Sedat Karademir

Differential expression of Caveolin-1 in hepatocellular carcinoma: correlation with differentiation state, motility and invasion

BackgroundCaveolin-1 is the main component of caveolae membrane structures and has different roles during tumorigenesis in different cancer types with varying expression profiles, indicating that the role of caveolin-1 varies according to tumor type. In this study, we investigated the role and expression of caveolin-1 in hepatocellular carcinogenesis.MethodsWe analyzed the expression of Caveolin-1 in 96 hepatocellular carcinoma (HCC), 29 cirrhosis, 20 normal liver tissues and 9 HCC cell lines by immunostaining and western blotting, respectively. After caveolin-1 was stably transfected to HepG2 and Huh7 cells, the effects of Caveolin-1 on the cellular motility, matrix invasion and anchorage-independent growth were studied. Also, caveolae structure was disrupted in endogenously caveolin expressing cells, SNU 449 and SNU 475 by addition of methyl-β-cyclodextrin and analyzed cellular motility and invasion.ResultsIn HCC cell lines, Caveolin-1 expression is correlated to differentiation and basal motility status of these cells. The percentage of Caveolin-1 positivity was found extremely low in normal liver tissue (5%) while it was increased in cirrhosis (45%) and in HCC (66%) (p = 0.002 and p = 0.001 respectively). Cav-1 expression in poorly differentiated HCC samples has been found significantly higher than well differentiated ones (p = 0.001). The caveolin-1 expression was found significantly higher in tumor cells than its peritumoral cirrhotic tissues in HCC samples (p < 0.001). Additionally, the patients with positive staining for Caveolin-1 had significantly higher portal vein invasion than those with negative staining (p = 0.02). Caveolin-1 overexpression increased motility and invasion of HepG2 and Huh7 cells. And disruption of caveolae results in a dramatic decline in both motility and invasion abilities in SNU-449 and SNU-475 cells. Furthermore, caveolin-1 overexpression resulted in down-regulation of E-cadherin while up-regulation of Vimentin. Also, it increased secreted MMP-2 and expression levels of MMP-9 and MT1-MMP. There was no significant difference in colony formation in soft agar between stable clones and parental ones.ConclusionIn conclusion, stepwise increase in Cav-1 expression in neoplastic stage with respect to pre-neoplastic stage during hepatocellular carcinogenesis and its ability to stimulate HCC cell motility and invasiveness indicate that this protein plays a crucial role in tumor progression.

Risk Factors for Early Bacterial Infections in Liver Transplantation

OBJECTIVE Our aim was to determine perioperative risk factors for early bacterial infection after liver transplantation. METHODS Retrospectively examining medical records using Centers for Disease Control and Prevention (CDC) definitions to identify nosocomial infections, we analyzed data on 367 adult patients. RESULTS The incidence of infection was 37.3% (n = 137): namely, surgical site (n = 74; 20.2%) [corrected], blood stream (n = 64; 17.4%), pulmonary (n = 49; 13.4%), urinary system (n = 26; 7.1%). Significant risk factors within the first 30 days were as follows: deceased donor, Model for End-Stage Liver Disease (MELD) >20, albumin level <2.8 g/dL, intraoperative erythrocyte transfusion >6 U, intraoperative fresh frozen plasma >12 U, bilioenteric anastomosis, postoperative intensive care unit stay >6 days, and postoperative length of stay >21 days. Significant risk factors detected within the first 90 days were as follows: MELD >20, preoperative length of stay >7 days, reoperation, postoperative length of intensive care unit stay >6 days, and postoperative length of stay >21 days. Variability was observed in risk factors according to localization of infection. As a result, except for MELD, type of donor, and biliary anastomosis, the others are preventable factors for early bacterial infection. In addition, the same risk factors showed variability according to the site of infection.

Immunohistochemical detection of pS2 protein and heat shock protein-70 in pancreatic adenocarcinomas. Relationship with disease extent and patient survival.

