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1.  PAR-3 Knockdown Enhances Adhesion Rate of PANC-1 Cells via Increased Expression of Integrinαv and E-Cadherin 
PLoS ONE  2014;9(4):e93879.
The balance between the adhesion of cancer cells to extracellular matrix and their migratory potential, as well as their proteolytic activity, are important parameters that determine cancer cells invasiveness and metastasis. Since thrombin has been implicated in cancer progression, we studied the role(s) of thrombin-activated receptors in the adhesion process. We stably knocked down proteinase-activated receptors (PARs) -1, or -3 in human pancreatic adenocarcinoma PANC-1 cells. PANC-1 cells exhibit rapid adhesion to cell culture treated plastic and much faster kinetics of adhesion to Matrigel coated surface. Knockdown of PAR-1 had no effect on cells' adhesiveness, while PAR-3 knockdowns (KDs) exhibited much faster adhesion kinetics. PAR-3 KDs also exhibited slower in vitro wound closure than vector-control and PAR-1 KD cells. To study the molecular mechanism(s) of PAR-3 KD cells' enhanced rate of adhesion, we assayed the expression of the molecules that mediate cell-surface and cell-cell adhesion. ITGαv, as well as ITGα6 and ITGα10 mRNAs, were greatly enriched (>40-fold) in a rapidly-adhering sub-population of PAR-3 KD cells. The whole population of both PAR-1 and -3 KDs exhibited enhanced expression of a number of integrins (ITGs) mRNAs. However, ITGαv mRNA and protein expression was increased in PAR-3 KD and markedly decreased in PAR-1 KD. PAR-3 KD cells also expressed more E-cadherin mRNA and protein. The enhanced adhesion kinetics of PAR-3 KDs was almost fully inhibited by calcium chelation, or by a HAV-motive decapeptide that affects E-cadherin intermolecular interactions. We propose that the enhanced rate of adhesion of PAR-3 KDs results from enhanced expression of E-cadherin, leading to a greater adhesion of free-floating cells to cells rapidly bound to the surface via their integrins, and particularly ITGαv.
doi:10.1371/journal.pone.0093879
PMCID: PMC3974847  PMID: 24699825
2.  Tissue Specific DNA Methylation in Normal Human Breast Epithelium and in Breast Cancer 
PLoS ONE  2014;9(3):e91805.
Cancer is a heterogeneous and tissue-specific disease. Thus, the tissue of origin reflects on the natural history of the disease and dictates the therapeutic approach. It is suggested that tissue differentiation, mediated mostly by epigenetic modifications, could guide tissue-specific susceptibility and protective mechanisms against cancer. Here we studied breast specific methylation in purified normal epithelium and its reflection in breast cancers. We established genome wide methylation profiles of various normal epithelial tissues and identified 110 genes that were differentially methylated in normal breast epithelium. A number of these genes also showed methylation alterations in breast cancers. We elaborated on one of them, TRIM29 (ATDC), and showed that its promoter was hypo-methylated in normal breast epithelium and heavily methylated in other normal epithelial tissues. Moreover, in breast carcinomas methylation increased and expression decreased whereas the reverse was noted for multiple other carcinomas. Interestingly, TRIM29 regulation in breast tumors clustered according to the PAM50 classification. Thus, it was repressed in the estrogen receptor positive tumors, particularly in the more proliferative luminal B subtype. This goes in line with previous reports indicating tumor suppressive activity of TRIM29 in estrogen receptor positive luminal breast cells in contrast to oncogenic function in pancreatic and lung cancers. Overall, these findings emphasize the linkage between breast specific epigenetic regulation and tissue specificity of cancer.
doi:10.1371/journal.pone.0091805
PMCID: PMC3961270  PMID: 24651077
3.  Serum DNA methylation for monitoring response to neoadjuvant chemotherapy in breast cancer patients 
Patients with large or nonoperable breast cancers often receive neoadjuvant chemotherapy to facilitate full resection of the tumor and enable conservation of the breast. However, currently available methods for evaluation of response during therapy are limited and the actual effect of the treatment is only recognized at surgery upon completion of chemotherapy. Timely assessment of response could allow individual tailoring of the treatment and save noneffective drugs and unnecessary toxicity. Here, we suggest that tumor derived DNA methylation in the serum may reflect changes in tumor burden and allow early recognition of responders versus nonresponders. In this pilot study, we collected 7 consecutive serum samples from 52 patients with locally advanced breast cancer during neoadjuvant chemotherapy. We selected RASSF1, which was methylated in more than 80% of the tumors, for serum analysis. Using the “methylation sensitive PCR and high resolution melting,” we detected RASSF1 methylation in the serum of 21 patients prior to therapy. In four patients who achieved complete pathological response, RASSF1 methylation in the serum became undetectable early during therapy. In contrast, in 17 patients that had partial or minimal pathological response, serum RASSF1 methylation persisted longer or throughout the treatment (complete versus partial response p = 0.02). These findings support further development of this assay for monitoring response during neoadjuvant therapy.
