Quantitative analysis of p16 methylation in Barrett's carcinogenesis.

p16 hypermethylation in Barrett's carcinogenesis has been evaluated in studies which did not take into account sample heterogeneity and yielded qualitative (methylated/unmethylated) instead of accurate quantitative (percentage of CpG methylation) data. We aimed to measure the degree of p16 methylation in pure samples representing all the steps of Barrett's tumorogenesis and to evaluate the influence of sample heterogeneity in methylation analysis. METHODS: 77 paraffin-embedded human esophageal samples were analyzed. Histological grading was established by two pathologists in: negative for dysplasia, indefinite for dysplasia, low-grade dysplasia, high-grade dysplasia and adenocarcinoma. Areas of interest were selected by laser-capture microdissection. p16 methylation was quantified by pyrosequencing. An adjacent section of the whole sample was also analyzed to compare methylation data. RESULTS: After microdissection, we obtained 15 samples of squamous epithelium, 36 non-dysplastic Barrett's esophagus, 3 indefinite for dysplasia, 24 low-grade dysplasia, 4 high-grade dysplasia and 12 adenocarcinoma. Squamous epithelium showed the lowest methylation rates: 6% (IQR 5-11) vs. 11%(7-39.50) in negative/indefinite for dysplasia, p<0.01; 10.60%(6-24) in low-grade dysplasia, p<0.05; and 44.50%(9-66.75) in high-grade dysplasia/adenocarcinoma, p<0.01. This latter group also exhibited higher methylation rates than Barrett's epithelium with and without low-grade dysplasia (p<0.05). p16 methylation rates of microdissected and non-microdissected samples did not correlate unless the considered histological alteration comprised >71% of the sample. CONCLUSIONS: p16 methylation is an early event in Barrett's carcinogenesis which increases with the severity of histological alteration. p16 methylation rates are profoundly influenced by sample heterogeneity, so selection of samples is crucial in order to detect differences.

Expansion of a CD26low Effector TH Subset and Reduction in Circulating Levels of sCD26 in Stable Allergic Asthma in Adults.

BACKGROUND AND OBJETIVE: The pathogenesis of asthma is dependent on the balance between regulatory and effector T cells, which display differential expression of CD25 and CD26. Therefore, alteration of circulating levels of sCD25 and sCD26 during allergic asthma could be conditioned by changes in leukocyte phenotype. Objectives: To analyze expression of CD25 and CD26 on T lymphocytes and their soluble derivatives (sCD25, sCD26) during stable phases of moderate-severe allergic asthma. METHODS: Cross-sectional study with 2 adult cohorts of allergic asthmatics. Clinical, anthropometric, pulmonary, hematological, and biochemical parameters were measured. Phenotyping was performed with flow cytometry in both circulating and cultured leukocytes. Dipeptidyl peptidase 4 (DPP4) activity was assayed in culture supernatants. RESULTS: In vitro studies revealed upregulation of CD26 on human T lymphocytes upon activation, especially under TH17-favoring conditions, and a correlation with soluble DPP4 activity (rs=0.641; P<.001). CD26 expression on lymphocytes was higher in asthmatics, while serum sCD26 was lower in women and patients. The latter finding could be associated with an expanded CD25low/CD26low/CD127low subset of effector CD4+ T cells in allergic asthma, with no changes in Treg percentages. However, women showed an increased Teff/Treg ratio, which could explain their greater susceptibility to asthma. CONCLUSIONS: Allergic asthma causes an increment in CD25lowCD26low helper T cells detected in stable stages. These changes are mirrored in serum and should be considered in the light of the downmodulating role of CD26 in major chemokines related to the pathogenesis of asthma such as CCL11 (eotaxin), CCL5 (RANTES), and CXCL12a (SDF-1α).

In vitro vitamin K3 effect on conjunctival fibroblast migration and proliferation.

