Differential expression of circulating vascular cell adhesion molecule-1 in subjects with coronary artery disease and cardiac syndrome X without known diabetes mellitus

Context: Inflammation is one of the mechanisms underlying cardiac syndrome X (CSX).

Objectives: Few studies have compared the expression of inflammatory or adhesion molecules between coronary artery disease (CAD) versus CSX.

Materials and methods: Ninety-two CSX and 145 CAD subjects without known diabetes mellitus underwent coronary angiogram for angina.

Results: Vascular cell adhesion molecule (VCAM)-1 (median, 507 versus 431?ng/ml, p?=?0.001) was significantly higher in the CAD group. In the binary regression, VCAM-1 was a significant differential factor for CAD versus CSX.

Discussion and conclusion: Adhesion molecules might be implicated in the differential expression of macro versus microvascular coronary disease.

Trial registration number: NCT01198730 at https://clinicaltrials.gov     

Tumour expressions of hypoxic markers predict the response to neo-adjuvant chemotherapy in children with inoperable rhabdomyosarcoma

Objective: The study was to assess whether tumour expressions of hypoxia-inducible factor (HIF)-1α, glucose transporter (GLUT)-1, carbonic anhydrase (CA) IX and vascular endothelial growth factor (VEGF) predict response to neo-adjuvant chemotherapy (naCHT) in children with inoperable rhabdomyosarcoma (RMS).

Methods: Immunohistochemical expressions of hypoxia markers were determined semi-quantitatively in tumour tissue microarray of 46 patients with embryonal RMS (RME) and 20 with alveolar (RMA), treated with CWS protocols (1992–2013).

Results: In paediatric RME, response to naCHT was influenced significantly by tumour expression of CA IX and GLUT-1. Patients with RMA with low expressions of analysed markers responded well to naCHT, while all poor-responders expressed highly hypoxia markers. Only 5.88% of RMA and 11.11% of RME tumours did not express any of the proteins. In both RME and RMA subgroups, most poor-responders demonstrated simultaneous high expression of ≥3 markers, while most patients expressing ≤2 markers responded well to naCHT. In the whole cohort, co-expression of ≥3 markers, was the only independent factor predicting poor-response to chemotherapy (odds ratio 14.706; 95% CI 1.72–125.75; p?=?0.014).

Conclusions: Immunohistochemical expression pattern of four endogenous markers of hypoxia, in tumour tissue at diagnosis, emerges as a promising tool to predict response to naCHT in children with inoperable RMS.     

TRH receptor mobility in the plasma membrane is strongly affected by agonist binding and by interaction with some cognate signaling proteins

Objectives: Extensive research has been dedicated to elucidating the mechanisms of signal transduction through different G protein-coupled receptors (GPCRs). However, relatively little is known about the regulation of receptor movement within the cell membrane upon ligand binding. In this study we focused our attention on the thyrotropin-releasing hormone (TRH) receptor that typically couples to G_q/11 proteins.

Methods: We monitored receptor diffusion in the plasma membrane of HEK293 cells stably expressing yellow fluorescent protein (YFP)-tagged TRH receptor (TRHR-YFP) by fluorescence recovery after photobleaching (FRAP).

Results: FRAP analysis indicated that the lateral movement of the TRH receptor was markedly reduced upon TRH binding as the value of its diffusion coefficient fell down by 55%. This effect was prevented by the addition of the TRH receptor antagonist midazolam. We also found that siRNA-mediated knockdown of G_q/11α, Gβ, β-arrestin2 and phospholipase Cβ1, but not of G_iα1, β-arrestin1 or G protein-coupled receptor kinase 2, resulted in a significant decrease in the rate of TRHR-YFP diffusion, indicating the involvement of the former proteins in the regulation of TRH receptor behavior. The observed partial reduction of the TRHR-YFP mobile fraction caused by down-regulation of G_iα1 and β-arrestin1 suggests that these proteins may also play distinct roles in THR receptor-mediated signaling.

Conclusion: These results demonstrate for the first time that not only agonist binding but also abundance of some signaling proteins may strongly affect TRH receptor dynamics in the plasma membrane.     

