The remodeling index as a practical tool for predicting progression rate during early stages of chronic liver disease

Chronic liver disease (CLD) constitutes a major cause of morbidity and mortality worldwide. Follow-up studies have documented that the majority of patients with CLD never reach the cirrhotic stage, while others display a higher progression rate leading to liver failure at relatively short intervals. This phenomenon has never been adequately explained. Recent evidence suggests that the renin-angiotensin system (RAS) is a major coordinator of chronic liver inflammation and subsequent fibrosis development, a process often termed hepatic remodeling. Combining these data with the “natural neutralizing antibodies theory” led us to the assumption that there could be an intrinsic anti-remodeling mechanism consisted of natural antibodies against components of the RAS. Varying degrees of activation of this defense mechanism could account for the variability in disease progression rate among patients with CLD. Identifying the main components of this mechanism allowed us to develop a ratio, designated remodeling index, as a measure of an individual's predilection towards cirrhosis. We believe that this index could be used as a safe, non-invasive and cost effective tool for assessing progression rate in normotensive patients with early CLD, thus alleviating the need for repeated liver biopsies.     

The Protein Kinase TPL2 Is Essential for ERK1/ERK2 Activation and Cytokine Gene Expression in Airway Epithelial Cells Exposed to Pathogen-Associated Molecular Patterns (PAMPs)

The epithelium forms a physical barrier important to the detection of pathogens. P. aeruginosa infections are frequently encountered in Cystic Fibrosis lungs, lead to ERK1/ERK2 activation and contribute to tissue destruction. We report here that in bronchial airway epithelial cells (BEAS-2B), diffusible material from P. aeruginosa and TLR2, TLR3 and TLR5 ligands activates ERK1/ERK2 via the protein kinase TPL2 and not the growth factor receptor EGFR. Activation of TPL2 by these agonists in airway epithelial cells requires the protein kinases TAK1 and IKKβ in accordance with the previously reported model of activation of TPL2 in macrophages. Inhibition of TPL2 activity with a pharmacological inhibitor (Compound 1) not only prevented ERK1/ERK2 activation but also decreased cytokine synthesis in response to pathogen-associated molecular patterns. These results suggest that inhibition of the protein kinase TPL2 is an attractive strategy to decrease inflammation in the lungs when it is not warranted.     

Inhibition of tyrosine phenol-lyase from Citrobacter freundii by 2-azatyrosine and 3-azatyrosine.

The interactions of 2-azatyrosine and 3-azatyrosine with tyrosine phenol-lyase (TPL) from Citrobacter freundii have been examined. 2-Aza-DL-tyrosine and 3-aza-DL-tyrosine were synthesized by standard methods of amino acid synthesis, while the L-isomers were prepared from 3-hydroxypyridine and 2-hydroxypyridine, respectively, with TPL (Watkins, E. B., and Phillips, R. S. (2001) Bioorg. Med. Chem. Lett. 11, 2099-2100). 3-Azatyrosine was examined as a potential transition state analogue inhibitor of TPL. Both compounds were found to be competitive inhibitors of TPL, with K(i) values of 3.4 mM and 135 microM for 3- and 2-aza-L-tyrosine, respectively. Thus, 3-azatyrosine does not act as a transition state analogue, possibly due to the lack of tetrahedral geometry at C-1. However, 2-aza-L-tyrosine is the most potent competitive inhibitor of TPL found to date. The K(i) value of 2-aza-L-tyrosine is half that of 2-aza-DL-tyrosine, indicating that the D-enantiomer is inactive as an inhibitor. Neither azatyrosine isomer was shown to be a substrate for beta-elimination, based on coupled assays with lactate dehydrogenase and on HPLC measurements. Both isomers of azatyrosine form equilibrium mixtures of external aldimine and quinonoid intermediates when they bind to TPL. However, 2-azatyrosine reacts about 10-fold faster to form a quinonoid intermediate than does 3-azatyrosine. Since 2-azatyrosine is in the zwitterion or phenolate ion form at all the pH values examined, the strong binding of this compound suggests that L-tyrosine may be bound to the active site of TPL as the phenolate anion.     