PURPOSE: To evaluate the dose effect of vitamin K3 on wound healing mechanisms. METHODS: Conjunctival fibroblasts were incubated for 24 hours. An artificial wound was made and the cells were incubated with fresh medium plus doses of vitamin K3 to be tested. Wound repair was monitored at 0, 18, 24, and 48 hours. Proliferation was measured in actively dividing cells by [(3)H]thymidine uptake. Six different groups were tested: group 1/no drugs added, group 2/ethanol 0.1%, group 3/vitamin K3 1 mg/L, group 4/vitamin K3 2 mg/L, group 5/vitamin K3 4 mg/L, and group 6/vitamin K3 6 mg/L. Each experiment was carried out in triplicate and 4 times. RESULTS: There were no differences among groups at the initial time. In vitro wound repair was slower in groups 4, 5, and 6. There were no differences between control and ethanol groups and between control and vitamin K3 1 mg/L groups. Fibroblast mitogenic activity was statistically decreased in all vitamin K groups; statistical differences were found among vitamin K3 1 mg/mL and higher doses too. In groups 5 and 6, cellular toxicity was presented. CONCLUSIONS: Vitamin K3 is able to inhibit fibroblast proliferation. Vitamin K3 2 mg/L or higher doses inhibit wound healing repair, exhibiting cellular toxicity at 4 and 6 mg/L.

Algorithm for phase extraction from a set of interferograms with arbitrary phase shifts.

The generalized analytical quadrature filter from a set of interferograms with arbitrary phase shifts is obtained. Both symmetrical and non symmetrical algorithms for any order are reported. The analytic expression is obtained through the convolution of a set of two-frame algorithms and expressed in terms of the combinatorial theory. Finally, the solution is applied to obtain several generalized tunable quadrature filters.

The CD14 (-159 C/T) SNP is associated with sCD14 levels and allergic asthma, but not with CD14 expression on monocytes.

LPS-ligation to CD14/TLR-4 on monocytes/macrophages triggers the production of IL-12-family cytokines. IL12/18 promote TH1-differentiation, counteracting the TH2-driven asthma. Therefore, CD14 modulation could alter the TH2-differentiation and should be taken into account when studying asthma. To analyse the alteration in CD14 levels and its association with CD14 (-159 C/T) SNP (rs2569190) in Caucasian adults with stable allergic asthma, we performed a cross-sectional study (277 healthy subjects vs. 277 patients) where clinical parameters, CD14 values and the CD14 (-159 C/T) SNP were studied. Apart from typical biomarkers, we found an increment of neuron-specific enolase (NSE) in allergic asthma, probably linked to monocyte activity. Indeed, we evidenced increased monocyte numbers, but lower CD14 expression and normalised sCD14 values in patients. Moreover, we noticed an association of the T allele (P = 0.0162) and TT genotype (P = 0.0196) of the CD14 SNP with a decreased risk of allergic asthma and augmented sCD14 levels. In conclusion, monocyte CD14 expression and normalized sCD14 values were reduced in stable state asthmatics, and this could be related to the presence of an expanded CD14low monocyte subset. This study also demonstrates that the CD14 (-159 C/T) polymorphism is a risk factor for moderate-severe allergic asthma in adult Caucasians.

Thoracic versus whole body bioimpedance measurements: the relation to hydration status and hypertension in peritoneal dialysis patients.

The whole body bioimpedance technique is a highly promising non-invasive, reproducible, fast and inexpensive bed-side method for monitoring hydration status. Using segmental bioimpedance measurements, it is possible to obtain information about the fluid change in each body segment (Song, Lee, Kim and Kim 1999 Perit. Dial. Int. 19 386-90). In this pilot study we have measured 25 male patients (30-65 yr, BMI 20-32 kg m(-2)) undergoing continuous ambulatory peritoneal dialysis (CAPD). Tetrapolar impedance measurements were obtained using the right-side technique (whole body), and a segmental impedance method focused in the thorax region. Blood pressure (BP) measurements were taken manually with a sphygmomanometer. Patients were classified as either stable (group 0) or unstable (group 1) using clinical parameters of overall cardiovascular risk. The Mahalanobis distance (dM2) was calculated for the mean blood pressure (BP(mean)), and the impedance parameter R normalized by body height H for the right-side (R(RS)/H) and the thorax segment (R(TH)/H). Differences between groups were significant (p < 0.0001) for R(TH)/H and for BP(mean), and less significant (p = 0.016) for R(RS)/H. Group 1 patients showed a small dM2 as compared with a reference patient (a critical patient with acute lung edema) with high BP(mean) and low values of R(TH)/H and R(RS)/H. Moreover, Group 0 patients showed a larger dM2 with respect to the reference patient, with lower BP(mean) and higher values of R(TH)/H and R(RS)/H. All patients classified as unstable by clinical assessment were correctly classified using R(TH)/H in conjunction with BP(mean) using dM2. Segmental-monofrequency non-invasive bioimpedance of the thoracic region could provide a simple, objective non-invasive method of support for facilitating the clinical assessment of CAPD patients.