doi:10.1002/ijc.27526
PMCID: PMC3904188  PMID: 22407753
locally advanced breast cancer; neoadjuvant therapy; serum DNA methylation; monitoring response
4.  Tumor differentiation as related to sentinel lymph node status in gastric cancer 
AIM: To investigate the influence of tumor grade on sentinel lymph node (SLN) status in patients with gastric cancer (GC).
METHODS: We retrospectively studied 71 patients with GC who underwent SLN mapping during gastric surgery to evaluate the relationship between SLN status and tumor grade.
RESULTS: Poorly differentiated tumors were detected in 50/71 patients, while the other 21 patients had moderately differentiated tumors. SLNs were identified in 58/71 patients (82%). In 41 of the 58 patients that were found to have stained nodes (70.7%), the tumor was of the poorly differentiated type (group I), while in the remaining patients with stained nodes 17/58 (29.3%), the tumor was of the moderately differentiated type (group II). Positive SLNs were found in 22/41 patients in group I (53.7%) and in 7/17 patients in group II (41.2%) (P = 0.325). The rate of positivity for the SLNs in the two groups (53.7% vs 41.2%) was not statistically significant (P = 0.514).
CONCLUSION: Most of our patients were found to have poorly differentiated adenocarcinoma of the stomach and there was no correlation between tumor grade and SLN involvement.
doi:10.4240/wjgs.v6.i1.1
PMCID: PMC3951807  PMID: 24627734
Gastric cancer; Sentinel lymph nodes; Tumor differentiation; Sentinel lymph node mapping; Prognosis
5.  Role of IL-10 in the progression of kidney disease 
Interleukin-10 (IL-10), a cytokine with anti-inflammatory and immunomodulatory functions, regulates the biology of B and T cells. The present review describes the role of IL-10 in normal renal physiology, during acute kidney injury and in the development of chronic renal failure. We further discuss IL-10-induced cellular and molecular pathways and their link to the progression of kidney injury.
doi:10.5500/wjt.v3.i4.91
PMCID: PMC3879528  PMID: 24392313
Transforming growth factor-β; Cystatin C; Interleukin-10 receptor; End-stage renal disease; Mesangial cell proliferation; Epithelial-to-mesenchymal transition; Allograft rejection
6.  Low Serum Concentration of Obestatin as a Predictor of Mortality in Maintenance Hemodialysis Patients 
BioMed Research International  2013;2013:796586.
Obestatin, a proposed anorexigenic gut hormone, has been shown to have a number of beneficial cardiotropic effects in experimental studies. We hypothesized that obestatin alteration in hemodialysis patients may link to clinical outcomes. This cross-sectional study with prospective followup for almost 4 years was performed on 94 prevalent hemodialysis patients. Obestatin, leptin, proinflammatory cytokines (tumor necrosis factor-α [TNF-α], interleukin-6, and various nutritional markers were measured. Patients with low obestatin levels, defined as a level less than median, had a worse all-cause mortality and cardiovascular mortality. The crude all-cause (HR 2.23, 95% CI 1.17 to 4.24) and cardiovascular mortality hazard ratios (HR 4.03, 95% CI 1.27 to 12.76) in these patients continued to be significant after adjustment for various confounders for all-cause mortality. Across the four obestatin-TNF-α categories, the group with low obestatin and high TNF-α (above median level) exhibited a worse outcome in both all-cause mortality and cardiovascular mortality. Clinical characteristics of patients in low obestatin high TNF-α group did not differ from other obestatin-TNF-α categorized groups. In summary, low serum obestatin concentration is an independent predictor of mortality in prevalent hemodialysis patients. Novel interactions were observed between obestatin and TNF-α, which were associated with mortality risk, especially those due to cardiovascular causes.
doi:10.1155/2013/796586
PMCID: PMC3786512  PMID: 24102059
7.  Knock-down of plasminogen-activator inhibitor-1 enhances expression of E-cadherin and promotes epithelial differentiation of human pancreatic adenocarcinoma cells 
Journal of cellular physiology  2012;227(11):3621-3628.
