Résumé
ABSTRACT Purpose: To compare cryopreserved human corneal endothelial cells (HCECs) grown in human serum-supplemented media (HS-SM) with cryopreserved HCECs grown in fetal bovine serum-supplemented media (FBS-SM). Methods: Three pairs of human corneas from donors aged 8, 28, and 31 years were obtained from the eye bank. From each pair, one cornea was used to start a HCEC culture using HS-SM; the other cornea was grown in FBS-SM. On reaching confluence, the six cell populations were frozen using 10% dimethyl sulfoxidecontaining medium. Thawed cells grown in HS-SM were compared with those grown in FBS-SM with respect to morphology, growth curves, immunohistochemistry, real time-reverse transcriptase polymerase chain reaction (RT-PCR) for endothelial cell markers, and detachment time. Results: No difference in morphology was observed for cells grown in the two media before or after cryopreservation. By growth curves, cell counts after thawing were similar in both media, with a slight trend toward higher cell counts in FBS-SM. Cells grown in both the media demonstrated a similar expression of endothelial cell markers when assessed by immunohistochemistry, although HCEC marker gene expression was higher in cells grown in HS-SM than in those grown in FBS-SM as assessed by RT-PCR. With FBS-SM, there was a tendency of longer detachment time and lower cell passages. Conclusions: HS-SM was similar to FBS-SM for cryopreservation of cultured HCECs as assessed by analysis of cell morphology, proliferation, and protein expression, although marker gene expression was higher in cells grown in HS-SM than in those grown in FBS-SM. Detachment time was longer with FBS-SM and in lower passages.
RESUMO Objetivo: Comparar células endoteliais de córnea humana (HCECs) criopreservadas e cultivadas em meio suplementado com soro humano (HS-SM) com HCEC criopreservadas e cultivadas em meio suplementado com soro bovino fetal (FBS-SM). Métodos: Três pares de córneas humanas de doadores com 8, 28 e 31 anos de idade foram obtidos do banco de olhos e, de cada par, uma córnea foi utilizado para iniciar uma cultura com HS-SM e outra com FBS-SM. Ao atingir a confluência, as populações de células foram congeladas utilizando-se dimetil-sulfóxido 10% no respectivo meio de cultura. Após descongeladas, as células cultivadas em HS-SM foram comparados com as cultivadas em FBS-SM por meio de morfologia, curva de crescimento, imuno-histoquímica, reação em cadeia de Reação em cadeia da polimerase da transcrição reversa em tempo real (RT-PCR) para marcadores de células endoteliais e tempo de descolamento. Resultado: Não foram observadas diferenças morfológicas antes ou após a criopreservação. Curva de crescimento mostrou contagens celulares semelhantes em ambos os meios, com discreta tendência para um maior número em FBS-SM. As células cultivadas em ambos os meios mostraram expressão semelhante de marcadores celulares endoteliais quando avaliadas por imuno-histoquímica, embora a expressão genética de marcadores para HCEC tenha sido maior em HS-SM quando avaliado por RT-PCR. Houve uma tendência de maior tempo de descolamento com FBS-SM e passagens iniciais. Conclusões: HS-SM foi semelhante ao FBS-SM na criopreservação de HCEC cultivadas in vitro quando avaliadas por morfologia celular, proliferação celular e expressão proteica, embora a expressão genética de marcadores endoteliais tenha sido maior em células cultivadas em HS-SM quando comparadas a células cultivadas em FBS-SM. O tempo de descolamento foi maior quando utilizado FBS-SM e em passagens iniciais.
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Adulte , Animaux , Bovins , Enfant , Humains , Techniques de culture cellulaire/méthodes , Cryoconservation/méthodes , Cellules endothéliales/cytologie , Endothélium de la cornée/cytologie , Sérum , Numération cellulaire , Milieux de culture conditionnés , Prolifération cellulaire/effets des médicaments et des substances chimiques , Cellules cultivées/effets des médicaments et des substances chimiques , Expression des gènes , Immunohistochimie , RT-PCR , Statistique non paramétrique , Facteurs tempsRésumé
OBJECTIVES: Experimental studies on sepsis have demonstrated that ethyl pyruvate is endowed with antioxidant and anti-inflammatory properties. This study aimed to investigate the effects of ethyl pyruvate on leukocyte-endothelial interactions in the mesenteric microcirculation in a live Escherichia coli-induced sepsis model in rats. METHODS: Male Wistar rats were administered an intravenous suspension of E. coli bacteria or were subjected to a sham procedure. Three hours after bacterial infusion, the rats were randomized into the following groups: a control group without treatment, a group treated with lactated Ringer’s solution (4 mL/kg, i.v.), and a group treated with lactated Ringer’s solution (4 mL/kg, i.v.) plus ethyl pyruvate (50 mg/kg). At 24 h after bacterial infusion, leukocyte-endothelial interactions were investigated using intravital microscopy, and the expression of P-selectin and intercellular adhesion molecule-1 was evaluated via immunohistochemistry. White blood cell and platelet counts were also determined at baseline and 3 h and 24 h after E. coli inoculation. RESULTS: The non-treated and lactated Ringer’s solution-treated groups exhibited increases in the numbers of rolling leukocytes (∼2.5-fold increase), adherent cells (∼3.0-fold), and migrated cells (∼3.5-fold) compared with the sham group. In contrast, treatment with Ringer’s ethyl pyruvate solution reduced the numbers of rolling, adherent and migrated leukocytes to the levels observed in the sham group. Additionally, the expression of P-selectin and intercellular adhesion molecule-1 was significantly increased on mesenteric microvessels in the non-treated group compared with the sham group (p<0.001). The expression of both adhesion molecules was reduced in the other groups, with ethyl pyruvate being more effective than lactated Ringer’s solution. Infusion of bacteria caused significant leukopenia (3 h), followed ...
