Admite-se que mais de 40% das crianças são acometidas pela desnutrição crônica em Moçambique (África Oriental). A doença pode estar relacionada, entre outros fatores, à qualidade da dieta que é oferecida à população, já que é bastante precária, pois exibe sérias deficiências de ferro, gordura e, principalmente, proteína animal em sua composição. Essa insuficiência proteica poderia acarretar em prejuízo ao desenvolvimento do organismo, pois a proteína animal é considerada uma boa fonte de aminoácidos essenciais, em decorrência de sua maior digestibilidade e absorção no intestino delgado, quando comparadas às fontes de origem vegetal. Na presente pesquisa foi reproduzida em laboratório, a dieta básica da população de Moçambique (DM), com o objetivo de avaliar seus efeitos nos componentes do plexo mioentérico e na mucosa dos segmentos do intestino delgado de ratos Wistar. Para isso, os animais foram divididos nos grupos Controle, com dieta AIN-93G com adição de 20% de caseína (NN21 e NN42); Dieta de Moçambique (DM21 e DM42) e Dieta Moçambique suplementada, acrescida de 20% de caseína (NM21 e NM42); e grupo Renutrido (RM42), composto por animais do grupo DM21 que, a partir do 22º dia, receberam a dieta NM até atingirem 42 dias de vida. Os segmentos foram coletados e submetidos às técnicas histoquímicas da NADH-diaforase e da NADPH-diaforase para evidenciação de neurônios do plexo mioentérico; histológicas (HE, Picro-sírius, Weigert) para avaliação da parede intestinal, mucosa, gânglios e seu tecido conjuntivo associado; de microscopia eletrônica de varredura (MEV) para observação da estrutura da mucosa; e de microscopia eletrônica de transmissão (MET) para a ultraestrutura dos componentes ganglionares. Estatisticamente, o peso corporal e o comprimento dos animais submetidos à dieta de Moçambique estavam abaixo dos valores encontrados para os animais controle. Na análise qualitativa, observou-se a presença de fibras elásticas, elaunínicas e oxitalânicas, assim como predominância de fibras colágenas do tipo I nos grupos NN42 e DM42, e do tipo III nos grupos NM42 e RM42 ao redor dos gânglios. A mucosa apresentou uma menor área no grupo DM21 com recuperação em DM42, com diminuição da altura das vilosidades nos dois grupos. Foram observadas alterações na organização do retículo endoplasmático rugoso e disposição dos materiais fibrilares e granulares do nucléolo dos animais DM. Sob MEV as vilosidades do grupo DM42 apresentaram superfície mais lisa, com poucas delimitações entre elas. A densidade dos neurônios reativos à NADH diminuiu de 21 para 42 dias em todos os grupos; porém, o DM21 e DM42 apresentou uma maior densidade. Os neurônios reativos à NADPH apresentaram a diminuição da densidade de 21 para 42 dias nos grupos DM e NM, quando comparados ao controle. Assim, conclui-se que a dieta vegetal de Moçambique levou à alterações na morfologia da mucosa, parede intestinal e neurônios entéricos, como uma forma de adaptação à dieta imposta / It is assumed that more than 40% of children are affected by chronic malnutrition in Mozambique (East Africa). The disease may be related, among other factors, the quality of diet that is offered to the population, since it is quite precarious, because it displays serious deficiencies of iron, fat and especially animal protein in their composition. This protein failure could result in damage to the development of the organism, as animal protein is considered a good source of essential amino acids, due to its higher digestibility and absorption in the small intestine when compared to vegetable sources. In this research has been reproduced in the laboratory, the staple diet of the population of Mozambique (DM), in order to evaluate its effects on components of the myenteric plexus and the mucosa of the small intestine segments of Wistar rats. For this, the animals were divided into control groups with AIN-93G diet with the addition of 20% casein (NN21 and NN42); Diet Mozambique (DM21 and DM42) and diet supplemented Mozambique, plus 20% casein (NM21 and NM42); and Refeeding group (RM42), consisting of the animals DM21 group, from the 22th day, given NM diet until they reached 42 days of life. The segments were collected and submitted to histochemical techniques of NADH-diaphorase and NADPH-diaphorase for disclosure of neurons of the myenteric plexus; histologic (HE, Sirius red, Weigert) for evaluation of the intestinal wall, mucosa, lymph nodes and its associated connective tissue; scanning electron microscopy (SEM) for observation of mucosal structure; and Transmission electron microscopy (TEM) ultrastructure to ganglion components. Statistically, body weight and length of the animals submitted to Mozambique diet were below the values found for control animals. Qualitative analysis showed the presence of elastic fibers, and elauninic