Nitric oxide induces the expression of the monocarboxylate transporter MCT4 in cultured astrocytes by a cGMP-independent transcriptional activation.

The monocarboxylate transporter MCT4 is a proton-linked carrier particularly important for lactate release from highly glycolytic cells. In the central nervous system, MCT4 is exclusively expressed by astrocytes. Surprisingly, MCT4 expression in primary cultures of mouse cortical astrocytes is conspicuously low, suggesting that an external, nonastrocytic signal is necessary to obtain the observed pattern of expression in vivo. Here, we demonstrate that nitric oxide (NO), delivered by various NO donors, time- and dose-dependently induces MCT4 expression in cultured cortical astrocytes both at the mRNA and protein levels. In contrast, NO does not enhance the expression of MCT1, the other astrocytic monocarboxylate transporter. The transcriptional effect of NO is not mediated by a cGMP-dependent mechanism as shown by the absence of effect of a cGMP analog or of a selective guanylate cyclase inhibitor. NO causes an increase in astrocytic lactate transport capacity which requires the enhancement of MCT4 expression as both are prevented by the use of a specific siRNA against MCT4. In addition, cumulated lactate release by astrocytes over a period of 24 h was also enhanced by NO treatment. Our data suggest that NO represents a putative intercellular signal to control MCT4 expression in astrocytes and in doing so, to facilitate lactate transfer to other surrounding cell types in the central nervous system. © 2011 Wiley-Liss, Inc.

Nitric oxide as a regulator of inflammatory processes

Nitric oxide (NO) plays an important role in mediating many aspects of inflammatory responses. NO is an effector molecule of cellular injury, and can act as an anti-oxidant. It can modulate the release of various inflammatory mediators from a wide range of cells participating in inflammatory responses (e.g., leukocytes, macrophages, mast cells, endothelial cells, and platelets). It can modulate blood flow, adhesion of leukocytes to the vascular endothelium and the activity of numerous enzymes, all of which can have an impact on inflammatory responses. In recent years, NO-releasing drugs have been developed, usually as derivatives of other drugs, which exhibit very powerful anti-inflammatory effects.

New insights into the anti-inflammatory actions of aspirin- induction of nitric oxide through the generation of epi-lipoxins

Aspirin has always remained an enigmatic drug. Not only does it present with new benefits for treating an ever-expanding list of apparently unrelated diseases at an astounding rate but also because aspirin enhances our understanding of the nature of these diseases processe. Originally, the beneficial effects of aspirin were shown to stem from its inhibition of cyclooxygenase-derived prostaglandins, fatty acid metabolites that modulate host defense. However, in addition to inhibiting cyclooxygenase activity aspirin can also inhibit pro-inflammatory signaling pathways, gene expression and other factors distinct from eicosanoid biosynthesis that drive inflammation as well as enhance the synthesis of endogenous protective anti-inflammatory factors. Its true mechanism of action in anti-inflammation remains unclear. Here the data from a series of recent experiments proposing that one of aspirin's predominant roles in inflammation is the induction of nitric oxide, which potently inhibits leukocyte/endothelium interaction during acute inflammation, will be discussed. It will be argued that this nitric oxide-inducing effects are exclusive to aspirin due to its unique ability, among the family of traditional anti-inflammatory drugs, to acetylate the active site of inducible cyclooxygenase and generate a family of lipid mediators called the epi-lipoxins that are increasingly being shown to have profound roles in a range of host defense responses.

