Person: Crooke Álvarez, Almudena
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First Name
Almudena
Last Name
Crooke Álvarez
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Óptica y Optometría
Department
Bioquímica y Biología Molecular
Area
Bioquímica y Biología Molecular
Identifiers
6 results
Search Results
Now showing 1 - 6 of 6
Item Melatonin Makes Me Feel Awake! Seven Years of Lab Experience (2000-2007)(Anales de la Real Academia Nacional de Farmacia, 2007) Pelaez, Teresa; Mediero Muñoz, Aránzazu; Alarma-Estrany, Pilar; Loma Lozano, Patricia; Guzmán Aránguez, Ana Isabel; Crooke Álvarez, Almudena; Colligris, Basilio; Peral Cerda, María Asunción; Pintor Just, Jesús JerónimoThe research on the effect of melatonin on intraocular pressure (IOP) is reviewed from the hystorical point of view of our laboratory. The original idea of melatonin modulating intraocular pressure has been improved by using selective compounds for MT2 and specially melatonin MT3 receptors. The selective compound 5-methoxyamino N-acetyltryptamine (5-MCA-NAT) has been an attractive compound due to its ability to reduce IOP about 40%, therefore being a good candidate to the treatment of the ocular hypertension linked to glaucoma. More compounds have been developed and tested permitting us to have a more accurate panorama of those receptors controlling the relevant process of intraocular pressure.Item Therapeutic potential of topical administration of siRNAs against HIF-1α for corneal neovascularization(Experimental Eye Research, 2022) Peral Cerda, María Asunción; Mateo Ibáñez, Jesús; Domínguez Godínez, Carmen Olalla; Carracedo Rodríguez, Juan Gonzalo; Gómez Pedrero, José Antonio; Crooke Álvarez, Almudena; Pintor Just, Jesús JerónimoGiven the implications of the problem of neovascularization on ocular health, as well as the growth in the number of cases, the purpose of the present study has been testing the efficacy of siRNAs (small interfering RNA) designed to silence Hypoxia Inducible Factor -1α (HIF-1α) and to demonstrate that their use stops neovascularization in a model of corneal burn. Corneal wounds in the limbic zone were made in the eyes of New Zealand white rabbits. Topical applications of siRNAs were done the next day to the wound for four consecutive days and eyes were examined with a slit lamp. Evaluation of neovascularization progress was done by analyzing images by ImageJTM and to determine the neovascular area in Matlab ® was used. At the same time, a rabbit corneal cell line was used for in vitro study of hypoxia exposure and Western blot analysis of the cell's extracts were done. Under normal cell culture oxygenation, the expression of HIF-1α was lower than that observed under hypoxic conditions. After 2 h of hypoxia, there was a significant increase in the HIF-1α expression, effect that was maintained up to 6 h. The increased in HIF-1α was mimicked by a cell permeable prolyl-4-hydroxylase inhibitor. Cobalt chloride showed no capacity to increase HIF-1α in vitro. The effect of three different siRNA on HIF-1α was tested after 4 h of hypoxia. siRNA#1 was able to silence 80% of HIF-1α expression, siRNA#2 and siRNA#3 reduce the expression in 45% and 40% respectively. In addition, the three siRNA were tested in a corneal model of neovascularization. scrambledsiRNA#2 was the most effective inhibitor of blood vessel production, followed by siRNA#3 and siRNA#1. Compared to the scrambled siRNA (100% of blood vessel generation), siRNA#2 blocked the presence of blood vessels by 83 ± 2%, siRNA#3 inhibited 45 ± 7% and siRNA#1 only inhibited 18 ± 5%. The necessary time to observe the 50% of effect showed values of NV50 of 10.2 ± 2.4 days for the scrambled siRNA, 9.1 ± 1.4 for siRNA#1, 6.5 ± 1.85 for siRNA#2 and 4.8 ± 1.8 days for siRNA#3. In conclusion, the topical application of siRNA towards HIF-1α seems to be an effective and reliable method to stop neovascularization.Item Corneal Re-epithelialization Stimulated by Diadenosine Polyphosphates Recruits RhoA/ROCK and ERK1/2 Pathways(Investigative Ophthalmology & Visual Science, 2008) Mediero Muñoz, Aránzazu; Guzmán Aránguez, Ana Isabel; Crooke Álvarez, Almudena; Peral Cerda, María Asunción; Pintor Just, Jesús