Search Results (14)

Retinal Müller glial cells (RMG) play a crucial role in retinal neuroinflammation, including autoimmune uveitis. Increasing evidence supports their function as active modulators of immune responses and potential atypical antigen-presenting cells (APCs). To further investigate this hypothesis, we conducted a differential proteome analysis of primary equine RMG from healthy controls and horses with equine recurrent uveitis (ERU), a spontaneous model of autoimmune uveitis. This analysis identified 310 proteins with differential abundance. Among these, the Major Histocompatibility Complex (MHC) class II and the enzyme Arginase 1 (ARG1) were significantly enriched in RMG from uveitis-affected horses, whereas Mannose Receptor C-type 2 (MRC2) and its interactor Thrombospondin 1 (THBS1) were more abundant in healthy RMG. The detection of MHC class II in equine RMG, consistent with previous studies, validates the robustness of our approach. Furthermore, the identification of ARG1 and MRC2, together with THBS1, provides new insights into the immunomodulatory and antigen-presenting properties of RMG. Immunohistochemical analyses confirmed the proteomic findings and revealed the spatial distribution of ARG1 and MRC2. ARG1 and MRC2 are thus markers for RMG in the neuroinflammatory or physiological milieu and highlight potential differences in the immune function of RMG, particularly in antigen presentation. Full article

Equine recurrent uveitis (ERU) is a spontaneously occurring autoimmune disease and one of the leading causes of blindness in horses worldwide. Its similarities to autoimmune-mediated uveitis in humans make it a unique spontaneous animal model for this disease. Although many aspects of ERU pathogenesis have been elucidated, it remains not fully understood and requires further research. CD4⁺ T cells have been a particular focus of research. In a previous study, we showed metabolic alterations in CD4⁺ T cells from ERU cases, including an increased basal oxygen consumption rate (OCR) and elevated compensatory glycolysis. To further investigate the underlying reasons for and consequences of these metabolic changes, we quantified reactive oxygen species (ROS) production in CD4⁺ T cells from ERU cases and compared it to healthy controls, revealing significantly higher ROS production in ERU-affected horses. Additionally, we aimed to define mitochondrial fuel oxidation of glucose, glutamine, and long-chain fatty acids (LCFAs) and identified significant differences between CD4⁺ T cells from ERU cases and controls. CD4⁺ T cells from ERU cases showed a lower dependency on mitochondrial glucose oxidation and greater metabolic flexibility for the mitochondrial oxidation of glucose and LCFAs, indicating an enhanced ability to switch to alternative fuels when necessary. Full article

Leptospirosis is one of the most common zoonotic infections and a major problem in terms of both veterinary medicine and public health. However, the disease is under-recognised and under-diagnosed worldwide, particularly in horses. Clinical leptospirosis in horses is mainly associated with recurrent uveitis (ERU), which has recently been studied more intensively, and reproductive disorders, the epidemiology of which is still relatively poorly understood. To enhance our comprehension of abortions caused by leptospirosis in horses and to identify the causative strains, a serological study was carried out with subsequent molecular characterisation of the isolate obtained. Using the microscopic agglutination test (MAT), serum samples from mares that aborted and foetal fluids (when available) were tested for antibodies against Leptospira spp. Furthermore, bacteria isolation from kidney cultures was conducted. Of 97 mare serum samples, 21 (21.64%) tested positive, with Grippotyphosa and Pomona being the most frequently detected serogroups. A significantly higher seroprevalence was found in aborting mares compared to the healthy horse population from the same geographical area, as well as a pronounced seasonal variation. Leptospiral antibodies were not detected in any of the foetal fluids, but isolation was successful in 1 case out of 39 (2.56%). Genotyping by multilocus sequence typing (MLST) and core genome multilocus sequence typing (cgMLST) identified the obtained isolate as Leptospira kirschneri, serogroup Pomona, serovar Mozdok. Further surveillance and molecular typing of Leptospira strains causing abortion in horses would be invaluable in understanding the prevalence and impact of leptospirosis on equine reproductive health in Europe. Full article