oxytalan, and predominance of type I collagen fibers in the NN42 and DM42 groups, and type III in the NM42 and RM42 groups around the ganglia. The mucosa showed a smaller area in DM21 group recovery DM42, with a decrease in villus height in both groups. There have been changes in the organization of the rough endoplasmic reticulum and arrangement of fibrillar and granular materials nucleolus of DM animals. Under SEM the villi of the DM42 group showed smoother surface, with few boundaries between them. The density of reactive NADH decreased from 21 to 42 days in all groups; however, the DM21 and DM42 had a higher density. Reactive neurons to NADPH had decreased from 21 to 42 days density in DM and NM groups when compared to the control. Thus, it is concluded that vegetable diet Mozambique led to changes in the morphology of the mucosa, the intestinal wall and enteric neurons, as a way to adapt to the imposed diet

Orientador: José Andrés Yunes / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-20T16:36:11Z (GMT). No. of bitstreams: 1 Conte_AnaLuizaOngaroSeidinger_D.pdf: 7461860 bytes, checksum: a44751dc1556a30853dcde70f024aed9 (MD5) Previous issue date: 2012 / Resumo: O gene supressor de tumor TP53 é um dos mais bem caracterizados genes cujo papel na etiologia é conhecido tanto em tumores hereditários como esporádicos. Mutações germinativas neste gene normalmente estão associadas à Síndrome de Li-Fraumeni, uma síndrome de predisposição ao câncer cujos portadores apresentam alto risco de desenvolvimento de uma variedade de tumores ao longo da vida. Uma mutação específica deste gene é altamente prevalente na população brasileira, chegando a estar presente em 0.3% da população saudável. Esta mutação, a R337H, não foi descrita como uma mutação associada a um amplo espectro tumoral e, sim, como uma mutação tecido-específica associada somente ao desenvolvimento do tumor do córtex da adrenal (TCA). Entretanto, algumas evidências indicavam que ela poderia contribuir para a etiologia de outros tumores, além do TCA. A R337H foi encontrada em 06 famílias com Síndrome de Li- Fraumeni like e em 2.4% de pacientes diagnosticadas com câncer de mama. Com o objetivo de obter maiores evidências sobre a contribuição da R337H na etiologia de outros tumores além do TCA, investigamos a prevalência desta mutação em 493 pacientes diagnosticados com diferentes tumores pediátricos. Além disto, buscamos investigar alterações cromossômicas que poderiam estar associadas ao processo de tumorigênese em portadores desta mutação. Os resultados mostraram que além do TCA e câncer de mama, a R337H está fortemente associada ao carcinoma de plexo coroide (69%) e ao osteossarcoma (7%) na faixa etária pediátrica. Em nossa instituição, foi observado um aumento da incidência relativa do carcinoma de plexo coroide em relação ao papiloma de plexo coroide. Uma deleção do gene TP53 em células do sangue periférico sob a forma de um mosaico foi identificada nos pacientes portadores da R337H que desenvolveram o TCA. Os dados mostraram que se trata de uma deleção intersticial do cromossomo 17, a qual especula-se ter ocorrido durante o período embrionário. Os resultados sobre a prevalência desta mutação em diferentes tumores pediátricos poderão contribuir para um adequado aconselhamento genético das famílias em que a R337H está presente. Adicionalmente, o mosaicismo encontrado nos portadores da R337H que desenvolveram o TCA nos permitiu formular uma hipótese sobre a tumorigênese adrenocortical associada à R337H e talvez possa também contribuir para a futura compreensão do comportamento oncogênico desta mutação / Abstract: The tumor suppressor gene TP53 is a well characterized gene whose role in the etiology of hereditary and sporadic cancer is well established. Germline mutations in this gene are usually associated with the Li-Fraumeni Syndrome (LFS), a cancer predisposition syndrome in which individuals are at a high risk to develop a wide spectrum of tumors during their lives. A specific mutation of TP53 gene is highly prevalent among the Brazilian population, reaching 0.3% of healthy individuals. This mutation, R337H, was not described as a mutation associated to a wide spectrum of tumors but as a tissue-specific mutation, predisposing individuals only to adrenocortical tumors (ACT). However, some evidences have indicated that it could contribute to the etiology of other tumors besides ACT. The R337H was found in six families diagnosed with LFS-like and also in 2.4% of patients diagnosed with breast cancer. In order to obtain more evidence regarding the role of R337H in the etiology of tumors besides ACT, we have investigated the prevalence of this mutation among 493 pediatric patients diagnosed with cancer. Moreover, we have looked for chromosomal changes potentially involved in tumorigenic process among