Serum MicroRNAs as Potential Biomarkers for Early Diagnosis of Hepatitis C Virus-Related Hepatocellular Carcinoma in Egyptian Patients

Circulating microRNAs are deregulated in liver fibrosis and hepatocellular carcinoma (HCC) and are candidate biomarkers. This study investigated the potential of serum microRNAs; miR-19a, miR-296, miR-130a, miR-195, miR-192, miR-34a, and miR-146a as early diagnostic biomarkers for hepatitis C virus (HCV)-related HCC. As how these microRNAs change during liver fibrosis progression is not clear, we explored their serum levels during fibrosis progression in HCV-associated chronic liver disease (CLD) and if they could serve as non-invasive biomarkers for fibrosis progression to HCC. 112 Egyptian HCV-HCC patients, 125 non-malignant HCV-CLD patients, and 42 healthy controls were included. CLD patients were subdivided according to Metavir fibrosis-scoring. Serum microRNAs were measured by qRT-PCR custom array. Serum microRNAs were deregulated in HCC versus controls, and except miR-130a, they were differentially expressed between HCC and CLD or late fibrosis (F3-F4) subgroup. Serum microRNAs were not significantly different between individual fibrosis-stages or between F1-F2 (early/moderate fibrosis) and F3-F4. Only miR-19a was significantly downregulated from liver fibrosis (F1-F3) to cirrhosis (F4) to HCC. Individual microRNAs discriminated HCC from controls, and except miR-130a, they distinguished HCC from CLD or F3-F4 patients by receiver-operating-characteristic analysis. Multivariate logistic analysis revealed a panel of four microRNAs (miR-19a, miR-195, miR-192, and miR-146a) with high diagnostic accuracy for HCC (AUC = 0.946). The microRNA panel also discriminated HCC from controls (AUC = 0.949), CLD (AUC = 0.945), and F3-F4 (AUC = 0.955). Studied microRNAs were positively correlated in HCC group. miR-19a and miR-34a were correlated with portal vein thrombosis and HCC staging scores, respectively. In conclusion, studied microRNAs, but not miR-130a, could serve as potential early biomarkers for HCC in high-risk groups, with miR-19a as a biomarker for liver fibrosis progression to cirrhosis to HCC. We identified a panel of four serum microRNAs with high accuracy in HCC diagnosis. Additional studies are required to confirm this panel and test its prognostic significance.     

2-Hydroxyestradiol slows progression of experimental polycystic kidney disease

Male gender is a risk factor for progression of polycystic kidney disease (PKD). 17β-Estradiol (E2) protects experimentally, but clinical use is limited by adverse effects. Novel E2 metabolites provide many benefits of E2 without stimulating the estrogen receptor, and thus may be safer. We hypothesized that E2 metabolites are protective in a model of PKD. Studies were performed in male control Han:SPRD rats, and in cystic males treated with orchiectomy, 2-methoxyestradiol, 2-hydroxyestradiol (2-OHE), or vehicle, from age 3 to 12 wk. Cystic rats exhibited renal functional impairment (～50% decrease in glomerular filtration and renal plasma flow rates, P < 0.05) and substantial cyst development (20.5 ± 2.0% of cortex area). 2-OHE was the most effective in limiting cysts (6.0 ± 0.7% of cortex area, P < 0.05 vs. vehicle-treated cystic rats) and preserving function, in association with suppression of proliferation, apoptosis, and angiogenesis markers. Downregulation of p21 expression and increased expression of Akt, the mammalian target of rapamycin (mTOR), and some of its downstream effectors were significantly reversed by 2-OHE. Thus, 2-OHE limits disease progression in a cystic rodent model. Mechanisms include reduced renal cell proliferation, apoptosis, and angiogenesis. These effects may be mediated, at least in part, by preservation of p21 and suppression of Akt and mTOR. Estradiol metabolites may represent a novel, safe intervention to slow progression of PKD.     

