Induction of antiproliferative effect by diosgenin through activation of p53, release of apoptosis-inducing factor (AIF) and modulation of caspase-3 activity in different human cancer cells

Previously, we demonstrated that a plant steroid, diosgenin, altered cell cycle distribution and induced apoptosis in the human osteosarcoma 1547 cell line. The objective of this study was to investigate if the antiproliferative effect of diosgenin was similar for different human cancer cell lines such as laryngocarcinoma HEp-2 and melanoma M4Beu cells. Moreover, this work essentially focused on the mitochondrial pathway. We found that diosgenin had an important and similar antiproliferative effect on different types of cancer cells. In addition, our new results show that diosgenininduced apoptosis is caspase-3 dependent with a fall of mitochondrial membrane potential, nuclear localization of AIF and poly (ADP-ribose) polymerase cleavage. Diosgenin treatment also induces p53 activation and cell cycle arrest in the different cell lines studied.

Maintenance of radiation-induced intestinal fibrosis:Cellular and molecular features

Recent advances in cell and molecular radiobiology clearly showed that tissue response to radiation injury cannot be restricted to a simple cell-killing process, but depends upon continuous and integrated pathogenic processes, involving cell differentiation and crosstalk between the various cellular components of the tissue within the extracellular matrix. Thus, the prior concept of primary cell target in which a single-cell type （whatever it＇s epithelial or endothelial cells） dictates the whole tissue response to radiation injury has to be replaced by the occurrence of coordinated multicellular response that may either lead to tissue recovery or to sequel development. In this context, the present review will focus on the maintenance of the radiation-induced wound healing and fibrogenic signals triggered by and through the microenvironment toward the mesenchymal cell compartment, and will highlight how sequential and sustained modifications in cell phenotypes will in cascade modify ceU-to-ceU interactions and tissue composition.

Hepatocellular carcinoma in patients with hepatitis C virus-related chronic liver disease

Hepatitis C virus (HCV) is a major cause of hepatocellular carcinoma (HCC) worldwide due to the high prevalence of HCV infection and the high rate of HCC occurrence in patients with HCV cirrhosis. A striking increase in HCC incidence has been observed during the past decades in most industrialized countries, partly related to the growing number of patients infected by HCV. HCC is currently the main cause of death in patients with HCV-related cirrhosis, a fact that justifies screening as far as curative treatments apply only in patients with small tumors. As a whole, treatment options are similar in patients with cirrhosis whatever the cause. Chemoprevention could be also helpful in the near future. It is strongly suggested that antiviral treatment of HCV infection could prevent HCC occurrence, even in cirrhotic patients, mainly when a sustained virological response is obtained.

Effect of sustained virological response on long-term clinical outcome in 113 patients with compensated hepatitis C-related cirrhosis treated by interferon alpha and ribavirin

AIM： To assess the long-term clinical benefit of sustained virological response （SVR） in patients with hepatitis C virus （HCV） cirrhosis treated by antiviral therapy using mostly ribavirin plus interferon either standard or pegylated.METHODS： One hundred and thirteen patients with uncomplicated HCV biopsy-proven cirrhosis, treated by at least one course of antiviral treatment ≥ 3 mo and followed ≥ 30 mo were included. The occurrence of clinical events [hepatocellular carcinoma （HCC）, decompensation and death] was compared in SVR and non SVR patients.RESULTS： Seventy eight patients received bitherapy and 63 had repeat treatments. SVR was achieved in 37 patients （33%）. During a mean follow-up of 7.7 years, clinical events occurred more frequently in non SVR than in SVR patients, with a significant difference for HCC （24/76 vs 1/37, P = 0.01）. No SVR patient died while 20/76 non-SVR did （P = 0.002）, mainly in relation to HCC （45%）.CONCLUSION： In patients with HCV-related cirrhosis, $VR is associated with a significant decrease in the incidence of HCC and mortality during a follow-up period of 7.7 years. This result is a strong argument to perform and repeat antiviral treatments in patients with compensated cirrhosis.

