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In Quebec, three pneumococcal conjugate vaccines (PCV) were used sequentially starting in December 2004. The objective of the study was to investigate the association between exposure to different PCV regimens and hospitalizations for lower respiratory tract infection (LRTI).Records with a main diagnosis of LRTI in children born in 2000-2012 and observed up to their second birthday were extracted from the provincial hospital administrative database. Main vaccine regimen in different birth cohorts was derived from the Quebec City Immunization Registry. Hospital admission risk was analyzed by Poisson regression models adjusting for age, season of birth, ambient air temperature, circulation of respiratory viruses, and the weekly hospital admission rate for all other causes excluding LRTI to control for temporal changes in hospital admission practices.In univariate analyses, hospitalizations for LRTI, pneumonia, and bronchiolitis were less frequent in cohorts exposed to PCVs than in unvaccinated cohorts with no difference between PCV regimens. For pneumonia, the difference in cumulative incidence was 16% (13%; 18%). In multivariate analyses, exposure to any PCV schedule was associated with a lower although statistically non-significant hospitalization risk for pneumonia as compared with unvaccinated cohorts. Again, differences between PCV regimens were minimal.Interpretation of results of this ecological study should be made with care as many factors could influence hospitalizations for respiratory infection in young children. Results are compatible with a modest effect of PCVs in reducing hospitalizations for pneumonia in children. No substantial differences between various PCV schedules were observed.RéSUMé: OBJECTIVES: Au Québec, trois vaccins pneumocoques conjugués (VPC) ont été utilisés de manière séquentielle depuis décembre 2004. L’objectif de l’étude était d’étudier l’association entre l’exposition aux différents calendriers vaccinaux de VPC et le risque d’hospitalisation pour infection respiratoire basse (IRB). MéTHODES: Les enregistrements avec un diagnostic principal d’IRB chez les enfants nés en 2000–2012 et observés jusqu’au 2ième anniversaire ont été extraits de la base provinciale de données hospitalières administratives. Le principal calendrier vaccinal utilisé pour chaque cohorte mensuelle de naissances a été identifié à l’aide du Registre de vaccination de la région de Québec. Le risque d’hospitalisation a été analysé par régression de Poisson en ajustant pour l’âge, la saison de naissance, la température ambiante, la circulation de virus respiratoires et le taux d’hospitalisation hebdomadaire de toutes causes excluant les IRB afin de contrôler les changements temporels dans les pratiques d’admission. RéSULTATS: Dans l’analyse univariée, les taux d’hospitalisation pour l’IRB, pour pneumonie et pour bronchiolite étaient plus faibles dans les cohortes exposées aux VPC que dans les cohortes non vaccinées et sans qu’existe de différences substantielles entre les différents calendriers vaccinaux. Pour la pneumonie, la différence du taux cumulatif à 2 ans était de 16 % [13 %; 18 %]. Dans l’analyse multivariée, l’exposition à n’importe quel calendrier vaccinal était associée à un risque moins élevé mais statistiquement non significatif d’hospitalisation et les différences entre les différents calendriers étaient faibles. CONCLUSIONS: L’interprétation des résultats d’une étude écologique doit être prudente, car de multiples facteurs peuvent influencer l’hospitalisation pour une infection respiratoire chez les jeunes enfants. Nos résultats sont compatibles avec un effet modeste du VPC dans la réduction des hospitalisations pour la pneumonie chez les enfants sans que des différences substantielles aient été observées entre les différents calendriers et vaccins.

Background Maintenance ART with dolutegravir-based dual regimens have proved their efficacy among HIV-1-infected subjects in randomized trials. However, real-life data are scarce, with limited populations and follow-up. Objectives We assessed virological failure (VF) and resistance-associated mutations (RAMs) on dolutegravir maintenance regimens in combination with rilpivirine or with lamivudine or emtricitabine (xTC) and analysed the factors associated with VF. Methods Between 2014 and 2018, all HIV-1-infected adults included in the Dat’AIDS cohort and starting dolutegravir/rilpivirine or dolutegravir/xTC as a maintenance dolutegravir-based dual regimen were selected. VF was defined as two consecutive HIV RNA values >50 copies/mL or a single value >400 copies/mL. We compared cumulative genotypes before initiation of a maintenance dolutegravir-based dual regimen with genotype at VF. Results We analysed 1374 subjects (799 on dolutegravir/rilpivirine and 575 on dolutegravir/xTC) with a median follow-up of 20 months (IQR = 11–31) and 19 months (IQR = 11–31), respectively. VF occurred in 3.8% (n = 30) of dolutegravir/rilpivirine subjects and 2.6% (n = 15) of dolutegravir/xTC subjects. Among subjects receiving dolutegravir/rilpivirine, two genotypes harboured emerging RAMs at VF: E138K on NNRTI (n = 1); and E138K+K101E on NNRTI and N155H on INSTI (n = 1). Among subjects receiving dolutegravir/xTC, no new RAM was detected. The only predictive factor of VF on dolutegravir/rilpivirine was the history of failure on an NNRTI-based regimen (adjusted HR = 2.97, 95% CI = 1.28–6.93). No factor was associated with VF on dolutegravir/xTC. Conclusions In this large real-life cohort, dolutegravir/rilpivirine and dolutegravir/xTC sustained virological suppression and were associated with a low rate of VF and RAM emergence. Careful virological screening is essential before switching to dolutegravir/rilpivirine in virologically suppressed patients with a history of NNRTI therapy.

Background In May 2014, a mass vaccination campaign with four-component meningococcal serogroup B (4CMenB) vaccine was launched in a localized region of Quebec, Canada experiencing high invasive meningococcal B disease endemicity. Active post-marketing surveillance identified several cases of nephrotic syndrome (NS) among ∼49,000 vaccinated individuals aged 2 months to 20 years. We report the epidemiologic investigation of this potential vaccine safety signal. Methods Active vaccine safety surveillance was conducted electronically, with participants completing an online questionnaire prompted at 7 days after each dose and 6 months following the last dose. Additional NS cases were sought from provincial hospitalization and emergency room databases. Results In the year following the first dose of 4CMenB vaccination, four confirmed NS cases (three hospitalized) were identified among vaccinated children 2–5-years-old with onset several months post-vaccination. None had renal biopsy but given their age, and positive response to steroids, idiopathic NS was presumptively diagnosed. Among vaccinated children 1–9-years-old, the NS incidence in the year post-vaccination was 17.7 per 100,000 (1 per 5650 vaccinees) with an NS hospitalization rate (i.e. excluding the outpatient case) that was 3.6-fold higher (95%CI = 0.7–11.8; p = 0.12) than the rest of the province for the same period, and 8.3-fold greater (95%CI = 1.1–62.0; p = 0.039) than during the eight years preceding the immunization campaign in the affected region. Conclusion Active safety surveillance identified an unexpected increase in NS incidence following 4CMenB vaccination. Further epidemiological investigation identified four vaccinated cases in total over a 12 month period of follow up. The greater risk in vaccinees had wide confidence intervals with he lower limit including or just above the nul value, an observation with no or marginal statistical significance. The temporal association with vaccination may be explained by other causes and/or chance clustering of a rare event unrelated to vaccination. To confirm or refute a potential link to vaccination, surveillance in other jurisdictions administering 4CMenB to children 1–9-years-old is needed.

