Multi-parametric Imaging Using Hybrid PET/MR to Investigate the Epileptogenic Brain

Neuroimaging analysis has led to fundamental discoveries about the healthy and pathological human brain. Different imaging modalities allow garnering complementary information about brain metabolism, structure and function. To ensure that the integration of imaging data from these modalities is robust and reliable, it is fundamental to attain deep knowledge of each modality individually. Epilepsy, a neurological condition characterised by recurrent spontaneous seizures, represents a field in which applications of neuroimaging and multi-parametric imaging are particularly promising to guide diagnosis and treatment. In this PhD thesis, I focused on different imaging modalities and investigated advanced denoising and analysis strategies to improve their application to epilepsy. The first project focused on fluorodeoxyglucose (FDG) positron emission tomography (PET), a well-established imaging modality assessing brain metabolism, and aimed to develop a novel, semi-quantitative pipeline to analyse data in children with epilepsy, thus aiding presurgical planning. As pipelines for FDG-PET analysis in children are currently lacking, I developed age-appropriate templates to provide statistical parametric maps identifying epileptogenic areas on patient scans. The second and third projects focused on two magnetic resonance imaging (MRI) modalities: resting-state functional MRI (rs-fMRI) and arterial spin labelling (ASL), respectively. The aim was to i) probe the efficacy of different fMRI denoising pipelines, and ii) formally compare different ASL data acquisition strategies. In the former case, I compared different pre-processing methods and assessed their impact on fMRI signal quality and related functional connectivity analyses. In the latter case, I compared two ASL sequences to investigate their ability to quantify cerebral blood flow and interregional brain connectivity. The final project addressed the combination of rs-fMRI and ASL, and leveraged graph-theoretical analysis tools to i) compare metrics estimated via these two imaging modalities in healthy subjects and ii) assess topological changes captured by these modalities in a sample of temporal lobe epilepsy patients

Structural changes of in vitro matured buffalo and bovine oocytes following cryopreservation.

The aim of this work was to evaluate chromatin and spindle organization of buffalo and bovine in vitro matured oocytes after vitrification/warming by Cryotop and after their exposure to cryoprotectants (CP). In vitro matured oocytes were vitrified/warmed and exposed to the vitrification/warming solutions containing ethylene glycol (EG), dimethyl sulfoxide (DMSO) and sucrose as CP. Two hours after warming, oocytes were fixed and immunostained for microtubules and nuclei and examined by fluorescence microscopy. Data were analyzed by Chi Square test. A higher percentage of Telophase II stage oocytes was found in the toxicity (26 and 34% in bovine and buffalo) and the vitrification groups (13 and 7% in bovine and buffalo) compared to the control, indicating occurrence of activation. An increased percentage of oocytes with abnormal spindle and chromosome organization was found in oocytes exposed to CP (24 and 13% in bovine; 32 and 30% in buffalo respectively) and in those vitrified (26 and 31% in bovine; 26 and 29% in buffalo respectively) compared to the control (0 in bovine and 2.5 % in buffalo)

HIV Aspartyl Peptidase Inhibitors Interfere with Cellular Proliferation, Ultrastructure and Macrophage Infection of Leishmania amazonensis

Submitted by Sandra Infurna ([email protected]) on 2019-01-08T13:43:09Z No. of bitstreams: 1 Ellenf_Altoe_etal_IOC_2009.pdf: 1452755 bytes, checksum: 77127a59920cef6bca71296107f6ec63 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2019-01-08T13:51:34Z (GMT) No. of bitstreams: 1 Ellenf_Altoe_etal_IOC_2009.pdf: 1452755 bytes, checksum: 77127a59920cef6bca71296107f6ec63 (MD5)Made available in DSpace on 2019-01-08T13:51:34Z (GMT). No. of bitstreams: 1 Ellenf_Altoe_etal_IOC_2009.pdf: 1452755 bytes, checksum: 77127a59920cef6bca71296107f6ec63 (MD5) Previous issue date: 2009Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Universidade Federal do Rio de Janeiro. Centro de Ciências da Saúde. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Microbiologia Geral,. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Universidade Federal do Rio de Janeiro. Centro de Ciências da Saúde. Instituto de Biofísica Carlos Chagas Filho. Rio de Janeiro, RJ, Brasil.Universidade Federal do Rio de Janeiro. Centro de Ciências da Saúde. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Microbiologia Geral,. Rio de Janeiro, RJ, Brasil.Universidade Federal do Rio de Janeiro. Centro de Ciências da Saúde. Instituto de Microbiologia Prof. Paulo de Góes. Departamento de Microbiologia Geral,. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Leishmania is the etiologic agent of leishmanisais, a protozoan disease whose pathogenic events are not well understood. Current therapy is suboptimal due to toxicity of the available therapeutic agents and the emergence of drug resistance. Compounding these problems is the increase in the number of cases of Leishmania-HIV coinfection, due to the overlap between the AIDS epidemic and leishmaniasis