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Animaux , Mâle , Rats , Communication cellulaire/effets des médicaments et des substances chimiques , Cellules endothéliales/effets des médicaments et des substances chimiques , Leucocytes/effets des médicaments et des substances chimiques , Veines mésentériques/effets des médicaments et des substances chimiques , Pyruvates/pharmacologie , Sepsie/traitement médicamenteux , Communication cellulaire/physiologie , Modèles animaux de maladie humaine , Infections à Escherichia coli , Cellules endothéliales/cytologie , Leucocytes/cytologie , Microcirculation , Veines mésentériques/cytologie , Rat WistarRésumé
Objective: To investigate the respiratory and postural adaptations associated with mouth and nasal breathing and to evaluate the associations of such adaptations in mouth breathers' self-perceived quality of life. Method: Cross-sectional study with mouth breathers (initial n=116 and final n=48) and nasal breathers (initial n=131 and final n=24) from elementary school, aged between 7 and 14 years. Chest expansion, using cirtometry, the breathing pattern and the use of accessory muscles, by means of clinical evaluations and photogrammetry, and flexibility tests were evaluated in both groups. Subsequently, the mouth breathers were asked to complete the quality of life questionnaire. Statistical tests: Chi-square, odds ratio, Mann-Whitney, and binomial tests were first applied followed by logistic regressions. Results: Thoracic breathing (p=0.04), using of accessory muscles (p=0.03) and reductions in flexibility (p=0.001) increased the chances of an individual being a mouth breather when compared to nasal breathers. Subsequently, using of accessory muscles decreased the chances of snoring among mouth breathers (p=0.03); the presence of shoulder asymmetry reduced the chances of experiencing quiet sleep (p=0.05) and increased the chances of coughing or being tired when playing or running (p=0.008). Finally, forward head position reduced the chances of waking up at night (p=0.04) and experiencing shortness of breath (p=0.05). Conclusions: Respiratory and postural adaptations increased the chances of individuals persisting with mouth breathing. Additionally, these adaptations could be associated with mouth breathers' self-perceived quality of life. .
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Animaux , Femelle , Mâle , Souris , Grossesse , Techniques cytologiques/méthodes , Cellules endothéliales/cytologie , Séparation immunomagnétique/méthodes , Prosencéphale/cytologie , Néovascularisation physiologique , Prosencéphale/vascularisation , Prosencéphale/embryologieRésumé
The purpose of this study was to elucidate the origin and cellular composition of retrocorneal membranes (RCMs) associated with chemical burns using immunohistochemical staining for primitive cell markers. Six cases of RCMs were collected during penetrating keratoplasty. We examined RCMs with hematoxylin and eosin (H&E), periodic acid-Schiff (PAS) staining and immunohistochemical analysis using monoclonal antibodies against hematopoietic stem cells (CD34, CD133, c-kit), mesenchymal stem cells (beta-1-integrin, TGF-beta, vimentin, hSTRO-1), fibroblasts (FGF-beta, alpha-smooth muscle actin), and corneal endothelial cells (type IV collagen, CD133, VEGF, VEGFR1). Histologic analysis of RCMs revealed an organized assembly of spindle-shaped cells, pigment-laden cells, and thin collagenous matrix structures. RCMs were positive for markers of mesenchymal stem cells including beta-1-integrin, TGF-beta, vimentin, and hSTRO-1. Fibroblast markers were also positive, including FGF-beta and alpha-smooth muscle actin (SMA). In contrast, immunohistochemical staining was negative for hematopoietic stem cell markers including CD34, CD133 and c-kit as well as corneal endothelial cell markers such as type IV collagen, CD133 except VEGF and VEGFR1. Pigment-laden cells did not stain with any antibodies. The results of this study suggest that RCMs consist of a thin collagen matrix and fibroblast-like cells and may be a possible neogenetic structure produced from a lineage of bone marrow-derived mesenchymal stem cells.
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Adulte , Sujet âgé , Femelle , Humains , Mâle , Adulte d'âge moyen , Antigènes CD/métabolisme , Cornée/cytologie , Cytokines/métabolisme , Cellules endothéliales/cytologie , Fibroblastes/cytologie , Cellules souches hématopoïétiques/cytologie , Immunohistochimie , Protéines et peptides de signalisation intercellulaire/métabolisme , Cellules souches mésenchymateuses/cytologie , Cellules souches/cytologieRésumé
OBJECTIVE: We aimed to assess the chemotactic response of endothelial progenitor cells to angiotensin-converting enzyme inhibitors in T2DM patients after acute myocardial infarction, as well as the associated prognosis. METHODS: Sixty-eight T2DM patients with acute myocardial infarction were randomized to either receive or not receive daily oral perindopril 4 mg, and 36 non-diabetic patients with acute myocardial infarction were enrolled as controls. The numbers of circulating CD45−/low+CD34+CD133+KDR+ endothelial progenitor cells, as well as the stromal cell-derived factor-α and high-sensitivity C reactive protein levels, were measured before acute percutaneous coronary intervention and on days 1, 3, 5, 7, 14, and 28 after percutaneous coronary intervention. Patients were followed up for 6 months. Chinese Clinical Trial Registry: ChiCTR-TRC-12002599. RESULTS: T2DM patients had lower circulating endothelial progenitor cell counts, decreased plasma vascular endothelial growth factor and α levels, and higher plasma high-sensitivity C reactive protein levels compared with non-diabetic controls. After receiving perindopril, the number of circulating endothelial progenitor cells increased from day 3 to 7, as did the plasma levels of vascular endothelial growth factor and stromal cell-derived factor-α, compared with the levels in T2DM controls. Plasma high-sensitivity C reactive protein levels in the treated group decreased to the same levels as those in non-diabetic controls. Furthermore, compared with T2DM controls, the perindopril-treated T2DM patients had lower cardiovascular mortality and occurrence of heart failure symptoms (p<0.05) and better left ventricle function (p<0.01). CONCLUSIONS: The use of angiotensin-converting enzyme inhibitors represents a novel approach for improving cardiovascular repair after acute myocardial infarction in T2DM patients. .
