Early identification of keratoconus using pachymetric indexes obtained with spectral domain optical coherence tomography.

PURPOSE: To evaluate the diagnostic ability of pachymetric indexes obtained with Spectral Domain Optical Coherence Tomography (SD-OCT) for early detection of keratoconus (Kc). METHODS: 64 patients with Kc in at least one eye (95 eyes, 46 men and 18 women, average age 27.84 ±13.50), 59 healthy control subjects (100 eyes, 28 men and 31 women, average age 27.15 ±16.14). All patients underwent detailed clinical examination, topography and anterior segment OCT. 37 subjects (37 eyes, 27 men and 10 women, average age 24.23 ± 14.24) having one eye with manifest Kc and the fellow eye without clinical signs of Kc were identified. We studied two groups of pachymetric indexes: C1-C2, M1-M2, pCLMI, Pmin-P2 (Group 1 indexes) and PPD, PSD, PSSD and PASD (Group 2 indexes). A ROC (Receiver Operating Characteristic) curve was developed to compare the diagnostic accuracy, relative sensitivity and specificity for each index. RESULTS: In manifest keratoconus, C1-C2, M1-M2, and pCLMI are significantly higher compared to the control group (P<0.0001); for suspect keratoconus, all Group 1 indexes are significantly higher compared to healthy subjects (P<0.0001) excluding M1-M2 obtained using a constant area circle (P = 0.02). Furthermore, for manifest and suspect keratoconus, PPD, PSD, PSSD and PASD are significantly higher compared to the control group (P <0.0001). CONCLUSION: The studied pachymetric indexes in patients with Kc have high diagnostic accuracy and are statistically significant when compared with healthy subjects (p<0.0001) and can provide a useful tool for keratoconus screening.

Long-lasting immunogenicity and safety of a 2009 pandemic influenza A(H1N1) MF59-adjuvanted vaccine when co-administered with a 2009-2010 seasonal influenza vaccine in young patients with type 1 diabetes mellitus.

AIMS: To evaluate the long-lasting immunogenicity and safety of a pandemic vaccine co-administered with a seasonal influenza vaccine in young subjects with Type 1 diabetes. METHODS: Eighty patients (mean age: 16.7 ± 5.5 years, disease duration: 10.2 ± 4.7 years) were randomly assigned to receive a single or a double dose (1 month apart) of MF59-adjuvanted influenza A(H1N1) vaccine, simultaneously with a single dose of a virosome-adjuvanted trivalent influenza vaccine for the 2009-2010 season. RESULTS: One month after immunization, the rate of seroconversion to 2009 pandemic A(H1N1) was 92.5% with an overall 100% proportion of vaccinees with protective antibody titres (≥ 1:40). No significant differences were observed between vaccinees who received the one-dose or the two-dose schedule. Seasonal vaccine induced a significant increase of both seroprotection rates and antibody levels. Local adverse events at the injection site of pandemic and seasonal vaccines were reported by 66.3% and 50% of subjects, respectively. Solicited systemic adverse events, mainly mild in intensity, were reported by 26.7% of vaccinees. No subjects had an influenza-like illness during the 6-month follow-up. CONCLUSIONS: One injection of 2009 pandemic influenza A(H1N1) MF59-adjuvanted vaccine is immunogenic and safe in young patients with Type 1 diabetes who are at increased risk of influenza morbidities. Pandemic vaccine can be safely co-administered with seasonal influenza vaccine.

Early co-circulation of different clades of influenza A/H1N1v pandemic virus in northern Italy.

