Introdução: Ainda que o uso de Sondas Nasoentéricas (SNE) pareça ser frequente em hospitais, são raros os estudos dimensionando a prevalência do procedimento e o tempo despendido entre indicação e seu uso, especialmente em Emergências. Nestes espaços assistenciais, os atrasos podem contribuir para a manutenção da superlotação e, também, configurarem-se como incidentes no processo assistencial. Objetivo: Conhecer o tempo entre a indicação da SNE e seu efetivo uso e entre as etapas intermediárias desse processo, além de dos fatores associados a atrasos. Método: Entre abril e junho de 2015, foram incluídos adultos admitidos em uma Emergência de um hospital universitário brasileiro em uma coorte prospectiva. Os pacientes foram acompanhados do momento da indicação ao efetivo uso da SNE, mesurando-se os tempos demandados em cada etapa e os possíveis fatores associados ao tempo despendido até a utilização da sonda (variáveis relacionadas à clínica do paciente ou ao processo de trabalho). Para avaliar os fatores associados a ao tempo despendido, adotou-se o modelo de Equações de Estimações Generalizadas (Generalized Estimating Equations – GEE), respeitando-se a distribuição Gamma da variável diferença de tempo. Para a comparação intrafatores foi adotado o teste de Bonferroni como teste Post Hoc. Resultados: Houve 150 inserções de SNE em 115 pacientes. A mediana de tempo entre a indicação e o uso da sonda foi 573 (IQR: 360 - 1093; mínimo: 63 – máximo: 3120) minutos. Analisando-se as diferentes subetapas do processo, foram identificados como fatores independentes de risco para maior tempo entre a indicação e o uso da SNE: inserção de SNE em paciente que não a utilizava antes do atendimento na Emergência, médico não entregar a prescrição de inserção de sonda ao enfermeiro, inserção de SNE na jornada de trabalho noturna, alocação do paciente na sala de cuidados intensivos, retardos na rotina médica, da nutrição e de enfermagem, uso de ventilação mecânica, noradrenalina e necessidade de jejum. Conclusão: O tempo entre a indicação e o uso de sonda nasoentérica foi elevado, excedendo 10 horas em metade dos casos. Fatores relacionados às condições clínicas dos pacientes e à gestão da assistência contribuiriam para o aumento neste tempo. / Background: Although the use of Nasoenteric tubes (NET) appears to be common in hospitals, few studies measure the prevalence of the procedure and the time between the indication and its use, especially in Emergencie Departement. In the Emergency Departament, delays may contribute to the maintenance of overcrowding and also being configured as incidents in the care process. Objective: Knowing the time spent between the indication of the NET and its effective use, as well as the times and factors associated to delays in sub-steps of process. Method: Between April and June 2015, patients adults admitted to an Emergency Departament of a Brazilian university hospital were included in a prospective Cohort. Patients were followed-up from the NET indication until its effective use. The spent time in each sub-step and the possible associated factors with the time taken to use the NET (variables related to the patient's clinical or work process) were analised. To evaluate the associated factors with time spent, the Generalized Estimating Equations model was adopted, respecting the Gamma distribution of variable "time difference". For intrafatores comparison was adopted Bonferroni test as a post hoc test. Results: There were 150 insertions NET in 115 patients. The median time between the NET indication and its use was 573 (IQR: 360-1093; minimum: 63 - maximum: 3120) minutes. Analyzing the different sub-steps of the process, were identified as independent risk factors for increased time between the NET indication and its use: NET insertion in patients who do not used before the Emergency Departement, doctor does not deliver the NET insert prescription to the nurse, NET insertion in the night working hours, patient allocation in intensive care room, delays in medical, nutrition and nursing routine, patients undergoing mechanical ventilation, norepinephrine and need fasting. Conclusion: The time between the NET indication and its use was high, exceeding 10 hours in half the cases. Factors related to the patients' clinical conditions and the management of the assistance would contribute to the increase at this time lapse. / Introducción: Aunque el usio de Sondas Nasoentericas (SNE) parezca frequenteen Hospitales, son raros los estudios dimensionando el predominio el procedimiento y el tiempo dispensado entre indicación y el uso, especialmente em Emergencias. En estos espacios asistenciales, los atrasos pueden contribuir para el mantenimiento de la super población, tambien se configura como incidentes em el proceso asistencial. Objetivo: Conocer el tiempo entre la indicación de la SNE e su efectivo uso y entrelas etapas intermediarias de este proceso, además de los factores asociados a atrasos. Método: Entre abril y junio de 2015, fueron incluídos adultos admitidos em uma emergência de um hospital universitário brasileño em uma Cohorte prospectiva. Los pacientes fueron acompanados del momento de la indicacional efectivo uso de la SNE, mesurando los tiempo demandados em cada etapa y los posibles factores asociados al tiempo dispoensado hasta la utilización de sonda (variables relacionadas a la paraclinica del paciente al proceso del trabajo). Para evaluar los factores asociados al tiempo dispensado, se adopto el modelo de Ecuaciones de Estimativos Generalizadas (Genneralized Estimating Equations – GEE), respetando la distribución Gamma de la variable diferencia del tiempo. Para la comparación intra factores fue adoptado la tesis de Bonferroni como test Post Hoc. Resultados: hubo 150 inserciones de SNE em 115 pacientes. A mediano tiempo entre la indicación y el uso de la sonda fueron 573 (IQR:360 – 1093; mínimo:63 – máximo: 3120) minutos. Se analizan lãs diferentes sub etapas del proceso, fueron identificados como factores independientes de riesgo para mayor tiempo entre la indicacion y el uso de la SNE: ionserción de SNE em paciente que no la utilizaba antes de la atencion em la Emergencia, médico no entregar la prescripción de inserción de sonda al enfermero, inserción de SNE em la jornada de trabajo nocturno, transferência del paciente em la sala de cuidados intensivos, retardos em la rutina medica, del la nutricion y de Enfermería, uso de ventilación mecânica, noradrenalina y la necesidad de la ayuna. Conclusión: El tiempo ente la indicación y el uso de la sonda nasoenterica fue elevado, exediendose 10 horas em la mitad de los casos. Factores relacionados a lãs condiciones clinicas de los pacientes y de la gestion de la Asistencia contribuyeron para el aumento en este tiempo.

