0.0018 and P = 0.04, respectively) and no difference was observed between the childhood and the adult cohort. The presence of the rs2241880 ATG16L1 and rs11209026 IL23R polymorphisms did not influence disease phenotype. CONCLUSION: Polymorphism 3020insC in NOD2/ CARD15 occurs statistically significantly more often in patients with childhood-onset CD than in patients with adult-onset CD. The ATG16L1 and IL23R variants are associated with susceptibility to CD, but not earlyonset disease.展开更多
Arachidonic acid cytochrome P-450 (CYP) hydroxylase 4A isoforms, including 4A1, 4A2, 4A3 and 4A8 in the rat kidney, catalyze arachidonic acid to produce 19/20-Hydroxyeicosatetraenoic acids (20-HETE), a biologically ac...Arachidonic acid cytochrome P-450 (CYP) hydroxylase 4A isoforms, including 4A1, 4A2, 4A3 and 4A8 in the rat kidney, catalyze arachidonic acid to produce 19/20-Hydroxyeicosatetraenoic acids (20-HETE), a biologically active metabolite, which plays an important role in the regulation of blood pressure. However, controversial results have been reported regarding the exact role of 20-HETE on blood pressure. In the present study, we used recombinant adeno- associated viral vector (rAAV) to deliver CYP 4A1 cDNA and antisense 4A1 cDNA into Sprague-Dawley (SD) rats and spontaneously hypertensive rats (SHR), respectively, to investigate the effects of long-term modifications of blood pressure and the potential for gene therapy of hypertension. The mean systolic pressure increased by 14.2±2.5 mm Hg in rAAV·4A1-treated SD rats and decreased by 13.7±2.2 mm Hg in rAAV·anti4A1-treated SHR rats 5 weeks after the injection compared with controls and these changes in blood pressure were maintained until the experiments ended at 24 weeks. In 4A1 treated animals CYP4A was overexpressed in various tissues, but preferentially in the kidney at both mRNA and protein levels. In anti-4A1-treated SHR, CYP4A mRNA in various tissues was probed, especially in kidneys, but 4A1 protein expression was almost completely inhibited. These results suggest that arachidonic acid CYP hydroxy- lases contribute not only to the maintenance of normal blood pressure but also to the development of hypertension. rAAV-mediated anti4A administration strategy has the potential to be used as targeted gene therapy in human hyperten- sion by blocking expression of CYP 4A in kidneys.展开更多
Objective: To explore the role of telomerase activity detected in biopsy samples for evaluating the efficacy of lapa- roscopic radiofrequency ablation (RFA) therapy in patients with hepatocellular carcinoma (HCC) and ...Objective: To explore the role of telomerase activity detected in biopsy samples for evaluating the efficacy of lapa- roscopic radiofrequency ablation (RFA) therapy in patients with hepatocellular carcinoma (HCC) and liver cirrhosis. Methods: From August 2001 to October 2004, 34 cirrhotic patients with HCC were treated by laparoscopic RFA under general anesthe- sia. A total of 34 tumors, with a mean maximum tumor diameter of 4.0 ± 1.0 cm, were all located on the liver surface or adja- cent to the gallbladder. Laparoscopic ultrasound-guided core biopsy for liver lesions was performed before and immediately after RFA therapy. In these biopsy samples, telomerase activity was detected by the ELISA-based telomeric repeat amplifica- tion protocol (ELISA-TRAP) assay, and pathological examination was routinely performed. Results: Laparoscopic RFA was successfully performed in all the 34 patients. A complete tumor necrosis was achieved in all patients on the contrast-enhanced helical CT scanning one month after laparoscopic RFA. The positive rates of telomerase activity and histopathologic diagnosis in biopsy samples were 91.2% (31/34) and 100% (34/34) respectively before RFA, and 26.5% (9/34) and 0% respectively after RFA. During a median follow-up period of 35 months (range, 18–51 months), the rates of local tumor recurrence at the ablation sites in post-RFA telomerase-positive and negative patients were 88.9% (8/9) and 4% (1/25) respectively (P < 0.01), and the rates of distant recurrence within the livers were 0% (0/9) and 12% (3/25) respectively (P > 0.05). Conclusion: For cirrhotic patients with HCC treated by laparoscopic RFA, detection of telomerase activity in biopsy samples may be useful for evaluating the therapeutic efficacy of RFA and predicting postoperative local tumor recurrence.展开更多
Combined hepatocellular-cholangiocarcinoma(CHC) is a mixed tumor containing elements of both hepatocellular carcinoma(HCC) and cholangiocarcinoma(CC). Its remarkable histological heterogeneity has been linked to putat...Combined hepatocellular-cholangiocarcinoma(CHC) is a mixed tumor containing elements of both hepatocellular carcinoma(HCC) and cholangiocarcinoma(CC). Its remarkable histological heterogeneity has been linked to putative hepatic progenitor cell(HPC) origin. However, detailed histological or phenotypic description is rarely documented. In the present study, we reassessed 68 cases previously diagnosed as hepatitis B-related CHCs by immunohistochemistry and double-fluorescence immunostaining, focusing on HPC associated phenotypic observation of intermediate area of the tumor. It was found that tumor cells showed remarkable heterogeneity in intermediate area. Tumor cells with intermediate morphology between hepatocytes and cholangiocytes were oval-shaped and small with scant cytoplasm and hyperchromatic nuclei, arranging in solid nests mostly. By Keratin 7(K7) staining, it appeared that the nests of tumor cells represented a maturation process from the undifferentiated small cells to mature hepatocytes through the "transitional" cells. Then, these small cells were further confirmed with intermediate phenotype as HPC by exploring immature hepatocellular marker and HPC/biliary markers co-localization. In conclusion, the HPC associated trait in CHC can be interpreted by HPC origin or gain of "stemness" by dedifferentiation. It is still too soon to give a final word that it is innate or acquired signature of HPC associated trait in CHC.展开更多