Nitric oxide and the resolution of inflammation: implications for atherosclerosis

The ubiquitous free radical, nitric oxide (NO), plays an important role in many biological processes including the regulation of the inflammatory response. Alterations in NO synthesis by endogenous systems likely influence inflammatory processes occurring in a wide range of diseases including many in the cardiovascular system (e.g. atherosclerosis). Progression of inflammatory conditions depends not only upon the recruitment and activation of inflammatory cells but also upon their subsequent removal from the inflammatory milieu. Apoptosis, or programmed cell death, is a fundamental process regulating inflammatory cell survival and is critically involved in ensuring the successful resolution of an inflammatory response. Apoptosis results in shutdown of secretory pathways and renders effete, but potentially highly histotoxic, cells instantly recognisable for non-inflammatory clearance by phagocytes (e.g., macrophages). However, dysregulation of apoptosis and phagocytic clearance mechanisms can have drastic consequences for development and resolution of inflammatory processes. In this review we highlight the complexities of NO-mediated regulation of inflammatory cell apoptosis and clearance by phagocytes and discuss the molecular mechanisms controlling these NO mediated effects. We believe that manipulation of pathways involving NO may have previously unrecognised therapeutic potential for limiting or resolving inflammatory and cardiovascular disease.

Unusual astrocyte reactivity caused by the food mycotoxin ochratoxin A in aggregating rat brain cell cultures.

Ochratoxin A (OTA), a mycotoxin and widespread food contaminant, is known for its patent nephrotoxicity and potential neurotoxicity. Previous observations in vitro showed that in the CNS, glial cells were particularly sensitive to OTA. In the search for the molecular mechanisms underlying OTA neurotoxicity, we investigated the relationship between OTA toxicity and glial reactivity, in serum-free aggregating brain cell cultures. Using quantitative reverse transcriptase-polymerase chain reaction to analyze changes in gene expression, we found that in astrocytes, non cytotoxic concentrations of OTA down-regulated glial fibrillary acidic protein, while it up-regulated vimentin and the peroxisome proliferator-activated receptor-gamma expression. OTA also up-regulated the inducible nitric oxide synthase and the heme oxygenase-1. These OTA-induced alterations in gene expression were more pronounced in cultures at an advanced stage of maturation. The natural peroxisome proliferator-activated receptor-gamma ligand, 15-deoxy-delta(12,14) prostaglandin J2, and the cyclic AMP analog, bromo cyclic AMP, significantly attenuated the strong induction of peroxisome proliferator-activated receptor-gamma and inducible nitric oxide synthase, while they partially reversed the inhibitory effect of OTA on glial fibrillary acidic protein. The present results show that OTA affects the cytoskeletal integrity of astrocytes as well as the expression of genes pertaining to the brain inflammatory response system, and suggest that a relationship exists between the inflammatory events and the cytoskeletal changes induced by OTA. Furthermore, these results suggest that, by inducing an atypical glial reactivity, OTA may severely affect the neuroprotective capacity of glial cells.

Patterns of co-association of C-reactive protein and nitric oxide in malaria in endemic areas of Iran

In addition to numerous immune factors, C-reactive protein (CRP) and nitric oxide (NO) are believed to be molecules of malaria immunopathology. The objective of this study was to detect CRP and NO inductions by agglutination latex test and Griess microassay respectively in both control and malaria groups from endemic areas of Iran, including Southeastern (SE) (Sistan & Balouchestan, Hormozgan, Kerman) and Northwestern (NW) provinces (Ardabil). The results indicated that CRP and NO are produced in all malaria endemic areas of Iran. In addition, more CRP and NO positive cases were observed amongst malaria patients in comparison with those in control group. A variable co-association of CRP/NO production were detected between control and malaria groups, which depended upon the malaria endemic areas and the type of plasmodia infection. The percentage of CRP/NO positive cases was observed to be lower in NW compare to SE region, which may be due to the different type of plasmodium in the NW (Plasmodium vivax) with SE area (P. vivax, Plasmodium falciparum, mixed infection). The fluctuations in CRP/NO induction may be consistent with genetic background of patients. Although, CRP/NO may play important role in malaria, their actual function and interaction in clinical forms of disease remains unclear.

Electroencephalogram changes during inhalation with nitric oxide in the pediatric intensive care patient--a preliminary report.