Background: Cataracts resulting from equine recurrent uveitis (ERU) or other forms of uveitis are usually associated with rapid progression. ERU is the most common ocular disease cause of blindness and cause of cataracts in horses. The necessity for the posterior capsulorhexis (PC) during phacoemulsification (PE) is controversial. This study aimed to evaluate vision and complications after PE combined with pars plana vitrectomy (PPV) in horses with uveitis-associated cataracts and compare the PE technique with and without posterior capsulorhexis. Methods: Thirty-two eyes of 28 horses with uveitis-associated cataracts aged 14 months to 19.6 years were treated with PE-PPV under identical conditions. Twenty-three eyes of 21 horses were affected by an ERU-associated (ERU group), and nine eyes of 7 horses were affected by cataracts related to uveitis with pathogenesis different to ERU (non-ERU group). PE-PPV was performed in 12 eyes of 10 horses (PC group) and 20 eyes of 18 horses without posterior capsulorhexis (NPC group). Follow-up examination was performed at a mean of 1.7 ± 1.8 years postoperatively (range: 1 month–6.4 years). Results: In the period up to 1 month postoperatively, 17/20 (85%) NPC-eyes and 8/12 (67%) PC-eyes (total: 25/32 [78%]) were visual. From 1–6 months postoperatively, 16/20 (80%) NPC-eyes and 7/12 (58.3%) PC-eyes (total: 23/32 [72%]), and from 6–12 months, 7/11 (63.6%) NPC-eyes and 3/8 (37.5%) PC-eyes (total: 10/19 [52.6%]) were visual. From 12–18 months postoperatively, 3/7 (42.9%) NPC-eyes and 2/9 (22.2%) PC-eyes (total: 5/16 [31.3%]), and from 18–24 months, 3/8 (37.5%) NPC-eyes and 1/8 (12.5%) PC-eyes (total: 4/16 [25%]) were visual. After 24 months postoperatively, 2/7 (28.6%) NPC-eyes and 1/8 (12.5%) PC-eyes (total: 3/15 [20%]) were visual. Despite the higher number of visual eyes in the NPC group at each time point, differences were not significant. No obvious differences regarding postsurgical vision were observed between the ERU- and non-ERU groups at each time point. In the overall population, a significant decrease in the number of eyes with postoperative active uveitis was observed during the follow-up examinations (p < 0.001). A significant increase in the number of eyes that were blind due to retinal detachment was observed in the overall patient population as the examination period progressed (p < 0.001). Retinal detachment was the sole long-term cause of blindness. Conclusions: In horses diagnosed with uveitis-associated cataracts and treated with PE-PPV, no persistent active uveitis was observed in the present study during follow-up examinations. However, the proportion of eyes that were blind due to retinal detachment increased. Whilst PE-PPV may prevent postsurgical persistent active uveitis and remove lens opacity, the prognosis for a visual outcome is guarded. A superior outcome in postsurgical vision was observed in the NPC group. However, caution is required when interpreting these results due to several factors that affect the independent comparison of the surgical groups. Full article

The perception of circulating granulocytes as cells with a predetermined immune response mainly triggered by pathogens is evolving, recognizing their functional heterogeneity and adaptability, particularly within the neutrophil subset. The involvement of these cells in the pathophysiology of autoimmune uveitis has become increasingly clear, yet their exact role remains elusive. We used an equine model for autoimmune-mediated recurrent pan-uveitis to investigate early responses of granulocytes in different inflammatory environments. For this purpose, we performed differential proteomics on granulocytes from healthy and diseased horses stimulated with IL8, LPS, or PMA. Compared to healthy horses, granulocytes from the recurrent uveitis model significantly changed the cellular abundance of 384 proteins, with a considerable number of specific changes for each stimulant. To gain more insight into the functional impact of these stimulant-specific proteome changes in ERU pathogenesis, we used Ingenuity Pathway Analysis for pathway enrichment. This resulted in specific reaction patterns for each stimulant, with IL8 predominantly promoting Class I MHC-mediated antigen processing and presentation, LPS enhancing processes in phospholipid biosynthesis, and PMA, clearly inducing neutrophil degranulation. These findings shed light on the remarkably differentiated responses of neutrophils, offering valuable insights into their functional heterogeneity in a T-cell-driven disease. Raw data are available via ProteomeXchange with identifier PXD013648. Full article

