O crescimento e a progressão tumorais estão associados à indução da angiogênese. O objetivo deste estudo foi avaliar a imunoexpressão do VEGF e a densidade de microvasos em adenocarcinomas gástricos. Espécimes cirúrgicos de 89 pacientes submetidos à ressecção gástrica com dissecção linfonodal a D2 foram analisados quanto à densidade microvascular tumoral. Testes imunohistoquímicos foram realizados utilizando-se os anticorpos CD31, CD34, Fator VIII e VEGF através do método da streptavidina-biotina. Os vasos foram contados nas áreas de maior vascularização tumoral e o VEGF foi graduado de 0 a 3, de acordo com a intensidade da imunocoloração. Os resultados imunohistoquímicos foram comparados com os dados patológicos e de extensão loco-regional da doença. Sessenta pacientes (67,4%) eram do sexo masculino e a média de idade foi 59,9 (±13,8) anos. Os tumores foram classificados em tipo intestinal em 62 casos (69,7%) e em difuso em 27 (30%). Onze pacientes (12,4%) apresentaram tumores precoces. A densidade microvascular apresentou média de 67,8 (±31,5) para o anticorpo CD31 e 94,2 (±39,0) para o CD34. A média do número de vasos corados pelo Fator VIII foi 9,2 (±8,2). Houve uma correlação entre os resultados imunohistoquímicos para CD31 e CD34, com r=0,57 e p=0,01. Segundo o VEGF os tumores foram: grau 0 = 16 (18%), I = 48 (53,9%), II = 16 (18%) e III = 9 (10,1%). Não houve associação entre a DMV e os dados patológicos: tipo histológico, grau de diferenciação, tamanho do tumor, invasão da parede gástrica, metástases linfonodais e metástase à distância. A imunoexpressão do VEGF foi associada com alta DMV (p=0,03) e com o estádio tumoral - TNM (p=0,003). Os resultados deste estudo permitiram concluir que a coloração imunohistoquímica com o Fator VIII não é segura como um marcador de células endoteliais; que os anticorpos CD31 e CD34 foram úteis na avaliação da DMV; e que a imunoexpressão do VEGF pode identificar um comportamento biológico mais agressivo / Tumor growth and progression, particularly in aggressive and malignant tumors, are associated with the induction of angiogenesis. The aim of this study was to evaluate the role of VEGF immunoexpression and microvascular density in gastric adenocarcinomas. Specimens from eightynine patients who underwent gastric resection with DII lymph node dissection were analyzed regarding microvascular density of the tumors. Immunohistochemistry for CD31, CD34, Factor VIII and VEGF were performed using the streptavidin-biotin-method. The vessels were counted in a hot spot area of the tumors and VEGF was categorized as grade 0 to III, according to the intensity. Immunohistochemical results were compared to pathological data and loco-regional extension of the disease. In the results of 89 patients, sixty (67.4%) were men, and the mean age was 59.9 ± 13.8 years. The tumors were classified as intestinal type in 62 (69.7%) and diffuse in 27 (30.3%). Eleven (12.4%) were early gastric tumors. Microvascular density showed a mean of 67.8 (± 31.5) for CD31 and 94.2 (± 39) for CD34 vessels. The mean number of vessels revealed by the antibody Fator VIII was 9.2 (± 8.2). There was a correlation between immunohistochemistry for CD31 and CD34, r = 0.57, p < 0.01. The tumors were classified for VEGF as: grade 0 = 16 (18%); I = 48 (53,9%); II = 16 (18%) and III = 9 (10,1%). There was no association between immunohistochemistry and pathological data including: histologic type, grade of differentiation, tumor size, tumor depth, lymph node metastasis and distant metastasis. However, VEGF immunoexpression was associated with high microvascular density (p = 0,03) and there was an association between the immunoexpression of VEGF and the tumor stage - TNM (p = 0,003). It was concluded that immunohistochemical staining for anti-Factor VIII was not reliable as a microvascular density marker; CD31 and CD34 were useful to demonstrate microvascular density and VEGF immunoexpression may identify tumors with more aggressive biological behavior

Bypass or resection of the stomach and oesophagus, has long been recognised to result in profound changes in the handling of ingested nutrients. This results in significant morbidity after radical surgery for oesophago-gastric cancer, in particular post-prandial hypoglycaemia, altered appetite, early satiety and noxious post-prandial symptoms. By profiling and challenging the gut hormone axis in healthy volunteers and patients who had undergone total or subtotal gastrectomy, or oesophagectomy, this thesis explores the possible causative mechanisms for the challenges faced by this patient population. In the surgical groups, an oral glucose tolerance test (OGTT) resulted in enhanced secretion of satiety and incretin gut hormones (GLP-1, GIP, PYY) and insulin, followed by hypoglycaemia in a cohort of patients. Continuous glucose monitoring of gastrectomy participants over two weeks of normal lifestyle identified an increased incidence of day and night time hypoglycaemia. RNAseq and mass spectrometry based peptidomics of human and murine enteroendocrine cells in the pre- and post-operative populations revealed no significant change in the underlying cellular pathways for nutrient sensing and gut hormone secretion, indicating that the altered hormone secretion is primarily driven by accelerated nutrient transit, rather than adaptive changes in the gut. Finally, specific blockade of the GLP-1 receptor in post-gastrectomy patients using Exendin 9-39 normalised insulin secretion and prevented reactive hypoglycaemia after an OGTT. In conclusion, profound changes in gut hormone secretion as a result of enhanced nutrient transit after foregut surgery likely underlie the early and late post-prandial symptoms seen in this group, and therapies specifically targeting the gut hormone axis, and GLP-1 in particular, could be the first targeted treatments for post-gastrectomy syndromes.