R337H carriers. Our results demonstrated that, besides ACT and breast cancer, the R337H mutation is strongly associated to choroid plexus carcinoma (69%) and osteosarcoma (7%) in the pediatric age. At our institution it was observed a raise in the relative incidence of choroid plexus carcinoma to choroid plexus papilloma. It was identified a mosaic of cells presenting TP53 deletion in peripheral blood of R337H carriers who developed ACT. Our data showed that is an interstitial deletion affecting the short arm of chromosome 17 that may have occurred during the embryonic life. The results regarding the R337H prevalence among different pediatric tumors may potentially contribute to a more accurate genetic counseling of families bearing this mutation. Additionally, the TP53 deletion found in R337H carriers who developed ACT prompted us to formulate a hypothesis on the adrenocortical tumorigenesis associated to this mutation. Finally, our findings may be useful to better understand the oncogenic behavior of this mutation / Doutorado / Genetica Animal e Evolução / Doutor em Genetica e Biologia Molecular

Abstract Collagen XVIII is a multidomain basement membrane proteoglycan with three tissue-specific isoforms. Endostatin, the C-terminal part of collagen XVIII, has antiangiogenic properties, while the frizzled-like domain of the longest isoform is suggested to be capable of inhibiting the Wnt/β-catenin signaling network. This study utilized several genetically modified mouse lines and electron microscopy to achieve new information on the biological role of collagen XVIII, its different isoforms, and the frizzled domain. Lack of collagen XVIII was found to affect the integrity of basement membranes of various tissues, leading to an abnormally loosened network structure. In the choroid plexus, the change in the basement membrane ultrastructure caused alterations in the production of the cerebrospinal fluid and predisposed to the development of hydrocephalus. In the kidney, broadening of the proximal tubular basement membrane was shown to be due specifically to the lack of the short isoform, while the lack of the two longer isoforms led to podocyte foot process effacement. Moreover, lack of collagen XVIII was found to cause softening of the kidney glomeruli and the levels of serum creatinine were elevated in the mutant animals, indicating altered kidney function. The hair follicle cycle was used as a model to study the possible role of the frizzled domain of collagen XVIII in the Wnt/β-catenin signaling cascade. The longer collagen XVIII isoforms were shown to be expressed in the basement membrane facing the dermal papilla and in the hair follicle bulge, containing the follicular stem cells. Lack of the long isoforms led to abnormalities in the progression of the first hair cycle, and the phenotype could be rescued via transgenic delivery of the frizzled domain of the longest isoform, suggesting its involvement in the regulation of the Wnt/β-catening signaling network during the cyclic growth of the hair. / Tiivistelmä Kollageeni XVIII on useista toiminnallisista osista koostuva tyvikalvojen proteoglykaani, jolla on kolme eri kudoksissa esiintyvää isomuotoa. Sen C-terminaalisella endostatiini-osalla on verisuonten kasvua estäviä vaikutuksia, kun taas pisimmän isomuodon frizzled-osan uskotaan estävän Wnt/β-kateniini signalointireitin toimintaa. Tässä tutkimuksessa saatiin uutta tietoa kollageeni XVIII:n, sen eri isomuotojen sekä frizzled-osan biologisesta merkityksestä useiden geenimuunneltujen hiirimallien sekä elektronimikroskopian avulla. Kollageeni XVIII:n puutoksen todettiin vaikuttavan tyvikalvojen rakenteen eheyteen useissa eri kudoksisssa, johtaen epänormaalisti löyhtyvään verkkorakenteeseen. Suonipunoksessa tämä tyvikalvon hienorakenteen muutos vaikutti aivo-selkäydinnesteen tuottumiseen ja altisti vesipään kehittymiselle. Munuaisessa proksimaalisen munuaistiehyen tyvikalvon levenemisen osoitettiin johtuvan lyhyen isomuodon puutoksesta, kun taas kahden pidemmän isomuodon puuttuminen aiheutti podosyyttien jalkalisäkkeiden leviämistä. Lisäksi kollageeni XVIII:n puuttumisen osoitettiin johtavan hiirimallien munuaiskerästen pehmenemiseen sekä veren kreatiniinitason kohoamiseen, viitaten munuaistoiminnan häiriöihin. Karvatuppien syklistä kasvua käytettiin mallina tutkittaessa kollageeni XVIII:n frizzled-osan mahdollisia vaikutuksia Wnt/β-kateniini signalointireittiin. Pidempien kollageeni XVIII isomuotojen osoitettiin tuottuvan karvanystyn tyvikalvossa sekä karvatupin kantasolut sisältävällä pullistuma-alueella. Pitkien isomuotojen puuttuminen johti karvojen ensimmäisen kasvukierron epänormaaliin etenemiseen. Tämä voitiin estää siirtogeenisen frizzled-osan avulla, mikä viittasi sen osallisuuteen Wnt/β-kateniini signalointireitin säätelyyn karvan syklisen kasvun aikana.