A methodology to quantify the differences between alternative methods of heart rate variability measurement.

This work proposes a systematic procedure to report the differences between heart rate variability time series obtained from alternative measurements reporting the spread and mean of the differences as well as the agreement between measuring procedures and quantifying how stationary, random and normal the differences between alternative measurements are. A description of the complete automatic procedure to obtain a differences time series (DTS) from two alternative methods, a proposal of a battery of statistical tests, and a set of statistical indicators to better describe the differences in RR interval estimation are also provided. Results show that the spread and agreement depend on the choice of alternative measurements and that the DTS cannot be considered generally as a white or as a normally distributed process. Nevertheless, in controlled measurements the DTS can be considered as a stationary process.

Accuracy of heart rate variability estimation by photoplethysmography using an smartphone: Processing optimization and fiducial point selection.

This work compares several fiducial points to detect the arrival of a new pulse in a photoplethysmographic signal using the built-in camera of smartphones or a photoplethysmograph. Also, an optimization process for the signal preprocessing stage has been done. Finally we characterize the error produced when we use the best cutoff frequencies and fiducial point for smartphones and photopletysmograph and compare if the error of smartphones can be reasonably be explained by variations in pulse transit time. The results have revealed that the peak of the first derivative and the minimum of the second derivative of the pulse wave have the lowest error. Moreover, for these points, high pass filtering the signal between 0.1 to 0.8 Hz and low pass around 2.7 Hz or 3.5 Hz are the best cutoff frequencies. Finally, the error in smartphones is slightly higher than in a photoplethysmograph.

Real time heart rate variability assessment from Android smartphone camera photoplethysmography: Postural and device influences.

The aim of this paper is to present a smartphone based system for real-time pulse-to-pulse (PP) interval time series acquisition by frame-to-frame camera image processing. The developed smartphone application acquires image frames from built-in rear-camera at the maximum available rate (30 Hz) and the smartphone GPU has been used by Renderscript API for high performance frame-by-frame image acquisition and computing in order to obtain PPG signal and PP interval time series. The relative error of mean heart rate is negligible. In addition, measurement posture and the employed smartphone model influences on the beat-to-beat error measurement of heart rate and HRV indices have been analyzed. Then, the standard deviation of the beat-to-beat error (SDE) was 7.81 ± 3.81 ms in the worst case. Furthermore, in supine measurement posture, significant device influence on the SDE has been found and the SDE is lower with Samsung S5 than Motorola X. This study can be applied to analyze the reliability of different smartphone models for HRV assessment from real-time Android camera frames processing.

Association of interleukin-1beta and interleukin-1 receptor antagonist genes with disease severity in MS.