Severely immunocompromised NOD.Cg-PrkdcIl2rg(NOG)mice are among the ideal animal recipients for generation of human cancer models.Transplantation of human solid tumors having abundant tumor-i nfiltrating lymphocytes(T...Severely immunocompromised NOD.Cg-PrkdcIl2rg(NOG)mice are among the ideal animal recipients for generation of human cancer models.Transplantation of human solid tumors having abundant tumor-i nfiltrating lymphocytes(TILs)can induce xenogeneic graft-versus-host disease(xGvHD)following engraftment and expansion of the TILs inside the animal body.Wilms’tumor(WT)has not been recognized as a lymphocyte-predominant tumor.However,3 consecutive generations of NOG mice bearing WT patient-derived xenografts(PDX)xenotransplanted from a single donor showed different degrees of inflammatory symptoms after transplantation before any therapeutic intervention.In the initial generation,dermatitis,auto-amputation of digits,weight loss,lymphadenopathy,hepatitis,and interstitial pneumonitis were observed.Despite antibiotic treatment,no response was noticed,and thus the animals were prematurely euthanized(day 47 posttransplantation).Laboratory and histopathologic evaluations revealed lymphoid infiltrates positively immunostained with anti-human CD3 and CD8 antibodies in the xenografts and primary tumor,whereas no microbial infection or lymphoproliferative disorder was found.Mice of the next generation that lived longer(91 days)developed sclerotic skin changes and more severe pneumonitis.Cutaneous symptoms were milder in the last generation.The xenografts of the last 2 generations also contained TILs,and lacked lymphoproliferative transformation.The systemic immunoinflammatory syndrome in the absence of microbial infection and posttransplant lymphoproliferative disorder was suggestive of xGvHD.While there are few reports of xGvHD in severely immunodeficient mice xenotransplanted from lymphodominant tumor xenografts,this report for the first time documented serial xGvHD in consecutive passages of WT PDX-bearing models and discussed potential solutions to prevent such an undesired complication.展开更多
The field of gene therapy has been increasingly studied in the last four decades, and its clinical application has become a reality in the last 15 years. Traditional Chinese medicine(TCM), an important component of co...The field of gene therapy has been increasingly studied in the last four decades, and its clinical application has become a reality in the last 15 years. Traditional Chinese medicine(TCM), an important component of complementary and alternative medicine, has evolved over thousands of years with its own unique system of theories, diagnostics and therapies. TCM is well-known for its various roles in preventing and treating infectious and chronic diseases, and its usage in other modern clinical practice. However, whether TCM can be applied alongside gene therapy is a topic that has not been systematically examined. Here we provide an overview of TCM theories in relation to gene therapy. We believe that TCM theories are congruent with some principles of gene therapy. TCM-derived drugs may also act as gene therapy vehicles, therapeutic genes, synergistic therapeutic treatments, and as co-administrated drugs to reduce side effects. We also discuss in this review some possible approaches to combine TCM and gene therapy.展开更多
AIM:To investigate the effect of the demethylating reagent 5-aza-2'-deoxycitidine(DAC) on telomerase activity in hepatocellular carcinoma(HCC) cell lines,SMMC-7721 and HepG2.METHODS:The related gene expression in ...AIM:To investigate the effect of the demethylating reagent 5-aza-2'-deoxycitidine(DAC) on telomerase activity in hepatocellular carcinoma(HCC) cell lines,SMMC-7721 and HepG2.METHODS:The related gene expression in cell lines was examined by real-time reverse transcription-polymerase chain reaction and Western blotting analysis.The telomerase activity was examined by telomeric repeat amplification protocol-enzyme-linked immunosorbent assay and DNA methylation was determined by methylation-specific polymerase chain reaction.RESULTS:The telomerase activity was significantly reduced in both cell lines treated with DAC,accompanied by downregulation of telomerase reverse transcriptase(hTERT).We also observed the effect of DAC on the methylation status of hTERT promoter and the expression of regulatory genes,such as c-myc,p15,p16,p21,E2F1,and WT1.The methylation status of hTERT promoter could be reversed in SMMC-7721 by DAC,but not in HepG2 cells.However,p16 expression could be reactivated by demethylation of its promoter,and c-Myc expression was repressed in both cell lines.Moreover,DAC could enhance the sensitivity to the chemotherapeutic agents,such as cisplatin,by induction of apoptosis of HCC cells.CONCLUSION:The DAC exerts its anti-tumor effects in HCC cells by inhibiting the telomerase activity.展开更多
Hepatocellular carcinoma(HCC) is the third leading cause of cancer-related deaths worldwide. Only 30%-40% of the patients with HCC are eligible for curative treatments, which include surgical resection as the first op...Hepatocellular carcinoma(HCC) is the third leading cause of cancer-related deaths worldwide. Only 30%-40% of the patients with HCC are eligible for curative treatments, which include surgical resection as the first option, liver transplantation and percutaneous ablation. Unfortunately, there is a high frequency of tumor recurrence after surgical resection and most HCC seem resistant to conventional chemotherapy and radiotherapy. Sorafenib, a multi-tyrosine kinase inhibitor, is the only chemotherapeutic option for patients with advanced hepatocellular carcinoma. Patients treated with Sorafenib have a significant increase in overall survival of about three months. Therefore, there is an urgent need to develop alternative treatments. Due to its role in cell growth and development, the insulin-like growth factor system is commonly deregulated in many cancers. Indeed, the insulin-like growth factor(IGF) axis has recently emerged as a potential target for hepatocellular carcinoma treatment. To this aim, several inhibitors of the pathway have been developed suchas monoclonal antibodies, small molecules, antisense oligonucleotides or small interfering RNAs. However recent studies suggest that, unlike most tumors, HCC development requires increased signaling through insulin growth factor Ⅱ rather than insulin growth factor Ⅰ. This may have great implications in the future treatment of HCC. This review summarizes the role of the IGF axis in liver carcinogenesis and the current status of the strategies designed to target the IGF-Ⅰ signaling pathway for hepatocellular carcinoma treatment.展开更多