OBJECTIVES: Although endogenous nitric oxide (NO) is an excitatory mediator in the central nervous system, inhaled NO is not considered to cause neurologic side effects because of its short half-life. This study was motivated by a recent case report about neurologic symptoms and our own observation of severe electroencephalogram (EEG) abnormalities during NO inhalation. DESIGN: Blind, retrospective analyses of EEGs which were registered before, during, and after NO inhalation. EEG was classified in a 5-point rating system by an independent electroencephalographer who was blinded to the patients' clinical histories. Comparisons were made with the previous evaluation documented at recording. Other EEG-influencing parameters such as oxygen saturation, hemodynamics, electrolytes, and pH were evaluated. SETTING: Pediatric intensive care unit of a tertiary care university children's hospital. PATIENTS: Eleven ventilated, long-term paralyzed, sedated children (1 mo to 14 yrs) who had EEG or clinical assessment before NO treatment and EEG during NO inhalation. They were divided into two groups according to the NO-indication (e.g., congenital heart defect, acute respiratory distress syndrome). MEASUREMENTS AND MAIN RESULTS: All 11 patients had an abnormal EEG during NO inhalation. EEG-controls without NO showed remarkable improvement. EEG abnormalities were background slowing, low voltage, suppression burst (n = 2), and sharp waves (n = 2) independent of patients' age, NO-indication, and other EEG-influencing parameters. CONCLUSIONS: These preliminary data suggest the occurrence of EEG-abnormalities after application of inhaled NO in critically ill children. We found no correlation with other potential EEG-influencing parameters, although clinical state, medication, or hypoxemia might contribute. Comprehensive, prospective, clinical assessment regarding a causal relationship between NO-inhalation and EEG-abnormalities and their clinical importance is needed.

Cells and mediators of inflammation (C-reactive protein, nitric oxide, platelets and neutrophils) in the acute and convalescent phases of uncomplicated Plasmodium vivax and Plasmodium falciparum infection

The haematological changes and release of soluble mediators, particularly C-reactive protein (CRP) and nitric oxide (NO), during uncomplicated malaria have not been well studied, especially in Brazilian areas in which the disease is endemic. Therefore, the present study examined these factors in acute (day 0) and convalescent phase (day 15) patients infected with Plasmodium falciparum and Plasmodium vivax malaria in the Brazilian Amazon. Haematologic parameters were measured using automated cell counting, CRP levels were measured with ELISA and NO plasma levels were measured by the Griess reaction. Our data indicate that individuals with uncomplicated P. vivax and P. falciparum infection presented similar inflammatory profiles with respect to white blood cells, with high band cell production and a considerable degree of thrombocytopaenia during the acute phase of infection. Higher CRP levels were detected in acute P. vivax infection than in acute P. falciparum infection, while higher NO was detected in patients with acute and convalescent P. falciparum infections. Although changes in these mediators cannot predict malaria infection, the haematological aspects associated with malaria infection, especially the roles of platelets and band cells, need to be investigated further.

Nitric oxide production by Peromyscus yucatanicus (Rodentia) infected with Leishmania (Leishmania) mexicana

Peromyscus yucatanicus (Rodentia: Cricetidae) is a primary reservoir of Leishmania (Leishmania) mexicana (Kinetoplastida: Trypanosomatidae). Nitric oxide (NO) generally plays a crucial role in the containment and elimination of Leishmania. The aim of this study was to determine the amount of NO produced by P. yucatanicus infected with L. (L.) mexicana. Subclinical and clinical infections were established in P. yucatanicus through inoculation with 1 x 10 2 and 2.5 x 10 6 promastigotes, respectively. Peritoneal macrophages were cultured alone or co-cultured with lymphocytes with or without soluble Leishmania antigen. The level of NO production was determined using the Griess reaction. The amount of NO produced was significantly higher (p ≤ 0.0001) in co-cultured macrophages and lymphocytes than in macrophages cultured alone. No differences in NO production were found between P. yucatanicus with subclinical L. (L.) mexicana infections and animals with clinical infections. These results support the hypothesis that the immunological mechanisms of NO production in P. yucatanicus are similar to those described in mouse models of leishmaniasis and, despite NO production, P. yucatanicus is unable to clear the parasite infection.