Polynitroxyl-albumin (PNA) enhances myocardial infarction therapeutic effect of tempol in rat hearts subjected to regional ischemia-reperfusion

Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl, TPL), a low molecular weight stable nitroxyl radical (nitroxide), has been demonstrated in many in vitro and in vivo models to have protective effects against oxidative stress. The beneficial effect of TPL, however, is limited because of its short life-time in tissues. We have previously shown that polynitroxylated macromolecules such as polynitroxyl-human serum albumin (PNA) enable maintaining a sustained concentration of TPL for longer periods in tissues. PNA itself has previously been shown to inhibit ischemia-reperfusion (I/R) injury in the gut and to potentiate the activity of TPL. The aim of the present study was (i) to select an optimum formulation of PNA + TPL for therapeutic applications using in vivo EPR spectroscopy and (ii) to evaluate the efficacy of the PNA + TPL formulation in preventing I/R injury to heart, in an in vivo rat model. Rats were subjected to 45 min occlusion of the left anterior descending (LAD) coronary artery followed by 120 min reperfusion. PNA (100 mg/ml) + TPL (10 mg/ml), human serum albumin (HSA, 100 mg/ml) + TPL (10 mg/ml), or saline were injected 5 min before ischemia (3 ml/kg BW, i.v.) and 5 min before reperfusion (3 ml/kg BW, i.v.), followed by a 4 ml/kg BW infusion over 2 h reperfusion. Myocardial risk and infarct regions were then estimated. The results showed that the infarct volume, expressed as a percentage of the risk region, in the group treated with PNA + TPL was 39.7 +/- 3.1%, which was significantly smaller than for the saline (51.3 +/- 3.5%) or HSA + TPL (48.4 +/- 1.4%) groups. The results demonstrate that the PNA + TPL combination is very effective in reducing myocardial ischemia-reperfusion injury.     

TPL2 signalling: From Toll-like receptors-mediated ERK1/ERK2 activation to Cystic Fibrosis lung disease

Cystic Fibrosis (CF) is the most common lethal genetic recessive disorder, with a carrier frequency of 1 in 27 among North American Caucasians. Mitogen-activated protein kinases (MAPKs) and pro-inflammatory cytokines have crucial functions in the innate immune response of epithelial cells. They determine the inflammation status and the host response to pathogenic infections. However, in CF, bacterial-driven inflammation leads to tissue destruction, reduction in lung function and mortality. Recognition of invading pathogens is mediated in part by Toll-like receptors (TLR) activation of intracellular signalling cascade leading to cytokines’ synthesis. The protein kinase Tumour Progression Locus 2 (TPL2) is a key molecule in relaying inflammatory stimuli to ERK1/ERK2 MAPKs. In this review, we summarized the recent findings on TPL2 signalling and how TPL2 can contribute to the excessive inflammation found in CF. Pharmacologically targeting this kinase could have a significant benefit for CF patients dealing with chronic bacterial infections such as Pseudomonas aeruginosa.This article is part of a Directed Issue entitled: Cystic Fibrosis: From o-mics to cell biology, physiology, and therapeutic advances.     

Inhibition of Tyrosine Kinase Receptor Tie2 Reverts HCV-Induced Hepatic Stellate Cell Activation

Background

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease (CLD) and is frequently linked to intrahepatic microvascular disorders. Activation of hepatic stellate cells (HSC) is a central event in liver damage, due to their contribution to hepatic renewal and to the development of fibrosis and hepatocarcinoma. During the progression of CLDs, HSC attempt to restore injured tissue by stimulating repair processes, such as fibrosis and angiogenesis. Because HSC express the key vascular receptor Tie2, among other angiogenic receptors and mediators, we analyzed its involvement in the development of CLD.