Expression of matrix metalloproteinases and tissue inhibitor metalloproteinases increases in X-irradiated rat ileum despite the disappearance of CD8a T cells

AIM： To investigate their expression and activity in the rat ileum after exposure to ionizing radiation along with that of the cellular effectors and molecular mediators involved in the regulation of MMPs. METHODS： Rats were exposed to a single 10-Gy dose of X-rays delivered to the abdomen. A combination of methods, such as zymography, immunohistochemistry and real time reverse transcriptase-polymerase chain reaction, were used to localize and quantify MMPs and the molecules involved in MMP activating and inhibitory pathways （plasmin/ plasminogen, TIMPs）, CD^8＋, as well as inflammatory （interleukin （IL）-1β, IL-8, tumor necrosis factor-s, TNF-α） and fibrogenic mediators （transforming growth factor- β1-3） within ileal tissue at 1, 3, and 7 d after irradiation. RESULTS： A marked increase in MMP-2 and -14 mRNA and protein levels associated with an increased activity of MMP-2 was observed in irradiated ileal tissue. MMP-2 and -14 expression was mainly observed in inflammatory, epithelial, and mesenchymal cells, whereas a slight increase in MMP-3 expression was detected in the few infiltrating macrophages at d i after irradiation. Conversely, MMP-1, -7, and -9 mRNA levels were not found to be affected by abdominal irradiation. Irradiation was found to induce disappearance of CD^8＋ cells. Furthermore, we have observed that TNF-α and IL-1β protein levels increased 6 h after irradiation, whereas those of IL-8 only increased after 3 d and was concomitant with neutrophil infiltration. In addition, the expressions of molecules involved in MMP activating and inhibitory pathways （urokinase-type plasminogen activator andtissue-type plasminogen activator; TIMP-1, TIMP-2, and plasminogen activator-inhibitor-i） were found to be increased after abdominal irradiation. CONCLUSION： This study showed that abdominal irradiation induces an acute remodeling of the ileum associated with an increased expression of MMPs and TIMPs that do not involve CD^8＋ T cells but involve mesenchymal and epithelial cells, although to a lesser extent, and probably even soluble inflammatory and fibrogenic mediators.

Heat and Mass Transfer Investigation of Hydrocarbon Droplet Evaporation under Rotatory Movement

The steady-state boundary layer equations of a rotatory movement around hydrocarbon droplets saturated of pure fuel are numerically solved. The transfer equations are schemed by using an implicit finite difference method. The system of algebraic equations is solved by using the Thomas algorithm. The model is compared to the Kreith one and a good agreement is observed between both models. The dimensionless physical parameters of the evaporation phenomena of droplet in rotation are calculated and presented under the effect of the convection.

Recent advances in hepatitis C virus research and understanding the biology of the virus

Since the identification of hepatitis C virus （HCV） genome in 1989m, a lot of progresses have been done about the understanding of HCV biology, natural history and therapeutic options. HCV is a member of the Flaviviridae viral family. Its genome is a positive simple strand RNA molecule which shows significant genetic variability. The HCV nomenclature has been recently re-examined by an international group of scientific experts in the field of HCV genetic variability or involved in HCV data base.

无机抗菌剂的添加对新型自酸蚀处理剂黏结性能和颜色稳定性的影响

目的:测试无机抗菌剂的添加对新型自酸蚀处理剂ESP黏结性能和颜色稳定性的影响。方法:6种无机抗菌剂分别加入ESP中,测试牛牙釉质粘结强度、边缘封闭性以及粘结样本颜色稳定性的变化。结果:抗菌剂添加比为0.5%对ESP牙釉质剪切强度无不利影响(P>0.05),比例增至2.5%时,黏结强度呈不同程度降低。抗菌剂添加比为0.5%时,仅有一种抗菌剂的添加使冠方微渗漏指数较对照组显著增加(P<0.05)。粘结样本经不同处理以老化1年后颜色参数变化最为显著,但与ESP空白对照相比,抗菌剂添加后的颜色变化仍在临床可接受的范围之内。结论:控制适当的添加比例,5种受试的无机抗菌剂的添加对ESP的黏结性能可无显著影响,对颜色稳定性的影响在临床可接受的范围之内。

