Longitudinal study of neonatal brain tissue volumes in preterm infants and their ability to predict neurodevelopmental outcome.

Premature birth has been associated with poor neurodevelopmental outcomes. However, the relation between such outcomes and brain growth in the neonatal period has not yet been fully elucidated. This study investigates longitudinal brain development between birth and term-equivalent age (TEA) by quantitative imaging in a cohort of premature infants born between 26 and 36 weeks gestational age (GA), to provide insight into the relation of brain growth with later neurodevelopmental outcomes. Longitudinal T2-weighted magnetic resonance images (MRI) of 84 prematurely born infants acquired shortly after birth and TEA were automatically segmented into cortical gray matter (CGM), unmyelinated white matter (UWM), subcortical gray matter (SGM), cerebellum (CB) and cerebrospinal fluid (CSF). General linear models and correlation analysis were used to study the relation between brain volumes and their growth, and perinatal variables. To investigate the ability of the brain volumes to predict children's neurodevelopmental outcome at 18-24 months and at 5 years of age, a linear discriminant analysis classifier was tested and several general linear models were fitted and compared by statistical tests. From birth to TEA, relative volumes of CGM, CB and CSF with respect to total intracranial volume increased, while relative volumes of UWM and SGM decreased. The fastest growing tissues between birth and TEA were found to be the CB and the CGM. Lower GA at birth was associated with lower growth rates of CGM, CB and total tissue. Among perinatal factors, persistent ductus arteriosus was associated with lower SGM, CB and IC growth rates, while sepsis was associated with lower CSF and intracranial volume growth rates. Model comparisons showed that brain tissue volumes at birth and at TEA contributed to the prediction of motor outcomes at 18-24 months, while volumes at TEA and volume growth rates contributed to the prediction of cognitive scores at 5 years of age. The family socio-economic status (SES) was not correlated with brain volumes at birth or at TEA, but was strongly associated with the cognitive outcomes at 18-24 months and 5 years of age. This study provides information about brain growth between birth and TEA in premature children with no focal brain lesions, and investigates their association with subsequent neurodevelopmental outcome. Parental SES was found to be a major determinant of neurodevelopmental outcome, unrelated to brain growth. However, further research is necessary in order to fully explain the variability of neurodevelopmental outcomes in this population.

Aortic Pseudoaneurysm Causing Compression of the Left Main Coronary Artery.

A 75-year-old man with a history of mechanical aortic valve replacement with aortic conduit for severe aortic insufficiency underwent routine screening computed tomography evaluation revealing right coronary anastomosis endoleak and proximal aortic root pseudoaneurysm.

Receptorphin: a conserved peptide derived from the sequence of the opioid receptor, with opioid displacement activity and potent antiproliferative actions in tumor cells.

BACKGROUND: In addition to endogenous opioids, a number of peptide sequences, derived from endogenous (hemorphins, alphaS1-casomorphin), and exogenous proteins (casomorphins, exorphins) have been reported, possessing opioid activity. In the present work, we report the identification of a new peptide, receptorphin (Tyr-Ile-Phe-Asn-Leu), derived from the sequence of the second transmembrane loop of the opioid receptor. This sequence is unique for the opioid receptor, and conserved in all species and receptor-types. RESULTS AND DISCUSSION: Receptorphin competes for opioid binding, presenting a kappa-receptor interaction, while it binds equally to delta- and mu- opioid and somatostatin-binding sites, and inhibits the cell proliferation of a number of human cancer cell lines, in a dose-dependent and reversible manner, at the picomolar or the nanomolar range. Receptorphin shows a preferential action on prostate cancer cells. CONCLUSION: Our work identifies, for the first time a peptide, in a receptor sequence, possessing ligand-agonistic activities. A hypothesis, based on receptorphin liberation after cell death, is presented, which could tentatively explain the time-lag observed during opioid antiproliferative action.

Microscopical examination of the localisation patterns of two novel rhodamine derivatives in normal and neoplastic colonic mucosa.

