Neuroimmune interactions in the cornea: effects of resident dendritic cell depletion on cold nerve terminal activity, basal tearing, and pain

La córnea es un tejido conectivo avascular que resulta crucial, no sólo como barrera primaria del ojo, sino también como transparente estructura refractiva. Este tejido avascular y con privilegio inmune resulta un modelo ideal para estudiar las interacciones que se producen entre sus densos nervios sensoriales (fácilmente accesibles tanto para su registro electrofisiológico focal como para su estudio morfológico) y sus pocos tipos de células inmunes residentes, que se distinguen de las que migran desde los vasos sanguíneos circundantes. Las diferentes funciones de las células inmunes residentes en la córnea se han evaluado en condiciones de enfermedad y patología corneales, sin embargo, su contribución al mantenimiento de la homeostasis sigue por determinar. El objetivo de la presente Tesis fue demostrar la posible interacción funcional entre las células dendríticas (CD) y los nervios sensoriales de la córnea en condiciones basales (es decir, en ausencia de cualquier inflamación o daño) y sus posibles consecuencias en los procesos protectores de la superficie ocular, como son el lagrimeo o el comportamiento nociceptivo. Para ello, se indujo una depleción de las CD residentes a corto y largo plazo en la córnea de ratones transgénicos CD11c-DTR+ (5 meses, ambos sexos) mediante inyecciones subconjuntivales de toxina diftérica (TD) para determinar los efectos de la ausencia de las CD en la actividad de los nervios sensoriales de frío corneales, el lagrimeo basal y el dolor. La actividad de los nervios sensoriales se estudió mediante registros electrofisiológicos ex vivo; la tasa de lagrimeo basal se midió con hilos comerciales de rojo de fenol y el dolor espontáneo se monitorizó midiendo la ratio de cierre del ojo. Por último, para evaluar la morfología y la distribución de las CD en la córnea, se llevaron a cabo montajes de córnea completa en las que se observaba la fluorescencia endógena de las células. La depleción local de las CD alteró la actividad de los impulsos nerviosos de las terminaciones (INT) de los termorreceptores de frío en respuesta a los cambios de temperatura: mientras que la depleción de las CD a corto plazo producía una sensibilización de los termorreceptores de frío, especialmente en su respuesta al calor, la depleción de CD a largo plazo producía una desensibilización de los termorreceptores de frío en su respuesta al frío y tuvo un impacto en la forma del INT. Además, la ausencia de las CD produjo dolor tanto a corto como a largo plazo, siendo este dolor mayor en condiciones agudas en las que, además, la tasa de lagrimeo basal se redujo significativamente hasta casi la mitad. La repoblación de las CD en la córnea produjo una recuperación casi total de la actividad y la forma de los INT de los termorreceptores de frío en condiciones naif, aunque todavía se observaron signos de dolor espontáneo. Por último, cuando analizamos el efecto de la propia inyección subconjuntival utilizando solución salina tamponada con fosfato en lugar de TD, observamos un aumento de las respuestas al frío y algunos cambios en la forma del INT, lo que sugiere una lesión leve debida al procedimiento de inyección. Sin embargo, estos cambios fueron opuestos a los observados en condiciones de depleción de las CD y ni la tasa de lagrimeo basal ni el dolor se vieron afectados en estos animales. En conclusión, nuestros datos experimentales demuestran que existe una interacción funcional entre las CD residentes y los nervios sensoriales corneales en condiciones basales. Esto sugiere un papel crucial de las CD en la actividad de los nervios corneales y en el mantenimiento de la homeostasis de la superficie ocular

A genetic compensatory mechanism regulated by Jun and Mef2d modulates the expression of distinct class IIa Hdacs to ensure peripheral nerve myelination and repair