INTRODUÇÃO: Não obstante os bons resultados e reprodutibilidade da derivação gástrica em Y de Roux (DGYR), discutem-se ainda aspectos técnicos que podem influenciar tanto o resultado na perda de peso, como a qualidade de vida pós-operatória (PO). Atribui-se hoje a efeitos restritivos, como volume da bolsa gástrica e sua velocidade de esvaziamento, assim como a fatores metabólicos, o resultado da cirurgia. As influências morfofuncionais não são totalmente compreendidas. OBJETIVO: Avaliar a influência do volume da bolsa gástrica e sua velocidade de esvaziamento sobre a perda de peso e a tolerância alimentar no PO de DGYR. MÉTODO: Pacientes em PO tardio de DGYR; foram avaliados a percentagem de perda de excesso de peso (%PEP), reganho de peso (%PEPreganho), tolerância alimentar por questionário de tolerância (ES), volume da bolsa (V) por tomografia computadorizada tridimensional (TC3D) e velocidade de esvaziamento da bolsa por cintilografia, avaliada por percentual de retenção (%Ret) em 1, 2 e 4 horas. Foram identificados pontos de corte para V e %Ret em 1 hora (%Ret1). A partir destes parâmetros a amostra foi categorizada por V e %Ret1, e os dados correlacionados com resultados em perda de peso e tolerância alimentar. RESULTADOS: Foram estudados 67 pacientes. A idade mediana foi 51 anos, 91,04% sexo feminino, e IMC mediano inicial de 51,44 kg/m2. O tempo de seguimento PO mediano foi de 47 meses. A amostra apresentou 60,27%PEPnadir como mediana da maior perda de peso PO e 16,13%PEPreganho. A mediana de V foi de 28mL; a %Ret em 1, 2, 4 horas foras foram 8%, 2%, 1%, respectivamente. A pontuação mediana do ES foi 21. Os pontos de corte considerados foram V=40mL, e %Ret1 de 12% e 25%. Foram comparados subgrupos por V (V <= 40mL, V > 40mL) e por %Ret1 (%Ret1 <= 12%, 12%<%Ret1 < 25%, e %Ret1>=25%). Na análise categorizada por V, houve associação entre V <= 40mL e maior velocidade de esvaziamento até 2 horas (V <= 40mL: %Ret1=6, %Ret2=2, p=0,009; V > 40mL: %Ret1=44, %Ret2=13,5, p=0,045). Na análise categorizada por %Ret1, observou-se associação entre maior velocidade de esvaziamento em 1 hora e melhor evolução ponderal tardia, representada por menor %PEPreganho (p=0,036), e maior %PEPatual (p=0,033) no grupo com %Ret1 <= 12%. Na avaliação do ES, associou-se melhor tolerância alimentar (ES > 24) e menor %Ret1 (p=0,003). CONCLUSÕES: Houve associação do volume da bolsa gástrica com a velocidade de esvaziamento. O esvaziamento mais rápido correlacionou-se com maior %PEP tardia, menor reganho de peso e melhor tolerância alimentar. Estes dados sugeriram que a presença de bolsa gástrica pequena, com menos de 40 mL, e com rápido esvaziamento, são parâmetros importantes para adequado resultado tardio na DGYR. O uso da TC3D e da cintilografia para avaliação morfofuncional da DGYR podem contribuir para investigação do reganho de peso e intolerância alimentar PO / INTRODUCTION: Despite the good results and good reproducibility of the Roux-en-Y gastric bypass (GBP), discussions about technical aspects still remain. Postoperative anatomy can influence the result in weight loss, as the postoperative (PO) quality-of-life. Surgical results are nowadays attributed to restriction, promoted by the small neo-stomach volume and its emptying, as well as metabolic factors. These anatomical and functional influences on surgical results are often poorly evaluated, and yet not fully understood. OBJECTIVE: To evaluate the influence of the gastric pouch volume and its emptying rate on weight loss, and food tolerance, in the late PO of GBP. METHOD: Patients followed-up at HC-FMUSP in late PO of GBP; percentage of excess weight loss (%EWL), weight regained (%EWLregain), food tolerance with specific questionnaire (Suter Score, SS), gastric pouch volumetry (V) by three-dimensional computed tomography (CT3D), and pouch emptying rate by scintigraphy, evaluated by retention percentage (%Ret) in 1, 2, 4 hours, were assessed. Cutoffs were identified for %Ret at 1 hour (%Ret1) and V. From these parameters the sample was categorized by V and %Ret1, looking for associations between V and %Ret and results in weight loss and food tolerance. RESULTS: Eighty patients were enrolled and 67 completed the study. The median age was 51 years, 91.04% were female, with initial median BMI of 51.44 kg/m2. The PO median follow-up time was 47 months. The sample showed median of 60.27% as EWLnadir as PO better weight loss, with 16.13% of EWLregain. The median V was 28mL; %Ret at 1, 2, 4 hours were 8%, 2%, 1%, respectively. The median