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Unlabelled: Podocin is a protein located in the glomerular slit diaphragm where it takes part in the regulation of glomerular filtration. Mutations of the NPHS2 gene that codes podocin are the main cause of autosomal recessive steroid resistant nephrotic syndrome (SRNS)., Objectives: To identify the NPHS2 mutations in Chilean children with SRNS, and to determine the prevalence of the most common variants in a group of healthy adults., Patients and Methods: Mutation analysis of NPHS2 in 34 Chilean children with SRNS. Once the two most common variants of NPHS2 were identified, screening for these mutations was performed on 233 healthy adults. The mutation analysis was performed by the direct sequencing of the eight coding exons by polymerase chain reaction amplification. The DNA sequencing was performed using a fluorometric method, and then evaluated with SeqPilot software. The relationship between the presence of NPHS2 variants and SRNS was calculated by comparing the allele frequency between patients with SRNS and those of the healthy volunteers using the exact Fisher test. A P<.05 was considered significant., Results: Pathogenic NPHS2 mutations were detected in 7 (21%) of the 34 patients studied, of which 6 were heterozygotes for p.R229Q and p.A284V. The presence of p.R229Q was 2.46% in the healthy volunteers., Conclusions: This study shows that p.R229Q and p.A284V are the most frequent variants in Chilean children with SRNS. It is the first time that this relationship has been reported in Chilean children. Based on this, a screening strategy is proposed for the genetic study in patients with SRNS and their families. A parallel or sequential search strategy for p.R229Q and p.A284V in these patients is proposed., (Copyright © 2015 Sociedad Chilena de Pediatría. Publicado por Elsevier España, S.L.U. All rights reserved.)

Purpose: To evaluate the autofluorescence findings in patients diagnosed with pseudoxanthoma elasticum., Material and Methods: A prospective study was conducted on 18 eyes of 9 patients who had ocular pathology andfollowed up in the pseudoxanthoma elasticum (PSX) unit of our hospital. We evaluated the best corrected visual acuity (BCVA), colour and autofluorescence photography (AF), and fluorescein angiography (FA) in patients with choroidal neovascularitation., Results: Of the 9 patients, 7 were women and 2 were men. The mean age was 40 +/- 14 years. The BCVA ranged from 1 to 0.01 (Mean 0.65 +/- 0.4). All patients showed PSX injuries. Angioid streaks (AS) 18 (100%), peau d'orange 16 (87.5%) and pigmented fibrotic plates 5 (31,5%). We observed different hypoautofluorescence patterns (RPE atrophy), of which 2 of them were AS patterns (irregular lines with hyperautofluorescence speckled in its interior and edges, or bands with lobulated lesions inside and hyperautofluorescence at the edges), and finally widespread areas of hypoautofluorescence, larger than observed by ophthalmoscope., Conclusions: Autofluorescence in patients with PSX is an easy method to evaluate the initial level of ophthalmoscopic involvement and its subsequent progression. The extensive changes in the retinal pigment epithelium (RPE) suggests the important role of this in the physiopathology of the disease., (2010 Sociedad Española de Oftalmología. Published by Elsevier Espana. All rights reserved.)

Background: The packaging of heterochromatin during spermatogenesis has been correlated with the expression of residual apoptotic bodies (which stain with merocyanine A) that will impact on sperm function in the fertilization process; as well as the joint expression of the transmembrane translocation phosphatidyl serine and oligonucleosomes., Objective: To evaluate the expression of bodies stained with merocyanine in the functional processes of sperm and their level of agreement with apoptotic Annexin V and TUNEL biomarkers., Material and Method: We performed a prospective, cross, including 11,000 cells belonging to semen samples from infertile men, were evaluated according to WHO criteria (1999), bounded by the lines of Tygerberg. The biomolecular transformation processing of the membrane and the expression of oligonucleosomes in the terminal cascade of apoptosis were quantified by cytometry flow, using an argon lasser as a reading source of 480 nm, discriminating the degree of cellularity, both negative and positive for each indicators., Results: Because of the study design was found low quantification in semen parameters, motility, morphology and sperm concentration. The average expression of cells [DNA-PI(+) / dUTP-FITC(+)] (quantification of TUNEL) and [Annexin-V(+) / PI(-)] was 36.5 +/- 17.4% and 31.2 +/- 17.4%, respectively. By comparing the expression of TUNEL without the effect of M540 bodies (36.3 +/- 1.7% vs. 36 +/- 1.7%) a significant difference was not determined., Conclusions: This study shows that there is a remnant of the primary processes of spermiation, which can take an important role in apoptotic and functional processes of the sperm. However, its expression does not affect measurement of biomarkers of apoptosis seminal, whose determination changed the diagnosis and functional perception of reproductive parameters in the sperm.