Dynamic variation of histone H3 trimethyl Lys4 (H3K4me3) and heterochromatin protein 1 (HP1) with employment length in nickel smelting workers

Objective: To investigate the dynamic variation in H3K4me3 and HP1 with employment length in nickel smelting workers.

Methods: Blood samples were collected from 140 nickel smelting workers and 140 age-matched office workers to test for H3K4me3, and HP1 levels.

Results: H3K4me3 was statistically significantly different (p?<?0.05) between the two groups and positively correlated with employment length (r_s?=_?0.267). HP1 was not correlated with employment length (p?=?0.066) but was significantly different between the two groups.

Conclusions: Chronic exposure to nickel can induce oxidative damage, and increase H3K4me3 expression and inhibit HP1 expression.     

Beta-2-glycoprotein-1 and alpha-1-antitrypsin as urinary markers of renal cancer in von Hippel–Lindau patients

Context: Von Hippel–Lindau disease (VHLD) is a rare inherited neoplastic syndrome. Among all the VHLD-associated tumors, clear cell renal cell carcinoma (ccRCC) is the major cause of death.

Objective: The aim of this paper is the discovery of new non-invasive biomarker for the monitoring of VHLD patients.

Materials and methods: We compared the urinary proteome of VHLD patients, ccRCC patients and healthy volunteers.

Results: Among all differentially expressed proteins, alpha-1-antitrypsin (A1AT) and APOH (beta-2-glycoprotein-1) are strongly over-abundant only in the urine of VHLD patients with a history of ccRCC.

Discussion and conclusion: A1AT and APOH could be promising non-invasive biomarkers.     

Propeptide big-endothelin,N-terminal-pro brain natriuretic peptide and mortality. The Ludwigshafen risk and cardiovascular health (LURIC) study

Context: The endothelin system (Big-ET-1) is a key regulator in cardiovascular (CV) disease and congestive heart failure (CHF).

Objectives: We have examined the incremental value of Big-ET-1 in predicting total and CV mortality next to the well-established CV risk marker N-Terminal Pro-B-Type Natriuretic Peptide (NT-proBNP).

Methods: Big-ET-1 and NT-proBNP were determined in 2829 participants referred for coronary angiography (follow-up 9.9 years).

Results: Big-ET-1 is an independent predictor of total, CV mortality and death due to CHF.

Discussion: The conjunct use of Big-ET-1 and NT-proBNP improves the risk stratification of patients with intermediate to high risk of CV death and CHF.

Conclusions: Big-ET-1improves risk stratification in patients referred for coronary angiography.     

Effect of different concentrations of oxygen on expression of sigma 1 receptor and superoxide dismutases in human colon adenocarcinoma cell lines

Context: Tumor cells due to distance from capillary vessels exist in different oxygenation conditions (anoxia, hypoxia, normoxia). Changes in cell oxygenation lead to reactive oxygen species production and oxidative stress. Sigma 1 receptor (Sig1R) is postulated to be stress responding agent and superoxide dismutases (SOD1 and SOD2) are key antioxidant enzymes. It is possible that they participate in tumor cells adaptation to different concentrations of oxygen.

Objective: Evaluation of Sig1R, SOD1, and SOD2 expression in different concentrations of oxygen (1%, 10%, 21%) in colon adenocarcinoma cell lines.

Materials and methods: SW480 (primary adenocarcinoma) and SW620 (metastatic) cell lines were cultured in standard conditions in Dulbecco’s modified Eagle’s medium for 5 days, and next cultured in Hypoxic Chamber in 1% O₂, 10% O₂, 21% O₂. Number of living cells was determined by trypan blue assay. Level of mRNA for Sig1R, SOD1, and SOD2 was determined by standard PCR method. Statistical analysis was conducted using Statistica 10.1 software.

Results: We observed significant changes in expression of Sig1R, SOD1, SOD2 due to different oxygen concentrations. ANOVA analysis revealed significant interactions between studied parameters mainly in hypoxia conditions in SW480 cells and between Sig1R and SOD2 in SW620 cells. It also showed that changes in expression of studied proteins depend significantly on type of the cell line.