利用无机抗菌剂研制抗菌自酸蚀处理剂的初步研究

目的评价利用无机抗菌剂研制抗菌自酸蚀处理剂的初步可行性。方法将6种无机抗菌剂以0.5% w/v和2.5% w/v的浓度分别加入自行研制的实验自酸蚀处理剂ESP中,进行变形链球菌体外药敏试验。并应用添加了抗菌剂的ESP制备黏结样本,测试无机抗菌剂的添加对材料固化后抗菌活性的影响。结果 ESP本身就具有一定的抗菌作用,在其中添加龙贝无机抗菌粉体或氧化锌晶须复合抗菌剂AT-83可使液体状态下ESP的最小抑菌浓度和最小杀菌浓度显著降低;加入受试的无机抗菌剂使材料固化后的抗菌活性显著增强,其中添加氧化锌晶须复合抗菌剂AT-83的效果最为显著。结论从抗菌性能方面证实了利用受试的无机抗菌剂研制抗菌自酸蚀处理剂的初步可行性。

Relationship between chromatin organization, mRNAs profile and human male gamete quality

Spermiogenesis is a complex process leading to the formation of motile spermatozoa characterized by a highly stable chromatin compaction that transfers the paternal genome into the oocyte. It is commonly held that these haploid cells are devoid of transcriptional and translational activities and that the transcripts represent remnants of stored mRNAs. Recently, the chromatin organization of mature spermatozoa has been revisited as a double nucleoprotamine-nucleohistone structure possessing less-condensed regions sensitive to nuclease activity, which could be implicated in the expression of genes involved in the early embryo development. The existence of a complex population of mRNAs in human sperm is well-documented, but their role is not yet elucidated. Evidence for a latent transcriptional capacity and/or a potential de novo translation in mature spermatozoa from fertile men are essential for understanding the last steps of sperm maturation, such as capacitation and acrosome reaction. As such, we have documented the relationship between sperm quality and the distribution of sperm RNAs by showing divergent levels of transcripts encoding for proteins involved in either nuclear condensation （protamines 1 and 2） or in capacitation （eNOS and nNOS, c-myc） or in motility and sperm survival （aromatase） between low and high motile sperm issued from the same sample. Therefore, analyzing the profile of mRNAs could be helpful either as a diagnostic tool for evaluating male fertility after spermatogenesis or for prognosis use for fertilization.

Genetic diversity of the hepatitis C virus: Impact and issues inthe antiviral therapy

The hepatitis C Virus (HCV) presents a high degree of genetic variability which is explained by the combination of a lack of proof reading by the RNA dependant RNA polymerase and a high level of viral replication. The re- sulting genetic polymorphism defines a classification in clades, genotypes, subtypes, isolates and quasispecies. This diversity is known to reflect the range of responses to Interferon therapy. The genotype is one of the pre- dictive parameters currently used to define the antiviral treatment strategy and the chance of therapeutic suc- cess. Studies have also reported the potential impact of the viral genetic polymorphism in the outcome of antivi- ral therapy in patients infected by the same HCV geno- type. Both structural and non structural genomic regions of HCV have been suggested to be involved in the Inter- feron pathway and the resistance to antiviral therapy. In this review, we first detail the viral basis of HCV diversity. Then, the HCV genetic regions that may be implicated in resistance to therapy are described, with a focus on the structural region encoded by the E2 gene and the non- structural genes NS3, NS5A and NS5B. Both mechanisms of the Interferon resistance and of the new antiviral drugs are described in this review.

