Evaluation of equine semen frozen in extenders free of egg yolk using two different freezing curves.

A chemically defined cryopreservation extender that maintains seminal parameters is relevant. Fifteen ejaculates from 5 stallions (n= 5; r=3) were diluted in 5 extenders: 1) EDTA-glucose based extender with egg-yolk and dimethylformamide (EY); 2) commercial equine extender (CE); 3) CE with dimethylformamide (CE-3); 4) bovine commercial extender with liposomes (OP); 5) bovine commercial extender with soybean lecithin (BIO), and frozen using a slow and a rapid temperature descent curve. Post-thaw evaluations were: sperm kinematic parameters, viability and acrosome status, membrane lipoperoxidation and DNA fragmentation. Sperm data were analysed using an ANOVA or Friedman test (results mean ± SD). Paired comparison between the two freezing curves was analysed using the Wilcoxon test. Total and progressive motility were significantly higher (P<0.05) in the EY and CE-3 samples using the slow curve, whereas for the fast curve, total and progressive motility were significantly higher (P<0.05) in the EY samples compared to all the extenders and the samples frozen in CE-3 were significantly higher than the remaining extenders (P<0.05). The percentages of live acrosome intact sperm and of live non-peroxidized sperm were significantly higher (P<0.05) in the EY extender when using either of the freezing curves and in turn, were significantly higher (P<0.05) in samples frozen in CE-3 compared to the remaining extenders. Intact DNA was significantly lower (P<0.05) in the BIO extender, using the rapid curve. To conclude, the commercial equine extender with 3% dimethylformamide, without egg-yolk, could be a suitable alternative for extenders with egg-yolk.

Comparison of different cryoprotectants for freezing donkey (Equus asinus) semen.

The aim of this study was to evaluate two cryoprotectants, dimethylformamide (DMF) and methylformamide (MF) in two concentrations (5 and 7%) in vitro in donkey semen using a rapid freezing technique and the effect on pregnancy rates in mares. Twenty-four ejaculates from 8 jacks (n=8; r=3) were divided into 4 extenders: BotuSemen Gold with 5% or 7% MF and 5% or 7% DMF, all containing 11% lactose, 20% egg-yolk and 0.5% Equex. Post-thaw evaluations included: sperm motility, membrane function and acrosome status. A linear mixed effect model was used to test the effect of different freezing media on semen parameters. No differences were observed between the 4 freezing media used, for any of the seminal parameters (P>0.05). However, samples with 5% DMF showed the highest percentages of sperm with acrosomes and functional membranes (DMF: 5%: 53.67±22.01; 7%: 33.92±23.4; MF: 5%: 44.5±20.46; 7%: 38.75±27.4) (Data: mean ± SD; P>0.05). Hence, thirty mares were inseminated: 15 with 5% DMF and 15 with 7% DMF. The pregnancy rate was 46% (7/15) and 0% (0/15) using the extender with 5% or 7% DMF, respectively (P=0.003). To conclude, the use of 5% or 7% of MF or DMF did not affect the in vitro parameters. Despite the lack of differences in vitro with the two DMF concentrations, in vivo results only showed pregnancies when using 5% DMF. Thus, the results of this study demonstrate the importance of accompanying in vitro semen evaluations with studies that evaluate post-insemination pregnancy rates.

Population Data Centre Profiles: Centre for Data Linkage.

The Centre for Data Linkage (CDL) was established at Curtin University, Western Australia, to develop infrastructure to enable cross-jurisdictional record linkage in Australia. The CDL's operating model makes use of the 'separation principle', with content data typically provided to researchers directly by the data custodian; jurisdictional linkage where available are used within the linkage process. Along with conducting record linkage, the team has also invested in establishing a research programme in record linkage methodology and in developing modern record linkage software which can handle the size and complexity of today's workloads. The Centre has been instrumental in the development of practical methods for privacy-preserving record linkage, with this methodology now regularly used for real-world linkages. While the promise of a nation-wide linkage system in Australia has yet to be met, distributed models provide a potential solution.

Privacy preserving linkage using multiple match-keys.