Conclusion: Changes of Sig1R and SOD2 expression point to mitochondria as main organelle responsible for survival of tumor cells exposed to hypoxia or oxidative stress. Studied proteins are involved in intracellular response to stress related with different concentrations of oxygen.     

A new nitrobenzoxadiazole-based GSTP1-1 inhibitor with a previously unheard of mechanism of action and high stability

Context: The nitrobezoxadiazole derivative NBDHEX is a potent inhibitor of glutathione transferase P1-1 (GSTP1-1) endowed with outstanding anticancer activity in different tumor models.

Objective: To characterize by in vitro biochemical and in silico studies the NBDHEX analogues named MC2752 and MC2753.

Materials and methods: Synthesis of MC2752 and MC2753, biochemical assays and in silico docking and normal-mode analyses.

Results: The presence of a hydrophobic moiety in the side chain of MC2753 confers unique features to this molecule. Unlike its parent drug NBDHEX, MC2753 does not require GSH to trigger the dissociation of the complex between GSTP1-1 and TRAF2, and displays high stability towards the nucleophilic attack of the tripeptide under physiological conditions.

Discussion and conclusion: MC2753 may represent a lead compound for the development of novel GSTP1-1 inhibitors not affected in their anticancer action by fluctuations of cellular GSH levels, and characterized by an increased half-life in vivo.     

Oxidative insults disrupt OPA1-mediated mitochondrial dynamics in cultured mammalian cells

Objective: To explore the impact of oxidative insults on mitochondrial dynamics. In mammalian cells, oxidative insults activate stress response pathways including inflammation, cytokine secretion, and apoptosis. Intriguingly, mitochondria are emerging as a sensitive network that may function as an early indicator of subsequent cellular stress responses. Mitochondria form a dynamic network, balancing fusion, mediated by optic atrophy-1 (OPA1), and fission events, mediated by dynamin-related protein-1 (DRP1), to maintain homeostasis.

Methods: Here, we examine the impact of oxidative insults on mitochondrial dynamics in 143B osteosarcoma and H9c2 cardiomyoblast cell lines via confocal microscopy, flow cytometry, and protein-based analyses.

Results: When challenged with hydrogen peroxide (H₂O₂), a ROS donor, both cell lines display fragmentation of the mitochondrial network and loss of fusion-active OPA1 isoforms, indicating that OPA1-mediated mitochondrial fusion is disrupted by oxidative damage in mammalian cells. Consistent with this, cells lacking OMA1, a key protease responsible for cleavage of OPA1, are protected against OPA1 cleavage and mitochondrial fragmentation in response to H₂O₂ challenge.

Discussion: Taken together, these findings indicate that oxidative insults damage OPA1-mediated mitochondrial dynamics in mammalian cells via activation of OMA1, consistent with an emerging role for mitochondrial dynamics as an early indicator of cellular stress signaling.     

Big endothelin-1 level is a useful marker for predicting the presence of isolated coronary artery ectasia

Context: Endothelin-1(ET-1) has been implicated in coronary artery disease (CAD) and may be associated with coronary artery ectasia (CAE).

Objective: To clarify the relationship between big ET-1 and isolated CAE.

Methods: We measured big ET-1 with ELISA in 216 patients (CAE, n?=?72; CAD, n?=?72; normal, n?=?72) and evaluated the link with isolated CAE.

Results: The level of plasma big ET-1 was significantly higher in patients with isolated CAE (p?<?0.001). Big ET-1 was strongly and independently associated with CAE by multivariate analysis (OR 95%CI: 1.026 (1.018–1.034), p?=?0.000).

Conclusions: Big ET-1 may be a useful predictor for the presence of isolated CAE.     

PPARgamma-mediated ALDH1A3 suppression exerts anti-proliferative effects in lung cancer by inducing lipid peroxidation

Context: The metabolic function of peroxisome proliferator-activated receptor gamma (PPARγ) in lung cancer remains unclear.

Objectives: To determine the relationship of PPARγ on ALDH1A3-induced lipid peroxidation to inhibit lung cancer cell growth.