Introdução: Ainda que o uso de Sondas Nasoentéricas (SNE) pareça ser frequente em hospitais, são raros os estudos dimensionando a prevalência do procedimento e o tempo despendido entre indicação e seu uso, especialmente em Emergências. Nestes espaços assistenciais, os atrasos podem contribuir para a manutenção da superlotação e, também, configurarem-se como incidentes no processo assistencial. Objetivo: Conhecer o tempo entre a indicação da SNE e seu efetivo uso e entre as etapas intermediárias desse processo, além de dos fatores associados a atrasos. Método: Entre abril e junho de 2015, foram incluídos adultos admitidos em uma Emergência de um hospital universitário brasileiro em uma coorte prospectiva. Os pacientes foram acompanhados do momento da indicação ao efetivo uso da SNE, mesurando-se os tempos demandados em cada etapa e os possíveis fatores associados ao tempo despendido até a utilização da sonda (variáveis relacionadas à clínica do paciente ou ao processo de trabalho). Para avaliar os fatores associados a ao tempo despendido, adotou-se o modelo de Equações de Estimações Generalizadas (Generalized Estimating Equations – GEE), respeitando-se a distribuição Gamma da variável diferença de tempo. Para a comparação intrafatores foi adotado o teste de Bonferroni como teste Post Hoc. Resultados: Houve 150 inserções de SNE em 115 pacientes. A mediana de tempo entre a indicação e o uso da sonda foi 573 (IQR: 360 - 1093; mínimo: 63 – máximo: 3120) minutos. Analisando-se as diferentes subetapas do processo, foram identificados como fatores independentes de risco para maior tempo entre a indicação e o uso da SNE: inserção de SNE em paciente que não a utilizava antes do atendimento na Emergência, médico não entregar a prescrição de inserção de sonda ao enfermeiro, inserção de SNE na jornada de trabalho noturna, alocação do paciente na sala de cuidados intensivos, retardos na rotina médica, da nutrição e de enfermagem, uso de ventilação mecânica, noradrenalina e necessidade de jejum. Conclusão: O tempo entre a indicação e o uso de sonda nasoentérica foi elevado, excedendo 10 horas em metade dos casos. Fatores relacionados às condições clínicas dos pacientes e à gestão da assistência contribuiriam para o aumento neste tempo. / Background: Although the use of Nasoenteric tubes (NET) appears to be common in hospitals, few studies measure the prevalence of the procedure and the time between the indication and its use, especially in Emergencie Departement. In the Emergency Departament, delays may contribute to the maintenance of overcrowding and also being configured as incidents in the care process. Objective: Knowing the time spent between the indication of the NET and its effective use, as well as the times and factors associated to delays in sub-steps of process. Method: Between April and June 2015, patients adults admitted to an Emergency Departament of a Brazilian university hospital were included in a prospective Cohort. Patients were followed-up from the NET indication until its effective use. The spent time in each sub-step and the possible associated factors with the time taken to use the NET (variables related to the patient's clinical or work process) were analised. To evaluate the associated factors with time spent, the Generalized Estimating Equations model was adopted, respecting the Gamma distribution of variable "time difference". For intrafatores comparison was adopted Bonferroni test as a post hoc test. Results: There were 150 insertions NET in 115 patients. The median time between the NET indication and its use was 573 (IQR: 360-1093; minimum: 63 - maximum: 3120) minutes. Analyzing the different sub-steps of the process, were identified as independent risk factors for increased time between the NET indication and its use: NET insertion in patients who do not used before the Emergency Departement, doctor does not deliver the NET insert prescription to the nurse, NET insertion in the night working hours, patient allocation in intensive care room, delays in medical, nutrition and nursing routine, patients undergoing mechanical ventilation, norepinephrine and need fasting. Conclusion: The time between the NET indication and its use was high, exceeding 10 hours in half the cases. Factors related to the patients' clinical conditions and the management of the assistance would contribute to the increase at this time lapse. / Introducción: Aunque el usio de Sondas Nasoentericas (SNE) parezca frequenteen Hospitales, son raros los estudios dimensionando el predominio el procedimiento y el tiempo dispensado entre indicación y el uso, especialmente em Emergencias. En estos espacios asistenciales, los atrasos pueden contribuir para el mantenimiento de la super población, tambien se configura como incidentes em el proceso asistencial. Objetivo: Conocer el tiempo entre la indicación de la SNE e su efectivo uso y entrelas etapas intermediarias de este proceso, además de los factores asociados a atrasos. Método: Entre abril y junio de 2015, fueron incluídos adultos admitidos em uma emergência de um hospital universitário brasileño em uma Cohorte prospectiva. Los pacientes fueron acompanados del momento de la indicacional efectivo uso de la SNE, mesurando los tiempo demandados em cada etapa y los posibles factores asociados al tiempo dispoensado hasta la utilización de sonda (variables relacionadas a la paraclinica del paciente al proceso del trabajo). Para evaluar los factores asociados al tiempo dispensado, se adopto el modelo de Ecuaciones de Estimativos Generalizadas (Genneralized Estimating Equations – GEE), respetando la distribución Gamma de la variable diferencia del tiempo. Para la comparación intra factores fue adoptado la tesis de Bonferroni como test Post Hoc. Resultados: hubo 150 inserciones de SNE em 115 pacientes. A mediano tiempo entre la indicación y el uso de la sonda fueron 573 (IQR:360 – 1093; mínimo:63 – máximo: 3120) minutos. Se analizan lãs diferentes sub etapas del proceso, fueron identificados como factores independientes de riesgo para mayor tiempo entre la indicacion y el uso de la SNE: ionserción de SNE em paciente que no la utilizaba antes de la atencion em la Emergencia, médico no entregar la prescripción de inserción de sonda al enfermero, inserción de SNE em la jornada de trabajo nocturno, transferência del paciente em la sala de cuidados intensivos, retardos em la rutina medica, del la nutricion y de Enfermería, uso de ventilación mecânica, noradrenalina y la necesidad de la ayuna. Conclusión: El tiempo ente la indicación y el uso de la sonda nasoenterica fue elevado, exediendose 10 horas em la mitad de los casos. Factores relacionados a lãs condiciones clinicas de los pacientes y de la gestion de la Asistencia contribuyeron para el aumento en este tiempo.