We investigated pS2 and HSP-70 protein expression in 36 pancreatic adenocarcinomas for their effect on disease extent and patient outcome. The cases were reviewed, histologically diagnosed, typed, graded, and staged. Lymphatic vessel, blood vessel and perineural invasion as well as lymph node, resection margin and adjacent organ involvements were re-evaluated. The standard streptavidin biotin immunperoxidase method was used for immunostaining with pS2 and HSP-70 antibodies. Cytoplasmic staining with both antibodies was scored semiquantitatively. The scores were compared with histopathological prognostic parameters using statistical methods. Standard prognostic parameters and staining scores were tested by survival analysis in terms of their effect on survival. All the tumors showed a positive cytoplasmic reaction with HSP-70 antibody. Seventy-seven percent of the tumors showed positive cytoplasmic staining with pS2 antibody (22.2% +, 13.9% ++ and 41.7% +++). There was a statistically significant difference between HSP-70 staining scores with N status and final stages of the tumors (Chi-square, p = 0.03 and p = 0.026, respectively), while neither direct nor inverse correlation was detected for both parameters. PS2 staining scores showed no statistically significant relationship with tumor grade T, M status, perineural invasion, lymph and blood vessel invasion. In tumors with extensive staining with pS2, tumor stage tended to be low (Chi square, p = 0.024, Kendall Tau-b, r: -0.336, p = 0.036). There was a statistically significant difference and inverse correlation between tumors with extensive pS2 staining and tumors with less intense staining in terms of lymph node metastasis (Chi-square, p = 0.041, Kendall Tau: p = 0.024, r = -0,373). In the R0 resection group, in univariate analysis, we found that with higher scores of HSP-70 staining, the prognosis of the patient tended to improve. (Cox regression, p = 0.013). In multivariate analysis, HSP-70 expression was found to be an independent prognostic factor. We found no relationship between pS2 staining and patient survival.

Comparison of different surgical repairs in the treatment of experimental duodenal injuries

BACKGROUND In this experimental study, we aimed to investigate the results of different surgical repair methods for delayed reconstruction of severe experimental duodenal defects. METHODS A large duodenal defect with irregular and tagged margins covering about 50% of the circumference was created in the second part of duodenum of male Wistar rats. The effectiveness of primary repair, jejunal serosal patch, Roux-en-Y duodenojejunostomy, or expanded polytetrafluoroethylene patch repair techniques were investigated on the basis of survival and histologic assessment. RESULTS No significant survival benefit was observed between jejunal serosal patch, Roux-en-Y duodenojejunostomy, or expanded polytetrafluoroethylene patch repair techniques. But these repair modalities were associated with better survival rates than no-treatment or primary repair techniques. Complete coverage of the expanded polytetrafluoroethylene grafts by neomucosa consisting of columnar epithelium with villus formation was observed in surviving rats about 16 weeks after surgery. CONCLUSIONS Expanded polytetrafluoroethylene patch can be used in the repair of experimental large duodenal defects, which can not be repaired primarily.

Cooperative interaction of MUC1 with the HGF/c-Met pathway during hepatocarcinogenesis.

BackgroundHepatocyte growth factor (HGF) induced c-Met activation is known as the main stimulus for hepatocyte proliferation and is essential for liver development and regeneration. Activation of HGF/c-Met signaling has been correlated with aggressive phenotype and poor prognosis in hepatocellular carcinoma (HCC). MUC1 is a transmembrane mucin, whose over-expression is reported in most cancers. Many of the oncogenic effects of MUC1 are believed to occur through the interaction of MUC1 with signaling molecules. To clarify the role of MUC1 in HGF/c-Met signaling, we determined whether MUC1 and c-Met interact cooperatively and what their role(s) is in hepatocarcinogenesis.ResultsMUC1 and c-Met over-expression levels were determined in highly motile and invasive, mesenchymal-like HCC cell lines, and in serial sections of cirrhotic and HCC tissues, and these levels were compared to those in normal liver tissues. Co-expression of both c-Met and MUC1 was found to be associated with the differentiation status of HCC. We further demonstrated an interaction between c-Met and MUC1 in HCC cells. HGF-induced c-Met phosphorylation decreased this interaction, and down-regulated MUC1 expression. Inhibition of c-Met activation restored HGF-mediated MUC1 down-regulation, and decreased the migratory and invasive abilities of HCC cells via inhibition of β-catenin activation and c-Myc expression. In contrast, siRNA silencing of MUC1 increased HGF-induced c-Met activation and HGF-induced cell motility and invasion.ConclusionsThese findings indicate that the crosstalk between MUC1 and c-Met in HCC could provide an advantage for invasion to HCC cells through the β-catenin/c-Myc pathway. Thus, MUC1 and c-Met could serve as potential therapeutic targets in HCC.