score of SS was 21. The cutoff points considered were V=40mL, and %Ret1=12% and 25%. Subgroups were compared by V (V <= 40mL, V > 40mL) and Ret1% (%Ret1 <= 12%, 12% < %Ret1 < 25%, and %Ret1 >= 25%). The categorized analysis by V found associations between V<=40mL and higher emptying speeds up to 2 hours (V <= 40mL: %Ret1=6, %Ret2=2, p=0.009; V > 40mL: %Ret1=44, %Ret2=13.5, p=0.045). In the categorized analysis by %Ret1, it was found association between higher emptying speed in 1 hour and higher late PO weight loss, represented by lower %EWLregain (p=0.036), and higher late %EWL (p=0.033) in the group with %Ret1<=12%, compared to the group %Ret1 >= 25%. Better food tolerance, assessed by Suter questionnaire (SS > 24), was associated with lower %Ret1 (p=0.003). CONCLUSIONS: There was an association between gastric pouch volume and emptying rate. Smaller gastric pouch have shown faster gastric emptying. The fastest gastric emptying was correlated with higher late %EWL, less weight regain and better PO food tolerance. These data suggest that the construction of small gastric pouch, with less than 40 mL, and with rapid emptying rate, are important parameters for good outcomes in late postoperative in GBP. The use of CT3D and scintigraphy for morphological and functional assessment of GBP can contribute in the evaluation of possible causes of late postoperative weight regain and food intolerance

O crescimento e a progressão tumorais estão associados à indução da angiogênese. O objetivo deste estudo foi avaliar a imunoexpressão do VEGF e a densidade de microvasos em adenocarcinomas gástricos. Espécimes cirúrgicos de 89 pacientes submetidos à ressecção gástrica com dissecção linfonodal a D2 foram analisados quanto à densidade microvascular tumoral. Testes imunohistoquímicos foram realizados utilizando-se os anticorpos CD31, CD34, Fator VIII e VEGF através do método da streptavidina-biotina. Os vasos foram contados nas áreas de maior vascularização tumoral e o VEGF foi graduado de 0 a 3, de acordo com a intensidade da imunocoloração. Os resultados imunohistoquímicos foram comparados com os dados patológicos e de extensão loco-regional da doença. Sessenta pacientes (67,4%) eram do sexo masculino e a média de idade foi 59,9 (±13,8) anos. Os tumores foram classificados em tipo intestinal em 62 casos (69,7%) e em difuso em 27 (30%). Onze pacientes (12,4%) apresentaram tumores precoces. A densidade microvascular apresentou média de 67,8 (±31,5) para o anticorpo CD31 e 94,2 (±39,0) para o CD34. A média do número de vasos corados pelo Fator VIII foi 9,2 (±8,2). Houve uma correlação entre os resultados imunohistoquímicos para CD31 e CD34, com r=0,57 e p=0,01. Segundo o VEGF os tumores foram: grau 0 = 16 (18%), I = 48 (53,9%), II = 16 (18%) e III = 9 (10,1%). Não houve associação entre a DMV e os dados patológicos: tipo histológico, grau de diferenciação, tamanho do tumor, invasão da parede gástrica, metástases linfonodais e metástase à distância. A imunoexpressão do VEGF foi associada com alta DMV (p=0,03) e com o estádio tumoral - TNM (p=0,003). Os resultados deste estudo permitiram concluir que a coloração imunohistoquímica com o Fator VIII não é segura como um marcador de células endoteliais; que os anticorpos CD31 e CD34 foram úteis na avaliação da DMV; e que a imunoexpressão do VEGF pode identificar um comportamento biológico mais agressivo / Tumor growth and progression, particularly in aggressive and malignant tumors, are associated with the induction of angiogenesis. The aim of this study was to evaluate the role of VEGF immunoexpression and microvascular density in gastric adenocarcinomas. Specimens from eightynine patients who underwent gastric resection with DII lymph node dissection were analyzed regarding microvascular density of the tumors. Immunohistochemistry for CD31, CD34, Factor VIII and VEGF were performed using the streptavidin-biotin-method. The vessels were counted in a hot spot area of the tumors and VEGF was categorized as grade 0 to III, according to the intensity. Immunohistochemical results were compared to pathological data and loco-regional extension of the disease. In the results of 89 patients, sixty (67.4%) were men, and the mean age was 59.9 ± 13.8 years. The tumors were classified as intestinal type in 62 (69.7%) and diffuse in 27 (30.3%). Eleven (12.4%) were early gastric tumors. Microvascular density showed a mean of 67.8 (± 31.5) for CD31 and 94.2 (± 39) for CD34 vessels. The mean number of vessels revealed by the antibody Fator VIII was 9.2 (± 8.2). There