Dynamics of sperm DNA fragmentation in the swine: ejaculate and temperature effects.

The dynamics of sperm DNA fragmentation was examined in 16 boar ejaculates using the sperm chromatin dispersion (SCD) test and a two-tail comet assay. The net sperm rich fraction was preserved at two different temperatures (Trial 1: 15 degrees C, n=10; Trial 2: 37 degrees C, n=6) and sub-samples were taken every day until a sperm motility of zero. Significant differences in the dynamics of DNA fragmentation were observed among the different ejaculates and also according to the storage temperature. After analyzing the dynamic response of the sperm DNA damage, when the sperm samples are incubated at 15 or 37 degrees C, each ejaculate could be classified and a considerable variation among individuals for an increase in DNA damage was observed. Thus, while in some ejaculates no rise in DNA fragmentation was observed, in others, the sperm DNA fragmentation process was triggered during the initial days of the experiment. In general, sperm incubation at 37 degrees C diminished sperm DNA quality. The two-tail comet assay indicated that at time zero existing DNA damage mainly consisted of double stranded DNA breakage. During storage, DNA damage affected one of the DNA strands until a second wave of DNA damage, in which there was both single and double stranded DNA damage.

Directional mapping of DNA nicking in ejaculated and cauda epididymidal spermatozoa of the short-beaked echidna (Tachyglossus aculeatus: Monotremata).

Prototherian spermatozoa are unique amongst the Mammalia in terms of their filiform morphology, tandem arrangement of chromosomes and formation of sperm bundles. In the present study, we provide observations of echidna spermatozoa and note that the superstructure of the bundle is engineered around the shape of the individual sperm head and that this in turn may be a consequence of the unusual circumferential and helicoidal condensation of the DNA during spermiogenesis. Frozen-thawed ejaculated echidna spermatozoa were incubated and examined for the presence of non-typical DNA conformation by means of in situ labelling of DNA breaks using Klenow polymerase and via alkaline single-cell comet assays for detection of fragmented DNA. Both techniques successfully revealed the presence of what appeared to be directional DNA nicking, co-localised with the presence of highly sensitive alkali sites along the length of the sperm nucleus. It was not possible to define whether these alternative DNA configurations were associated with a failure of the sperm nucleus to condense appropriately during spermiogenesis or were evidence of DNA fragmentation following post-thaw incubation or a sequential structural chromatin rearrangement necessary for fertilisation.

Assessing sperm DNA fragmentation in the field: an adaptation of sperm chromatin dispersion technology.

The sperm chromatin dispersion (SCD) test is a new technique that allows assessment of sperm DNA fragmentation (SDF) in different species. The application of this technique, like other techniques, is restricted to the laboratory. Our investigation was aimed at exploring the possibilities of extending SCD methodology for use in the field, where electric powered facilities such as freezers, microscopes or heaters are not available. Our results showed that SCD methodology, with minor modifications to the standard protocol, can be performed readily in the field, offering reliable information about SDF. An Light Emitting Diode (LED)-equipped microscope attached to a laptop, a gas heater and a CO(2) spray for cooling are sufficient to assess the quality of sperm DNA. The results obtained after assessing 10 different semen samples under different conditions (30 degrees C in the laboratory and at 17 degrees C and 4 degrees C in the field) showed that except after processing the slides at 4 degrees C, the results of SDF in different animals showed no significant differences. With the modifications suggested here, the SCD technique can be used to assess SDF in the wild. In particular, the DNA quality of spermatozoa obtained from animals post mortem can be assessed in the field.

Dynamics of sperm DNA fragmentation in domestic animals III. Ram.

From a biological viewpoint spermatozoa are ejaculated by the male and received into the female while maintaining roughly constant temperature, which in most mammals is below the temperature of the soma. When ejaculated spermatozoa are used for artificial reproductive purposes a temperature excursion episode is produced, because the spermatozoa are often stored as frozen or chilled samples and the biological temperature is only recovered after insemination. In this study we have analyzed the effects of cooling (to 15 degrees C) and freezing ram spermatozoa on the subsequent sperm DNA fragmentation index (sDFI) during a varying period of storage at 37 degrees C. The aim was to emulate in vivo processes that cooled or frozen-thawed spermatozoa experience after insemination. The study was performed using commercial semen samples derived from rams regularly used for reproductive purposes. Semen samples were studied after a cooling or cryopreservation episode followed by biological temperature recovery and incubation up to 48 h. The results indicated that when spermatozoa experience a severe (frozen) or mild (cooled) temperature excursion episode, major effects on sperm viability and DNA fragmentation are induced and cause the subsequent rapid decline of ram sperm quality. This effect could be detected just at the onset of the biological temperature recovery. Sperm DNA damage in cooled samples was observed after 5 h of incubation at 37 degrees C, while this time was reduced to less than 60 min in frozen-thaw samples. The dynamics of sDFI in different animals, analyzed under the same experimental conditions, was different from one sample to another, regardless of the method used for storage. Sperm viability was better preserved in cooled rather than in frozen samples. While for the frozen-thawed samples sperm viability was almost abolished after 5 h of incubation, a stable proportion of viable spermatozoa (ranging from 20% to 60%) was observed in the cooled samples at the corresponding time points. Finally, with respect to the prevalence of sDFI in ram, the level commonly found was lower than 5% at the onset of the experiment. However, sDFI was higher than 5% in 25% of the samples and in 15% of rams this index exceeded 10%.

Sperm DNA fragmentation in a random sample of the Spanish boar livestock.