Bakgrund: Distal radiusfraktur är den vanligaste frakturen i Sverige där kvinnor drab-bas oftare än män. Majoriteten av de patienter som opereras vårdas på en dagkirurgisk enhet och sköter sin egenvård i hemmet. Kvinnor upplever smärta annorlunda än män vilket kan ha betydelse i valet av smärtlindringsstrategi. Inför operationen erhåller patienten antingen en plexusblockad eller generell anestesi med infiltrationsanestesi i smärtlindrande syfte. I anestesisjuksköterskans kompetensbeskrivning ingår det att ansvara för planering och övervakning vid dessa olika anestesimetoder. Vilken av de två anestesimetoder som används varierar på sjukhusen runt om i Sverige. Det finns ett begränsat forskningsunderlag på vilken anestesimetod som är att föredra. Syfte: Syftet var att undersöka smärtupplevelsen hos kvinnor efter dagkirurgisk operation av distal radiusfraktur. En jämförelse mellan anestesimetoderna plexusblockad och infiltrationsanestesi med Ropivacain. Metod: En kvantitativ enkätundersökning. Data har samlats in genom ett bekvämlighetsurval där enkäten bestående av strukturerade frågor har delats ut av personal på sju postoperativa avdelningar. Resultat: Inga statistiskt signifikanta skillnader har hittats mellan grupperna som erhållit antingen plexusblockad eller infiltrationsanestesi med Ropivacain. Dock visade sig en betydande andel uppleva svår eller värsta tänkbara smärta kvällen eller natten efter operation. Båda grupperna uppvisar även problem med sömn och vila postoperativt. Slutsatser: Resultatet från magisteruppsatsen antyder att det finns förbättringspotential postoperativt för kvinnor med radiusfraktur som upplever smärta. Dessutom bör grundläggande behov, särskilt vila, sömn och aptit, tas i beaktande. / Background: Distal radius fracture is the most common fracture in Sweden and women are affected more often than men. The majority of the patients who require surgery are cared for in a day surgery unit. They are then responsible for their own care at home. Women experience pain differently than men and this can be important when deciding upon a pain relief strategy. Prior to surgery, the patient receives either a plexus blockade or general anesthesia with infiltration anesthesia for pain relief purposes. The nurse anesthetist´s (CRNA) competence description includes being responsible for planning and monitoring of these different anesthesia methods. The choice of anesthesia method used varies in hospitals around Sweden. There is a limited amount of research on which anesthesia method is preferable. Method: A quantitative survey. Data has been collected using convenience sampling where a questionnaire consisting of structured questions was distributed by hospital staff in seven postoperative departments. Results: No statistically significant differences were found between the groups that received either plexus block or ropivacaine infiltration anesthesia. However, a significant proportion were found to experience either severe pain or the worst possible pain in the evening or night after surgery. Both groups show problems with sleep and rest postoperatively, while the infiltration group tends to be more inconvenienced by decreased appetite than the patients in the plexus group. Conclusions: The results of the master's thesis suggest that there is potential for improvement for postoperative female patients with a radius fracture experiencing pain. Additionally basic needs, in particular rest, sleep and appetite, should be taken into account.