INTRODUCTION: Available and practical methods for privacy preserving linkage have shortcomings: methods utilising anonymous linkage codes provide limited accuracy while methods based on Bloom filters have proven vulnerable to frequency-based attacks. OBJECTIVES: In this paper, we present and evaluate a novel protocol that aims to meld both the accuracy of the Bloom filter method with the privacy achievable through the anonymous linkage code methodology. METHODS: The protocol involves creating multiple match-keys for each record, with the composition of each match-key depending on attributes of the underlying datasets being compared. The protocol was evaluated through de-duplication of four administrative datasets and two synthetic datasets; the 'answers' outlining which records belonged to the same individual were known for each dataset. The results were compared against results achieved with un-encoded linkage and other privacy preserving techniques on the same datasets. RESULTS: The multiple match-key protocol presented here achieved high quality across all datasets, performing better than record-level Bloom filters and the SLK, but worse than field-level Bloom filters. CONCLUSION: The presented method provides high linkage quality while avoiding the frequency based attacks that have been demonstrated against the Bloom filter approach. The method appears promising for real world use.

Evaluation of approximate comparison methods on Bloom filters for probabilistic linkage.

INTRODUCTION: The need for increased privacy protection in data linkage has driven the development of privacy-preserving record linkage (PPRL) techniques. A popular technique using Bloom filters with cryptographic analyses, modifications, and hashing variations to optimise privacy has been the focus of much research in this area. With few applications of Bloom filters within a probabilistic framework, there is limited information on whether approximate matches between Bloom filtered fields can improve linkage quality. OBJECTIVES: In this study, we evaluate the effectiveness of three approximate comparison methods for Bloom filters within the context of the Fellegi-Sunter model of recording linkage: Sørensen-Dice coefficient, Jaccard similarity and Hamming distance. METHODS: Using synthetic datasets with introduced errors to simulate datasets with a range of data quality and a large real-world administrative health dataset, the research estimated partial weight curves for converting similarity scores (for each approximate comparison method) to partial weights at both field and dataset level. Deduplication linkages were run on each dataset using these partial weight curves. This was to compare the resulting quality of the approximate comparison techniques with linkages using simple cut-off similarity values and only exact matching. RESULTS: Linkages using approximate comparisons produced significantly better quality results than those using exact comparisons only. Field level partial weight curves for a specific dataset produced the best quality results. The Sørensen-Dice coefficient and Jaccard similarity produced the most consistent results across a spectrum of synthetic and real-world datasets. CONCLUSION: The use of Bloom filter similarity comparisons for probabilistic record linkage can produce linkage quality results which are comparable to Jaro-Winkler string similarities with unencrypted linkages. Probabilistic linkages using Bloom filters benefit significantly from the use of similarity comparisons, with partial weight curves producing the best results, even when not optimised for that particular dataset.

Traumatismos dentoalveolares: reporte de casos clínicos / Dentoalveolar trauma: report of clinical cases

Los traumatismos dentoalveolares que afectan a escolares pueden involucrar a los tejidos blandos y/o duros de las piezas primarias y permanentes jóvenes. Las causas por las que se producen son múltiples e incluyen caídas, accidentes en el hogar, actividad deportiva en el colegio o el club, accidentes de tránsito. El objetivo de este trabajo es presentar la resolución y el seguimiento a dos años de dos situaciones clínicas de traumatismos dentoalveolares en pacientes escolares que concurrieron a la Universidad de Buenos Aires, Facultad de Odontología, Odontología Integral Niños (AU)

Dentoalveolar trauma in schoolchildren can involve soft and/or hard tissues of deciduous and young permanent teeth. It has multiple causes, including falls, home accidents, sports accidents at school or at a club, and car accidents. The aim of the present work was to show treatmentand two-year clinical follow up of dentoalveolar trauma in schoolchildren seen at the Universidad de Buenos Aires, Facultad de Odontología, Odontología Integral Niños (AU)

Histopathology of the gut in rheumatic diseases.