Background: Erythrasma is a superficial infection caused by Corynebacterium minutissimum and affects the major skin folds and the interdigital regions of the feet. It is characterized by erythematous, brown, scaly patches and maceration, and exhibits coral-red fluorescence under Wood light., Objective: The aim of this study was to determine the frequency of erythrasma in patients with interdigital lesions., Methods: An open, prospective, longitudinal, observational study was performed in a hospital in Mexico City between March and December, 2006. All patients with interdigital lesions were examined with a Wood lamp and direct examination was performed with 20 % potassium hydroxide. Cultures were done in Sabouraud dextrose agar and brain heart infusion agar, and smears were analyzed. General characteristics and concomitant diseases were recorded., Results: We examined 73 patients, of whom 24 (32.8 %) were diagnosed with erythrasma based on coral-red fluorescence under Wood light and identification of corynebacteria by Gram staining. The disease was more common in women (83.33 %) and the mean age of the patients was 43.5 years. The main clinical findings were scaling and maceration, and the fourth interdigital web was the most commonly affected. Corynebacterium could not be isolated in any of the cases. Mycology was positive in 15 cases (62.5 %) and the following microorganisms were isolated: Candida (16.6 %), dermatophytes (12.5 %), and Trichosporon (4.1 %)., Conclusions: Interdigital erythrasma is a common condition and can be easily confused with interdigital tinea. It persists if not treated appropriately. Rapid diagnosis is easily obtained by examination with a Wood lamp, while culture is difficult and unnecessary for diagnosis. The coexistence of erythrasma with dermatophytes and Candida should be considered when the interdigital webs are affected.

Species within the Mycobacterium genus are of major medical interest, since, together with environmental and opportunistic species, there are two species (Mycobacterium tuberculosis and Mycobacterium leprae) that remain an important public health challenge. Despite efforts to control tuberculosis (TB), this disease remains one of the most prominent health problems worldwide. In the last few years, mycobacteriology has experienced major technological advances. Nevertheless, the early diagnosis of mycobacterial infection and, especially of TB, is still based on microscopic examination of properly stained samples. At present, this procedure is still the simplest, fastest and most cost-effective method for preliminary diagnostic guidance. Effective control of TB is based on rapid detection of M. tuberculosis, followed by immediate implementation of the appropriate antituberculosis therapy. Because of the emergence of multidrug resistant strains, the development of rapid diagnostic methods, both for identification of M. tuberculosis and susceptibility testing, has become a pressing need. The availability of molecular epidemiology methods that are easy to implement and standardized and that would allow identification of related cases is of key importance to identify epidemic outbreaks and control the spread of TB. Despite the evident progress in the molecular diagnosis of mycobacterial infections, the available techniques are still inadequate. In this review, we describe the state of the art of the main molecular techniques for direct detection of mycobacteria in clinical samples, their identification, detection of resistance to the most important antituberculosis agents, and molecular epidemiology. In each case, we describe the advantages and limitations of current techniques. In the near future, clinical mycobacteriology will probably evolve to the universal use of genetic techniques for direct diagnosis and detection of resistance. The molecular epidemiology of TB will be performed, in its various applications, by faster and more automated techniques than those currently available.