Perivascular cells of microvascular niches are the prime candidates for being a reservoire of mesenchymal stem cell (MSC)-like cells in many tissues and organs that could serve as a potential source of cells and a target of novel cell-based therapeutic approaches. In the present study, by utilising typical markers of pericytes (neuronal-glial antigen 2, NG2, a chondroitin sulphate proteoglycan) and those of MSCs (CD146 and CD105) and primitive pluripotent cells (sex-determining region Y-box 2, Sox2), the phenotypic traits and the distribution of murine and rat retinal perivascular cells were investigated in situ. Our findings indicate that retinal microvessels of juvenile rodents are highly covered by NG2-positive branching processes of pericytic (perivascular) cells that are less prominent in mature capillary networks of the adult retina. In the adult rodent retinal vascular bed, NG2 labeling is mainly confined to membranes of the cell body resulting in a pearl-chain-like distribution along the vessels. Retinal pericytes, which were identified by their morphology and NG2 expression, simultaneously express CD146. Furthermore, CD146-positive cells located at small arteriole-tocapillary branching points appear more intensely stained than elsewhere. Evidence for a differential expression of the two markers around capillaries that would hint at a clonal heterogeneity among pericytic cells, however, is lacking. In contrast, the expression of CD105 is exclusively restricted to vascular endothelial cells and Sox2 is detected neither in perivascular nor in endothelial cells. In dissociated retinal cultures, however, simultaneous expression of NG2 and CD105 was observed. Collectively, our data indicate that vascular wall resident retinal pericytes share some phenotypic features (i.e. CD146 expression) with archetypal MSCs, which is even more striking in dissociated retinal cultures (i.e. CD105 expression). These findings might have implications for the treatment of retinal pathologies.

Streptococcus suis (S. suis) is an important opportunistic pathogen, which can cause septicemia and meningitis in pigs and humans. Previous in vivo observations in S. suisinfected pigs revealed lesions at the choroid plexus (CP). In vitro experiments with primary porcine CP epithelial cells (PCPEC) and human CP epithelial papilloma (HIBCPP) cells demonstrated that S. suis can invade and traverse the CP epithelium, and that the CP contributes to the inflammatory response via cytokine expression. Here, next generation sequencing (RNA-seq) was used to compare global transcriptome profiles of PCPEC and HIBCPP cells challenged with S. suis serotype (ST) 2 infected in vitro, and of pigs infected in vivo. Identified differentially expressed genes (DEGs) were, amongst others, involved in inflammatory responses and hypoxia. The RNA-seq data were validated via quantitative PCR of selected DEGs. Employing Gene Set Enrichment Analysis (GSEA), 18, 28, and 21 enriched hallmark gene sets (GSs) were identified for infected HIBCPP cells, PCPEC, and in the CP of pigs suffering from S. suis ST2 meningitis, respectively, of which eight GSs overlapped between the three different sample sets. The majority of these GSs are involved in cellular signaling and pathways, immune response, and development, including inflammatory response and hypoxia. In contrast, suppressed GSs observed during in vitro and in vivo S. suis ST2 infections included those, which were involved in cellular proliferation and metabolic processes. This study suggests that similar cellular processes occur in infected human and porcine CP epithelial cells, especially in terms of inflammatory response.

T cells infiltrate the degenerating brain and influence central nervous system (CNS) inflammation and neuronal health. In mice, the choroid plexus and the meninges have been implicated in regulating T cell entry and egress from the CNS, respectively. Further, antigen presenting cells in the mouse meninges present CNS-derived antigens to T cells and may represent a method for the peripheral immune system to sense and respond to CNS immune triggers. However, whether these processes occur in the human choroid plexus and meninges has not been comprehensively studied. Further, the antigens towards which T cells in the degenerating human brain and its borders respond remain unknown. Therefore, I implemented a multi-omics approach using fresh postmortem tissue from patients diagnosed with amyotrophic lateral sclerosis (ALS), Alzheimer’s disease (AD), Parkinson’s disease (PD), and non-neurodegenerative controls to identify not only the T cell-associated changes that occur in the degenerating human CNS and surrounding tissues but also identified a library of putative antigen targets for disease-associated T cell populations. Specifically, using single cell RNA and TCR sequencing information from paired postmortem choroid plexus, leptomeninges, and brain I lineage traced T cells using their TCR information and found that T cell access to leptomeninges and brain is likely limited and controlled by anti-inflammatory macrophage activity at the blood/CSF barrier (BCSFB). Once past the BCSFB, I present evidence that T cells access the CNS where they interact with MHC expressed by microglia. T cells also accumulate in the leptomeninges where they become tissue resident memory T cells. These tissue resident memory T cells likely serve as a reservoir for a rapid antigen-driven immune response to future CNS inflammatory insults. Finally, by performing immunopeptidomics to identify peptides presented by MHC in the same patients’ CNS and border tissues, I identified a library of putative antigenic triggers that may drive high levels of T cell clonal expansion in the brain and surrounding tissues. Altogether, this thesis serves as a resource for understanding the trajectory of T cells as they travel into the degenerating human brain and as a foundation for the development of antigen-specific precision medicines to treat neurodegeneration.