Vaccination has a huge public health impact. Maintaining vaccine coverage is key to avoid the devastating consequences of resurgence. In the Province of Quebec, vaccine coverage in young children are sub-optimal, mostly due to ambivalence toward vaccine safety and efficacy. We previously conducted a regional study in the Quebec’s Eastern Townships region, the PromoVac Study, to test a new educational intervention, based on motivational interviewing techniques, aimed at promoting infant vaccination. This first study evidenced that the intervention led to a marked increase in mothers’ intention to vaccinate, and vaccine coverage in their infants. The current study protocol aims at scaling up these results at a provincial level using a randomized controlled trial design. This pragmatic, randomized, controlled, parallel-group clinical trial will compare the effectiveness of the motivational interviewing to an educational intervention, including the distribution of an information flyer as standard of care on vaccination coverage in four maternity wards across the Province of Quebec (PromovaQ). Adult mothers of children born in participating maternity wards were recruited between March 2014 and February 2015. Vaccination coverage will be assessed at 3-years of age, thus the trial is expected to be completed in March 2019. Statistical analyses will be conducted under the intention-to-treat principle. Vaccine coverage will be analyzed using Chi-squared distribution testing and logistic regression to identify determinant factors. Secondary outcomes will include vaccine hesitation and intention scores, mother’s knowledge, attitudes and beliefs about immunization, and psychosocial determinants of intention to vaccinate. In the case results of this Provincial RCT be confirmed, serious consideration should then be given by Ministry of Health authorities to the possible implementation of MI-based strategies across provincial maternity wards. To ensure adequate input and secure implementation, study design and results will be reviewed with relevant stakeholders, including the children’s families, and provincial and regional decision-makers. Results will be adapted and shared with all stakeholders. ClinicalTrials.gov NCT02666872 (Retrospectively registered as January 28, 2016).

OBJECTIVES Here, we examine (1) if the sex-related differences in iron (Fe) and copper (Cu) isotope ratios, represented as δ56 Fe and δ65 Cu values, respectively observed in humans exist in bulk occipital bone and incisors of male and female non-human primates, and (2) if the variation of Fe and Cu isotope ratios, known to vary in human blood as a factor of age are similar in non-human primate bone. MATERIALS AND METHODS Isotope ratios were measured from the skeletal elements of 20 rhesus macaques (Macaca mulatta) with known life history traits. The metals were purified by column chromatography and their isotope ratios measured by MC-ICP-MS. Data were analyzed using generalized additive models (GAM). RESULTS When accounting for age and sex independently, we found a significant relationship between δ65 Cu values and occipital bone, but not in incisors. There were no significant relationships observed between δ56 Fe values, occipital bone, or incisors. Similarly, there were no significant relationships observed between δ56 Fe values, δ65 Cu values, and age. DISCUSSION We suggest that Cu and Fe isotope ratios have the potential to be useful supplementary tools in future research in biological anthropology, but additional studies are needed to further verify the relationship between sex, age, δ65 Cu, and δ56 Fe values in primates.

Our organs consist of multiple cell types that ensure proper architecture and function. How different cell types coexist and interact to maintain their homeostasis in vivo remain elusive. The skin epidermis comprises mostly epithelial cells, but also harbors Langerhans cells (LCs) and Dendritic Epidermal T cells (DETCs). In response to injury or infection, LCs and DETCs become activated and play critical immunological roles. During homeostasis, they coexist with epithelial cells in the basal layer of the epidermis. Whether, and how, distributions of LCs and DETCs are regulated during homeostasis is unclear. Here, we addressed this question by tracking LCs, DETCs and epithelial basal cells over time within the skin of live adult mice. We show that LCs and DETCs maintain their overall position despite continuous turnover of neighboring basal epithelial stem cells. Moreover, LCs and DETCs rapidly and maximally explore basal epithelial cell junctions through their dendritic extensions. Altering the epithelial cell density triggers corresponding changes in the immune cell density, but not vice versa, suggesting that epithelial cells determine immune tissue composition in the epidermis. Moreover, LCs and DETCs are organized in a tiling pattern that is actively maintained. When LCs or DETCs are ectopically removed, neighboring epidermal LCs or DETCs, respectively, move into the emptied spaces and re-establish the tiling pattern. Finally, LCs require the GTPase Rac1 to maintain their positional stability, density and tiling pattern. Overall, we discovered that epidermal cells regulate the density of immune cells during homeostasis, and that immune cells actively maintain a non-random spatial distribution, reminiscent of neuronal self-avoidance. We propose that these cellular mechanisms provide the epidermis with an optimal response to environmental insults.

Background Many countries are grappling with growing numbers of parents who delay or refuse recommended vaccinations for their children. This has created a need for strategies to address vaccine hesitancy (VH) and better support parental decision-making regarding vaccination. Aim To assess vaccination intention (VI) and VH among parents who received an individual motivational-interview (MI) based intervention on infant immunisation during post-partum stay at a maternity ward between March 2014 and February 2015. Methods This non-controlled pre-/post-intervention study was conducted using the results from parents enrolled in the intervention arm of the PromoVaQ randomised control trial (RCT), which was conducted in four maternity wards across the Province of Quebec. Participants (n = 1,223) completed pre- and post-intervention questionnaires on VI and VH using Opel’s score. Pre-/post-intervention measures were compared using McNemar’s test for categorical variables and Wilcoxon signed-rank test for continuous variables. Results Pre-intervention: overall VI was 78% and significantly differed across maternity wards (74%, 77%, 84%, 79%, p = 0.02). Post-intervention: VI rose significantly across maternity wards (89%, 85%, 95%, 93%) and the overall increase in VI was 12% (78% vs 90%, p Conclusions Compared with pre-intervention status, participants who received the MI-based intervention on immunisation displayed lower hesitancy and greater intention to vaccinate their infant at 2 months of age.

BACKGROUND For patients who have experienced adverse events following immunization (AEFI) or who have specific medical conditions, there is limited evidence regarding the best approach to immunization. The Special Immunization Clinics (SICs) Network was established to standardize patient management and assess outcomes after reimmunization. The study objective was to describe the first 2 years of the network's implementation. METHODS Twelve SICs were established across Canada by infectious diseases specialists and allergists. Inclusion criteria were as follows: local reaction ≥ 10 cm, allergic symptoms < 24 hours postimmunization, neurologic symptoms and other AEFI or medical conditions of concern. Eligible patients underwent a standardized evaluation, causality assessment was performed, immunization recommendations were made by expert physicians and patients were followed up to capture AEFI. After individual consent, data were transferred to a central database for analysis. RESULTS From June 2013 to May 2015, 151 patients were enrolled. Most were referred for prior AEFI (132/151, 87%): 42 (32%) for allergic-like reactions, 31 (23%) for injection-site reactions, 20 (15%) for neurologic symptoms and 39 (30%) for other systemic symptoms. Nineteen patients (13%) were seen for underlying conditions that complicated immunization. Reimmunization was recommended for 109 patients, 60 of whom (55%) were immunized and followed up. Eleven patients (18%) experienced recurrence of their AEFI; none were serious (eg, resulting in hospitalization, permanent disability or death). CONCLUSIONS The most frequent reasons for referral to a SIC were allergic-like events and injection site reactions. Reimmunization was safe in most patients. Larger studies are needed to determine outcomes for specific types of AEFI.