The gastrointestinal tract regulates the trafficking of macromolecules between the environment and the host through an epithelial barrier mechanism and is an important part of the immune system controlling the equilibrium between tolerance and immunity to non-self-antigens. Various evidence indicates that intestinal inflammation occurs in patients with rheumatic diseases. In many rheumatic diseases intestinal inflammation appears to be linked to dysbiosis and possibly represents the common denominator in the pathogenesis of different rheumatic diseases. The continuative interaction between dysbiosis and the intestinal immune system may lead to the aberrant activation of immune cells that can re-circulate from the gut to the sites of extraintestinal inflammation as observed in patients with ankylosing spondylitis. The exact contribution of genetic factors in the development of intestinal inflammation in rheumatic diseases needs to be clarified.

Pathogenesis of polymyalgia rheumatica.

Polymyalgia rheumatica (PMR) is a chronic, inflammatory disorder of unknown cause, almost exclusively occurring in people aged over 50 and often associated with giant cell arteritis. The evidence that PMR occurs almost exclusively in individuals aged over 50 may indicate that age-related immune alterations in genetically predisposed subjects contribute to development of the disease. Several infectious agents have been investigated as possible triggers of PMR even though the results are inconclusive. Activation of the innate and adaptive immune systems has been proved in PMR patients as demonstrated by the activation of dendritic cells and monocytes/macrophages and the altered balance between Th17 and Treg cells. Disturbed B cell distribution and function have been also demonstrated in PMR patients suggesting a pathogenesis more complex than previously imagined. In this review we will discuss the recent findings regarding the pathogenesis of PMR.

Interleukin (IL)-9/IL-9R axis drives γδ T cells activation in psoriatic arthritis patients.

Cytokines such as tumour necrosis factor (TNF)-α, interleukin (IL)-12, interferon (IFN)-γ, IL-23 and, more recently, IL-9, have been implicated in the initiation/maintenance of inflammation in psoriasis and psoriatic arthritis (PsA). In the present study we aimed to characterize the role of γδ T cells in peripheral blood and synovial fluid of PsA patients and to investigate their response to in-vitro stimulation with antigen or cytokines (IL-9 and IL-23). γδ T cells isolated from peripheral blood mononuclear cells and synovial fluid were analysed by flow cytometry to evaluate the phenotype and cytokine production. IL-23R and IL-9R gene expression were also evaluated by reverse transcription-polymerase chain reaction (RT-PCR). Peripheral blood mononuclear cells (PBMC), sorted γδ T cells and γδ cell lines were also stimulated in vitro with isopentenyl pyrophosphate (IPP), recombinant IL-9 or recombinant IL-23. Our results show an expansion of γδ T cells with a predominant effector memory phenotype in peripheral blood and synovium of untreated PsA patients, which reverses significantly after treatment with anti-TNF-α or anti-IL-12/IL-23R monoclonal antibodies (mAbs). Moreover, in PsA patients γδ T cells activation is driven prevalently by IL-9/IL-9R interaction, and not only by IL-23/IL-23R. Together these findings indicate γδ T cells and IL-9 as new players in the pathogenesis of PsA.

Interleukin-9 and T helper type 9 cells in rheumatic diseases.

Interleukin (IL)-9 is a 28-30 kDa monomeric glycosylated polypeptide belonging to the IL-7/IL-9 family of proteins that bind to a composite receptor consisting of the private receptor IL-9R and the IL-2 receptor, gamma (IL-2RG), a common gamma subunit shared by the receptors of many different cytokines. The IL-9R is expressed widely and IL-9 impacts a number of effector cells, such as effector T cells, B cells, innate lymphoid cells, mast cells, polymorphonuclear cells, epithelial cells and smooth muscle cells, playing an important role in regulating inflammatory immunity. The critical role of IL-9 in promoting cellular and humoral immune responses makes it an important focus of potential therapeutic interventions. Recently, a defined subset of T helper type cells, Th9 cells, has been identified by the potent production of IL-9. The involvement of the Th9 cell subset has been described in many types of inflammatory diseases, namely atopic diseases, helminth infections, experimental autoimmune encephalomyelitis and ulcerative colitis. In this review, we summarize the IL-9 biological activities, highlighting roles for IL-9 and Th9 cells in rheumatoid and psoriatic arthritis, systemic vasculitis, systemic lupus erythematosus and systemic sclerosis.