Au cours des vingt dernières années, l’anesthésie régionale est devenue, autant en médecine vétérinaire qu’humaine, un outil essentiel à l’élaboration de protocoles analgésiques péri-opératoires. Parmi l’éventail de techniques mises au point en anesthésie canine, le bloc paravertébral du plexus vertébral (PBPB) et sa version modifiée sont d’un grand intérêt pour toute procédure du membre thoracique, dans sa portion proximale. Toutefois, l’essentiel des données publiées à ce jour provient d’études colorimétriques, sans évaluation clinique, et peu d’information est disponible sur les techniques de localisation nerveuse envisageables à ce site. Notre étude visait à décrire une approche échoguidée du PBPB modifié, puis à caractériser ses paramètres pharmacocinétiques et pharmacodynamiques après administration de lidocaïne (LI) ou lidocaïne adrénalinée (LA). Huit chiens ont été inclus dans un protocole prospectif, randomisé, en aveugle et croisé, réparti sur trois périodes. L’impact pharmacodynamique du bloc effectué avec LI ou LA a été évalué régulièrement pour 180 min suivant son exécution. Le traitement à l’adrénaline n’a pas démontré d’impact significatif (P = 0,845) sur la durée du bloc sensitif, tel qu’évalué par un stimulus douloureux mécanique appliqué aux dermatomes ciblés. À l’opposé, l’atteinte proprioceptive évaluée par la démarche a été trouvée prolongée (P = 0,027) et le bloc moteur mesuré par le pic de force verticale (PVF) au trot sur la plaque de force s’est avéré plus marqué (PVF réduit; P = 0,007) sous LA. À l’arrêt comme au trot, le nadir de la courbe PVF-temps a été trouvé retardé (P < 0,005) et la pente ascendante de retour aux valeurs normales adoucie (P = 0,005). Parallèlement aux évaluations cliniques, des échantillons plasmatiques ont été collectés régulièrement afin de quantifier et décrire le devenir pharmacocinétique de la lidocaïne. Parmi les trois élaborés, un modèle bi-compartimental doté d’une double absorption asynchrone d’ordre zéro a finalement été sélectionné et appliqué aux données expérimentales. Sous LA, la Cmax a été trouvée significativement diminuée (P < 0,001), les phases d’absorption prolongées [P < 0,020 (Dur1) et P < 0,001 (Dur2)] et leurs constantes réduites [P = 0,046(k01) et P < 0,001 (k02)], le tout en concordance avec les effets proprioceptifs et moteurs rapportés. Bien que l’extrapolation du dosage soit maintenant théoriquement envisageable à partir du modèle mis en lumière ici, des études supplémentaires sont encore nécessaires afin d’établir un protocole de PBPB d’intérêt clinique. L’analyse sur plaque de force pourrait alors devenir un outil de choix pour évaluer l’efficacité du bloc dans un cadre expérimental. / Over the last decade, regional anaesthesia has become a gold standard for peri-surgical management in veterinary medicine. Among the many techniques developed for analgesia in dogs, the paravertebral brachial plexus block (PBPB) is of great interest when targeting the proximal half of the thoracic limb. Yet, most available data on this technique is based on colorimetric protocols rather than clinical evaluation, and there are very few published results for PBPB execution using nerve location techniques. Through this work, we wished to describe an ultrasound-guided approach of the PBPB and characterize its pharmacokinetic/ pharmacodynamic parameters when executed with either lidocaine alone (LI) or combined to adrenaline (LA). Eight dogs were included in a prospective, randomised, blinded, crossover protocol performed over three distinct periods. Pharmacodynamic impact of LI and LA was compared for 180 minutes after block administration. No significant difference (P = 0.845) was noted between treatments regarding length of the sensitive block, as evaluated regularly through a mechanical painful stimulus applied to selected dermatomes. On the opposite, gait examination showed a longer proprioceptive deficit using LA (P = 0.027). Motor block measured with dynamic force plate analysis showed a lower peak vertical force with LA than LI (P = 0.007). For both dynamic and static evaluations, nadir was clearly delayed (P < 0.005) and the ascending slope back to baseline significantly softened (P = 0.005) in the LA group. Throughout block execution and evaluation, blood samples were collected regularly in order to quantify and describe lidocaine kinetics. Models where developed and compared. A two-compartment model with dual zero-order absorption processes was selected as the best fit for our experimental data. Cmax proved to be significantly reduced with LA (P < 0.001), thus reducing potential toxicity. Absorption phase was prolonged [P < 0.020 (Dur1) and P < 0.001 (Dur2)] and zero-order absorption constant rates lowered [P = 0.046(k01) and P < 0.001 (k02)] following adrenaline addition, in accordance with the previously noted prolonged gait and motor effects. Though dosage extrapolation is now possible using the model developed and tested here, further studies would be needed to establish a PBPB protocol of more clinical interest. Then, force plate analysis could become a key tool for block quality assessment, as both dynamic and static measurements proved to be the reliable ways to collect ground reaction force (GRF) data.