Throughout its 40-year history,the field of gene therapy has been marked by many transitions.It has seen great strides in combating human disease,has given hope to patients and families with limited treatment options,...Throughout its 40-year history,the field of gene therapy has been marked by many transitions.It has seen great strides in combating human disease,has given hope to patients and families with limited treatment options,but has also been subject to many setbacks.Treatment of patients with this class of investigational drugs has resulted in severe adverse effects and,even in rare cases,death.At the heart of this dichotomous field are the viral-based vectors,the delivery vehicles that have allowed researchers and clinicians to develop powerful drug platforms,and have radically changed the face of medicine.Within the past 5 years,the gene therapy field has seen a wave of drugs based on viral vectors that have gained regulatory approval that come in a variety of designs and purposes,these modalities range from vector-based cancer therapies,to treating monogenic diseases with life-altering outcomes.At present,the three key vector strategies are based on adenoviruses,adeno-associated viruses,and lentiviruses.They have led the way in preclinical and clinical successes in the past two decades.However,despite these successes,many challenges still limit these approaches from attaining their full potential.To review the viral vector-based gene therapy landscape,we focus on these three highly regarded vector platforms and describe mechanisms of action and their roles in treating human disease.展开更多
Objective: To explore the effects of thiazolidinediones(TZDs) pioglitazone on proliferation and differentiation of human preadipocytes. Methods:Omental adipose tissue biopsies were obtained from 15 patients who were u...Objective: To explore the effects of thiazolidinediones(TZDs) pioglitazone on proliferation and differentiation of human preadipocytes. Methods:Omental adipose tissue biopsies were obtained from 15 patients who were undergoing elective open-abdominal surgery. The primary culture and differentiated induction of human preadipocytes were performed, and the human preadipo-cytes were treated with pioglitazone at different concentrations at proper moments. Dynamic morphological changes of the human preadipocytes were observed, and their proliferation and differentiation were assessed with Colorimetric MTT Assay and Oil Red O Staining. Results:After 24 hours and 72 hours with pioglitazone, 0.1 μmol/L(μmol/ml) pioglitazone increased the MTT values of the human preadipocytes by 25.3% and 34.8%,respectively(P < 0.05), while 1 μmol/L pioglitazone by 27.4% and 26.6%(P < 0.05), compared with the control group without pioglitazone. The human preadipocytes with pioglitazone cumulated more adipose in the endochylema than those without pioglitazone obviously. 0.1 μmol/L pioglitazone increased the differentiation degree of the human preadipocytes differentiated for 8-10 days by 44.81% and 1 μmol/L pioglitazone by 53.76%(P < 0.05). Conclusion:Thiazolidinediones pioglitazone may significantly promote the proliferation and differentiation of the human omental preadipocytes.展开更多
Background:Bladder cancer poses a great burden on society and its high rate of recurrence and treatment failure necessitates use of appropriate animal models to study its pathogenesis and test novel treatments.Orthoto...Background:Bladder cancer poses a great burden on society and its high rate of recurrence and treatment failure necessitates use of appropriate animal models to study its pathogenesis and test novel treatments.Orthotopic models are superior to other types since they provide a normal microenvironment.Four methods are described for developing bladder cancer models inside the animal’s bladder.Direct intramural injection is one of these methods and is widely used.However,its efficacy in model development has not yet been studied.We aimed to evaluate the efficacy and success rate of the direct intramural injection method of developing an orthotopic model for the study of bladder cancer.Method:Tumor cell lines were prepared in four microtubes.Aliquots of 200×10^(3) cells were injected through a 27 gauge needle into the ventral wall of the bladders of 4male and 4 female BALB/c mice following a midline 1 cm laparotomy incision.In addition,1 million cells from each microtube were injected into the flanks of control mice.To prevent infection and alleviate pain,5 mg/kg enrofloxacin and 2.5 mg/kg flunixin meglumine,respectively,were injected subcutaneously.Results:Tumors formed in all mice,resulting in 100% take rate and zero post-operation mortality.Surgery time was≤15 min per mouse.In two mice,tumors were found in the peritoneal space as well.Conclusion:Direct intramural injection is a rapid,reliable,and reproducible method for developing orthotopic models of bladder cancer.It can be done on both male and female mice and only requires readily available surgical tools.However,needle track can result in cell spillage and peritoneal tumors.展开更多
Objective: To induce changes in biological character of human liver cancer cell line SMMC-7721 by blocking the expression of telomerase genes hTRT and to explore its value in cancer gene therapy. Methods: The vehicle ...Objective: To induce changes in biological character of human liver cancer cell line SMMC-7721 by blocking the expression of telomerase genes hTRT and to explore its value in cancer gene therapy. Methods: The vehicle for eukaryotic expression of antisense hTRT was constructed and then transfected into SMMC-7721 cells. The effects of antisense hTRT gene on telomerase activity, cancer cell growth and malignant phenotypes were analyzed. Results: The obtained transfectants that could express antisense hTRT gene stably showed marked decrease in telomerase activity; the shortening of telomere was obvious; cells presented contact growth inhibition; in nude mice transplantation, the rate of tumor induction dramatically decreased. Conclusion: Antisense hTRT gene expression can significantly inhibit telomerase activity of cancer cells and decrease malignant phenotypes in vitro and in vivo. Therefore, as a telomerase inhibitor, antisense hTRT gene may be a new pathway for cancer therapy.展开更多