Metallothionein-1 and nitric oxide expression are inversely correlated in a murine model of Chagas disease

Chagas disease, caused by Trypanosoma cruzi, represents an endemic among Latin America countries. The participation of free radicals, especially nitric oxide (NO), has been demonstrated in the pathophysiology of seropositive individuals with T. cruzi. In Chagas disease, increased NO contributes to the development of cardiomyopathy and megacolon. Metallothioneins (MTs) are efficient free radicals scavengers of NO in vitro and in vivo. Here, we developed a murine model of the chronic phase of Chagas disease using endemic T. cruzi RyCH1 in BALB/c mice, which were divided into four groups: infected non-treated (Inf), infected N-monomethyl-L-arginine treated (Inf L-NAME), non-infected L-NAME treated and non-infected vehicle-treated. We determined blood parasitaemia and NO levels, the extent of parasite nests in tissues and liver MT-I expression levels. It was observed that NO levels were increasing in Inf mice in a time-dependent manner. Inf L-NAME mice had fewer T. cruzi nests in cardiac and skeletal muscle with decreased blood NO levels at day 135 post infection. This affect was negatively correlated with an increase of MT-I expression (r = -0.8462, p < 0.0001). In conclusion, we determined that in Chagas disease, an unknown inhibitory mechanism reduces MT-I expression, allowing augmented NO levels.

Regulation of cell death receptor S-nitrosylation and apoptotic signaling by Sorafenib in hepatoblastoma cells.

Nitric oxide (NO) plays a relevant role during cell death regulation in tumor cells. The overexpression of nitric oxide synthase type III (NOS-3) induces oxidative and nitrosative stress, p53 and cell death receptor expression and apoptosis in hepatoblastoma cells. S-nitrosylation of cell death receptor modulates apoptosis. Sorafenib is the unique recommended molecular-targeted drug for the treatment of patients with advanced hepatocellular carcinoma. The present study was addressed to elucidate the potential role of NO during Sorafenib-induced cell death in HepG2 cells. We determined the intra- and extracellular NO concentration, cell death receptor expression and their S-nitrosylation modifications, and apoptotic signaling in Sorafenib-treated HepG2 cells. The effect of NO donors on above parameters has also been determined. Sorafenib induced apoptosis in HepG2 cells. However, low concentration of the drug (10nM) increased cell death receptor expression, as well as caspase-8 and -9 activation, but without activation of downstream apoptotic markers. In contrast, Sorafenib (10µM) reduced upstream apoptotic parameters but increased caspase-3 activation and DNA fragmentation in HepG2 cells. The shift of cell death signaling pathway was associated with a reduction of S-nitrosylation of cell death receptors in Sorafenib-treated cells. The administration of NO donors increased S-nitrosylation of cell death receptors and overall induction of cell death markers in control and Sorafenib-treated cells. In conclusion, Sorafenib induced alteration of cell death receptor S-nitrosylation status which may have a relevant repercussion on cell death signaling in hepatoblastoma cells.

The JNK binding domain of islet-brain 1 inhibits IL-1 induced JNK activity and apoptosis but not the transcription of key proapoptotic or protective genes in insulin-secreting cell lines.