Methods

Tie2 expression was monitored in HSC cultures that were exposed to media from HCV-expressing cells (replicons). The effects of Tie2 blockade on HSC activation by either neutralizing antibody or specific signaling inhibitors were also examined.

Results

Media from HCV-replicons enhanced HSC activation and invasion and upregulated Tie2 expression. Notably, the blockade of Tie2 receptor (by a specific neutralizing antibody) or signaling (by selective AKT and MAPK inhibitors) significantly reduced alpha-smooth muscle actin (α-SMA) expression and the invasive potential of HCV-conditioned HSC.

Conclusions

These findings ascribe a novel profibrogenic function to Tie2 receptor in the progression of chronic hepatitis C, highlighting the significance of its dysregulation in the evolution of CLDs and its potential as a novel therapeutic target.     

Unexpected structure of the Ca2+-regulatory region from soybean calcium-dependent protein kinase-alpha

Calcium-dependent protein kinases (CDPKs) are an extensive class of multidomain Ca(2+)-regulated enzymes from plants and protozoa. In vivo the so-called calmodulin-like domain (CLD) of CDPK binds intramolecularly to the junction domain (JD), which exhibits both kinase-inhibitory and CLD binding properties. Here we report the high resolution solution structure of the calcium-regulatory region from soybean CDPK-alpha determined in the presence of a peptide encompassing the JD. The structure of both lobes of CLD resembles that of related helix-loop-helix Ca(2+)-binding proteins. NMR chemical shift mapping studies demonstrate that the JD induces significant structural changes in isolated Ca(2+)-CLD, particularly the C-terminal domain, although a stable complex is not formed. A CLD solution structure calculated on the basis of NMR data and long range fluorescence resonance energy transfer distances reveals an activated state with both lobes positioned side by side, similar to calcineurin B rather than calmodulin, highlighting the possible pitfall of assigning function purely from sequence information.     

The nitroxide Tempol modulates anthracycline resistance in breast cancer cells

The occurrence of multidrug resistance (MDR) is the major obstacle to successful anthracycline-based cancer chemotherapy. In the present study, we assessed the effects of Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl, TPL), a piperidine nitroxide with growth-inhibitory properties in tumor cell lines, on a number of molecular mechanisms involved in the resistance of human breast adenocarcinoma cell lines to doxorubicin (DOX). Cytotoxicity studies in MCF-7 wildtype and their MDR variant MCF-7 Adr(R) cells showed a synergistic effect between TPL and DOX when exposure to TPL preceded or was simultaneous with DOX treatment in MCF-7 Adr(R) cells. This effect of TPL seems to be due in part to its ability to increase peroxide levels and to deplete cellular glutathione pools. In addition, TPL increased DOX accumulation in MCF-7 Adr(R) cells by interfering with P-glycoprotein-mediated DOX efflux, as evidenced using a specific antibody that recognizes the active form of the protein. TPL was also found to affect the expression levels of proteins involved in response to drug treatment (e.g., p53, bcl2, bax, p21). Taken together, our results indicate that TPL is a potential new agent that may improve the clinical effect of DOX in tumors exhibiting a MDR phenotype.     

Structure and mechanism of tryptophan indole-lyase and tyrosine phenol-lyase

Tyrosine phenol-lyase (TPL) and tryptophan indole-lyase (Trpase) catalyse the reversible hydrolytic cleavage of L-tyrosine or L-tryptophan to phenol or indole, respectively, and ammonium pyruvate. These enzymes are very similar in sequence and structure, but show strict specificity for their respective physiological substrates. We have mutated the active site residues of TPL (Thr(124), Arg(381), and Phe(448)) to those of Trpase and evaluated the effects of the mutations. Tyr(71) in Citrobacter freundii TPL, and Tyr(74) in E. coli Trpase, are essential for activity with both substrates. Mutation of Arg(381) of TPL to Ala, Ile, or Val (the corresponding residues in the active site of Trpase) results in a dramatic decrease in L-Tyr beta-elimination activity, with little effect on the activity of other substrates. Arg(381) may be the catalytic base with pK(a) of 8 seen in pH-dependent kinetic studies. T124D TPL has no measureable activity with L-Tyr or 3-F-L-Tyr as substrate, despite having high activity with SOPC. T124A TPL has very low but detectable activity, which is about 500-fold less than wild-type TPL, with L-Tyr and 3-F-L-Tyr. F448H TPL also has very low activity with L-Tyr. None of the mutant TPLs has any detectable activity with L-Trp as substrate. H463F Trpase also exhibits low activity with L-Trp, but retains high activity with other substrates. Thus, additional residues remote from the active site may be needed for substrate specificity. Both Trpase and TPL may react by a rare S(E)2-type mechanism.     