INTRODUCTION: The spatial diffusion over time of pandemic influenza A/HINI virus (A/HIN1v) was surveyed in Northern Italy (nearly 10 million inhabitants)from April to December 2009, and the molecular characteristics of circulating viruses were analyzed to identify the appearance of drift variants. About 45% of analyzed samples were laboratory-confirmed cases of A/HINlv. Sporadic cases occurred until the middle of June 2009, then, case numbers began to increase delineating distinct epidemiological phases of viral circulation. METHODS: RNA was extracted using RNeasy Mini kit (QIAGEN GmbH, Germany). Virological diagnosis of A/HINlv infection was carried out by real-time RT-PCR assay. Sequence analysis of hemagglutinin (HA) gene was performed through a RT-PCR assay specific for a 995 bp fragment (nt. 64-1,058) in the HAl domain. The nucleotide sequences were obtained by automated DNA sequencing. The HAl sequences were aligned with other sequences collected from GenBank database by ClustalX software. The multiple sequence alignment was used to perform a basic phylogenetic analysis and a phylogenetic tree from HA sequences was constructed. RESULTS: The HA gene sequences ofA/HINlv analyzed segregated into three genetically distinct clades and were characterized by the appearance of amino acid variations that were progressively fixed in the field viral population under scrutiny. CONCLUSIONS: These data suggest an early co-circulation of genetically distinct A/HNINv variants and emphasize the importance of a close molecular surveillance to detect rapidly the spread of new viral variants and to define their epidemiological impact.

[Clinic use of Laser Doppler Flowmetry (LDF) in the study of subfoveal choroidal circulation]. / Impiego clinico del Laser Doppler Flussometria (LDF) nello studio della circolazione coroideale sottofoveale.

Today our knowledge of the choroidal circulation is limited: we know its anatomy, but, on the other hand, its physiopathology remains to be fully. The choroid is involved in a number of important ocular diseases. The Laser Doppler Flowmetry (LDF) is a technique that allows non-invasive measurement of haemodynamic parameters of the subfoveal choroidal circulation. It is easy to use in daily clinic activity. The aim of this mini-review is to describe LDF studies of the choroidal circulation performed in healthy subjects under different environmental conditions, in subjects with ocular diseases, as well as studies of the effects of various drugs can induce on this circulation.

Differential long-term subcellular responses in heart and liver to adriamycin stress. Exogenous L-carnitine cardiac and hepatic protection.

In order to evaluate the heart and liver responses after adriamycin (ADR) toxic aggression, with and without exogenous L-carnitine (CAR) protection, female Sprague-Dawley rats, body weight 40-60 g, were randomized into four groups: CON, ADR, CAR and CAR-ADR. ADR was injected i.v. at a dose of 15-18 mg/kg body wt (0.1 ml). CAR was administered i.v. at a dose of 20 mg (0.1 ml) before each subdose of ADR, and then orally at 180 mg/kg body wt daily for 12 weeks. Long-term cardiac and hepatic subcellular damage were determined by transmission electron microscopic analysis of ultrathin sections. The ADR-induced long-term cardiac subcellular pathology included loss, disruption and disassembly of myofibrils, and mitochondrial swelling and condensation. On the other hand, the ADR-induced subcellular hepatic alterations consisted of polymorphic mitochondria, cytoplasmic vacuolization and accumulation of lipid droplets. Apparently, cardiac tissue was more affected by ADR toxic aggression than hepatic tissue. However, these alterations were of less severity in protected groups, in both heart and liver, suggesting CAR as a possible hepatoprotector agent against ADR toxicity. Because of the liver-L-carnitine-heart relationship, studying ADR-hepatotoxicity could be helpful in the further understanding of severe ADR-cardiotoxicity.

Valores séricos de marcadores hepáticos en ratas: aporte de un patrón de referencia / Hepatic marker seric values of hepatic markers in rat: contribution a reference pattern

La cepa Sprague Dawley representa un modelo animal usado para evaluar el potencial terapéutico y tóxico de drogas y permite la elaboración de diseños experimentales para estudiar órgano toxicidad, síendo el hígado uno de los órganos mas estudiados. Una manera indirecta de evaluar la condición del indiviuo y el órgano afectado, es la determinación sérica de metabolitos específicos, lo cual exige el establecimiento de rangos de referencia de marcadores universales para la detección de daño orgánico. Aplicando un diseño no experimental, se determinaron los valores séricos normales de y-GT, TGP, Bt, Bd y Tg, en ratas hembras Sprague Dawley, de 150-250 g de peso corporal (p.c), empleando métodos optimizados para humanos. Se extrajeron 3 cc de sangre, mediante punción directa de la arteria aorta a 40 ratas, posterior a su sacrificio con vapores de cloroformo. Las muestras fueron centrifugadas 15 min. separando el suero para la determinación de los parámetros químicos establecidos, con el uso de estuches comerciales de técnica manual. Se calculó el promedio y la desviación estándar de los valores obtenidos arrojando los siguientes rangos de valores de referencia: y-GT (16.0 UI/Lñ7.2), TGP (32.5 UI/Lñ9.5), Bt (0.36 mg/dlñ0.18), Bd (0,08 mg/dlñ0.04) y Tg (88 mg/dlñ30) y un promedio de p.c de 203 gñ21. Las similitudes y diferencias entre los valores obtenidos y los reportados por otros investigadores, sugieren que cambios ambientales, alimenticios y de cautiverio pudieran modificar los niveles de metabolitos séricos en estos animales experimentales