Tracking stem cell fate in time and space After injury and during homeostasis, tissues rely on the balance of cell loss and renewal. Rompolas et al. visualized individual stem cells over their lifetime in the epidermis of live mice. Tracking stem cells over multiple generations revealed that tissue homeostasis in the mouse epidermis is not maintained by asymmetric cell division as previously thought, but through the coordination of sibling cell fate and lifetimes. Furthermore, differentiating stem cells reused the existing spatial organization of the epidermis. Science , this issue p. 1471

Periconceptional folic acid significantly reduces the risk of neural tube defects. It is difficult to achieve optimal levels of folate by diet alone, even with fortification of flour, especially because flour consumption in Canada is slightly decreasing. Intermittent concerns have been raised concerning possible deleterious effects of folate supplementation, including the masking of symptoms of vitamin B12 deficiency and an association with cancer, especially colorectal cancer. Both concerns have been disproved. The Canadian Paediatric Society endorses the following steps to enhance folate intake in women of child-bearing age: encouraging the consumption of folate-rich foods such as leafy vegetables, increasing the level of folate food fortification, taking a supplement containing folate and B12, and providing free folate supplementation to disadvantaged women of child-bearing age. These recommendations are consistent with those of the Society of Obstetricians and Gynaecologists of Canada.La consommation d’acide folique pendant la période périconcep- tionnelle réduit considérablement le risque d’anomalie du tube neural. Il est difficile d’atteindre un taux optimal de folate à partir du seul régime alimentaire, malgré l’enrichissement de la farine, surtout que la consommation de ce produit diminue légèrement au Canada. Les effets délétères possibles des suppléments de folate ont suscité sporadiquement des inquiétudes, y compris le camouflage des symptômes de carence en vitamine B

Histoplasmosis is a common fungal infection, normally infecting people exposed to demolition sites or bat/bird droppings in the central and eastern states. When a child presents with a chest mass and eosinophilia in a non-endemic region the likelihood of an infectious process like pulmonary histoplasmosis is unknown. A seven year old immunocompetent child with a mediastinal mass and eosinophilia presented with acute cough, fever, non-bloody emesis, and four pound weight loss. A neoplastic work up was negative. Further evaluation showed a positive M band (chronic histoplasmosis infection) and negative H band (acute infection). Tissue obtained by thoracotomy demonstrated necrotizing granulomatous inflammation with calcification consistent with histoplasmosis. Patient recovered after completion of a twelve week course of itraconazole. A mediastinal mass in a symptomatic child has a 50% risk of cancer as the primary diagnosis. The Infectious Disease Society of America guidelines recommend treatment of histoplasmosis granulomatous disease with itraconazole if symptomatic and surgery only for obstruction. Thus our patient did not have a clear indication for surgery. In a child with a mediastinal mass, despite low risk factors should they be evaluated for a fungal infection prior to invasive surgery? This case demonstrates that histoplasmosis can cause a granuloma in a non-endemic region and that an infectious etiology ought to be considered when working up a symptomatic child with a chest mass as it may prevent unnecessary surgery. Keywords: Histoplasmosis, Fungal, Cough, Chest mass, Eosinophilia, Thoracotomy, Mediastinal

Purpose To compare the frequency and the severity of influenza and respiratory syncytial viruses (RSV) infections among children

Decreased active and passive range of motion (ROM) accompanied by pain in the shoulder is a common presentation for patients with frozen shoulder, and it can be difficult to restore normal function. Through the fascial distortion model, physicians can apply a manual technique to rapidly and effectively increase ROM and decrease pain. A 28-year-old man presented 18 months after sustaining a shoulder hyperextension injury. On active and passive ROM examination, he had approximately 90° of shoulder abduction and moderately reduced internal rotation associated with 8/10 pain. After 2 applications of the fascial distortion model, his shoulder restored to full abduction and internal rotation with no pain.

Fibroblasts are an essential cellular and structural component of our organs. Despite several advances, the critical behaviors that fibroblasts utilize to maintain their homeostasis in vivo have remained unclear. Here, by tracking the same skin fibroblasts in live mice, we show that fibroblast position is stable over time and that this stability is maintained despite the loss of neighboring fibroblasts. In contrast, fibroblast membranes are dynamic during homeostasis and extend to fill the space of lost neighboring fibroblasts in a Rac1-dependent manner. Positional stability is sustained during aging despite a progressive accumulation of gaps in fibroblast nuclei organization, while membrane occupancy continues to be maintained. This work defines positional stability and cell occupancy as key principles of skin fibroblast homeostasis in vivo, throughout the lifespan of mice, and identifies membrane extension in the absence of migration as the core cellular mechanism to carry out these principles.

SUMMARYYoung age, adverse environmental conditions and infectious agents are established risk factors of lower respiratory tract infection (LRTI), whereas pneumococcal conjugate vaccines may be protective. To explore their relative role as predictors of hospitalizations under the continental climate prevailing in the province of Quebec, Canada, an ecological study was performed. Records with a main diagnosis of LRTI in children born during 2007–2010 and observed up to their second-year anniversary were extracted from the provincial hospital administrative database. Respiratory virus surveillance data and statistics on ambient air temperature were obtained. Vaccine use in different birth cohorts was derived from the Quebec City Immunization Registry. Additive and multiplicative Poisson regression models were applied to estimate attributable fractions. Age, month of birth, ambient temperature, and respiratory syncytial virus (RSV), human metapneumovirus (hMPV) and influenza-positive test proportions were significant predictors of LRTI hospitalizations. No substantial differences were observed in cohorts exposed to the 7-valent or 10-valent pneumococcal conjugate vaccines. In the additive model, the fraction of hospitalizations explained by temperature variation was 37%, whereas RSV circulation explained 28%, hMPV 4% and influenza 1%. Complex interplay between biological, environmental and social mechanisms may explain the important role of ambient air temperature in predicting LRTI hospitalization risk in young children.

Paradoxical immune reconstitution inflammatory syndrome is a well-described entity even in immunocompetent children, principally in association with Mycobacterium tuberculosis infections. Central nervous system involvement is a potential life-threatening form, sometimes refractory to standard treatment. We report the case of an HIV-negative refugee teenager, who presented with brain tuberculomas and pseudoabscesses responsive only to thalidomide.

SUMMARYStreptococcus pneumoniaeis an important cause of community-acquired pneumonia and pneumococcal conjugate vaccines (PCVs) may reduce this burden. This study's goal was to analyse trends in lower respiratory tract infections (LRTI) hospitalisations before and during a routine vaccination programme targeting all newborns with PCV was started in the province of Quebec, Canada in December 2004. The study population included hospital admissions with a main diagnosis of LRTI among 6–59 month-old Quebec residents from April 2000 to December 2014. Trends in proportions and rates were analysed using Cochran-Armitage tests and Poisson regression models. We observed a general downward trend in all LTRI hospitalisations rate: from 11·55/1000 person-years in 2000–2001 to 9·59/1000 in 2013–2014, a 17·0% reduction, which started before the introduction of PCV vaccination. Downward trends in hospitalisation rates were more pronounced for all-cause of pneumonia (minus 17·8%) than for bronchiolitis (minus 15·4%). There was also a decrease in the mean duration of hospital stay. There was little evidence that all-cause pneumonia decreased over the study period due mainly to the introduction of PCVs. Trends may be related to changes in clinical practice. This study casts doubt on the interpretation of ecological analyses of the implementation of PCV vaccination programmes.

Cells in healthy tissues acquire mutations with surprising frequency. Many of these mutations are associated with abnormal cellular behaviours such as differentiation defects and hyperproliferation, yet fail to produce macroscopically detectable phenotypes1–3. It is currently unclear how the tissue remains phenotypically normal, despite the presence of these mutant cells. Here we use intravital imaging to track the fate of mouse skin epithelium burdened with varying numbers of activated Wnt/β-catenin stem cells. We show that all resulting growths that deform the skin tissue architecture regress, irrespective of their size. Wild-type cells are required for the active elimination of mutant cells from the tissue, while utilizing both endogenous and ectopic cellular behaviours to dismantle the aberrant structures. After regression, the remaining structures are either completely eliminated or converted into functional skin appendages in a niche-dependent manner. Furthermore, tissue aberrancies generated from oncogenic Hras, and even mutation-independent deformations to the tissue, can also be corrected, indicating that this tolerance phenomenon reflects a conserved principle in the skin. This study reveals an unanticipated plasticity of the adult skin epithelium when faced with mutational and non-mutational insult, and elucidates the dynamic cellular behaviours used for its return to a homeostatic state.

