Higher importance of interleukin 6 than classic tumor markers (carcinoembryonic antigen and squamous cell cancer antigen) in the diagnosis of esophageal cancer patients.

It has been suggested that interleukin 6 (IL-6) plays a potential role in the growth and progression of tumors, including esophageal cancer (EC). The aim of the study was to compare clinical significance of serum IL-6 with classic tumor markers - carcinoembryonic antigen (CEA) and squamous cell cancer antigen (SCC-Ag) - in EC patients in relation to its histological types - squamous cell carcinoma of esophagus (ESCC) and adenocarcinoma (AD) of esophagus. The study included 53 EC patients and 90 healthy subjects. Serum IL-6 and CEA levels were determined using immunoenzyme assays, while SCC-Ag - chemiluminescent assay. The diagnostic criteria and prognostic values for markers were defined. The levels of all proteins tested in EC, ESCC, and AD were higher than in healthy subjects. The percentage of elevated results was substantially higher for IL-6 (86%) than for CEA (30%) and SCC-Ag (24%) in EC, similarly as in ESCC (87%, 23%, and 33%) and AD (87%, 39%, and 13%, respectively) patients. Concentrations of IL-6 depended on distant metastases and patients' survival in EC and were significantly higher in ESCC patients with more advanced tumor stage and nodal metastases. The IL-6 area under receiver operating characteristic curve (0.92) was larger than for CEA (0.84) and SCC-Ag (0.62) in EC, likewise in ESCC (0.92, 0.87, 0.77) and AD (0.91, 0.79, 0.57, respectively). Our findings indicate better usefulness of IL-6 than classic tumor markers in the diagnosis of EC, especially in patients with ESCC.

Vascular endothelial growth factor C and D expression correlates with lymph node metastasis and poor prognosis in patients with resected esophageal cancer.

Vascular endothelial growth factors C (VEGF-C) and D (VEGF-D) are important lymphangiogenic factors in human cancers. We studied the expression of VEGF-C and VEGF-D using immunohistochemistry in 73 resected esophageal cancer specimens, and correlated the results with patient clinicopathologic features and survival. High expression of VEGF-C was identified in 40 (54.7%) patients, and it correlated positively with histological grade (p=0.038), tumor stage (p=0.01), depth of tumor invasion (p=0.036) and lymph node metastasis (p=0.001). In 48 of 73 (65.7%) tumors, the VEGF-D protein was also expressed at high levels. VEGF-D immunoreactivity significantly correlated with tumor location (p=0.027), size of tumor (p=0.015), histological grade (p=0.02), depth of invasion (p=0.001) and lymph node metastasis (p=0.018). In logistic multivariate analysis, high expression of VEGF-C (OR 1.941, 95% CI 1.263-7.289, p=0.024) was associated with lymph node metastasis. Calculating the prognostic relevance revealed that both VEGF-C and VEGF-D correlated with decreased overall survival (p=0.01, p=0.003), disease free survival (p=0.02, p=0.006), and cancer-specific survival (p=0.03, p=0.005). In conclusion, our results suggest that high levels of both VEGF-C and VEGF-D proteins are associated with lymph node involvement, and that VEGF-C expression is an independent predictor of risk for lymph node metastasis in esophageal cancer. In locally advanced disease, overexpression of VEGF-C and VEGF-D may be useful in identifying patients who are more likely to have a poor prognosis even after curative resection.

Neuronal spike-train responses in the presence of threshold noise.

The variability of neuronal firing has been an intense topic of study for many years. From a modelling perspective it has often been studied in conductance based spiking models with the use of additive or multiplicative noise terms to represent channel fluctuations or the stochastic nature of neurotransmitter release. Here we propose an alternative approach using a simple leaky integrate-and-fire model with a noisy threshold. Initially, we develop a mathematical treatment of the neuronal response to periodic forcing using tools from linear response theory and use this to highlight how a noisy threshold can enhance downstream signal reconstruction. We further develop a more general framework for understanding the responses to large amplitude forcing based on a calculation of first passage times. This is ideally suited to understanding stochastic mode-locking, for which we numerically determine the Arnol'd tongue structure. An examination of data from regularly firing stellate neurons within the ventral cochlear nucleus, responding to sinusoidally amplitude modulated pure tones, shows tongue structures consistent with these predictions and highlights that stochastic, as opposed to deterministic, mode-locking is utilised at the level of the single stellate cell to faithfully encode periodic stimuli.