Tissue characterisation by fluorescence imaging, using exogenous fluorophores, is a promising method for cancer detection. Histochemical alterations in the composition of mucins, when neoplastic transformations occur, could be exploited to derive more selective fluoroprobes indicative of early malignant transformation. The aim of this work was to develop and examine tumour selective fluoroprobes for colon cancer diagnosis, as well as to determine the morphological components where selective dye accumulation has occurred. Two novel fluoroprobes: rhodamine B-L-leucine amide and rhodamine B-phenylboronic acid were synthesised and examined together with Mayer's mucicarmine, alexa 350-wheat germ agglutinin (WGA) and tetramethyl rhodamine-concanavalin A (ConA). Fluorescence microscopy studies were performed with deparaffinised human colon sections, using an epifluorescence microscope equipped with a colour CCD camera. The intense accumulation of the novel fluoroprobes was localised in the amorphous material in the lumen of neoplastic crypts. To gain insight into the localisation patterns, mucicarmine, alexa 350-WGA and tetramethyl rhodamine-ConA were used. Alexa 350-WGA reacted primarily with mucin secreted in the malignant crypt lumen suggesting that this material is rich in sialic acid and N-acetylglucosaminyl residues. These derivatives clearly and consistently distinguished non-neoplastic from neoplastic human colon tissue sections. The intense accumulation at the altered mucins indicates that they could be used as fluoroprobes of biochemical alterations for carcinoma detection.

Texture analysis of fluorescence microscopic images of colonic tissue sections.

The aim of this study was to assess the potential of texture analysis for the characterization of fluorescence images from colonic tissue sections stained with a novel and selective fluoroprobe, Rhodamine B-phenylboronic acid. Fluorescence microscopy images of colonic healthy mucosa (n = 35) and adenocarcinomas (n = 35) were digitally captured and subjected to image texture analysis. Textural features derived from the grey level co-occurrence matrix were calculated. A modified version of the multiple discriminant analysis criterion was used to choose an appropriate subset of features. A minimum Mahalanobis distance, linear discriminant classifier and a simple evaluation 'score' method were used to classify image feature data into the two categories. A subset of four textural features was selected and used for the description and classification of each image field. They were found appropriate to correctly classify 95% of the images into the two classes, using two different classifiers. These features contained information about local homogeneity and grey level linear dependencies of the image. This study demonstrated that texture analysis techniques could provide valuable diagnostic decision support in a complex domain such as colorectal tissue.

Relatively low red cell folate levels and acute coronary syndromes.

BACKGROUND: Low folate levels are related to increased risk for coronary artery disease in humans, while experimental work has shown that folate deficiency is thrombogenic. We hypothesized that relatively low folate levels are related to the development of acute coronary syndromes in patients with previously stable coronary artery disease. METHODS: One hundred and forty-one men were studied: 53 consecutive patients with acute coronary syndromes, 41 with stable coronary artery disease and 47 control participants. Known clinical and lipid risk factors were identified in all subjects and in addition plasma B12, plasma and red cell folate levels were measured. RESULTS: Red cell folate levels were significantly lower in patients with acute coronary syndromes (510+/-178 nmol/l) than in both stable coronary artery disease patients (638+/-264 nmol/l, P< 0.005) and controls (615+/-193 nmol/l, P< 0.05 respectively). Plasma folate and B12 levels were similar in all three groups. Multiple logistic regression analysis identified red cell folate levels as the only independent predictor of acute coronary events in the whole population of patients with known coronary artery disease and in the subgroup of non-smokers (P=0.010 and P=0.031). CONCLUSIONS: The present study suggests that relatively low red cell folate levels are associated with acute coronary syndromes and are an independent predictor of acute coronary events.

Agents facilitating the electric field-induced fusion of intact rabbit erythrocytes.

In the recent years, the role of specific membrane active agents in the electrofusion process has started to draw attention, and it has been found that the presence of various substances in the cell medium can affect the fusion process either in a positive or negative way. In this work, the effect of several proteins, bivalent cations and antibiotics was tested with respect to their ability to protect intact erythrocytes from hemolysis and facilitate the fusion process. The effect of different sugars was also studied. Among the different proteins, pronase and proteinase were found to be the most effective. With respect to bivalent cations, Ca2+ and Mn2+ were more effective while Mg2+ was less important. From the antibiotics, penicillin caused a negative effect while streptomycin acted positively. Finally, glucose medium was found to be the most effective compared to all sugars tested. The results indicated that there are strong differentiations of the induced effects caused by each substance, and some possible mechanisms of action of these agents on the erythrocyte membrane were discussed.