Materials and methods: In silico analysis using microarray dataset was performed to screen the positive correlation between PPARγ and all ALDH isoforms. NUBIscan software and ChIP assay were used to identify the binding sites (BSs) of PPARγ on ALDH1A3 promoter. The expression of ALDH1A3 under thiazolidinedione (TZD) treatment was evaluated by QPCR and Western Blot in HBEC and H1993 cell lines. Upon treatment of TZD, colony formation assay was used to check cell growth inhibition and 4-hydroxy-2-nonenal (4HNE) production as lipid peroxidation marker was determined by Western Blot in PPARγ positive cell H1993 and PPARγ negative cell H1299.

Results: Compared to other ALDH isoforms, ALDH1A3 showed the highest positive correlation to PPARγ expression. ALDH1A3 upregulated PPARγ expression while PPARγ activation suppressed ALDH1A3. Among 2 potential screened PPARγ response elements, BS 1 and 2 in the promoter of ALDH1A3 gene, PPARγ bound directly to BS2. Ligand activation of PPARγ suppressed mRNA and protein expression of ALDH1A3. Growth inhibition was observed in H1993 (PPARγ positive cell) treated with PPARγ activator and ALDH inhibitor compared to H1299 (PPARγ negative cell). PPARγ activation increased 4HNE which is known to be suppressed by ALDH1A3.

Conclusions: ALDH1A3 suppression could be one of PPARγ tumor suppressive function. This study provides a better understanding of the role of PPARγ in lung cancer.     

The pharmacologic characterization of allosteric molecules: Gq protein activation

Context: Drugs such as positive allosteric modulators (PAMs) produce complex behaviors when acting on tissues in different physiological contexts in vivo.

Objective: This study describes the use of functional assays of varying receptor sensitivity to unveil the various behaviors of PAMs and thus quantify allosteric effect through system independent scales.

Materials and methods: Muscarinic receptor activation with acetylcholine (ACh) was used to the demonstrate activity of the PAM agonist 1–(4-methoxybenzyl)-4-oxo-1,4-dihydroquinoline-3-carboxylic acid, Benzyl quinolone carboxylic acid (BQCA) in terms of direct agonism, potentiation of ACh affinity, and ACh efficacy. Concentration–response curves were fit to the functional allosteric model to yield indices of agonism (τ_B), effects on affinity (α cooperativity), and efficacy (β cooperativity).

Results: It is shown that a highly sensitive functional assay revealed the direct efficacy of BQCA as an agonist and relatively insensitive cells (produced by chemical alkylation of muscarinic receptor with phenoxybenzamine) revealed a positive allosteric effect of BQCA on ACh efficacy. A wide range of functional assay sensitivities produced a complex pattern of behavior for BQCA all of which was accurately quantified through the system-independent parameters of the functional allosteric model.

Conclusions: The study of complex allosteric molecules in a range of functional assays of varying sensitivity allows the measurement of the complete array of activities of these molecules on receptors and also better predicts which will be seen with these in vivo where a range of tissue sensitivities is encountered.     

Aberrant methylation of nucleotide excision repair genes is associated with chronic arsenic poisoning

Objective: To define whether aberrant methylation of DNA repair genes is associated with chronic arsenic poisoning.

Methods: Hundred and two endemic arsenicosis patients and 36 healthy subjects were recruited. Methylight and bisulfite sequencing (BSP) assays were used to examine the methylation status of ERCC1, ERCC2 and XPC genes in peripheral blood lymphocytes (PBLs) and skin lesions of arsenicosis patients and NaAsO₂-treated HaCaT cells.

Results: Hypermethylation of ERCC1 and ERCC2 and suppressed gene expression were found in PBLs and skin lesions of arsenicosis patients and was correlated with the level of arsenic exposure. Particularly, the expression of ERCC1 and ERCC2 was associated with the severity of skin lesions. In vitro studies revealed an induction of ERCC2 hypermethylation and decreased mRNA expression in response to NaAsO₂ treatment.

Conclusion: Hypermethylation of ERCC1 and ERCC2 and concomitant suppression of gene expression might be served as the epigenetic marks associated with arsenic exposure and adverse health effects.     

Proteomics and phosphoproteomics study of LCMT1 overexpression and oxidative stress: overexpression of LCMT1 arrests H2O2-induced lose of cells viability

Objectives: Protein phosphatase 2A (PP2A), a major serine/threonine phosphatase, is also known to be a target of ROS. The methylation of PP2A can be catalyzed by leucine carboxyl methyltransferase-1 (LCMT1), which regulates PP2A activity and substrate specificity.

