O treinamento de caminhada (TC) é recomendado para o tratamento de indivíduos com claudicação intermitente (CI) porque melhora a capacidade de caminhada. Além disso, seria interessante que o TC também promovesse modificações benéficas no sistema cardiovascular, pois os eventos cardiovasculares são a principal causa de morte nestes indivíduos. No entanto, os efeitos cardiovasculares do TC em indivíduos com CI foi pouco estudado. Assim, o objetivo deste estudo foi verificar o efeito do TC sobre o risco, a função e a regulação cardiovasculares em indivíduos com CI. Quarenta e dois indivíduos com CI foram divididos de forma aleatória em 2 grupos: controle (GC, n=20, sessões de 30 min de alongamento) e treinamento de caminhada (GT, n=22, 15 séries de 2 min de caminhada em intensidade correspondente à frequência cardíaca (FC) do limiar de dor intercalados por 2 min de repouso passivo). Nos dois grupos, a intervenção foi realizada 2 vezes/semana durante 12 semanas consecutivas. No início e ao final do estudo, os indivíduos realizaram as seguintes avaliações: glicemia e perfil lipídico de jejum; índice de massa corporal; capacidade de caminhada; consumo de oxigênio no 1º estágio e no pico do teste de esforço; índice tornozelo-braço (ITB) de repouso; janela isquêmica após teste ergoespirométrico; pressão arterial (PA) em repouso (auscultatória) e de 24 horas (oscilométrica); débito cardíaco (DC - reinalação de CO2); FC (ECG); volume sistólico (VS); resistência vascular (RV) sistêmica, do antebraço e da perna (plestismografia); componentes de alta (AF) e baixa (BF) frequência da variabilidade da FC; e sensibilidade barorreflexa espontânea (SBR). Mudanças significantes (P<0,05) ao longo do tempo e entre os grupos foram verificadas pela análise de variância ANOVA de dois fatores para medidas repetidas. O TC aumentou significantemente a capacidade de caminhada (&Delta;=+302±85m) e a SBR (&Delta;=+2.13±1.07 ms/mmHg), e diminuiu o VO2) no 1º estágio do teste (&Delta;=-1,8±0,4ml.kg-1.min-1), a janela isquêmica (&Delta;=- 0,40±0,38mmHg.min.m-1), a PA média (&Delta;=-5±2mmHg), a variabilidade da PA média de 24h (&Delta;=-0,8±0,2mmHg), o DC (&Delta;=-0.37±0.24L/min), a FC (&Delta;=- 4±2bpm), a RV do antebraço (&Delta;=-8.5±2.8 U) e a razão BF/AF (&Delta;=-1.24±0.99). A glicemia, o perfil lipídico, o índice de massa corporal, o VO2) pico, o ITB de repouso e a RV sistêmica e da perna não foram modificadas pelo TC. Não houve mudança em nenhuma variável no GC. Em conclusão, o TC melhorou a capacidade de caminhada, a economia de caminhada e a janela isquêmica. Além disso, o TC melhorou a função (PA, DC, FC e RV antebraço) e a regulação (BF/AF e SBR) cardiovasculares em indivíduos com CI. Estas alterações fornecem suporte adicional para a utilização do TC no tratamento de indivíduos com CI / Walking training (WT) is recommended for the treatment of patients with intermittent claudication (IC) because it improves walking capacity. Moreover, it would be interesting that WT also promotes beneficial changes on cardiovascular system, since cardiovascular events are the main causes of death in these patients. Nevertheless, the effects of WT on cardiovascular system in patients with IC have been poorly studied. Thus, the objective of this study was to investigate the effects of WT on cardiovascular risk, function and regulation in patients with IC. Forty-two IC patients were randomly divided into 2 groups: Control (CG, n=20, 30 min of stretching exercises) and walking training (TG, n=22, 15 sets of 2:2-min walk:rest at the heart rate (HR) of pain threshold). In both groups, the intervention was performed twice/week for 12 consecutive weeks. At the beginning and end of the study, the following measured were done: fasting glycemia and lipid profile; body mass index; walking capacity; VO2 at the first stage and the peak of a treadmill test; ankle brachial index (ABI); ischemic window after maximal test; resting (auscultatory) and 24-hour (oscillometric) blood pressure (BP); cardiac output (CO - CO2 