Alkali injury-induced pathological lymphangiogenesis in the iris facilitates the infiltration of T cells and ocular inflammation.

Inflammatory lymphangiogenesis is intimately linked to immune regulation and tissue homeostasis. However, current evidence has suggested that classic lymphatic vessels are physiologically absent in intraocular structures. Here, we show that neolymphatic vessels were induced in the iris after corneal alkali injury (CAI) in a VEGFR3-dependent manner. Cre-loxP-based lineage tracing revealed that these lymphatic endothelial cells (LECs) originate from existing Prox1+ lymphatic vessels. Notably, the ablation of iridial lymphangiogenesis via conditional deletion of VEGFR3 alleviated the ocular inflammatory response and pathological T cell infiltration. Our findings demonstrate that iridial neolymphatics actively participate in pathological immune responses following injury and suggest intraocular lymphangiogenesis as a valuable therapeutic target for the treatment of ocular inflammation.

EE-Explorer: A Multimodal Artificial Intelligence System for Eye Emergency Triage and Primary Diagnosis.

PURPOSE: To develop a multimodal artificial intelligence (AI) system, EE-Explorer, to triage eye emergencies and assist in primary diagnosis using metadata and ocular images. DESIGN: A diagnostic, cross-sectional, validity and reliability study. METHODS: EE-Explorer consists of 2 models. The triage model was developed from metadata (events, symptoms, and medical history) and ocular surface images via smartphones from 2038 patients presenting to Zhongshan Ophthalmic Center (ZOC) to output 3 classifications: urgent, semiurgent, and nonurgent. The primary diagnostic model was developed from the paired metadata and slitlamp images of 2405 patients from ZOC. Both models were externally tested on 103 participants from 4 other hospitals. A pilot test was conducted in Guangzhou to evaluate the hierarchical referral service pattern assisted by EE-Explorer for unspecialized health care facilities. RESULTS: A high overall accuracy, as indicated by an area under the receiver operating characteristic curve (AUC) of 0.982 (95% CI, 0.966-0.998), was obtained using the triage model, which outperformed the triage nurses (P < .001). In the primary diagnostic model, the diagnostic classification accuracy (CA) and Hamming loss (HL) in the internal testing were 0.808 (95% CI 0.776-0.840) and 0.016 (95% CI 0.006-0.026), respectively. In the external testing, model performance was robust for both triage (average AUC, 0.988, 95% CI 0.967-1.000) and primary diagnosis (CA, 0.718, 95% CI 0.644-0.792; and HL, 0.023, 95% CI 0.000-0.048). In the pilot test in the hierarchical referral settings, EE-explorer demonstrated consistently robust performance and broad participant acceptance. CONCLUSION: The EE-Explorer system showed robust performance in both triage and primary diagnosis for ophthalmic emergency patients. EE-Explorer can provide patients with acute ophthalmic symptoms access to remote self-triage and assist in primary diagnosis in unspecialized health care facilities to achieve rapid and effective treatment strategies.

Effect of Tauroursodeoxycholic Acid on Inflammation after Ocular Alkali Burn.

Inflammation is the main cause of corneal and retinal damage in an ocular alkali burn (OAB). The aim of this study was to investigate the effect of tauroursodeoxycholic acid (TUDCA) on ocular inflammation in a mouse model of an OAB. An OAB was induced in C57BL/6j mouse corneas by using 1 M NaOH. TUDCA (400 mg/kg) or PBS was injected intraperitoneally (IP) once a day for 3 days prior to establishing the OAB model. A single injection of Infliximab (6.25 mg/kg) was administered IP immediately after the OAB. The TUDCA suppressed the infiltration of the CD45-positive cells and decreased the mRNA and protein levels of the upregulated TNF-α and IL-1ß in the cornea and retina of the OAB. Furthermore, the TUDCA treatment inhibited the retinal glial activation after an OAB. The TUDCA treatment not only ameliorated CNV and promoted corneal re-epithelization but also attenuated the RGC apoptosis and preserved the retinal structure after the OAB. Finally, the TUDCA reduced the expression of the endoplasmic reticulum (ER) stress molecules, IRE1, GRP78 and CHOP, in the retinal tissues of the OAB mice. The present study demonstrated that the TUDCA inhibits ocular inflammation and protects the cornea and retina from injury in an OAB mouse model. These results provide a potential therapeutic intervention for the treatment of an OAB.