Quasispecies evolution in NS5A region of hepatitis C virus genotype 1b during interferon or combined interferon-ribavirin therapy

AIM: To evaluate the implication of substitutions in the hepatitis C virus (HCV) non-structural 5A (NS5A) protein in the resistance of HCV during mono-interferon (IFN) or combined IFN-ribavirin (IFN-R) therapy. Although NS5A has been reported to interact with the HCV RNA- dependent RNA polymerase, NS5B, as well as with many cellular proteins, the function of NS5A in the life cycle of HCV remains unclear. METHODS: HCV quasispecies were studied by clon- ing and sequencing of sequential isolates from patients infected by HCV genotype 1b. Patients were treated by IFN-α2b for 3 mo followed by IFN-α2b alone or com- bined IFN-R therapy for 9 additional months. Patients were categorized intro two groups based on their re- sponse to the treatments: 7 with sustained virological re- sponse (SVR) (quasispecies = 150) and 3 non-respond- ers (NR) to IFN-R (quasispecies = 106). RESULTS: Prior to treatment, SVR patients displayed a lower complexity of quasispecies than NR patients. Most patients had a decrease in the complexity of quasispe- cies during therapy. Analysis of amino acids substitu- tions showed that the degree of the complexity of the interferon sensitivity-determining region (ISDR) and the V3 domain of NS5A protein was able to discriminate thetwo groups of patients. Moreover, SVR patients displayed more variability in the NS5A region than NR patients. CONCLUSION: These results suggest that detailed mo- lecular analysis of the NS5A region may be important for understanding its function in IFN response during HCV 1b infection.

Liver insulin-like growth factor 2 methylation in hepatitis C virus cirrhosis and further occurrence of hepatocellular carcinoma

AIM: To assess the predictive value of the insulinlike growth factor 2 (Igf2) methylation profile for the occurrence of Hepatocellular Carcinoma (HCC) in hepatitis C (HCV) cirrhosis. METHODS: Patients with: (1) biopsy-proven compensated HCV cirrhosis; (2) available baseline frozen liver sample; (3) absence of detectable HCC; (4) regular screening for HCC; (5) informed consent for genetic analysis were studied. After DNA extraction from liver samples and bisulfite treatment, unbiased PCR and DHPLC analysis were performed for methylation analysis at the Igf2 locus. The predictive value of the Igf2 methylation profile for HCC wasassessed by Kaplan-Meier and Cox methods. RESULTS: Among 94 included patients, 20 developed an HCC during follow-up (6.9 ± 3.2 years). The methylation profile was hypomethylated, intermediate and hypermethylated in 13, 64 and 17 cases, respectively. In univariate analysis, two baseline parameters were associated with the occurrence of HCC: age (P = 0.01) and prothrombin (P = 0.04). The test of linear tendency between the three ordered levels of Igf2 methylation and probability of HCC occurrence was significant (Log Rank, P = 0.043; Breslow, P = 0.037; Tarone-Ware, P = 0.039). CONCLUSION: These results suggest that hypomethylation at the Igf2 locus in the liver could be predictive for HCC occurrence in HCV cirrhosis.

无机抗菌剂对自酸蚀处理剂全身毒性和细胞毒性影响的研究

目的测试无机抗菌剂的添加对新型实验自酸蚀处理剂(ESP)全身毒性和细胞毒性的影响。方法将6种无机抗菌剂分别加入ESP中,进行经口途径大鼠短期全身毒性实验。并采用受试材料与NIH3T3细胞直接接触培养的方法,观测其对细胞增殖、活力和形态的影响。结果抗菌剂添加比为5.0%时,为期2周的大鼠全身毒性实验中未发现临床毒性体征,各组间大鼠体重相对增长率无显著差异。经含质量浓度为0.5%不同抗菌剂的处理剂处理固化后的各实验组细胞活力比率和细胞增殖率与ESP空白对照组相比无显著差异。仅样本覆盖区及样本边缘邻近区域细胞可观察到毒性表现,远离样本覆盖区的细胞密度未受明显影响。结论以适当的质量浓度添加受试的无机抗菌剂对受试ESP的全身毒性和细胞毒性无显著影响。

Genome-wide differences in hepatitis C-vs alcoholism-associated hepatocellular carcinoma