Methods: In the previous study, we have showed that LCMT1-dependent PP2Ac methylation arrests H₂O₂-induced cell oxidative stress damage. To explore the possible protective mechanism, we performed iTRAQ-based comparative quantitative proteomics and phosphoproteomics studies of H₂O₂-treated vector control and LCMT1-overexpressing cells.

Results: A total of 4480 non-redundant proteins and 3801 unique phosphopeptides were identified by this means. By comparing the H₂O₂-regulated proteins in LCMT1-overexpressing and vector control cells, we found that these differences were mainly related to protein phosphorylation, gene expression, protein maturation, the cytoskeleton and cell division. Further investigation of LCMT1 overexpression-specific regulated proteins under H₂O₂ treatment supported the idea that LCMT1 overexpression induced ageneral dephosphorylation of proteins and indicated increased expression of non-erythrocytic hemoglobin, inactivation of MAPK3 and regulation of proteins related to Rho signal transduction, which were known to be linked to the regulation of the cytoskeleton.

Discussion: These data provide proteomics and phosphoproteomics insights into the association of LCMT1-dependent PP2Ac methylation and oxidative stress and indirectly indicate that the methylation of PP2A plays an important role against oxidative stress.     

A comparison of HMGB1 concentrations between cerebrospinal fluid and blood in patients with neurological disease

Aims: To determine whether a correlation exists between paired cerebrospinal fluid (CSF) and serum levels of a novel inflammatory biomarker, high-mobility group box 1 (HMGB1), in different neurological conditions.

Methods: HMGB1 was measured in the serum and CSF of 46 neurological patients (18 idiopathic intracranial hypertension [IIH], 18 neurological infection/inflammation [NII] and 10 Rasmussen’s encephalitis [RE]).

Results: Mean serum (±?SD) HMGB1 levels were 1.43?±?0.54, 25.28?±?27.9 and 1.89?±?1.49?ng/ml for the patients with IIH, NII and RE, respectively. Corresponding mean (±?SD) CSF levels were 0.35?±?0.22, 4.48?±?6.56 and 2.24?±?2.35?ng/ml. Both CSF and serum HMGB1 was elevated in NII. Elevated CSF HMGB1 was demonstrated in RE. There was no direct correlation between CSF and serum levels of HMGB1.

Conclusion: Serum HMGB1 cannot be used as a surrogate measure for CSF levels. CSF HMGB1 was elevated in NII and RE, its role as a prognostic/stratification biomarker needs further study.     

Prognostic value of PCT,copeptin, MR-proADM,MR-proANP and CT-proET-1 for severe acute dyspnea in the emergency department: the BIODINER study

Context: Acute dyspnea is a frequent complaint in patients attending the emergency department (ED).

Objective: To evaluate the accuracy of PCT, MR-proANP, MR-proADM, copeptin and CT-proET1 for the risk-stratification of severe acute dyspnea patients presenting to the ED.

Methods: Multicenter prospective study in adult patients with a chief complaint of acute dyspnea. Pro-hormone type biomarkers concentrations were measured on arrival. Combined primary endpoint was a poor outcome.

Results: Three hundred and ninety-four patients were included, 137 (35%) met the primary endpoint. MR-proADM was the only biomarker associated with the primary endpoint (odds ratio 1.43 [95%CI: 1.13–1.82], p?=?0.003) as were the presence of paradoxical abdominal breathing (odds ratio 2.48 [95%CI: 1.31–4.68]) or cyanosis (odds ratio 3.18 [1.46–6.89])

Conclusions: In patients with severe acute dyspnea in the ED, pro-hormone type biomarkers measurements have a low added value to clinical signs for the prediction of poor outcome.     

New markers of cell migration and inflammation in children with chronic kidney disease

Context: Chronic kidney disease (CKD) is characterized by immunocompetent cell migration and inflammation. Monocyte chemoattractant protein (MCP)-1 and macrophage colony-stimulating factor (MCSF) stimulate monocyte migration and transition into macrophages with subsequent release of neopterin.

