Análise da expressão de neurotrofinas durante a regeneração de nervo periférico de rato por enxerto venoso Enxertos de veias têm sido empregados para preencher lacunas em nervos periféricos transeccionados para melhor recuperação funcional. No entanto, vários inconvenientes, como a constrição do enxerto secundário foram observados. Uma nova alternativa para esta técnica foi desenvolvida. Simplesmente invertendo a veia de dentro para fora, chamado do Inside- out vein graft. As neurotrofinas são uma família de fatores neurotróficos conhecidos por desempenhar um papel significativo na regeneração de nervos periféricos. A família da neurotrofina é constituído por fator de crescimento nervoso (NGF), fator neurotrófico derivado do cérebro (BDNF), Neurotrofina-3 (NT-3) e Neurotrofina-4 (NT-4). No campo da neurobiologia, vários autores têm utilizado a técnica de PCR a fim de obter mais informações sobre os nervos regenerados. Neste estudo, foi utilizada a técnica de biologia molecular para explorar o papel e o nível das neurotrofinas durante a regeneração de nervos periféricos com enxerto de veia. O nervo isquiático de ratos foi seccionado e reparado com enxerto de veia invertida (IOVG) e técnicas de enxerto de veia padrão (SVG). No grupo controle, os ratos foram operados e o nervo isquiático foi mantido intacto. Os animais foram sacrificados após 6 e 12 semanas e os enxertos foram colhidos para observar o nível das neurotrofinas. Músculos EDL e Sóleo foram excisados e pesados para determinar a diferença de peso entre os grupos. Um pequeno segmento dos cotos distais de ambos os grupos SVG e IOVG também foram excisados e foram processados histologicamente para examinar a quantidade de axónios regenerados. Além disso, um outro pequeno segmento do coto distal foi processado para RT-PCR para analisar o nível das neurotrofinas nesta área.A tecnica do walk track analysis foi realizada para determinar o índice funcional do nervo isquiático nos grupos. Em 6 semanas, não ocorreu crescimento neuronal significativo no coto distal dos dois tipos de enxertos, porém um crescimento foi observado em 12 semanas. Não houve diferença significativa na massa muscular entre IOVG e SVG em ambos os períodos de tempo. No entanto, um aumento significativo na massa muscular foi observado a partir de 6 a 12 semanas nos grupos IOVG e SVG. Um aumento significativo na produção de NT-3 foi observado no grupo de SVG em ambos, enxerto e o coto distal quando comparados a partir de 6 a 12 semanas, no entanto, não houve aumento observado no nível de neurotrofinas dos outros tipos (NGF e NT-4) . Surpreendentemente, não houve aumento significativo da NT-3 no grupo IOVG. Conclui-se que, entre as neurotrofinas avaliadas neste estudo, não há nenhuma diferença significativa no seu nível de RNAm entre os dois grupos, exceto NT-3. Finalmente, uma vez que o nível de RNAm de NT-3 aumenta significativamente entre 6 e 12 semanas no grupo SVG e não no IOVG, observado por estas duas técnicas de nível molecular, estudos adicionais necessitam serem feitos para decifrar o mecanismo exato. / Vein grafts have been employed to bridge the gap in transected peripheral nerves to produce better functional recovery. However several disadvantages such as secondary graft constriction were observed and a new alternative to this technique was developed by simply reversing the vein inside out. Both inside out and standard vein grafts were successfully used in recovering the sensory segmental defect in humans. Neurotrophins are a family of eurotrophic factors known to play an important role in the regeneration of peripheral nerves. The neurotrophin family consists of Nerve Growth Factor (NGF), Brain Derived Neurotrophic Factor (BDNF), Neurotrophin-3 (NT-3) and Neurotropinh-4 (NT-4). In the neurobiology field, several authors have been using PCR technique in order to gain more information regarding regenerated nerves. In this study, we employed this molecular biology technique to explore the role and level of the neurotrophins during the peripheral nerve regeneration with vein graft. The sciatic nerve of rats were sectioned and repaired with Inside out vein graft (IOVG) and standard vein graft techniques (SVG). In the control group the rats were sham operated wherein the sciatic nerve was kept intact. The animals were euthanized at 6 and 12 weeks and the grafts were harvested to observe the level the neurotrophins. EDL and Sol muscles were excised and measured to determine any weight difference between the groups. A small segment of the distal stumps from both the SVG and IOVG groups were also excised and were subjected to histological process to examine the amount of regenerated axon. In addition, another small segment of the distal stump was processed for RT-PCR to further examine the level of the neurotrophins in this area. At 6 weeks, no significant neuronal growth was observed in the distal stump of both graft types but a distinct growth was seen at 12 weeks. Walk