Introduction : L’utilisation des méthodes anatomiques classiques rend difficile la localisationprécise des micro-fibres nerveuses et ne permet pas de déterminer leur nature ni leur fonction.La chirurgie pelvienne est associée à des séquelles urinaires et sexuelles fréquentes causéespar lésion iatrogène des nerfs pelviens. La connaissance de l’anatomie et de la physiologie del’innervation intra-pelvienne est fondamentale pour tenter de réduire le taux de troublesfonctionnels postopératoires.Objectifs : i) étudier l’anatomie topographique et la nature de fibres nerveuses intrapelvienneen utilisant la reconstruction tridimensionnelle des coupes histologiques immunomarquéesii) standardiser la technique de Dissection Anatomique Assistée par Ordinateur(DAAO) et vérifier sa faisabilité sur des sujets adultes et iii) établir grâce à cette technique deDAAO des modèles pédagogiques tridimensionnels afin d’améliorer la compréhension desdysfonctions sphinctériennes et sexuelles survenant après la chirurgie (rectale et prostatique)et d’adapter éventuellement la technique opératoire.Matériel et méthodes : Des coupes histologiques sériées de pelvis ont été réalisées chez septfoetus masculins, sept foetus féminins et six cadavres adultes masculins. Les coupes ont ététraitées par des méthodes histologiques (Hématoxyline-Eosine et trichrome de Masson) etimmuno-histochimiques pour détecter les fibres nerveuses (anti-S100), les fibres somatiques(anti-PMP22), les fibres adrénergiques (anti-TH), cholinergiques (anti-VAChT), sensitives(anti-SP/CGRP) et nitrergiques (anti-nNOS) ainsi que l’actine lisse des sphincters. Les lamesont ensuite été numérisées par un scanner de haute résolution optique et les imagesbidimensionnelles ont été reconstruites en trois dimensions grâce au logiciel WinSurf.Résultats: La reconstruction tri-dimensionnelle des coupes histologiques immuno-marquéesa permis d'identifier l’anatomie topographique et structurelle de l’innervation intra-pelvienne.Les structures nerveuses afférentes du plexus hypogastrique inférieur (PHI) : nerfssplanchniques pelviens (NSP) et nerfs hypogastriques (NH) véhiculent de façon mixte l’influxsympathique et parasympathique pelvien.Les fibres nerveuses issues de la partie distale du PHI sont responsables de la continenceurinaire et de la fonction sexuelle. Elles sont regroupées et associées au pédicule vasculairepour former la bandelette neuro-vasculaire (BNV). Trois efférences principales sont issues decette BNV : i) des fibres antérieures destinées au sphincter urétral, ii) des fibres antérolatérales,par rapport à la prostate et au vagin, constituant le nerf caverneux destiné aux corpscaverneux du pénis/clitoris et iii) des fibres nerveuses postéro-latérales, par rapport à laprostate et au vagin, formant le « nerf spongieux » destiné aux corps spongieux/bulbesvestibulaires.Les communications autonomiques-somatiques entre le PHI supra-lévatorien et le nerfpudendal infra-lévatorien existent à trois niveaux ; proximal, intermédiaire et distal. Lacommunication distale caverno-pudendale est responsable de l’activité érectile segmentaireobservée au sein du nerf dorsal du pénis/clitoris.Conclusion: La DAAO est une méthode originale de recherche anatomique qui a étéprogressivement améliorée au sein de notre unité de recherche. Cette évolution illustre le faitque l'anatomie descriptive est encore une science dynamique. Notre étude a permis dedévelopper des modèles anatomo-physiologiques d’innervation pelvienne contribuant àaméliorer du point de vue morphologique, chirurgical et pédagogique la compréhension derégions anatomiques complexes comme le petit bassin. / Introduction: Classic anatomical methods have limitations in micro determination of nervefibre location. Furthermore, the precise detection of the nerve fibres nature is not possible bymeans of dissection. Pelvic surgery is associated with urinary and sexual consequence causedby iatrogenic damage of the pelvic nerves. Anatomic and physiologic knowledge of the intrapelvicinnervation is essential to reduce the rate of postoperative functional complication.Objectives: i) to study the topographic anatomy and the nature of intra-pelvic nerve fibersusing three-dimensional reconstruction of histological immuno-labeled sections ii)standardize the technique of Computer-Assisted Anatomic Dissection (CAAD) and check itsfeasibility on adult subjects iii) reconstructe three dimensional teaching models to