Veillonella spp. are anaerobic gram-negative cocci associated to oral health. Different types of cultures have been reported for the isolation of these microorganisms. Veillonella spp. colonies produce a red fluorescence, which is made visible through ultraviolet light and disappears in contact with oxygen. This feature would be very useful for rapid presumptive identification. The aims of this study were: 1. to compare the Rogosa selective medium for Veillonella with the cultures recommended by different authors in order to determine best saliva recovery, since this sample is generally used to determine the presence and predominance of this bacteria; 2. to detect red fluorescence production on these different culture media as a rapid method for identification. Selective medium for Veillonella, Schaedler agar for anaerobic bacteria with vitamin K, thioglycollate agar, brain heart infusion agar, Brucella agar, trypticase soy agar, and Columbia agar, all of them with and without the addition of vancomycin, and laked blood were used for this study. The tested sample was a saliva pool. Both, Veillonella colonies, and the total number of microorganisms were counted, and expressed as CFU/ml of saliva. The greatest Veillonella recovery in saliva was obtained with the selective medium for Veillonella with vancomycin and laked blood. The production of fluorescence was only observed in this medium.

We evaluated RNA/DNA ratio as an index of physiological condition during larval development of a hybrid between the fishes Colossoma macropomum (cachama) and Piaractus brachypomus (morocoto). The samples were obtained by induced reproductive technology and the eggs were maintained in acrylic conical incubator with a continuous waterflow. Embryonic development, from egg fertilization to cell division and hatch out, took 12 hours 20 minutes at 29.5 degrees C, dissolved oxygen contents of 6.0 ppm and pH 7.5. Nucleic acids quantification was determined by fluorometry with ethidium bromide and Hoechst 33258 dyes. We observed significant changes of RNA/DNA ratios during all stages of the embryonic larval development. Therefore, RNA/DNA relation is an useful technique to evaluate physiological condition in short period and could be utilized as nutritional condition and/or instantaneous growth for routine check to verify the health status in early life of cultivated species.

Introduction: Pyroglutamyl aminopeptidase (pGluAP) is an omega peptidase widely distributed in fluid and tissues, which hydrolyses biological active peptides including thyrotropin-releasing-hormone (TRH). In the last years, different endocrine and extraendocrine functions have been attributed to TRH., Objectives: The aim of present work is to study the influence of high dietary cholesterol on soluble and membrane bound pGluAP activity in frontal cortex of male and female mice. Material and methods. Soluble and membrane bound pGluAP activities of frontal cortex of mice feeding during 15 days, a standard diet enriched with cholesterol (1%) and cholic acid (0.5%) were measured using fluorimetric assays., Results: Significant increases were observed in membrane bound pGluAP activity in males and females. No changes were detected in soluble pGluAP activity in frontal cortex of females but a significant increase was observed in this enzymatic activity in males., Conclusions: High dietary cholesterol induces a significant increase in tissue pGluAP activity. These increases may produce changes in TRH metabolism which may be related with alterations in its neuromodulatory functions and with the possible relationship between TRH and other neurotransmitter systems.

In this work, different aminopeptidase activity levels in whole human saliva are described. Aminopeptidase activities were studied by measuring the rate of hydrolysis of the artificial substrates Ala-, pGlu-, Pro-, Arg-, Asp- y Cis-2-naphthylamides (fluorimetrically detected at 412 rim with excitation at 345 nm). The presence of these enzyme activities in the saliva suggests that the active levels of saliva peptides can be controlled by homeostatic mechanism similar to those that have been described in other tissues, such as plasma, the central nervous system, and immunocompetent cells.