Severe Asthma Standard-of-Care Background Medication Reduction With Benralizumab: ANDHI in Practice Substudy

Background: The phase IIIb, randomized, parallel-group, placebo-controlled ANDHI double-blind (DB) study extended understanding of the efficacy of benralizumab for patients with severe eosinophilic asthma. Patients from ANDHI DB could join the 56-week ANDHI in Practice (IP) single-arm, open-label extension substudy. Objective: Assess potential for standard-of-care background medication reductions while maintaining asthma control with benralizumab. Methods: Following ANDHI DB completion, eligible adults were enrolled in ANDHI IP. After an 8-week run-in with benralizumab, there were 5 visits to potentially reduce background asthma medications for patients achieving and maintaining protocol-defined asthma control with benralizumab. Main outcome measures for non-oral corticosteroid (OCS)-dependent patients were the proportions with at least 1 background medication reduction (ie, lower inhaled corticosteroid dose, background medication discontinuation) and the number of adapted Global Initiative for Asthma (GINA) step reductions at end of treatment (EOT). Main outcomes for OCS-dependent patients were reductions in daily OCS dosage and proportion achieving OCS dosage of 5 mg or lower at EOT. Results: For non-OCS-dependent patients, 53.3% (n = 208 of 390) achieved at least 1 background medication reduction, increasing to 72.6% (n = 130 of 179) for patients who maintained protocol-defined asthma control at EOT. A total of 41.9% (n = 163 of 389) achieved at least 1 adapted GINA step reduction, increasing to 61.8% (n = 110 of 178) for patients with protocol-defined EOT asthma control. At ANDHI IP baseline, OCS dosages were 5 mg or lower for 40.4% (n = 40 of 99) of OCS-dependent patients. Of OCS-dependent patients, 50.5% (n = 50 of 99) eliminated OCS and 74.7% (n = 74 of 99) achieved dosages of 5 mg or lower at EOT. Conclusions: These findings demonstrate benralizumab's ability to improve asthma control, thereby allowing background medication reduction

L-carnitine during in vitro culture enhances the cryotolerance of buffalo (Bubalus bubalis) in vitro derived embryos.

In buffalo, in vitro embryo production (IVEP) technology is the best tool to improve the genetic merit through the maternal lineage. A major limitation of IVEP technology in buffalo species is the poor cryotolerance of the embryos, likely due to their high lipid content (Gasparrini 2002 Theriogenology 57, 237–256). It was previously demonstrated that supplementing bovine culture media with L-carnitine, a cofactor of β-oxidation, improves in vitro embryo development (Sutton-McDowall et al. 2012 Theriogenology 77, 1632–1641). The aim of this work was to evaluate whether L-carnitine supplementation during in vitro culture (IVC) improves blastocyst development and cryotolerance of in vitro produced buffalo embryos. After a preliminary dose response trial, we selected the concentration of 0.25 mM for the experiment. Cumulus–oocytes complexes (n = 288, over 4 replicates), recovered from slaughtered animals, were matured and fertilized in vitro according to our standard procedures (Gasparrini et al. 2006 Theriogenology 65, 275–287). On Day 1 (Day 0 = IVF), zygotes were cultured in SOF supplemented with 8 mg mL–1 BSA, in the absence (control, n = 143) or presence of 0.25 mM L-carnitine (n = 145). In vitro culture was carried out at 38.5°C under 5% CO2, 7% O2, and 88% N2. Cleavage rate was evaluated on Day 5, when the cleaved embryos were transferred into fresh medium for further 2 days. On Day 7 after IVF, embryo outcome was assessed and all the embryos were vitrified by cryotop in 16.5% ethylene glycol, 16.5% dimethyl sulfoxide (DMSO), and 0.5 M sucrose (De Rosa et al. 2007 Ital. J. Anim. Sci. 6(Suppl 2), 747–750). The resistance to cryopreservation was evaluated by assessing the survival rate, on the basis of morphological criteria, after 24 h culture. Data were analyzed by chi-square test. No differences were found in cleavage rates between the control (81.5%) and the L-carnitine group (78.8%). The blastocyst yields (calculated in relation to the cleaved embryos) were not significantly influenced by the L-carnitine treatment (40.2 and 52.9%, in the control and the L-carnitine groups, respectively). However, buffalo embryos cultured in the presence of L-carnitine showed an increased resistance to cryopreservation, as indicated by the higher survival rates recorded after 24 h culture (78.7 and 96.4%, in the control and the L-carnitine groups, respectively; P < 0.01). In conclusion, these results demonstrated that L-carnitine supplementation of culture medium improves the resistance to cryopreservation of in vitro produced buffalo embryos. We speculate that the increased cryotolerance observed in the presence of L-carnitine may be due to a better utilization of the endogenous lipid stores, resulting in improved embryo quality