A single-cell transcriptomic atlas of the human ciliary body.

The ciliary body critically contributes to the ocular physiology with multiple responsibilities in the production of aqueous humor, vision accommodation and intraocular immunity. Comparatively little work, however, has revealed the single-cell molecular taxonomy of the human ciliary body required for studying these functionalities. In this study, we report a comprehensive atlas of the cellular and molecular components of human ciliary body as well as their interactions using single-cell RNA sequencing (scRNAseq). Cluster analysis of the transcriptome of 14,563 individual ciliary cells from the eyes of 3 human donors identified 14 distinct cell types, including the ciliary epithelium, smooth muscle, vascular endothelial cell, immune cell and other stromal cell populations. Cell-type discriminative gene markers were also revealed. Unique gene expression patterns essential for ciliary epithelium-mediated aqueous humor inflow and ciliary smooth muscle contractility were identified. Importantly, we discovered the transitional states that probably contribute to the transition of ciliary macrophage into retina microglia and verified no lymphatics in the ciliary body. Moreover, the utilization of CellPhoneDB allowed us to systemically infer cell-cell interactions among diverse ciliary cells including those that potentially participate in the pathogenesis of glaucoma and uveitis. Altogether, these new findings provide insights into the regulation of intraocular pressure, accommodation reflex and immune homeostasis under physiological and pathological conditions.

Alleviating experimental allergic eye disease by inhibiting pro-lymphangiogenic VEGFR3 signal.

PURPOSE: Ocular allergy leads to acute and chronic inflammation that may deteriorate the conjunctiva and other ocular tissue. The conjunctiva is covered with abundant lymphatic vessels but how the conjunctival lymphatic system patriciates in the development of allergic eye disease (AED) remains to be elucidated. METHODS AND RESULTS: By using ovalbumin (OVA)+pertussis toxin (PTX) as a sensitizer followed by daily OVA challenges, we induced optimized AED manifestations in mice. We show that conjunctival lymphatics underwent significant expansion after 28 days of chronic OVA challenge, and this process can be prevented by inducible genetic ablation of lymphatic Vegfr3. Through transcriptomic profile analysis in combination with histopathological examinations, we found that pro-lymphangiogenic VEGFR3 signal promoted allergy-induced activation of T helper 2 (Th2) type immune responses, including antigen presentation, and Th2 cells, B cells and mast cell-related pathways in the conjunctiva, thereby critically contributing to the immunoglobulin E (IgE) production and AED manifestations. As a result, ocular allergy can be alleviated by genetic inhibition of lymphatic Vegfr3. Interestingly, pro-lymphangiogenic VEGFR3 signal did not appear to affect the obstruction of meibomian glands (MGs) or the activation of Th17 type and neutrophil pathways that are associated with AED. CONCLUSIONS: These data reveal the key role of pro-lymphangiogenic VEGFR3 signaling in the development of AED and provide experimental evidence that VEGFR3 inhibition may be useful in treating ocular allergy in patients.

Th17 Activation and Th17/Treg Imbalance in Prolonged Anterior Intraocular Inflammation after Ocular Alkali Burn.