Many adult tissues are dynamically sustained by the rapid turnover of stem cells. Yet, how cell fates such as self-renewal and differentiation are orchestrated to achieve long-term homeostasis remains elusive. Studies utilizing clonal tracing experiments in multiple tissues have argued that while stem cell fate is balanced at the population level, individual cell fate - to divide or differentiate – is determined intrinsically by each cell seemingly at random ( 1 2 3 4 5). These studies leave open the question of how cell fates are regulated to achieve fate balance across the tissue. Stem cell fate choices could be made autonomously by each cell throughout the tissue or be the result of cell coordination ( 6 7). Here we developed a novel live tracking strategy that allowed recording of every division and differentiation event within a region of epidermis for a week. These measurements reveal that stem cell fates are not autonomous. Rather, direct neighbors undergo coupled opposite fate decisions. We further found a clear ordering of events, with self-renewal triggered by neighbor differentiation, but not vice-versa. Typically, around 1-2 days after cell delamination, a neighboring cell entered S/G2 phase and divided. Functional blocking of this local feedback showed that differentiation continues to occur in the absence of cell division, resulting in a rapid depletion of the epidermal stem cell pool. We thus demonstrate that the epidermis is maintained by nearest neighbor coordination of cell fates, rather than by asymmetric divisions or fine-tuned cell-autonomous stochastic fate choices. These findings establish differentiation-dependent division as a core feature of homeostatic control, and define the relevant time and length scales over which homeostasis is enforced in epithelial tissues.

Tissue repair is fundamental to our survival as tissues are challenged by recurrent damage. During mammalian skin repair, cells respond by migrating and proliferating to close the wound. However, the coordination of cellular repair behaviours and their effects on homeostatic functions in a live mammal remains unclear. Here we capture the spatiotemporal dynamics of individual epithelial behaviours by imaging wound re-epithelialization in live mice. Differentiated cells migrate while the rate of differentiation changes depending on local rate of migration and tissue architecture. Cells depart from a highly proliferative zone by directionally dividing towards the wound while collectively migrating. This regional coexistence of proliferation and migration leads to local expansion and elongation of the repairing epithelium. Finally, proliferation functions to pattern and restrict the recruitment of undamaged cells. This study elucidates the interplay of cellular repair behaviours and consequent changes in homeostatic behaviours that support tissue-scale organization of wound re-epithelialization.

In Canada, several new vaccines were recently approved for clinical use or are expected to be soon. Decision-makers are faced with the choice whether or not to include these vaccines in publicly funded vaccination programs. The aim of this study was to assess Canadian pediatricians' and family physicians' opinions regarding 7 new vaccines, and perceived priority for the introduction of new programs. A self-administered, anonymous, mail-based questionnaire was sent during fall 2009 to a random sample of 1182 family physicians and to all 1852 Canadian pediatricians. Responses to 8 statements regarding frequency and severity of the diseases, efficacy and safety of the vaccines as well as feasibility of immunization programs were used to calculate priority scores to rank the 7 potential new vaccination programs (calculated scores ranging from 0 to 100). Overall response rate was 43%. The majority of respondents perceived the health and economic burden of diseases prevented by the seven new vaccines as important and considered new vaccines to be safe and effective. More than 90% of physicians strongly agreed or agreed that the new vaccines would be or are currently well accepted by the public and by the health professionals who administer vaccines, except for the HPV and rotavirus vaccines (respectively 30% and 29% strongly agreed or agreed). Mean priority scores were: 77.4 out of 100 for the measles, mumps, rubella and varicella (MMRV) combined vaccine; 75.6 for the hexavalent (DTaP-IPV-Hib-HBV) vaccine; 73.1 for the new pneumococcal conjugate vaccines; 69.8 for the meningococcal ACYW135; 68.9 for the combined hepatitis A and B; 63.5 for the human papillomavirus vaccine and 56.9 for the rotavirus vaccine. Health professionals' opinion is an important element to consider in the decision-making process regarding implementation of new immunization programs. Without health professional support, the introduction of a new vaccination program may be unsuccessful. In this study, the MMRV and the hexavalent (DTaP-IPV-Hib-HBV) vaccines received the highest ratings.

OBJECTIVE: Young children are generally considered immunologically naive with respect to influenza exposure opportunities; thus, a 2-dose schedule is recommended when a child is first immunized with conventional influenza vaccine lacking adjuvant. We estimated the effectiveness of a single pediatric dose of AS03-adjuvanted vaccine against hospitalization for confirmed pandemic influenza A/H1N1 (pH1N1) infection in children aged 6 months to 9 years during the fall 2009 vaccination campaign. METHODS: In a matched case-control design, case subjects were children hospitalized for pH1N1 infection in the Fall of 2009, in Quebec, Canada. Controls were nonhospitalized children, matched by age and region of residence. Vaccination status in case subjects and controls was ascertained in relation to the case subject's date of illness onset. Vaccine effectiveness was estimated through conditional logistic regression. RESULTS: The overall effectiveness of a single pediatric dose of vaccine administered ≥14 days before illness onset was 85% (95% confidence interval [CI]: 61% to 94%), varying according to age category but with wide and overlapping CIs: 92% (95% CI: 51% to 99%) in 6–23 month-old children, 89% (95% CI: 34% to 98%) in 2–4 year-olds, and 79% (95% CI: −31% to 96%) in 5–9 year-olds. Overall vaccine effectiveness for immunization ≥10 days before illness onset was slightly lower at 80% (95% CI: 60% to 90%), with similar variation according to age. CONCLUSION: In children aged 6 months to 9 years, a single pediatric dose of the AS03-adjuvanted pH1N1 vaccine was highly protective against hospitalization beginning at 10 and 14 days after vaccination.

Rotavirus is the leading cause of dehydration and hospitalization due to gastroenteritis (GE) in young children. Almost all children are affected by the age of 5 years. Two safe and effective rotavirus vaccines are available for clinical use in Canada. In the context where rotavirus vaccination is recommended, but not publicly funded, we have assessed paediatricians’ knowledge, attitudes and beliefs (KAB) regarding rotavirus disease and its prevention by vaccination. A self-administered anonymous questionnaire based upon the Health Belief Model and the Analytical framework for immunization programs was mailed to all 1852 Canadian paediatricians. The response rate was 50%. The majority of respondents rated consequences of rotavirus infection for young patients as moderate. Sixty-six percent considered that rotavirus disease occur frequently without vaccination and 62% estimated that the disease generates a significant economic burden. Sixty-nine percent of respondents considered rotavirus vaccines to be safe and 61%, to be effective. The reduction of severe GE cases was seen as the main benefit of rotavirus vaccination, while the risk of adverse events was the principal perceived barrier. Fifty-three percent (53%) indicated a strong intention to recommend rotavirus vaccines. In multivariate analysis, main determinant of paediatricians’ intention to recommend rotavirus vaccines was the perceived health and economic burden of rotavirus diseases (partial R2 = 0.49, p

Nucleoside analogs are efficacious cancer chemotherapeutics due to their incorporation into tumor cell DNA. However, they exhibit vastly different antitumor efficacies, suggesting that incorporation produces divergent effects on DNA replication. Here we have evaluated the consequences of incorporation on DNA replication and its fidelity for three structurally related deoxyguanosine analogs: ganciclovir (GCV), currently in clinical trials in a suicide gene therapy approach for cancer, D-carbocyclic 2'-deoxyguanosine (CdG) and penciclovir (PCV). GCV and CdG elicited similar cytotoxicity at low concentrations, whereas PCV was 10-100-fold less cytotoxic in human tumor cells. DNA replication fidelity was evaluated using a supF plasmid-based mutation assay. Only GCV induced a dose-dependent increase in mutation frequency, predominantly GC--TA transversions, which contributed to cytotoxicity and implicated the ether oxygen in mutagenicity. Activation of mismatch repair with hydroxyurea decreased mutations but failed to repair the GC--TA transversions. GCV slowed S-phase progression and CdG also induced a G2/M block, but both drugs allowed completion of one cell cycle after drug treatment followed by cell death in the second cell cycle. In contrast, PCV induced a lengthy early S-phase block due to profound suppression of DNA synthesis, with cell death in the first cell cycle after drug treatment. These data suggest that GCV and CdG elicit superior cytotoxicity due to their effects in template DNA, whereas strong inhibition of nascent strand synthesis by PCV may protect against cytotoxicity. Nucleoside analogs based on the carbohydrate structures of GCV and CdG is a promising area for antitumor drug development.