JerónimoPurpose. To investigate the role of ERK1/2 and RhoA/ROCK intracellular pathways in the modification of corneal re-epithelialization when stimulated by the diadenosine polyphosphates Ap4A and Ap3A. Methods. In wounded confluent SIRC (Statens Seruminstitut rabbit cornea) cell monolayers and in the presence or absence of Ap4A or Ap3A 100 μM, a battery of P2 receptor antagonists and inhibitors of tyrosin kinases, MAPK, and cytoskeleton pathways (AG1478 100 μM, U0126 100 μM, Y27632 100 nM, and (−)-blebbistatin 10 μM; n = 8 each) were assayed. Also, the activation of ERK1/2 and ROCK-I was examined by Western blot assay after treatment with Ap4A and Ap3A (100 μM), with or without suramin, RB-2, U0126, and Y27632. The intracellular distribution of pERK and ROCK-I was examined in the presence of Ap4A or Ap3A (100 μM) with U0126 and Y27632 (100 nM). Results. In the presence of Ap4A, U0126, Y27632, AG1478, and (−)-blebbistatin, reduced the migration rate compared to the effect of Ap4A alone (P < 0.0001, P < 0.001, P < 0.01, and P < 0.1 versus Ap4A, respectively). In the presence of Ap3A 100 μM, U0126 and Y27632 accelerated the migration rate when compared with the effect of Ap3A alone, whereas AG1478 and (−)-blebbistatin (P < 0.0001 versus Ap3A) slowed the migration rate. Western blot assays demonstrated that both dinucleotides activated the ERK1/2 pathway but only Ap4A activated the ROCK-I pathway. The intracellular distribution of pERK1/2 and ROCK-I reflected cross-talk between these two pathways. Conclusions. The activation of the Ap4A/P2Y2 receptor, accelerates corneal epithelial cell migration during wound healing with the activation of MAPK and cytoskeleton pathways, whereas activation of the Ap3A/P2Y6 receptor signals only the MAPK pathway.Item Effect of Melatonin and Analogues on Corneal Wound Healing: Involvement of Mt2 Melatonin Receptor(Current Eye Research, 2015) Crooke Álvarez, Almudena; Guzmán Aránguez, Ana Isabel; Mediero Muñoz, Aránzazu; Alarma Estrany, Pilar; Carracedo Rodríguez, Juan Gonzalo; Peláez García, María Teresa; Peral Cerda, María Asunción; Pintor Just, Jesús JerónimoPurpose: We have investigated the effect of melatonin and its analogues on rabbit corneal epithelial wound healing. Methods: New Zealand rabbits were anaesthetised and wounds were made by placing Whatman paper discs soaked in n-heptanol on the cornea. Melatonin and analogues (all 10 nmol) were instilled. Wound diameter was measured every 2 hours by means of fluorescein application with a Topcon SL-8Z slit lamp. Melatonin antagonists (all 10 nmol) were applied 2 hours before the application of the n-heptanol-soaked disc and then every 6 hours together with melatonin. To confirm the presence of MT2 receptors in corneal epithelial cells immunohistochemistry, Western blot and RT-PCR assays in native tissue and in rabbit corneal epithelial cells were performed. The tear components were extracted then processed by HPLC to quantify melatonin in tears. Results: Migration assays revealed that melatonin and particularly the treatment with the MT2 agonist IIK7, accelerated the rate of healing (p < 0.001). The application of the non-selective melatonin receptor antagonist luzindole and the MT2 antagonist DH97 (but not prazosin), prevented the effect of melatonin on wound healing (both p < 0.001). Immunohistochemistry, Western blot and RT-PCR assays showed the presence of MT2 melatonin receptor in corneal epithelial cells. In addition, we have identified melatonin in tears and determined its daily variations. Conclusions: These data suggest that MT2 receptors are implicated in the effect of melatonin on corneal wound healing regulating migration rate. This suggests the potential use of melatonin and its analogues to enhance epithelial wound healing in ocular surface disease.Item PhDAY 2020 -FOO (Facultad de Óptica y Optometría)(2020) Pintor Just, Jesús Jerónimo; Carpena Torres, Carlos; Peral Cerda, María Asunción; Pérez de Lara, María Jesús; Toral, Fernando; Crooke Álvarez, Almudena; Pastrana Robles, Cristina; Carracedo Rodríguez, Juan Gonzalo; Cayuela López, Ana; Sorzano Sánchez, Óscar; Charbel, Carla; Garzón Jiménez, Nuria; Carballo Álvarez, Jesús; Diz