Expression of transforming growth factor-beta-1 and p27Kip1 in pancreatic adenocarcinomas: relation with cell-cycle-associated proteins and clinicopathologic characteristics

BackgroundThe purpose of our study was to investigate the immunohistochemical expression of TGF-β1 and p27 in pancreatic adenocarcinomas and to compare the findings with the clinicopathological features and survival. We also aimed to evaluate the expression of TGF-β1 and p27 in the context of other cell cycle and proliferation markers such as cyclin D1 and Ki-67.MethodsWe examined TGF-β1 and p27 expression immunohistochemically in 63 cases of invasive ductal adenocarcinoma of the pancreas. Standard streptavidin-biotin immunperoxidase method was used for immunostaining and the stained slides were examined microscopically using semiquantitative criteria.ResultsTGF-β1 stained the cytoplasms of the tumor cells in 43 cases [68.3%]. There was a statistically significant difference among TGF-β1 staining scores in terms of clinicopathologic factors such as blood vessel invasion, stage and distant metastasis [p < 0.05]. Of the 63 tumors evaluated 23 [36.5%] were positive for p27 within the nucleus. An inverse correlation was found between p27 immunoreactivity and grade [p < 0.05]. But no significant correlation was found between p27 and other parameters. Among the patients with survival data 27 patients had RO resections and these cases were considered in survival analysis. In the univariate analysis, neither TGF-β1 nor p27 expression was related with patient survival.ConclusionOur findings suggest that in pancreatic carcinoma, TGF-β1 expression is related to tumor growth and metastasis. But it is not associated with cell cycle proteins. p27 expression is reduced in pancreatic adenocarcinomas and decreased protein levels of p27 may play a role in the differentiation of pancreatic cancer.

The effect of apoptotic activity, survivin, Ki-67, and P-glycoprotein expression on prognosis in pancreatic carcinoma.

Objectives: The pathogenetic mechanisms that regulate the aggressive behavior of pancreatic cancer still remain to be clarified. Alterations in the apoptotic pathway and proliferative activity of tumor cells as well as mechanisms contributing to the intrinsic drug resistance of pancreatic tumors have been investigated. Survivin is a recently described antiapoptotic protein, which, when overexpressed, is associated with worse prognosis in a majority of tumors. P-glycoprotein, a product of multidrug resistance gene-1 (MDR-1) was reported to be expressed in drug-resistant tumors. The purpose of this study was to investigate whether apoptosis, its regulation by survivin, tumor cell proliferation, and P-glycoprotein expression have a significant role on the biologic behavior of pancreatic adenocarcinoma. Methods: Tumors of 45 patients with pancreatic adenocarcinoma were studied for the detection of survivin, P-glycoprotein, and Ki-67 expression by immunohistochemical method and apoptotic index by TUNEL method. Immunohistochemical staining was scored and Ki-67 and apoptotic indices were expressed as percentage of stained cells. Results: Immunohistochemistry for survivin and P-glycoprotein revealed positive staining in 7 (15.4%) and 36 (79.5%) of the 45 tumors, respectively. The mean Ki-67 proliferative index was 43.75 ± 25.30%. The mean apoptotic index evaluated with the TUNEL method was 37.12 ± 34.55% for the whole group. We found no significant association between apoptotic index, expressions of survivin and P-glycoprotein, and clinicopathologic variables and survival. Conclusions: Apoptotic activity, survivin, and P-glycoprotein expression failed to predict the disease extent and biologic behavior in pancreatic adenocarcinoma in our cases.