A collection of 180 chilled boar semen samples, randomly chosen from stocks currently used for routine characterization of standard seminal quality, were studied for DNA fragmentation status using the sperm chromatin dispersion test and the DNA fragmentation index (DFI: percent of abnormal cell versus normal cells for DNA fragmentation) was determined. Values for sperm motility, acrosome status, coiled tails and abnormal head morphology, including presence and position of cytoplasmic droplets were also obtained. The DFI in the whole sample presented a wide range of variation with values oscillating between practically 0% and 47.95% and do not fit to a normal distribution. The most frequent classes (83.3%) presented a DFI lower than a 5%. Significant correlations between sperm DNA fragmentation and sperm motility, acrosome status, frequency of distal droplets, coiled tails and abnormal head morphology, were not observed. However, the presence of proximal cytoplasmic droplets showed a significant correlation with the level of DNA fragmentation observed in the ejaculated spermatozoa.

Efecto de la congelación sobre la fragmentación del ADN espermático en semen de caballo / No disponible

No disponible

[Perioperative transfusion of erythrocyte concentrates during elective surgery: introduction of a protocol for indications]. / Transfusión perioperatoria de concentrado de hematíes en cirugía programada: introducción de un protocolo de indicaciones.

PURPOSE: The aim of this paper is, first, to know the actual situation of the perioperatory red cell transfusion for elective surgery in our hospital. In a second phase and prospectively, we tested guidelines for red cell perioperatory transfusion in order to observe the change of transfusions. Then, we compared the results between the basal and postintervention periods. PATIENTS AND METHODS: We performed an aleatory assay with two periods, basal and interventionist. Basal period: 151 patients undergoing elective surgery with perioperatory blood requested and general anesthesia. Intervention period: We applied a transfusion guidelines protocol for perioperatory red cell transfusion from the Hospital's Transfusion Committee, also a questionnaire to evaluate the medical indication; We studied 164 patients with clinical features like the basal period. Study/results variables: preoperative blood request, perioperatively transfusion, number of packed red-cell units transfused, crossmatch--to--transfusion ratio, haemoglobin level pre and posttransfusion. RESULTS: No significant drop of the cross match-transfusion ratio was observed after intervention. There is a slight reduction of the crossmatch--to--transfusion ratio, although these value is high (4.48), due to an increase of the transfusion keeping the percentage of appropriate transfusions. The most frequent reason (53%) of inadequate transfusion is the active bleeding. CONCLUSIONS: 1) The transfusional activity of the Marina Alta Hospital supposes approximately 17% of the request and 6% of the global transfusion. 2) The introduction of a protocol of perioperative transfusion instructions suppose a small decrease of the crossmatch--to--transfusion ratio, without statistical significance. This slight reduction is due to an increase of transfusion in the post-intervention period, since in this period there is a group of older age patients and with greater percentage of associated pathology. 3) The rate of appropriate transfusions in both periods is similar. 4) The preoperative request of red cells is inappropriate. 5) The most frequent reason of inappropriate transfusion is active bleeding.

[Anatomical correction in transposition of the great arteries and double-outlet right ventricle. Our initial experience]. / Corrección anatómica en la transposición de grandes arterias y en la doble salida de ventrículo derecho. Experiencia inicial.

INTRODUCTION AND OBJECTIVES: Anatomical correction of transposition of great arteries and double outlet right ventricle with subpulmonary ventricular septal defect is a surgical approach that has not been generally adopted in our clinical environment. Our aim with this paper is reporting on our initial experience with this technique. METHODS: The clinical data and additional investigations are reviewed from 15 infants with transposition of the great arteries and 2 with double outlet right ventricle who underwent anatomical repair, with a postoperative follow-up of one year. RESULTS: The survival rate has been 76% (13 out of 17 cases). The 13 survivors are in good hemodynamic condition, with no treatment whatsoever. Thirteen patients are in sinus rhythm with normal repolarization patterns and 5 patients show a right bundle branch block. Neither aortic nor pulmonary gradients have been detected on Doppler examination, and slight valvular insufficiencies are found at aortic level in 4 patients, pulmonary in 2 and mitral in one. CONCLUSION: Anatomical correction is the method of choice for transposition of the great arteries and double outlet right ventricle with pulmonary-related ventricular septal defect.

[Visceral leishmaniasis in Alicante. Analysis of 21 cases]. / Leishmaniasis visceral en Alicante. Análisis de 21 casos.

We present 21 cases of visceral Leishmaniasis diagnosed in our hospital during the past 8 years. The diagnostic method used was the visualization of the parasite in bone marrow aspiratory puncture. All cases presented fever at admission, hepatosplenomegaly, anemia elevated sedimentation rate and polyclonal gammapathy. Two of our patients were diagnosed of AIDS during the course of the disease. Cure was observed in all cases after one cycle treatment with pentavalent antimonials except for the two AIDS cases one of whom died due to cerebral toxoplasmosis. We point out visceral Leishmaniasis as an opportunistic infection in patients with AIDS and its resistance to the usual treatment.

[Diabetes mellitus and changes in hemostasis: study of the fibrinolytic response of insulin-dependent patients without vascular disease]. / Diabetes mellitus y alteraciones de la hemostasia: estudio de la respuesta fibrinolítica en pacientes insulinodependientes sin vasculopatía.

Diabetes mellitus (DM) is characterized by a hypercoagulability state and many of these disorders are corrected with adequate metabolic control. The goal of this study was to assess diverse hemostatic and fibrinolytic parameters in 12 insulin-dependent (IDDM) metabolically controlled patients without vascular lesions and in a group of 12 healthy volunteers. A significant difference was observed in the euglobulin lysis time (ELT), after a stress test, since only 3 patients had an adequate fibrinolytic response. These results suggest that the fibrinolytic alterations found in DM are not secondary to metabolic disorders caused by the disease and we can consider that the existence of subclinical alterations of the vascular endothelium would be responsible for these alterations.