Tratamiento de luxación discal sin reducción: responsabilidad del odontopediatra / Treatment of disk displacement without reduction: the pediatric dentist´s responsibility

Actualmente, es frecuente la consulta por trastornos temporomandibulares (TTM) en la clínica odontopediátrica, siendo reconocida la múltiple causalidad de los mismos. Se presenta la resolución de una situación clínica de una paciente de 13 años de edad que acudió a la Cátedra de Odontología Integral Niños (OIN), presentando dolor muscular y articular del lado derecho, limitando la apertura bucal a 25 mm confortable y a 28 mm forzada, con desvío de la mandíbula hacia la izquierda. Refería haber sido atendida en una guardia médica el día anterior con bloqueo en apertura, luego de 8 meses de evolución, con síntomas de chasquido y doloir, sin haber realizado consulta alguna. Se indicó tratamiento sintomático y se solicitaron estudios complementarios. El diagnóstico fue de luxación discal sin reducción. Con la evaluación integral, se hallaron factores concomitantes, tales como maloclusión, hiperlaxitud, respiración bucal y parafunciones. En el abordaje terapéutico, se colocó un intermediario oclusal para reposicionamiento mandibular, se realizó tratamiento fonoaudiológico miofuncional y reeducación postural global (RPG). La paciente evolucionó favorablemente; a los 6 meses resolvió el ruido articular y al año estaba asintomática, con 37 mm de apertura, comenzando su tratamiento de ortodoncia y manteniendo controles durante 4 años. El compromiso de la familia y la participación de un equipo de trabajo, permitió la resolución integral del caso. El odontopediatra tiene la responsabilidad de alertar a padres y pacientes sobre factores de riesgo, diagnosticar TTM y orientar los tratamientos, involucrándose cuando corresponde su intervención, o derivando en forma oportuna...

Stimulation of adenosine A2A receptors reduces intracellular cholesterol accumulation and rescues mitochondrial abnormalities in human neural cell models of Niemann-Pick C1.

Niemann Pick C 1 (NPC1) disease is an incurable, devastating lysosomal-lipid storage disorder characterized by hepatosplenomegaly, progressive neurological impairment and early death. Current treatments are very limited and the research of new therapeutic targets is thus mandatory. We recently showed that the stimulation of adenosine A2A receptors (A2ARs) rescues the abnormal phenotype of fibroblasts from NPC1 patients suggesting that A2AR agonists could represent a therapeutic option for this disease. However, since all NPC1 patients develop severe neurological symptoms which can be ascribed to the complex pathology occurring in both neurons and oligodendrocytes, in the present paper we tested the effects of the A2AR agonist CGS21680 in human neuronal and oligodendroglial NPC1 cell lines (i.e. neuroblastoma SH-SY5Y and oligodendroglial MO3.13 transiently transfected with NPC1 small interfering RNA). The down-regulation of the NPC1 protein effectively resulted in intracellular cholesterol accumulation and altered mitochondrial membrane potential. Both effects were significantly attenuated by CGS21680 (500 nM). The protective effects of CGS were prevented by the selective A2AR antagonist ZM241385 (500 nM). The involvement of calcium modulation was demonstrated by the ability of Bapta-AM (5-7 µM) in reverting the effect of CGS. The A2A-dependent activity was prevented by the PKA-inhibitor KT5720, thus showing the involvement of the cAMP/PKA signaling. These findings provide a clear in vitro proof of concept that A2AR agonists are promising potential drugs for NPC disease.

[Finger or thumb sucking. New interpretations and therapeutic implications]. / Il problema del succhiamento del dito. Nuove interpretazioni e implicazioni terapeutiche.