The class IIa histone deacetylases (HDACs) have pivotal roles in the development of different tissues. Of this family, Schwann cells express Hdac4, 5 and 7 but not Hdac9. Here we show that a transcription factor regulated genetic compensatory mechanism within this family of proteins, blocks negative regulators of myelination ensuring peripheral nerve developmental myelination and remyelination after injury. Thus, when Hdac4 and 5 are knocked-out from Schwann cells in mice, a JUN-dependent mechanism induces the compensatory overexpression of Hdac7 permitting, although with a delay, the formation of the myelin sheath. When Hdac4,5 and 7 are simultaneously removed, the Myocyte-specific enhancer-factor d (MEF2D) binds to the promoter and induces the de novo expression of Hdac9, and although several melanocytic lineage genes are misexpressed and Remak bundle structure is disrupted, myelination proceeds after a long delay. Thus, our data unveil a finely tuned compensatory mechanism within the class IIa Hdac family, coordinated by distinct transcription factors, that guarantees the ability of Schwann cells to myelinate during development and remyelinate after nerve injury

Conocimiento objetivo y subjetivo de los consumidores españoles sobre el pescado

Product knowledge has been recognised as an important factor influencing consumer decision making. The aim of this study was to assess Spanish consumers’ knowledge about fish as well as their relationship with certain socio-demographic characteristics. Objective and subjective knowledge were measured by means of 5 and 4 items respectively. Overall results revealed the existence of a certain lack of knowledge about fish, although some differences were observed depending on gender, age, education level, region of residence and growing up region (coastal or interior). A positive relationship between objective and subjective knowledge was also detected

An experimental model of neuro–immune interactions in the eye: Corneal sensory nerves and resident dendritic cells

This article belongs to the Special Issue Immune Pathogenesis and Regulation of Ocular Inflammation.The cornea is an avascular connective tissue that is crucial, not only as the primary barrier of the eye but also as a proper transparent refractive structure. Corneal transparency is necessary for vision and is the result of several factors, including its highly organized structure, the physiology of its few cellular components, the lack of myelinated nerves (although it is extremely innervated), the tightly controlled hydration state, and the absence of blood and lymphatic vessels in healthy conditions, among others. The avascular, immune-privileged tissue of the cornea is an ideal model to study the interactions between its well-characterized and dense sensory nerves (easily accessible for both focal electrophysiological recording and morphological studies) and the low number of resident immune cell types, distinguished from those cells migrating from blood vessels. This paper presents an overview of the corneal structure and innervation, the resident dendritic cell (DC) subpopulations present in the cornea, their distribution in relation to corneal nerves, and their role in ocular inflammatory diseases. A mouse model in which sensory axons are constitutively labeled with tdTomato and DCs with green fluorescent protein (GFP) allows further analysis of the neuro-immune crosstalk under inflammatory and steady-state conditions of the eye.This work was funded by grants SAF2017–83674–C2–1–R (JG) and SAF2017–83674–C2–2–R (MCA) from the Spanish Agencia Estatal de Investigación and European Regional Development Funds “Una manera de hacer Europa”, and grant PID2020–115934RB–I00 (JG/MCA) funded by MICIN/AEI/1013039/5011100011033. Funding by the Excellence Program grant PROMETEO/2018/114 (JG) and predoctoral fellowships ACIF/2017/169 (LF–R) from the Generalitat Valenciana and PRE2018–083980 (AI–P) from MICIN/AEI is also acknowledged.Peer reviewe

Cyclosporine A Decreases Dryness-Induced Hyperexcitability of Corneal Cold-Sensitive Nerve Terminals

Cyclosporine A (CsA) is used for the treatment of dry eye (DE) with good clinical results, improving tear secretion and decreasing subjective symptoms. These effects are attributed to the improved tear film dynamics, but there are no data on the effect of CsA on the abnormal sensory nerve activity characteristic in DE. Our purpose was to evaluate the CsA effect on the enhanced activity of corneal cold thermoreceptors in a tear-deficient DE animal model using in vitro extracellular recording of cold thermoreceptors nerve terminal impulses (NTIs) before and in the presence of CsA. NTI shape was also analyzed. Blinking frequency and tearing rate were also measured in awake animals before and after topical CsA. CsA increased the tearing and blinking of treated animals. CsA significantly decreased the peak response to cold of cold thermoreceptors. Neither their spontaneous NTIs discharge rate nor their cooling threshold were modified. CsA also seemed to reverse some of the changes in NTI shape induced by tear deficiency. These data suggest that, at least in part, the beneficial clinical effects of CsA in DE can be attributed to a direct effect on sensory nerve endings, although the precise mechanisms underlying this effect need further studies to be fully clarified