Objective: The aim of the study was to analyze these parameters in children with various stages of CKD.

Material and methods: The study group consisted of 41 CKD children, 19 patients on haemodialysis (HD), 22 children on automated peritoneal dialysis (APD) and 23 controls. Serum concentrations of MCP-1, MCSF and neopterin were assessed by ELISA. Correlations to matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) were analyzed.

Results: MCP-1, MCSF and neopterin were significantly elevated in all patients versus controls and the highest values concerned HD children. A single HD session lessened the concentrations of all parameters, yet they rose back before the next HD session. All markers correlated with MMPs and TIMPs in different combinations.

Conclusions: Systemic inflammation and cell migration are triggered by CKD and additionally aggravated by chronic dialysis, with the more evident negative impact of HD than APD. Discrepancies in MCP1, MCSF and neopterin serum concentrations suggest they may serve as new markers of cellular and inflammatory responses in children with CKD.     

Bone turnover markers and novel biomarkers in lung cancer bone metastases

Context: Lung cancer still remains the leading cause of cancer-related mortality worldwide. Bone is one of preferred metastatic sites for lung cancer cells. So far, both accurate diagnosis and effective treatment of lung cancer bone metastases are difficult.

Objective: This review aimed to evaluate roles of bone turnover markers (BTMs), microRNAs (miRNAs), dickkopf1 (DKK1) and insulin like growth factor binding protein 3 (IGFBP-3) in lung cancer bone metastases.

Methods: We searched articles about these four biomarkers in lung cancer bone metastases mainly in PubMed.

Result: The levels of bone specific alkaline phosphatase (BALP), cross-linked carboxy-terminal telopeptide of type-I collagen (ICTP) and N-terminal telopeptides of type-I collagen (NTX) were reported to be significantly increased in lung cancer patients with bone metastases. ALP, NTX and bone sialoprotein were thought to be associated with prognosis of lung cancer patients with bone metastases. MiRNA-335, miRNA-33a, miRNA-21, DKK1 and IGFBP-3 were revealed to be novel biomarkers in lung cancer bone metastases.

Discussion and conclusion: Current researches have revealed that BTMs, miRNAs, DKK1 and IGFBP-3 may be useful in diagnosis, prognosis evaluation or treatment of lung cancer bone metastases. More studies about these biomarkers in lung cancer bone metastases are needed.     

Notch1 regulates tongue cancer cells proliferation,apoptosis and invasion

Objectives: Notch1 regulates tumor biology in a complex, context-dependent manner. The roles of Notch1 in tongue cancer are still controversial. The aim of this study is to investigate the roles of Notch1 in tongue cancer.

Materials and Methods: The expression of Notch1 was tested between tongue cancer and normal samples by using immunohistochemistry. Tongue cancer cells were transfected with siRNA or plasmid, respectively. Cell proliferation, apoptosis, migration and invasion ability were tested in appropriate ways. The subcutaneous tumor model was established to observe the tumor growth.

Results: Notch1 was upregulated in tongue carcinoma tissues and the expression of Notch1 was related with tumor stage and differentiation. Overexpression of Notch1 could increase tongue cancer cells proliferation, invasion and migration. But inhibited the expression of Notch1 could decrease cells proliferation, invasion and migration and promote cell apoptosis in vitro and in vivo.

Conclusion: Our results prove that the oncogenic role of Notch1 in tongue cancer and provide the direction of targeted therapy of tongue cancer.     

Soluble TREM-1 as a diagnostic and prognostic biomarker in patients with septic shock: an observational clinical study

Objectives: The impact of TREM-1-mediated inflammation was investigated in different inflammatory settings.

Methods: Secondary analyses of an observational clinical pilot study, including 60 patients with septic shock, 30 postoperative controls and 30 healthy volunteers.

Results: Plasma levels of sTREM-1 were found to identify patients with septic shock more effectively than procalcitonin and C-reactive protein. Moreover, sTREM-1 was identified to be an early predictor for survival in patients with septic shock.

Conclusion: Due to its diagnostic as well as prognostic value in sepsis syndrome, implementation of sTREM-1 measurements in routine diagnostics should be taken into account.