Suicide gene therapy with herpes simplex virus thymidine kinase and ganciclovir is notable for producing multi-log cytotoxicity in a unique pattern of delayed cytotoxicity in S-phase. Because hydroxyurea, a ribonucleotide reductase inhibitor that activates mismatch repair, can increase sensitivity to ganciclovir, we evaluated the role of MLH1, an essential mismatch repair protein, in ganciclovir cytotoxicity. Using HCT116TK (HSV-TK-expressing) colon carcinoma cells that express or lack MLH1, cell survival studies demonstrated greater ganciclovir sensitivity in the MLH1 deficient cells, primarily at high concentrations. This could not be explained by differences in ganciclovir metabolism, as the less sensitive MLH1-expresssing cells accumulated more ganciclovir triphosphate and incorporated more of the analog into DNA. SiRNA suppression of MLH1 in U251 glioblastoma or SW480 colon carcinoma cells also enhanced sensitivity to high concentrations of ganciclovir. Studies in a panel of yeast deletion mutants confirmed the results with MLH1, and further suggested a role for homologous recombination repair and several cell cycle checkpoint proteins in ganciclovir cytotoxicity. These data suggest that MLH1 can prevent cytotoxicity with ganciclovir. Targeting mismatch repair-deficient tumors may increase efficacy of this suicide gene therapy approach to cancer treatment.

Peripheral and CNS inflammation leads to aberrations in developmental and postnatal neurogenesis, yet little is known about the mechanism linking inflammation to neurogenic abnormalities. Specific miRs regulate peripheral and CNS inflammatory responses. miR-155 is the most significantly upregulated miR in primary murine microglia stimulated with lipopolysaccharide (LPS), a proinflammatory Toll-Like Receptor 4 ligand. Here, we demonstrate that miR-155 is essential for robust IL6 gene induction in microglia under LPS stimulation in vitro. LPS-stimulated microglia enhance astrogliogenesis of cocultured neural stem cells (NSCs), whereas blockade of IL6 or genetic ablation of microglial miR-155 restores neural differentiation. miR-155 knock-out mice show reversal of LPS-induced neurogenic deficits and microglial activation in vivo. Moreover, mice with transgenic elevated expression of miR-155 in nestin-positive neural and hematopoietic stem cells, including microglia, show increased cell proliferation and ectopically localized doublecortin-positive immature neurons and radial glia-like cells in the hippocampal dentate gyrus (DG) granular cell layer. Microglia have proliferative and neurogenic effects on NSCs, which are significantly altered by microglial miR-155 overexpression. In addition, miR-155 elevation leads to increased microglial numbers and amoeboid morphology in the DG. Our study demonstrates that miR-155 is essential for inflammation-induced neurogenic deficits via microglial activation and induction of IL6 and is sufficient for disrupting normal hippocampal development.

Suicide gene therapy with the herpes simplex virus thymidine kinase (HSV-TK) cDNA and ganciclovir can elicit cytotoxicity to transgene-expressing and nonexpressing bystander cells via transfer of ganciclovir phosphates through gap junctions. HeLa cells do not exhibit bystander cytotoxicity, although we showed recently that they transfer low levels of ganciclovir phosphates to bystander cells. Here, we attempted to induce bystander cytotoxicity using hydroxyurea, an inhibitor of ribonucleotide reductase, to decrease the endogenous dGTP pool, which should lessen competition with ganciclovir triphosphate for DNA incorporation. Addition of hydroxyurea to cocultures of HSV-TK-expressing and bystander cells synergistically increased ganciclovir-mediated cytotoxicity to both cell populations while producing primarily an additive effect in cultures of 100% HSV-TK-expressing cells. Whereas HSV-TK-expressing cells in coculture were ∼50-fold less sensitive to ganciclovir compared with cultures of 100% HSV-TK-expressing cells, addition of hydroxyurea restored ganciclovir sensitivity. Quantification of deoxynucleoside triphosphate pools showed that hydroxyurea decreased dGTP pools without significantly affecting ganciclovir triphosphate levels. Although hydroxyurea significantly increased the ganciclovir triphosphate:dGTP value for 12 to 24 hours in HSV-TK-expressing and bystander cells from coculture (1.4- to 4.9-fold), this value was increased for

The combination of cytosine deaminase (CD) and herpes simplex virus thymidine kinase (HSV-TK) suicide gene protocols has resulted in enhanced antitumor activity in cultured tumor cells and animal models. In this study, we show that concurrent addition of prodrugs 5-fluorocytosine (5-FC) and ganciclovir (GCV) was less efficacious than sequential treatment in human DU145 prostate carcinoma cells infected with an adenovirus containing a CD/HSV-TK fusion gene. If cells were incubated for 24 hours with 5-FC followed by a 24-hour GCV treatment, GCV triphosphate levels were 2-fold higher, incorporation of GCV monophosphate into DNA was 2.5-fold higher, and growth inhibition was increased 4-fold compared with simultaneous treatment. As expected, cellular dTTP levels were reduced during the 5-FC preincubation. However, dGTP pools also declined parallel to the dTTP decrease. Similar results were obtained when 5-fluorouracil or 5-fluoro-2′-deoxyuridine was used instead of CD/5-FC. These data allowed us to propose a novel hypothesis for the synergistic interaction between CD/5-FC and HSV-TK/GCV treatments. We suggest that the CD/5-FC–mediated reduction of dTTP results in a concurrent decrease of dGTP due to allosteric regulation of ribonucleotide reductase. Because dGTP is the endogenous competitor of GCV triphosphate, depleted dGTP at the time of GCV addition results in increased GCV in DNA and cell kill. In fact, addition of deoxyguanosine during the 5-FC incubation reverses the dGTP depletion, reduces the amount of GCV monophosphate incorporated into DNA, and prevents the CD/5-FC–mediated enhancement of HSV-TK/GCV cytotoxicity. Understanding this mechanistic interaction may help recognize better strategies for creating more efficacious clinical protocols. (Cancer Res 2006; 66(6): 3230-7)

To improve bystander cell killing with HSV-TK/GCV, we have utilized dFdCyd to reduce endogenous dGTP, which competes with GCVTP for incorporation into DNA. In this study we demonstrate the ability of dFdCyd to enhance GCV-mediated bystander cytotoxicity in cultured SW620 human colon carcinoma cells as well as in a murine xenograft model. In vitro, dFdCyd reduced cellular dGTP levels and produced a fourfold increase in the GCVTP:dGTP ratio. This elevated GCVTP:dGTP ratio resulted in a twofold increase in GCVMP incorporation into DNA in bystander cells cocultured with HSV-TK-expressing cells. The combination of GCV and dFdCyd was determined to be synergistic by isobologram analysis of bystander cytotoxicity. Tumors in mice treated with GCV and dFdCyd exhibited a significant growth delay requiring 40 days to obtain approximately 10 times their initial size compared to tumors in PBS- or single-drug-treated animals, which grew rapidly, increasing to a similar size in just 19 to 24 days. In addition, complete tumor regression was observed only in animals treated with both drugs. Furthermore, dFdCyd alone or in combination with GCV produced no evidence of toxicity or significant weight loss. These data suggest that dFdCyd may improve the clinical efficacy of HSV-TK/GCV gene therapies.