Calcium window currents, periodic forcing, and chaos: understanding single neuron response with a discontinuous one-dimensional map.

Thalamocortical (TC) neurones are known to express the low-voltage activated, inactivating Ca2+ current I(T). The triggering of this current underlies the generation of low threshold Ca2+ potentials that may evoke single or bursts of action potentials. Moreover, this current can contribute to an intrinsic slow (<1 Hz) oscillation whose rhythm is partly determined by the steady state component of I(T) and its interaction with a leak current. This steady state, or window current as it is so often called, has received relatively little theoretical attention despite its importance in determining the electroresponsiveness and input-output relationship of TC neurones. In this paper, we introduce an integrate-and-fire spiking neuron model that includes a biophysically realistic model of I(T). We briefly review the subthreshold bifurcation diagram of this model with constant current injection before moving on to consider its response to periodic forcing. Direct numerical simulations show that as well as the expected mode-locked responses there are regions of parameter space that support chaotic behavior. To reveal the mechanism by which the window current generates a chaotic response to periodic forcing we consider a piecewise linear caricature of the dynamics for the gating variables in the model of I(T). This model can be analyzed in closed form and is shown to support an unstable set of periodic orbits. Trajectories are repelled from these organizing centers until they reach the threshold for firing. By determining the condition for a grazing bifurcation (at the border between a spiking and nonspiking event) we show how knowledge of the unstable periodic orbits (existence and stability) can be combined with the grazing condition to determine an effective one-dimensional map that captures the essentials of the chaotic behavior. This map is discontinuous and has strong similarities with the universal limit mapping in grazing bifurcations derived in the context of impacting mechanical systems [A. B. Nordmark, Phys. Rev. E 55, 266 (1997)].

Virodhamine relaxes the human pulmonary artery through the endothelial cannabinoid receptor and indirectly through a COX product.

BACKGROUND AND PURPOSE: The endocannabinoid virodhamine is a partial agonist at the cannabinoid CB(1) receptor and a full agonist at the CB(2) receptor, and relaxes rat mesenteric arteries through endothelial cannabinoid receptors. Its concentration in the periphery exceeds that of the endocannabinoid anandamide. Here, we examined the influence of virodhamine on the human pulmonary artery. EXPERIMENTAL APPROACH: Isolated human pulmonary arteries were obtained during resections for lung carcinoma. Vasorelaxant effects of virodhamine were examined on endothelium-intact vessels precontracted with 5-HT or KCl. KEY RESULTS: Virodhamine, unlike WIN 55,212-2, relaxed 5-HT-precontracted vessels concentration dependently. The effect of virodhamine was reduced by endothelium denudation, two antagonists of the endothelial cannabinoid receptor, cannabidiol and O-1918, and a high concentration of the CB(1) receptor antagonist rimonabant (5 muM), but only slightly attenuated by the NOS inhibitor L-NAME and not affected by a lower concentration of rimonabant (100 nM) or by the CB(2) and vanilloid receptor antagonists SR 144528 and capsazepine, respectively. The COX inhibitor indomethacin and the fatty acid amide hydrolase inhibitor URB597 and combined administration of selective blockers of small (apamin) and intermediate and large (charybdotoxin) conductance Ca(2+)-activated K(+) channels attenuated virodhamine-induced relaxation. The vasorelaxant potency of virodhamine was lower in KCl- than in 5-HT-precontracted preparations. CONCLUSIONS AND IMPLICATIONS: Virodhamine relaxes the human pulmonary artery through the putative endothelial cannabinoid receptor and indirectly through a COX-derived vasorelaxant prostanoid formed from the virodhamine metabolite, arachidonic acid. One or both of these mechanisms may stimulate vasorelaxant Ca(2+)-activated K(+) channels.

Severe respiratory distress caused by central airway obstruction treated with self-expandable metallic stents.