Sujets)
Sujet âgé , Femelle , Humains , Mâle , Adulte d'âge moyen , Inhibiteurs de l'enzyme de conversion de l'angiotensine/usage thérapeutique , /complications , Cellules endothéliales/effets des médicaments et des substances chimiques , Mobilisation de cellules souches hématopoïétiques , Infarctus du myocarde/traitement médicamenteux , Périndopril/usage thérapeutique , Cellules souches/effets des médicaments et des substances chimiques , Protéine C-réactive/analyse , /sang , /sang , Cellules endothéliales/cytologie , Études de suivi , Infarctus du myocarde/sang , Infarctus du myocarde/complications , Pronostic , Études prospectives , Cellules souches/cytologie , Facteur de croissance endothéliale vasculaire de type A/sangRésumé
OBJECTIVES: The onset of chronic subdural hematoma may be associated with direct or indirect minor injuries to the head or a poorly repaired vascular injury. Endothelial progenitor cells happen to be one of the key factors involved in hemostasis and vascular repair. This study was designed to observe the levels of endothelial progenitor cells, white blood cells, platelets, and other indicators in the peripheral blood of patients diagnosed with chronic subdural hematoma to determine the possible relationship between the endothelial progenitor cells and the occurrence, development, and outcomes of chronic subdural hematoma. METHOD: We enrolled 30 patients with diagnosed chronic subdural hematoma by computer tomography scanning and operating procedure at Tianjin Medical University General Hospital from July 2009 to July 2011. Meanwhile, we collected 30 cases of peripheral blood samples from healthy volunteers over the age of 50. Approximately 2 ml of blood was taken from veins of the elbow to test the peripheral blood routine and coagulation function. The content of endothelial progenitor cells in peripheral blood mononuclear cells was determined by flow cytometry. RESULTS: The level of endothelial progenitor cells in peripheral blood was significantly lower in preoperational patients with chronic subdural hematomas than in controls. There were no significant differences between the two groups regarding the blood routine and coagulation function. However, the levels of circulating endothelial progenitor cells were significantly different between the recurrent group and the non-recurrent group. CONCLUSIONS: The level of circulating endothelial progenitor cells in chronic subdural hematoma patients was significantly lower than the level in healthy controls. Meanwhile, the level of endothelial progenitor cells in recurrent patients was significantly lower than the level in patients without recurrence. Endothelial progenitor cells may ...
Sujets)
Sujet âgé , Femelle , Humains , Mâle , Adulte d'âge moyen , Cellules endothéliales/cytologie , Hématome subdural chronique/sang , Cellules souches/cytologie , Coagulation sanguine , Marqueurs biologiques/analyse , Études cas-témoins , Numération cellulaire , Loi du khi-deux , Cytométrie en flux , Récidive , Valeurs de référence , Facteurs sexuelsRésumé
Devices for the rotational culture of cells and the study of biological reactions have been widely applied in tissue engineering. However, there are few reports exploring the effects of rotational culture on cell morphology, nitric oxide (NO) production, and cell cycle of the endothelial cells from human umbilical vein on the stent surface. This study focuses on these parameters after the cells are seeded on the stents. Results showed that covering of stents by endothelial cells was improved by rotational culture. NO production decreased within 24 h in both rotational and static culture groups. In addition, rotational culture significantly increased NO production by 37.9% at 36 h and 28.9% at 48 h compared with static culture. Flow cytometry showed that the cell cycle was not obviously influenced by rotational culture. Results indicate that rotational culture may be helpful for preparation of cell-seeded vascular grafts and intravascular stents, which are expected to be the most frequently implanted materials in the future.