In the pathophysiology of autoimmune-mediated uveitis, granulocytes have emerged as possible disease mediators and were shown to be pre-activated in equine recurrent uveitis (ERU), a spontaneous disease model. We therefore used granulocytes from ERU horses to identify early molecular mechanisms involved in this dysregulated innate immune response. Primary granulocytes from healthy and ERU horses were stimulated with IL8, and cellular response was analyzed with differential proteomics, which revealed significant differences in protein abundance of 170 proteins in ERU. Subsequent ingenuity pathway analysis identified three activated canonical pathways “PKA signaling”, “PTEN signaling” and “leukocyte extravasation”. Clustered to the leukocyte extravasation pathway, we found the membrane-type GPI-anchored protease MMP25, which was increased in IL8 stimulated ERU granulocytes. These findings point to MMP25 as a possible regulator of granulocyte extravasation in uveitis and a role of this molecule in the impaired integrity of the blood-retina-barrier. In conclusion, our analyses show a clearly divergent reaction profile of pre-activated granulocytes upon IL8 stimulation and provide basic information for further in-depth studies on early granulocyte activation in non-infectious ocular diseases. This may be of interest for the development of new approaches in uveitis diagnostics and therapy. Raw data are available via ProteomeXchange with identifier PXD013648. Full article

In the equine clinic of the LMU in Munich, therapeutic vitrectomies have been routinely performed in horses for three decades. The vitreous samples obtained during vitrectomies were usually tested for anti-Leptospira antibodies and for more than 20 years also by PCR for leptospiral DNA. If the indication for surgery was ophthalmologically inconclusive, an aqueous humor was collected preoperatively and examined for evidence of leptospiral infection. In this study, medical records from 2002 to 2017 were analyzed. Records for 1387 eyes affected by equine recurrent uveitis (ERU) and 237 eyes affected by another type of uveitis met the inclusion criteria. A total of 216 samples from healthy eyes were used as controls. In 83% of intraocular samples from ERU eyes, antibody titers of 1:100 or higher were detectable by microscopic agglutination test (MAT). Similarly, 83% of intraocular samples had anti-Leptospira antibodies detected by ELISA. In 72% of the intraocular specimens, leptospiral DNA was detectable by PCR. No antibodies were detectable in the samples from eyes with another type of uveitis or in the samples from healthy eyes. A PCR was positive in only one sample from a healthy eye. These results with a very high number of intraocular specimens demonstrate the great importance of an intraocular leptospiral infection for ERU. It can be concluded that for a reliable diagnosis of intraocular leptospiral infection or to reliably exclude an infection multiple tests should be applied. Full article

Uveitis is a sight-threatening eye disease in equids known worldwide that leads to considerable pain and suffering. By far the most common type of uveitis in Germany and neighboring countries is classical equine recurrent uveitis (ERU), which is caused by chronic intraocular leptospiral infection and is the main cause of infectious uveitis in horses. Other infectious causes are extremely rare and are usually clinically distinguishable from ERU. ERU can be treated very effectively by vitreous cavity lavage (vitrectomy). For proper indications of this demanding surgery, it is necessary to differentiate ERU from other types of uveitis in which vitrectomy is not helpful. This can be conducted on the basis of anamnesis in combination with ophthalmologic findings and by aqueous humor examination. During vitrectomy, vitreous material is obtained. These vitreous samples have historically been used for numerous etiologic studies. In this way, a chronic intraocular leptospiral infection has been shown to be the cause of typical ERU and, among other findings, ERU has also been recognized as a biofilm infection, providing new insights into the pathogenesis of ERU and explaining some thus far unexplainable phenomena of ERU. ERU may not only have transmissible aspects to some types of uveitis in humans but may also serve as a model for a spontaneously occurring biofilm infection. Vitreous material obtained during therapeutically indicated vitrectomy can be used for further studies on in vivo biofilm formation, biofilm composition and possible therapeutic approaches. Full article