Energy source during in vitro culture (IVC) and sex ratio of bovine embryos

Most systems for producing mammalian embryos in vitro use glucose as an energy source despite putative toxic effects. It is known that female embryos are more sensitive to negative effects of glucose during IVC. The aim of this work was to evaluate whether replacing glucose with myo-inositol and citrate during IVC affects sex ratio. Abattoir-derived oocytes were matured and fertilized in vitro using standard procedures. After 20–22 h of gametes co-incubation, zygotes were denuded and cultured in SOF containing either 1.5 mM glucose or 2.77 mM myo-inositol and 0.34 mM citrate, for 7 days. The percentages of blastocysts were recorded and the embryos (on average 122 per group) were sexed by PCR as previously described (Alomar, 2008, Anim. Reprod. Sci. 107 48-61.). Differences in blastocyst rates and in the percentages of female embryos between groups were analyzed by Chi-Square test. The results of this study showed that myo-inositol-citrate increased both blastocyst yield (37.4 vs 29.5 %, respectively; P<0.01) and the percentage of female embryos compared to glucose (61.5 vs 45.6 % respectively; P<0.05). In conclusion, these results suggest to use myo-inositol and citrate in culture media to switch embryo sex ratio towards females

The influence of cumulus cells during in vitro fertilization of buffalo (Bubalus bubalis) denuded oocytes that have undergone vitrification.

The aim of this work was to evaluate whether providing a support of cumulus cells during IVF of buffalo denuded oocytes submitted to vitrification-warming enhances their fertilizing ability. In vitro matured denuded oocytes were vitrified by Cryotop in 20% EG + 20% of DMSO and 0.5 M sucrose and warmed into decreasing concentrations of sucrose (1.25 M-0.3M). Oocytes that survived vitrification were fertilized: 1) in the absence of a somatic support (DOs); 2) in the presence of bovine cumulus cells in suspension (DOs+susp); 3) on a bovine cumulus monolayer (DOs+monol); and 4) with intact bovine COCs in a 1:1 ratio (DOs+COCs). In vitro matured oocytes were fertilized and cultured to the blastocyst stage as a control. An increased cleavage rate was obtained from DOs+COCs (60.9%) compared to DOs, DOs+susp (43.6 and 38.4, respectively; P < 0.01) and DOs+monol (47.5%; P < 0.05). Interestingly, cleavage rate of DOs+COCs was similar to that of fresh control oocytes (67.8%). However, development to blastocysts significantly decreased in all vitrification groups compared to the control (P < 0.01). In conclusion the co-culture with intact COCs during IVF completely restores fertilizing capability of buffalo denuded vitrified oocytes, without improving blastocyst development

Structural changes of in vitro matured buffalo and bovine oocytes following cryopreservation.

The aim of this work was to evaluate chromatin and spindle organization of buffalo and bovine in vitro matured oocytes after vitrification/warming by Cryotop and after their exposure to cryoprotectants (CP). In vitro matured oocytes were vitrified/warmed and exposed to the vitrification/warming solutions containing ethylene glycol (EG), dimethyl sulfoxide (DMSO) and sucrose as CP. Two hours after warming, oocytes were fixed and immunostained for microtubules and nuclei and examined by fluorescence microscopy. Data were analyzed by Chi Square test. A higher percentage of Telophase II stage oocytes was found in the toxicity (26 and 34% in bovine and buffalo) and the vitrification groups (13 and 7% in bovine and buffalo) compared to the control, indicating occurrence of activation. An increased percentage of oocytes with abnormal spindle and chromosome organization was found in oocytes exposed to CP (24 and 13% in bovine; 32 and 30% in buffalo respectively) and in those vitrified (26 and 31% in bovine; 26 and 29% in buffalo respectively) compared to the control (0 in bovine and 2.5% in buffalo)

Structural changes of in vitro matured buffalo and bovine oocytes following cryopreservation

The aim of this work was to evaluate chromatin and spindle organization of buffalo and bovine in vitro matured oocytes after vitrification/warming by Cryotop and after their exposure to cryoprotectants (CP). In vitro matured oocytes were vitrified/warmed and exposed to the vitrification/warming solutions containing ethylene glycol (EG), dimethyl sulfoxide (DMSO) and sucrose as CP. Two hours after warming, oocytes were fixed and immunostained for microtubules and nuclei and examined by fluorescence microscopy. Data were analyzed by Chi Square test. A higher percentage of Telophase II stage oocytes was found in the toxicity (26 and 34% in bovine and buffalo) and the vitrification groups (13 and 7% in bovine and buffalo) compared to the control, indicating occurrence of activation. An increased percentage of oocytes with abnormal spindle and chromosome organization was found in oocytes exposed to CP (24 and 13% in bovine; 32 and 30% in buffalo respectively) and in those vitrified (26 and 31% in bovine; 26 and 29% in buffalo respectively) compared to the control (0 in bovine and 2.5% in buffalo)