Decellularization of human umbilical arteries / Descelularización de las arterias umbilicales humanas

SUMMARY: Arterial obstruction in small diameter (<6 mm) vessels are many times treated with grafts, however autologous aren't always available and synthetic have a high rate of complications. Decellularization of umbilical arteries may provide a solution, but the ideal method is debatable. We compare effectiveness between SDS and Triton X-100. Umbilical cords obtained from full term pregnancies with normal development and no evident complications in the newborn, were micro-dissected within 12 h and stored in phosphate buffered saline without freezing. Arteries were then processed for decellularization using 0.1 % and 1 % SDS, and 1 % Triton X100 protocols. Evaluation of cellular and nuclear material, collagen fibers, elastic fibers, and glycosoaminoglycans of the extracellular matrix (ECM) were evaluated as well as morphometric analysis under histological and immunohistochemical techniques. Triton X-100 was ineffective, preserving nuclear remains identified by immunofluorescence, had the most notable damage to elastic fibers, and decrease in collagen. SDS effectively eliminated the nuclei and had a less decrease in elastic fibers and collagen. Laminin was preserved in all groups. No significant differences were identified in luminal diameters; however the middle layer decreased due to decellularization of muscle cells. In conclusion, 0.1 % SDS decellularization was the most effective in eliminating cells and preserving the main components of the ECM.

RESUMEN: La obstrucción arterial en vasos de pequeño diámetro (<6 mm) se trata muchas veces con injertos, sin embargo, los autólogos no siempre están disponibles y los sintéticos tienen una alta tasa de complicaciones. La descelularización de las arterias umbilicales puede proporcionar una solución, pero el método ideal es discutible. Comparamos la efectividad entre los métodos SDS y Triton X-100. Cordones umbilicales obtenidos a partir de embarazos a término con evolución normal y sin complicaciones evidentes del recién nacido, se microdiseccionaron en 12 horas y se almacenaron en solución salina con fosfato sin congelación. Las arterias se procesaron luego para la descelularización usando los protocolos de SDS al 0,1 % y 1 %, y Triton X-100 al 1 %. Se realizó la evaluación de material celular y nuclear, fibras de colágeno, fibras elásticas y glucosoaminoglicanos de la matriz extracelular (MEC), así como el análisis morfométrico bajo técnicas histológicas e inmunohistoquímicas. Triton X-100 fue ineficaz, conservando los restos nucleares identificados por inmunofluorescencia, tuvo el daño más notable a las fibras elásticas y la disminución del colágeno. SDS efectivamente eliminó los núcleos y tuvo una disminución menor en las fibras elásticas y el colágeno. Laminina fue preservado en todos los grupos. No se identificaron diferencias significativas en los diámetros luminales; sin embargo, la capa media disminuyó debido a la descelularización de las células musculares. la descelularización con SDS al 0,1 % fue la más efectiva para eliminar células y preservar los principales componentes de la MEC.

Human umbilical artery smooth muscle exhibits a 2-APB-sensitive capacitative contractile response evoked by vasoactive substances and expresses mRNAs for STIM, Orai and TRPC channels

After depletion of intracellular Ca2+ stores the capacitative response triggers an extracellular Ca2+ influx through store-operated channels (SOCs) which refills these stores. Our objective was to explore if human umbilical artery smooth muscle presented this response and if it was involved in the mechanism of serotonin- and histamine-induced contractions. Intracellular Ca2+ depletion by a Ca2+-free extracellular solution followed by Ca2+ readdition produced a contraction in artery rings which was inhibited by the blocker of Orai and TRPC channels 2-aminoethoxydiphenyl borate (2-APB), suggesting a capacitative response. In presence of 2-APB the magnitude of a second paired contraction by serotonin or histamine was significantly less than a first one, likely because 2-APB inhibited store refilling by capacitative Ca2+ entry. 2-APB inhibition of sarcoplasmic reticulum Ca2+ release was excluded because this blocker did not affect serotonin force development in a Ca2+-free solution. The PCR technique showed the presence of mRNAs for STIM proteins (1 and 2), for Orai proteins (1, 2 and 3) and for TRPC channels (subtypes 1, 3, 4 and 6) in the smooth muscle of the human umbilical artery. Hence, this artery presents a capacitative contractile response triggered by stimulation with physiological vasoconstrictors and expresses mRNAs for proteins and channels previously identified as SOCs.

