Lucas Ruiz, Fernando

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Lucas Ruiz, Fernando

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Lucas Ruiz, Fernando

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Universidad de Murcia. Departamento de Oftalmología, Optometría, Otorrinolaringologíay Anatomía Patológica

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Open Access
Pan-retinal ganglion cell markers in mice, rats, and rhesus macaques
(Kunming Institute of Zoology; the Chinese Academy of Sciences; and the China Zoological Society, 2022-12-16) Nadal-Nicolás, Francisco Manuel; Galindo Romero, Caridad; Lucas Ruiz, Fernando; Marsh-Amstrong, Nicholas; Li, Wei; Vidal Sanz, Manuel; Agudo Barriuso, Marta; Oftalmología, Optometría, Otorrinolaringología y Anatomía Patológica; Facultad de Medicina
Univocal identification of retinal ganglion cells (RGCs) is an essential prerequisite for studying their degeneration and neuroprotection. Before the advent of phenotypic markers, RGCs were normally identified using retrograde tracing of retinorecipient areas. This is an invasive technique, and its use is precluded in higher mammals such as monkeys. In the past decade, several RGC markers have been described. Here, we reviewed and analyzed the specificity of nine markers used to identify all or most RGCs, i.e., pan-RGC markers, in rats, mice, and macaques. The best markers in the three species in terms of specificity, proportion of RGCs labeled, and indicators of viability were BRN3A, expressed by vision-forming RGCs, and RBPMS, expressed by vision- and non-vision-forming RGCs. NEUN, often used to identify RGCs, was expressed by non-RGCs in the ganglion cell layer, and therefore was not RGC-specific. γ-SYN, TUJ1, and NF-L labeled the RGC axons, which impaired the detection of their somas in the central retina but would be good for studying RGC morphology. In rats, TUJ1 and NF-L were also expressed by non-RGCs. BM88, ERRβ, and PGP9.5 are rarely used as markers, but they identified most RGCs in the rats and macaques and ERRβ in mice. However, PGP9.5 was also expressed by non-RGCs in rats and macaques and BM88 and ERRβ were not suitable markers of viability.
Open Access
7,8-Dihydroxiflavone protects adult rat axotomized retinal ganglion cells through MAPK/ERK and PI3K/AKT activation
(MDPI, 2021-10-08) Galindo Romero, Caridad; Vidal-Villegas, Beatriz; Asís-Martínez, Javier; Lucas Ruiz, Fernando; Gallego Ortega, Alejandro; Vidal Sanz, Manuel; Oftalmología, Optometría, Otorrinolaringología y Anatomía Patológica; Facultad de Óptica y Optometría
We analyze the 7,8-dihydroxyflavone (DHF)/TrkB signaling activation of two main intracellular pathways, mitogen-activated protein kinase (MAPK)/ERK and phosphatidylinositol 3 kinase (PI3K)/AKT, in the neuroprotection of axotomized retinal ganglion cells (RGCs). Methods: Adult albino Sprague-Dawley rats received left intraorbital optic nerve transection (IONT) and were divided in two groups. One group received daily intraperitoneal DHF (5 mg/kg) and another vehicle (1%DMSO in 0.9%NaCl) from one day before IONT until processing. Additional intact rats were employed as control (n = 4). At 1, 3 or 7 days (d) after IONT, phosphorylated (p)AKT, p-MAPK, and non-phosphorylated AKT and MAPK expression levels were analyzed in the retina by Western blotting (n = 4/group). Radial sections were also immunodetected for the above-mentioned proteins, and for Brn3a and vimentin to identify RGCs and Müller cells (MCs), respectively (n = 3/group). Results: IONT induced increased levels of p-MAPK and MAPK at 3d in DHF- or vehicle-treated retinas and at 7d in DHF-treated retinas. IONT induced a fast decrease in AKT in retinas treated with DHF or vehicle, with higher levels of phosphorylation in DHF-treated retinas at 7d. In intact retinas and vehicle-treated groups, no p-MAPK or MAPK expression in RGCs was observed. In DHF- treated retinas p-MAPK and MAPK were expressed in the ganglion cell layer and in the RGC nuclei 3 and 7d after IONT. AKT was observed in intact and axotomized RGCs, but the signal intensity of p-AKT was stronger in DHF-treated retinas. Finally, MCs expressed higher quantities of both MAPK and AKT at 3d in both DHF- and vehicle-treated retinas, and at 7d the phosphorylation of p-MAPK was higher in DHF-treated groups. Conclusions: Phosphorylation and increased levels of AKT and MAPK through MCs and RGCs in retinas after DHF-treatment may be responsible for the increased and long-lasting RGC protection afforded by DHF after IONT.
Open Access
Systemic and Intravitreal Antagonism of the TNFR1 Signaling Pathway Delays Axotomy-Induced Retinal Ganglion Cell Loss
(Frontiers Media, 2019-10-15) Lucas Ruiz, Fernando; Galindo Romero, Caridad; Salinas Navarro, Manuel Ángel; González Riquelme, María Josefa; Vidal Sanz, Manuel; Agudo Barriuso, Marta; Anatomía Humana y Psicobiología
Here, we have blocked the signaling pathway of tumor necrosis factor α (TNFα) in a mouse model of traumatic neuropathy using a small cell permeable molecule (R7050) that inhibits TNFα/TNF receptor 1 (TNFR1) complex internalization. Adult pigmented mice were subjected to intraorbital optic nerve crush (ONC). Animals received daily intraperitoneal injections of R7050, and/or a single intravitreal administration the day of the surgery. Some animals received a combinatorial treatment with R7050 (systemic or local) and a single intravitreal injection of brain derived neurotrophic factor (BDNF). As controls, untreated animals were used. Retinas were analyzed for RGC survival 5 and 14 days after the lesion i.e., during the quick and slow phase of axotomy-induced RGC death. qPCR analyses were done to verify that Tnfr1 and TNFα were up-regulated after ONC. At 5 days post-lesion, R7050 intravitreal or systemic treatment neuroprotected RGCs as much as BDNF alone. At 14 days, RGC rescue by systemic or intravitreal administration of R7050 was similar. At this time point, intravitreal treatment with BDNF was significantly better than intravitreal R7050. Combinatory treatment was not better than BDNF alone, although at both time points, the mean number of surviving RGCs was higher. In conclusion, antagonism of the extrinsic pathway of apoptosis rescues axotomized RGCs as it does the activation of survival pathways by BDNF. However, manipulation of both pathways at the same time, does not improve RGC survival.
