Esta tesis se presenta en forma de compendio de publicaciones según la normativa aprobada por la Comisión de Doctorado de la Universitat Autònoma de Barcelona.
El núcleo principal se basa en artículos originales publicados en revistas indexadas.
Los trabajos evalúan los niveles de fosforilación y nitrosilación de tirosinas de las interleuquinas del vítreo de pacientes afectos de retinopatía diabética. Para ello se han analizado muestras de vítreo de los mismos y se han comparado con pacientes no diabéticos
Artículo 1
Tyrosine Phosphorylation of Vitreous Inflammatory and Angiogenic Peptides and Proteins in Diabetic Retinopathy
Investigative Ophthalmology and Visual Science
2009; 50: 1378–1382 DOI:10.1167/iovs.08-2736 FACTOR IMPACTO ………………3,431
Objetivo
Estudiar el grado de fosforilación de tirosinas en un amplio conjunto de péptidos y proteínas relacionados con procesos de angiogénesis y de inflamación en muestras de humor vítreo de pacientes diabéticos tipo 2 con Retinopatía diabética proliferativa.
Comparar los resultados obtenidos con las muestras de vítreo de pacientes sin retinopatía diabética, de sexo y edades similares.
Material y Método
Estudio comparativo entre muestras obtenidas durante la VPP realizada por complicaciones derivadas de la RDP con las obtenidas en las VPP por agujero macular idiopático en los sujetos control. Estudio comparativo entre muestras obtenidas durante la VPP realizada por complicaciones derivadas de la RDP con las obtenidas en las VPP por agujero macular idiopático en los sujetos control.
Obtención de las muestras de humor vítreo: Vitrectomía Pars Plana con intercambio de aire para la obtención de muestra no diluida , se realiza una centrifugación inmediata y se congela a – 80º C
Se aplicaron técnicas de inmunoblot en un sistema de mini-array para la cuantificación de una amplia gama de quimioquinas, péptidos vasoactivos y proteínas con cuantificación de bandas software multi Gauge v 3.0
Los resultados se expresaron como el porcentaje de variación en comparación con sujetos control.
Muestra: 8 pacientes afectos de Retinopatía diabética proliferativa
Control: 8 pacientes con Agujero macular idiopático
La cantidad total de proteínas analizadas fue similar en los pacientes y sujetos control (48).
Fosfosforilación de tirosinas sin cambios significativos (p< 0,05) en pacientes diabéticos respecto al grupo de control:
Crecimiento regular del oncogén α (GRO α )
IL-2 , IL-3, IL-4
Interferón (FN- ϒ)
Proteína quimiotáctica de monocitos (MCP-1-2-3)
Factor de necrosis tumoral (TNF α , β)
Factor de crecimiento epidérmico (EGF)
Factor de crecimiento similar a la insulina (IGF)
Trombopoyetina
Factor de crecimiento vascular endotelial (VEGF)
Factor de crecimiento derivado de las plaquetas (PDGF 88)
Fosfosforilación de tirosinas con disminución moderada 15%-20% (p< 0,05) en pacientes diabéticos respecto al grupo de control:
Crecimiento regular del oncogén (GRO)
Citoquina humana I-309
Interleuquina (IL-13)
Factor estimulante de las colonias de monocitos
Quimioquina derivada de los macrófagos
Factor de las células madre / This thesis is presented in the form of a compendium of published articles in accordance with the doctorate commission of the Universitat Autónoma de Barcelona.
The main body of this thesis comprises original research articles published in indexed journals.
The main subject of the research is the evaluation of Phosphorylation and Nitrosylation of interleuquines in the vitreous of patients with diabetic retinopathy. Samples from diabetic patients were analysed and compared with a control group of not diabetic patients.
Article 1
Tyrosine Phosphorylation of Vitreous Inflammatory
and Angiogenic Peptides and Proteins in
Diabetic Retinopathy
Invest Ophthalmol .Vis Sci. 2009;50:1378–1382) DOI:10.1167/iovs.08-2736
Impact factor 3,431
.To evaluate the degree of phosphorylation of vitreous
proteins in patients with type 2 diabetes mellitus and diabetic
retinopathy compared with a group of control subjects without
diabetes and of similar age and sex.
METHODS. In samples obtained after vitrectomy for diabetic
retinopathy in patients and for macular hole in control subjects,
immunoblot techniques were applied to a mini-array
system for quantification of a wide range of chemokines and
vasoactive peptides and proteins. Antiphosphotyrosine antibody
was used for tyrosine phosphorylation evaluation and
results were expressed as the percentage of variation compared
with that in control subjects.
RESULTS. Samples from eight patients with type 2 diabetes and
from eight control subjects were analyzed. The total quantity
of proteins analyzed was similar in both patients and control
subjects. Tyrosine phosphorylation was very significantly decreased
(_20%, P _ 0.05) in diabetic patients with respect to
the control group in growth-related oncogene, human cytokine
I-309, interleukin-13, monocyte colony-stimulating factor,
macrophage-derived chemokine, stem cell factor, transforming
growth factor-_1, angiogenin, and oncostatin M. A significant
decrease in phosphorylation (between 20% and 40%, P _ 0.05)
was observed in epithelial neutrophil-activating peptide 78;
granulocyte colony-stimulating factor; granulocyte-monocyte–
stimulating colony factor; IL-5, -6, -7, -8, -10, and -12p40p70;
monokine induced by interferon-_; macrophage inflammatory
protein 1-; and normal T expressed and secreted cytokine
(RANTES) in comparison with that in the control subjects. The
greatest decrease in phosphorylation status was found in IL-1-_
and -1.