Arias, Elena; Fernández Jiménez, Elena; Lledó Mayans, Victoria Eugenia; Gómez Pedrero, José Antonio; Durán Prieto, Elena; López Alonso, José Manuel; Fernández Torres, Miguel Ángel; Guzmán Aránguez, Ana Isabel; Gómez Manzanares, Ángela; Vázquez Molini, Daniel; Martínez Antón, Juan Carlos; Bernárdez Vilaboa, Ricardo; Mayorga Pinilla, Santiago; Álvarez Fernández-Balbuena, Antonio; Benítez, AntoJ.; Igalla El-Youssfi, Asmae; León Álvarez, Alejandro; Palomo Álvarez, Catalina; Awad Alkozi, Hanan; Sánchez Naves, Juan; Martínez Alberquilla, Irene; García Montero, María; Ruiz Alcocer, Javier; Madrid Costa, David; Martínez Florentín, Gema; Papas, Eric B.; Medrano Muñoz, Sandra Milena; Molina, Nancy; Jurado, Sandra; Oliveiros López, Juan; Platero Alvarado, Nadiuska Cristine; Garrido Mercado, Rafaela; Pérez Garmendia, Carlos; Antona Peñalba, Beatriz; Barrio De Santos, Ana Rosa; González Pérez, Mariano; Pérez Garmendia, Carlos; Serramito Blanco, María; Privado Aroco, Ana; Almalki, Wael; Bodas Romero, Julia; Ouzzani, Mohamed; Paune, Jaume; Calderón García, Raquel; Pitarch Velasco, Aída; Cebrián, José Luis; Sánchez Pérez, María Isabel; García Rojo, Marta María; Bonnin Arias, Cristina Natalia; Sánchez Ramos, Celia; Gutiérrez Jorrín, Sara Carmen; Rodríguez Alonso, Xabier; Laucirica Sáenz, Gorka; Arranz Márquez, Esther; Alonso Castellanos, Miriam; Teus Guezala, Miguel Ángel; Hernández Verdejo, José Luis; Mármol Errasti, Esther; Martín García, Beatriz; Arriola Villalobos, Pedro; Gómez De Liaño Sánchez, María Rosario; Mínguez Caro, N; Orduña Azcona, Javier; Navarro Gil, Francisco Javier; Huete Toral, Fernando; Rodríguez Pomar, Candela; Martínez Águila, Alejandro; Martín Gil, Alba; Tomé de la Torre, Miguel ÁngelPor cuarto año consecutivo los doctorandos de la Facultad de Óptica y Optometría de la Universidad Complutense de Madrid cuentan con un congreso propio organizado por y para ellos, el 4º PhDAY- FOO. Se trata de un congreso gratuito abierto en la que estos jóvenes científicos podrán presentar sus investigaciones al resto de sus compañeros predoctorales y a toda la comunidad universitaria que quiera disfrutar de este evento. Apunta en tu agenda: el 15 de octubre de 2020. En esta ocasión será un Congreso On-line para evitar que la incertidumbre asociada a la pandemia Covid-19 pudiera condicionar su celebración.Item Silencing of P2Y2 receptors reduces intraocular pressure in New Zealand rabbits(British Journal of Pharmacology, 2012) María Jesús Perez de Lara; Concepción Santano; Martín Gil, Alba; Crooke Álvarez, Almudena; Peral Cerda, María Asunción; Pintor Just, Jesús JerónimoBACKGROUND AND PURPOSE: P2 receptors are involved in the regulation of ocular physiological processes like intraocular pressure (IOP). In the present study, the involvement of P2Y2 receptors in the hypertensive effect of nucleotides was investigated by use of antagonists and of a siRNA designed for the P2Y2 receptor. EXPERIMENTAL APPROACH: Agonists of the P2Y2 receptor a as well as P2 antagonists were applied to eyes of New Zealand rabbits, and the changes in IOP were followed for up to 6 h. Cloning of the P2Y2 receptor cDNA was done using a combination of degenerate reverse transcription PCR (RT-PCR) and rapid amplification of cDNA ends (RACE). siRNA was synthesized and tested by immunohistochemistry. KEY RESULTS: Single doses of 2-thioUTP, UTP-γ-S and UTP increased IOP. This behaviour was concentration-dependent and partially antagonized by reactive blue 2. Silencing the P2Y2 receptor was observed in the ciliary body by immunohistochemistry labelling, where a reduction in the immunofluorescence was observed. This reduction in the expression of the P2Y2 receptor was concomitant with a reduction in IOP, which was measurable 24 h after treatment with the siRNA, maximal after 2 days, followed by a slow increase towards control values for the following 5 days. Application of the P2Y2 agonists after pretreatment of the animals with this siRNA did not produce any change in IOP. CONCLUSIONS AND IMPLICATIONS: P2Y2 receptors increase IOP in New Zealand rabbits. The application of a siRNA for this receptor significantly reduced IOP, suggesting that this technology might be used for the treatment of glaucoma.