Resumen de Papel del sistema STIM/Orai en la hipercontractilidad del tejido vascular peneano asociada al envejecimiento y sus implicaciones en la disfunción eréctil
- español
Las alteraciones funcionales y estructurales del tejido vascular producidas por el envejecimiento y la diabetes condicionan la aparición de situaciones patológicas como la disfunción eréctil. Dada la asociación de estos dos factores de riesgo cardiovascular con la desregulación de la homeostasis del calcio, se ha propuesto al mecanismo de entrada de calcio SOCE y sus principales efectores, STIM y Orai, como dianas terapéuticas emergentes en la fisiopatología cardiovascular. Los objetivos de esta tesis doctoral se centran en evaluar el impacto del envejecimiento sobre el sistema STIM/Orai en el tejido vascular peneano de ratas y humanos, analizar la relevancia de este sistema en el tejido vascular peneano de pacientes con disfunción eréctil, y caracterizar la influencia de la diabetes sobre STIM/Orai en el tejido vascular peneano de ratas y pacientes con disfunción eréctil. La evaluación de las respuestas funcionales mostró una reversión de la hipercontractilidad asociada al envejecimiento tras la inhibición de STIM/Orai con YM-58483 en los tejidos peneanos de ratas y humanos. Además, la inhibición de STIM/Orai moduló las respuestas neurogénicas y favoreció las respuestas de relajación dependiente de endotelio en ratas viejas. El análisis de la expresión proteica mostró una asociación entre el envejecimiento y la sobreexpresión de Orai 3 en los tejidos vasculares peneanos de ratas y humanos. También se observó una reversión de la hipercontractilidad asociada a la disfunción eréctil en tejido peneano humano tras la inhibición de STIM/Orai. Así mismo, el tratamiento con YM-58483 moduló las respuestas neurogénicas y mejoró la relajación endotelial en los tejidos peneanos de pacientes con disfunción eréctil. El análisis de expresión proteica mostró una relación entre la disfunción eréctil y la sobreexpresión de Orai 1 y Orai 3 en los tejidos peneanos humanos. La capacidad de YM-58483 de reducir significativamente las contracciones también fue observada en presencia de diabetes en tejido vascular peneano de ratas y pacientes con disfunción eréctil. Del mismo modo, la inhibición de STIM/Orai con YM-58483 también moduló las respuestas neurogénicas en cuerpo cavernoso de ratas diabéticas. Por otro lado, el antagonismo de STIM/Orai potenció las relajaciones inducidas por tadalafilo y las dependientes de endotelio en los tejidos cavernosos de ratas diabéticas. La diabetes se asoció con una disminución de la expresión proteica de STIM-1 en cuerpo cavernoso de ratas, y con una sobreexpresión de Orai 1 y Orai 3 en tejido cavernoso de pacientes con disfunción eréctil. En conjunto, estos resultados sugieren que el sistema STIM/Orai podría ser considerado una potencial diana terapéutica en el envejecimiento funcional del tejido peneano. Además, la modulación de STIM/Orai, al minimizar las alteraciones funcionales del tejido peneano humano, podría tener una relevancia terapéutica en la disfunción eréctil, especialmente en la disfunción eréctil diabética, que presenta una menor respuesta al tratamiento convencional.