Discordances between the different types of atrial arrangement and the positions of the thoraco-abdominal organs.

Numerous investigations have addressed the most appropriate anatomical structures to be used for the identification of segmental arrangement ("situs"). Variations from normal have been termed "heterotaxy syndromes". The assumption is frequently made that, in such instances, all the organs in a given individual case follow the same rules for segmental morphology. This leads to confusion. The starting point for cardiac analysis is the structure of the atriums. On this basis, we can define viscero-atrial discordance when, in usual arrangement, mirror imagery, and heterotaxy, the atriums and their appendages are no longer concordant with the remaining thoracic and abdominal organs. With this in mind, we examined a large number of autopsied specimens to identify all possible variations of thoracic structures and abdominal organs relative to atrial arrangement. We analyzed 1100 malformed hearts. Of these, 1046 had usual arrangement with normally situated heart (95%); 10 cases had usual arrangement with right-sided heart (0.90%); 25 showed mirror imagery (2.27%), and 19 were found with heterotaxy (1.72%). There was a total of 54 hearts of usual arrangement with right-sided heart, mirror imagery and heterotaxy, (4.90%) and, amongst them, 17 showed viscero-atrial discordances (31.48%). In those having usual arrangement with right-sided heart, there were 2 cases (20%); 8 in those with mirror imagery (32%); and 7 in heterotaxy (36.8%). In the 1046 cases of usual arrangement with normally situated heart, there were 12 examples of viscero-atrial discordances (1.14%). In all, we found a total of 29 cases of viscero-atrial discordances. Taken together, the discordances are best dealt with by describing separately, in every case of normal or abnormal atrial arrangement, the thoracic structures, the atrial appendages, and the abdominal organs. In the final analysis, the diagnosis of the cardiac arrangement rests on the location and morphology of the atriums and their appendages.

Influenza vaccination of HIV-1-positive and HIV-1-negative former intravenous drug users.

The immunogenicity of an anti-influenza vaccine was assessed in 409 former intravenous drug user volunteers and its effect on the levels of HIV-1 RNA, proviral DNA and on CD4+ lymphocyte counts in a subset HIV-1-positive subjects was measured. HIV-1-positive individuals (n = 72) were divided into three groups on the basis of their CD4+ lymphocyte counts, while the 337 HIV-1-negative participants were allocated into group four. Haemagglutination inhibiting (HI) responses varied from 45.8 to 70% in the HIV-1-positive subjects and were significantly higher in group four (80.7% responses to the H1N1 strain, 81.6% to the H3N2 strain, and 83% to the B strain). The percentage of subjects with HI protective antibody titres (> or = 1:40) increased significantly after vaccination, especially in HIV-1 uninfected subjects. Immunization caused no significant changes in CD4+ counts and in neither plasma HIV-1 RNA nor proviral DNA levels. Therefore, vaccination against influenza may benefit persons infected by HIV-1.

Comparison of RT-PCR with other diagnostic assays for rapid detection of influenza viruses.

To compare the effectiveness of reverse transcription-polymerase chain reaction (RT-PCR), shell vial culture and cytospin assay as laboratory techniques for rapid diagnosis of influenza infections, a retrospective study was carried out on 270 aliquots of oropharyngeal swabs collected from October 1993 to March 1996 and already characterized by standard isolation procedures, and a prospective study in which 65 clinical samples taken from patients with influenza-like syndrome between October 1996 and March 1997 were tested. In the retrospective study, using conventional isolation as the gold standard, the sensitivity of RT-PCR and cytospin assay for virus A was 100% (95% confidence interval (CI), 89.1-100) and for virus B it was 100% (95% CI, 56.1-100) compared with 77.5% (95% CI, 61.1-88.6) and 71.4% (95% CI, 30.3-94.9) for shell vial culture. The specificity of all the three assays was 100% (95% CI, 98.0-100) for virus A and 100% (95% CI, 98.2-100) for virus B. In the prospective study the sensitivity of RT-PCR was greater than that of the other tests considered, both rapid and standard. It is suggested that RT-PCR should be employed in combination with conventional culture techniques in routine diagnosis of influenza infections in order to obtain results more rapidly and to improve virus detection even in circumstances in which standard isolation could be problematic.