AIM:To look at a comprehensive picture of etiology- dependent gene abnormalities in hepatocellular carcinoma in Western Europe. METHODS:With a liver-oriented microarray,transcript levels were compared in nodules and cirrhosis from a training set of patients with hepatocellular carcinoma (alcoholism,12;hepatitis C,10)and 5 controls.Loose or tight selection of informative transcripts with an abnormal abundance was statistically valid and the tightly selected transcripts were next quantified by qRTPCR in the nodules from our training set(12+10) and a test set(6+7). RESULTS:A selection of 475 transcripts pointed to significant gene over-representation on chromosome 8 (alcoholism)or-2(hepatitis C)and ontology indicated a predominant inflammatory response(alcoholism)or changes in cell cycle regulation,transcription factors and interferon responsiveness(hepatitis C).A stringent selection of 23 transcripts whose differences betweenetiologies were significant in nodules but not in cirrhotic tissue indicated that the above dysregulations take place in tumor but not in the surrounding cirrhosis.These 23 transcripts separated our test set according to etiologies. The inflammation-associated transcripts pointed to limited alterations of free iron metabolism in alcoholic vs hepatitis C tumors. CONCLUSION:Etiology-specific abnormalities(chromo- some preference;differences in transcriptomes and related functions)have been identified in hepatocellular carcinoma driven by alcoholism or hepatitis C.This may open novel avenues for differential therapies in this disease.

Relaxant Effects of Quercetin and Rutin on Human Isolated Bronchus

Increasing epidemiological evidence supports the view, that quercetin has protective roles in a multitude of disease states in human who have a high intake of polyphenols. To investigate the ability of quercetin and its rutinoside, rutin, to modulate the relaxation of human airways smooth muscle and to determine the mechanism (s) of such relaxation, isolated human bronchus rings were suspended in individual organ baths, precontracted with acetylcholine or with histamine and the relaxing effects of quercetin and rutin were determined by measurement of isometric tension. Quercetin induced concentration-dependent relaxant responses on acetylcholine or histamine precontracted human bronchial rings and with almost equal effectiveness. In terms of potency (pD2) and efficacy (Emax), quercetin is more potent than rutin on relaxant responses of human bronchus. K+ and Ca2+ concentration-dependent contraction curves were inhibited after incubation with increasing concentrations of quercetin. Quercetin potentiated in a concentration-dependent manner the relaxant effects of isoprenaline or sodium nitroprusside. Rutin had no effect on K+-induced contraction and on relaxant activity of isoprenaline or sodium nitroprusside. Our results suggest that the bronchodilator effects of quercetin are modulated by an increase in cyclic nucleotide levels as well as an alteration in availability of Ca2+ to the contractile machinery.

Tissue toxicity induced by ionizing radiation to the normal intestine:Understanding the pathophysiological mechanisms to improve the medical management

At the present time, more than one-half of all cancer patients are treated with radiation therapy. Despite a good therapeutic index, radiotherapy can disable normal tissue injury to normal tissues in long-term cancer survivors.Thus, an important challenge to modern radiation therapy is to increase the tolerance of normal tissues,

以波生坦作为一线药物治疗特发性肺动脉高压的远期疗效

Aims: Data on long-term efficacy of bosentan in unselected idiopathic pulmonary arterial hypertension(IPAH) patients are lacking. We aimed to describe the long-term outcome of consecutive IPAH patients treated first-line with bosentan. Methods and results: A retrospective analysis of 103 consecutive New York Heart Association functional class III/IV IPAH patients treated with bosentan at our centre between November 1999 and May 2004 was performed. The 6-minute walk distance(6MWD) and haemodynamics were assessed at baseline and after 4 and 12 months. Mean follow-up was 24±15 months. At 4 months, significant improvements in exercise capacity and haemodynamics were observed and persisted up to 1 year. Overall survival estimates were 90 and 87%and event-free status(survival without transplantation, prostanoid initiation, or hospitalization for right heart failure) estimates were 61 and 44%at 1 and 2 years, respectively. Forty-five(44%) patients required prostanoid therapy during follow-up. The 6MWD and the right atrial pressure at baseline and the 6MWD, the increase in 6MWD, and the decrease in pulmonary resistance after 4 months of treatment were associated with long-term outcomes. Conclusion: In our series of consecutive IPAH patients treated with bosentan, improvements in exercise capacity and haemodynamics were similar to those observed in previous randomized trials. However, on the basis of local criteria, many patients required the addition of prostanoid therapy during follow-up.