The Contribution of TRPA1 to Corneal Thermosensitivity and Blink Regulation in Young and Aged Mice

The role of TRPA1 in the thermosensitivity of the corneal cold thermoreceptor nerve endings was studied in young and aged mice. The contribution of the TRPA1-dependent activity to basal tearing and thermally-evoked blink was also explored. The corneal cold thermoreceptors’ activity was recorded extracellularly in young (5-month-old) and aged (18-month-old) C57BL/6WT (WT) and TRPA1−/− knockout (TRPA1-KO) mice at basal temperature (34 °C) and during cooling (15 °C) and heating (45 °C) ramps. The blink response to cold and heat stimulation of the ocular surface and the basal tearing rate were also measured in young animals using orbicularis oculi muscle electromyography (OOemg) and phenol red threads, respectively. The background activity at 34 °C and the cooling- and heating-evoked responses of the cold thermoreceptors were similar in WT and TRPA1-KO animals, no matter the age. Similar to the aged WT mice, in the young and aged TRPA1-KO mice, most of the cold thermoreceptors presented low frequency background activity, a low cooling threshold, and a sluggish response to heating. The amplitude and duration of the OOemg signals correlated with the magnitude of the induced thermal change in the WT but not in the TRPA1-KO mice. The basal tearing was similar in the TRPA1-KO and WT mice. The electrophysiological data suggest that the TRPA1-dependent nerve activity, which declines with age, contributes to detecting the warming of the ocular surface and also to integrating the thermally-evoked reflex blink

A genetic compensatory mechanism regulated by Jun and Mef2d modulates the expression of distinct class IIa Hdacs to ensure peripheral nerve myelination and repair

The class IIa histone deacetylases (HDACs) have pivotal roles in the development of different tissues. Of this family, Schwann cells express Hdac4, 5, and 7 but not Hdac9. Here, we show that a transcription factor regulated genetic compensatory mechanism within this family of proteins, blocks negative regulators of myelination ensuring peripheral nerve developmental myelination and remyelination after injury. Thus, when Hdac4 and 5 are knocked-out from Schwann cells in mice, a JUN-dependent mechanism induces the compensatory overexpression of Hdac7 permitting, although with a delay, the formation of the myelin sheath. When Hdac4, 5, and 7 are simultaneously removed, the myocyte-specific enhancer-factor d (MEF2D) binds to the promoter and induces the de novo expression of Hdac9, and although several melanocytic lineage genes are misexpressed and Remak bundle structure is disrupted, myelination proceeds after a long delay. Thus, our data unveil a finely tuned compensatory mechanism within the class IIa Hdac family, coordinated by distinct transcription factors, that guarantees the ability of Schwann cells to myelinate during development and remyelinate after nerve injury.This work has been funded by grants from the Ministerio de Economía y Competitividad (BFU2016-75864R and PID2019-109762RB-I00), ISABIAL (UGP18-257 and UGP-2019–128) to H Cabedo, and Conselleria Educació Generalitat Valenciana (PROMETEO 2018/114) to J Gallar and H Cabedo. Predoctoral fellowships ACIF/2 017/169 from Generalitat Valenciana (to L Frutos-Rincón) and FPU16/00283from Ministerio de Universidades are also acknowledged. The Instituto de Neurociencias is a 'Center of Excellence Severo Ochoa' (Ministerio de Economía y Competitividad SEV-2013-0317).Peer reviewe