Indiana University-Purdue University Indianapolis (IUPUI) / Gastric residual volume (GRV) is a clinical assessment to evaluate gastric emptying and enteral feeding tolerance. Factors such as the tube size, tube material, tube port configuration, placement of the tube in the gastric fluid, the amount of fluid and person completing the assessment may influence the accuracy of residual volume assessment. Little attention has been paid to assessing the accuracy of GRV measurement when the actual volume being aspirated is known, and no studies have compared the accuracy in obtaining RV using the three different techniques reported in the literature that are used to obtain aspirate in practice (syringe, suction, and gravity drainage). This in vitro study evaluated three different methods for aspirating feeding formula through two different tube sizes (10 Fr [small] and 18 Fr [large]), tube materials (polyvinyl chloride and polyurethane), using four levels of nursing experience (student, novice, experienced and expert) blinded to the five fixed fluid volumes of feeding formula in a simulated stomach, to determine if the RV can be accurately obtained. The study design consisted of a 3x2x2x4x5 completely randomized factorial ANOVA (with a total of 240 cells) and 479 RV assessments were made by the four nurse participants. All three methods (syringe, suction and gravity) used to aspirate RV did not perform substantially well in aspirating fluid, and on average, the methods were able to aspirate about 50% of the volume available. The syringe and suction techniques were comparable and produced higher proportions of RVs, although the interrater reliability of RV assessment was better with the syringe method. The gravity technique generally performed poorly. Overall, the polyvinyl chloride material and smaller tubes were associated with higher RV assessments. RV assessment is a variable assessment and the three methods did not perform well in this in vitro study. These findings should be further explored and confirmed using larger samples. This knowledge will be important in establishing the best technique for assessing RV to maximize EN delivery in practice and will contribute to future research to test strategies to optimize EN intake in critically ill patients.

Thesis (MMed)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: Background Occult blood loss from the gastrointestinal tract (GIT), causing iron deficiency often with anaemia, can be diagnostically and therapeutically challenging. This is because the endoscopic and radiologic tests may be negative due to the slow, chronic and intermittent nature of the gastrointestinal bleeding, making timing key in detection and localisation of the bleed. These limitations can be approached using two different radioactive isotopes. Firstly, we tested the sensitivity of extending Cr-51 RBC for 21 days relative to 5 days to detect GIT bleeding and its use to optimise timing of a Tc-99m RBC study for GIT blood loss localisation. Finally, we tested if the information provided by the Tc-99m RBC study aided gastroenterologic intervention for anatomical localisation of a lesion. Method In this retrospective review, after obtaining institutional and ethics committee approval, records of patients referred for evaluation of possible GIT blood loss were reviewed. In each; daily appearance of radiochromium in stool was measured in the whole body counter. In those cases exceeding 50 ml/day, a technetium-99m (Tc-99m) localization study was performed. These studies were correlated with clinical findings. Results A total of 59 Cr-51 RBC studies were carried out in 36 females and 21 males (n = 57). In 32 (54%) the radiochromium results were positive with 75% of the bleeding incidences occurring after 5 days of stool collection. Of 17 cases in whom Tc-99m RBC imaging studies were performed, 14 (82%) were positive with specific anatomical sites successfully defined in twelve. In all patients with blood loss of >100 ml/24h, Tc-99m RBC were positive and localised. Ten of the 17 Tc-99m RBC studies were further investigated and half diagnosed with small-bowel angiodysplasia. Conclusion This sequential twin isotope method is practical in revealing otherwise silent intestinal haemorrhage. Although it has good patient acceptability and clinical as well as diagnostic utility in management, further studies are required to clearly establish a cut-off level of blood loss for performing imaging studies and the impact of the findings on the overall patient management. / AFRIKAANSE OPSOMMING: Agtergrond Die evaluasie van okkulte bloedverlies uit die gastro-intestinale kanaal (GIT), met gevolglike ystertekort anemie, kan diagnosties en terapeuties uitdagend wees. Dit is