Indirect role of Ca2+ in the assembly of extracellular matrix proteins

We have investigated through molecular dynamics the binding properties at the interface between the C-type lectin subdomain (CLD) of aggrecan and the fibronectin type III domains (^4-5FnIII) of tenasin, in particular the mechanistically unknown, but essential role of Ca²⁺ in the binding. The binding between the CLD and ^4-5FnIII is critical in cross-linking aggrecan, hyaluronan, and tenascin to form extended protein networks found in tissues such as cartilage. None of the structurally resolved Ca²⁺ ions in the complex of the CLD and ^4-5FnIII is directly bridging the two proteins. However, one of the Ca²⁺ ions (Ca2) is found to play the role of maintaining the structure of the L4 loop at the CLD binding surface, thus facilitating a high affinity binding between the CLD and ^4-5FnIII. Removal of this Ca²⁺ ion causes a drastic structural change in the L4 loop, which presumably hinders the binding of the CLD to the ^4-5FnIII. The other bound Ca²⁺ ions (Ca1 and Ca3) have no significant effect on the structure of the CLD binding surface, and thus are not expected to affect the binding. Our results might also suggest that the role of Ca2 in maintaining the structure of the L4 loop is most important during the binding process. Once the complex is formed, the dependence of the complex on the structuring role of Ca2 is reduced. In response to tensile force, the CLD and ^4-5FnIII separate by breaking first the electrostatic interactions at the interface, followed by the hydrophobic ones. The sequence of the unbinding events and the maximum force required to separate the two proteins are independent of the presence of the Ca²⁺ ions, underlying the indirect role of Ca²⁺ in the binding.     

The cyclophilin-like domain of Ran-binding protein-2 modulates selectively the activity of the ubiquitin-proteasome system and protein biogenesis

The ubiquitin-proteasome system (UPS) plays a critical role in protein degradation. The 19S regulatory particle (RP) of the 26S proteasome mediates the recognition, deubiquitylation, unfolding, and channeling of ubiquitylated substrates to the 20S proteasome. Several subunits of the 19S RP interact with a growing number of factors. The cyclophilin-like domain (CLD) of Ran-binding protein-2 (RanBP2/Nup358) associates specifically with at least one subunit, S1, of the base subcomplex of the 19S RP, but the functional implications of this interaction on the UPS activity are elusive. This study shows the CLD of RanBP2 promotes selectively the accumulation of a subset of reporter substrates of the UPS, such as the ubiquitin (Ub)-fusion yellow fluorescent protein (YFP) degradation substrate, Ub(G76V)-YFP, and the N-end rule substrate, Ub-R-YFP. Conversely, the degradation of endoplasmic reticulum and misfolded proteins, and of those linked to UPS-independent degradation, is not affected by CLD. The selective effect of CLD on the UPS in vivo is independent of, and synergistic with, proteasome inhibitors, and CLD does not affect the intrinsic proteolytic activity of the 20S proteasome. The inhibitory activity of CLD on the UPS resides in a purported SUMO binding motif. We also found two RanBP2 substrates, RanGTPase-activating protein and retinitis pigmentosa GTPase regulator interacting protein-1alpha1, whose steady-state levels are selectively modulated by CLD. Hence, the CLD of RanBP2 acts as a novel auxiliary modulator of the UPS activity; it may contribute to the molecular and subcellular compartmentation of the turnover of properly folded proteins and modulation of the expressivity of several neurological diseases.     