Incarcerated inferior lumbar (Petit's) hernia.

Petits hernia is an uncommon abdominal wall defect in the inferior lumbar triangle. Colonic incarceration through the inferior lumbar triangle, which causes mechanical obstructive symptoms, necessitates particular diagnostic and management strategy. We present a rare case of inferior lumbar hernia, leading to mechanical bowel obstruction, successfully treated with prosthetic mesh reinforcement repair.

Genome-Wide Transcriptional Reorganization Associated with Senescence-to-Immortality Switch during Human Hepatocellular Carcinogenesis

Senescence is a permanent proliferation arrest in response to cell stress such as DNA damage. It contributes strongly to tissue aging and serves as a major barrier against tumor development. Most tumor cells are believed to bypass the senescence barrier (become “immortal”) by inactivating growth control genes such as TP53 and CDKN2A. They also reactivate telomerase reverse transcriptase. Senescence-to-immortality transition is accompanied by major phenotypic and biochemical changes mediated by genome-wide transcriptional modifications. This appears to happen during hepatocellular carcinoma (HCC) development in patients with liver cirrhosis, however, the accompanying transcriptional changes are virtually unknown. We investigated genome-wide transcriptional changes related to the senescence-to-immortality switch during hepatocellular carcinogenesis. Initially, we performed transcriptome analysis of senescent and immortal clones of Huh7 HCC cell line, and identified genes with significant differential expression to establish a senescence-related gene list. Through the analysis of senescence-related gene expression in different liver tissues we showed that cirrhosis and HCC display expression patterns compatible with senescent and immortal phenotypes, respectively; dysplasia being a transitional state. Gene set enrichment analysis revealed that cirrhosis/senescence-associated genes were preferentially expressed in non-tumor tissues, less malignant tumors, and differentiated or senescent cells. In contrast, HCC/immortality genes were up-regulated in tumor tissues, or more malignant tumors and progenitor cells. In HCC tumors and immortal cells genes involved in DNA repair, cell cycle, telomere extension and branched chain amino acid metabolism were up-regulated, whereas genes involved in cell signaling, as well as in drug, lipid, retinoid and glycolytic metabolism were down-regulated. Based on these distinctive gene expression features we developed a 15-gene hepatocellular immortality signature test that discriminated HCC from cirrhosis with high accuracy. Our findings demonstrate that senescence bypass plays a central role in hepatocellular carcinogenesis engendering systematic changes in the transcription of genes regulating DNA repair, proliferation, differentiation and metabolism.

BENEFICIAL EFFECTS OF PENTOXIFYLLINE PRETREATMENT IN NON-HEART-BEATING DONORS IN RATS

BACKGROUND Pentoxifylline (PTX) pretreatment of recipients was shown to protect against liver graft failure from ischemia-reperfusion injury after orthotopic rat liver transplantation. It has also been shown that PTX protects against normothermic ischemia-reperfusion injury to the liver in lobar ischemia model in the rat. Whether PTX can benefit the liver procured from non-heart-beating donors (NHBDs) with up to 9 hr of cold ischemia is unknown. METHODS Donor and recipient rats were pretreated with intraperitoneal PTX (50 mg/kg) 1 hr before cardiac arrest and transplantation, respectively. Grafts were transplanted 0, 30, and 60 min after cardiac arrest with additional 1 and 9 hr of cold ischemia in both PTX-pretreated or untreated (control) groups (10 rats per group). PTX (25 mg/kg/day) was continuously given to the surviving rats for 5 days postoperatively. Recipient survival rates, serum enzyme levels, and histopathological examination of postreperfusion liver biopsies were all analyzed. RESULTS The survival rates, serum enzyme levels, and postreperfusion histology were significantly improved in groups pretreated with PTX compared to the controls. CONCLUSION Donor and recipient PTX pretreatment significantly improves the viability of the liver grafts procured from NHBDs.