High levels of plasminogen activator inhibitor-1 (PAI-1), which is produced by stromal, endothelial and cancer cells and has multiple complex effects on cancers, correlate with poor cancer prognosis. To more definitively study the role of endogenously produced PAI-1 in human pancreatic adenocarcinoma (PAC) PANC-1 cell line biology, we used anti-PAI-1 shRNA to create stable PAI-1 deficient cells (PD-PANC-1s). PD-PANC-1s exhibited a heterogeneous morphology. While the majority of cells exhibited a cuboidal shape similar to the parental PANC-1 or the vector-infected control cells, numerous large cells with long filopodia and a neuronal-like appearance were observed. Although both Vector-control cells and PD-PANC-1s expressed mRNAs that are characteristic of mesenchymal, neural and epithelial phenotypes, epithelial marker RNAs were up-regulated (e.g. E-cadherin, 32-fold) whereas mesenchymal marker RNAs were down-regulated (e.g. Thy1, 9-fold) in PD-PANC-1s, suggesting mesenchymal-to-epithelial transition. Neural markers exhibited both up- and down-regulation. Immunocytochemistry indicated that epithelial-like PD-PANC-1s expressed E-cadherin and β-catenin in significantly more cells, while neural-like cells exhibited robust expression of organized β-3-tubulin. PAI-1 and E-cadherin were rarely co-expressed in the same cells. Indeed, examination of PAI-1 and E-cadherin mRNAs expression in additional cell lines yielded clear inverse correlation. Indeed, infection of Colo357 PAC cells (that exhibit high expression of E-cadherin) with PAI-1-expressing adenovirus led to a marked decrease in E-cadherin expression and to enhanced migration of cells from clusters. Our results suggest that endogenous PAI-1 suppresses expression of E-cadherin and differentiation in PAC cells in vitro, supporting its negative impact on tumor prognosis.
doi:10.1002/jcp.24068
PMCID: PMC3469200  PMID: 22331587
Plasminogen activator inhibitor 1; E-cadherin; Differentiation; Human pancreatic adenocarcinoma
8.  Gene expression subtraction of non-cancerous lung from smokers and non-smokers with adenocarcinoma, as a predictor for smokers developing lung cancer 
Background
Lung cancer is the commonest cause of cancer death in developed countries. Adenocarcinoma is becoming the most common form of lung cancer. Cigarette smoking is the main risk factor for lung cancer. Long-term cigarettes smoking may be characterized by genetic alteration and diffuse injury of the airways surface, named field cancerization, while cancer in non-smokers is usually clonally derived. Detecting specific genes expression changes in non-cancerous lung in smokers with adenocarcinoma may give us instrument for predicting smokers who are going to develop this malignancy.
Objectives
We described the gene expression in non-cancerous lungs from 21 smoker patients with lung adenocarcinoma and compare it to gene expression in non-cancerous lung tissue from 10 non-smokers with primary lung adenocarcinoma.
Methods
Total RNA was isolated from peripheral non-cancerous lung tissue. The cDNA was hybridized to the U133A GeneChip array. Hierarchical clustering analysis on genes obtained from smokers and non-smokers, after subtracting were exported to the Ingenuity Pathway Analysis software for further analysis.
Results
The genes subtraction resulted in disclosure of 36 genes with high score. They were subsequently mapped and sorted based on location, cellular components, and biochemical activity. The gene functional analysis disclosed 20 genes, which are involved in cancer process (P = 7.05E-5 to 2.92E-2).
Conclusion
Detected genes may serve as a predictor for smokers who may be at high risk of developing lung cancer. In addition, since these genes originating from non-cancerous lung, which is the major area of the lungs, a sample from an induced sputum may represent it.
doi:10.1186/1756-9966-27-45
PMCID: PMC2570656  PMID: 18811983
9.  Fatal Rickettsia conorii subsp. israelensis Infection, Israel 
Emerging Infectious Diseases  2008;14(5):821-824.