We investigate retrospectively the efficacy of self-expandable metallic stents (SEMS) for severe respiratory distress (SRD) in patients with central airway obstruction (CAO). Thirty three patients with CAO were treated with SEMS using fiberoptic bronchoscopy method. We found an intraluminal obstruction present in 7, extraluminal compression in 10, and combined stenosis in 16 cases. Tumor infiltration occupied more than 90% of the endoluminal diameter in 21, 70% in 9, and 50% in 3 cases. Obstruction was caused by primary cancer of lung in 23, thyroid in 5, and esophagus in 5 patients. Up to 3 stents per patient were placed. Double stenting (esophagus and trachea) was required in five patients. All patients exhibited symptomatic and arterial blood gas improvement. The mean follow-up was 65 (5 to 752) days. SEMS are useful for the treatment of SRD caused by CAO. The overall effect is related to the degree of tumor progression itself.

Auditory thalamus responses to guinea-pig vocalizations: a comparison between rat and guinea-pig.

Although neuronal responses to species-specific vocalizations have long been described, very few between-species comparisons have been made. In a previous study, a differential representation of species-specific vocalizations was found in the auditory cortex (ACx): marmoset ACx neurons responded more, and more selectively, to marmoset calls than did cat ACx neurons [Wang, X., Kadia, S.C., 2001. Differential representation of species-specific primate vocalizations in the auditory cortices of marmoset and cat. J. Neurophysiol. 86, 2616-2620]. The present study analyzed responses of guinea-pig and rat auditory thalamus neurons to four well-defined guinea-pig vocalizations. Neurons of guinea-pigs (n = 96) and rats (n = 87) displayed similar response strength to guinea-pig vocalizations, and did not exhibit a preference for the natural over the time-reversed version of the calls in both species. This difference with the study by Wang and Kadia might suggest that, in mammals, the selectivity for the natural version of species-specific vocalizations is prominent only at the cortical level.

Therapy increases poly-ADP-ribose and p53-Ser392-P levels in recurrent squamous cell lung cancer.

p53 protein is a critical regulator of the cell cycle and apoptosis and its levels and functions change in response to many stimuli. To assess whether the cytotoxic drugs induce DNA changes, affect phosphorylation and stability of p53 protein, we determined poly-ADP-ribose levels, the expression of p53 protein and its carboxyl-terminal Ser-392 phosphate levels in fiberoptic bronchoscopy biopsy samples taken from patients suffering from recurrent squamous cell lung cancer before and after radiotherapy and chemotherapy. All 14 patients included in this study were in IA-IIIA clinical stage prior to surgery. Radiation/chemotherapy decreased G2/M cell numbers but increased S-phase cells by almost 50% compared to ploidy status before therapy, while median p53 expression was doubled (109% increase). p53 phosphorylated on Ser-392 was also increased by approximately 70% in patients treated with radiotherapy and with chemotherapy and correlated with elevated poly-ADP-ribose levels. Our data suggest that apart from changes in p53 quantity, posttranslational phosphorylation/dephosphorylation-mediated alterations may play an important role in neoplastic cell proliferation as well as in antiproliferative activity of drugs inducing DNA damage and apoptosis.

Prognostic molecular markers in non-small cell lung cancer.

Although TNM stage is the most significant prognostic parameter in lung cancer, additional parameters are required for explaining variability of survival. Hence molecular alterations in lung cancer have been extensively studied. Most prominent among them are alterations in the p53-p21 pathway, controlling the G1/S transition. They are the most commonly observed aberrations in non-small cell lung cancer (NSCLC). The results of p53 mutations on an individual patient's changes for survival are rather controversial. In a recent study however, after analyzing p53 abnormalities both by direct sequencing and immunohistochemistry together with evaluation of bcl-2 protein expression, we have found that p53 alterations were significantly associated with poor overall survival. Recently, a more sensitive yeast functional assay for altered p53 protein has been developed, with about 70% positivity in NSCLC patients and a correlation with shortened survival. The clinical significance of p21WAF1, the protein encoded by the target gene of p53 transcription, is still controversial; however expression has been associated with favorable prognosis in squamous cell carcinoma type. The 'Rb pathway' involving two oncogenes (cyclins D and E) and two tumor suppressor genes (Rb and p16) represents another major source of molecular alterations in lung cancer. Loss of Rb does not seem to significantly influence prognosis, white loss of p16 has been show repeatedly to be a factor for poor survival. Hypermethylation of the promoter region has been proposed as an alternative mechanism for inactivation of the p16 gene. The relation between cyclin D and E expression and prognosis, still is matter of controversy. Ras mutations are reported especially in adenocarcinoma; considered alone they bear no clear relation with prognosis, in opposition when considering them together with other molecular alterations. As a conclusion, a variety of molecular markers have been implicated in the prognosis of NSCLC. However, conflicting results were reported in the literature. Thus further investigations will be required, especially the use of newer molecular assays and the development of appropriate markers or panels of molecular markers.