was a correlation between immunohistochemistry for CD31 and CD34, r = 0.57, p < 0.01. The tumors were classified for VEGF as: grade 0 = 16 (18%); I = 48 (53,9%); II = 16 (18%) and III = 9 (10,1%). There was no association between immunohistochemistry and pathological data including: histologic type, grade of differentiation, tumor size, tumor depth, lymph node metastasis and distant metastasis. However, VEGF immunoexpression was associated with high microvascular density (p = 0,03) and there was an association between the immunoexpression of VEGF and the tumor stage - TNM (p = 0,003). It was concluded that immunohistochemical staining for anti-Factor VIII was not reliable as a microvascular density marker; CD31 and CD34 were useful to demonstrate microvascular density and VEGF immunoexpression may identify tumors with more aggressive biological behavior

INTRODUÇÃO: Não obstante os bons resultados e reprodutibilidade da derivação gástrica em Y de Roux (DGYR), discutem-se ainda aspectos técnicos que podem influenciar tanto o resultado na perda de peso, como a qualidade de vida pós-operatória (PO). Atribui-se hoje a efeitos restritivos, como volume da bolsa gástrica e sua velocidade de esvaziamento, assim como a fatores metabólicos, o resultado da cirurgia. As influências morfofuncionais não são totalmente compreendidas. OBJETIVO: Avaliar a influência do volume da bolsa gástrica e sua velocidade de esvaziamento sobre a perda de peso e a tolerância alimentar no PO de DGYR. MÉTODO: Pacientes em PO tardio de DGYR; foram avaliados a percentagem de perda de excesso de peso (%PEP), reganho de peso (%PEPreganho), tolerância alimentar por questionário de tolerância (ES), volume da bolsa (V) por tomografia computadorizada tridimensional (TC3D) e velocidade de esvaziamento da bolsa por cintilografia, avaliada por percentual de retenção (%Ret) em 1, 2 e 4 horas. Foram identificados pontos de corte para V e %Ret em 1 hora (%Ret1). A partir destes parâmetros a amostra foi categorizada por V e %Ret1, e os dados correlacionados com resultados em perda de peso e tolerância alimentar. RESULTADOS: Foram estudados 67 pacientes. A idade mediana foi 51 anos, 91,04% sexo feminino, e IMC mediano inicial de 51,44 kg/m2. O tempo de seguimento PO mediano foi de 47 meses. A amostra apresentou 60,27%PEPnadir como mediana da maior perda de peso PO e 16,13%PEPreganho. A mediana de V foi de 28mL; a %Ret em 1, 2, 4 horas foras foram 8%, 2%, 1%, respectivamente. A pontuação mediana do ES foi 21. Os pontos de corte considerados foram V=40mL, e %Ret1 de 12% e 25%. Foram comparados subgrupos por V (V <= 40mL, V > 40mL) e por %Ret1 (%Ret1 <= 12%, 12%<%Ret1 < 25%, e %Ret1>=25%). Na análise categorizada por V, houve associação entre V <= 40mL e maior velocidade de esvaziamento até 2 horas (V <= 40mL: %Ret1=6, %Ret2=2, p=0,009; V > 40mL: %Ret1=44, %Ret2=13,5, p=0,045). Na análise categorizada por %Ret1, observou-se associação entre maior velocidade de esvaziamento em 1 hora e melhor evolução ponderal tardia, representada por menor %PEPreganho (p=0,036), e maior %PEPatual (p=0,033) no grupo com %Ret1 <= 12%. Na avaliação do ES, associou-se melhor tolerância alimentar (ES > 24) e menor %Ret1 (p=0,003). CONCLUSÕES: Houve associação do volume da bolsa gástrica com a velocidade de esvaziamento. O esvaziamento mais rápido correlacionou-se com maior %PEP tardia, menor reganho de peso e melhor tolerância alimentar. Estes dados sugeriram que a presença de bolsa gástrica pequena, com menos de 40 mL, e com rápido esvaziamento, são parâmetros importantes para adequado resultado tardio na DGYR. O uso da TC3D e da cintilografia para avaliação morfofuncional da DGYR podem contribuir para investigação do reganho de peso e intolerância alimentar PO / INTRODUCTION: Despite the good results and good reproducibility of the Roux-en-Y gastric bypass (GBP), discussions about technical aspects still remain. Postoperative anatomy can influence the result in weight loss, as the postoperative (PO) quality-of-life. Surgical results are nowadays attributed to restriction, promoted by the small neo-stomach volume and its emptying, as well as metabolic factors. These anatomical and functional influences on surgical results are often poorly evaluated, and yet not fully understood. OBJECTIVE: To evaluate the influence of the gastric pouch volume and its emptying rate on weight loss, and food tolerance, in the late PO of GBP. METHOD: Patients followed-up at HC-FMUSP in late PO of GBP; percentage of excess weight loss (%EWL), weight regained (%EWLregain), food tolerance with specific questionnaire (Suter Score, SS), gastric pouch volumetry (V) by three-dimensional computed tomography (CT3D), and pouch emptying rate by scintigraphy, evaluated by retention percentage (%Ret) in 1, 2, 4 hours, were assessed. Cutoffs were identified for %Ret at 1 hour (%Ret1) and V. From these parameters the sample was categorized by V and %Ret1, looking for associations between V and %Ret and results in weight loss and food tolerance. RESULTS: Eighty patients were enrolled and 67 completed the study. The median age was 51 years, 91.04% were female, with initial median BMI of 51.44 kg/m2. The PO median follow-up time was 47 months. The sample showed median of 60.27% as EWLnadir as PO better weight loss, with 16.13% of EWLregain. The median V was 28mL; %Ret at 1, 2, 4 hours were 8%, 2%, 1%, respectively. The median score of SS was 21. The cutoff points considered were V=40mL, and %Ret1=12% and 25%. Subgroups were compared by V (V <= 40mL, V > 40mL) and Ret1% (%Ret1 <= 12%, 12% < %Ret1 < 25%, and %Ret1 >= 25%). The categorized analysis by V found associations between V<=40mL and higher emptying speeds up to 2 hours (V <= 40mL: %Ret1=6, %Ret2=2, p=0.009; V > 40mL: %Ret1=44, %Ret2=13.5, p=0.045). In the categorized analysis by %Ret1, it was found association between higher emptying speed in 1 hour and higher late PO weight loss, represented by lower %EWLregain (p=0.036), and higher late %EWL (p=0.033) in the group with %Ret1<=12%, compared to the group %Ret1 >= 25%. Better food tolerance, assessed by Suter questionnaire (SS > 24), was associated with lower %Ret1 (p=0.003). CONCLUSIONS: There was an association between gastric pouch volume and emptying rate. Smaller gastric pouch have shown faster gastric emptying. The fastest gastric emptying was correlated with higher late %EWL, less weight regain and better PO food tolerance. These data suggest that the construction of small gastric pouch, with less than 40 mL, and with rapid emptying rate, are important parameters for good outcomes in late postoperative in GBP. The use of CT3D and scintigraphy for morphological and functional assessment of GBP can contribute in the evaluation of possible causes of late postoperative weight regain and food intolerance

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.

Obesity has grown to epidemic proportions, and in lack of efficient life-style and medical treatments, the bariatric surgeries are performed in rising numbers. The most common surgery is the Gastric Bypass (GBP) surgery, with the Biliopancreatic diversion with duodenal switch (DS) as an option for the most extreme cases with a BMI&gt;50 kg/m2. In paper I 20 GBP-patients were examined during the first post-operative year regarding the natriuretic peptide, NT-ProBNP, which is secreted from the cardiac ventricles. Levels of NT-ProBNP quickly increased during the first post-surgery week, and later established itself on a higher level than pre-surgery. In paper II we report of 5 patient-cases after GBP-surgery with severe problems with postprandial hypoglycaemia that were successfully treated with GLP-1-analogs. The effect of treatment could be observed both symptomatically and in some cases using continuous glucose measuring systems (CGMS). In paper III three groups of subjects; 15 post-GBP patients, 15 post-DS, and 15 obese controls were examined for three days using CGMS during everyday life. The post-GBP group had high glucose variability as measured by MAGE and CONGA, whereas the post-DS group had low variability. Both post-operative groups exhibited significant time in hypoglycaemia, about 40 and 80 minutes per day &lt;3.3mmol/l and 20 and 40 minutes &lt; 2.8mmol/l, respectively, longer time for DS-group. Remarkably, only about 20% of these hypoglycaemic episodes were accompanied with symptoms. In Paper IV the hypoglycaemia counter regulatory system was investigated; 12 patients were examined before and after GBP-surgery with a stepped hypoglycaemic hyperinsulinemic clamp. The results show a downregulation of symptoms, counter regulatory hormones (glucagon, cortisol, epinephrine, norepinephrine, growth hormone), incretin hormones (GLP-1 and GIP), and sympathetic nervous response. In conclusion patients post bariatric surgery exhibit a downregulated counter regulatory response to hypoglycaemia, accompanied by frequent asymptomatic hypoglycaemic episodes in everyday life. Patients suffering from severe hypoglycaemic episodes can often be treated successfully with GLP-1-analogues.

