ROS-Induced JNK and p38 Signaling Is Required for Unpaired Cytokine Activation during Drosophila Regeneration

Upon apoptotic stimuli, epithelial cells compensate the gaps left by dead cells by activating proliferation. This has led to the proposal that dying cells signal to surrounding living cells to maintain homeostasis. Although the nature of these signals is not clear, reactive oxygen species (ROS) could act as a signaling mechanism as they can trigger pro-inflammatory responses to protect epithelia from environmental insults. Whether ROS emerge from dead cells and what is the genetic response triggered by ROS is pivotal to understand regeneration of Drosophila imaginal discs. We genetically induced cell death in wing imaginal discs, monitored the production of ROS and analyzed the signals required for repair. We found that cell death generates a burst of ROS that propagate to the nearby surviving cells. Propagated ROS activate p38 and induce tolerable levels of JNK. The activation of JNK and p38 results in the expression of the cytokines Unpaired (Upd), which triggers the JAK/STAT signaling pathway required for regeneration. Our findings demonstrate that this ROS/JNK/p38/Upd stress responsive module restores tissue homeostasis. This module is not only activated after cell death induction but also after physical damage and reveals one of the earliest responses for imaginal disc regeneration

Ciencia, Tecnología y Salud en la Atención de los Adolescentes

El Centro de Investigación en Ciencias Médicas (CICMED) de la Universidad Autónoma del Estado de México, institución que se distingue por su preocupación en el desarrollo de proyectos de investigación, actividades académicas y de vinculación encaminadas a la detección y tratamiento de aquellos problemas que aquejan a los adolescentes, se ha distinguido por la promoción de la salud del adolescente, además de un trabajo intenso a través de redes de investigación con otras instituciones con lo cual se han enriquecido nuestros proyectos de investigación, permitiendo establecer convenios con instituciones tanto nacionales como internacionales. Los temas que se tratan en este libro, han dado lugar a una gran cantidad de reflexiones, mitos, estrategias y propuestas para la cuidado del adolescente, motivo por el cual se invitó tanto a docentes como a investigadores a participar en este libro y poder mostrar lo que se está haciendo actualmente en relación con la atención de la salud del adolescente desde diferentes perspectivas temáticas, con el propósito de difundir ampliamente los hallazgos que como investigadores se han encontrado a través del trabajo clínico y de campo, aportar información relevante para la prevención y tratamiento de la problemática más frecuente en el adolescente y abrir un espacio de intercambio y actualización, basado en el trabajo interdisciplinario para entender mejor y ampliar la visión de los diferentes factores que inciden en la salud integral del adolescente. El adolescente concebido como la persona que está en la transición de la niñez a la vida adulta, ha existido siempre. Sin embargo, la adolescencia entendida como un proceso bio-psico-social con entidad propia y, en consecuencia, con características necesidades y problemas específicos, es una realidad que apenas ha comenzado a tenerse en cuenta recientemente. La adolescencia es el período de tiempo en el que se producen los cambios desde la inmadurez propia del niño a la madurez propia del adulto. Entendiendo al individuo maduro como: aquel que es capaz de orientar su propia vida según el sentido de la existencia, con criterios propios. Desde el punto de vista intelectual o mental, el individuo maduro es aquel que es capaz de juzgar con independencia y objetividad, con sentido crítico. Desde la vertiente afectiva, posee autocontrol de las emociones y es capaz de aceptar los fracasos sin grandes conmociones interiores. Desde el aspecto social, está dispuesto a colaborar en las tareas colectivas, es tolerante con los demás y es capaz de asumir su propia responsabilidad. Esta etapa de la vida de los seres humanos es importante ya que depende del desarrollo armónico de ésta, obliga al futuro adulto en su mundo globalizado lleno de exigencias y riesgos, que los profesionales de la salud reconsideren su actuar con estas personas, dado que hoy en día no hay especialistas o posgraduados que atiendan de manera específica a los adolescentes. En este sentido la Universidad Autónoma del Estado de México, a través del Centro de Investigación en Ciencias Médica adquiere la responsabilidad y el compromiso de motivar a padres, educadores, profesionales de la salud y a los mismos adolescentes para que se conviertan en promotores de la salud física y mental que bajo la misma meta en la escuela y el hogar promuevan una conducta saludable. Por lo que brindamos nuestro agradecimiento a los autores por su valiosa participación y confianza al dejar en nuestras manos su trabajo profesional, comprometido y entusiasta, con el propósito de aportar a la sociedad una perspectiva general sobre el adolescente y sus trastornos para ubicarnos después en un contexto de salud integral