Background: To study distinctive clinical features of pheochromocytoma in the setting of multiple endocrine neoplasia type 2A (MEN 2A) in comparison with sporadic pheochromocytoma., Patients and Method: All patients diagnosed of sporadic pheochromocytoma (n = 29) and in the setting of MEN 2A (n = 16) between 1976 and 1998 in a tertiary hospital were included in the study. The following variables were compared: age at diagnosis, symptoms, presence and characteristics of hypertension, size of the tumor, localization and malignancy. The accuracy of diagnostic tests was also evaluated., Results: The pheochromocytoma associated to MEN 2A had a higher prevalence in our study (35.5%) and the diagnosis was performed earlier than for sporadic pheochromocytoma (29.1 [7.8] vs. 47,5 [10. 9] years; p < 0.001). Other distinctive features in comparison with sporadic pheochromocytoma were: bilaterality (81.25 vs. 3.44%; p < 0. 001), absence of symptoms (44 vs 11%; p < 0.05) and paroxysmal hypertension. Concerning the diagnostic tests, it must be noted the low sensitivity (60%) of vanillylmandelic acid in MEN 2A., Conclusions: The pheochromocytoma in the setting of MEN 2A is diagnosed at younger age and frequently asymptomatic and bilateral. In view of the high prevalence of MEN 2A in our country, a genetic analysis should be performed in all patients with pheochromocytoma, especially when it is bilateral.

LIFE system (lung imaging fluorescence endoscope) is a new system of detection for early lung cancer using a He-Cd laser through a conventional fiberoptic bronchoscope. This system is able to capture and processing the autofluorescence from the bronchial mucosa and to show it in a video screen in real time. In the screen the normal tissue appears in green colour and the neoplastic tissue in brown colour. The first studies performed with LIFE system show the same specificity of the conventional bronchoscopy but the sensibility is 50% higher. In this paper we present the first case of detection of a occult squamous cell carcinoma with sputum and bronchial aspiration cytology positive for carcinoma but chest X ray, CT scanner and bronchoscopy negative for lung cancer. The LIFE system was able to detect carcinoma in the carina.

Introduction: The aminopeptidases are considered to be enzymes which can regulate the activity of various neuropeptides. However, their precise function has not yet been fully determined., Objective: With a view to studying the function of these enzymes at the cerebral level, the subcellular distribution of soluble leucine aminopeptidase from left and right cerebral hemisphere, in rats of different pre-natal and post-natal ages was determined as described in this paper., Materials and Methods: The activity of soluble leucine aminopeptidase was determined fluorimetrically in the sub-cellular fractions obtained from the left and right cerebral hemispheres of rats of different pre-natal and post-natal ages: fetuses (of 19-21 days gestation), 1 week, 1, 5 and 24 months of age., Results: No differences were observed between the cerebral hemispheres at any of the ages studied. However, significant differences were seen in the evolutionary behaviour of the two hemispheres. At all ages, except 24 months, there were significant differences between the fractions. In the fetuses, most activity was seen at the level of the synaptosomal raw fraction and least in the microsomal. At 1-week-old, the highest levels were seen at a microsomal level. In rats of 1 and 5 months of age, the highest levels were detected at synaptosomal level. With regard to the evolution with age, in the homogenate, nuclear fraction, cytosol and mitochondrial fraction a similar profile was observed, with a decrease after birth and an increase at one month, being stable at the other ages. At a synaptosomal level the profile is similar, although the differences are much more marked, with a drastic decrease in activity at two years of age. By contrast, the microsomal fraction showed a very different evolutionary profile with an increase after birth, becoming stable at later ages., Conclusions: The activity of soluble leucine aminopeptidase shows a heterogeneous subcellular distribution together with significant differences during cerebral development and ageing. The behaviour of this enzyme may reflect the functional state of its neuropeptide substrates.