AIM: Finger sucking is frequently found in children and sometimes in adults too. Often reasons are found to explain why certain children feel the need to suck their thumbs or fingers, but these explanations are always derived from clinical observation without any rational support, searching for motives for this bad habit from family life; strict parents, jealousy at the birth of a sibling, difficulty accepting the end of breastfeeding etc. Some think that a child should continue to suck after actual nutrition has ended and the substitution of the mother's nipple with a thumb or finger could fill this need. Research however, using anatomical and neurophysiological data, explains why an infant forms the habit of thumb sucking from a neurological standpoint abandoning the psychological explanations proposed until now. METHODS: Forty thumb-sucking subjects with ages ranging from 5 to 25 years and a control group of 20 non-thumb-sucking subjects with correct swallowing patterns were analyzed evaluating postural and neurological effects of thumb sucking. Instruments used for this evaluation were a device for electromyography to measure the muscular tone and a baropodometer, stabilometer and a scoliosometer to analyse posture. RESULTS: Analyzed subjects had an evident improvement at the baropodometer and stabilometer and a scoliosometer examination to analyze posture and at the electromyography analysis. CONCLUSION: The results obtained confirm that a thumb-sucking subject puts the thumb in the mouth to stimulate the nasal-palatal receptors of trigeminus and obtain muscular balance and a release of physical and psychological tension.

Arterial stiffness indexes and serum cytokine levels in seronegative spondyloarthritis: relationships between stiffness markers and metabolic and immunoinflammatory variables.

OBJECTIVE: The aim of this study was to investigate the relationship between immunoinflammatory markers and indexes of arterial stiffness in patients with seronegative spondyloarthritis (SpA). METHOD: We enrolled consecutive patients with inflammatory seronegative SpA referred to a rheumatology outpatient clinic. Control subjects were patients admitted in the same period for any cause other than chronic inflammatory disease or acute cardiovascular and cerebrovascular events. Carotid-femoral pulse wave velocity (PWV) was measured and the aortic pressure waveform was used to calculate the augmentation index (Aix). We also evaluated plasma levels of C-reactive protein (CRP), interleukin (IL)-1ß, tumour necrosis factor (TNF)-α, and interleukin (IL)-6 as markers of immunoinflammatory activation. RESULTS: This study enrolled 53 patients with SpA and 55 control subjects. After adjustment for blood glucose, cholesterol, and triglyceride levels, and systolic (SBP) and diastolic blood pressure (DBP), patients with seronegative SpA showed higher mean PWV and Aix compared to controls. Moreover, in patients with seronegative SpA, we observed higher mean plasma levels of IL-6, IL-1ß, and TNF-α in subjects with mean PWV > 8 m/s in comparison with those with PWV < 8 m/s. Multivariate analysis revealed a significant association between PWV > 8 m/s and male gender, age, diabetes, hypertension, low density lipoprotein cholesterol (LDL-C) > 120 mg/dL, total cholesterol (TC) > 200 mg/dL, coronary artery disease (CAD), microalbuminuria, carotid plaque, and plasma levels of IL-6, IL-1ß, and TNF-α. CONCLUSIONS: These findings emphasize the role of inflammatory variables and metabolic factors in indexes of high arterial stiffness. Thus, an inflammatory-metabolic background may influence the pathogenesis of increased arterial stiffness in seronegative inflammatory arthritis.

Cervical paragangliomas: single centre experience with 44 cases.

Paragangliomas (PGL) are rare lesions of the neuroendocrine system; in the neck, they usually affect the carotid glomus (carotid body tumours-CBT). This retrospective analysis reports our experience in management of these lesions in patients treated by surgical resection. Between 2000 and 2014, 33 patients were surgically treated at our institution, obtaining a series of 44 cervical PGLs. Tumour characteristics, family history, diagnostic procedures, surgical treatment, short- and long-term outcomes were reviewed. A female prevalence was found (76% of cases). Familial cases occurred in 9 patients (20%); 6 presented with bilateral lesions and 1 had multiple paragangliomas. Lymph node metastasis was not found in any patient. All lesions were classified into three groups according to the latero-lateral diameter. Complete resection of the PGL was performed in all patients. Mortality was null; transitory cranial nerve deficit occurred in 20% of cases with permanent palsy in 6.7%. No perioperative stroke/TIA were observed. Surgical resection of PGL should be considered as the only therapeutic option because it can ensure complete removal of the disease. Patients with bilateral lesions and positive family history should be referred for genetic analysis. Preoperative planning of the surgical procedure by integrated diagnostic imaging and a full mastery of vascular surgery techniques are mandatory to minimise the risk of the most common postoperative complications. Long-term follow-up is recommended, particularly in patients with familial disease or sporadic lesions treated in an advanced stage.