Open Access
Mesenchymal stromal cell therapy for damaged retinal ganglion cells, is gold all that glitters?
(Wolters Kluwer – Medknow Publications, 2019-11-01) Lucas Ruiz, Fernando; Galindo Romero, Caridad; García Bernal, David; Norte Muñoz, María; Rodríguez Ramírez, Kristy T.; Salinas Navarro, Manuel Ángel; Millán Rivero, José E.; Vidal Sanz, Manuel; Agudo Barriuso, Marta; Anatomía Humana y Psicobiología
Mesenchymal stromal cells are an excellent source of stem cells because they are isolated from adult tissues or perinatal derivatives, avoiding the ethical concerns that encumber embryonic stem cells. In preclinical models, it has been shown that mesenchymal stromal cells have neuroprotective and immunomodulatory properties, both of which are ideal for central nervous system treatment and repair. Here we will review the current literature on mesenchymal stromal cells, focusing on bone marrow mesenchymal stromal cells, adipose-derived mesenchymal stromal cells and mesenchymal stromal cells from the umbilical cord stroma, i.e., Wharton’s jelly mesenchymal stromal cells. Finally, we will discuss the use of these cells to alleviate retinal ganglion cell degeneration following axonal trauma.
Open Access
Shared and differential retinal responses against optic nerve injury and ocular hypertension
(Frontiers Media SA., 2017-04-26) Vidal Sanz, Manuel; Galindo Romero, Caridad; Valiente-Soriano, Francisco J.; Nadal-Nicolás, Francisco Manuel; Ortín-Martínez, Arturo; Rovere, Giuseppe; Salinas Navarro, Manuel Ángel; Lucas Ruiz, Fernando; Sánchez-Migallón, María C.; Sobrado Calvo, Paloma; Avilés Trigueros, Marcelino; Villegas Pérez, Maria Paz; Agudo Barriuso, Marta; Oftalmología, Optometría, Otorrinolaringología y Anatomía Patológica; Facultad de Medicina
Glaucoma, one of the leading causes of blindness worldwide, affects primarily retinal ganglion cells (RGCs) and their axons. The pathophysiology of glaucoma is not fully understood, but it is currently believed that damage to RGC axons at the optic nerve head plays a major role. Rodent models to study glaucoma include those that mimic either ocular hypertension or optic nerve injury. Here we review the anatomical loss of the general population of RGCs (that express Brn3a; Brn3a+RGCs) and of the intrinsically photosensitive RGCs (that express melanopsin; m+RGCs) after chronic (LP-OHT) or acute (A-OHT) ocular hypertension and after complete intraorbital optic nerve transection (ONT) or crush (ONC). Our studies show that all of these insults trigger RGC death. Compared to Brn3a+RGCs, m+RGCs are more resilient to ONT, ONC, and A-OHT but not to LP-OHT. There are differences in the course of RGC loss both between these RGC types and among injuries. An important difference between the damage caused by ocular hypertension or optic nerve injury appears in the outer retina. Both axotomy and LP-OHT induce selective loss of RGCs but LP-OHT also induces a protracted loss of cone photoreceptors. This review outlines our current understanding of the anatomical changes occurring in rodent models of glaucoma and discusses the advantages of each one and their translational value.
Open Access
Restorative potential of ciliary body cells in a retinal ganglion cell degeneration model
(Nature Research, 2025-05-03) Fernández-Nogales, Marta; Herrera, Macarena; Herrera, Eloisa ; Lucas Ruiz, Fernando; Valiente Soriano, Francisco Javier; Nadal-Nicolás, Francisco Manuel; Agudo Barriuso, Marta; Lucas Ruiz, Fernando; Oftalmología, Optometría, Otorrinolaringología y Anatomía Patológica; Facultades de la UMU::Facultad de Medicina
The ciliary body (CB) has been proposed as a niche of neural stem cells because, in vitro, cells from this area are able to form neurospheres, proliferate and differentiate. Here, we explore the potential of CB cells to differentiate and replace degenerated retinal ganglion cells (RGCs) in vivo. CB cells and cells from the subventricular zone (SVZ) were isolated from adult or postnatal C57BL/6Tg(CAG-EGFP) mice, respectively, and intravitreally injected into intact retinas, immediately after optic nerve crush or 45 days after the lesion of adult C57/BL/6 mice. Retinas were analysed in whole mounts or cross sections at different time points. Controls were matched untreated retinas. Neither cell type caused gliosis or toxicity when injected into intact retinas. When CB or SVZ cells were injected right after axotomy, they formed an epimembrane without integrating in the retina. However, when CB cells were administered in retinas depleted of RGCs, they integrated into the ganglion cell layer and expressed RGC and neuronal markers. Although SVZ cells were also able to integrate into RGC depleted retinas they did so more slowly than CB cells. These results shed light in the long-standing question of whether cells in the CB have the potential to transdifferentiate in vivo and point to the CB as a suitable source of cells that could be used in cell-replacement therapies for neurodegenerative diseases of the retina.