Potencial de la gramática del MuSIASEM en la representación del análisis de la sostenibilidad

Una alta capacidad para representar el análisis de la sostenibilidad es la exigencia central de cualquier metodología que pretenda encontrar en dicho análisis una herramienta para el desarrollo social y un diagnóstico de relación de las sociedades humanas con el entorno. Los requerimientos de dicha capacidad representadora se cifran en un lenguaje lo suficientemente recursivo y sintético en sus rasgos y principios como para ser flexible y en ello ofrecer pertinencia y operacionalidad para diversidad de escenarios y situaciones que, en todo caso, se caracterizan por la complejidad del tejido entre sus múltiples niveles y dimensiones. A través de una gramática el MUSIASEM ha cifrado en ello su potencial analítico y predictivo. El presente artículo argumenta las ventajas y alcances de las gramáticas como alternativa a los modelos en la representación del análisis de la sostenibilidad, explica cómo está fundamentado y construido el caso de la gramática del MUSIASEM y expone aplicaciones ya elaboradas a los temas de alimentos, suelos, energía, construcción y energía

ROS-Induced JNK and p38 Signaling Is Required for Unpaired Cytokine Activation during Drosophila Regeneration

Upon apoptotic stimuli, epithelial cells compensate the gaps left by dead cells by activating proliferation. This has led to the proposal that dying cells signal to surrounding living cells to maintain homeostasis. Although the nature of these signals is not clear, reactive oxygen species (ROS) could act as a signaling mechanism as they can trigger pro-inflammatory responses to protect epithelia from environmental insults. Whether ROS emerge from dead cells and what is the genetic response triggered by ROS is pivotal to understand regeneration of Drosophila imaginal discs. We genetically induced cell death in wing imaginal discs, monitored the production of ROS and analyzed the signals required for repair. We found that cell death generates a burst of ROS that propagate to the nearby surviving cells. Propagated ROS activate p38 and induce tolerable levels of JNK. The activation of JNK and p38 results in the expression of the cytokines Unpaired (Upd), which triggers the JAK/STAT signaling pathway required for regeneration. Our findings demonstrate that this ROS/JNK/p38/Upd stress responsive module restores tissue homeostasis. This module is not only activated after cell death induction but also after physical damage and reveals one of the earliest responses for imaginal disc regeneration

ROS-Induced JNK and p38 Signaling Is Required for Unpaired Cytokine Activation during <i>Drosophila</i> Regeneration

<div><p>Upon apoptotic stimuli, epithelial cells compensate the gaps left by dead cells by activating proliferation. This has led to the proposal that dying cells signal to surrounding living cells to maintain homeostasis. Although the nature of these signals is not clear, reactive oxygen species (ROS) could act as a signaling mechanism as they can trigger pro-inflammatory responses to protect epithelia from environmental insults. Whether ROS emerge from dead cells and what is the genetic response triggered by ROS is pivotal to understand regeneration of <i>Drosophila</i> imaginal discs. We genetically induced cell death in wing imaginal discs, monitored the production of ROS and analyzed the signals required for repair. We found that cell death generates a burst of ROS that propagate to the nearby surviving cells. Propagated ROS activate p38 and induce tolerable levels of JNK. The activation of JNK and p38 results in the expression of the cytokines Unpaired (Upd), which triggers the JAK/STAT signaling pathway required for regeneration. Our findings demonstrate that this ROS/JNK/p38/Upd stress responsive module restores tissue homeostasis. This module is not only activated after cell death induction but also after physical damage and reveals one of the earliest responses for imaginal disc regeneration.</p></div

p38 and JNK are activated independently.