Equine recurrent uveitis (ERU) is typically caused by chronic intraocular leptospiral infection in warm-blooded horses in central Europe. The most effective therapy for leptospiral-induced ERU is the surgical removal of diseased vitreous (vitrectomy). Since vitrectomy is a highly specialized and invasive surgery, the indication must be determined very carefully. In order to obtain evidence of intraocular leptospiral infection by laboratory diagnostics in questionable leptospiral ERU-cases, sampling of aqueous humor is required, because serum tests using microscopic agglutination test (MAT) are too unspecific. The SNAP Lepto is a cross-species rapid test for the detection of anti-Lipl32 antibodies that has a high sensitivity (0.97) and specificity (1.00) for the detection of anti-leptospiral antibodies using aqueous humor or vitreous samples, which is comparable to MAT. To evaluate sensitivity and specificity of SNAP Lepto using serum, serum samples from 90 horses with confirmed leptospiral ERU and from 103 ocularly healthy horses were tested by both MAT and SNAP Lepto. Sensitivity was similar for both tests (0.82 vs. 0.79), but specificity was lower for MAT (0.52 vs. 0.95). Sensitivity and specificity are therefore lower in serum samples compared to intraocular samples, however, the SNAP Lepto is far superior to MAT and suitable as a screening method using equine serum. Full article

Equine recurrent uveitis (ERU) causes painful inflammatory attacks and oftentimes blindness in the affected eyes. The disease is considered a late sequela of systemic leptospirosis. The most effective therapy is the surgical removal of the vitreous (vitrectomy), which is not only therapeutic, but provides vitreous material that can be assessed diagnostically. For example, the lipL32 gene, culturable Leptospira spp., and anti-Leptospira antibodies have all been detected in vitreous samples obtained from eyes with chronic ERU. Despite this clear evidence of leptospiral involvement, the systemic administration of antibiotics in infected horses is ineffective at resolving ERU. This syndrome of chronic recurrent inflammation, which is unresponsive to antibiotic therapy, combined with apparent bacteria evading the immune response, is consistent with a biofilm-associated infection. The purpose of this study, therefore, was to detect the in vivo biofilm formation of Leptospira spp. in vitreous samples collected during vitrectomy and examined using a Warthin-Starry silver stain and immunohistochemistry. All known steps of biofilm formation were visualized in these samples, including individual Leptospira spp., leptospiral microcolonies and dense roundish accumulations of Leptospira spp. In many instances spirochetes were surrounded by an extracellular substance. Taken together, data from the present study show that ERU is a biofilm-associated intraocular leptospiral infection, which best explains the typical clinical course. Full article

Leptospirosis has been reported in both humans and animals in Israel but has not been reported in horses. In 2018, an outbreak of Leptospira spp. serogroup Pomona was reported in humans and cattle in Israel. In horses, leptospirosis may cause equine recurrent uveitis (ERU). This report describes the first identification of Leptospira serogroup Pomona as the probable cause of ERU in horses in Israel, followed by an epidemiological investigation of equine exposure in the area. Serologic exposure to Leptospira was determined by microscopic agglutination test (MAT) using eight serovars. In 2017, serovar Pomona was identified in a mare with signs of ERU. Seven of thirteen horses from that farm were seropositive for serogroup Pomona, of which three had signs of ERU. During the same time period, 14/70 horses from three other farms were positive for serogroup Pomona. In 2015, two years prior to this diagnosis, 259 horses from 21 farms were sampled and one horse tested seropositive for serovar Icterohaemorrhagiae. In 2018, one year later, 337 horses were sampled on 29 farms, with none testing seropositive. Although horses are not considered a major host of Leptospira spp., it appears that horses may be infected, and clinically affected, in the course of an outbreak in other species. The identification of leptospirosis in stabled horses may impose a significant zoonotic risk to people. Full article