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Humains , Cycle cellulaire , Cellules endothéliales/cytologie , Monoxyde d'azote/métabolisme , Ingénierie tissulaire/méthodes , Techniques de culture cellulaire , Division cellulaire , Cellules cultivées , Endothélium vasculaire/cytologie , Cellules épithéliales/cytologie , Cellules endothéliales de la veine ombilicale humaine , Modèles statistiques , Facteurs tempsRésumé
An increased plasma concentration of von Willebrand factor (vWF) is detected in individuals with many infectious diseases and is accepted as a marker of endothelium activation and prothrombotic condition. To determine whether ExoU, a Pseudomonas aeruginosa cytotoxin with proinflammatory activity, enhances the release of vWF, microvascular endothelial cells were infected with the ExoU-producing PA103 P. aeruginosa strain or an exoU-deficient mutant. Significantly increased vWF concentrations were detected in conditioned medium and subendothelial extracellular matrix from cultures infected with the wild-type bacteria, as determined by enzyme-linked immunoassays. PA103-infected cells also released higher concentrations of procoagulant microparticles containing increased amounts of membrane-associated vWF, as determined by flow cytometric analyses of cell culture supernatants. Both flow cytometry and confocal microscopy showed that increased amounts of vWF were associated with cytoplasmic membranes from cells infected with the ExoU-producing bacteria. PA103-infected cultures exposed to platelet suspensions exhibited increased percentages of cells with platelet adhesion. Because no modulation of the vWF mRNA levels was detected by reverse transcription-polymerase chain reaction assays in PA103-infected cells, ExoU is likely to have induced the release of vWF from cytoplasmic stores rather than vWF gene transcription. Such release is likely to modify the thromboresistance of microvascular endothelial cells.
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Humains , Protéines bactériennes/métabolisme , Cellules endothéliales/microbiologie , Endothélium vasculaire/microbiologie , Pseudomonas aeruginosa/métabolisme , Facteur de von Willebrand/métabolisme , Cellules cultivées , Cellules endothéliales/cytologie , Cellules endothéliales/métabolisme , Endothélium vasculaire/cytologie , Endothélium vasculaire/métabolisme , Adhésivité plaquettaireRésumé
O exercício físico mobiliza Células Progenitoras Endoteliais (CPE) para o sangue periférico. Entretanto, esse efeito parece depender de características do exercício, como duração e intensidade. O objetivo do presente trabalho foi verificar, por meio de revisão sistemática, o impacto de uma única sessão de exercício aeróbico sobre a mobilização de CPE em indivíduos sadios e os potenciais mecanismos envolvidos. A busca bibliográfica foi realizada nas bases de dados eletrônicas SciELO, LILACS, Cochrane, ClinicalTrials.gov, SPORTDiscus e Medline, em maio de 2011. Dos 178 estudos inicialmente identificados, 12 atenderam aos critérios de inclusão e foram classificados quanto à qualidade mediante critérios da escala PEDro. A magnitude e a duração da resposta de mobilização das CPE foram maiores após a realização de exercícios de longa/ultralonga duração e estão correlacionadas com níveis plasmáticos de fator de crescimento endotélio vascular (VEGF). O pico de mobilização dessas células em resposta a uma sessão de exercício máximo ou submáximo, com duração de até uma hora, ocorre no período imediatamente após o esforço até uma hora após sua interrupção. Um possível mecanismo é a biodisponibilidade do Óxido Nítrico (NO). A idade dos indivíduos e a intensidade do exercício parecem interferir na resposta de mobilização das CPE. Exercícios de longa/ultralonga duração promovem mobilização mais acentuada das CPE quando comparados a exercícios máximos ou submáximos. Os mecanismos envolvem a liberação do VEGF em exercícios de longa/ultralonga duração e a biodisponibilidade de NO em exercícios máximo e submáximo com até uma hora de duração.
Physical exercise mobilizes endothelial progenitor cells (EPCs) to peripheral blood. However, this effect seems to depend on exercise characteristics, such as duration and intensity. The aim of this systematic review was to verify the impact of a single bout of aerobic exercise on the mobilization of EPCs in healthy individuals, and the potential mechanisms involved. The bibliographic search was conducted on the following electronic databases in May 2011: SciELO, LILACS, Cochrane, ClinicalTrials.gov, SPORTDiscus and Medline. Of the 178 articles initially identified, 12 met the inclusion criteria and were classified regarding quality according to the PEDro scale. The magnitude and duration of the EPC mobilization response were higher after long/ultralong duration exercises, and they are correlated with vascular endothelial growth factor (VEGF) plasma levels. The EPC mobilization peak in response to a maximal or submaximal single bout of exercise lasting up to one hour occurs immediately after the exercise or within the first hour after it. One possible mechanism is nitric oxide (NO) bioavailability. The individuals' age and exercise intensity seem to interfere with the EPC mobilization response. Long/ultralong duration exercises promote more pronounced EPC mobilization as compared with maximal or submaximal exercises. The mechanisms involve VEGF release in long/ultralong duration exercises and NO bioavailability in maximal or submaximal exercises lasting less than one hour.
El ejercicio físico moviliza Células Progenitoras Endoteliales (CPE) hacia la sangre periférica. Entre tanto, ese efecto parece depender de características del ejercicio, como duración e intensidad. El objetivo de este estudio fue verificar, por medio de revisión sistemática, el impacto de una única sesión de ejercicio aeróbico sobre la Movilización de CPE en individuos sanos y los potenciales mecanismos envueltos. La búsqueda bibliográfica fue realizada en las bases de datos electrónicas SciELO, LILACS, Cochrane, ClinicalTrials.gov, SPORTDiscus y Medline, en mayo de 2011. De los 178 estudios inicialmente identificados, 12 respondieron a los criterios de inclusión y fueron clasificados en cuanto a la calidad mediante criterios de la escala PEDro. La magnitud y la duración de la respuesta de Movilización de las CPE fueron mayores después de la realización de ejercicios de larga/ultra larga duración y están correlacionadas con niveles plasmáticos de factor de crecimiento endotelio vascular (VEGF). El pico de Movilización de esas células en respuesta a una sesión de ejercicio máximo o submáximo, con duración de hasta una hora, ocurre en el período inmediatamente después del esfuerzo y hasta una hora después de su interrupción. Un posible mecanismo es la biodisponibilidad del Óxido Nítrico (NO). La edad de los individuos y la intensidad del ejercicio parecen interferir en la respuesta de Movilización de las CPE. Ejercicios de larga/ultra larga duración promueven Movilización más acentuada de las CPE cuando son comparados a ejercicios máximos o submáximos. Los mecanismos envuelven la liberación del VEGF en ejercicios de larga/ultra larga duración y la biodisponibilidad de NO en ejercicios máximo y submáximo con hasta una hora de duración.