omdat endoskopiese en radiologiese ondersoeke negatief mag wees as gevolg van die stadige, chroniese en intermitterende aard van die gastro-intestinale bloeding, wat die presiese tydstip van opsporing en lokalisering van die bloeding krities belangrik maak. Hierdie beperkings kan aangespreek word deur twee verskillende radioaktiewe isotope te gebruik. Eerstens is die sensitiwiteit van die verlenging van die Cr-51 RBS studie tot 21 dae in plaas van 5 dae om die GIT bloeding op te spoor, getoets, asook die gebruik daarvan om die optimale tyd vir ‘n Tc-99m RBS studie om die GIT bloedverlies te lokaliseer, vas te stel. Laastens is getoets of die inligting van die Tc-99m RBS studie wel bygedra het tot die gastroenterologiese ingreep om die letsel anatomies te lokaliseer. Metode Na institusionele en etiese komitee toestemming is inligting van pasiënte wat vir die evaluering van ‘n moontlike GI bloedverlies verwys is, in hierdie retrospektiewe oorsig nagegaan. Die daaglikse voorkoms van radioaktiewe chroom in stoelgangmonsters is in ‘n heelliggaamteller gemeet. In gevalle waar dit 50 ml/dag oorskry het, is ‘n tegnesium 99m (Tc 99m) studie gedoen. Hierdie studies is met die kliniese bevindinge gekorreleer. Resultate ‘n Totaal van 59 Cr-51 RBS studies is in 36 vroue en 21 mans (n = 57) gedoen. Die gemerkte chroomstudies was positief in 32 (54%), met 75% van die bloedings wat meer as 5 dae na versameling van die stoelgang plaasgevind het. In veertien (82%) van die 17 gevalle waar Tc-99m RBS studies gedoen is, was die studies positief. Spesifieke anatomiese gebiede van bloeding kon in 12 hiervan suksesvol bevestig word. Tc-99m RBS studies was positief in al die pasiënte met ‘n bloedverlies van >100 ml/24h, en kon gelokaliseer word. Tien van die 17 Tc-99m RBS studies is verder ondersoek en die helfte daarvan gediagnoseer met dunderm angiodisplasie. Gevolgtrekking Die opeenvolgende twee isotoopmetode om andersins asimptomatiese dermbloeding op te spoor, is prakties uitvoerbaar. Alhoewel die studies goed deur pasiënte aanvaar is, en ook van kliniese en diagnostiese waarde in die hantering van die pasiënte is, is verdere studies nodig om die afsnypunt vir die hoeveelheid bloedverlies om beeldingstudies uit te voer, sonder twyfel vas te stel, asook om die impak van die bevindings op ‘n groter pasiëntpopulasie vas te stel.

<b>Background:</b> Comparative assessments of the vascular and upper gastrointestinal risks of different regimens of non-steroidal anti-inflammatory drugs (NSAIDs) are required. <b>Methods:</b> Meta-analyses were conducted, using individual participant data where possible, of placebo-controlled trials of a selective cyclo-oxygenase [COX]-2 inhibitor ('coxib') or traditional NSAID, or randomised trials of a coxib versus traditional NSAIDs. A prespecified subdivision of traditional NSAID regimens of those with antiplatelet activity (mainly naproxen) and those without (mainly diclofenac) was made. Primary outcomes were major vascular events (MVEs; nonfatal myocardial infarction, nonfatal stroke or vascular death) and upper gastrointestinal complications (UGICs; perforation, obstruction or bleed). <b>Findings:</b> Searches identified 788 trials: 200 comparisons of a coxib vs placebo (88,604 participants, mean follow-up 0.60 years), 206 comparisons of a traditional NSAID vs placebo (43,482 participants, 0.46 years) and 149 comparisons of a coxib vs traditional NSAID (137,466 participants, mean follow-up 0.95 years). Compared to placebo, allocation to a coxib increased the risk of MVEs (rate ratio 1.38, 95&percnt; CI 1.14-1.66), vascular mortality (1.58, 1.11-2.24) and UGICs (1.81, 1.17-2.81). Overall, in the population studied, coxibs were associated with three additional major vascular events (one fatal) and two (rarely fatal) upper gastrointestinal complications per 1000 person-years exposure. There was no evidence of heterogeneity by duration of follow-up, coxib type, dose (other than for celecoxib), or patient characteristics, for the primary outcomes. The risk of MVEs for traditional NSAIDs without antiplatelet activity (mostly diclofenac 75mg bd or ibuprofen 800mg tds) were comparable to coxibs (1.40, 1.15-1.72); but the risk of UGICs (1.98, 1.39-2.84) was significantly greater. For traditional NSAIDs with antiplatelet activity (mostly naproxen 500mg bd) there were no significant excess of MVEs (0.84, 0.66-1.08), but UGICs were substantially increased (4.06, 2.85-5.78). Both coxibs and traditional NSAIDs increased risk of hospitalisation for heart failure by about two-fold. <b>Interpretation:</b> The vascular and upper gastrointestinal risks of coxibs and high-dose tNSAID regimens can be predicted, allowing the choice of analgesia to be tailored for particular patients.