rebreathing); heart rate (HR - ECG); stroke volume (SV); systemic, forearm and leg vascular resistance (VR - plethysmography); low- (LF) and high-frequency (HF) components of HR variability; and spontaneous baroreflex sensitivity (SBS). Significant changes (P<0.05) over time and between groups were assessed by 2-way ANOVA for repeated measures. WT significantly increased walking capacity (&Delta;=+302±85m) and SBS (&Delta;=+2.13±1.07 ms/mmHg), and decreased VO2 at the first stage of treadmill test (&Delta;=-1.8±0.4ml.kg-1.min-1), ischemic window (&Delta;=-0.40±0.38mmHg.min.m-1), mean BP (&Delta;=-5±2mmHg), ambulatory mean BP variability (&Delta;=-0,8±0,2 mmHg), CO (&Delta;=-0.37±0.24 L/min), HR (&Delta;=- 4±2bpm), forearm VR (&Delta;=-8.5±2.8 U) and LF/HF (&Delta;=-1.24±0.99). Glycemia, lipid profile, body mass index, VO2 peak, ABI, systemic and leg VR were unchanged following WT. There was no significant change for any variable in CG. In conclusion, WT enhanced walking capacity, walking economy and ischemic window. In addition, WT improved cardiovascular function (BP, CO, HR and forearm VR) and autonomic regulation (LF/HF, SBS) in patients with IC. These changes provide further support for the use of regular WT in treating patients with IC

Tyrimo objektas. 27–90 metų asmenys, sergantys pirmojo ir antrojo tipo diabetu ir periferinių arterijų liga, gydyti Vilniaus universiteto kraujagyslių chirurgijos centre Vilniaus miesto universitetinėje ligoninėje 1997–2011 m. Tyrimo tikslas. Nustatyti Vilniaus universiteto kraujagyslių chirurgijos centre Vilniaus miesto universitetinėje ligoninėje gydytų pacientų, sergančių diabetu, periferinių arterijų ligos ypatumus, PAL padarinius ir PAL padarinių rizikos veiksnius. Tyrimo medžiaga ir metodai. Išanalizuotos 925 pacientų, sergančių 1 ir 2 tipo diabetu ir 1997–2011 metais gydytų Vilniaus universiteto kraujagyslių chirurgijos centre Vilniaus miesto universitetinėje ligoninėje nuo periferinių arterijų ligos istorijos. Statistinė analizė atlikta SPSS 19.0 for Windows programų paketu. Pasirinktas statistinio reikšmingumo lygmuo &#945;=0,05. Rezultatai. Ištyrėme 387 moterų ir 538 vyrų, duomenis. Vidutinis tiriamųjų amžius buvo 67,99±9,47 metų. 95,6 proc. tiriamųjų sirgo 2 tipo diabetu. Vidutinė sirgimo diabetu trukmė buvo 12,95±9,91 metų. Diabetinę angiopatiją turėjo 47,8 proc. tiriamųjų, nefropatiją – 34,9 proc., retinopatiją – 14,6 proc., polineuropatiją – 33,7 proc.. Statistiškai reikšmingai dažniau nefropatija, retinopatija ir polineuropatija buvo nustatyta pacientams, sergantiems 1 tipo diabetu. Pacientai dažniausiai sirgo širdies ir kraujagyslių sistemos ligomis. 21,2 proc. tiriamųjų jau buvo patyrę galūnių amputacijas. Vidutinė hospitalizacijos trukmė buvo 17,3±10,80... [toliau žr. visą tekstą] / Object of the research. 27–90 years old patients having the type 1 and type 2 diabetes mellitus and peripheral artery disease, who were treated at the Vilnius University Vascular Surgery Center in the Vilnius town University Hospital in the year 1997–2011. The aim of our research was. To analyse aspects of peripheral artery disease in patients with diabetes mellitus, who were cured at Vilnius University Vascular Surgery Center in the Vilnius town University Hospital as well as to determine the consequenses of PAD and the risk factors PAD. Material and methods of the research. 925 cases of the type 1 and type 2 diabetes mellitus were explored at the Vilnius University Vascular Surgery center in the Vilnius town University hospital in the year 1997–2011, who received treatment for the peripheral artery disease. The statistic survey has been done using the program pack SPSS 19.0 for Windows. Statistic importance level &#945;=0.05. Results. Data about 378 women and 538 men have been explored. An average age of all the patients was 67.99±9.47 years. 95.6% of patients had the type 2 diabetes mellitus. An average duration of having disease was 12.95±9.91 years, 47.8% of cases had diabetic angiopathy, 34.9% of patients had nephropaty, 14.6% of cases had retinopathy, 33.7% of all the cases had polyneuropathy. According to the statistic importance rates retinopathy, nephropathy, polyneuropathy are more often among patients with the type 1 diabetes mellitus. Cardiovascular diseases were... [to full text]