AIM: To elucidate the biological function of HBV core antigen (HBcAg) on pathogenesis of hepatitis B, a novel gene C12 coding for protein with unknown function interacting with HBcAg in hepatocytes was identified and ...AIM: To elucidate the biological function of HBV core antigen (HBcAg) on pathogenesis of hepatitis B, a novel gene C12 coding for protein with unknown function interacting with HBcAg in hepatocytes was identified and characterized. METHODS: HBcAg bait plasmid pGBKT7-HBcAg was constructed and transformed into yeast AH109, then the transformed yeast was mated with yeast Y187 containing liver complementary DNA (cDNA) library plasmid in 2×YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade)containing X-α-gal for screening twice. After extracting and sequencing of plasmid from blue colonies, we isolated a cDNA clone encoding a novel protein designated as C12that directly interacted with HBcAg. The interaction between HBcAg and C12 was verified again by re-mating.pEGFP-N1-C12 fluorescent protein fusion gene was transfected in 293 and L02 cell, and observed by fluorescent microscope. MTT reduction assay was used to study the action of C12 protein effect on metabolism of mammal cell. Yeast two-hybrid and cDNA microarray were performed to search binding protein and differential expression genes regulated by C12 protein.RESULTS: C12 gene was screened and identified by yeast two-hybrid system 3. The interaction between HBcAg and the novel protein coded by the new gene C12 was further confirmed by re-mating. After 48 h, fluorescence of fusion protein could be observed steadily in the 293 and L02 cell plasma. Under MTT assay, we found that the expression of C12 did not influence the growth of liver cells. Seventeen differential expression genes in HepG2 cells transfected with C12 protein expression plasmid by cDNA microarray,of which 16 genes were upregulated and 1 gene was downregulated by C12 protein. Twenty-one colonies containing 16 different genes coding for C12 protein binding proteins were isolated by yeast two-hybrid, there were 2 new genes with unknown function.CONCLUSION: The novel protein C12 is located in cell plasma, and its overexpression has no significant effect on the metabolism of liver cell. It interacts with many proteins in hepatocytes and may be involved in many processes of gene expression.展开更多
LIM domain kinases(LIMKs),which modulate cytoskeletal dynamics,are found throughout the central nervous system.The synaptic junctions between neurons show structural alteration or plasticity during neurodevelopment,le...LIM domain kinases(LIMKs),which modulate cytoskeletal dynamics,are found throughout the central nervous system.The synaptic junctions between neurons show structural alteration or plasticity during neurodevelopment,learning and memory formation,or after injury and disease;in some cases completely new connections form,while others are lost.This remodeling is under the control of the actin cytoskeleton and therefore is influenced by the activity of LIMKs(Scott and Olson,2007).Recent reports discussed in this perspective shed light on the potential role of LIMK in hippocampal dendritic spines and photoreceptor presynaptic terminals and illustrate the importance of LIMK in nerve cell function.展开更多
BACKGROUND Metastatic skin cancers are relatively rare dermatological malignancies.They usually present as nodules,erythematous lesions,scar-like lesions or other lesion types.Signet-ring cell carcinoma(SRCC)is an unc...BACKGROUND Metastatic skin cancers are relatively rare dermatological malignancies.They usually present as nodules,erythematous lesions,scar-like lesions or other lesion types.Signet-ring cell carcinoma(SRCC)is an uncommon histological type of gastric cancer that usually behaves aggressively and has a poor prognosis.Skin metastasis may be the first sign of clinically silent visceral cancer or recurrence of an internal malignancy.CASE SUMMARY Herein we report on the case of a 55-year-old man with edema of a lower extremity as the primary symptom which progressed from local to generalized pitting edema in the year following skin involvement.Pathological evidence from gastroscopic specimens and subcutaneous tissue biopsy showed typical signetring cells and gland-like structures.Consistently,immunohistochemical analysis revealed positive pan-cytokeratin expression in tumor cells.A diagnosis of gastric SRCC with skin metastasis was established.Moreover,lymphoscintigraphy showed an obvious accumulation of radiotracer on the anterior and posterior sides of the right leg which indicated lymphedema.We reviewed the relevant literature on subcutaneous metastases of gastric SRCC.CONCLUSION This rare case emphasizes the importance of physical examination as it may help elucidate the etiology of edema.展开更多
Hepatic fibrosis/cirrhosis is a significant health burden worldwide,resulting in liver failure or hepatocellular carcinoma(HCC)and accounting for many deaths each year.The pathogenesis of hepatic fibrosis/cirrhosis is...Hepatic fibrosis/cirrhosis is a significant health burden worldwide,resulting in liver failure or hepatocellular carcinoma(HCC)and accounting for many deaths each year.The pathogenesis of hepatic fibrosis/cirrhosis is very complex,which makes treatment challenging.Endogenous mesenchymal stromal cells(MsCs)have been shown to play pivotal roles in the pathogenesis of hepatic fibrosis.Paradoxically,exogenous MsCs have also been used in clinical trials for liver cirrhosis,and their effectiveness has been observed in most completed clinical trials.There are still many issues to be resolved to promote the use of MsCs in the clinic in the future.In this review,we will examine the controversial role of MsCs in the pathogenesis and treatment of hepatic fibrosis/cirrhosis.We also investigated the clinical trials involving MsCs in liver cirrhosis,summarized the parameters that need to be standardized,and discussed how to promote the use of MsCs from a clinical perspective.展开更多
基金supported by grants from the National Natural Science Foundation of China(No.81071943)the Foundation of Natural Sciences of Hubei Province of China(No.2007ABA072)
文摘P-450-dependent epoxygenase pathway of arachidonic acid and the products of epoxyeicosatrienoic acids(EETs) have been demonstrated to be involved in angiogenesis and tumor progression.This study examined the expression of EETs and the role of the pathway in the angiogenesis of multiple myeloma(MM).MM cell lines of U266 and RPMI8226 were cultured,and the EETs levels(11,12-EET and 14,15-EET) in the supernatant were determined by ELISA.Human umbilical vein endothelial cells(HUVECs) were cultured and used for analysis of the angiogenesis activity of the two MM cell lines,which was examined both in vitro and in vivo by employing MTT,chemotaxis,tube formation and matrigel plug assays.11,12-EET and 14,15-EET were found in the supernatant of the cultured MM cells.The levels of the two EETs in the supernatant of U266 cells were significantly higher than those in the RPMI8226 cell supernatant(P<0.05),and the levels paralleled the respective angiogenesis activity of the two different MM cell lines.17-octadecynoic acid(17-ODYA),as a specific inhibitor of P450 enzyme,suppressed HUVECs proliferation and tube formation induced by MM cells.Furthermore,17-ODYA decreased the EET levels in the supernatant of MM cells.These results suggest that EETs may play an important role in the angiogenesis of MM,and the inhibitor 17-ODYA suppresses this effect.