Ocular alkali burn (OAB) is a sight-threatening disease with refractory ocular inflammation causing various blinding complications. Th17 lymphocytes account for the pathogeneses of the autoimmune disease and chronic inflammation, but their role in prolonged anterior intraocular inflammation after OAB is still unknown. A rat OAB model was established for this purpose. Anterior intraocular inflammation was observed in both the acute and late phases of OAB, and histological examination confirmed the presence of inflammatory cell infiltration and fibrin exudation in the anterior segment. Luminex xMAP technology and qPCR were used to evaluate the intraocular levels of cytokines. The levels of IL-1ß, IL-6, and TNF-α were significantly elevated during the acute phase. The expression of IL-17A gradually increased from day 7 onwards and remained at a relatively high level. Immunofluorescence was performed to identify Th17 cells. CD4 and IL-17A double positive cells were detected in the anterior chamber from days 7 to 28. Flow cytometry showed that the frequency of Th17 cells increased in both lymph nodes and spleen, while the frequency of Treg cells remained unchanged, resulting in an elevated Th17/Treg ratio. The present study suggests that Th17 activation and Th17/Treg imbalance account for prolonged anterior intraocular inflammation after OAB.

Inhibition of endoplasmic reticulum stress by 4-phenylbutyrate alleviates retinal inflammation and the apoptosis of retinal ganglion cells after ocular alkali burn in mice.

OBJECTIVE: Retinal ganglion cell (RGC) apoptosis is one of the most severe complications that causes permanent visual impairment following ocular alkali burn (OAB). Currently, very few treatment options exist for this condition. This study was conducted to determine the effect of 4-phenylbutyric acid (4-PBA) on endoplasmic reticulum (ER) stress after OAB using a well-established OAB mouse model. METHODS: Ocular alkali burn was induced in C57BL/6 mouse corneas using 1 M NaOH. 4-PBA (10 mg/kg; 250 µL per injection) or saline (250 µL per injection) was injected intraperitoneally once per day for 3 days before the establishment of the OAB model. The apoptosis of retinal ganglion cells (RGCs) was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay, and the histological damage was examined by hematoxylin and eosin and immunofluorescence assay on retinal flat mounts. The key inflammatory response and the expression of ER stress-related markers in the retinal tissues were assessed by real-time PCR, western blotting and histologic analyses. RESULTS: 4-PBA significantly alleviated the apoptosis of RGCs and prevented the structural damage of the retina, as determined by the evaluation of RGC density and retinal thickness. Inhibition of ER stress by 4-PBA decreased the expression of vital proinflammatory cytokines, tumor necrosis factor alpha, and interleukin-1 beta; and suppressed the activation of retinal microglial cells and nuclear factor-kappa B (NF-κB). 4-PBA reduced the expression of the ER stress molecules, glucose-regulated protein 78, activated transcription factor 6, inositol-requiring enzyme-1 (IRE1), X-box-binding protein 1 splicing, and CCAAT/enhancer-binding protein homologous protein, in the retinal tissues and RGCs of OAB mice. CONCLUSIONS: The present study demonstrated that the inhibition of ER stress by 4-PBA alleviates the inflammatory response via the IRE1/NF-κB signaling pathway and protects the retina and RGCs from injury in an OAB mouse model. Such findings further suggest that 4-PBA might have potential therapeutic implications for OAB treatment.

Clinical Features of Endogenous Endophthalmitis Secondary to Minimally Invasive Upper Urinary Tract Calculus Removal.

PURPOSE: To evaluate endogenous endophthalmitis clinical features following minimally invasive removal of upper urinary tract calculi. METHODS: Medical records of twelve patients (17 eyes) with endogenous endophthalmitis secondary to minimally invasive upper urinary tract calculus removal were retrospectively reviewed. RESULTS: Diabetes mellitus was found in 7 patients (58%). 10 patients (83%) suffered from fever. The stone extraction and ocular symptom onset interval ranged from 2 to 22 days. All eyes presented as vitritis and fluffy yellow-white retinal exudates. Hypopyon was only found in 3 eyes (18%). 5 patients (42%) were misdiagnosed as uveitis which led to mismanagement. Ocular fluids were culture positive for only C. albicans in 12 eyes (71%). 10 of 12 eyes (83%) with silicon oil tamponade obtained a final BCVA≥0.05. CONCLUSIONS: C. albicans was the most common endogenous endophthalmitis pathogen after urinary calculus removal by minimally invasive surgery. Pars plana vitrectomy with silicon oil tamponade may be helpful to achieve a favorable visual outcome. Routine ophthalmologic evaluation by the uveitis or vitreoretinal specialist may be necessary within 2 weeks after the urological procedures.