In response to ingestion of nutrients, enteroendocrine L cells secrete the incretin hormone, glucagon-like peptide-1 (GLP-1), to enhance glucose-dependent insulin release. Therapies related to GLP-1 are approved for type 2 diabetes. The GLP-1 receptor (GLP-1R) is expressed in cells of the gastrointestinal tract and elsewhere. In pancreatic beta cells, GLP-1R are coupled to the Gs/cAMP/PKA pathway. The expression and function of GLP-1R in gastrointestinal smooth muscle are not known. Aim. To test the hypothesis that GLP-1 regulates smooth muscle function by acting on GLP-1R expressed on smooth muscle. Methods. Smooth muscle cells (SMC) were isolated and cultured. Expression of GLP-1R mRNA was measured by RT-PCR. Expression of GLP-1R protein was measured by western blot. The effect of GLP-1 (7-36) amide on Gαs activation, cAMP formation, and PKA activity was examined in cultured SMC. The effect of GLP-1 on basal activity and on acetylcholine-induced contraction was measured in intact colon via organ bath. Results. Amplification of GLP-1R mRNA suggested expression of GLP-1R mRNA in mucosal and non-mucosal colon cells, which was confirmed in pure SMC cultures. Similar patterns of protein expression were obtained with western blot. Addition of GLP-1 caused relaxation of phasic activity and agonist-induced tonic contractions in intact colon, suggesting a role of smooth muscle Gs-coupled GLP-1R in mediating relaxation. In SMC, GLP-1 (7-36) amide activated Gas, increased cAMP levels, and stimulated PKA activity. Conclusion. Colonic SMC express GLP-1R, and GLP-1 inhibits both basal and acetylcholine-induced contraction. The GLP1-R is coupled to the heterotrimeric G protein, Gas.

<p>Interindividualne razlike u sastavu i aktivnosti crevne mikroflore mogu uticati na metabolizam lekova kao i na njihov konačan terapijski odgovor. Simvastatin je lek iz grupe statina i karakteri&scaron;e ga izuzetno mala rastvorljivost u vodi, mala bioraspoloživost (&lt;5%) i velike interindividualne razlike u terapijskom odgovoru čiji uzroci nisu u potpunosti obja&scaron;njeni. Poslednjih godina velika pažnja se posvećuje ispitivanjima žučnih kiselina u razvoju novih farmaceutskih formulacija zbog svoje uloge u solubilizaciji i modifikaciji prodora lekova kroz biolo&scaron;ke membrane. Zbog svega navedenog, u fokusu na&scaron;eg istraživanja su bile potencijalne interakcije između simvastatina, probiotskih bakterija i žučnih kiselina o kojima se vrlo malo zna, a od izuzetne su važnosti, zbog mogućeg uticaja na farmakokinetske i farmakodinamske osobine simvastatina, pa samim tim i na konačan terapijski odgovor kod pacijenta.Cilj istraživanja je bio da se ispita prodor i metabolizam simvastatina u probiotskim bakterijama kao i uticaj različitih žučnih kiselina na transport ovog leka u bakterijske ćelije. Takođe, cilj je bio da se ispita uticaj soli žučnih kiselina na distribucioni koeficijent simvastatina, kao i interakcije žučnih kiselina sa simvastatinom na nivou transportnih proteina probiotskih bakterija kako bi se objasnila priroda očekivanih interakcija.Identifikacija i kvantifikacija uzoraka vr&scaron;ena je metodom tečne hromatografije sa masenom spektrometrijom (LC-MS/MS). Kori&scaron;ćenjem programskih paketa VolSurf+ i Molinspiration, za identifikovane metabolite su izračunati molekulski deskriptori koji opisuju fizičko-hemijske i farmakokinetske osobine molekula. Određivanje distribucionog koeficijenta vr&scaron;eno je Shake-flask metodom. Interakcije žučnih kiselina sa simvastatinom na nivou transportnih proteina probiotskih bakterija ispitane su doking studijama pomoću SwissDock programa. Prilikom dvadesetčetvoročasovne inkubacije sa probiotskim bakterijama uočen je statistički značajan pad koncentracije simvastatina u ekstracelularnom sadržaju. Ukupan sadržaj simvastatina, kao zbir ekstracelulamog i intracelularnog sadržaja, je tokom čitavog ispitivanog perioda bio statistički značajno niži u odnosu na kontrolnu grupu bez probiotika navodeći na zaključak da se deo simvastatina tokom vremena metabolisao pod dejstvom enzima ispitivanih bakterija. Detektovano je i identifikovano 8 metaboličkih produkata simvastatina. Na osnovu izračunatih vrednosti molekulskih deskriptora, očekuje se da će metabolit M-452, koji predstavlja hidroksilovani produkt simvastatinske kiseline, pokazati najbolje rezultate u pogledu fizičko-hemijskih osobina i bioraspoloživosti u biolo&scaron;kom sistemu. Žučne kiseline nisu dovele do statistički značajne modifikacije transporta simvastatina u/iz probiotskih bakterija. Ipak, u nekim vremenskim tačkama primećena je ne&scaron;to veća koncentracija leka u ekstracelulamom prostoru u grupama sa žučnim kiselinama. Ove razlike se mogu delimično objasniti rezultatima određivanja distribucionog koeficijenta koji su pokazali da ispitivane žučne kiseline dovode do statistički značajnog smanjenja distribucionog koeficijenta simvastatina usled povećanja rastvorljivosti u vodenoj fazi. Rezultatima doking studija procenjeno je da ispitivane žučne kiseline imaju veći afinitet prema čak 80% multidrug transportera ispitivanih bakterija u odnosu na simvastatin &scaron;to govori o mogućnosti ostvarivanja interakcija žučnih kiselina sa ovim lekom na nivou transportnih proteina probiotskih bakterija. Na osnovu dobijenih rezultata možemo zaključiti da probiotske bakterije imaju ogroman uticaj na sudbinu simvastatina u biolo&scaron;kom sistemu. Uzimajući u obzir činjenicu da probiotske bakterije ulaze u sastav normalne crevne flore i da svaki organizam poseduje specifičan bakterijski sastav, trebalo bi posvetiti vi&scaron;e pažnje ispitivanju njegovog uticaja na farmakokinetiku lekova. Neophodna su dalja in vivo ispitivanja kako bi se utvrdila potencijalna farmakolo&scaron;ka aktivnost identifikovanih metabolita simvastatina nastalih pod dejstvom enzimske aktivnosti probiotskih bakterija. Povećanje rastvorljivosti simvastatina pomoću žučnih kiselina otvara mogućnost za dalja istraživanja u cilju razvoja novih farmaceutskih formulacija sa pobolj&scaron;anom bioraspoloživosti i farmakokinetskim osobinama.