Sujets)
Adolescent , Adulte , Sujet âgé , Sujet âgé de 80 ans ou plus , Humains , Adulte d'âge moyen , Jeune adulte , Mouvement cellulaire/physiologie , Cellules endothéliales/physiologie , Exercice physique/physiologie , Monoxyde d'azote/métabolisme , Facteur de croissance endothéliale vasculaire de type A/sang , Cellules endothéliales/cytologie , Facteurs tempsRésumé
We previously reported the successful establishment of embryonic stem cell (ESC)-like multipotent spermatogonial stem cells (mSSCs) from neonatal mouse testis. Here, we examined the ability of mSSCs to differentiate into vascular endothelial cells and smooth muscle cells, and compared to that of mouse ESCs. We used real-time reverse transcriptase polymerase chain reaction and immunohistochemistry to examine gene expression profiles of mSSCs and ESCs during in vitro vascular differentiation. Both mSSCs and ESCs exhibited substantial increase in the expression of mesodermal markers, such as Brachyury, Flk1, Mesp1, Nkx2.5, and Islet1, and a decrease in the expression of pluripotency markers, such as Oct3/4 and Nanog during the early stage of differentiation. The mRNA levels of vascular endothelial (VE)-cadherin and CD31 gradually increased in both differentiated mSSCs and ESCs. VE-cadherin- or CD31-positive cells formed sprouting branch-like structures, as observed during embryonic vascular development. At the same time, vascular smooth muscle cell-specific markers, such as myocardin and alpha-smooth muscle actin (SMA), were also highly expressed in differentiated mSSCs and ESCs. Immunocytochemical analysis revealed that the differentiated cells expressed both alpha-SMA and SM22-alpha proteins, and exhibited the intracellular fibril structure typical of smooth muscle cells. Overall, our findings showed that mSSCs have similar vascular differentiation abilities to those of ESCs, suggesting that mSSCs may be an alternative source of autologous pluripotent stem cells for vascular regeneration.
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Animaux , Humains , Mâle , Souris , Animaux nouveau-nés , Marqueurs biologiques/métabolisme , Différenciation cellulaire/physiologie , Cellules souches embryonnaires/cytologie , Cellules endothéliales/cytologie , Expression des gènes , Analyse de profil d'expression de gènes , Immunohistochimie , Muscles lisses vasculaires/cytologie , Myocytes du muscle lisse/cytologie , Cellules souches pluripotentes/cytologie , Réaction de polymérisation en chaine en temps réel , Spermatogonies/cytologie , Testicule/cytologieRésumé
We investigated tissue responses to endoskeleton stent grafts for saccular abdominal aortic aneurysms (AAAs) in canines. Saccular AAAs were made with Dacron patch in 8 dogs, and were excluded by endoskeleton stent grafts composed of nitinol stent and expanded polytetrafluoroethylene graft. Animals were sacrificed at 2 months (Group 1; n = 3) or 6 months (Group 2; n = 5) after the placement, respectively. The aortas embedding stent grafts were excised en bloc for gross inspection and sliced at 5 to 8 mm intervals for histopathologic evaluation. Stent grafts were patent in all except a dog showing a thrombotic occlusion in Group 2. In the 7 dogs with patent lumen, the graft overhanging the saccular aneurysm was covered by thick or thin thrombi with no endothelial layer, and the graft over the aortic wall was completely covered by neointima with an endothelial layer. Transgraft cell migration was less active at an aneurysm than at adjacent normal aorta. In conclusion, endoskeleton stent grafts over saccular aneurysms show no endothelial coverage and poor transgraft cell migration in a canine model.
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Animaux , Chiens , Alliages/composition chimique , Angiographie , Anévrysme de l'aorte abdominale/anatomopathologie , Mouvement cellulaire , Modèles animaux de maladie humaine , Cellules endothéliales/cytologie , Néointima/étiologie , Polytétrafluoroéthylène/composition chimique , Endoprothèses , Thrombose/étiologie , TomodensitométrieRésumé
Introduction: Oral submucous fibrosis (OSMF) is a chronic, progressive, scarring disease that predominantly affects the people of south-east Asian origin. Aim: The present study is aimed at comparing and correlating the mast cell density (MCD) and micro vascular density (MVD) in normal mucosa and different grades of OSMF and to analyze their role in the disease progression. Materials and Methods: MCD was assessed immunohistochemically using anti mast cell tryptase and MVD was assessed using anti-factor VIII related von Willebrand factor. Results: The one way comparison of MVD and MCD in normal mucosa and among different grades of OSMF showed a significant increase in MCD and MVD among OSMF cases. Correlation analysis using Pearson correlation coefficient, showed positive correlation between MCD and MVD i.e. as MCD increases there is an exponential increase in MVD. Conclusion: The increase in MVD and MCD reveals their role in the pathogenesis of OSMF, a lesion characterized by progressive fibrosis in early stages and failure of degradation or remodeling in the advanced stages.