Assignment of the locus for congenital lactase deficiency to 2q21, in the vicinity of but separate from the lactase-phlorizin hydrolase gene.

Congenital lactase deficiency (CLD) is an autosomal recessive, gastrointestinal disorder characterized by watery diarrhea starting during the first 1-10 d of life, in infants fed lactose-containing milks. Since 1966, 42 patients have been diagnosed in Finland. CLD is the most severe form of lactase deficiency, with an almost total lack of lactase-phlorizin hydrolase (LPH) activity on jejunal biopsy. In adult-type hypolactasia, the most common genetic enzyme deficiency in humans, this enzyme activity is reduced to 5%-10%. Although the activity of intestinal LPH has been found to be greatly reduced in both forms, the molecular pathogenesis of lactase deficiencies is unknown. On the basis of the initial candidate-gene approach, we assigned the CLD locus to an 8-cM interval on chromosome 2q21 in 19 Finnish families. At the closest marker locus, a specific allele 2 was present in 92% of disease alleles. On the basis of a genealogical study, the CLD mutation was found to be enriched in sparsely populated eastern and northern Finland, because of a founder effect. The results of both the genealogical study and the haplotype analysis indicate that one major mutation in a novel gene causes CLD in the Finnish population. Consequently, the critical region could be restricted further, to an approximately 350-kb interval, by ancient-haplotype and linkage-disequilibrium analyses. Surprisingly, the LPH gene was shown to lie outside the critical CLD region, excluding it as a causative gene for CLD. The LPH locus was found to reside >2 Mb from the critical CLD region.     

Conformational changes in the Ca2+-regulatory region from soybean calcium-dependent protein kinase-alpha: fluorescence resonance energy transfer studies

Calcium-dependent protein kinases are key proteins involved in plant and protozoal Ca2+ signaling. These unique molecules include a calcium regulatory calmodulin-like domain (CLD), which binds to another small regulatory domain named the junction domain (JD). Both CLD and JD are part of the same polypeptide as the protein kinase domain. The CLD consists of N- and C-terminal lobes, each having two helix-loop-helix Ca2+-binding motifs. In this study, fluorescence resonance energy transfer using a series of Trp and Cys site-directed mutants was undertaken to probe the relative motions of the two lobes of CLD between the apo- and Ca2+-saturated forms, as well as bound to a peptide encoding the JD sequence. Using an IAEDANS-modified Cys, a total of 15 Trp --> Cys distances were measured in these three states from the five donor-acceptor combinations F334W-Cys436, L371W-Cys436, L403W-Cys436, F334W-L403C, and L371W-L403C. Consistent with recently reported NMR diffusion measurements and with 1H,15N heteronuclear correlation NMR spectra, the distances derived from fluorescence resonance energy transfer measurements in apoCLD indicate partial unfolding and a subsequent contraction on binding Ca2+, which is maintained on addition of the JD peptide. Interpretation of the distances suggests that the Ca2+-saturated form is open with the two lobes sitting side-by-side although highly flexible. The transition to the JD-CLD state appears to be accompanied by a rotation of the N- and C-terminal domains with respect to each other inducing a slightly more closed overall complex. The observed differences between the behavior of CLD and the well studied related protein calmodulin are likely because of different physiological requirements for activation in vivo.     