Somatostatin and opioid receptors in mammary tissue. Role in cancer cell growth.

Somatostatin and opioid systems, are the two main inhibitory systems in mammals. Both classes of substances have been identified in normal and malignant mammary gland, as well as their cognitive receptors. They have been implied in the inhibition of cell growth of cancer cells and cell lines, in a dose-dependent and reversible manner. Somatostatin acts through homologous receptors (SSTRs), belonging to five distinct classes (SSTR1-5). We, and others have identified SSTR2 and 3 as been the only SSTRs present in the breast. Furthermore, opioids act through the three classes of opioid receptors (mu, delta,kappa). In the breast, kappa opioid receptor subtypes (kappa 1-kappa 3) are the most widely expressed. We further have shown that opioids, in addition to their binding to opioid receptors, compete for binding to SSTRs. This functional interaction, together with other identified modes of opioid action in the breast (modulation of steroid receptors, proteases' secretion, interaction with cytoskeletal elements), will be discussed, taking into consideration also the possible local production of casomorphins (casein-derived opioids), which are very potent antiproliferative agents.

The use of soft and flexible guidewires in the treatment of chronic total coronary occlusions by activated guidewire angioplasty.

Activated guidewire angioplasty (AGA) is a new technique which has been designed to assist in angioplasty of total occlusions. The purpose of this study was to determine the safety and efficacy of using flexible relatively soft guidewires (floppy wires) in conjunction with this technique and also to determine the predictors of lesion crossing and final success by this technique in patients with chronic total coronary occlusions. 73 patients with 73 chronic total coronary occlusions in whom coronary angioplasty using conventional techniques had failed were treated with AGA using floppy guidewires. The success of crossing these lesions was 65.7% (48/73) resulting in a final angioplasty success of 56.1% (41/73). Angioplasty success was reduced compared with crossing success in seven arteries in which complications occurred during balloon angioplasty. Multiple stepwise logistic regression analysis identified the location of the occlusion (right coronary artery, p = 0.005) as independent predictor of crossing success of this technique and the male gender (p = 0.03), the duration of occlusion (p = 0.05), the lesion length (p = 0.01) and the location of the occlusion (right coronary artery, p = 0.02) as independent predictors of final procedural success of the method.

Use of vibrational angioplasty for the treatment of chronic total coronary occlusions: preliminary results.

The aim of the study was to investigate the feasibility and clinical safety of vibrational angioplasty in the treatment of chronic total coronary occlusions and evaluate the clinical and angiographic factors that are predictive of the procedural success and complications of the procedure. Seventy-eight patients with chronic total occlusions (>3 months) resistant to conventional techniques were treated by vibrational angioplasty using a variety of conventional guidewires. Lesions were successfully crossed in 67 (85.9%) cases and antegrade flow was achieved in 59 (75.5%). Major complications (myocardial infarction and tamponade) occurred in two (2.5%) patients, but no fatalities ensued. Angiographically detectable dissections were seen in 23 (29.5%) patients but only resulted in vessel compromise and reclosure in 5 cases. Multiple stepwise logistic regression analysis identified the duration (<6 months, P = 0.008) and the length of the occlusion (<15 mm, P = 0.03) as independent predictors of final success and the age of the patient (<55 years, P = 0.006) as the only independent predictor of procedural complications. Vibrational angioplasty is a safe technique useful in the treatment of chronic coronary occlusive disease. Patients in whom the procedure is likely to prove most successful may be easily identified by clinical and angiographic features (duration and length of occlusion).

Loading of intact rabbit erythrocytes with fluorophores and the enzyme pronase by means of electroporation.