Background: A comparative evaluation of the intracellular penetration of norfloxacin, ofloxacin, levofloxacin, fleroxacin and lomefloxacin was carried out in human polymorphonuclear leucocytes (PMNs)., Methods: The fluorometric method was used based on the natural fluorescence of some fluoroquinolones., Results: All the fluoroquinolones accumulated within the human polymorphonuclear leucocytes achieving a quotient of between 4 to 6 in intracellular and extracellular concentration (I/C) with no significant differences being observed between the two. Penetration was rapid and was not saturable within the intervals of extracellular concentrations tested. The intracellular penetration of these fluoroquinolones was dependent on temperature and cell viability was altered in only levofloxacin and norfloxacin. The I/E values obtained significantly decreased when the cells were incubated at 4 degrees C. These values showed an important increase to pH 5 and decreased to pH 9, with this pH being statistically significant only in ofloxacin., Conclusions: All the fluoroquinolones evaluated penetrated rapidly into the phagocytic cells achieving intracellular concentrations which were several fold higher than the extracellular concentrations, with no notable differences being observed being the two, thus providing an additional advantage for their use in infections produced by microorganisms capable of surviving intracellularly.

We report 93 patients with catecholamine producing tumors that were analyzed at the Hormone Laboratory of the Institute of Cardiology. They are 75 pheochromocytoma patients and 18 children with neuroblastoma. Fluorimetric methods were used to measure urinary and plasma catecholamines on neuroblastoma and pheochromocytoma patients. Dopamine high excretion (mean value 2889 micrograms/24 hs), was constantly observed in the neuroblastoma children as were adrenaline and noradrenaline in the benign and malignant pheochromocytoma patients. The mean values for the malignant tumours were 53 for adrenaline and 1436 micrograms/24 hs for noradrenaline. Structural and biochemical differences of the catecholamine producing tumours are reflected on the clinical manifestations which are observed in the patients bearing such neoplasms.

The seric activity of the enzyme NAG (N-acetyl-beta-D-glucosaminidase) was studied in 3 groups of patients with jaundice of different etiology (extrahepatic, intrahepatic and acute hepatitis) and in a control group, in order to observe the discrimination capacity of this enzyme in each group. After the use of a fluorimetric method, the results were exposed, applying the statistical test, comparing different variables; the results are discussed and we conclude that NAG is an enzyme of easy detection and low cost to be included in the group of tests to study the biological pattern of jaundice.

The concentrations of free amino-acids have been determined in 54 samples of human milk: 18 in colostrum, 18 in transition milk and 18 in mature milk, all of them taken from women in the Eastern part of Andalucía. The amino-acids have been determined through high pressure ion-exchange chromatography and fluorimetric detection. Glutamic acid is the major free amino-acid, representing 21-52% of the total amount of these components in human milk. Its level increase significantly with advancing lactation. Taurine represents 14-19 of total free amino-acids in human milk, remaining its levels practically unchanged during the whole lactation. The fact that the major content of essential free amino-acids in present in human colostrum could suppose a contribution of these substances which are directly absorbable during a period of life when the nutritional requirements of the neonate are incremented.

The application of the histamine liberation test to the diagnosis and follow-up of allergic processes has been pursued by many authors. Since Shore first described a manual fluorimetric technique, efforts have been directed at automating the system and reducing the sample volume. According to the latest contributions by Moneo et al., from 300 to 400 determinations can be carried out in a single work day, and from 40 to 50 determinations with 10 ml of blood. In the present work we present our experienced in updating the technique and the system of preparation of a histamine standard, which we consider of great practical use in the application of this technique routine. Using the standard solution as reference, we calculated total and basal histamine levels in 342 presumed healthy individuals; the mean values were 69.5 ng/ml for total histamine and 8% for basal histamine in males, and 71 ng/ml for total histamine and 6% for basal histamine in females. Age range was from 17 to 57 years, and there were no differences between sexes. The maximum value was 176 ng/ml and the minimum 32 ng/ml. The reaction of O-phthaldialdehyde with histamine is not chemically specific. In the presence of light, aldehydes self-polymerize; moreover, O-phthaldialdehyde can react with other products in addition to histamine. The first problem is avoided by protecting the reagents from light, and the second by using specific absorption and emission filters which do not pick up signals from reaction side products. Nevertheless, at the quantitative level these effects are negligible when compared to other causes which, although not part of the chemical reaction, are also involved in the process (antigen standardization, etc). By means of this technique, from 300 to 400 determinations can be carried out in a single work day, making it possible to use this technique in the daily work routine.