Hepatocyte apoptosis fragment product cytokeratin-18 M30 level and non-alcoholic steatohepatitis risk diagnosis:an international registry study

Background:Liver biopsy for the diagnosis of non-alcoholic steatohepatitis(NASH)is limited by its inherent invasiveness and possible sampling errors.Some studies have shown that cytokeratin-18(CK-18)concentrations may be useful in diagnosing NASH,but results across studies have been inconsistent.We aimed to identify the utility of CK-18 M30 concentrations as an alternative to liver biopsy for non-invasive identification of NASH.Methods:Individual data were collected from 14 registry centers on patients with biopsy-proven non-alcoholic fatty liver disease(NAFLD),and in all patients,circulating CK-18 M30 levels were measured.Individuals with a NAFLD activity score(NAS)≥5 with a score of≥1 for each of steatosis,ballooning,and lobular inflammation were diagnosed as having definite NASH;individuals with a NAS≤2 and no fibrosis were diagnosed as having non-alcoholic fatty liver(NAFL).Results:A total of 2571 participants were screened,and 1008(153 with NAFL and 855 with NASH)were finally enrolled.Median CK-18 M30 levels were higher in patients with NASH than in those with NAFL(mean difference 177 U/L;standardized mean difference[SMD]:0.87[0.69–1.04]).There was an interaction between CK-18 M30 levels and serum alanine aminotransferase,body mass index(BMI),and hypertension(P<0.001,P=0.026 and P=0.049,respectively).CK-18 M30 levels were positively associated with histological NAS in most centers.The area under the receiver operating characteristics(AUROC)for NASH was 0.750(95%confidence intervals:0.714–0.787),and CK-18 M30 at Youden’s index maximum was 275.7 U/L.Both sensitivity(55%[52%–59%])and positive predictive value(59%)were not ideal.Conclusion:This large multicenter registry study shows that CK-18 M30 measurement in isolation is of limited value for non-invasively diagnosing NASH.

对Rho激酶的抑制可通过改变细胞骨架及结缔组织生长因子的表达,调节小肠平滑肌细胞的辐射诱发纤维化显型

Background: Late radiation enteritis in humans is associated with accumulatio n of extracellular matrix and increased connective tissue growth factor (CTGF) e xpression that may involve intestinal muscular layers. Aims: We investigated the molecular pathways involved in maintenance of radiation induced fibrosis by gen e profiling and postulated that alteration of the Rho pathway could be associate d with radiation induced fibrogenic signals and CTGF sustained expression. Patie nts and methods: Ileal biopsies from individuals with and without radiation ente ritis were analysed by cDNA array, and primary cultures of intestinal smooth mus cle cells were established. Then, the effect of pharmacological inhibition of p1 60 Rho kinase, using Y- 27632, was studied by real time reverse transcription- polymerase chain reaction, western blot, and electrophoretic mobility shift ass ay. Results: Molecular profile analysis of late radiation enteritis showed alter ations in expression of genes coding for the Rho proteins. To investigate furthe r the involvement of the Rho pathway in intestinal radiation induced fibrosis, p rimary intestinal smooth muscle cells were isolated from radiation enteritis. Th ey retained their fibrogenic differentiation in vitro, exhibited a typical cytos keletal network, a high constitutive CTGF level, increased collagen secretory ca pacity, and altered expression of genes coding for the Rho family. Rho kinase bl ockade induced a simultaneous decrease in the number of actin stress fibres, α smooth muscle actin, and heat shock protein 27 levels. It also decreased CTGF l evels, probably through nuclear factor κ B inhibition, and caused decreased exp ression of the type I collagen gene. Conclusion: This study is the first showing involvement of the Rho/Rho kinase pathway in radiation fibrosis and intestinal smooth muscle cell fibrogenic differentiation. It suggests that specific inhibit ion of Rho kinase may be a promising approach for the development of antifibroti c therapies.