<p>Akrilamid je toksična hemijska supstanca koja ima vrlo &scaron;iroku primenu u hemijskoj industriji, a 2002. godine otkriveno je njegovo prisustvo u namirnicima bogatim skrobom koje se pripremaju na visokim temperaturama. U poslednjh desetak godina primećen je veliki porast gastrointestinalnih tegoba u ljudskoj populaciji. Cilj istraživanja bio je ispitati patohistolo&scaron;ke promene u tkivu želuca pacova soja Wistar izazvanih peroralnim aplikovanjem akrilamida i na taj način povući paralelu sa mogućim gastrointestinalnim tegobama nastalim kao posledica konzumiranja hrane bogate akrilamidom. U istraživanju je ispitivano 6 grupa od po 5 eksperimentalnih životinja (pacovi soja Wistar). Dve kontrolne grupe kojima je peroralno aplikovana destilovana voda i koje su žrtvovane posle 24h i 72h; dve eksperimentalne kojima je peroralno aplikovan akrilamid u dnevnoj dozi od 25 mg/kg i koje su žrtvovane posle 24h i 72h; dve eksperimentalne grupe kojima je peroralno aplikovan akrilamid u dnevnoj dozi od 50 mg/kg i koje su žrtvovane posle 24h i 72h. Na histolo&scaron;kom materijalu tkiva želuca primenjena je kvalitativna histolo&scaron;ka analiza pod svetlosnim mikroskopom, semikvantitativna procena tipa mucina u epitelnim ćelijama sluznice želuca, prisustvo limfocita i granulocita u sluznici želuca, stereolo&scaron;ka merenja pojedinih kompartmana zida želuca, linearna merenja broja i veličine ganglijskih ćelija u Maissner-ovom i Auerbach-ovom nervnom pleksusu, kao i broj mastocita u lamini propriji sluznice i podsluznici želuca. Dobijene vrednosti merenih parametara su potom statistički obrađene. Nastale promene na tkivu želuca pacova soja Wistar se ogledaju u vidu blagog direktnog o&scaron;tećenja povr&scaron;nog epitela sa propratnom blagom inflamatornom reakcijom i blagom degranulacijom mastocita. U Maissner-ovom i Auerbach-ovom nervnom pleksusu su smanjene volumenske gustine nervnih vlakana i ganglijskih ćelija, kao i broj i veličina ganglijskih ćelija. Direktno toksično delovanje na epitel dovodi do posledične obnove epitela, te je potvrđeno prisustvo nezrelijih oblika mukoproduktivnih ćelija koje sadrže kisele, AB pozitivne mucine. Ispitani inflamatorni i degenerativni parametri pokazuju pozitivnu korelaciju u odnosu na dozu i/ili dužinu ekspozicije akrilamidu. Primena akrilamida peroralno pokazala je da postoje patohistolo&scaron;ke promene na tkivu želuca u vidu direktnog toksičnog o&scaron;tećenja epitela, inflamatorne reakcije i o&scaron;tećenja nervnih pleksusa. Poznavanjem mehanizma delovanja ove toksične materije moguće je primeniti adekvatnu prevenciju u ishrani i izvr&scaron;iti odgovarajući izbor terapijskih metoda.</p> / <p>Acrylamide is a toxic chemical substance with wide implementation in chemical industry. In 2002 it was discovered the presence of acrylamide in foods rich in starch which are prepared at high temperatures. In the last ten years there is a large increase in gastrointestinal illnesses in human population. The aim of this study was to investigate the histopathological changes in the gastric tissue in Wistar rats induced with injection of oral acrylamide and thus draw a parallel with possible gastrointestinal problems arising as a result of the consumption of foods rich in acrylamide. The research was carried out 6 groups of 5 experimental animals (Wistar rats). Two control groups that are orally concomitant application of distilled water and which were sacrificed after 24h and 72h; two experimental groups which are orally administrated acrylamide in a daily dose of 25 mg / kg and that were sacrificed after 24h and 72h; two experimental groups which were orally administrated acrylamide in a daily dose of 50 mg / kg and that were sacrificed after 24h and 72h. On histological gastric tissue material is applied qualitative histological analysis by light microscopy, semi-quantitative assessment of the type of mucin in epithelial cells of the stomach lining, the presence of lymphocytes and granulocytes in gastric mucosa, stereological measurements of individual compartments of the stomach wall, linear measuring the number and size of ganglion cells in the Maissner and Auerbach&#39;s nerve plexus, and the number of mast cells in the lamina propria of the mucosa and in the submucosis of the stomach. Obtained values of measured parameters were statistically processed. Histological changes in the stomach tissue of Wistar rats are seen as a direct slight damage of the surface epithelium, with accompanynig mild inflammatory reaction and the degranulation of mast cells. The Meissner&#39;s and Auerbach&#39;s nerve plexus decreased volume density of nerve fibers and ganglion cells, as well as the number and size of the ganglion cells. Directly toxic effect on epithelium leads to the result of the reconstruction of the epithelium, which is confirmed by the presence of immature form of mucoproductive cells which contain acid, AB positive mucins. Examined inflammatory and degenerative parameters show a positive correlation with respect to dose and / or a time of exposition to acrylamide. Acrylamide oral application revealed that there are histologic changes in the stomach tissue in the form of a direct toxic damage to the epithelium, inflammatory reaction and damage to the nerve plexus. Knowing the mechanism of action of these toxic substances allows to apply adequate prevention in nutrition and make an appropriate choice of therapeutic methods.</p>