<p>(A) <i>Hep</i>^<i>r75</i> hemizygous disc cut (wedge) and stained with P-p38. Sketch of wing discs with square indicate location of images. (B) <i>Hep</i>^<i>r75</i> hemizygous disc after <i>ptc>rpr</i> induction and stained for P-p38. Dead domain is outlined white. TP-3; TO-PRO-3. (C) Wild type and <i>p38a</i>^<i>1-/-</i> discs, cultured for 7h showing <i>TRE-red</i> activation close to the cut edges. (D) Mean pixel intensity for <i>TRE-red</i> measured in discs with physical injury in wild type (88.24 ± 22.58; S.D.) and <i>p38a</i>^<i>1-/-</i> (70.80 ± 19.14; S.D.). P = 0.33 n.s.</p

ROS produced after physical injury and after cell death.

<p>(A) Cut disc cultured ex vivo (white wedge indicates cut edges) and thermal LUT of CellROX Green. (B) Cut disc cultured ex vivo, imaged at just after cut (0–5’) and 30’ later. Thermal scale indicates pixel intensity. (C) Sketch of wing imaginal discs with the area (black square) shown in A, B and E. (D) Fixed disc stained for nuclei to show disc contour (TP-3: TO-PRO-3) and caspase-3 after <i>ptc>rpr</i> activation for 11h at 29°C. (E) <i>ptc>rpr</i> disc cultured ex vivo; basal images at the bottom, apical at the top. Left, cell death (TO-PRO-3). Right, thermal LUT taken from the ROS channel (CellROX Green) of the same preparation. Note that most dead cells (TO-PRO-3 positive) show high ROS (red in thermal image) whereas living cells (TO-PRO-3 negative) had low ROS (green-cyan in thermal image). (F) Mean pixel intensity (grey value) of the indicated zones in control discs without cell death (<i>ptc>rpr</i> OFF) and discs with cell death (<i>ptc>rpr</i> ON). The pixel intensity in the <i>ptc</i> domain in the absence of cell death (<i>ptc>rpr</i> OFF) was 1.76 ± 0.55 (SD; from 48 regions of interest [ROI] in n = 5 discs). The mean pixel intensity for the apoptotic region (basal; <i>ptc>rpr</i> ON) was 33.14±8.18 (SD), measured in 27 ROI on confocal images taken from n = 6 discs. Living cells adjacent to the apoptotic zone showed a mean grey value of 8.51±2.12 (SD; 15 ROI from 6 discs taken from cells near the <i>ptc</i> domain). White rectangles in E: example ROI for Basal Apoptotic Zone (1) and Apical Living Zone (2). The ROI’s for the <i>ptc</i> Zone, in discs in which <i>ptc>rpr</i> is OFF, were placed as (1). ***<i>P</i><0.001. Thermal scale indicates sample values from raw images.</p

ROS control JNK activity.

<p>(A) Test of JNK reporters. All images in A correspond to the same disc after <i>ptc>rpr</i> induction. Top row: apical sections. Bottom row: basal sections. Note that <i>puc</i> is more abundant in apical than basal sections, particularly in the notum (n; arrowhead) and wing pouch (wp; arrow). Cell death (TUNEL) and high <i>TRE-red</i> are more abundant in basal sections. (B) Zoom of a digital cross section of the zone marked with a white line in A. Endogenous <i>puc-lacZ</i> is found in the outer layer of peripodial membrane cells (pm). <i>Puc-lacZ</i> cells in the disc columnar epithelium are apical (white), most apoptotic cells are basal (red), and <i>TRE-red</i> positive cells are apical and basal (blue). (C) Three digital cross section in an apical <i>puc-lacZ</i> zone of the wing pouch (wp) and notum (n1, n2). Each example contains three to four cells with co-localization of ß-galactosidase and EdU. (D) <i>TRE-red</i> reporter in <i>ptc>rpr</i> discs of larvae fed with standard food or NAC-supplemented food (NAC). TP-3: TO-PRO-3. (E) Mean pixel intensities of <i>TRE-red</i> reporter in <i>ptc>rpr</i> discs with standard or NAC food. The pixel intensity for standard food was 26.06 ± 7.22 (S.D.; n = 15) and for NAC 18.12 ± 8.32 (S.D.; n = 25). (F) <i>TRE-red</i> reporter expression in physically injured discs, cultured for 7 h ex vivo in Schneider’s culture medium with or without NAC. Outline: disc contour. Wedges: cut. (G) Mean pixel intensities of <i>TRE-red</i> reporter in ex vivo cultured discs with or without NAC. The pixel intensity for standard culture was 88.98 ± 22.25 (S.D.; n = 6) and for NAC 23.98 ± 10.26 (S.D.; n = 16). **<i>P</i><0.01, ***<i>P</i><0.001.</p

ROS are required for tissue repair.