OBJECTIVE: To investigate whether polymorphisms in the interleukin (IL)-1beta and IL-1 receptor antagonist (IL-1RA) genes are associated with both susceptibility to and clinical characteristics of MS. BACKGROUND: Genetic susceptibility to MS is determined by many partially identified genes. The genes encoding various cytokines are logical candidates for MS susceptibility and phenotype. METHODS: Genotypes were determined from 148 patients with clinically definite MS and 98 healthy controls. All the patients were unrelated, Dutch, and white. Patient files were reviewed for disease type, initial symptoms, age at onset of disease, and rate of disease progression. RESULTS: No significant differences in genotypes, allele frequencies, or carrier frequencies were found between MS patients and healthy controls. Stratification for disease type (relapsing-remitting, primary progressive, or secondary progressive) did not provide significant differences between patients and controls. However, a specific IL-1RA/IL-1beta combination was associated with disease severity. MS patients with the IL-1RA allele 2+/IL-1beta allele 2- combination had a higher rate of progression on the Expanded Disability Status Scale when compared with the other possible combinations (p = 0.007). CONCLUSIONS: IL-1RA and IL-1beta are disease severity genes rather than disease susceptibility genes. Furthermore, these gene polymorphisms may define subgroups of patients with a worse prognosis.

A novel robust index to assess beat-to-beat variability in heart rate time-series analysis.

A new index is proposed to estimate the variance of the differentiated heart rate (RR) time series from its truncated histogram. The index is more robust to artifacts than the standard deviation of the differentiated RR time series (rMSDD) and, unlike the pNN50, does not saturate for very high or very low heart rate variability.

Variations in breathing patterns increase low frequency contents in HRV spectra.

This paper shows that variations in breathing patterns broaden heart rate variability (HRV) spectral bands and increase the power amplitude of low-frequency bands. Because of these influences, spectral markers for HRV signals, such as the quotient between spectral power at different frequency bands, should be compared only under controlled breathing conditions or after considering the effect of variations in breathing patterns.

Bias and uncertainty in heart rate variability spectral indices due to the finite ECG sampling frequency.

Spectral analysis of the heart rate variability is becoming a usual tool as a marker of the autonomic nervous system. The final output of the spectral analysis is a set of indices that are always estimators due to technical limitations. In this work, the bias and the uncertainty in the VLF, LF, HF and LF/HF indices due to the finite sampling frequency of the ECG are analysed. The results indicate that for low sampling frequency (125 Hz), the bias and uncertainty in the HF and LF/HF indices can blur the results of the analysis, especially if the RR time series has low variability. The HF index is overestimated and, accordingly, the LF/HF index is underestimated. Then, results from RR time series with low sampling frequency must be used with care. The uncertainty of the spectral indices is proportional to the inverse of the sampling frequency and the bias is proportional to the inverse of the square sampling frequency.

A new index for the analysis of heart rate variability dynamics: characterization and application.

A new index for the analysis of the heart rate variability (HRV) dynamics is presented. The proposed index (acceleration change index (ACI)) characterizes the sign of the differences of a time series. A theoretical study shows an expression that relates ACI and the autocorrelation function of the time series. This formula is tested and validated with different simulated time series (uncorrelated noise, sinusoidal and fractional Brownian motion). Next, ACI is applied to RR time series from healthy subjects showing that ACI decreases with periodic controlled breathing, increases during exercise, and it has a lower value at night than during the day. In a preliminary study, ACI has been shown to be lower in healthy subjects than in patients who had suffered a myocardial infarction one month previously. ACI can be employed as a fast and robust new marker of the HRV dynamics.

[Nitric oxide and inflammatory bowel disease]. / Oxido nítrico y enfermedad inflamatoria intestinal.

Nitric oxide (NO) is an important biological mediator with effects on homeostasis, neurotransmission and immune function. Chronic inflammation of the intestinal mucosa in patients in ulcerative colitis and Crohn's disease has been reported to be associated with enhanced production of NO and nitric oxide synthase (NOS) activity. Whereas small amounts of NO produced by endothelial constitutive calcium-dependent NOS may act to preserve intestinal mucosa integrity, large amounts of NO synthesised by inducible calcium-independent NOS may play a key role in further aggravation of the inflammation and may be associated with the development of intestinal mucosal injury and amplification of immune response in patients with inflammatory bowel disease (IBD). In this article we review NO pathways, mechanisms of action, functions, regulation, immunogenetics and the role played in IBD. A deeper knowledge of the NO physiopathology may allow new therapeutical approaches in IBD patients. In fact, the development of selective inhibitors of NOS isoforms could provide a novel therapeutic option in the management of IBD patients.