improveunderstanding of urinary and sexual dysfunction occurring after surgery (of rectal and ofprostate) to ameliorate the operative technique.Materials and methods: serial histological sections of pelvic portion were performed inseven male foetuses, seven female foetuses and six adult male cadavers. The sections weretraited by histological methods (Hematoxylin-Eosin and Masson's trichrome) and immunohistochemicalmarker of the nerve fibers (anti-S100), the somatic fibers (anti-PMP22), theadrenergic (anti-TH), cholinergic (anti-VAChT), sensory (anti-SP/CGRP) and nitrergic (antinNOS)fibers and the actin smooth muscles. The slides were then digitized by a scanner ofhigh optical resolution and two-dimensional images were reconstructed in three dimensionsusing WinSurf software.Results: The three dimensional reconstruction of histological immuno-labelled sectionsidentified structural and topographic anatomy of intra-pelvic innervation. The afferencenerves of the inferior hypogastric plexus (IHP): pelvic splanchnic nerves (PSN) andhypogastric nerves (HN) contain both sympathetic and parasympathetic fibers.The nerve fibers from the distal part of the IHP is responsible for urinary continence andsexual function. They are grouped and associated with the vascular pedicle to form theneurovascular bundles (NVB). Efferences of this NVB are distributed in three maindirections: i) anterior fibers the urethral sphincter, ii) anterolateral fibers, compared withprostate / vagina, which constitute the cavernous nerve for the corpora cavernosa of the penis/ clitoris and iii) " spongious nerve”, which is the continuity of posterior-lateral nerve fibers,compared with prostate / vagina, for the corpus spongiosum / vestibular bulbs.Autonomic-somatic communications between supra-lavator IHP and infra-levator pudendalnerve are present at three levels; proximal, intermediate and distal communications. Cavernopudendaldistal communication provides segmental erectile activity of the dorsal nerve of thepenis / clitoris.Conclusion: The CAAD is an original method in anatomical research which has beenprogressively improved. This illustrates the fact that descriptive anatomy is still a dynamicscience. Our study has developed anatomico-physiological models of intra-pelvic innervationcontributing to a better understanding of complex anatomical

As lesões térmicas corporais (LTC) são um sério problema de saúde, atingindo principalmente crianças. A extensão e a profundidade da lesão são fatores que alteram várias estruturas. Alterações gastrintestinais também são relatadas, sendo a principal delas, a atrofia das mucosas, provocando ulcerações e a perda da barreira seletiva. Na presente pesquisa avaliou-se através de técnicas histoquímicas, imunohitoquimicas e de microscopia de luz, as alterações ocorridas nos componentes do plexo mioentérico e na espessura da mucosa do jejuno em três porções: oral (O), média (M) e aboral (A), de ratos adultos com 30% da superfície corpórea exposta ao escaldamento, 4 dias (q4) e 10 dias (q10) após a LTC. Verificou-se em q10 o não restabelecimento da massa corpórea, a diminuição da área do jejuno, bem como espessura de sua mucosa. No plexo mioentérico, a área média do perfil celular dos neurônios NADPH não variou, tendo estes uma menor densidade em q10, sendo estes corpos altamente reativos em q4 e q10. Varicosidades grandes destacaram-se em q4 e q10, quando pela SP e VIP. / Burn is a determinant factor to alter body structures as the striated muscle. It also determines gastrintestinal mucosal atrophy what produce loss of selective barrier. With histochemical, immunohistochemical and light microscopy methods the myenteric plexus (MP) of the jejunum was evaluated in rats submitted to burn injury. The scalding was performed in 30% of the body surface. The MP and the mucosa of the oral (O), middle (M) and aboral (A) parts of the jejunum were analyzed four (q4) and ten (q10) days post-lesion. The loss of weight due the burn is not recovered in q10 where the jejunal surface area and the thickness of the mucosa decreased. The neuronal profile of nitregic neurons was similar in q4, q10. The density of nitregic neurons was lower in q10 showing that the time post injury is an important factor able to alter this parameter. The q4 and q10 groups exhibited neuronal bodies highly reactive to NADPH. The immunoreactivity to SP and VIP in q4 and q10 was expressed mainly in large varicosities.