Effect of cryoprotectant and equilibration temperature on cryopreservation of Lama glama spermatozoa.

The aim of this study was to determine the effect of two equilibration temperatures (5 °C and room temperature) and two cryoprotectants (glycerol and dimethylformamide, both at 7%) on llama sperm cryopreservation. Llama ejaculates were divided into four aliquots. A lactose-EDTA-egg yolk (LEEY) extender with either 7% glycerol (LEEY-G) or 7% dimethylformamide (LEEY-DMF) was added to two of the aliquots, which were equilibrated for 20 min at room temperature and subsequently frozen. The other two aliquots were extended in LEEY, cooled to 5 °C, then LEEY-G or LEEY-DMF was added, equilibrated for 20 min at 5 °C and frozen. No significant differences (P > 0.05) were observed in membrane function and chromatin condensation between any of the freeze-thawing protocols. Post-thaw motility was greater (P < 0.05) in LEEY-DMF than LEEY-G. DNA fragmentation was not different between raw and frozen semen with LEEY-DMF but was high in all samples with glycerol. Our results indicate that 7% glycerol would be detrimental for llama spermatozoa, but further studies are needed to evaluate effectiveness if used at lower concentrations. Dimethylformamide preserved motility and DNA integrity of frozen-thawed llama spermatozoa and could be used to replace glycerol at the concentrations used in this study.

The in vitro addition of methotrexate and/or methylprednisolone determines peripheral reduction in Th17 and expansion of conventional Treg and of IL-10 producing Th17 lymphocytes in patients with early rheumatoid arthritis.

The aim of our study was to evaluate methotrexate (MTX) and methylprednisolone (MP) effect on peripheral Th17 and Treg subsets in patients with rheumatoid arthritis (RA). We enrolled 15 patients (10 early RA and 5 long-standing disease) with active RA and 10 age-matched healthy donors as controls. Frequencies of Th17 and Treg were quantified using flow cytometry before and after in vitro addition of MTX, MP or both drugs. Our results showed a reduction in the overall Th17 population followed by an increase in Th17 IL-10(+) and Treg, after in vitro treatment of PBMCs with the drugs in patients with early RA. Long-standing disease patients showed a less evident increase in Treg cells and less enhancement of IL-10 Th17 cells. We suggest that the treatment with MTX and MP could ameliorate RA disease activity by normalizing the distribution/imbalance of Th17/Treg and indicate a new regulatory role of IL-17(+) cells in RA patients.

In vitro equine embryo production using air-dried spermatozoa, with different activation protocols and culture systems.

The aim of this work was to evaluate the use of air-dried spermatozoa for in vitro production of equine embryos and verify if sperm extract activation and in vivo culture improve in vitro embryo production. Cooled spermatozoa (control) and air-dried spermatozoa stored for 2, 14 or 28 days were used for ICSI sperm extract, or ionomycin was used for oocyte activation, and embryos were in vitro or in vivo (in mare's oviduct) cultured for 7 days. With in vitro culture, cleavage rate was higher when activating with sperm extract (P < 0.05). No differences in embryo development were seen between the two activation treatments nor between storage periods (P > 0.05). Blastocysts were obtained with cooled spermatozoa, and morulae were achieved using in vivo culture with 28-day storage spermatozoa and ionomycin-activated oocytes. When in vivo culture was performed, sperm DNA fragmentation was assessed using the sperm chromatin dispersion test and did not show statistical correlation with cleavage nor embryo recovery rates. In conclusion, equine embryos can be produced using air-dried spermatozoa stored for several weeks. Sperm extract activation increased cleavage rates but did not improve embryo development. In vivo culture allowed intrauterine stage embryos to be achieved.