Three case reports expand the clinical spectrum of HPeV-3 infections and highlight the need to characterize this new pathogen., A third serotype of human parechovirus (HPeV) has been recently isolated from stool specimens of a young Japanese child with transient paralysis. We report 3 additional cases of neonatal sepsis caused by HPeV-3 in the fall of 2001 in Canadian infants 7–27 days old. All children were hospitalized with high fever, erythematous rash, and tachypnea for a median of 5 days. The viruses isolated from nasopharyngeal aspirates grew slowly on tertiary monkey kidney cells and were successfully passaged on Vero cells. The predicted amino acid identity of the VP0-VP3-VP1 region of the three viruses was 74.6%–74.8%, 73.4%–73.6%, and 97.0%–97.1% when compared to HPeV-1, -2, and –3 prototype strains, respectively. Although different, our isolates were closely related; amino acid identity was 99.6%–100% for the last 3 proteins.

Objective. Although universal immunization against Bordetella pertussis (whooping cough) infection has resulted in dramatic reductions in the incidence of pertussis, outbreaks continue to occur in countries with excellent vaccine coverage. Treatment of infection may ameliorate symptom severity during the catarrhal phase of pertussis but has no effect on established paroxysms, emesis, or apnea if given during the paroxysmal or convalescent phases. Erythromycin, recommended for treatment of pertussis to prevent transmission of infection, is poorly tolerated because of gastrointestinal side effects. We compared the safety and efficacy of erythromycin with azithromycin for treatment of pertussis in a large, randomized, controlled trial that enrolled children from primary care practices in 1 American and 11 Canadian urban centers. Methods. Children who were 6 months to 16 years of age and had cough illness that was suspected to be or was culture confirmed as pertussis were randomized to azithromycin (10 mg/kg on day 1 and 5 mg/kg on days 2–5 as a single dose) or erythromycin estolate (40 mg/kg/day in 3 divided doses for 10 days) with stratification by center. The primary outcome measure was bacteriologic cure of infection as determined by cultures of nasopharyngeal aspirates. Culture-positive participants had a second aspirate collected at the end of therapy (days 5–7 for azithromycin, days 10–12 for erythromycin) and 1 week after therapy. Bacteriologic cure was defined as negative cultures at the end of therapy. Bacteriologic relapse was defined as a positive culture 1 week after completion of therapy and after a negative end-of-therapy culture. Secondary outcomes were pertussis diagnosed by serology and polymerase chain reaction (PCR), treatment-associated adverse events, compliance, and presence of clinical symptoms at the end of the treatment course. Serology was performed using standard enzyme-linked immunosorbent assay methods. A participant was considered to have pertussis when the PCR was positive or a 4-fold increase in pertussis toxin antibody between baseline and follow-up visits was observed. PCR was performed using a 1046-bp ClaI DNA fragment from B pertussis. Adverse events (nausea, vomiting, diarrhea, any gastrointestinal complaint, or other) were determined by a parent-completed diary that was reviewed with study personnel during study visits. Compliance was measured by review of the parent medication diary during study visits and observation of medication containers by the pharmacist at study completion. Symptoms were determined by history collected by study personnel at enrollment and subsequently from the diary. The design of the study was an equivalence trial, aimed at demonstrating that the bacteriologic failure rates with the 2 therapies did not differ by >8%. For the safety analysis, all participants who received at least 1 dose of study drug were included. In the per-protocol efficacy analysis, all culture-positive participants with end-of-treatment cultures were considered. Results. A total of 477 children were enrolled and randomly assigned to either azithromycin (n = 239) or erythromycin (n = 238). Of these children, 114 (24%) grew B pertussis from nasopharyngeal specimens (azithromycin group: 58 of 239 [24%]; erythromycin group: 56 of 238 [23%]); these children composed the efficacy cohort for the per-protocol and intention-to-treat analyses. Serology and PCR added 52 children to the number considered to have pertussis for a total of 35% (166 of 477) of all children who presented with cough illness. In the safety analysis (antibiotic side effects, compliance) and comparison of cough symptoms after treatment, all randomized children are reported in their assigned treatment group. At end of therapy, bacterial eradication was demonstrated in all 53 patients in the azithromycin group and all 53 patients in the erythromycin group with follow-up cultures available (eradication 100%; 95% confidence interval [CI]: 93.3–100). No bacterial recurrence was demonstrated in children with 1 week posttreatment nasopharyngeal cultures available (51 and 53 participants in the azithromycin and erythromycin arms, respectively [0%, 95% CI: 0–7.0; and 0%, 95% CI: 0–6.7]). No serious adverse events attributable to study drug were observed. Gastrointestinal adverse events were reported less frequently in azithromycin (18.8%; 45 of 239) than in erythromycin estolate (41.2%; 98 of 238) recipients (90% CI on difference: −29.0% to −15.7%) as a result of less nausea (2.9% vs 8.4%; 95% CI: −8.9% to −2.0%), less vomiting (5.0% vs 13.0%; 95% CI: −4.9% to −1.4%), and less diarrhea (7.1% vs 11.8%; 95% CI: −9.0% to −0.3%). Children who were randomized to azithromycin were much more likely to have complied with antimicrobial therapy over the treatment period. In the azithromycin group, 90% of children took 100% of prescribed doses, whereas only 55% of children in the erythromycin group took 100% of prescribed doses. Conclusions. In this large, multicenter, randomized trial, we found that azithromycin is as effective as erythromycin estolate for the treatment of pertussis in children. Gastrointestinal adverse events were much more common with erythromycin treatment than azithromycin. Compliance with therapy was markedly better with azithromycin than with erythromycin in this study.

L’interet et la faisabilite de la cryoconservation des spermatozoides testiculaires humains sont maintenant bien demontres. Toutefois, la realisation de la congelation de ces spermatozoides reste empirique, elle est basee essentiellement sur les techniques utilisees pour les spermatozoides de l’ejaculât. Les spermatozoides testiculaires sont consideres comme des gametes encore «immatures» et sont toujours retrouves en quantite limitee. Une meilleure connaissance de leurs caracteristiques cryobiologiques contribuerait a mieux definir le type d’echantillon a congeler (tissu, suspension cellulaire contenant des spermatozoides.…), a faire evoluer les processus de congelation/decongelation et la nature des milieux cryoprotecteurs et a adapter les conditions de stockage. Compte tenu des caracteristiques fonctionnelles de ces spermatozoides et du fait qu’ils sont utilises exclusivement en ICSI, de nouveaux tests d’evaluation de la resistance a la congelation bases notamment sur leur qualite nucleaire devraient etre developpes.

Gap junctional intercellular communication and expression of gap junction proteins (connexins) are decreased frequently in neoplastic cells including human ovarian carcinoma cells. In order to test the hypothesis that these changes contribute to the neoplastic phenotype of ovarian carcinoma cells, we transfected human ovarian carcinoma SKOV-3 cells with connexin43. Stable, connexin43-expressing transfectants were characterized for cell proliferation in vitro in normal, low-serum, and serum-free culture medium, for tumorigenicity in nude mice, and for sensitivity to adriamycin in vitro. Transfected clones expressed higher levels of connexin43 and gap junctional intercellular communication, reduced proliferation and greater dependence upon serum for growth in vitro, decreased tumor formation, increased sensitivity to adriamycin, and reduced expression of p-glycoprotein. These data suggest that gap junctional intercellular communication and/or connexin43 expression suppresses the neoplastic phenotype of ovarian carcinoma cells and their downregulation is involved in neoplastic transformation of ovarian epithelial cells. The increased sensitivity to adriamycin and elevated expression of p-glycoprotein by the transfected cells also suggest that gap junctional intercellular communication and connexin43 expression are involved in drug sensitivity and might be manipulated to enhance the clinical response.