track analysis showed poor motor function recovery in the experimental groups during both time intervals. Morphometric analysis demonstrated no significant differences in the amount of myelination between both the groups. There was no significant difference in the muscle mass between IOVG and SVG in both time periods. However, a significant increase in both the muscle mass was observed from 6 to 12 weeks in the IOVG and SVG groups. A significant increase in the production of NT-3 was observed in SVG group in both the distal stump and graft segment when compared from 6 to 12 weeks; however there was no observed increase in the level of other neurotrophins (NGF and NT-4). Surprisingly, no significant increase of NT-3 was noticed in the IOVG group. We conclude that amongst the neurotrophins evaluated in this study, there is no significant difference in their mRNA level between both groups except NT-3. Also, since mRNA level of NT-3 increases significantly between 6 and 12 weeks in SVG and not in IOVG, it suggests that the mechanism by which these two techniques operate at a molecular level may differ and further studies need to be done to decipher the exact mechanism.

Background: Myocardial failure and dysfunction is not uncommon during critical illness and following cardiac surgery. For optimal treatment, a better understanding of the effects of inotropic drugs is needed. In this thesis, two non-adrenergic mediated inotropes, milrinone and levosimendan were studied in different models of myocardial dysfunction. The study aims were to assess the following: the effects of milrinone on blood flow in coronary artery bypass grafts during CABG surgery; the effects of milrinone on left ventricular diastolic function during post-ischaemic myocardial dysfunction; whether milrinone or levosimendan are protective or injurious during acute myocardial ischaemia, and if levosimendan potentiates myocardial function when added to milrinone in an experimental model of post-ischaemic (stunned) myocardium. Material and Methods: In Study I, 44 patients undergoing coronary artery bypass surgery(CABG) were included as subjects. Milrinone or saline was administrated in a single dose during cardio-pulmonary bypass (CPB) and coronary graft flow measurements were recorded after 10 and 30 min following CPB. In Study II; 24 patients undergoing CABG had estimations of peak ventricular filling rates made before and after CPB with administration of milrinone or saline as a single dose during CPB, performed by assessment of the rate of change in diastolic cross-sectional left ventricular area. In Study III, energy-metabolic effects of milrinone and levosimendan were measured in an anaesthetized porcine model during 45 minutes of regional myocardial ischemia. Microdialysis sampling of metabolites of local ischemic metabolism allowed assessment of glycolytic activity and the degree of myocardial calcium overload. In Study IV, in a porcine model of postischaemic myocardial stunning, ventricular pressure-volume relationships were analyzed when milrinone or a combination of milrinone and levosimendan were given together. Results: In Study I, there was a clear increase in non-sequential saphenous vein graft blood flow with milrinone at 10 minutes (64.5 ± 37.4 compared to placebo 43.6 ± 25.7 ml/min (mean ± SD).). A decreasing but still measureable flow increase was seen for milrinone at 30 minutes. In Study II, an increase in early left ventricular filling rate (ventricular cross-sectional area rate of change,dA/dt) was seen in the milrinone treated group. Pre-bypass milrinone group dA/dt 22.0 ± 9.5 changed to post-bypass values dA/dt 27.8 ± 11.5 cm2/sec). Placebo group pre-bypass dA/dt was 21.0 ± 8.7 and post-bypass 17.1 ± 7.1 cm2/sec. A milrinone effect was demonstrated in an adjusted regression model (p = 0.001). In Study III, neither milrinone nor levosimendan led to a change in energy-metabolic activity during ischemia as reflected by interstitial glucose, pyruvate, lactate orglycerol. Neither drug exacerbated the relative myocardial calcium overload during ischemia. In Study IV, milrinone improved active relaxation (tau) in post-ischemic stunned myocardium, but did not markedly improve systolic function by preload recruitable stroke work. Levosimendan added to milrinone showed minimal effect on active relaxation but a positive effect on systolic function in combination with milrinone. Conclusions: We conclude that milrinone treatment leads to an increase in blood flow in newly implanted coronary saphenous vein grafts, and improves ventricular relaxation post-cardiopulmonary bypass. Neither milrinone nor levosimendan, in this porcine model, negatively influence myocardial energy metabolism or calcium overload during acute ischaemia. Addition of levosimendan to milrinone treatment during post-ischaemic ventricular dysfunction may provide additive inotropic effects on systolic function but probably not for active relaxation.