Streptococcus thermophilus est la bactérie lactique la plus utilisée après Lactococcus lactis dans l'industrie laitière pour la fabrication de yaourts et de fromages à pâte cuite (Emmental, gruyère), filée (Mozarella) ou pressée (Cheddar). Il s'agit du seul streptocoque à avoir le statut de bactérie GRAS (Generally Recognized As Safe). Dans un premier temps le potentiel probiotique de 30 souches de S. thermophilus de différentes origines a été testé par l'étude de leurs capacités à résister aux différentes conditions de stress rencontrées pendant leur passage dans le tractus gastro-intestinal (bas pH, sels biliaires et stress oxydant), de leur capacité à adhérer aux cellules épithéliales intestinales et de leurs propriétés immunomodulatrices. La majorité des souches réduit la production d'interleukine IL-8 (pro-inflammatoire) alors qu'elles induisent la production d'interleukine IL-10 (anti-inflammatoire) et l'IL-12 (pro-inflammatoire). Sur la base du rapport IL-10/IL-12, qui permet d'apprécier le potentiel anti-inflammatoire d'une souche, nous avons observé que certaines d'entre-elles pourraient avoir un fort potentiel anti-inflammatoire. L'Analyse en Composantes Principales (ACP) nous a permis de séparer les souches en 6 catégories différentes présentant des propriétés distinctes. A l'intérieur de chaque classe, une variabilité entre les souches a été observée et des caractéristiques intéressantes identifiées. Cependant, aucune des classes ne peut être considérée comme contenant le probiotique « parfait ». Suite à cette étude et sur la base de sa résistance aux stress gastriques et de ses capacités d'adhésion, et sachant que la séquence de son génome est disponible, la souche LMD-9 a été sélectionnée pour la deuxième partie de ce travail, à savoir la construction d'un outil basé sur la technologie R-IVET pour étudier l'état physiologique de S. thermophilus dans le tractus digestif. L'outil R-IVET mis au point se compose de deux éléments: un vecteur plasmidique portant le gène cre codant une recombinase spécifique dépourvu de son promoteur et une cassette chromosomique composée d'un gène de résistance à la spectinomycine flanqué par des sites loxP, reconnus par la recombinase Cre. La fonctionnalité de l'outil R-IVET a ensuite été testée par le clonage de trois promoteurs différents de S. thermophilus (PprtS, Pshsp and Plac) en amont de cre. Le système a été valide in vitro avec les trois promoteurs et in vivo avec le promoteur Plac / Streptococcus thermophilus is a lactic acid bacterium used after Lactococcus lactis in the dairy industry for the production of yogurt and cheeses like Emmental, Gruyere, Mozarella and Cheddar. It is the only streptococcus to have the GRAS (Generally Recognized As Safe) status. In this work, the probiotic potential of 30 S. thermophilus strains from different origins was tested by studying their ability to resist different stress conditions encountered during their passage through the GIT (low pH, bile salts and oxidative stress), their ability to adhere to intestinal epithelial cells and their immunomodulatory properties. Majority of the strains reduced the production of interleukin IL-8 (pro-inflammatory) and induced the production of interleukin IL-10 (anti-inflammatory) and IL-12 (pro-inflammatory). On the basis of the ratio IL-10/IL-12 which allows to evaluate the anti-inflammatory potential of a probiotic, several strains appeared to have a high the anti-inflammatory potential. Principal Component Analysis (PCA) allowed us to classify strains in 6 different categories with different properties. Within each class, variability and interesting features were observed, but none of the classes could be considered as containing the perfect probiotic. Following this study and on the basis of its resistance to gastric stress and its adhesion capacity and knowing that its genome sequence is available, the strain LMD-9 was selected for the second part of this work, namely the construction of a tool based on R-IVET to study the physiological state of S. thermophilus in the digestive tract. This tool is composed of two elements: a plasmid vector (pULNcreB), carrying the gene cre encoding a site-specific recombinase without its promoter and a chromosomal cassette composed of a gene of resistance to spectinomycin flanked by loxP sites which are recognized by the recombinase Cre. The functionality of the tool R-IVET was then tested by cloning three different promoters of S. thermophilus (PprtS, Pshsp and Plac) upstream of cre. System was valid in vitro with all the three promoters and in vivo by using the lactose operon promoter Plac