The application of electric field pulses in cell suspensions is known to alter membrane integrity, resulting in increased membrane permeabilization. This field-induced membrane poration provides the means to load cells with a variety of external substances, useful for clinical applications. In this work, intact rabbit erythrocytes were successfully loaded with low molecular weight fluorescent probes and with the high molecular weight enzyme pronase, which has been shown to mimic the effects of insulin. Attachment of the enzyme onto the cell surface was also achieved by modifying the applied pulse parameters. Both applications were efficient and accompanied by high cell survival rates. In this way, biological carriers loaded with active substances were produced, offering the potentials for useful clinical applications, either for diagnostic or therapeutic purposes.

Identification of a novel opioid peptide (Tyr-Val-Pro-Phe-Pro) derived from human alpha S1 casein (alpha S1-casomorphin, and alpha S1-casomorphin amide).

A new casomorphin pentapeptide (alpha S1-casomorphin) has been isolated from the sequence of human alpha S1-casein [alpha S1-casein-(158-162)], with the sequence Tyr-Val-Pro-Phe-Pro. This peptide was found to bind with high affinity to all three subtypes of the kappa-opioid receptor (kappa 1-kappa 2). When amidated at the C-terminus, alpha S1-casomorphin amide binds to the delta- and kappa 3-opioid sites. Both alpha S1-casomorphin and its amide inhibit in a dose-dependent and reversible manner the proliferation of T47D human breast cancer cells. This anti-proliferative activity was greater for alpha S1-casomorphin, which was the most potent opioid in inhibiting T47D cell proliferation. In T47D breast cancer cells, other casomorphins have been found to bind to somatostatin receptors in addition to opioid sites. In contrast, alpha S1-casomorphin and its amide do not interact with somatostatin receptors in our system.

Photodynamic effects of hypericin on photosynthetic electron transport and fluorescence of Anacystis nidulans (Synechococcus 6301).

We investigated the photodynamic action of hypericin, a natural naphthodianthrone, on photosynthetic electron transport and fluorescence of the cyanobacterium Anacystis nidulans (Synechococcus 6301). The most drastic effect was the inactivation of photosynthetic oxygen evolution in the presence of the electron acceptor phenyl-p-benzoquinone in aerobic cells which required 1 hypericin/5 chlorophyll a for half-maximal effect. Anaerobic A. nidulans was only partially inactivated and variable chlorophyll a fluorescence remained unperturbed suggesting that photoreaction center II was not a target. Further, hypericin, stimulated photoinduced oxygen uptake in the presence of methylviologen in aerobic cells. This action was less specific than the inactivation of oxygen evolution (1 hypericin/0.5-0.7 chlorophyll a for half-maximal effect). Results point to the involvement of molecular oxygen in two ways. Type I mechanism (Henderson BW and Dougherty TJ (1992) Photochem Photobiol 55: 145-157) in which ground state oxygen reacts with excited substrate triplets appears probable for the inactivation of oxygen evolution. On the other hand, Type II mechanism in which excited oxygen singlets react with ground state substrate molecules appears probable in the stimulation of methylviologen mediated oxygen uptake.

Interpretation of diagnostic implications of fluorescence parameters for atherosclerosis in fibrous, calcified and normal arteries.

Fluorescence spectroscopy presents great interest for the diagnosis of atherosclerosis. Nevertheless there are some difficulties in the interpretation of diagnostic information. This could be overcome by precise methods of extraction of the diagnostic parameters and convenient statistical analysis, which are the subject of this work. Fluorescence excitation-emission matrices from different categories of coronary arteries were developed and used to derive the optimum excitation wavelength on the one hand and to assign the spectra to specific chromophores on the other hand. Simple dimensionless functions (Fi) were formed by the ratio of the intensities at selected wavelength and the logistic model was used for statistical analysis. Decision surfaces were drawn and it was estimated that the probability of correct classification is 88%. The algorithm correctly diagnoses 97% of healthy from diseased samples and 80% of fibrous from calcified coronary arteries.

Wide geographic variations of sensitivity to MOTT sensitins in Greece.