No description available.

Análise da expressão de neurotrofinas durante a regeneração de nervo periférico de rato por enxerto venoso Enxertos de veias têm sido empregados para preencher lacunas em nervos periféricos transeccionados para melhor recuperação funcional. No entanto, vários inconvenientes, como a constrição do enxerto secundário foram observados. Uma nova alternativa para esta técnica foi desenvolvida. Simplesmente invertendo a veia de dentro para fora, chamado do Inside- out vein graft. As neurotrofinas são uma família de fatores neurotróficos conhecidos por desempenhar um papel significativo na regeneração de nervos periféricos. A família da neurotrofina é constituído por fator de crescimento nervoso (NGF), fator neurotrófico derivado do cérebro (BDNF), Neurotrofina-3 (NT-3) e Neurotrofina-4 (NT-4). No campo da neurobiologia, vários autores têm utilizado a técnica de PCR a fim de obter mais informações sobre os nervos regenerados. Neste estudo, foi utilizada a técnica de biologia molecular para explorar o papel e o nível das neurotrofinas durante a regeneração de nervos periféricos com enxerto de veia. O nervo isquiático de ratos foi seccionado e reparado com enxerto de veia invertida (IOVG) e técnicas de enxerto de veia padrão (SVG). No grupo controle, os ratos foram operados e o nervo isquiático foi mantido intacto. Os animais foram sacrificados após 6 e 12 semanas e os enxertos foram colhidos para observar o nível das neurotrofinas. Músculos EDL e Sóleo foram excisados e pesados para determinar a diferença de peso entre os grupos. Um pequeno segmento dos cotos distais de ambos os grupos SVG e IOVG também foram excisados e foram processados histologicamente para examinar a quantidade de axónios regenerados. Além disso, um outro pequeno segmento do coto distal foi processado para RT-PCR para analisar o nível das neurotrofinas nesta área.A tecnica do walk track analysis foi realizada para determinar o índice funcional do nervo isquiático nos grupos. Em 6 semanas, não ocorreu crescimento neuronal significativo no coto distal dos dois tipos de enxertos, porém um crescimento foi observado em 12 semanas. Não houve diferença significativa na massa muscular entre IOVG e SVG em ambos os períodos de tempo. No entanto, um aumento significativo na massa muscular foi observado a partir de 6 a 12 semanas nos grupos IOVG e SVG. Um aumento significativo na produção de NT-3 foi observado no grupo de SVG em ambos, enxerto e o coto distal quando comparados a partir de 6 a 12 semanas, no entanto, não houve aumento observado no nível de neurotrofinas dos outros tipos (NGF e NT-4) . Surpreendentemente, não houve aumento significativo da NT-3 no grupo IOVG. Conclui-se que, entre as neurotrofinas avaliadas neste estudo, não há nenhuma diferença significativa no seu nível de RNAm entre os dois grupos, exceto NT-3. Finalmente, uma vez que o nível de RNAm de NT-3 aumenta significativamente entre 6 e 12 semanas no grupo SVG e não no IOVG, observado por estas duas técnicas de nível molecular, estudos adicionais necessitam serem feitos para decifrar o mecanismo exato. / Vein grafts have been employed to bridge the gap in transected peripheral nerves to produce better functional recovery. However several disadvantages such as secondary graft constriction were observed and a new alternative to this technique was developed by simply reversing the vein inside out. Both inside out and standard vein grafts were successfully used in recovering the sensory segmental defect in humans. Neurotrophins are a family of eurotrophic factors known to play an important role in the regeneration of peripheral nerves. The neurotrophin family consists of Nerve Growth Factor (NGF), Brain Derived Neurotrophic Factor (BDNF), Neurotrophin-3 (NT-3) and Neurotropinh-4 (NT-4). In the neurobiology field, several authors have been using PCR technique in order to gain more information regarding regenerated nerves. In this study, we employed this molecular biology technique to explore the role and level of the