L’[alpha]-casozépine, décapeptide issu de l’hydrolysat trypsique de la caséine [alpha]s1, possède in vitro une affinité pour le site benzodiazépine des récepteurs GABAA centraux (Lecouvey et al., 1997 ; Miclo et al., 2001). Des résultats attestent du potentiel anxiolytique de ce décapeptide seul ou au sein de son hydrolysat, in vivo, en administration intrapéritonéale et per os chez le rat Wistar (Guesdon et al., 2006 ; Miclo et al., 2001 ; Violle et al., 2007) et chez l’Homme en administration orale (Kim et al., 2007 ; Messaoudi et al., 2005). Pour déterminer la sensibilité de l’[alpha]-casozépine à différentes attaques protéolytiques, des cinétiques d’hydrolyses gastriques, pancréatiques et intestinales, ont été réalisées in vitro avec les enzymes et systèmes enzymatiques suivants ; pepsine, trypsine, [alpha]-chymotrypsine, CorolasePP®, vésicules membranaires de bordure en brosse d’entérocytes de rats et épithélium entérocytaire reconstitué par les cellules de la lignée Caco-2. Pour chacune des conditions d’hydrolyses étudiées, l’ensemble des fragments peptidiques libérés ont été séparés par chromatographie liquide haute performance en phase inversée et caractérisés par spectrométrie de masse. Ces analyses mettent en évidence la résistance partielle de l’[alpha]-casozépine à certaines enzymes et systèmes enzymatiques. L’hydrolyse pepsique de l’[alpha]-casozépine libère notamment deux fragments : 91YLGYLEQ97 et 91YLGYL95, nommés F97 et F95 qui s’avèrent être plus résistants que [alpha]-casozépine à certaines enzymes employées. Ainsi pour permettre une meilleure compréhension de la relation entre la structure de l’[alpha]-casozépine et la fonction benzodiazépine mimétique obtenu in vivo, ces peptides tronqués dans leur partie carboxy-terminale ont été testés in vivo chez le rat Wistar en injection intrapéritonéale. Ils s’avèrent être également détenteurs d’activité anxiolytique. Cependant aucun transfert de l’[alpha]-casozépine et des fragments F97 et F95 n’a pu être observé au travers de l’épithélium entérocytaire reconstitué par le modèle cellulaire de la lignée Caco-2 et ce malgré les différentes conditions de cultures et d’analyses testées / [alpha]-Casozepine, a tryptic decapeptide from bovine [alpha]s1-casein, have in vitro an affinity for the benzodiazepine site of the central receptor GABAA (Lecouvey et al., 1997; Miclo et al., 2001). Some results display the anxiolytic potential of this peptide alone or within its hydrolysat, in vivo, in intra peritoneal administration and per os in Wistar rats (Guesdon et al., 2006; Miclo et al., 2001; Violle et al., 2007) and in human in oral administration (Kim et al., 2007; Messaoudi et al., 2005). To determine the sensitivity of [alpha]-casozepine in various proteolytic attacks, kinetics of gastric, pancreatic and intestinal hydrolyses, were realized in vitro with enzymes and following enzymatic systems; pepsin, trypsin, [alpha]-chymotrypsin, CorolasePPTm, brush border membranar vesicles rats enterocytes and enterocytes epithelium reconstituted by the Caco-2 cell line. For each of the conditions of studied hydrolysis, all the peptides fragments released were separated by liquid chromatography high-performance in inverted phase and characterized by mass spectrometry. These analyses underline the partial resistance of [alpha]-casozepine in some enzymes and enzymatic systems. [alpha]-Casozepine pepsic hydrolysis release in particular two peptide fragments: 91YLGYLEQ97 and 91YLGYL95, named F97 and F95 who turn out to be more resistant than [alpha]-casozepine in some used enzymes. So to allow a better understanding of the relation between structure of [alpha]-casozepine and the benzodiazepine mimetic function obtained in vivo, these peptides truncated in their carboxy-terminal party were tested in vivo with Wistar rat in intra peritoneal injection. They turn out to be also holders of anxiolytic activity. However no transfer of [alpha]-casozepine and fragments F97 and F95 was observed through the enterocyte epithelium reconstituted by the cellular model Caco-2 and this in spite of the various conditions of cultures and tested analyses