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Études cas-témoins , Prolifération cellulaire , Cellules endothéliales/cytologie , Humains , Immunohistochimie , Mastocytes/cytologie , Microcirculation , Muqueuse de la bouche/vascularisation , Muqueuse de la bouche/cytologie , Néovascularisation pathologique/anatomopathologie , Fibrose buccale sous-muqueuse/classification , Fibrose buccale sous-muqueuse/anatomopathologie , Valeurs de référenceRésumé
In an oxygen-depleted environment, endothelial cells initiate an adaptive pattern of synthesis, which may enable them to survive hypoxic crises. Using high-resolution two-dimensional gel electrophoresis in conjunction with mass spectroscopy, we obtained a 24 differential display of proteins in the pancreatic endothelial cell line, MS-1, at four time points following induction of hypoxia. The induction of Wee1 under hypoxia was confirmed both at the mRNA and protein levels. The phosphorylation of cell division cycle 2, which is downstream of Wee1, was also increased after hypoxic exposure. In addition, pre-exposure to hypoxia attenuated a decrease in hydrogen peroxide-induced cell number. The induction of bax (a pro-apoptotic protein) and reduction of bcl (an anti-apoptotic protein) after hypoxia stimulus were also attenuated by hypoxic pre-exposure. Moreover, hydrogen peroxide-induced morphologic damage did not appear in the wild-type Wee1-expressing cells. Taken together, our results suggest that Wee1 may have important role in hypoxia-induced pathophysiological situations in endothelial cells.
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Animaux , Souris , Protéine-kinase CDC2/métabolisme , Protéines du cycle cellulaire/génétique , Hypoxie cellulaire , Lignée cellulaire , Survie cellulaire , Cellules endothéliales/cytologie , Régulation de l'expression des gènes , Peroxyde d'hydrogène/métabolisme , Protéines nucléaires/génétique , Pancréas/cytologie , Phosphorylation , Protein-tyrosine kinases/génétiqueRésumé
Cardiac lymphatic system in the remodeling after acute myocardial infarction (AMI) has been overlooked. We wanted to investigate the role of bone marrow-derived endothelial progenitor cells (EPCs) and their contribution to lymphatic distribution in myocardial remodeling after AMI. Mouse (C57bl/6J) MI models were created by ligation of the left anterior descending coronary artery and were treated with phosphate buffered saline (PBS) or EPCs. Real-time RT-PCR with 2- to 4-week myocardial tissue samples revealed that lymphangiogenetic factors such as vascular endothelial growth factor (VEGF)-C (8.5 fold, P < 0.05), VEGF-D (6.1 fold, P < 0.05), Lyve-1 (15 fold, P < 0.05), and Prox-1 (11 fold, P < 0.05) were expressed at significantly higher levels in the PBS group than the EPC group. The PBS group also showed a significantly higher density of lymphatic vessels in the peri-infarction area. Echocardiography showed that from 2 weeks after the treatment, left ventricle (LV) dimensions at both systole and diastole were significantly smaller in the EPC group than in the PBS group (P < 0.01) and LV fractional shortening was higher in the EPC group accordingly (P < 0.01). Lymphangiogenic markers increased in a mouse MI model. EPC transplantation decreased lymphangiogenesis and adverse ventricular remodeling after AMI. These novel findings suggest that new lymphatic vessels may be formed in severely damaged myocardium, and may be involved in adverse myocardial remodeling after AMI.
Sujets)
Animaux , Souris , Transplantation cellulaire , Cellules endothéliales/cytologie , Protéines à homéodomaine/génétique , Immunohistochimie , Lymphangiogenèse/génétique , Souris de lignée C57BL , Souris transgéniques , Infarctus du myocarde/métabolisme , Réaction de polymérisation en chaine en temps réel , Transplantation de cellules souches , Protéines suppresseurs de tumeurs/génétique , Facteur de croissance endothéliale vasculaire de type A/génétique , Facteur de croissance endothéliale vasculaire de type D/génétiqueRésumé
Gene transfer of basic fibroblast growth factor (bFGF) has been shown to induce significant endothelial migration and angiogenesis in ischemic disease models. Here, we investigate what factors are secreted from skeletal muscle cells (SkMCs) transfected with bFGF gene and whether they participate in endothelial cell migration. We constructed replication-defective adenovirus vectors containing the human bFGF gene (Ad/bFGF) or a control LacZ gene (Ad/LacZ) and obtained conditioned media, bFGF-CM and LacZ-CM, from SkMCs infected by Ad/bFGF or Ad/LacZ, respectively. Cell migration significantly increased in HUVECs incubated with bFGF-CM compared to cells incubated with LacZ-CM. Interestingly, HUVEC migration in response to bFGF-CM was only partially blocked by the addition of bFGF-neutralizing antibody, suggesting that bFGF-CM contains other factors that stimulate endothelial cell migration. Several proteins, matrix metalloproteinase-1 (MMP-1), plasminogen activator inhibitor-1 (PAI-1), and cathepsin L, increased in bFGF-CM compared to LacZ-CM; based on 1-dimensional gel electrophoresis and mass spectrometry. Their increased mRNA and protein levels were confirmed by RT-PCR and immunoblot analysis. The recombinant human bFGF protein induced MMP-1, PAI-1, and cathepsin L expression in SkMCs. Endothelial cell migration was reduced in groups treated with bFGF-CM containing neutralizing antibodies against MMP-1 or PAI-1. In particular, HUVECs treated with bFGF-CM containing cell-impermeable cathepsin L inhibitor showed the most significant decrease in cell migration. Cathepsin L protein directly promotes endothelial cell migration through the JNK pathway. These results indicate that cathepsin L released from SkMCs transfected with the bFGF gene can promote endothelial cell migration.