</p> / <p>Interindividual differences in the composition and activity of the gut microflora may affect the metabolism of drugs as well as their final therapeutic response. Simvastatin is drug from the group of statins and has extremely low water solubility, low bioavailability (&lt;5%) and high interindividual differences in therapeutic response whose causes are not fully understood. In recent years, great attention has been paid to studies of bile acids in the development of new pharmaceutical formulations because of their role in the drug solubilization and modification of drug transport through biological membranes. Accordingly, interactions between simvastatin, probiotic bacteria and bile acids were the focus of our research due to great importance and potential influence on the pharmacokinetic and pharmacodynamic properties of simvastatin, and therefore the final therapeutic response in the patients. The aim of the study was to investigate the simvastatin transport and metabolism in probiotic bacteria as well as the effect of various bile acids on drug transport into the bacterial cell. Additonally, the aim was to investigate the influence of bile salts on the distribution coefficient of simvastatin, and the interactions of bile acids with simvastatin at the level of probiotic transport proteins in order to elucidate the nature of expected interactions. Identification and quantification of samples were performed with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Molecular descriptors that describe the physico-chemical and pharmacokinetic properties of identified metabolites were calculated using the software packages VolSurf+ and Molinspiration. Determination of the distribution coefficient was performed using Shake-flask method. Interaction of bile acids with simvastatin at the level of bacterial transport proteins were studied using docking studies with SwissDock program. During the twenty-four hours of incubation with probiotic bacteria, simvastatin concentrations in the extracellular contet showed a statistically significant decrease. The total amount of simvastatin, as the sum of the extracellular and intracellular amount, during the whole study period, was significantly lower in comparison with control group without probiotics, suggesting that the part of simvastatin was metabolized by the enzymatic activity of studied bacteria. Accordingly, eight metabolic products of simvastatin were detected and identified. Based on the calculated values of molecular descriptors, it is expected that the metabolite M-452, which is the hydroxylated product of simvastatin acid, will show the best results in terms of physico-chemical properties and bioavailability in biological system. Bile acids did not show a significant influence on simvastatin transport into probiotic bacteria. However, in some time points, slightly higher drug concentrations in the extracellular medium in groups with bile acids were observed. These differences can be partly explained by the results of the determination of the distribution coefficients which showed that investigated bile acids lead to a statistically significant decrease in simvastatin distribution coefficient due to increased solubility in the aqueous phase. The results of docking studies estimated that studied bile acids have stronger affinities for the 80% of bacterial multidrug transporters compared to simvastatin indicating the possibility of achieving the interactions of bile acids with simvastatin at the level of transport proteins of probiotic bacteria. Based on the obtained results it could be concluded that probiotic bacteria have great influence on the fate of simvastatin in a biological system. Taking into account the fact that probiotic bacteria are the normal part of gut microflora and that each individual has specific bacterial fingerprint, more attention should be paid on studying its influence on drug pharmakocinetics. Further in vivo studies are required in order to determine potential pharmacological activity of identified simvastatin metabolites. Increased water solubility of simvastatin with bile acids may open the possibility for further investigations with the aim of development of new pharmaceutical formulation with improved bioavailability and pharmacokinetic properties.</p>

Les tumeurs stromales gastro-intestinales (GIST) sont les tumeurs mésenchymateuses les plus fréquentes du système digestif. Elles ont pour origine les cellules interstitielles de Cajal (ICC) ou les cellules précurseurs mésenchymateuses communes aux ICCs et aux cellules musculaires lisses (SMC). Les GISTs sont des tumeurs qui sont chimiorésistantes et radiorésistantes. L'identification de mutations activatrices des gènes KIT (75-80%) ou/et PDGFRA (5-10%) a ouvert la voie à un traitement systémique chez les patients GIST sous forme d'Imatinib, un inhibiteur de tyrosine-kinase. Si ce traitement aboutit à une réponse clinique d'amélioration, un certain nombre d'effet secondaire sont néanmoins observés, comme les résistances au traitement. Afin d'améliorer le traitement initial, la physiopathologie du GIST doit progresser. La musculature de l'appareil digestif est une structure complexe composée de SMCs, de neurones entériques, de fibroblastes et d'ICCs. Au cours du développement, le mésoderme splanchnique donnera lieu au moins à deux types de cellules, les SMCs et les ICCs. Récemment, notre laboratoire a montré que la protéine de liaison à l'ARN RBPMS2 (pour RNA Binding Protein with Multiple Splicing 2) est impliquée dans le développement et le remodelage des SMCs digestives. Les travaux que j'ai réalisés au cours de ma thèse avaient pour objectifs d'étudier l'expression et la fonction de RBPMS2 dans les tumeurs GISTs humains. Nous avons analysé l'expression de RBPMS2 dans les GIST humains et nous avons démontré que RBPMS2 était fortement exprimé dans les tumeurs GISTs de manière indépendante de l'activité KIT. Nous avons également analysé la fonction de RBPMS2 en culture et avons montré que l'expression ectopique de RBPMS2 dans les SMCs humaines adultes et différenciées culture conduisaient à l'augmentation de leur taux de prolifération et altèreraient leur différenciation. Ces résultats suggèrent que RBPMS2 et les voies de signalisation qu´il contrôle pourraient être des cibles thérapeutiques potentielles dans la thérapie des tumeurs GISTs. / Gastrointestinal stromal tumors (GIST) are the most common mesenchymal neoplasm of the GI tract. They are supposed to arise from the interstitial cells of Cajal (ICCs) or from a mesenchymal precursor