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology and Toxicology Student: Jan Juhász Supervisor: Prof. MUDr. Radomír Hrdina, CSc. Title of diploma thesis: Treatment of lower extremity peripheral artery disease The lower extremities ischemia is a disease caused most often by atherosclerosis during which the lumen in lower limb arteries becomes narrow. Its prevalence is increasing, especially in the developed countries. The disease can be asymptomatic and symptomatic. The symptoms are very unpleasant and decrease patient's quality of life. Advanced stages of the disease may be life threatening. Therefore, it is vital to timely and correctly diagnose the illness. During the therapy, it is possible to use pharmacological as well as non-pharmacological procedures, and, preferably, a combination of the two types of treatment. The pharmacotherapy can be divided into several parts. The prevention of atherosclerotic complications makes use of preventive measures and antiplatelet therapy to reduce the cardiovascular risk. The symptoms therapy focuses on improving patients' quality of life by prescribing the vasoactive medications cilostazol, naftidrofuryl or pentoxifylin. The critical limb ischemia therapy uses prostaglandin analogues alprostadil, iloprost, limaprost or...

A Doença da Artéria Periférica (DAP) é o resultado do processo aterosclerótico das artérias dos membros superiores, inferiores, aorta abdominal e seus ramos viscerais. Nosso objetivo foi detectar a DAP correlacionando-a com fatores de risco (FR) e a filtração glomerular estimada (FGe) nos pacientes com &#8805;30 anos, assistidos na Unidade docente - assistencial de Angiologia (UDA) do Hospital Universitário Pedro Ernesto da Faculdade de Ciências Médicas - UERJ, com o intuito de descrever os FR e associação com doença renal crônica à DAP, a partir da aferição do índice tornozelo-braço (ITB); determinar a alteração da FGe por equações, relacionando-a à progressão da DAP. Foi usado um questionário padrão e o ITB para identificar os pacientes com e sem DAP. Correlacionou-se as variáveis laboratoriais, como os níveis séricos de colesterol, triglicerídeos, HDL-c, LDL-c, glicemia, homocisteína com a FGe e com o ITB. As análises estatísticas foram feitas pelo programa Statistical Package for the Social Sciences (SPSS) 16.0 Os resultados apontaram para a importância do ITB no diagnóstico da DAP, com configuração de graus de obstrução leve, discreta, moderada e grave para os sintomáticos, e a identificação dos assintomáticos, possibilitando intervenção nos fatores de risco demarcados e o controle de suas complicações. O tabagismo mostou-se como o FR com razão de risco mais importante para DAP. A hipertensão sistólica e a diastólica foram variáveis clínicas mais significativas que o diabetes mellitus. Os marcadores séricos tradicionais para DAP: colesterol total, triglicerídeos e glicemia mostraram significância estatística. A homocisteína foi o marcador mais significativo em relação à DAP. Ocorreu associação entre redução do ITB com a elevação dos níveis pressóricos, das glicemias, da homocisteína, assim como diminuição das médias da FGe. Foi conclído que nos pacientes com DAP a hiperhomocisteinemia está associada à diminuição da FGe, ambas passíveis de prevenção, contribuindo na redução da morbimortalidade da DAP. A estreita associação da DAP com a FGe diminuída representou relevante contribuição do estudo. / Peripheral Artery (DAP) is the result of the atherosclerotic process involving the arteries of the superior and inferior limbs, abdominal aorta and its visceral branches. The objective was detect PAD, using the ankle brachial index (ABI), in patients &#8805; 30 years old attended in the Unidade Docente - Assistencial of Angiology (UDA), correlating it with