Fatal Rickettsia conorii subsp. israelensis Infection, Israel
Underdiagnosis of fatal spotted fever may be attributed to nonspecific clinical features and insensitive acute-phase serologic studies. We describe the importance of molecular and immunohistochemical methods in establishing the postmortem diagnosis of locally acquired Israeli spotted fever due to Rickettsia conorii subsp. israelensis in a traveler returning to Israel from India.
doi:10.3201/eid1405.071278
PMCID: PMC2600240  PMID: 18439372
PCR; Israel; epidemiology; Rickettsia infections; Rickettsia conorii; genetics; pathogenicity; travel; dispatch
10.  The Immunohistochemistry Profile of Lymphocytic Gastritis in Celiac Disease and Helicobacter Pylori Infection: Interplay between Infection and Inflammation 
Mediators of Inflammation  2007;2007:81838.
Lymphocytic gastritis (LG) is associated with helicobacter pylori (Hp) and celiac disease (CD). We aimed to clarify the relationship between Hp infection and CD by defining a unique histopathology profile of LG in these two diseases. Forty patients who underwent upper endoscopy were divided into four groups: eight controls, ten active CD patients without Hp, twelve CD negative with Hp, and ten active CD with Hp infection. Antral samples were assessed by immunohistochemical staining for CD20, CD3, CD4, CD8, CD57, CNA42, and Ki67 for lymphoid aggregates, intraepithelial lymphocytes (IELs) number, density of lamina propria (LP) lymphocytes, and inflammatory glandular involvement. Only IELs positive for CD3 and CD8 were increased significantly in CD patients with or without Hp infection. Hp did not contribute to the number of CD8 IELs. In complicated cases with Hp and suspicious for CD, the number of CD8+ IELs hints toward a CD rather than Hp infection.
doi:10.1155/2007/81838
PMCID: PMC2222270  PMID: 18274643
11.  Induction of the chemokine stromal-derived factor-1 following DNA damage improves human stem cell function 
Journal of Clinical Investigation  2000;106(11):1331-1339.
The chemokine stromal-derived factor-1 (SDF-1) controls many aspects of stem cell function. Details of its regulation and sites of production are currently unknown. We report that in the bone marrow, SDF-1 is produced mainly by immature osteoblasts and endothelial cells. Conditioning with DNA-damaging agents (ionizing irradiation, cyclophosphamide, and 5-fluorouracil) caused an increase in SDF-1 expression and in CXCR4-dependent homing and repopulation by human stem cells transplanted into NOD/SCID mice. Our findings suggest that immature osteoblasts and endothelial cells control stem cell homing, retention, and repopulation by secreting SDF-1, which also participates in host defense responses to DNA damage.
PMCID: PMC381461  PMID: 11104786
12.  The chemokine SDF-1 stimulates integrin-mediated arrest of CD34+ cells on vascular endothelium under shear flow 
Journal of Clinical Investigation  1999;104(9):1199-1211.
The chemokine SDF-1 plays a central role in the repopulation of the bone marrow (BM) by circulating CD34+ progenitors, but the mechanisms of its action remain obscure. To extravasate to target tissue, a blood-borne cell must arrest firmly on vascular endothelium. Murine hematopoietic progenitors were recently shown in vivo to roll along BM microvessels that display selectins and integrins. We now show that SDF-1 is constitutively expressed by human BM endothelium. In vitro, human CD34+ cells establish efficient rolling on P-selectin, E-selectin, and the CD44 ligand hyaluronic acid under physiological shear flow. ICAM-1 alone did not tether CD34+ cells under flow, but, in the presence of surface-bound SDF-1, CD34+ progenitors rolling on endothelial selectin rapidly developed firm adhesion to the endothelial surface, mediated by an interaction between ICAM-1 and its integrin ligand, which coimmobilized with SDF-1. Human CD34+ cells accumulated efficiently on TNF-activated human umbilical cord endothelial cells in the absence of SDF-1, but they required immobilized SDF-1 to develop firm integrin-mediated adhesion and spreading. In the absence of selectins, SDF-1 also promoted VLA-4–mediated, Gi protein–dependent tethering and firm adhesion to VCAM-1 under shear flow. To our knowledge, this is the first demonstration that SDF-1 expressed on vascular endothelium is crucial for translating rolling adhesion of CD34+ progenitors into firm adhesion by increasing the adhesiveness of the integrins VLA-4 and LFA-1 to their respective endothelial ligands, VCAM-1 and ICAM-1.
PMCID: PMC409822  PMID: 10545519

Results 1-12 (12)