Prevalence, Risk Factors, and Causes of Visual Impairment in an Elderly Chinese Uygur Population in Southern Xinjiang.

PURPOSE: To investigate the prevalence, risk factors, and major causes of visual impairment (VI) in an elderly Chinese Uygur population in southern Xinjiang. METHODS: This was a population-based cross-sectional study. Participants aged 50 years and older from Haohan Country, Xinjiang Uygur Autonomous Region, were enrolled from August 2018 to December 2018 using cluster sampling. Participants underwent examinations including presenting visual acuity (PVA), pinhole vision, slit-lamp, intraocular pressure, and direct ophthalmoscopy. Participants' education and demographic information was collected by a questionnaire. The prevalence, risk factors, and major causes of vision loss were evaluated. RESULTS: A total of 1465 participants (85.4% response rate) were enrolled. The mean age of the subjects was 59.1 ± 9.7 years. The prevalence of mild VI, moderate VI, severe VI, and blindness in the better eye was 13.3%, 12.8%, 2.9%, and 3.4%, respectively. The prevalence of low vision and blindness in this study was higher than that in Altay & Tacheng and Changji in northern Xinjiang, lower than that in Luxi, and similar to that in Tibet. The multiple logistic regression analysis showed that age, education level, and body mass index (BMI) were significantly associated with low vision and blindness (P ≤ 0.001, <0.05, 0.002, respectively). The major causes of low vision were cataract (42.6%), refractive error (19.6%), and glaucoma (12.6%), whereas the primary causes of blindness were cataract (34%), glaucoma (34%), and retinitis pigmentosa (10%). CONCLUSIONS: VI is an important public health issue among elderly Uygur individuals in the area, especially for those with low education levels. Cataract is the leading cause of low vision and blindness.

αB-Crystallin Alleviates Endotoxin-Induced Retinal Inflammation and Inhibits Microglial Activation and Autophagy.

αB-Crystallin, a member of the small heat shock protein (sHSP) family, plays an immunomodulatory and neuroprotective role by inhibiting microglial activation in several diseases. However, its effect on endotoxin-induced uveitis (EIU) is unclear. Autophagy may be associated with microglial activation, and αB-crystallin is involved in the regulation of autophagy in some cells. The role of αB-crystallin in microglial autophagy is unknown. This study aimed to explore the role of αB-crystallin on retinal microglial autophagy, microglial activation, and neuroinflammation in both cultured BV2 cells and the EIU mouse model. Our results show that αB-crystallin reduced the release of typical proinflammatory cytokines at both the mRNA and protein level, inhibited microglial activation in morphology, and suppressed the expression of autophagy-related molecules and the number of autophagolysosomes in vitro. In the EIU mouse model, αB-crystallin treatment alleviated the release of ocular inflammatory cytokines and the representative signs of inflammation, reduced the apoptosis of ganglion cells, and rescued retinal inflammatory structural and functional damage, as evaluated by optical coherence tomographic and electroretinography. Taken together, these results indicate that αB-crystallin inhibits the activation of microglia and supresses microglial autophagy, ultimately reducing endotoxin-induced neuroinflammation. In conclusion, αB-crystallin provides a novel and promising option for affecting microglial autophagy and alleviating symptoms of ocular inflammatory diseases.

METTL3 attenuates proliferative vitreoretinopathy and epithelial-mesenchymal transition of retinal pigment epithelial cells via wnt/ß-catenin pathway.