The preoperative study of mediastinal lymph nodes metastasis in lung cancer by endoscopic ultrasonography (EUS) and helical computed tomography (CT).

OBJECTIVE: Accurate staging of mediastinal lymph nodes in patients with lung cancer is fundamental for their treatment and prognosis. The aim of this study was to compare the value of EUS and CT staging in patients with non-small-cell lung cancer (NSCLC) with postsurgical stage. METHODS: Ninety two patients with NSCLC underwent EUS and CT for preoperative detection of metastases to the mediastinal lymph nodes. EUS examinations were done with the ultrasonic linear array scanning echoendoscope (FG 32 UA, Hitachi/Pentax), CT-Toshiba Exvision GX scanner, with 24-s spiral acquisition, pitch 1:1 (7 mm collimation, 4 mm reconstruction index), during i.v. administration of non-ionic iodinated contrast media. RESULTS: The frequency of mediastinal involvement was 22.7%. The regions most accessible by EUS evaluation were subaortic, subcarinal and paraoesophageal lymph nodes. On a per-patient basis, EUS and CT results were: sensitivity 70.0 and 60.0%, specificity 80.6 and 72.6%, accuracy 77.2 and 68.5%. On a per-sites basis, the sensitivity of EUS evaluation was 78.8%, specificity 89.9%, accuracy 87.7%, comparing with CT-63.6, 84.0, 79.9%, respectively. When the EUS and CT images were analysed in combination, the sensitivity increased to 86.4%. CONCLUSION: We believe that EUS and CT should be used together for preoperative non-invasive staging of mediastinal lymph nodes in patients with NSCLC.

G1 phase of the cell cycle control and lung cancer: biological and clinical implications. Minireview.

Neoplastic diseases, including lung cancer are characterized by an uncoordinated cell growth. Cellular proliferation follows an orderly progression through the cell cycle, which is governed by protein complexes composed of cyclins and cyclin-dependent kinases. These complexes exert their regulatory function by phosphorylation of key proteins involved in cell cycle transitions. Abnormalities of cyclins and cyclin-dependent kinases have been reported and proposed to be oncogenic events. It appears that the molecular networking of these proteins and complexes exerts an impact on two fundamental cell cycle regulators; p53 and pRb. Interactions between these two nuclear proteins are being delineated, implying potential links between p53 and Rb in cell cycle control, apoptosis, and tumor progression. Furthermore, the detection of alterations in p53 and Rb pathways appears to be of clinical significance.

Prognostic significance of p53 and bcl-2 abnormalities in operable nonsmall cell lung cancer.

The association of p53 abnormalities and bcl-2 protein expression with clinical data and prognosis in 102 patients with resected nonsmall cell lung cancer (NSCLC) was investigated. Deoxyribonucleic acid analysis of exons 5-8 of the p53 gene showed mutations (p53-M) in 47% of resected NSCLC, serum p53 antibodies (p53-Abs) were detected in 25%, p53 protein overexpression (p53-PE) in 54%, and bcl-2 protein overexpression (bcl-2-PE) in 48%. A statistically significant association was found between p53-PE, serum p53-Abs and the presence of a p53 gene alteration. No significant associations were found between results of the p53-M, p53-Abs, bcl-2-PE tests and clinicopathological parameters. In the case of the p53-PE test there were significantly fewer positive results for adenocarcinoma than for squamous cell carcinoma and large cell carcinoma. Survival analysis showed that both p53 abnormalities and negative staining for bcl-2, when analysed separately, were associated with poor overall survival. In a multivariate analysis, only the positive result of the p53-M test remained an independent, statistically significant, unfavourable prognostic factor for survival. When the p53 mutation test was removed from the model, positive results of the p53-PE test and the p53-Abs test became statistically significant, unfavourable prognostic factors. To conclude, among p53 and bcl-2 abnormalities, only p53 gene mutations seem to have a strong and independent effect on prognosis. When deoxyribonucleic acid sequence information is not available, p53 protein expression and the presence of p53 antibodies in serum may be used to obtain important prognostic information.