neurotrophins during the peripheral nerve regeneration with vein graft. The sciatic nerve of rats were sectioned and repaired with Inside out vein graft (IOVG) and standard vein graft techniques (SVG). In the control group the rats were sham operated wherein the sciatic nerve was kept intact. The animals were euthanized at 6 and 12 weeks and the grafts were harvested to observe the level the neurotrophins. EDL and Sol muscles were excised and measured to determine any weight difference between the groups. A small segment of the distal stumps from both the SVG and IOVG groups were also excised and were subjected to histological process to examine the amount of regenerated axon. In addition, another small segment of the distal stump was processed for RT-PCR to further examine the level of the neurotrophins in this area. At 6 weeks, no significant neuronal growth was observed in the distal stump of both graft types but a distinct growth was seen at 12 weeks. Walk track analysis showed poor motor function recovery in the experimental groups during both time intervals. Morphometric analysis demonstrated no significant differences in the amount of myelination between both the groups. There was no significant difference in the muscle mass between IOVG and SVG in both time periods. However, a significant increase in both the muscle mass was observed from 6 to 12 weeks in the IOVG and SVG groups. A significant increase in the production of NT-3 was observed in SVG group in both the distal stump and graft segment when compared from 6 to 12 weeks; however there was no observed increase in the level of other neurotrophins (NGF and NT-4). Surprisingly, no significant increase of NT-3 was noticed in the IOVG group. We conclude that amongst the neurotrophins evaluated in this study, there is no significant difference in their mRNA level between both groups except NT-3. Also, since mRNA level of NT-3 increases significantly between 6 and 12 weeks in SVG and not in IOVG, it suggests that the mechanism by which these two techniques operate at a molecular level may differ and further studies need to be done to decipher the exact mechanism.

Background: Current guidelines give a class I recommendation to use of embolic protection devices (EPD) for saphenous vein graft (SVG) intervention; however, studies have shown conflicting results. The objective of this meta-analysis is to compare all-cause mortality, major adverse cardiovascular events, myocardial infarction (MI), or target vessel revascularization in SVG intervention with and without EPD. Methods and Results: Literature was searched through October 2016. Eight studies (n=52 893) comparing SVG intervention performed with EPD (n=11 506) and without EPD (n=41 387) were included. There was no significant difference in all-cause mortality (odds ratio [OR], 0.79; confidence interval [CI], 0.55–1.12; P=0.19), major adverse cardiovascular events (OR, 0.73, CI, 0.51–1.05; P=0.09), target vessel revascularization (OR, 1.0; CI, 0.95–1.05; P=0.94), periprocedural MI (OR, 1.12; CI, 0.65–1.90, P=0.69), and late MI (OR, 0.80; CI, 0.52–1.23; P=0.30) between the 2 groups. Sensitivity analysis excluding CathPCI Registry study showed no difference in periprocedural MI, late MI, and target vessel revascularization; however, it favored EPD use in all-cause mortality and major adverse cardiovascular events. Further sensitivity analysis including only observational studies revealed no difference in all-cause mortality, major adverse cardiovascular events, target vessel revascularization, and late MI. Additional analysis after excluding CathPCI Registry study revealed no difference in outcomes. Conclusions: This study including 52 893 patients suggests no apparent benefit in routine use of EPD during SVG intervention in the contemporary real-world practice. Further randomized clinical trials are needed in current era to evaluate long-term outcomes in routine use of EPD, and meanwhile, current guideline recommendations on EPD use should be revisited.