Several studies have suggested that large bodies of water are a main source of infection with mycobacteria other than tuberculosis (MOTT). If this is correct, there should be a gradient in the infection rate with MOTT between mountainous and seaside areas. To test this hypothesis, we performed skin testing with tuberculin and sensitins in 19,470 Greek Armed Forces recruits. Initially, several MOTT sensitins were used, but when it became clear that the Mycobacterium scrofulaceum sensitin was the most appropriate, the study was continued with it alone in 17,403 recruits. Finally, in order to evaluate the geographical distribution of sensitivity to sensitins, we studied the results of 8,507 of these recruits living in or near their birthplace. They were divided into three geophysical areas: seaside 3,389 recruits; mountains 2,692 recruits; and inland plains 2,426 recruits. MOTT sensitivity rates were 4.1% in mountainous areas and 7.1% in seaside areas. All small Aegean islands had high MOTT rates (above 8%). In inland plains, high MOTT rates (above 8%) were observed among those living near big rivers. This geographical distribution of MOTT sensitivity supports the theory that large bodies of water are a main source of infection with MOTT.

Purification to homogeneity of an active opioid receptor from rat brain by affinity chromatography.

Active opioid binding proteins were solubilized from rat brain membranes in high yield with sodium deoxycholate in the presence of NaCl. Purification of opioid binding proteins was accomplished by opioid antagonist affinity chromatography. Chromatography using the delta-opioid antagonist N,N-diallyl-Tyr-D-Leu-Gly-Tyr-Leu attached to omega-aminododecyl-agarose (Affi-G) (procedure A) yielded a partially purified protein that binds selectively the delta-opioid agonist [3H]Tyr-D-Ser-Gly-Phe-Leu-Thr ([3H]DSLET), with a Kd of 19 +/- 3 nM and a Bmax of 5.1 +/- 0.4 nmol/mg of protein. Subsequently, Lens culinaris agglutinin-Sepharose 4B chromatography of the Affi-G eluate resulted in isolation of an electrophoretically homogeneous protein of 58 kDa that binds selectively [3H]DSLET with a Kd of 21 +/- 3 nM and a Bmax of 16.5 +/- 1.0 nmol/mg of protein. Chromatography using the nonselective antagonist 6-aminonaloxone coupled to 6-aminohexanoic acid-Sepharose 4B (Affi-NAL) (procedure B) resulted in isolation of a protein that binds selectively [3H]DSLET with a Kd of 32 +/- 2 nM and a Bmax of 12.4 +/- 0.5 nmol/mg of protein, and NaDodSO4/PAGE revealed a major band of apparent molecular mass 58 kDa. Polyclonal antibodies (Anti-R IgG) raised against the Affi-NAL protein inhibit the specific [3H]DSLET binding to the Affi-NAL eluate and to the solubilized membranes. Moreover, the Anti-R IgG inhibits the specific binding of radiolabeled Tyr-D-Ala-Gly-N-methyl-Phe-Gly-ol (DAMGO; mu-agonist), DSLET (delta-agonist), and naloxone to homogenates of rat brain membranes with equal potency. Furthermore, immunoaffinity chromatography of solubilized membranes resulted in the retention of a major protein of apparent molecular mass 58 kDa. In addition, immunoblotting of solubilized membranes and purified proteins from the Affi-G and Affi-NAL matrices revealed that the Anti-R IgG interacts with a protein of 58 kDa.

Factors related to the presence of autoantibodies in patients with chronic mental disorders.

Serum samples from 307 patients with various chronic mental disorders were examined for the presence of several autoantibodies. Autoantibodies detected included antinuclear antibodies (ANA) in 122/307 (39.7%), rheumatoid factor (RF) in 23/307 (7.5%), anticardiolipin antibodies (anti-CL) in 23/304 (7.6%, IgM in 12 patients, IgG in 13 patients). Isolated cases with IgG anti-dsDNA, anti-Ro(SSA), and anti-Ro(SSA)/anti-La(SSB) were also identified. The analysis of data revealed that the aging process in patients studied contributed significantly to the incidence of ANA (p less than 0.0001) and RF (p less than 0.01). In addition, the chronic administration of chlorpromazine (CPZ) was associated with the presence of ANA (p less than 0.03) as well as with the presence of IgM and/or IgG anti-CL antibodies (p less than 0.003). Finally, the diagnosis of schizophrenia correlated with the presence of ANA (p less than 0.001). This study represents the autoantibody profile of patients with chronic mental disorders and emphasizes the multifactorial origin of autoantibody response in psychiatric patients.