文摘AIM: To assess whether the polymorphisms of NOD2/ CARD15 , autophagy-related 16-like 1 (ATG16L1 ), and interleukin-23 receptor (IL23R ) genes play a more critical role in the susceptibility of childhood-onset than in adult-onset Crohn’s disease (CD). METHODS: Polymorphisms R702W, G908R, and 3020insC of NOD2/CARD15 ; rs2241880 A/G of ATG16L1 , and rs11209026 (R381Q) of IL23R gene were assessed in 110 childhood-onset CD, 364 adult-onset CD, and 539 healthy individuals. Analysis of polymorphisms R702W, G908R, and 3020insC of NOD2/CARD15 genotyping was performed by allele specific polymerase chain reaction (PCR) or by PCR-restriction fragment length polymor-phism analysis. The polymorphisms rs2241880 A/G of the ATG16L1 , and rs11209026 (R381Q) of the IL23R gene in the children’s cohort were genotyped by PCR and melting curve analysis whereas adult group genotyping was performed using the Affymetrix Genome-Wide Human SNP Array 5.0 (500K). RESULTS: The 3020insC allele in NOD2/CARD15 was significantly higher in childhood than in adult-onset CD (P = 0.0067). Association with at least 1 NOD2/CARD15 variant was specific for ileal disease (with or without co- lonic involvement). Even if the frequency of G allele of the rs2241880 ATG16L1 polymorphism was increased in both paediatric and adult CD patients compared to con- trols (P = 0.017 and P = 0.001, respectively), no difference was observed between the childhood and the adult cohort. The rare Q allele of IL23R rs11209026 polymorphism was underrepresented in both paediatric and adult CD cases (P = 0.0018 and P = 0.04, respectively) and no difference was observed between the childhood and the adult cohort. The presence of the rs2241880 ATG16L1 and rs11209026 IL23R polymorphisms did not influence disease phenotype. CONCLUSION: Polymorphism 3020insC in NOD2/ CARD15 occurs statistically significantly more often in patients with childhood-onset CD than in patients with adult-onset CD. The ATG16L1 and IL23R variants are associated with susceptibility to CD, but not earlyonset disease.
基金This project Was supported by the National Natural Science Foundation of China(NSFC,No.39870307)National Basic Research Program of China(973 Program,No.G2000056901)KC was the recipient of an Fonds de la recherche en sante du Quebec(FRSQ,Quebec-Canada)-NSFC(China exchange grant).
文摘Arachidonic acid cytochrome P-450 (CYP) hydroxylase 4A isoforms, including 4A1, 4A2, 4A3 and 4A8 in the rat kidney, catalyze arachidonic acid to produce 19/20-Hydroxyeicosatetraenoic acids (20-HETE), a biologically active metabolite, which plays an important role in the regulation of blood pressure. However, controversial results have been reported regarding the exact role of 20-HETE on blood pressure. In the present study, we used recombinant adeno- associated viral vector (rAAV) to deliver CYP 4A1 cDNA and antisense 4A1 cDNA into Sprague-Dawley (SD) rats and spontaneously hypertensive rats (SHR), respectively, to investigate the effects of long-term modifications of blood pressure and the potential for gene therapy of hypertension. The mean systolic pressure increased by 14.2±2.5 mm Hg in rAAV·4A1-treated SD rats and decreased by 13.7±2.2 mm Hg in rAAV·anti4A1-treated SHR rats 5 weeks after the injection compared with controls and these changes in blood pressure were maintained until the experiments ended at 24 weeks. In 4A1 treated animals CYP4A was overexpressed in various tissues, but preferentially in the kidney at both mRNA and protein levels. In anti-4A1-treated SHR, CYP4A mRNA in various tissues was probed, especially in kidneys, but 4A1 protein expression was almost completely inhibited. These results suggest that arachidonic acid CYP hydroxy- lases contribute not only to the maintenance of normal blood pressure but also to the development of hypertension. rAAV-mediated anti4A administration strategy has the potential to be used as targeted gene therapy in human hyperten- sion by blocking expression of CYP 4A in kidneys.
基金National Basic Research Program (973 Program) of China(No. G20000057001)National Natural Science Foundation of China (No.30471994)+1 种基金Shanghai Pujiang Talent Program (No. 05PJ14010)Major Basic Research Project of Shanghai (No. 04DZ14006)
文摘Objective: To explore the role of telomerase activity detected in biopsy samples for evaluating the efficacy of lapa- roscopic radiofrequency ablation (RFA) therapy in patients with hepatocellular carcinoma (HCC) and liver cirrhosis. Methods: From August 2001 to October 2004, 34 cirrhotic patients with HCC were treated by laparoscopic RFA under general anesthe- sia. A total of 34 tumors, with a mean maximum tumor diameter of 4.0 ± 1.0 cm, were all located on the liver surface or adja- cent to the gallbladder. Laparoscopic ultrasound-guided core biopsy for liver lesions was performed before and immediately after RFA therapy. In these biopsy samples, telomerase activity was detected by the ELISA-based telomeric repeat amplifica- tion protocol (ELISA-TRAP) assay, and pathological examination was routinely performed. Results: Laparoscopic RFA was successfully performed in all the 34 patients. A complete tumor necrosis was achieved in all patients on the contrast-enhanced helical CT scanning one month after laparoscopic RFA. The positive rates of telomerase activity and histopathologic diagnosis in biopsy samples were 91.2% (31/34) and 100% (34/34) respectively before RFA, and 26.5% (9/34) and 0% respectively after RFA. During a median follow-up period of 35 months (range, 18–51 months), the rates of local tumor recurrence at the ablation sites in post-RFA telomerase-positive and negative patients were 88.9% (8/9) and 4% (1/25) respectively (P < 0.01), and the rates of distant recurrence within the livers were 0% (0/9) and 12% (3/25) respectively (P > 0.05). Conclusion: For cirrhotic patients with HCC treated by laparoscopic RFA, detection of telomerase activity in biopsy samples may be useful for evaluating the therapeutic efficacy of RFA and predicting postoperative local tumor recurrence.