No description available.

The final publication is available at http://dx.doi.org/10.1016/j.jtcvs.2013.11.054. Motoaki Ohnaka, Akira Marui, Kenichi Yamahara, Kenji Minakata, Kazuhiro Yamazaki, Motoyuki Kumagai, Hidetoshi Masumoto, Shiro Tanaka, Tadashi Ikeda, Ryuzo Sakata, Effect of microRNA-145 to prevent vein graft disease in rabbits by regulation of smooth muscle cell phenotype, The Journal of Thoracic and Cardiovascular Surgery, Volume 148, Issue 2, August 2014, Pages 676-682.e2, ISSN 0022-5223. / 京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18544号 / 医博第3937号 / 新制||医||1006(附属図書館) / 31444 / 京都大学大学院医学研究科医学専攻 / (主査)教授 木村 剛, 教授 野田 亮, 教授 瀬原 淳子 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第21638号 / 医博第4444号 / 新制||医||1034(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 木村 剛, 教授 浅野 雅秀, 教授 山下 潤 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

yes / Type 2 diabetes (T2DM) confers increased risk of endothelial dysfunction, coronary heart disease, and vulnerability to vein graft failure after bypass grafting, despite glycaemic control. This study explored the concept that endothelial cells (EC) cultured from T2DM and nondiabetic (ND) patients are phenotypically and functionally distinct. Cultured human saphenous vein- (SV-) EC were compared between T2DM and ND patients in parallel. Proliferation, migration, and in vitro angiogenesis assays were performed; western blotting was used to quantify phosphorylation of Akt, ERK, and eNOS. The ability of diabetic stimuli (hyperglycaemia, TNF-α, and palmitate) to modulate angiogenic potential of ND-EC was also explored. T2DM-EC displayed reduced migration (~30%) and angiogenesis (~40%) compared with ND-EC and a modest, nonsignificant trend to reduced proliferation. Significant inhibition of Akt and eNOS, but not ERK phosphorylation, was observed in T2DM cells. Hyperglycaemia did not modify ND-EC function, but TNF-α and palmitate significantly reduced angiogenic capacity (by 27% and 43%, resp.), effects mimicked by Akt inhibition. Aberrancies of EC function may help to explain the increased risk of SV graft failure in T2DM patients. This study highlights the importance of other potentially contributing factors in addition to hyperglycaemia that may inflict injury and long-term dysfunction to the homeostatic capacity of the endothelium.

EINLEITUNG: Die externe Stabilisierung von Venengrafts soll die Scherkräfte auf die Venenwand vermindern und dadurch die Ausbildung einer Neointimaproliferation reduzieren. In experimentellen Modellen wurde diese Hypothese im Kurzzeitversuch überprüft. Es fanden sich in diesem Zeitraum widersprüchliche Ergebnisse. Ziel unserer Untersuchung war es, in einem neuen Modell der arterialisierten segmentalen Vena-jugularis-Transposition auf die infrarenale Aorta den Einfluss einer externen Fibrinkleberstabilisierung auf die Neointimabildung des Venengrafts im Langzeitversuch darzustellen. MATERIAL UND METHODEN: Männlichen Wistar-Ratten wurden Segmente der Vena jugularis entnommen und in Flussrichtung, nach Entfernen eines Aortensegmentes, in die infrarenale Aorta eingebracht. Somit entspricht dieses einem Venenbypassmodell mit komplett arterialisiertem Venengraft. Vor