cell, common of ICCs and smooth muscle cells (SMCs). GISTs are highly resistant to conventional chemotherapy and radiotherapy. However, a targeted therapy is now proposed. These tumors have activating mutations in two closely related genes, the KIT (75-80%) or/and the PDGFRA (5-10%). Targeting these mutated activated proteins with Imatinib mesylate, a small-molecule tyrosine kinase inhibitor, has proven efficient in GIST treatment. However, resistance to Imatinib finally develops and new-targeted therapies are necessary. The musculature of the gastrointestinal (GI) tract is a highly complex structure composed of visceral SMCs, enteric neurons, fibroblast-like cells and ICCs. During the development, the splanchnic mesoderm will give rise at least to two cell types, ICCs and SMCs. Recently our laboratory showed that the RNA Binding Protein with Multiple Splicing 2 (RBPMS2) is involved into the development and remodeling of SMC.My PhD works investigate the expression and function of RBPMS2 in human GISTs. We analyzed the expression of RBPMS2 in human GISTs and we found that RBPMS2 was abnormally highly expressed in the tumoral cells of GISTs. We also analyzed the function of RBPMS2 into human adult SMC cell culture and demonstrated that ectopic expression of RBPMS2 in mature and differentiated SMC cultures increases their proliferation rate and alters their differentiation. These findings suggest that RBPMS2 could be a potential target for cancer therapy.

Após a constatação da escassez de estudos realizados com vegetais crus na busca por novas estirpes de bactérias láticas (BAL) produtoras de bacteriocinas e diante do potencial tecnológico da aplicação destas cepas tanto como agentes de conservação em alimento, bem como cultura probiótica em alimentos funcionais, este estudo objetivou isolar e identificar cepas de bactérias láticas potencialmente bacteriocinogênicas de amostras de rúcula obtidas no comércio local de São Paulo, SP - Brasil, identificar e caracterizar as bacteriocinas produzidas pelos isolados e avaliar o potencial probiótico dos isolados testando sua sobrevivência no modelo dinâmico do trato gastrointestinal TNO gastro-Intestinal Model - TIM-1 disponível no TNO (The Netherlands Organization for Applied Scientific Research) divisão Quality of Life (Zeist, Holanda). A produção de bacteriocinas neste modelo também foi avaliada, comparando-se com L. sakei 2a, também produtora de bacteriocinas e ainda avaliou-se a interferência na viabilidade de E. faecium LMA1. A cepa Lactococcus lactis subsp. lactis MK02R de rúcula produziu uma bacteriocina sensível à enzimas proteolíticas, termoestável e não influenciada pelo pH, sendo capaz de inibir Enterococcus faecium, Lactobacillus sakei, Listeria innocua, Lactobacillus delbrueckii e Listeria Monocytogenes de diferentes grupos sorológicos. Os ensaios genéticos utilizando primers Nisf e Nisr confirmaram que a bacteriocina MK02R é uma nisina, apresentando uma alteração dos aminoácidos no peptídeo líder em relação às nisinas A, Z, Q, F e U, porém com a estrutura do peptídeo maduro idêntica ao da nisina F. Estes resultados foram confirmados por espectrometria de massas de amostras purificadas por HPLC. L. lactis MK02R resistiu à passagem no modelo dinâmico TIM-1, apresentando uma alta capacidade de sobreviver nas condições simuladas do trato gastrointestinal humano. Entretanto, não foi capaz de causar a redução no número de E. faecium LMA1. Em contrapartida, L. sakei 2a, mesmo apresentando uma sobrevivência menor, foi capaz de causar uma redução de 70% na população de E. faecium LMA1 no ambiente simulado do TGI. Não foi detectada atividade residual da ação antimicrobiana das bacteriocinas produzidas por L. lactis MK02R ou L. sakei 2a após a passagem pelo modelo dinâmico TIM-1. Estes resultados evidenciam a possível aplicação de L. lactis MK02R como um agente de controle biológico na conservação de alimentos e também como uma cultura potencialmente probiótica. / Given the scarcity of studies performed with raw vegetables addressing the search for new bacteriocinogenic strains of lactic acid bacteria (LAB) and considering the technological application of these strains as food preservatives and probiotic cultures in functional foods, this study was aimed at isolation and identification of bacteriocinogenic LAB strains from samples of rocket salad obtained in the local market of São Paulo, SP - Brazil, subsequent characterization of the bacteriocins produced by these LABs and evaluation of their probiotic potential by testing their survival in the dynamic gastrointestinal model TNO gastro- Intestinal-Model - TIM-1, available at the TNO (Netherlands Organization for Applied Scientific Research) Quality of Life division (Zeist, Netherlands). The studies in the TIM-1 model were also done with another bacteriocinogenic strain L. sakei 2a for comparison, evaluating their interference on the viability of E. faecium LMA1. The bacteriocin produced by strain Lactococcus lactis subsp. lactis MK02R isolated from rocket salad was sensitive to proteolytic enzymes, heat-stable and not influenced by the pH. The bacteriocin inhibited the growth of Enterococcus faecium, Lactobacillus sakei, Listeria innocua, Lactobacillus delbrueckii the primers Nisf and Nisr indicated that the bacteriocin produced by the strain MK02R is a nisin, with a change in the amino acid sequence of the leader peptide when compared to nisin A, Z, Q, U and F, but with the structure of the mature peptide homologous to that of nisin F. These results were confirmed by mass spectrometry of purified samples obtained by HPLC. L. lactis MK02R withstood the test in the dynamic model TIM-1, presenting capability to survive in the simulated conditions of the human gastrointestinal tract. However, the strain was not able to cause a reduction in the number of E. faecium LMA1. On the other hand, L. sakei 2a, even presenting lower survival, was able to cause 70% reduction in the population of E. faecium LMA1 in the gut simulated environment. No residual antimicrobial activity of bacteriocin produced by L. lactis MK02R or L. sakei 2a was detected after the transit through the dynamic model TIM-1. These results demonstrate the possible application of L. lactis MK02R both as a biocontrol agent in food preservation and as a potentially probiotic culture.