Proliferative vitreoretinopathy (PVR) is a refractory vitreoretinal fibrosis disease, and epithelial-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells is the key pathological mechanism of PVR. However, few studies focused on the role of METTL3, the dominating methyltransferase for m6A RNA modification in PVR pathogenesis. Immunofluorescence staining and qRT-PCR were used to determine the expression of METTL3 in human tissues. Lentiviral transfection was used to stably overexpress and knockdown METTL3 in ARPE-19 cells. MTT assay was employed to study the effects of METTL3 on cell proliferation. The impact of METTL3 on the EMT of ARPE-19 cells was assessed by migratory assay, morphological observation and expression of EMT markers. Intravitreal injection of cells overexpressing METTL3 was used to assess the impact of METTL3 on the establishment of the PVR model. We found that METTL3 expression was less in human PVR membranes than in the normal RPE layers. In ARPE-19 cells, total m6A abundance and the METTL3 expression were down-regulated after EMT. Additionally, METTL3 overexpression inhibited cell proliferation through inducing cell cycle arrest at G0/G1 phase. Furthermore, METTL3 overexpression weakened the capacity of TGFß1 to trigger EMT by regulating wnt/ß -catenin pathway. Oppositely, knockdown of METTL3 facilitated proliferation and EMT of ARPE-19 cells. In vivo, intravitreal injection of METTL3-overexpressing cells delayed the development of PVR compared with injection of control cells. In summary, this study suggested that METTL3 is involved in the PVR process, and METTL3 overexpression inhibits the EMT of ARPE-19 cells in vitro and suppresses the PVR process in vivo.

Changing Trends in Firework-Related Eye Injuries in Southern China: A 5-Year Retrospective Study of 468 Cases.

Firework-related eye injury is a horrible medical problem and creates huge health and social burdens. Herein, we explored the changing trends and demographic and clinical features of firework-related eye injury, in an effort to inform strategies to prevent this injury. We reviewed the data of 468 hospitalized patients who underwent surgery for firework-related eye injury at the Zhongshan Ophthalmic Center between January 2013 and December 2017. During this period, the trend in firework-related eye injuries was stable (mean, 93 ± 14 cases per year), and fireworks were the major cause of explosive eye injury. The average age of the patients was 24 ± 18 years and 87% of the patients were male, with boys under 10 years of age comprising the largest group (27% of patients). There were an average of 24 ± 7 cases per year from urban areas and 70 ± 8 cases from rural areas (P < 0.05). Furthermore, 21 ± 5% of cases occurred during Spring Festival. After treatment, the best corrected visual acuity was increased compared to that before treatment, and the intraocular pressure tended to become normal by the final visit (P < 0.05). The top three diagnoses were cataract (39%), retinal detachment (18%), and choroidal detachment (14%). Additionally, the most common surgery was cataract extraction (25%), followed by pars plana vitrectomy (19%) and pars plana vitrectomy plus pars plana lensectomy (10%). Over the five-year study period, day surgery hospitalization increased from 1% to 32%. This was associated with a corresponding decrease in the length of hospitalization, without adverse events, demonstrating that day surgery is feasible in firework-related eye injury cases. The present study results suggest that greater attention should be paid to firework-related eye injury, and a variety of measures should be taken to prevent this kind of ocular tragedy.

Intraocular foreign body injury in children: clinical characteristics and factors associated with endophthalmitis.