The objective of this study was to describe and compare the fecal bacterial microbiota of horses with equine recurrent uveitis (ERU) and healthy horses using next-generation sequencing techniques. Fecal samples were collected from 15 client-owned horses previously diagnosed with ERU on complete ophthalmic examination. For each fecal sample obtained from a horse with ERU, a sample was collected from an environmentally matched healthy control with no evidence of ocular disease. The Illumina MiSeq sequencer was used for high-throughput sequencing of the V4 region of the 16S rRNA gene. The relative abundance of predominant taxa, and alpha and beta diversity indices were calculated and compared between groups. The phyla Firmicutes, Bacteroidetes, Verrucomicrobia, and Proteobacteria predominated in both ERU and control horses, accounting for greater than 60% of sequences. Based on linear discriminant analysis effect size (LEfSe), no taxa were found to be enriched in either group. No significant differences were observed in alpha and beta diversity indices between groups (p > 0.05 for all tests). Equine recurrent uveitis is not associated with alteration of the gastrointestinal bacterial microbiota when compared with healthy controls. Full article

Equine recurrent uveitis (ERU) is considered one of the most important eye diseases in horses and typically appears with relapsing inflammatory episodes without systemic effects. Various disorders have been described as an initial trigger, including infections. Independent of the initiating cause, there are numerous indications that ERU is an immune-mediated disease. We investigated whether neutrophil extracellular traps (NETs) are part of the ERU pathogenesis. Therefore, vitreous body fluids (VBF), sera, and histological sections of the eye from ERU-diseased horses were analyzed for the presence of NET markers and compared with horses with healthy eyes. In addition, NET formation by blood derived neutrophils was investigated in the presence of VBF derived from horses with healthy eyes versus ERU-diseased horses using immunofluorescence microscopy. Interestingly, NET markers like free DNA, histone-complexes, and myeloperoxidase were detected in higher amounts in samples from ERU-diseased horses. Furthermore, in vitro NET formation was higher in neutrophils incubated with VBF from diseased horses compared with those animals with healthy eyes. Finally, we characterized the ability of equine cathelicidins to induce NETs, as potential NET inducing factors in ERU-diseased horses. In summary, our findings lead to the hypothesis that ERU-diseased horses develop more NETs and that these may contribute to the pathogenesis of ERU. Full article

Retinal pigment epithelium (RPE) builds the outer blood-retinal barrier of the eye. Since one typical feature of the autoimmune disease, equine recurrent uveitis (ERU), is the breakdown of this barrier, we recently performed comparative analysis of healthy and uveitic RPE. We identified for the first time peripherin 2, which is responsible for visual perception and retina development, to be localized in RPE. The purpose of this study was therefore to validate our findings by characterizing the expression patterns of peripherin 2 in RPE and retina. We also investigated whether peripherin 2 expression changes in ERU and if it is expressed by the RPE itself. Via immunohistochemistry, significant downregulation of peripherin 2 in uveitic RPE compared to the control was detectable, but there was no difference in healthy and uveitic retina. A further interesting finding was the clear distinction between peripherin 2 and the phagocytosis marker, rhodopsin, in healthy RPE. In conclusion, changes in the expression pattern of peripherin 2 selectively affect RPE, but not retina, in ERU. Moreover, peripherin 2 is clearly detectable in healthy RPE due to both phagocytosis and the expression by the RPE cells themselves. Our novel findings are very promising for better understanding the molecular mechanisms taking place on RPE in uveitis. Full article