Sujets)
Humains , Anticorps neutralisants/immunologie , Cathepsine L/génétique , Mouvement cellulaire , Cellules cultivées , Test des comètes , Dependovirus/génétique , Cellules endothéliales/cytologie , Facteur de croissance fibroblastique de type 2/génétique , Techniques de transfert de gènes , Immunotransfert , JNK Mitogen-Activated Protein Kinases , Opéron lac/génétique , Spectrométrie de masse , Matrix metalloproteinase 1/biosynthèse , Muscles squelettiques/métabolisme , Néovascularisation physiologique , Inhibiteur-1 d'activateur du plasminogène/biosynthèse , ARN messager/biosynthèse , RT-PCRRésumé
The purpose of this study is to determine 1) whether morphine postconditiong (MPostC) can attenuate the intercellular adhesion molecules-1 (ICAM-1) expression after reoxygenation injury and 2) the subtype(s) of the opioid receptors (ORs) that are involved with MPostC. Human umbilical vein endothelial cells (HUVECs) were subjected to 6 hr anoxia followed by 12 hr reoxygenation. Three morphine concentrations (0.3, 3, 30 microM) were used to evaluate the protective effect of MPostC. We also investigated blockading the OR subtypes' effects on MPostC by using three antagonists (a micro-OR antagonist naloxone, a kappa-OR antagonist nor-binaltorphimine, and a delta-OR antagonist naltrindole) and the inhibitor of protein kinase C (PKC) chelerythrine. As results, the ICAM-1 expression was significantly reduced in the MPostC (3, 30 microM) groups compared to the control group at 1, 6, 9, and 12 hours reoxygenation time. As a consequence, neutrophil adhesion was also decreased after MPostC. These effects were abolished by coadministering chelerythrine, nor-binaltorphimine or naltrindole, but not with naloxone. In conclusion, it is assumed that MPostC could attenuate the expression of ICAM-1 on endothelial cells during reoxygenation via the kappa and delta-OR (opioid receptor)-specific pathway, and this also involves a PKC-dependent pathway.
Sujets)
Animaux , Humains , Benzophénanthridines/pharmacologie , Cellules endothéliales/cytologie , Endothélium vasculaire/cytologie , Molécule-1 d'adhérence intercellulaire/génétique , Morphine/pharmacologie , Naloxone/pharmacologie , Naltrexone/analogues et dérivés , Antagonistes narcotiques/pharmacologie , Stupéfiants/pharmacologie , Isoformes de protéines/métabolisme , Protéine kinase C/antagonistes et inhibiteurs , Récepteurs aux opioïdes/métabolisme , Lésion d'ischémie-reperfusion/métabolisme , Transduction du signal/physiologie , Veines ombilicales/cytologieRésumé
Surface properties of a biomaterial could be critical in determining biomaterial's biocompatibility due to the fact that the first interactions between the biological environment and artificial materials are most likely occurred at material's surface. In this study, the surface properties of a new nanocomposite [NC] polymeric material were modified by combining plasma treatment and collagen immobilization in order to enhance cell adhesion and growth. Methods: NC films were plasma treated in reactive O[2] plasma at 60 W for 120 s. Afterward, type I collagen was immobilized on the activated NC by a safe, easy, and effective one-step process. The modified surfaces of NC were characterized by water contact angle measurement, water uptake, scanning electron microscopy [SEM], and Fourier transformed infrared spectroscopy in attenuated total reflection mode [ATR-FTIR]. Furthermore, the cellular behaviors of human umbilical vascular endothelial cells [HUVEC] such as attachment, growth and proliferation on the surface of the NC were also evaluated in vitro by optical microscopy and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide test. Results: The outcomes indicated that plasma treatment and collagen immobilization could improve hydrophilicity of NC. SEM micrograph of the grafted film showed a confluent layer of collagen with about 3-5 jum thicknesses. In vitro tests showed that collagen-grafted and plasma-treated surfaces both resulted in higher cell adhesion and growth state compared with untreated ones. Conclusion: Plasma surface modification and collagen immobilization could enhance the attachment and proliferation of HUVEC onto NC, and the method would be usefully applied to enhance its biocompatibility
Sujets)
Humains , Collagène/pharmacologie , Cellules endothéliales/cytologie , Cellules endothéliales/effets des médicaments et des substances chimiques , Nanocomposites/composition chimique , Adhérence cellulaire/effets des médicaments et des substances chimiques , Prolifération cellulaire/effets des médicaments et des substances chimiques , Propriétés de surface/effets des médicaments et des substances chimiques , Veines ombilicales/cytologie , EauRésumé
Molecular plant components have long been aimed at the angiogenesis and anti-angiogenesis pathways, and have been tested as sources for antineoplasic drugs with promising success. The present work deals with the anti-angiogenic effects of Methyl Jasmonate. Jasmonate derivatives were demonstrated to selectively damage the mitochondria of cancer cells. In vitro, 1-10 mM Methyl Jasmonate induced the cell death of the human umbilical vein endothelial cells (HUVEC) and the Murine melanoma cells (B16F10), while micromolar concentrations were ineffective. In vivo, comparable concentrations were toxic and reduced the vessel density of the Chorioallantoic Membrane of the Chicken Embryo (CAM). However, 1-10 µM concentrations produced a complex effect. There was increased capillary budding, but the new vessels were leakier and less organised than corresponding controls. It is suggested that not only direct toxicity, but also the drug effects upon angiogenesis are relevant to the antineoplasic effects of Methyl Jasmonate.