BACKGROUND/AIMS: To analyse the clinical characteristics of intraocular foreign body (IOFB) injury in children and evaluate factors associated with endophthalmitis development. METHODS: Patients aged <18 years with IOFB attending Zhongshan Ophthalmic Center between January 2003 and July 2016 were included retrospectively. Demographic features, clinical characteristics and factors associated with endophthalmitis development were analysed. RESULTS: Consecutive subjects (n=484, 484 eyes) were included: mean age 10.12±4.54 years, 86.4% male. Fireworks (28.5%) were the most common cause of injury. Injury of uncertain cause (17.5%), fireworks (36.7%) and metal splatter (31.7%) accounted for most injuries in the 0-3, 4-12 and 13-17 years age groups, respectively. Plant branches (16.7%) and fireworks (31.1%) were the most common causes of injuries in females and males, respectively. Endophthalmitis occurred in 116 patients (24.0%), with metallic IOFB (OR=0.338, p=0.001), intraocular haemorrhage (OR=0.100, p<0.001) and uveal tissue prolapse (OR=0.206, p<0.001) conferring lower risk, while zone II wound (OR=4.336, p<0.001) and traumatic lens rupture (OR=2.567, p=0.028) were associated with higher risk. CONCLUSION: Clinical characteristics of children with IOFB injury differ from those of adults. Fireworks are the most common cause of injury, indicating a ban on children igniting fireworks should be advocated. Safety education and protective measures should vary by age and sex. Endophthalmitis is associated with zone II wound, traumatic lens rupture, IOFB material, intraocular haemorrhage and uveal tissue prolapse, which has clinical relevance.

Risk Factors Associated with Failure of Direct Cyclopexy in the Treatment of Small to Moderate Tramatic Cyclodialysis Clefts.

BACKGROUND: To evaluate the risk factors associated with failure to correct hypotony using direct cyclopexy in patients with traumatic cyclodialysis cleft. METHODS: In a series of 116 patients with traumatic cyclodialysis who underwent direct cyclopexy at Zhongshan Ophthalmic Center from January 2008 to August 2018, the clinical correlation between the risk factors and failure of the operation were retrospectively studied, after adjusting for other potential confounders. RESULTS: The curative ratio after one procedure was 82.76%, whereas 20 (17.24%) eyes experienced treatment failure after the first surgery. The degree of anterior chamber angle closure was significantly wider in patients with a failed first surgery than in patients for whom one procedure was a success (p = .046). The risk of failure to achieve closure increased as the angle-closure exceeded 5 clock hour (odds ratio, 10.39; 95% confidence interval, 1.75-61.72; p = .010). An analysis of the recurrent position indicated that an angle closure exceeding 5 clock hour may impede accurate cleft location and is thus associated with an increased risk of failure to correct hypotony. CONCLUSION: Exceeding the threshold of 5 clock hour in anterior chamber angle closure may impede accurate cleft location and, thus, present a higher risk of failure to correct hypotony using direct cyclopexy. These patients may need injection of a viscoelastic agent into the anterior chamber by paracentesis to deepen the anterior chamber and to delineate the clefts using gonioscopy pre- or intraoperatively.

Development of a rabbit model of persistent hypotony without ciliary body injury.

In order to study the pathophysiological alterations of the ciliary body (CB) during persistent hypotony, it is necessary to develop an animal model without CB injury. In this study, we successfully established a modified model of persistent hypotony without CB injury in New Zealand rabbits. A 23-gauge pars plana vitrectomy (PPV) was performed and a trocar-formed fistula was allowed to remain in situ, to produce a continuous outflow of intraocular fluid. Both eyes underwent PPV with normal intraocular pressure (IOP); eyes with no surgical intervention were used as controls. The IOP was monitored and used to evaluate the reliability of the model. Secondary changes of hypotony were evaluated by slit-lamp biomicroscopy and B scans while morphological changes of the CB were observed by haematoxylin and eosin staining. The mean IOP in the hypotony groups were consistently lower than 6 mmHg. Furthermore, there were no significant differences in IOP between the PPV control group and normal eyes. Collectively, our data indicate that this model successfully simulates the secondary changes of hypotony, including a reduction in corneal size, corneal oedema, anterior chamber inflammation, morphological alterations of the CB, cataract, retinal detachment, and choroidal detachment. The morphological structure of the CB tissue changed dramatically after persistent hypotony, indicating that normal IOP may be required in order to maintain normal function in the CB. This model of persistent hypotony potentially represents a valuable tool for future studies aiming to investigate the pathophysiological mechanisms underlying CB dysfunction and other secondary changes that occur during hypotony.

Effect of vitrectomy with intrasurgical regulation of intraocular pressure in a rabbit model of central retinal artery occlusion.