The stress-activated protein kinase c-Jun NH2-terminal kinase (JNK) is a central signal for interleukin-1beta (IL-1beta)-induced apoptosis in insulin-producing beta-cells. The cell-permeable peptide inhibitor of JNK (JNKI1), that introduces the JNK binding domain (JBD) of the scaffold protein islet-brain 1 (IB1) inside cells, effectively prevents beta-cell death caused by this cytokine. To define the molecular targets of JNK involved in cytokine-induced beta-cell apoptosis we investigated whether JNKI1 or stable expression of JBD affected the expression of selected pro- and anti-apoptotic genes induced in rat (RIN-5AH-T2B) and mouse (betaTC3) insulinoma cells exposed to IL-1beta. Inhibition of JNK significantly reduced phosphorylation of the specific JNK substrate c-Jun (p&lt;0.05), IL-1beta-induced apoptosis (p&lt;0.001), and IL-1beta-mediated c-fos gene expression. However, neither JNKI1 nor JBD did influence IL-1beta-induced NO synthesis or iNOS expression or the transcription of the genes encoding mitochondrial manganese superoxide dismutase (MnSOD), catalase (CAT), glutathione peroxidase (GPx), glutathione-S-transferase rho (GSTrho), heat shock protein (HSP) 70, IL-1beta-converting enzyme (ICE), caspase-3, apoptosis-inducing factor (AIF), Bcl-2 or Bcl-xL. We suggest that the anti-apoptotic effect of JNK inhibition by JBD is independent of the transcription of major pro- and anti-apoptotic genes, but may be exerted at the translational or posttranslational level.

Highly parallel genetic approaches in Mendelian and complex diseases

The recent advance in high-throughput sequencing and genotyping protocols allows rapid investigation of Mendelian and complex diseases on a scale not previously been possible. In my thesis research I took advantage of these modern techniques to study retinitis pigmentosa (RP), a rare inherited disease characterized by progressive loss of photoreceptors and leading to blindness; and hypertension, a common condition affecting 30% of the adult population. Firstly, I compared the performance of different next generation sequencing (NGS) platforms in the sequencing of the RP-linked gene PRPF31. The gene contained a mutation in an intronic repetitive element, which presented difficulties for both classic sequencing methods and NGS. We showed that all NGS platforms are powerful tools to identify rare and common DNA variants, also in case of more complex sequences. Moreover, we evaluated the features of different NGS platforms that are important in re-sequencing projects. The main focus of my thesis was then to investigate the involvement of pre-mRNA splicing factors in autosomal dominant RP (adRP). I screened 5 candidate genes in a large cohort of patients by using long-range PCR as enrichment step, followed by NGS. We tested two different approaches: in one, all target PCRs from all patients were pooled and sequenced as a single DNA library; in the other, PCRs from each patient were separated within the pool by DNA barcodes. The first solution was more cost-effective, while the second one allowed obtaining faster and more accurate results, but overall they both proved to be effective strategies for gene screenings in many samples. We could in fact identify novel missense mutations in the SNRNP200 gene, encoding an essential RNA helicase for splicing catalysis. Interestingly, one of these mutations showed incomplete penetrance in one family with adRP. Thus, we started to study the possible molecular causes underlying phenotypic differences between asymptomatic and affected members of this family. For the study of