risk factors (RF) and estimated glomerular filtration rate (eGFR), with intention to describe the RF and association with chronic kidney disease. It was used a standard questionnaire and the ABI to identify patients with and without PAD. Laboratorial tests, as total cholesterol, triglycerides, HDL-c, LDL-c, glycemia, creatinine and homocysteine were correlated to ABI. Statistical analyses were done using the Statistical Package for the Social Sciences (SPSS) 16.0 program. The results had pointed to the importance of the ABI in the diagnosis of PAD, with degrees of mild, discrete, moderate and serious stenosis for the symptomatic patients, and the identification of the asymptomatic ones, making possible intervention in the RF and control of their complications. Tabagism was confirmed as the RF with most important odds ratio for PAD. The systolic and diastolic hypertension showed to be more significant than diabetes mellitus, as diseases associated to PAD. In laboratorial evaluation, the traditional blood markers for PAD: total cholesterol, triglycerides and glucose had shown statistics significance. Homocysteine was the marker most significant in PAD. Association between reduction of ABI with systolic and diastolic hypertension and glycemias occurred, as well as reduction of the averages of the eGFR. The conclusion was, in patients with PAD, hyperhomocysteinemia and decrease of eGFR are possible of prevention, contributing in the reduction of the morbimortality of PAD. The narrow association of decrease eGFR in patients with PAD represented excellent contribution of this study.

A Doença da Artéria Periférica (DAP) é o resultado do processo aterosclerótico das artérias dos membros superiores, inferiores, aorta abdominal e seus ramos viscerais. Nosso objetivo foi detectar a DAP correlacionando-a com fatores de risco (FR) e a filtração glomerular estimada (FGe) nos pacientes com &#8805;30 anos, assistidos na Unidade docente - assistencial de Angiologia (UDA) do Hospital Universitário Pedro Ernesto da Faculdade de Ciências Médicas - UERJ, com o intuito de descrever os FR e associação com doença renal crônica à DAP, a partir da aferição do índice tornozelo-braço (ITB); determinar a alteração da FGe por equações, relacionando-a à progressão da DAP. Foi usado um questionário padrão e o ITB para identificar os pacientes com e sem DAP. Correlacionou-se as variáveis laboratoriais, como os níveis séricos de colesterol, triglicerídeos, HDL-c, LDL-c, glicemia, homocisteína com a FGe e com o ITB. As análises estatísticas foram feitas pelo programa Statistical Package for the Social Sciences (SPSS) 16.0 Os resultados apontaram para a importância do ITB no diagnóstico da DAP, com configuração de graus de obstrução leve, discreta, moderada e grave para os sintomáticos, e a identificação dos assintomáticos, possibilitando intervenção nos fatores de risco demarcados e o controle de suas complicações. O tabagismo mostou-se como o FR com razão de risco mais importante para DAP. A hipertensão sistólica e a diastólica foram variáveis clínicas mais significativas que o diabetes mellitus. Os marcadores séricos tradicionais para DAP: colesterol total, triglicerídeos e glicemia mostraram significância estatística. A homocisteína foi o marcador mais significativo em relação à DAP. Ocorreu associação entre redução do ITB com a elevação dos níveis pressóricos, das glicemias, da homocisteína, assim como diminuição das médias da FGe. Foi conclído que nos pacientes com DAP a hiperhomocisteinemia está associada à diminuição da FGe, ambas passíveis de prevenção, contribuindo na redução da morbimortalidade da DAP. A estreita associação da DAP com a FGe diminuída representou relevante contribuição do estudo. / Peripheral Artery (DAP) is the result of the atherosclerotic process involving the arteries of the superior and inferior limbs, abdominal aorta and its visceral branches. The objective was detect PAD, using the ankle brachial index (ABI), in patients &#8805; 30 years old attended in the Unidade Docente - Assistencial of Angiology (UDA), correlating it with risk factors (RF) and estimated glomerular filtration rate (eGFR), with intention to describe the RF and association with chronic kidney disease. It was used a standard questionnaire and the ABI to identify patients with and without PAD. Laboratorial tests, as total cholesterol, triglycerides, HDL-c, LDL-c, glycemia, creatinine and homocysteine were correlated to ABI. Statistical analyses were done using the Statistical Package for the Social Sciences (SPSS) 16.0 program. The results had pointed to the importance of the ABI in the diagnosis of PAD, with degrees of mild, discrete, moderate and serious stenosis for the symptomatic patients, and the identification of the asymptomatic ones, making possible intervention in the RF and control of their complications. Tabagism was confirmed as the RF with most important odds ratio for PAD. The systolic and diastolic hypertension showed to be more significant than diabetes mellitus, as diseases associated to PAD. In laboratorial evaluation, the traditional blood markers for PAD: total cholesterol, triglycerides and glucose had shown statistics significance. Homocysteine was the marker most significant in PAD. Association between reduction of ABI with systolic and diastolic hypertension and glycemias occurred, as well as reduction of the averages of the eGFR. The conclusion was, in patients with PAD, hyperhomocysteinemia and decrease of eGFR are possible of prevention, contributing in the reduction of the morbimortality of PAD. The narrow association of decrease eGFR in patients with PAD represented excellent contribution of this study.

Gene therapy techniques delivering exogenous angiogenic growth factors, such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (FGF-2), are currently being investigated as potential treatments for ischemia resulting from a variety of conditions, such as peripheral artery disease (PAD) and chronic wounds. Despite these intense efforts, a viable clinical option to promote therapeutic neovascularization remains elusive. Electroporation is a simple in vivo method to deliver normally impermeable molecules, such as plasmid DNA, to a variety of tissues including skin and muscle. This study investigated intradermal injection of plasmids encoding angiogenic growth factors with electroporation as a novel therapeutic approach to increase perfusion in areas of ischemia. Two common animal models of ischemia were employed: a skin flap model, used to study wound healing, and a hindlimb ischemia model, used to investigate potential therapies for PAD. In the skin flap model, delivery of plasmid VEGF with electroporation significantly increased VEGF expression for 5 days after delivery compared to injection of the plasmid alone. While the increase in VEGF expression was short-term, it significantly increased expression of the downstream angiogenic growth factor endothelial nitric oxide synthase, as well as perfusion and healing in the distal area of the skin flap. To facilitate the translation of electroporation to the clinic, a novel electrode configuration was previously designed for cutaneous delivery of plasmids to a large surface area. The design of the Multielectrode Array allows for delivery to a large surface area without the need to increase the applied voltage. Conditions for plasmid delivery with this electrode were optimized and it was then utilized to deliver plasmid FGF-2 (pFGF) to the hindlimb ischemia model. FGF-2 expression, perfusion, and angiogenesis were assessed. FGF-2 expression was significantly higher for 10 days after treatment with pFGF with electroporation compared to injection of pFGF alone. This increase in FGF-2 expression induced a significant increase in perfusion and angiogenesis in the ischemic limb. The research presented here suggests intradermal injection of plasmids encoding angiogenic factors by electroporation is a novel potential therapeutic approach to increase perfusion to areas of ischemia and promote wound healing.