Rapid detection of gammaT cell receptor gene rearrangements in acute lymphoblastic leukemia by electrophoresis and silver staining: implications for detection of minimal residual disease.

Minimal residual disease (MRD) in acute lymphoblastic leukemia (ALL) was studied using polymerase chain reaction (PCR). GammaT cell receptor (TCRG) genes are ideal targets for PCR-based detection of MRD due to their molecular characteristics. Polyacrylamide gel electrophoresis (PAGE) analysis of PCR products followed by silver staining was performed for 72 children with ALL at the onset of disease. Silver staining is an effective technique to detect gene rearrangements without the use of ethidium bromide. Moreover, this method may show heteroduplex bands of a clonal nature when both TCRG alleles are rearranged. PCR products subjected to a rapid staining protocol were recovered from the gel, reamplified by a second PCR and directly sequenced. After sequencing, we identified the junctional region and obtained patient-specific probes. In more than half of the patients we detected TCRG rearrangements that were used as molecular markers for residual disease.

Carnitine promotes heat shock protein synthesis in adriamycin-induced cardiomyopathy in a neonatal rat experimental model.

In order to evaluate carnitine protective strategy and its relationship with heat shock protein induction, female Sprague-Dawley neonatal rats, body weight 40 g, were randomized into four groups: control, adriamycin, carnitine and carnitine-adriamycin. Adriamycin was injected i.v. at a dose of 27 mg/kg (0.1 ml). Carnitine was administered i.v. (20 mg/0.1 ml) before each subdose of adriamycin and then per os (180 mg/kg) daily for 12 weeks. Body weight was recorded weekly. Ventricular wall thickness and cellular damage percentage were morphometrically and ultrastructurally determined, respectively. The determinations were realized monthly until the third month after treatment. The heat shock protein 25 content in the supernatant of the homogenized heart tissue was determined by Western blot analysis. Eight and 12 weeks after treatment, body weight and ventricular wall thickness decreased much more in adriamycin groups than in control and carnitine ones. At the same time, electron microscopic analysis of adriamycin left ventricular wall samples showed loss of myofibrils, swollen mitochondria and vacuoles. Carnitine-adriamycin treated rats resemble control groups more than adriamycin treated samples. Moreover, de-novo synthesis of heat shock protein was three times more induced in carnitine-adriamycin rats than in adriamycin ones. Carnitine may enhance the cell-protecting mechanism based on an induction of shock protein, and this first cellular response could reduce the severity of late adriamycin-cardiomiopathy.

Efecto de la D-Carnitina sobre la arquitectura ventricular de corazón de embrión de pollo: estudio estructural y ultraestructural / Effects of D-Carnitine in the myoarchitecture of the chick embryo heart ventricular region

Objetivo:El presente trabajo estudia, a nivel estructural y ultraestructural, el efecto de D-carnitina sobre la arquitectura ventricular de corazón de embrión de pollo de 5 días de desarrollo .Métodos: Se disecaron corazones a partir de embriones de cinco díasde desarrollo tratados con D-carnitina (6mg./Kg. de peso). Los controles fueron obtenidos a partir de embriones tratados con solución salina isotónica. Las muestras controles y tratadas fueron fijadas, post-fijadas, deshidratadas, infiltradas, incluidas y polimerizadas. Para el análisis en microscopia de luz (ML), se realizaron cortes gruesos teñidos con azul detoluidina y para microscopia electrónica de transmisión (MET)cortes finos contrastados con acetato de uranilo y citrato de plomo. Para microscopia electrónica de barrido (MEB), las muestras , fueron desecadas y cubiertas con oro para ser observadas en un microscopio de barrido (Hitachi-300) a 20 Kvolt de aceleración. Resultados:En los corazones tratados con D-carnitina disminuye:el espacio subepicárdico, el grosor del miocardio ventricular y del septum interventricular primitivo. La región ventricular carece de apariencia cónica, pierde la estrechez caudal y el ápice exhibe un patrón trabecular homogéneo. Se observa una arquitectura ventricular de apariencia esponjosa. Resalta la infiltración grasa ó presencia de gotas de lípidos en el interior de los miocitos. Conclusiones:D-carnitina cambia el patrón de estructuración normal del corazón embrionario. Se sugiere que L-carnitina actúa como factor regulador de la relación existente entre la anatomía y el metabolismo cardíaco