hypertension, I joined a European consortium to perform genome-wide association studies (GWAS). Thanks to the use of very informative genotyping arrays and of phenotipically well-characterized cohorts, we could identify a novel susceptibility locus for hypertension in the promoter region of the endothelial nitric oxide synthase gene (NOS3). Moreover, we have proven the direct causality of the associated SNP using three different methods: 1) targeted resequencing, 2) luciferase assay, and 3) population study. - Le récent progrès dans le Séquençage à haut Débit et les protocoles de génotypage a permis une plus vaste et rapide étude des maladies mendéliennes et multifactorielles à une échelle encore jamais atteinte. Durant ma thèse de recherche, j'ai utilisé ces nouvelles techniques de séquençage afin d'étudier la retinite pigmentale (RP), une maladie héréditaire rare caractérisée par une perte progressive des photorécepteurs de l'oeil qui entraine la cécité; et l'hypertension, une maladie commune touchant 30% de la population adulte. Tout d'abord, j'ai effectué une comparaison des performances de différentes plateformes de séquençage NGS (Next Generation Sequencing) lors du séquençage de PRPF31, un gène lié à RP. Ce gène contenait une mutation dans un élément répétable intronique, qui présentait des difficultés de séquençage avec la méthode classique et les NGS. Nous avons montré que les plateformes de NGS analysées sont des outils très puissants pour identifier des variations de l'ADN rares ou communes et aussi dans le cas de séquences complexes. De plus, nous avons exploré les caractéristiques des différentes plateformes NGS qui sont importantes dans les projets de re-séquençage. L'objectif principal de ma thèse a été ensuite d'examiner l'effet des facteurs d'épissage de pre-ARNm dans une forme autosomale dominante de RP (adRP). Un screening de 5 gènes candidats issus d'une large cohorte de patients a été effectué en utilisant la long-range PCR comme étape d'enrichissement, suivie par séquençage avec NGS. Nous avons testé deux approches différentes : dans la première, toutes les cibles PCRs de tous les patients ont été regroupées et séquencées comme une bibliothèque d'ADN unique; dans la seconde, les PCRs de chaque patient ont été séparées par code barres d'ADN. La première solution a été la plus économique, tandis que la seconde a permis d'obtenir des résultats plus rapides et précis. Dans l'ensemble, ces deux stratégies se sont démontrées efficaces pour le screening de gènes issus de divers échantillons. Nous avons pu identifier des nouvelles mutations faux-sens dans le gène SNRNP200, une hélicase ayant une fonction essentielle dans l'épissage. Il est intéressant de noter qu'une des ces mutations montre une pénétrance incomplète dans une famille atteinte d'adRP. Ainsi, nous avons commencé une étude sur les causes moléculaires entrainant des différences phénotypiques entre membres affectés et asymptomatiques de cette famille. Lors de l'étude de l'hypertension, j'ai rejoint un consortium européen pour réaliser une étude d'association Pangénomique ou genome-wide association study Grâce à l'utilisation de tableaux de génotypage très informatifs et de cohortes extrêmement bien caractérisées au niveau phénotypique, un nouveau locus lié à l'hypertension a été identifié dans la région promotrice du gène endothélial nitric oxide sinthase (NOS3). Par ailleurs, nous avons prouvé la cause directe du SNP associé au moyen de trois méthodes différentes: i) en reséquençant la cible avec NGS, ii) avec des essais à la luciférase et iii) une étude de population.