Central retinal artery occlusion (CRAO) is an ophthalmic emergency that causes severe and permanent visual impairment. The effects of conventional treatments on recanalizing retinal arteries and improving visual outcome are equivocal. This study was designed to determine the possible benefits of pars plana vitrectomy (PPV) with intrasurgical regulation of intraocular pressure using intraocular vascular counterpulsation (IVT). CRAO was induced by 532-nm argon green laser activation of auricular intravenous injected rose bengal, a photosensitive dye, in the central retinal arteries (CRA) of eighty-four New Zealand white albino rabbits. CRAO rabbits were randomly assigned to photocoagulation, vitrectomy and counterpulsation groups. Depending on the time intervals between surgery and CRAO induction, vitrectomy and counterpulsation groups were further divided into 2 h (2h), 6 h (6h) and 24 h (24h) subgroups. The proportion of eyes with complete recanalization was significantly higher in the 2h counterpulsation subgroup after three days (P = 0.032) and in all counterpulsation subgroups after one week (P = 0.020). After one month, the 2h and 6h counterpulsation subgroups showed greater oscillatory potential (OPs) responses (F = 3.519, P = 0.049). The 2h counterpulsation subgroup also exhibited greater b-wave amplitude in photopic 3.0 Flicker(F = 4.530, P = 0.044). Histologic evaluation revealed less destruction in the inner retina for the 2h and 6h counterpulsation subgroups. Expression of HSP70 was higher in the 2h and 6h counterpulsation subgroups (F = 48.915,P < 0.001). Levels of HSP90 were lower in all counterpulsation subgroups (F = 30.065,P < 0.001). Levels of TNF-α were lower in the 2h counterpulsation subgroup (F = 14.762,P < 0.001). These results indicate that PPV with IVT was effective to recanalize retinal arteries after CRAO. Early intervention provided better morphologic and functional prognosis for inner retina. The protective effect was related with higher retinal levels of HSP70 and lower levels of HSP90 and TNF-α.

The anti-inflammatory effect of minocycline on endotoxin-induced uveitis and retinal inflammation in rats.

Purpose: Uveitis is a serious inflammatory disease of the uvea, frequently leading to visual impairment and irreversible blindness. Here, we investigated the anti-inflammatory effect of minocycline on rat endotoxin-induced uveitis (EIU) and retinal inflammation. Methods: For in vivo studies, the rat EIU model was induced with intravitreal injection of lipopolysaccharide (LPS). Minocycline was administered intraperitoneally 2 h before and after the LPS injection. The severity of the ocular inflammation was evaluated with slit-lamp photography, aqueous humor cell counting, protein quantitative determination, and histological analysis. Retinal microglia were labeled with a fluorescent dye 4Di-10ASP. Microglial activity and inflammatory cytokine production were analyzed with immunofluorescence and real-time PCR. For the in vitro studies, BV-2 microglia cells were stimulated with LPS or cotreated with minocycline for 6 h. Toll-like receptor (TLR) 2/4 levels were determined with real-time PCR and western blotting. Results: The LPS-challenged eyes displayed severe inflammation in all ocular structures, including a large number of anterior chamber cells, fibrin exudation, hypopyon, and infiltrated inflammatory cells in the vitreous and retina. Immunostaining of the retinal whole-mounts also revealed numerous retinal microglia were activated promptly, and then more and more peripheral leukocytes were recruited and infiltrated in the LPS-injected retinas. Additionally, the production of tumor necrosis factor-α (TNF-α), chemokine (C-C motif) ligand 2 (CCL-2), interleukin-1 beta (IL-1ß), and IL-6 was dramatically increased. However, minocycline treatment strongly inhibited microglia activation, decreased inflammatory cytokine production, prevented peripheral inflammatory cell recruitment, and significantly attenuated ocular inflammation. Finally, we demonstrated the mechanism of the microglia inactivation effect of minocycline is via suppression of TLR4 signaling. Conclusions: This study indicates minocycline is far beyond an antibiotic. It not only attenuates rat EIU but also inhibits retinal inflammation through inactivating microglia, inhibiting inflammatory cell recruitment and inflammatory cytokine production.