Early single-photon emission computed tomography in mild head trauma. A controlled study.

Although computed tomography (CT) and magnetic resonance imaging scans often appear normal after mild head trauma, many patients experience attentional or other cognitive disturbances that are difficult to quantify by neuropsychological testing in the absence of a premorbid profile. Within 2 days of mild head trauma, 14 patients with normal-appearing brain CTs were studied with single-photon emission computed tomography (SPECT). They were compared to 15 normal control subjects and to 12 patients with mild human immunodeficiency virus (HIV) encephalopathy. Ten of 14 head trauma patients were separated from the normal control subjects by both independent readers, blinded to the clinical diagnosis. None of the SPECT results from normal control subjects were "read" as trauma. Trauma could not be differentiated from HIV encephalopathy. The observed percentage agreement between raters was 0.83 (kappa = 0.72). SPECT is more sensitive than CT in detecting brain injury after mild head trauma.

In vitro evaluation of a 4-leaf coil design for magnetic stimulation of peripheral nerve.

The performance of a 4-leaf magnetic coil was evaluated during magnetic stimulation of a peripheral nerve in vitro. The site of stimulation was below the coil center, and a 90 degrees rotation of the coil was equivalent to a change in current polarity. A hyperpolarizing magnetic stimulus failed to slow or block a propagating action potential.

Mesenchymal tissue of the interventricular septum. Structural and ultrastructural study.

Light and electron microscopic studies of frontal and sagittal sections of embryonic chick hearts (Stages 25, 28-29), reveal mesenchymal tissue in the cephalic portion of the interventricular septum. The endocardium of this cephalic portion contains reoriented and invaginated cells with pseudopodia; in addition there are cells immediately subjacent to the endocardium. Similar cellular events take place during the formation of mesenchymal tissue in the atrioventricular and conotruncal regions. In these regions the mesenchymal tissue originates by means of an endocardial activation process. The structural characteristics of the formation of the cephalic portion of the interventricular septum suggest that local mesenchymal tissue is contributed by the endocardium. However, based upon the close anatomic relationship observed by us between the mesenchymal tissues of the atrioventricular canal, conotruncal region and the cephalic portion of the interventricular septum; we do not discard a contribution by migration of cells from atrioventricular and conotruncal regions to the interventricular septum.

Ultrastructural study of the myocardial wall of the atrio-ventricular canal during the development of the embryonic chick heart.

In the early stages of chick embryo development (days 3-5) the myocardium of the atrio-ventricular canal (AV) is continuous with the atrial and ventricular muscles; however, interruption of muscular continuity is observed at later stages (from day 6 to day 8). The most relevant event occurring at the AV canal region is the dissociation of the myocytes due to the loss of their cellular attachments, rather than an invasion of connective tissue cells (endocardial and sub-epicardial) located on both sides of the myocardium. In this study, particular attention was paid to the sequential changes that take place in the myocardium of this region, these being (1) a reduction in the number of desmosomes and intercalated discs with the subsequent appearance of large, inter-cellular spaces between myocytes; (2) migration of these cells through a complex extra-cellular matrix, to which it appears to be closely related, suggesting that the macromolecules of this matrix may be being synthesized by the myocytes, and may take part in the process of cardiac cell separation; (3) incorporation of the myocytes in the developing tricuspid valve, where they co-exist with fibroblasts. The results of the study correspond remarkably well to those previously carried out on the left AV canal myocardium, suggesting that the behaviour of the muscle is the same, at all points around the AV canal.