Endurance training enhances vasodilation induced by nitric oxide in human skin.

Résumé Il a été démontré que l'exercice physique modifiait le contrôle de la thermorégulation cutané, ce qui se manifeste par une augmentation de la perfusion de la microcirculation de la peau. Pour une même augmentation de température, ce phénomène est plus important chez les sportifs d'endurance que chez les sujets sédentaires. Dans cette étude, nous posons l'hypothèse qu'une composante de cette adaptation peut provenir d'une plus haute capacité des vaisseaux sanguins à répondre à un stimulus vasodilatateur. Pour la tester, nous avons recruté des hommes sains, non fumeurs, soit entraînés (surtout sport d'endurance) ou sédentaires que nous avons partagé en deux classes d'âges (18-35 ans [jeunes] et >50 ans[âgés]). Le flux sanguin cutané était mesuré par un laser-Doppler au niveau de la peau de l'avant-bras. Nous avons alors mesuré la vasodilatation obtenue par les stimuli suivant : Iontophorèse à l'acétylcholine (ACh, un vasodilatateur dépendant de l'endothélium), iontophorèse au nitroprussiate de sodium (SNP, un donneur d'oxyde nitrique) et par libération d'une interruption momentanée du flux artériel huméral (hyperémie réactive). Chez les sujets entraînés, l'effet de l'hyperémie réactive et de l'ACh n'ont pas montré de différence. Par contre, l'augmentation de la perfusion, suivant la iontophorèse de SNP, exprimé en unité de perfusion (PU), était plus importante chez les sujets entraînés que chez les sujets sédentaires (jeunes: 398±54 vs 350±87, p<0.05; âgés: 339±72 vs 307±66, p<0.05). Pour conclure, l'entraînement d'endurance augmente l'effet vasodilatateur de l'oxyde nitrique de la microcirculation cutanée humaine, au moins au niveau de la peau de l'avant-bras. Ces observations ont un intérêt physiologique considérable au vu des résultats d'études récentes qui montrent que le NO sert d'intermédiaire dans la vasodilatation cutanée produite par un stress thermique. Donc, l'augmentation de la bioactivité du NO dans la microcirculation cutanée pourrait être un des mécanismes par lequel l'entraînement physique modifierait le contrôle de la thermorégulation du flux sanguin cutané. Abstract Endurance training modifies the thermoregulatory control of skin blood flow, as manifested by a greater augmentation of skin perfusion for the same increase in core temperature in athletes, in comparison with se-dentary subjects. In this study, we tested the hypothesis that a component of this adaptation might reside in a higher ability of cutaneous blood vessels to respond to vasodilatory stimuli. We recruited healthy nonsmoking males, either endurance trained or sedentary, in two different age ranges (18-35 y and >50 y). Skin blood flow was measured in the forearm skin, using a laser Doppler imager, allowing to record the vasodilatory responses to the following stimuli: iontophoresis of acetylcholine (an endothelium-dependent vasodilator), iontophoresis of sodium nitroprusside (a nitric oxide donor), and release of a temporary interruption of arterial inflow (reactive hyperemia). There was no effect of training on reactive hyperemia or the response to acetylcholine. In contrast, the increase in perfusion following the iontophoresis of sodium nitroprusside, ex-pressed in perfusion units, was larger in trained than in sedentary subjects (younger: 398±54 vs 350±87, p<0.05; older 339±72 vs 307±66, p<0.05). In conclusion, endurance training enhances the vasodilatory effects of nitric oxide in the human dermal microcirculation, at least in forearm skin. These observations have considerable physiologic interest in view of recent data indicating that nitric oxide mediates in part the cutaneous vasodilation induced by heat stress in humans. Therefore, the augmentation of nitric oxide bioactivity in the dermal microcirculation might be one mechanism whereby endurance training modifies the thermoregulatory control of skin blood flow.

A peptide inhibitor of c-Jun N-terminal kinase for the treatment of endotoxin-induced uveitis.

PURPOSE: To evaluate the effect of XG-102 (formerly D-JNKI1), a TAT-coupled dextrogyre peptide that selectively inhibits the c-Jun N-terminal kinase, in the treatment of endotoxin-induced uveitis (EIU). METHODS: EIU was induced in Lewis rats by LPS injection. XG-102 was administered at the time of LPS challenge. The ocular biodistribution of XG-102 was evaluated using immunodetection at 24 hours after either 20 microg/kg IV (IV) or 0.2 microg/injection intravitreous (IVT) administrations in healthy or uveitic eyes. The effect of XG-102 on EIU was evaluated using clinical scoring, infiltration cell quantification, inducible nitric oxide synthase (iNOS) expression and immunohistochemistry, and cytokines and chemokines kinetics at 6, 24, and 48 hours using multiplex analysis on ocular media. Control EIU eyes received vehicle injection IV or IVT. The effect of XG-102 on c-Jun phosphorylation in EIU was evaluated by Western blot in eye tissues. RESULTS: After IVT injection, XG-102 was internalized in epithelial cells from iris/ciliary body and retina and in glial and microglial cells in both healthy and uveitic eyes. After IV injection, XG-102 was concentrated primarily in inflammatory cells of uveitic eyes. Using both routes of administration, XG-102 significantly inhibited clinical signs of EIU, intraocular cell infiltration, and iNOS expression together with reduced phosphorylation of c-Jun. The anti-inflammatory effect of XG-102 was mediated by iNOS, IFN-gamma, IL-2, and IL-13. CONCLUSIONS: This is the first evidence that interfering with the JNK pathway can reduce intraocular inflammation. Local administration of XG-102, a clinically evaluated peptide, may have potential for treating uveitis.