We have previously demonstrated with several cell lines in vitro that hydroxyurea (HU) synergistically enhances ganciclovir (GCV)-mediated cytotoxicity in bystander cells. In this study, we evaluated the role of DNA synthesis inhibition on enhanced bystander killing and assessed whether addition of HU would improve the efficacy of the HSV-TK/GCV system in vivo. Compared with GCV treatment alone, addition of HU resulted in increased DNA synthesis inhibition and delayed progression through S phase following removal of drug. In a xenograft tumor model, 1:10 and 1:1 mixtures of HSVtk- and LacZ-expressing SW620 cells were injected s.c. in the flanks of nude mice and treated i.p. (100 mg/kg GCV, 1500 mg/kg HU) daily for 5 days. Tumors from mice treated with GCV alone grew rapidly and increased to 10 times their initial size in 15.7 +/- 1.8 and 16.0 +/- 0.9 days for 1:10 and 1:1 mixtures, respectively. However, when both GCV and HU were administered in combination, a single complete tumor regression was observed in both the 1:10 and 1:1 groups. In the remaining mice treated with GCV/HU, it took 23.2 +/- 2.1 (1:10) and 26.4 +/- 3.8 days (1:1) to obtain a similar 10-fold increase in tumor size.

Individual sperm from men with balanced translocations have different chromosomal contents. Thus, an estimation of the overall sperm chromosomal imbalance of such patients could help to give the couple an adapted genetic counselling. We report here the study of a balanced translocation carrier, t(17;22) (q11;q12) whose reproductive history reported four miscarriages. Moreover, he had an abnormal semen analysis with oligoteratozoospermia. The meiotic segregation pattern was examined in 700 sperm, using fluorescence in-situ hybridization (FISH). Nineteen percent of the sperm had balanced translocations or were normal. All other sperm were unbalanced (81%) and their distribution was observed as follows: the frequencies of adjacent 1, adjacent 2 and 3:1 segregations were 12.9, 5.8 and 46.8% respectively. Among the segregations scored, 13.7% were related to second meiotic division abnormalities. Less than 2% of the total sperm scored were not explained. The 3:1 segregation was present at a very high rate, which is very unusual. In cases of balanced translocations, we believe that no general features can be drawn. Thus, the FISH technique may be very helpful for genetic counselling, which remains an important step and must be done with care.

We investigate a new experimental method for detecting molecules from the ultraviolet to the near infrared range using ultrashort laser pulses. Two types of sources are used: a white light continuum generated by 200khz Ti: Sapphire regenerative amplifier system, and a near-gaussian femtosecond pulse (100 fs) generated by an optical parametric amplifier and tunable from 0.45 to 2.4 μm. Up to now, this technique has only been performed in the visible domain. Both broadband sources allow the detection of the oxygen and the water vapor bands. Moreover, a new extraction method has been implemented, which provides the molecules' concentration by using a nonlinear fit technique.

OBJECTIVES: Given the improved efficacy of the nasal live-attenuated influenza virus vaccine (LAIV) compared with the injectable vaccine in children, we aimed to determine its safety in individuals with cystic fibrosis (CF). METHODS: A cohort of 168 study participants, aged 2 to 18 years with CF, vaccinated with LAIV between October 1, 2012, and January 30, 2013, was followed prospectively for 56 days after initial vaccination in 3 pediatric CF clinics across the province of Quebec. Days 0 to 28 post-LAIV were considered the at-risk period for all outcomes of interest, and days 29 to 56 post-LAIV were considered the non–at-risk period. Incident respiratory deteriorations were defined as an unscheduled medical visit, hospitalization, or a new course of oral antibiotics for respiratory complaints. Using a self-controlled design, incidence rate ratios (IRR) were used to compare at-risk and non–at-risk periods. RESULTS: Comparing at-risk to non–at-risk periods, there was no significant increase in the rate of incident respiratory deteriorations (IRR, 0.72; 95% confidence interval, 0.11–4.27) or all-cause hospitalizations (IRR, 1.16; 95% confidence interval, 0.30–4.81). A greater proportion of participants reported experiencing at least 1 minor respiratory and/or systemic adverse event after immunization during the at-risk period compared with the non–at-risk period (77% vs 54%, respectively). During the first week after LAIV, 13 of 168 (8%) children reported some wheezing, with the vast majority, 9 of 13 (69%), on the day of vaccination. CONCLUSIONS: There was no increased risk of respiratory deterioration or all-cause hospitalization associated with LAIV in our study population. LAIV seems well tolerated in children and adolescents with CF.

Au cours de ces dernieres annees les tentatives d’ICSI realisees avec des spermatozoides testiculaires se sont considerablement accrues. Cependant, l’une des difficultes techniques tient au fait que la mobilite des spermatozoides est extremement reduite. Un accroissement de la mobilite avait ete observe apres une incubation des prelevements testiculaires pendant quelques heures. Dans l’optique d’augmenter les chances de succes, une culture in vitro des spermatozoides, prealable a l’ICSI, a ete envisagee. Nous rapportons ici la revue de la litterature sur ce sujet. Dans les azoospermies obstructives, la culture in vitro presente peu d’interet puisque, excepte une eventuelle “maturation” des spermatozoides pendant cette periode, un pourcentage appreciable de spermatozoides mobiles est souvent observe rapidement apres le traitement de la biopsie. Dans les azoospermies non obstructives la culture in vitro parait justifiee, puisque lorsque l’on retrouve des spermatozoides dans la biopsie, ils sont generalement immobiles. Cependant, la plupart des auteurs s’accordent a reconnaitre que les resultats restent aleatoires: au terme de la culture, soit il existe un gain de mobilite, soit au contraire la plupart des spermatozoides sont morts. Quelque soit le type d’azoospermie, les meilleurs resultats sont obtenus apres 3–4 jours de culture. La supplementation des milieux de culture avec de la FSH recombinante s’avere aussi efficace. La culture in vitro peut etre associee a la congelation des spermatozoides et la sequence culture in vitro/congelation donnerait de meilleurs resultats que la sequence congelation/culture in vitro. Les quelques travaux sur l’utilisation en ICSI des spermatozoides maintenus en culture in vitro pendant 1 a 2 jours rapportent des resultats satisfaisants en terme de taux de fecondation et de grossesse. La culture in vitro de spermatozoides testiculaires peut constituer une voie de recherche interessante pour ameliorer les resultats en ICSI, dans les cas ou la quantite de spermatozoides retrouvee et/ou leur mobilite sont tres faibles. De plus, elle nous parait etre un bon modele d’etude des mecanismes d’acquisition de la mobilite des spermatozoides.

Microdeletions of the long arm of the Y chromosome (Yq) are a common cause of male infertility. Since large structural rearrangements of the Y chromosome are commonly associated with a 45,XO/46,XY chromosomal mosaicism, we studied whether submicroscopic Yq deletions could also be associated with the development of 45,XO cell lines. We studied blood samples from 14 infertile men carrying a Yq microdeletion as revealed by polymerase chain reaction (PCR). Patients were divided into two groups: group 1 (n = 6), in which karyotype analysis demonstrated a 45,X/46,XY mosaicism, and group 2 (n = 8) with apparently a normal 46,XY karyotype. 45,XO cells were identified by fluorescence in-situ hybridization (FISH) using X and Y centromeric probes. Lymphocytes from 11 fertile men were studied as controls. In addition, sperm cells were studied in three oligozoospermic patients in group 2. Our results showed that large and submicroscopic Yq deletions were associated with significantly increased percentages of 45,XO cells in lymphocytes and of sperm cells nullisomic for gonosomes, especially for the Y chromosome. Moreover, two isodicentric Y chromosomes, classified as normal by cytogenetic methods, were detected. Therefore, Yq microdeletions may be associated with Y chromosomal instability leading to the formation of 45,XO cell lines.