- English
Ageing and diabetes are main risk factors for developing cardiovascular diseases. Functional and structural disorders produced by these risk factors in vascular tissue determine the appearance of pathological situations such as erectile dysfunction. Ageing and diabetes have been associated with dysregulation of vascular calcium homeostasis. In this sense, Store-Operated Calcium Entry (SOCE) mechanism and its key players, Stromal Interaction molecule (STIM), as sensor protein, and Orai calcium channels, have been suggested as emerging therapeutic targets in cardiovascular pathophysiology. The main objectives of this doctoral thesis are: 1) to evaluate the impact of ageing on STIM/Orai system in both rat and human vascular penile tissue, 2) to analyse the relevance of STIM/Orai system and the impact of its modulation in vascular penile tissue from patients with erectile dysfunction, and 3) to characterize the influence of diabetes in STIM/Orai signalling in vascular penile tissue from rats and erectile dysfunction patients. Rat cavernosal tissue samples were obtained from 3 months-old and 20 months-old animals, and rats with insulin-dependent diabetes induced by a single intraperitoneal administration of streptozotocin. Both human corpus cavernosum and penile resistance arteries were obtained from organ donors of different ages without history of erectile dysfunction, as well as from erectile dysfunction patients undergoing penile prosthesis insertion. Corpus cavernosum and penile arteries were functionally evaluated in organ chambers and wire myographs, respectively. Protein expression of STIM-1, Orai 1 and Orai 3 was determined and quantified by Western blot and/or immunofluorescence. STIM/Orai inhibition with YM-58483 (20 µM) reversed ageing-induced hypercontractility in penile vascular tissue. This enhancement of both norepinephrineinduced and thromboxane analogue, U46619-induced contractions was attenuated in rat penile tissues but completely reversed in human penile tissues. Another STIM/Orai inhibitor, 2-aminoethoxydiphenyl borate (2-APB) (100 µM), reduced norepinephrineinduced contractions in corpus cavernosum from aged rats, as well. YM-58483 treatment also reduced neurogenic contractile responses and favoured neurogenic relaxation responses (nitrergic). Additionally, it partially improved endothelium-dependent relaxations in cavernosal tissues from aged rats. Ageing was associated with overexpression of Orai 3 in both rat and human penile vascular tissues. STIM/Orai antagonism with YM-58483 (20 µM) completely reversed erectile dysfunction-related adrenergic hypercontractility in both human corpus cavernosum and penile resistance arteries. Although YM-58483 treatment also reduced norepinephrineinduced contractions in cavernosal tissues from subjects without erectile dysfunction, the magnitude of the effect was lower than in patients with erectile dysfunction. STIM/Orai inhibition with 2-APB (100 µM) reduced norepinephrine-induced contractions in corpus cavernosum and penile resistance arteries from patients with erectile dysfunction too. STIM/Orai inhibition with YM-58483 reduced both neurogenic and U46619-induced contractions while it improved both neurogenic and endothelium-dependent relaxations in penile tissues from patients with erectile dysfunction. On the other hand, the addition of YM-58483 concentration-dependently relaxed precontracted corpus cavernosum and penile resistance arteries. These relaxations were significantly more pronounced in tissues from patients with erectile dysfunction. Erectile dysfunction was related to overexpression of Orai 1 and Orai 3 in both human corpus cavernosum and penile resistance arteries. STIM/Orai inhibition with YM-58483 (20 µM) reduced norepinephrine-induced contractions more pronouncedly in corpus cavernosum from diabetic rats, and in both human corpus cavernosum and penile resistance arteries from diabetic patients with erectile dysfunction in comparison with their respective non-diabetic groups. 2-APB treatment (100 µM) reduced norepinephrine-induced contractile responses in corpus cavernosum from diabetic rats as well. U46619-induced contractions were also reduced in cavernosal tissue from diabetic rats by STIM/Orai inhibition with YM-58483. Furthermore, YM-58483 was able to inhibit neurogenic contractile responses and to improve neurogenic relaxation responses in corpus cavernosum from diabetic rats. On the other hand, STIM/Orai antagonism with YM-58483 (10 µM) improved tadalafil-induced and endothelium-dependent relaxations in cavernosal tissues from diabetic rats. Diabetes was associated with a dysregulation in STIM-1 expression in rat corpus cavernosum while overexpression of Orai 1 and Orai 3 was detected in human penile tissue from diabetic patients with erectile dysfunction. Overall, these outcomes suggest that STIM/Orai system could be considered as a potential therapeutic target for preserving functional status of aged penile tissue. Furthermore, STIM/Orai modulation, minimizing functional disorders of human penile tissue, could have therapeutic relevance in erectile dysfunction, particularly in diabetic erectile dysfunction where response to conventional treatment is reduced.