Detection of P53 abnormalities in non-small cell lung cancer by yeast functional assay.

We assessed the status of P53 in 32 surgically treated non-small cell lung cancers (NSCLC) by using yeast functional assay. For functional assay, total RNA extracted from fresh-frozen specimens was reverse transcribed and P53 cDNAs were PCR-amplified using Pfu DNA polymerase (Stratagene). The transcriptional competence of the P53 cDNA was then tested in a yeast reporter strain. 20 of the 32 (69%) NSCLC patients contained mutant P53 in the yeast functional assay with the higher frequency in squamous cell carcinoma (14/17, 82%) than in adenocarcinoma (5/10, 50%) and large cell carcinoma (3/5, 60%) (p<0.01, chi2 test). No significant difference was observed with respect to the TNM. Preliminary survival analysis showed that patients scoring positive for the yeast test had shorter disease-free survival (median = 10 months) than those that scored negative (median > 21 months). Our results suggest that yeast functional assay is not only an improved method to examine the status of P53, but might hopefully improve understanding of the role of mutant P53 in the clinical evaluation of NSCLC.

Strong association between P53 protein accumulation, serum antibodies and gene mutation in non-small cell lung cancer.

DNA sequencing is the gold-standard method, but it is not feasible on a routine clinical basis. Immunohistochemistry is the most widely used method for assessing P53 alterations in human cancers. It can be performed during routine analysis, but some mutations may not lead to P53 protein accumulation, or P53 overexpression may be detected in the absence of mutations of the P53 gene. Recent studies have demonstrated the appearance of P53 antibodies in the serum of patients with lung cancer, probably due to the accumulation of mutant P53 protein in tumor cells. Using a logistic regression model applied to data for 102 non-small cell lung cancer (NSCLC) patients, we show a strong association between results of P53 mutation (P53-M) test, TNM stage and results of anti-P53 antibody in serum (P53-Abs) and P53 protein expression (P53-PE) tests. According to that model, we propose a procedure allowing for prediction of result of P53-M test. The percentage of correct predictions for independent data of 15 NSCLC patients was 80%. We conclude that in the absence of P53-M test result, a reasonably precise prediction of the test can be obtained using TNM stage and results of P53-Abs and P53-PE tests. The prediction can in turn be used to evaluate prognosis of NSCLC patients.

Vascular endothelial growth factor and basic fibroblast growth factor in patients with squamous cell oesophageal cancer.

It is suggested that vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) play an important role in tumor-induced angiogenesis. The purpose of this study was to estimate the correlation between VEGF and bFGF levels and tumor pathological status according to pTNM classification in patients with squamous cell oesophageal cancer. A group of 25 healthy controls and 32 consecutive patients with oesophageal cancer were included in this study. Serum VEGF and bFGF levels were determined by enzyme-linked immunosorbent assay (Quantikine R&D Systems). Serum VEGF and bFGF levels were significantly elevated in the patient groups (VEGF: 146.0 pg/ml, 79.0-386.3 pg/ml vs. 38.0 pg/ml, 6.5-135.1 pg/ml, p<0.005, and bFGF: 5.2 pg/ml, 1.2-10.6 pg/ml vs. 2.06 pg/ml, 0.07-4.0 pg/ml, p<0.02 Fisher test). The highest correlation between serum VEGF and bFGF levels were found in patients with advanced cancers, especially with: T4, N1, and M1 factors. The VEGF and bFGF levels were significantly higher in patients with pT4 (p<0.01). Patients with N1 lymph node invasion, compared with N0 factor, have higher levels of angiogenetic factors (p<0.04). Also in patients with advanced cancers with liver metastases the serum levels VEGF and bFGF were significantly higher (M1 vs. M0, VEGF p<0.001 and bFGF p<0.05). Consecutive monitoring of VEGF and bFGF serum levels may be a useful prognostic marker for patients with squamous cell oesophageal cancer.