基金supported by grants from National Natural Science Foundation of China(No.81302130)Military Medical Scientific Youth Cultivation Project(No.13QNP054)
文摘Combined hepatocellular-cholangiocarcinoma(CHC) is a mixed tumor containing elements of both hepatocellular carcinoma(HCC) and cholangiocarcinoma(CC). Its remarkable histological heterogeneity has been linked to putative hepatic progenitor cell(HPC) origin. However, detailed histological or phenotypic description is rarely documented. In the present study, we reassessed 68 cases previously diagnosed as hepatitis B-related CHCs by immunohistochemistry and double-fluorescence immunostaining, focusing on HPC associated phenotypic observation of intermediate area of the tumor. It was found that tumor cells showed remarkable heterogeneity in intermediate area. Tumor cells with intermediate morphology between hepatocytes and cholangiocytes were oval-shaped and small with scant cytoplasm and hyperchromatic nuclei, arranging in solid nests mostly. By Keratin 7(K7) staining, it appeared that the nests of tumor cells represented a maturation process from the undifferentiated small cells to mature hepatocytes through the "transitional" cells. Then, these small cells were further confirmed with intermediate phenotype as HPC by exploring immature hepatocellular marker and HPC/biliary markers co-localization. In conclusion, the HPC associated trait in CHC can be interpreted by HPC origin or gain of "stemness" by dedifferentiation. It is still too soon to give a final word that it is innate or acquired signature of HPC associated trait in CHC.
基金supported by the grant received from Tehran University of Medical Sciences(TUMS-38292)。
文摘Severely immunocompromised NOD.Cg-PrkdcIl2rg(NOG)mice are among the ideal animal recipients for generation of human cancer models.Transplantation of human solid tumors having abundant tumor-i nfiltrating lymphocytes(TILs)can induce xenogeneic graft-versus-host disease(xGvHD)following engraftment and expansion of the TILs inside the animal body.Wilms’tumor(WT)has not been recognized as a lymphocyte-predominant tumor.However,3 consecutive generations of NOG mice bearing WT patient-derived xenografts(PDX)xenotransplanted from a single donor showed different degrees of inflammatory symptoms after transplantation before any therapeutic intervention.In the initial generation,dermatitis,auto-amputation of digits,weight loss,lymphadenopathy,hepatitis,and interstitial pneumonitis were observed.Despite antibiotic treatment,no response was noticed,and thus the animals were prematurely euthanized(day 47 posttransplantation).Laboratory and histopathologic evaluations revealed lymphoid infiltrates positively immunostained with anti-human CD3 and CD8 antibodies in the xenografts and primary tumor,whereas no microbial infection or lymphoproliferative disorder was found.Mice of the next generation that lived longer(91 days)developed sclerotic skin changes and more severe pneumonitis.Cutaneous symptoms were milder in the last generation.The xenografts of the last 2 generations also contained TILs,and lacked lymphoproliferative transformation.The systemic immunoinflammatory syndrome in the absence of microbial infection and posttransplant lymphoproliferative disorder was suggestive of xGvHD.While there are few reports of xGvHD in severely immunodeficient mice xenotransplanted from lymphodominant tumor xenografts,this report for the first time documented serial xGvHD in consecutive passages of WT PDX-bearing models and discussed potential solutions to prevent such an undesired complication.
文摘The field of gene therapy has been increasingly studied in the last four decades, and its clinical application has become a reality in the last 15 years. Traditional Chinese medicine(TCM), an important component of complementary and alternative medicine, has evolved over thousands of years with its own unique system of theories, diagnostics and therapies. TCM is well-known for its various roles in preventing and treating infectious and chronic diseases, and its usage in other modern clinical practice. However, whether TCM can be applied alongside gene therapy is a topic that has not been systematically examined. Here we provide an overview of TCM theories in relation to gene therapy. We believe that TCM theories are congruent with some principles of gene therapy. TCM-derived drugs may also act as gene therapy vehicles, therapeutic genes, synergistic therapeutic treatments, and as co-administrated drugs to reduce side effects. We also discuss in this review some possible approaches to combine TCM and gene therapy.
基金Supported by The National Natural Science Foundation of China,No.30901722,30973433,81000970,81030041,31171321 and 81101622
文摘AIM:To investigate the effect of the demethylating reagent 5-aza-2'-deoxycitidine(DAC) on telomerase activity in hepatocellular carcinoma(HCC) cell lines,SMMC-7721 and HepG2.METHODS:The related gene expression in cell lines was examined by real-time reverse transcription-polymerase chain reaction and Western blotting analysis.The telomerase activity was examined by telomeric repeat amplification protocol-enzyme-linked immunosorbent assay and DNA methylation was determined by methylation-specific polymerase chain reaction.RESULTS:The telomerase activity was significantly reduced in both cell lines treated with DAC,accompanied by downregulation of telomerase reverse transcriptase(hTERT).We also observed the effect of DAC on the methylation status of hTERT promoter and the expression of regulatory genes,such as c-myc,p15,p16,p21,E2F1,and WT1.The methylation status of hTERT promoter could be reversed in SMMC-7721 by DAC,but not in HepG2 cells.However,p16 expression could be reactivated by demethylation of its promoter,and c-Myc expression was repressed in both cell lines.Moreover,DAC could enhance the sensitivity to the chemotherapeutic agents,such as cisplatin,by induction of apoptosis of HCC cells.CONCLUSION:The DAC exerts its anti-tumor effects in HCC cells by inhibiting the telomerase activity.