Peripheral Artery Disease is a very serious disease characterized by an arterial occlusion due to atherosclerotic plaques. In response to an arterial occlusion, arteriogenesis occurs, causing smooth muscle cells to transition from a contractile to synthetic state. Also following an arterial occlusion, functional impairment was seen in the collateral circuit. An immunofluorescence protocol was developed in order to assess the impact of collateral enlargement (arteriogenesis) on smooth muscle phenotype at various time points. Smooth muscle α-actin was used to mark all smooth muscle cells, Ki-67 was used to label proliferating smooth muscle cells, and a fluorescent nuclear stain was used to quantify the number of cells present. Samples of the profunda femoris and gracilis were dissected from each mouse hind limb (one ligated, one sham) at three different time points: 3 days, 7 days, and 14 days after a femoral artery ligation surgery. Smooth muscle cell phenotype and luminal cross-sectional area were assessed in the profunda femoris and the midzone of the gracilis collaterals. Smooth muscle cells were proliferating at 3 and 7 days following the occlusion in the gracilis collaterals and significant collateral vessel growth was observed over the two week period. No proliferation was observed in the profunda femoris and although there was an increasing trend in vessel size over the two week period, the averages were not significantly different. The phenotypic transition of the smooth muscle cells was not the cause of vascular impairment in the collateral circuit. This shows that further research is needed to characterize impairment in the collateral circuit.

Rationnel : L’implication établie des progéniteurs endothéliaux circulants dans les phénomènes de néovascularisation chez l’adulte a stimulé la recherche de thérapeutiques angiogènes basées sur la greffe de ces cellules. Deux types cellulaires au phénotype endothélial sont actuellement définis entre autres par leur cinétique d’apparition en culture : les progéniteurs précoces (CFU-EC ou CAC) et tardifs (ECFC). Notre équipe a montré que l’injection thérapeutique de cellules mononucléées de moelle osseuse (BM-MNC) permettait la néovascularisation du site ischémié chez des patients atteints d’artériopathie des membres inférieurs, et que les néovaisseaux formés avaient le phénotype d’ECFC. Nous avons dans un premier temps mesuré les concentrations de différentes protéines modulant l’angiogenèse, chez des patients atteints de pathologies ischémiques et cardiovasculaires, ou impliquant des anomalies vasculaires associées à la fibrose. Ainsi, le transforming growth factor - β1 (TGF-β1) dans la fibrose pulmonaire idiopathique (FPI), la thrombospondine-1 (TSP-1) dans l’artériopathie des membres inférieurs (AMI), et le placental growth factor (PlGF) chez les patients atteints de pathologies cardiovasculaires [syndrome coronarien aigu (SCA), ou patients devant subir une chirurgie de la valve ou un pontage coronarien], se sont distingués comme potentiel biomarqueur plasmatique dans ces pathologies, et ont été étudiés dans la biologie des ECFC humaines.Résultats : Le taux plasmatique de TGF-β1 est augmenté chez les patients atteints de FPI par rapport à la population contrôle ; il a un effet pro-angiogène in vivo (vascularisation des implants de Matrigel®) et in vitro (prolifération et migration des ECFC) via les récepteurs ALK-1, ALK-5 et TGF-βRII. Le taux plasmatique de TSP-1 est augmenté chez les patients artéritiques par rapport à la population contrôle. Par ailleurs les néovaisseaux formés de patients artéritiques ayant été traités par injection locale de BM-MNC expriment la TSP-1. Dans les modèles murins de Matrigel®-plugs et d’ischémie du membre inférieur (IMI), la TSP-1 induit une diminution de la vascularisation des implants ainsi qu’une diminution de la revascularisation du membre ischémié. In vitro, la TSP-1 augmente l’adhésion via un mécanisme N-Terminal dépendant, et diminue le potentiel angiogène (prolifération et migration) des ECFC