Moléculas de origem vegetal são, há muito, conhecidas como substâncias ativas sobre as vias de angiogênese e antiangiogênese e foram testadas como fonte de drogas antineoplásicas com sucesso promissor. Este trabalho trata dos efeitos antiangiogênicos do Metiljasmonato, um protótipo da família dos derivados do ácido jasmônico, que danificam seletivamente a mitocôndria de células neoplásicas. In vitro, metiljasmonato 1-10 mM promoveu a morte celular de células endoteliais humanas de cordão umbilical (HUVEC) e de melanoma murino (B16F10); concentrações micromolares foram inócuas. In vivo, concentrações equivalentes foram tóxicas e reduziram a densidade de vasos em membranas corioalantoicas de embrião de galinha (CAM). Entretanto, concentrações entre 1-10 µM produziram um efeito complexo. Ocorreu aumento no brotamento capilar, mas os novos vasos apresentaram-se frágeis e menos organizados que os controles correspondentes. Sugere-se que, além da toxicidade direta contra as células tumorais, a ação do metiljasmonato sobre a angiogênese seja relevante para seu efeito antineoplásico.
Sujets)
Animaux , Embryon de poulet , Humains , Acétates , Inhibiteurs de l'angiogenèse/pharmacologie , Antinéoplasiques/pharmacologie , Chorioallantoïde/effets des médicaments et des substances chimiques , Cyclopentanes/pharmacologie , Cellules endothéliales/effets des médicaments et des substances chimiques , Oxylipines/pharmacologie , Lignée cellulaire tumorale , Chorioallantoïde/vascularisation , Cellules endothéliales/cytologie , Veines ombilicales/cytologie , Veines ombilicales/effets des médicaments et des substances chimiquesRésumé
PURPOSE: Endothelial cells maintain the homeostasis of blood, which consists of plasma and cellular components, and regulate the interaction between blood and the surrounding tissues. They also have essential roles in vascular permeability, the circulation, coagulation, inflammation, wound healing, and tissue growth. The senescence of endothelial cells is closely related to the aging of the adjacent tissues and to age-related vascular disease. Recently, the expression of moesin was found to be decreased in elderly human dermal microvascular endothelial cells (HDMECs), and an association between moesin and senescence has been suggested. This study examined the functional role of moesin in cellular senescence. MATERIALS AND METHODS: To study the effects of decreased moesin expression on cellular senescence and metabolism, HDMECs were transfected with short hairpin-RNA (shRNA) lentivirus to silence moesin gene expression. In addition, specimens from young and old human skin were stained with anti-moesin and anti-p16 antibodies as an in vivo study. RESULTS: Using shRNAl-entivirus, moesin knock-down HDMECs developed characteristics associated with aging and expressed senescence associated-beta-galactosidase during early passages. They also showed increased p16 expression, decreased metabolic activity, and cell growth retardation. Human skin tissue from elderly persons showed decreased moesin expression and increased p16 expression. CONCLUSION: These findings suggest that there is a functional association between moesin expression and cellular senescence. Further study of the functional mechanism of moesin in the cytoskeleton and cellular senescence is needed. In addition, this study provides a useful model for developing anti-aging treatments.
Sujets)
Sujet âgé de 80 ans ou plus , Enfant , Humains , Antigènes CD31/métabolisme , Technique de Western , Vieillissement de la cellule/génétique , Lignée cellulaire , Cellules endothéliales/cytologie , Immunohistochimie , Protéines des microfilaments/génétique , Microscopie de contraste de phase , Microvaisseaux/cytologie , Petit ARN interférent/génétique , RT-PCR , Peau/vascularisationRésumé
Angiogenesis is critical and indispensable for tumor progression. Since VEGF is known to play a central role in angiogenesis, the disruption of VEGF-VEGF receptor system is a promising target for anti-cancer therapy. Previously, we reported that a hexapeptide (RRKRRR, RK6) blocked the growth and metastasis of tumor by inhibiting VEGF binding to its receptors. In addition, dRK6, the D-form derivative of RK6, retained its biological activity with improved serum stability. In the present study, we developed a serum-stable branched dimeric peptide (MAP2-dRK6) with enhanced anti-VEGF and anti-tumor activity. MAP2-dRK6 is more effective than dRK6 in many respects: inhibition of VEGF binding to its receptors, VEGF- and tumor conditioned medium-induced proliferation and ERK signaling of endothelial cells, and VEGF-induced migration and tube formation of endothelial cells. Moreover, MAP2-dRK6 blocks in vivo growth of VEGF-secreting colorectal cancer cells by the suppression of angiogenesis and the subsequent induction of tumor cell apoptosis. Our observations suggest that MAP2-dRK6 can be a prospective therapeutic molecule or lead compound for the development of drugs for various VEGF-related angiogenic diseases.