Dopplervelocimetria do ducto venoso na predicão da acidemia fetal / Ductus venosus Doppler velocimetry to predict acidemia at birth in pregnancies with placental insufficiency

OBJETIVOS: Investigar a possibilidade da predicão da acidemia no nascimento mediante dopplervelocimetria do ducto venoso e definir qual o melhor parâmetro e seus pontos de corte nessa predicão em gestacões com insuficiência placentária. MÉTODOS: Trata-se de estudo transversal e prospectivo que analisou 47 gestacões únicas com insuficiência placentária e idade gestacional superior a 26 semanas, realizado no Hospital São Paulo (UNIFESP) e na Maternidade-Escola Assis Chateaubriand (UFC). A insuficiência placentária foi diagnosticada quando o índice de pulsatilidade da artéria umbilical encontrava-se acima do percentil 95 para a idade gestacional estimada. Fetos com anomalias estruturais ou cromossômicas foram excluídos. O doppler foi realizado a menos de 24 horas do parto. A amostra de sangue da artéria umbilical foi coletada imediatamente após o nascimento para análise da gasometria. Diagnosticou-se acidemia quando o pH encontrava-se abaixo de 7,20 na ausência de trabalho de parto e abaixo de 7,15 quando parto vaginal. Foram consideradas patológicas as acidemias metabólicas ou mistas. Construiu-se curva ROC para as velocidades S, D e A e para o IPV e as relacões S/A e (S-A)/S do DV (variáveis independentes) e acidemia (variável dependente). O teste de MacNemar foi utilizado para comparar os parâmetros entre si. RESULTADOS: As velocidades absolutas S, D e A mostraram ser pobres preditoras da acidemia no nascimento. O IPV mostrou ser bom preditor de acidemia (área sob a curva ROC 0,79, p=0,003). As relacões S/A e (S-A)/S também mostraram ser boas preditoras da acidemia (área sob a curva ROC 0,818, p=0,001). Os pontos de corte calculados foram: IPV = 0,76, S/A = 2,67 e (S-A)/S = 0,63. CONCLUSÕES: Os índices ângulo-independentes do doppler do DV mostraram excelente correlacão com acidemia no nascimento nesta populacão. Não houve diferenca estatisticamene significativa entre estes parâmetros.

Aferição da taxa de eritroblastos e dos parâmetros do equilíbrio ácido-básico no sangue da veia umbilical no nascimento: capacitação da enfermagem na aplicação da metodologia / Measurement of the erythroblast level and acid-base paramethers in the umbilical vein blood at delivery: nursing qualification in the methodology aplication

Objetivo: Identificar os fatores que dificultam a aferição da taxa dos eritroblastos e dos parâmetros do estado ácido-básico no sangue da veia umbilical e avaliar as repercussões de um treinamento sobre a efetividadeda metodologia. Pacientes e Métodos: Ensaio clínico observacional, com 1204 parturientes divididas em três amostras...

Vascular prostacyclin production in andean women with pregnancy-induced hypertension

Although prostacyclin (PGI2) production in the umbilical artery is known to be reduced in prognancy-induced hypertension (PIH), little information is available about its production in maternal vascular tissues. We measured 6-keto-prostaglandin F 1* generation in the umbiblical and comentum arteries of 24 Andean women divided into three groups: 1) 8 normal pregnant women, 2) 8 cases with clinical evidence of severe PIH, and 3) 8 normotensive non-pregnant women.The normal pregnant group (232 ñ 172 pg mg-1 2h-1) and the non-pregnant control group (237 ñ 146 pg mg-1 2h-1 0 showed similar PGI2 production in the omentum arteries, whereas the PIH group showed lower PGI2 generation (P<0.05) than the normal patients both in the umbilical (697 ñ 377 vs 1528 ñ 291 pg mg-1 2h-1) and omentum (132 ñ 73 vs 232 ñ 172 pg mg-1 2h-1) arteries. PGI2 production was 6.8 times lower in the omentum arteries than in the umbilical arteries. The data confirm and extend the view of the occurrence of reduced PGI2 production in the maternal-fetal vascular tissues of women with severe PIH.