基金Supported by Ministerio de Ciencia e Innovacion BIO2009/09295 and SAF2012-40003FEDER funding,funds from the"UTE project CIMA"+1 种基金the project RNAREG(CSD2009-00080)from The Ministry of Science and Innovation under the programme CONSOLIDER INGENIO 2010
文摘Hepatocellular carcinoma(HCC) is the third leading cause of cancer-related deaths worldwide. Only 30%-40% of the patients with HCC are eligible for curative treatments, which include surgical resection as the first option, liver transplantation and percutaneous ablation. Unfortunately, there is a high frequency of tumor recurrence after surgical resection and most HCC seem resistant to conventional chemotherapy and radiotherapy. Sorafenib, a multi-tyrosine kinase inhibitor, is the only chemotherapeutic option for patients with advanced hepatocellular carcinoma. Patients treated with Sorafenib have a significant increase in overall survival of about three months. Therefore, there is an urgent need to develop alternative treatments. Due to its role in cell growth and development, the insulin-like growth factor system is commonly deregulated in many cancers. Indeed, the insulin-like growth factor(IGF) axis has recently emerged as a potential target for hepatocellular carcinoma treatment. To this aim, several inhibitors of the pathway have been developed suchas monoclonal antibodies, small molecules, antisense oligonucleotides or small interfering RNAs. However recent studies suggest that, unlike most tumors, HCC development requires increased signaling through insulin growth factor Ⅱ rather than insulin growth factor Ⅰ. This may have great implications in the future treatment of HCC. This review summarizes the role of the IGF axis in liver carcinogenesis and the current status of the strategies designed to target the IGF-Ⅰ signaling pathway for hepatocellular carcinoma treatment.
基金supported by grants from the University of Massachusetts Medical School(an internal grant)and by the National Institutes of Health(R01NS076991-01,1P01All00263-01,4P01HL131471-02,UG3 HL147367-01,1U19AI149646,and R01HL097088).
文摘Throughout its 40-year history,the field of gene therapy has been marked by many transitions.It has seen great strides in combating human disease,has given hope to patients and families with limited treatment options,but has also been subject to many setbacks.Treatment of patients with this class of investigational drugs has resulted in severe adverse effects and,even in rare cases,death.At the heart of this dichotomous field are the viral-based vectors,the delivery vehicles that have allowed researchers and clinicians to develop powerful drug platforms,and have radically changed the face of medicine.Within the past 5 years,the gene therapy field has seen a wave of drugs based on viral vectors that have gained regulatory approval that come in a variety of designs and purposes,these modalities range from vector-based cancer therapies,to treating monogenic diseases with life-altering outcomes.At present,the three key vector strategies are based on adenoviruses,adeno-associated viruses,and lentiviruses.They have led the way in preclinical and clinical successes in the past two decades.However,despite these successes,many challenges still limit these approaches from attaining their full potential.To review the viral vector-based gene therapy landscape,we focus on these three highly regarded vector platforms and describe mechanisms of action and their roles in treating human disease.
文摘Objective: To explore the effects of thiazolidinediones(TZDs) pioglitazone on proliferation and differentiation of human preadipocytes. Methods:Omental adipose tissue biopsies were obtained from 15 patients who were undergoing elective open-abdominal surgery. The primary culture and differentiated induction of human preadipocytes were performed, and the human preadipo-cytes were treated with pioglitazone at different concentrations at proper moments. Dynamic morphological changes of the human preadipocytes were observed, and their proliferation and differentiation were assessed with Colorimetric MTT Assay and Oil Red O Staining. Results:After 24 hours and 72 hours with pioglitazone, 0.1 μmol/L(μmol/ml) pioglitazone increased the MTT values of the human preadipocytes by 25.3% and 34.8%,respectively(P < 0.05), while 1 μmol/L pioglitazone by 27.4% and 26.6%(P < 0.05), compared with the control group without pioglitazone. The human preadipocytes with pioglitazone cumulated more adipose in the endochylema than those without pioglitazone obviously. 0.1 μmol/L pioglitazone increased the differentiation degree of the human preadipocytes differentiated for 8-10 days by 44.81% and 1 μmol/L pioglitazone by 53.76%(P < 0.05). Conclusion:Thiazolidinediones pioglitazone may significantly promote the proliferation and differentiation of the human omental preadipocytes.
基金Tehran University of Medical Sciences and Health ServicesGrant/Award Number:98-3-101-45499。
文摘Background:Bladder cancer poses a great burden on society and its high rate of recurrence and treatment failure necessitates use of appropriate animal models to study its pathogenesis and test novel treatments.Orthotopic models are superior to other types since they provide a normal microenvironment.Four methods are described for developing bladder cancer models inside the animal’s bladder.Direct intramural injection is one of these methods and is widely used.However,its efficacy in model development has not yet been studied.We aimed to evaluate the efficacy and success rate of the direct intramural injection method of developing an orthotopic model for the study of bladder cancer.Method:Tumor cell lines were prepared in four microtubes.Aliquots of 200×10^(3) cells were injected through a 27 gauge needle into the ventral wall of the bladders of 4male and 4 female BALB/c mice following a midline 1 cm laparotomy incision.In addition,1 million cells from each microtube were injected into the flanks of control mice.To prevent infection and alleviate pain,5 mg/kg enrofloxacin and 2.5 mg/kg flunixin meglumine,respectively,were injected subcutaneously.Results:Tumors formed in all mice,resulting in 100% take rate and zero post-operation mortality.Surgery time was≤15 min per mouse.In two mice,tumors were found in the peritoneal space as well.Conclusion:Direct intramural injection is a rapid,reliable,and reproducible method for developing orthotopic models of bladder cancer.It can be done on both male and female mice and only requires readily available surgical tools.However,needle track can result in cell spillage and peritoneal tumors.