We report the cytologic features of 15 cases of angiosarcoma from various sites and include 14 fine-needle aspiration (FNA) biopsy specimens and 1 pleural fluid specimen. Six were initial diagnoses with histologic confirmation; an additional case in the liver was an initial diagnosis without tissue confirmation. One case represented lymph node metastasis from a primary prostatic epithelioid angiosarcoma. In 10 cases, immunohistochemical staining for factor VIII-related antigen, CD34, CD31, or Ulex europaeus agglutinin I was performed on the cytology or histology specimen. The aspirates varied in cellularity, and the degree of nuclear atypia ranged from relatively bland in a case of low-grade angiosarcoma of the prostate to highly pleomorphic in a lymph node metastasis from a facial cutaneous angiosarcoma. Vasoformative features such as intracellular RBCs, well-formed vessels, attempts at microacinar/lumen formation, and intracytoplasmic lumens were variably present. The background was bloody in all specimens, with necrosis in rare cases. This cytologic series emphasizes that the cytologic features are heterogeneous but that the diagnosis can be suggested by fine-needle aspiration (FNA) when vasoformative features are present. The diagnosis can be made conclusively by FNA with immunocytochemical confirmation of endothelial differentiation.

Objective.We wished to compare outcomes of respiratory syncytial virus (RSV) infection in children with bronchopulmonary dysplasia (BPD) with those with other pulmonary disorders: cystic fibrosis, recurrent aspiration pneumonitis, pulmonary malformation, neurogenic disorders interfering with pulmona

Background: Interferon regulatory factor (IRF)-1 and IRF-2 are nuclear transcription factors that respond to interferon-γ. IRF-1 acts as the effector arm of the interferon-γresponse in tumor cells, whereas IRF-2 binds to the same DNA consensus sequence and opposes IRF-1 activity. This effect is intact in human and murine tumor models, including melanomas; previous work in our laboratory demonstrated the tumor-suppressing activity of IRF-1 expression in in vivo models and the opposing effect of IRF-2. The expression of IRF-1 and -2 in human solid tumors had not been previously investigated. Methods: Formalin-fixed, paraffin-embedded, archival tissue specimens from 38 human melanomas were obtained and stained with polyclonal anti-IRF-1 and anti-IRF-2 antibodies, using an avidin-biotin-peroxidase complex technique with epitope retrieval. Results: Twenty-nine specimens showed granular cytoplasmic staining with the anti-IRF-1 or anti-IRF-2 antibodies. IRF-1 staining was correlated with less advanced disease. Superficial spreading and in situ lesions exhibited more frequent IRF-1 staining, compared with nodular or metastatic disease. Only more advanced lesions showed neither IRF-1 nor IRF-2 staining. Conclusions: Immunohistochemical staining of archival tissue identified IRF-1 and -2 in human melanomas; this had not been previously demonstrated. IRF-1 staining was correlated with the morphologic characteristics of less advanced disease. Tumor-suppressing effects of IRF-1 may account for the less aggressive biologic features of IRF-1-expressing melanomas, as we would predict from the experimental data.

The present study was undertaken to evaluate the frequency and nature of mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene in infertile patients undergoing intracytoplasmic sperm injection. A total of 90 patients were screened for a panel of 10 mutations in the CFTR gene frequently involved in congenital absence of the vas deferens (CAVD); the patients included 14 with azoospermia and CAVD, 39 patients with azoospermia without CAVD (n = 39) and 37 patients with severe oligozoospermia. The length of the polymorphic polypyrimidine tract (allele 5T, 7T and 9T) in the intron 8/exon 9 splice-acceptor site was also determined. In 10 out of 14 patients with CAVD, CFTR mutations were found; nine patients had one DeltaISOdiaDeltaF508 mutation and one patient had two CFTR mutations (N1303K/R117H). Allele 5T was present in eight of these patients. In six patients, 5T was the non-DeltaISOdiaDeltaF508 allele and in two patients there was no known CFTR mutation. None of the CFTR mutations were observed in patients with azoospermia without CAVD or with severe oligozoospermia and the frequency of allele 5T was 3.6% (three out of 78 alleles) and 1.35% (one out of 74 alleles) respectively. Our observation suggests that the CFTR gene is not involved in either spermatogenesis or in the pathology of the genital tract, except for CAVD.

The objective was to evaluate the efficiency of a biological material-free medium and the role of seminal plasma (SP) in the cryopreservation of human spermatozoa. Normal semen samples and low-quality semen samples were used for this study. After centrifugation of 300 microL fractions of whole semen, pellets were resuspended either in autologous SP or in a chemically defined medium (BM) supplemented or not with 3% bovine serum albumin (BSA); after 15 min at 37 degrees C, the samples were diluted (V/V) with cryoprotective medium (30 mM NaCl; 22 mM sodium citrate, 19.4 mM fructose; 80 mM glutamine; 14%, V/V, glycerol) and maintained for 15 min at room temperature before freezing. Assessment of viability and motility was performed using fresh semen (T0), after centrifugation and resuspension prior to adding the cryoprotectant (T15), after adding the cryoprotectant (T30) and after freezing and thawing (Tpost). In all three resuspending media used, sperm viability and motility (forward and total) decreased (p < 0.05) during both the equilibration period especially before addition of the cryoprotective medium (between T0 and T15) and during the freeze-thaw process comparison between T30 and Tpost. The recovery of viable and motile spermatozoa (post-thaw values/values of fresh samples) was higher (p < 0.05) in normal semen than in low-quality semen. In both groups, the recovery was slightly, but significantly, higher with SP than with BM and the presence of BSA has no beneficial effect. To conclude, these data suggest that SP may reduce the deleterious effects of cryopreservation. Nevertheless cryopreservation of spermatozoa in a medium containing neither SP nor biological substances could offer an acceptable cryoprotection of spermatozoa to be used in assisted fertilization procedures, especially for intracytoplasmic sperm injection.

L’objet de ce travail est d’analyser l’evolution des caracteristiques spermiques en fonction de l’âge. Pour cette etude, nous avons analyse le spermogramme de 2126 hommes âges de 20 a 64 ans qui ont consulte, dans le service de reproduction humaine de Clermont-Ferrand de 1980 a 1994, pour indications feminines d’une fecondation in vitro. La recherche d’une liaison, entre l’âge du patient au moment du spermogramme et 10 parametres spermiques, par l’etude d’une regression lineaire, n’a pas permis de mettre en evidence de correlation significative entre l’âge et les parametres etudies. La comparaison des valeurs moyennes par classes d’âge de 5 ans, a ete effectuee pour le volume, la concentration en spermatozoides, le pourcentage de formes mobiles (normale, diminuee), le pourcentage de formes typiques et mobiles et le pourcentage de formes anormales. Ces valeurs ne different pas significativement selon les classes d’âge etudiees. Entre 20 et 60 ans, les parametres essentiels de spermogramme semblent, en moyenne, independants de l’âge, ce qui permet de penser que la qualite de l’ejaculat est conservee jusqu’a 50/60ans. Cependant, un biais de recrutement de la population etudiee et un grand etalement des valeurs pour un âge donne peuvent expliquer que nous n’ayons pas decele des fluctuations des caracteristiques spermiques liees a l’âge.