Circulating anti-p53 antibodies in esophageal cancer patients.

Circulating anti-p53 protein antibodies (p53-Abs) have been detected in some cancer patients. The aim of the study was to determine the presence of circulating anti-p53 protein antibodies and their clinical significance in patients with esophageal carcinoma. Serum specimens from 75 consecutive patients with squamous cell carcinomas and 10 healthy subjects were studied. Enzyme linked immunosorbent assay (ELISA--Pharma Cell) was used to detect p53-Abs. At the time of diagnosis 20 (26.6%) of 75 analyzed patients had positive result in the p53-Abs test, but not any of the healthy subjects. The positive rate was 25% (1/4) cases in stage I, 41% (10/24) cases in stage IIA, 0% (0/8) cases in stage IIB, 28% (8/28) cases in stage III and 9% (1/11) cases in stage IV. In respect of tumour differentiation, cases graded as G1, G2 and G3 were positive in 28.5% (4/14), 25.9% (7/27) and 26.4% (9/34), respectively. There was no correlation between presence of p53-Abs and stage, rumour differentiation, lymph nodes metastases, tumour size, patient age and sex. In conclusion, the results of the present study indicate that serum p53-Abs did not correlate with cliniocopathologic feature of esophageal carcinoma.

Detection of p16 abnormalities in early-stage non-small cell lung cancer.

We investigated the clinical significance of p16 abnormalities (mutations by sequencing and hypermethylation by methylation-specific PCR) in 52 radically resected stage I-II non-small cell lung cancers (NSCLCs). P16 abnormalities were detected in 20 (38%) patents (point mutations in 4 (8%) and promoter hypermethylation in 16 (31%) cases. No correlation was found between p16 abnormalities and various clinicopathologic factors, including sex, histological type of tumor and TNM (I vs. II) stage. The multivariate analysis of survival was performed using the Cox proportional hazard model. When the types of 16 inactivation were analyzed, p16 hypermethylation rather than point mutation was associated with poor Prognosis. The presented results prompt the conclusion that hypermethylation of p16 is the major mechanism for p16 gene inactivation in early stage NSCLC and could be a useful molecular marker for the prognosis.

p53 gene mutation and protein expression in operable non-small cell lung cancer in Poland.

We investigated the association of p53 abnormalities (gene mutations by DNA sequencing and protein over-expression by immunostaining) with clinical data and prognosis in 74 patients with resected non-small cell lung cancer (NSCLC). DNA analysis of exons 5-8 of the p53 gene showed 34 mutations in 74 resected primary NSCLC (45.9%). Immunohistochemical study of the p53 protein revealed that 41 of 74 (55.4%) samples had positive staining. We found strong agreement between the results of the p53 protein expression test (p53-PE) and the p53 gene mutation test (p53-M) (Cohen's kappa = 0.65, 95% CI 0.48-0.82). Joint distribution of the results (analysed using the bivariate Dale model) was mainly influenced by, histological type of tumour. A positive result for the p53-PE test significantly increased (estimated odds ratio 84.5; 95% CI 8.89-803.03) the odds of observing a positive result in the p53-M test. In the univariate analysis (log rank test), positive results in the p53-M test and the p53-PE test were significantly associated with overall survival (P < 0.001 and P = 0.005, respectively). In the multivariate analysis (Cox's proportional hazard model), a positive result for the p53-M test significantly increased relative risk for overall survival (RR 9.56; 95% CI 2.62-34.87; P < 0.001). When the result of the p53-M test was accounted for, a positive result for the p53-PE test did not offer any additional prognostic information due to the strong dependence of results of the tests. However, when the result of the p53-M test was removed from the model, a positive result for the p53-PE test became a significant unfavourable prognostic factor (P = 0.009). We conclude that p53 gene mutation and protein expression analyses are in a strong agreement. Joint distribution of the results depends mainly on histological type of tumour. When considered separately, both tests are unfavourable prognostic factors in NSCLC. When the result of the p53-M test is taken into account, the p53-PE test does not offer any additional prognostic information.