via sa liaison au récepteur CD47, ce qui active la voie de signalisation SDF-1/CXCR4. Le taux plasmatique de PlGF est augmenté chez les patients atteints de SCA par rapport à 2 populations contrôles ; il est également augmenté chez les patients ayant subit une chirurgie cardiaque. Les PlGF-1 et -2 potentialisent la tubulogenèse des ECFC in vitro via la phosphorylation du récepteur VEGFR1. Cet effet est aboli lorsque le VEGFR1 est inhibé par ARN interférence ou par le composé chimique « 4321 ». De plus ce composé « 4321 » inhibe la vascularisation des implants de Matrigel®, ainsi que la revascularisation du membre ischémié dans le modèle d’IMI.Conclusions : Le TGF-β1 joue un rôle dans le remodelage vasculaire de la FPI via les ECFC ; la TSP-1 est un potentiel biomarqueur de l’angiogenèse induite par les ECFC dans l’AMI ; l’inhibition de la voie PlGF/VEGFR1 module la tubulogenèse induite par les ECFC, cellules impliquées dans la formation de nouveaux vaisseaux. Nous avons ainsi mis en évidence 3 protéines modulant l’angiogenèse dans 3 contextes pathologiques différents, caractérisés par un remodelage vasculaire et où les ECFC sont impliquées dans leurs mécanismes physiopathologiques. Ces 3 protéines se présentent donc comme de potentiels biomarqueurs plasmatiques, modulant les propriétés angiogènes des ECFC et pouvant influencer leur efficacité en tant que produit de thérapie cellulaire. Ces protéines jouent un rôle probable dans l’équilibre homéostatique au décours des pathologies concernées. / Rationale: The pro-angiogenic capacities of endothelial progenitor cells are now well established, and their involvement in neovascularization events in adults has stimulated the research in the field of angiogenic therapy based on transplant of these cells. Current data converge towards the notion of two cell types with endothelial phenotype, defined at least by their kinetics of appearance in culture: early endothelial progenitor cells (CFU-EC or CAC) and late (ECFC). Our team has shown that the therapeutic injection of bone marrow mononuclear cells (BM-MNC) led to neovascularization of the ischemic site in patients with critical limb ischemia, and that the new vessels formed bore the phenotype of ECFC. We initially measured the concentrations of different proteins modulating angiogenesis in patients with ischemic and cardiovascular diseases, or involving vascular abnormalities associated with fibrosis. Thus, the transforming growth factor - ß1 (TGF-ß1) in idiopathic pulmonary fibrosis, the thrombospondin-1 (TSP-1) in peripheral artery disease, and the placental growth factor (PlGF) in patients with cardiovascular diseases [acute coronary syndrome (ACS), patients undergoing valve surgery or coronary artery bypass surgery], emerged as potential plasmatic biomarkers in these pathological settings, and have been studied in the biology of human ECFC.Results: TGF-ß1 plasma level is increased in patients with idiopathic pulmonary fibrosis (IPF) compared to the control population; it exerts a pro-angiogenic effect in vivo (vascularization of Matrigel ®-plugs) and in vitro (proliferation and migration of ECFC) via ALK-1, ALK-5 and TGF-ßRII receptors. TSP-1 plasma level is increased in patients with peripheral artery disease (PAD) compared to the control population. In addition, the new vessels formed in PAD patients treated by local injection of BM-MNC express TSP-1. In murine models of Matrigel ®-plugs and hindlimb ischemia, TSP-1 induces a decrease in plugs vascularization and impaired revascularization of ischemic limb. In vitro, TSP-1 increases ECFC adhesion via an N-terminal dependent mechanism and reduces their angiogenic potential (proliferation and migration) via its binding to CD47 receptor, which activates the SDF-1/CXCR4 signaling pathway. PlGF plasma level is increased in ACS patients compared with the control population and stable angina patients and is also increased in patients undergoing cardiac surgery. PlGF-1 and -2 potentiate ECFC tubulogenesis in vitro via phosphorylation of the