基金Supported by National 973 Project of China(No.G2000057001)
文摘Objective: To induce changes in biological character of human liver cancer cell line SMMC-7721 by blocking the expression of telomerase genes hTRT and to explore its value in cancer gene therapy. Methods: The vehicle for eukaryotic expression of antisense hTRT was constructed and then transfected into SMMC-7721 cells. The effects of antisense hTRT gene on telomerase activity, cancer cell growth and malignant phenotypes were analyzed. Results: The obtained transfectants that could express antisense hTRT gene stably showed marked decrease in telomerase activity; the shortening of telomere was obvious; cells presented contact growth inhibition; in nude mice transplantation, the rate of tumor induction dramatically decreased. Conclusion: Antisense hTRT gene expression can significantly inhibit telomerase activity of cancer cells and decrease malignant phenotypes in vitro and in vivo. Therefore, as a telomerase inhibitor, antisense hTRT gene may be a new pathway for cancer therapy.
文摘AIM: To elucidate the biological function of HBV core antigen (HBcAg) on pathogenesis of hepatitis B, a novel gene C12 coding for protein with unknown function interacting with HBcAg in hepatocytes was identified and characterized. METHODS: HBcAg bait plasmid pGBKT7-HBcAg was constructed and transformed into yeast AH109, then the transformed yeast was mated with yeast Y187 containing liver complementary DNA (cDNA) library plasmid in 2×YPDA medium. Diploid yeast was plated on synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade) and synthetic dropout nutrient medium (SD/-Trp-Leu-His-Ade)containing X-α-gal for screening twice. After extracting and sequencing of plasmid from blue colonies, we isolated a cDNA clone encoding a novel protein designated as C12that directly interacted with HBcAg. The interaction between HBcAg and C12 was verified again by re-mating.pEGFP-N1-C12 fluorescent protein fusion gene was transfected in 293 and L02 cell, and observed by fluorescent microscope. MTT reduction assay was used to study the action of C12 protein effect on metabolism of mammal cell. Yeast two-hybrid and cDNA microarray were performed to search binding protein and differential expression genes regulated by C12 protein.RESULTS: C12 gene was screened and identified by yeast two-hybrid system 3. The interaction between HBcAg and the novel protein coded by the new gene C12 was further confirmed by re-mating. After 48 h, fluorescence of fusion protein could be observed steadily in the 293 and L02 cell plasma. Under MTT assay, we found that the expression of C12 did not influence the growth of liver cells. Seventeen differential expression genes in HepG2 cells transfected with C12 protein expression plasmid by cDNA microarray,of which 16 genes were upregulated and 1 gene was downregulated by C12 protein. Twenty-one colonies containing 16 different genes coding for C12 protein binding proteins were isolated by yeast two-hybrid, there were 2 new genes with unknown function.CONCLUSION: The novel protein C12 is located in cell plasma, and its overexpression has no significant effect on the metabolism of liver cell. It interacts with many proteins in hepatocytes and may be involved in many processes of gene expression.
文摘LIM domain kinases(LIMKs),which modulate cytoskeletal dynamics,are found throughout the central nervous system.The synaptic junctions between neurons show structural alteration or plasticity during neurodevelopment,learning and memory formation,or after injury and disease;in some cases completely new connections form,while others are lost.This remodeling is under the control of the actin cytoskeleton and therefore is influenced by the activity of LIMKs(Scott and Olson,2007).Recent reports discussed in this perspective shed light on the potential role of LIMK in hippocampal dendritic spines and photoreceptor presynaptic terminals and illustrate the importance of LIMK in nerve cell function.
基金The National Nature Science Foundation of China,No.81900363.
文摘BACKGROUND Metastatic skin cancers are relatively rare dermatological malignancies.They usually present as nodules,erythematous lesions,scar-like lesions or other lesion types.Signet-ring cell carcinoma(SRCC)is an uncommon histological type of gastric cancer that usually behaves aggressively and has a poor prognosis.Skin metastasis may be the first sign of clinically silent visceral cancer or recurrence of an internal malignancy.CASE SUMMARY Herein we report on the case of a 55-year-old man with edema of a lower extremity as the primary symptom which progressed from local to generalized pitting edema in the year following skin involvement.Pathological evidence from gastroscopic specimens and subcutaneous tissue biopsy showed typical signetring cells and gland-like structures.Consistently,immunohistochemical analysis revealed positive pan-cytokeratin expression in tumor cells.A diagnosis of gastric SRCC with skin metastasis was established.Moreover,lymphoscintigraphy showed an obvious accumulation of radiotracer on the anterior and posterior sides of the right leg which indicated lymphedema.We reviewed the relevant literature on subcutaneous metastases of gastric SRCC.CONCLUSION This rare case emphasizes the importance of physical examination as it may help elucidate the etiology of edema.
文摘Hepatic fibrosis/cirrhosis is a significant health burden worldwide,resulting in liver failure or hepatocellular carcinoma(HCC)and accounting for many deaths each year.The pathogenesis of hepatic fibrosis/cirrhosis is very complex,which makes treatment challenging.Endogenous mesenchymal stromal cells(MsCs)have been shown to play pivotal roles in the pathogenesis of hepatic fibrosis.Paradoxically,exogenous MsCs have also been used in clinical trials for liver cirrhosis,and their effectiveness has been observed in most completed clinical trials.There are still many issues to be resolved to promote the use of MsCs in the clinic in the future.In this review,we will examine the controversial role of MsCs in the pathogenesis and treatment of hepatic fibrosis/cirrhosis.We also investigated the clinical trials involving MsCs in liver cirrhosis,summarized the parameters that need to be standardized,and discussed how to promote the use of MsCs from a clinical perspective.