Soluble interleukin-2 receptor as a marker for progression of coronary artery calcification in type 1 diabetes

INTRODUCTION: Soluble interleukin-2 receptor (sIL2r), a marker of T cell activation, is elevated in inflammatory processes, such as rheumatoid arthritis, hepatitis and neoplasm. We explored a potential association between plasma sIL2r levels and progression of coronary artery calcification (CAC), a marker for subclinical atherosclerosis, in a prospectively followed cohort of type 1 diabetic and non-diabetic subjects, aged 20-59 years, with no history of coronary artery disease. MATERIALS AND METHODS: CAC progression was assessed by electron beam tomography over 2.6 years (range 1.6-3.2). Plasma sIL2r levels were measured in a nested case-control substudy of 98 subjects (67 diabetic, 31 non-diabetic) with and 173 subjects (84 diabetic, 89 non-diabetic) without significant CAC progression. Log-transformed sIL2r levels were analyzed by conditional logistic regression to compare subjects with and without significant CAC progression. RESULTS: SIL2r was a significant predictor for CAC progression after adjusting for presence of baseline CAC, age, gender, diabetes status, baseline calcium volume score and adiponectin (OR 1.99, 95% CI 1.09-3.61, p = 0.02 for a doubling of sIL2r level). Addition of BMI, LDL, HDL, hypertension, smoking status, HbA1c, CRP, fibrinogen, homocysteine and PAI-1 to regression models weakened but did not remove sIL2r as a predictor of CAC progression. There was no indication that this effect was different by diabetes status (p = 0.6 for diabetes-sIL2r interaction). DISCUSSION: Elevated plasma sIL2r is associated with CAC progression independent of traditional coronary artery disease risk factors in type 1 diabetic and non-diabetic young adults. SIL2r should be considered as a novel marker of inflammation leading to coronary artery disease.     

CCR2 expression correlates with prostate cancer progression

Although the primary role of chemokines and their receptors is controlling the trafficking of leukocytes during inflammatory responses, they also play pleoitropic roles in cancer development. There is emerging evidence that cancer cells produce chemokines that induce tumor cell proliferation or chemotaxis in various cancer types. We have previously reported that MCP-1 acts as a paracrine and autocrine factor for prostate cancer (PCa) growth and invasion. As the cellular effects of MCP-1 are mediated by CC chemokine receptor 2 (CCR2), we hypothesized that CCR2 may contribute PCa progression. Accordingly, we first determined CCR2 mRNA and protein expression in various cancer cell lines, including PCa and other cancer types. All cells expressed CCR2 mRNA and protein, but in PCa, more aggressive cancer cells such as C4-2B, DU145, and PC3 expressed a higher amount of CCR2 compared with the less aggressive cancer cells such as LNCaP or non-neoplastic PrEC and RWPE-1 cells. Further, we found a positive correlation between CCR2 expression and PCa progression by analyzing an ONCOMINE gene array database. We confirmed that CCR2 mRNA was highly expressed in PCa metastatic tissues compared with the localized PCa or benign prostate tissues by real-time RT-PCR. Finally, CCR2 protein expression was examined by immunohistochemical staining on tissue microarray specimens from 96 PCa patients and 31 benign tissue controls. We found that CCR2 expression correlated with Gleason score and clinical pathologic stages, whereas lower levels of CCR2 were expressed in normal prostate tissues. These results suggest that CCR2 may contribute to PCa development.     

A role for the NG2 proteoglycan in glioma progression

Many human gliomas carry markers characteristic of oligodendrocyte progenitor cells (such as Olig-2, PDGF alpha receptor and NG2 proteoglycan), suggesting these progenitors as the cells of origin for glioma initiation. This review considers the potential roles of the NG2 proteoglycan in glioma progression. NG2 is expressed not only by glioma cells and by oligodendrocyte progenitors, but also by pericytes associated with the tumor microvasculature. The proteoglycan may therefore promote tumor vascularization and recruitment of normal progenitors to the tumor mass, in addition to mediating expansion of the transformed cell population. Along with potentiating growth factor signaling and serving as a cell surface receptor for extracellular matrix components, NG2 also has the ability to mediate activation of β-1 integrins. These molecular interactions allow the proteoglycan to contribute to critical processes such as cell proliferation, cell motility and cell survival.Key words: NG2 proteoglycan, glioma progression, cell motility, cell proliferation, cell survival, tumor vascularization