[Risk factors in adult periodontitis: polymorphism in the interleukin-1 gene family]. / Risicofactoren voor adulte parodontitis: polymorfismen in de interleukine-1 genfamilie.

Interleukin (IL)-1 alpha, IL-1 beta and IL-1 receptor antagonist (ra) play a major role in regulation of the inflammatory response in periodontal tissues. The aim of this study was to investigate the distribution of genetic variation in the IL-1 gene family among periodontitis patients and controls, taking into account smoking and microbiology as additional variables. There were 53 non-smoking and 52 smoking patients with severe adult periodontitis and 53 periodontal healthy controls genotyped for genetic variation in the IL-1 gene family. The presence of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans was established by culture techniques. A higher frequency of genotype+ (IL-1A*2 + IL-1B*2 + IL-1RN*2) was found in non-smoking periodontitis patients in whom P. gingivalis and A. actinomycetemcomitans could not be detected (42.1% vs. 11.3% in controls; p = 0.0068; or 5.7, 95% ci: 1.6-19.8). This data provide evidence that polymorphisms in genes of the IL-1 family are associated with severe adult periodontitis and may be a risk factor for severe periodontitis.

Heart rate variability analysis using a seismocardiogram signal.

Seismocardiography is a simple and non invasive method of recording cardiac activity from the movements of the body caused by heart pumping. In this preliminary study we use a smartphone to record this acceleration and estimate the heart rate. We compare the heart rate variability parameters from the seismocardiogram and ECG reference signal. The results show a great similarity and are strongly influenced by the instability in the sampling frequency of the device. The differences between RR series are lower than 10 ms.

Characterization of the prostaglandin E2 pathway in a rat model of esophageal adenocarcinoma.

UNLABELLED: Accumulating evidence indicates that the cyclooxygenase-2 (COX-2)/prostaglandin E2 (PGE2) pathway plays a key role in esophageal carcinogenesis. A better understanding of the pathway downstream of COX-2 may reveal novel targets for the prevention of esophageal adenocarcinoma (EAC). The objective of this study was to characterize the profile of genes involved in PGE2 metabolism and signaling in an experimental model of EAC. Esophagojejunostomy with gastric preservation was performed in wistar rats to induce gastroduodenal reflux. Rats were sacrificed 2 or 4 months after surgery. Nine non-operated rats were used to obtain normal (control) esophageal tissues. RESULTS: All rats that underwent esophagojejunostomy developed inflammation. In addition, 90% of the animals showed intestinal metaplasia; of those, 40% progressed to AC. This process was accompanied by a significant increase in esophageal PGE2 levels and the induction of both mRNA and protein levels of COX-2, COX-1, prostaglandin E synthase, 15-hydroxyprostaglandin dehydrogenase, and PGE2 receptors EP3, EP4 and especially EP2, which rose to particularly high levels in experimental rats. In addition, exposure to a selective COX-2 inhibitor (SC58125) or an EP1/EP2 antagonist (AH6809), but not an EP4 antagonist (AH23848B), significantly reduced cell proliferation of esophageal explants in 24 hour-organ culture experiments. Our data suggest that, in addition to COX-2, other components of the PGE2 pathway, including COX-1, may play important roles in the development of EAC induced by gastroduodenal reflux in the rat. Although it must be confirmed in vivo, the EP2 receptor may represent a promising selective target in the prevention of Barrett's associated AC.

Drowsiness detection by thoracic effort signal analysis in real driving environments.

Detection of drowsiness while driving is a leading objective in advanced driver assistance systems. This work presents a new index to assess the alertness state of drivers based on the respiratory dynamics derived from an inductive band. More than 100 hours of driving in real environments from 13 healthy subjects were analyzed. The proposed method has a sensitivity of 93.7% and specificity of 86.3% in detecting full awake drivers while it has a sensitivity of 83.1% and specificity of 95.3% in detecting drowsy drivers. The results show that the proposed index may be promising to assess the alertness state of real drivers.