CONCLUSIONS. Diabetic retinopathy is associated with a decrease
in tyrosine phosphorylation of many vitreous proteins which
may indicate an alteration in protein functionality or action
even before significant quantitative variations.
Article 2
Diabetic Retinopathy Is Associated with
Decreased Tyrosine Nitrosylation of
Vitreous Interleukins IL-1 , IL-1 , and IL-7
Ophthalmic Res 2011;46:169–174
DOI: 10.1159/000323812
Impact factor 1.29
Objective: To simultaneously evaluate tyrosine nitrosylation
and phosphorylation levels of vitreous interleukins of patients
with diabetic retinopathy, in which abnormal tyrosine
phosphorylation has been previously described.
ResearchDesign and Methods: Specific immunoprecipitation of interleukins
IL-1 _ , IL-1 _ , IL-2 and IL-7 was carried out in samples
obtained during vitrectomy performed for proliferative diabetic
retinopathy in patients (n = 12) and for macular hole in
controls (n = 12). Tyrosine nitrosylation and phosphorylation
levels of the immunoprecipitated interleukins were analysed
by Western blot with the respective specific antibodies and
correlated. The results were also correlated with the total
amount of immunoprecipitated interleukin protein. The
mean phosphorylation/nitrosylation ratios of these proteins
in vitreous humour of both the control group and diabetic patients were determined
Results: Diabetes was associated with decreased tyrosine nitrosylation of IL-1, IL-1 and IL-7and an increased tyrosine phosphorylation/nitrosylation ratio
with respect to controls in IL-1 (1.58 8 0.22 vs. 2.74 80.39, respectively; p 0.05) and IL-7 (2.15 8 0.01 vs. 3.26 80.57, respectively; p 0.05). No significant changes were observedin nitrotyrosine or in the tyrosine phosphorylation/
nitrosylation ratio of IL-2.
Conclusions: Proliferative diabetic
retinopathy is associated with concomitant and simultaneous
changes in both tyrosine phosphorylation and tyrosine
nitrosylation status of specific pro-inflammatory interleukins
present in the vitreous fluid such as IL-1 , IL-1 and
IL-7. These changes could be related to the increase in
pro-inflammatory activity detected in diabetes-induced retinopathy.
Main conclusions
Diabetic retinopathy is associated with alterations in the tyrosine phosphorylation status of protein inflammatory interleukins group, which could be the pathophysiological basis of retinal involvement.
Tyrosine nitrosylation is not the result of changes in phosphorylation.
Alterations in interleukin-nitrosylation may have an implication in the pathogenesis of diabetic retinopathy.
The phospho-dephosphorylation mechanisms and nitro-denitrosilatión could be therapeutic targets in the future.
These findings open an future research, which could be based on experimental models in which they assessed the effects that changes in the mechanisms of phosphorylation / tyrosine nitrosylation on the occurrence or progression of diabetic retinopathy.
| Identifer | oai:union.ndltd.org:TDX_UAB/oai:www.tdx.cat:10803/83996 |
| Date | 11 November 2011 |
| Creators | Nadal Reus, Jeroni |
| Contributors | Camara Hermoso, Julio de la, Reverter Calatayud, Jordi Lluís, Universitat Autònoma de Barcelona. Departament de Cirurgia |
| Publisher | Universitat Autònoma de Barcelona |
| Source Sets | Universitat Autònoma de Barcelona |
| Language | Spanish |
| Detected Language | English |
| Type | info:eu-repo/semantics/doctoralThesis, info:eu-repo/semantics/publishedVersion |
| Format | 99 p., application/pdf |
| Source | TDX (Tesis Doctorals en Xarxa) |
| Rights | info:eu-repo/semantics/openAccess, ADVERTIMENT. L'accés als continguts d'aquesta tesi doctoral i la seva utilització ha de respectar els drets de la persona autora. Pot ser utilitzada per a consulta o estudi personal, així com en activitats o materials d'investigació i docència en els termes establerts a l'art. 32 del Text Refós de la Llei de Propietat Intel·lectual (RDL 1/1996). Per altres utilitzacions es requereix l'autorització prèvia i expressa de la persona autora. En qualsevol cas, en la utilització dels seus continguts caldrà indicar de forma clara el nom i cognoms de la persona autora i el títol de la tesi doctoral. No s'autoritza la seva reproducció o altres formes d'explotació efectuades amb finalitats de lucre ni la seva comunicació pública des d'un lloc aliè al servei TDX. Tampoc s'autoritza la presentació del seu contingut en una finestra o marc aliè a TDX (framing). Aquesta reserva de drets afecta tant als continguts de la tesi com als seus resums i índexs. |
Page generated in 0.1068 seconds