Estudos com farinha de banana verde (FBV), rica em amido resistente, mostram efeitos positivos sobre a saciedade, resposta glicêmica e melhora do funcionamento intestinal. Entretanto, pouco se sabe sobre a capacidade da FBV em estimular seletivamente o crescimento e/ou atividades de microbiota intestinal benéfica e os efeitos fisiológicos do consumo habitual. No presente trabalho foi investigado o efeito da ingestão regular e descontinuada de FBV sobre a microbiota intestinal em voluntários saudáveis, bem como as interações com hormônios relacionados à fome e saciedade, funcionamento intestinal e homeostase da glicose. Para tanto foi realizado estudo de intervenção, duplo cego paralelo controlado com placebo, no qual voluntários saudáveis consumiram FBV ou maltodextrina, veiculadas através de sopa prontas congeladas, três vezes por semana e durante seis semanas. Os resultados evidenciaram que a FBV pode aumentar a saciedade, promover redução no aporte energético de refeições subsequentes (14%) e melhorar o funcionamento intestinal. Ao mesmo tempo reduz a secreção plasmática de insulina no jejum e o Índice HOMA2-RI em 20%, sinalizando aumento na sensibilidade à insulina. A análise da microbiota intestinal utilizando o rDNA 16S mostrou que existem dois grupos distintos de indivíduos, os quais respondem diferentemente ao consumo de FBV. O consumo de FBV por voluntários, cujo microbioma era mais abundante no gênero Prevotella, apresentou aumento de genes envolvidos em vias metabólicas relacionadas à degradação anaeróbia de carboidratos (794 Kegg orthologs, FDR=0,05), como as vias do metabolismo de amido e glicose, do butirato, propionato. Paralelamente outros genes indicaram redução de algumas vias metabólicas, incluindo a biossíntese de lipopolissacarídeos. Este mesmo grupo de voluntários apresentou gêneros microbianos positivamente relacionados com conteúdo de ácidos graxos de cadeia curta (AGCC), em padrão distinto do outro grupo de voluntários que consumiu FBV e do grupo Controle. Foi possível demonstrar que o consumo de FBV pode promover a modulação do microbioma em indivíduos saudáveis com enterótipos distintos, trazendo efeitos benéficos para a saúde humana. / Studies with Unripe Banana Flour, rich in resistant starch, shave shown positive effects on satiety, glycemic response and improved intestinal function. Nevertheless, little is known about its capacity to selectively stimulate the intestinal microbiota\'s activity, or the physiological effects of its habitual consumption. This study investigated the effects of the regular, discontinued ingestion of UBF on the intestinal microbiome in healthy volunteers, as well as effects on hormones related to satiety, intestinal function and glucose homeostasis. To achieve these goals, a double blind, parallel, placebo controlled study was designed, in which healthy volunteers ingested UBF or maltodextrin added to a standardized frozen soup meal, 3 times a week for 6 weeks. The results showed that UBF can improve satiety, promote a reduction in energy intake at subsequent meals (14%) and improve intestinal function. At the same time, it reduces plasmatic secretion of fasting insulin and e the HOMA2-RI index by 20%, signaling an increase in insulin sensitivity. The analysis of the microbiome using the 16S rDNA gene showed that there are two clusters of individuals, which respond differently to the dietary intervention. The UBF consumption by volunteers with a Prevotella dominant microbiome showed an increase in genes related to anaerobic carbohydrate degradation (794 Kegg orthologs, FDR=0,05), such as members of the starch and glucose metabolism, propanoate metabolism and butyrate metabolism. At the same time, other genes were reduced, including the biosynthesis of lipopolysaccharides. The same volunteers presented several microbial groups positively correlated with the short chain fatty acids (SCFA) present in the fecal samples analyzed. This was a distinct pattern to that observed for the remaining volunteers. We demonstrated that the consumption of UBF can promote the overall health of the human host as well as the modulation of the intestinal microbiome in healthy individuals and that this effect is dependent on the enterotype present.