VEGFR1 receptor. This effect was abolished when the ECFC VEGFR1 is inhibited by RNA interference or by the chemical compound "4321". In addition this compound "4321" inhibits the vascularization of Matrigel ®-plugs, and revascularization of the ischemic limb in the hindlimb ischemia model.Conclusions: TGF-ß1 is involved in the IPF vascular remodeling through ECFC; TSP-1 is a potential biomarker of angiogenesis induced by ECFC in PAD; the inhibition of the PlGF/VEGFR1 pathway modulates ECFC tubulogenesis, cells involved in the formation of new vessels. We thus identified three proteins that modulate angiogenesis in three different pathological settings characterized by a vascular remodeling and where ECFC are involved in their pathophysiology. These three proteins therefore state as potential plasmatic biomarkers, modulating ECFC angiogenic properties and are able to influence their efficacy as a cell therapy product. These plasmatic biomarkers likely play a role in the homeostasis of those pathologies progress.

<p>Cardiovascular disease (CVD) is the leading cause of death worldwide and is expected to increase direct medical costs in the U.S. to $749 billion by the year 2035. Diagnosis of CVD through imaging techniques can improve our understanding of CVD progression and its associated risks through visualization of anatomical features and biological constituents. Non-invasive imaging relies on optimal image quality for visualization of such tissue structures that can be difficult to identify and segment. While various imaging modalities are used to determine tissue characteristics, many lack the spatial resolution that optics-based imaging can provide, which can assess hemodynamic parameters in preclinical models of ischemic disease. Acoustic-based imaging can complement optics-based imaging by providing anatomical and location-specific information of tissues with greater penetration depth. Even with all the advancements in imaging technology, however, limitations still exist in non-invasively, efficiently, and accurately capturing biologically relevant information with adequate spatial and temporal resolution. Furthermore, reproducible feature extraction is difficult due to a lack of standardization in the field, making it difficult to implement when image quality varies. In this work, we implement spatial frequency domain imaging (SFDI), ultrasound, and photoacoustic imaging in preclinical models of 1) peripheral artery disease, 2) traumatic brain injury, and 3) myocardial ischemia to capture imaging biomarkers of vascular and cardiac health in longitudinal studies. We also implement deep learning on preclinical ultrasound and photoacoustic images of the cardiac left ventricle to automatically extract regions of interest to calculate radial strain and oxygen saturation. Eventually findings from this work may help improve clinical cardiovascular disease diagnosis, prognosis, and treatment.</p>

Functional vasodilation in arterioles is impaired with chronic ischemia. We sought to examine the impact of chronic ischemia and age on skeletal muscle resistance artery function. To examine the impact of chronic ischemia, the femoral artery was resected from young (2-3mo) and adult (6-7mo) mice and the profunda femoris artery diameter was measured at rest and following gracilis muscle contraction 14 days later using intravital microscopy. Functional vasodilation was significantly impaired in ischemic mice (14.4±4.6% vs. 137.8±14.3%, p<0.0001 n=8) and non-ischemic adult mice (103.0±9.4% vs. 137.8±14.3%, p=0.05 n=10). In order to analyze the cellular mechanisms of the impairment, a protocol was developed to apply pharmacological agents to the experimental preparation while maintaining tissue homeostasis. Endothelial and smooth muscle dependent vasodilation were impaired with ischemia, 39.6 ± 13.6% vs. 80.5 ± 11.4% and 43.0 ± 11.7% vs. 85.1 ± 10.5%, respectively. From this data, it can be supported that smooth muscle dysfunction is the reason for the observed impairment in arterial vasodilation.