Nine-year analysis of isolated pathogens and antibiotic susceptibilities of microbial keratitis from a large referral eye center in southern China.

Purpose: To analyze the genus profile of isolated pathogens and antibiotic susceptibility trends of microbial keratitis over nine years at a large referral eye center in southern China. Methods: Data of corneal specimens from January 2010 to August 2018 of patients clinically diagnosed with infectious keratitis were obtained from the center's microbiology database. Results with positive cultures along with antibiotic susceptibility were reviewed and analyzed. Results: We collected and reviewed 7,229 specimens, including 3,092 with positive cultures. Among them, 1,630 (52.72%) were bacterial, 1781 (57.60%) were fungal, and 319 (10.32%) were coinfected. A significant decreasing trend was observed in the isolates of Gram-positive cocci (r =-0.711, P=0.032), among which the proportion of coagulase-negative staphylococcus (CNS) was also reduced (r =-0.883, P=0.002). In contrast, an increasing trend in the proportion of Gram-negative bacilli was observed (r=0.661, P=0.053). The susceptibility rates of Gram-positive cocci to cephalosporins were near 90%, which was relatively high compared to fluoroquinolones. Fluoroquinolones represented the antibiotics to which Gram-negative bacilli were the most susceptible. Their susceptibility to moxifloxacin was 78.79%. The overall performance of aminoglycosides and vancomycin was both around 70%. The susceptibility of Gram-positive cocci to several antibiotics including levofloxacin (r=-0.717, P=0.03), tobramycin (r= -0.933, P<0.001), cefazolin (r= -0.964, P<0.001), ceftazidime (r=-0.929, P=0.003), chloramphenicol (r=-0.929, P=0.003), and cefuroxime (r=-0.829, P=0.042) decreased over time. The susceptibility of Gram-negative bacilli to ofloxacin increased over time (r=0.854, P=0.004), whereas that to cefazolin (r=-0.833, P=0.005) and chloramphenicol (r=-0.886, P=0.019) decreased over time. Conclusion: From 2010 to 2018 in Zhongshan Ophthalmic Center, most isolates from infectious keratitis were Gram-positive cocci (mainly CNS), which decreased over time, with an increase in Gram-positive bacilli. More than half of the antibiotics showed reducing trend of susceptibilities, and the antibiotic resistance situation in southern China was not encouraging.

Inhibition of Notch1 Signaling Alleviates Endotoxin-Induced Inflammation Through Modulating Retinal Microglia Polarization.

Microglial cells are resident immune cells and play an important role in various cerebral and retinal inflammatory diseases. Notch1 signaling is involved in the microglia polarization and the control of cerebral inflammatory reactions. However, its role in endotoxin-induced uveitis (EIU) remains unknown. This study aimed to investigate the role of Notch1 signaling on retinal microglia polarization and inflammation in the cultured retinal microglial cells and EIU rat model. We found that Notch1 signaling blockade with N-[N-(3, 5-difluorophenacetyl)-1-alany1-S-phenyglycine t-butyl ester (DAPT) shifted retinal microglia phenotype from pro-inflammatory M1 phenotype (COX2+ and iNOS+) to anti-inflammatory M2 phenotype (Arg-1+) and reduced the release of pro-inflammatory cytokines both in vivo and in vitro. Moreover, DAPT treatment contributed to prevent retinal ganglion cells from apoptosis, reduce the intraocular infiltrating cells, and attenuate the impairment of retinal function. Taken together, these results suggest that inhibition of Notch1 signaling could alleviate the inflammatory response in EIU rat mainly through regulating the polarization of retinal microglia. Therefore, Notch1 signaling might be a promising therapeutic target in the treatment of ocular inflammatory diseases.