<p>(A) Design for chemical antioxidant intake and cell death induction. At 17°C and 24 h before cell death induction, larvae were transferred to a vial with standard food supplemented with antioxidant. Cell death (<i>sal</i>^<i>E/Pv</i><i>>rpr</i> ON) was induced by shifting temperature to 29°C for 11 h (blue stripes in the disc). Larvae were transferred to 17°C where they regenerated and emerged into adults, in which wings were scored. Blue color in the wing: area emerged from <i>sal</i>^<i>E/Pv</i>. Controls <i>sal</i>^<i>E/Pv</i><i>>rpr</i> OFF were kept at 17°C to avoid cell death. (B) Percentage of regenerated wings after cell death (<i>sal</i>^<i>E/Pv</i><i>>rpr</i> ON) in the absence of antioxidant (Std Food), or in the presence of antioxidants (NAC, Trolox or Vitamin C). (C) Examples of <i>sal</i>^<i>E/Pv</i><i>>rpr</i> ON wings with the indicated food supplement. In controls without antioxidants (Std Food), the complete wing recovered. For each antioxidant an example of incomplete regeneration after cell death induction is shown. (D) Mitosis number in <i>ptc>rpr</i> discs from larvae fed with and without NAC and with or without <i>rpr</i>-ablation (ON versus OFF). <i>Ptc>rpr</i> OFF: 41.86 ± 9.84 (S.D.); NAC <i>ptc>rpr</i> OFF: 39.9 ± 4.68 (S.D.); <i>ptc>rpr</i> ON: 49.73 ± 8.18 (S.D.); NAC <i>ptc>rpr</i> ON: 29.52 ± 9.41 (S.D.) (E) Design for ectopic expression of enzymatic antioxidant transgenes and simultaneous cell death induction when shifted to 29°C for 11 h. The Gal4/UAS (red) activate Cat, Sod or Cat+Sod transgenes. Blue striped area: <i>sal</i>^<i>E/Pv</i><i>-LHG lexO-rpr</i>. Adult wings were scored for complete regeneration of the missing zone. Red coloration indicates zone influenced by the enzymatic antioxidant; purple: zone influenced by enzymatic antioxidant and cell death. <i>sal</i>^<i>E/Pv</i><i>-LHG</i> and <i>nub-Gal4</i> are under the control of <i>tubGal80</i>^<i>TS</i>. (F) Percentage of regenerated wings in Cat, Sod or Cat and Sod ectopically expressed transgenes. (G) Wings from individuals after cell death and transgene activation (ON). For Cat, Sod or Sod:Cat and example of incomplete regeneration is shown. TP-3: TO-PRO-3. ***<i>P</i><0.001</p

ROS stimulate p38 phosphorylation.

<p>(A) P-p38 staining of intact (uncut, controls) and cut discs cultured for the indicated times after injury. White lines: wound edges; white arrowhead: small incision. (B) Discs cultured with or without NAC, cut and stained for P-p38. (C) Mean pixel intensities of P-p38 fluorescence from cut discs cultured with standard medium (95.29 ± 17.52; S.D.) or NAC-supplemented (22.45±2.56; S.D.). (D) Apical and basal images of P-p38 after <i>ptc>rpr</i> induction. (E) Apical and basal images of <i>ptc>rpr</i> after NAC supplementation showing reduction of P-p38 localization. (F) Mean pixel intensities of P-p38 fluorescent labeling from <i>ptc>rpr</i> discs fed with standard (52.17±19.96; S.D.) or NAC-supplemented food (7,85 ± 2,42; S.D.). (G) Genetic ROS scavenging using <i>ci>Sod</i>:<i>Cat</i>, activated in the anterior compartment (<i>ci</i>, black in the sketch). <i>Sal</i>^<i>EPv</i><i>>rpr</i> cell death (blue in the sketch) in the same disc results in inhibition of P-p38 only in anterior compartment. TP-3: TO-PRO-3 nuclei staining. Outlined white in D, E and G: apoptotic zone. ***<i>P</i><0.001.</p