Prevalence and clinical impact of alcohol withdrawal syndrome in alcohol-associated hepatitis and the potential role of prophylaxis: a multinational, retrospective cohort study.

Background: The prevalence and impact of alcohol withdrawal syndrome (AWS) in patients with alcohol-associated hepatitis (AH) are unknown. In this study, we aimed to investigate the prevalence, predictors, management, and clinical impact of AWS in patients hospitalized with AH. Methods: A multinational, retrospective cohort study enrolling patients hospitalized with AH at 5 medical centres in Spain and in the USA was performed between January 1st, 2016 to January 31st, 2021. Data were retrospectively retrieved from electronic health records. Diagnosis of AWS was based on clinical criteria and use of sedatives to control AWS symptoms. The primary outcome was mortality. Multivariable models controlling for demographic variables and disease severity were performed to determine predictors of AWS (adjusted odds ratio [OR]) and the impact of AWS condition and management on clinical outcomes (adjusted hazard ratio [HR]). Findings: In total, 432 patients were included. The median MELD score at admission was 21.9 (18.3-27.3). The overall prevalence of AWS was 32%. Lower platelet levels (OR = 1.61, 95% CI 1.05-2.48) and previous history of AWS (OR = 2.09, 95% CI 1.31-3.33) were associated with a higher rate of incident AWS, whereas the use of prophylaxis decreased the risk (OR = 0.58, 95% CI 0.36-0.93). The use of intravenous benzodiazepines (HR = 2.18, 95% CI 1.02-4.64) and phenobarbital (HR = 2.99, 95% CI 1.07-8.37) for AWS treatment were independently associated with a higher mortality. The development of AWS increased the rate of infections (OR = 2.24, 95% CI 1.44-3.49), the need for mechanical ventilation (OR = 2.49, 95% CI 1.38-4.49), and ICU admission (OR = 1.96, 95% CI 1.19-3.23). Finally, AWS was associated with higher 28-day (HR = 2.31, 95% CI 1.40-3.82), 90-day (HR = 1.78, 95% CI 1.18-2.69), and 180-day mortality (HR = 1.54, 95% CI 1.06-2.24). Interpretation: AWS commonly occurs in patients hospitalized with AH and complicates the hospitalization course. Routine prophylaxis is associated with a lower prevalence of AWS. Prospective studies should determine diagnostic criteria and prophylaxis regimens for AWS management in patients with AH. Funding: This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.

Widespread vitiligo and poliosis following ipilimumab-nivolumab combination therapy.

INTRODUCTION: Combined immune checkpoint inhibitor therapy has been successfully used in the treatment of several malignancies. Adverse effects with the combination therapy may be more severe than the ones seen with single immune checkpoint inhibitors. CASE PRESENTATION: We report a unique case of a 59-year-old man of dark skin complexion who underwent treatment with intravenous ipilimumab-nivolumab every 3 weeks for metastatic malignant melanoma. After three cycles of this therapy, he developed extensive skin depigmentation that within 6 weeks, affected nearly the entire skin surface, along with progressive poliosis. MANAGEMENT AND OUTCOME: Ipilimumab-nivolumab therapy was subsequently discontinued due to grade 3 enterocolitis requiring high-dose steroids and intravenous infliximab. About six months later, imaging studies showed a relapse of malignant melanoma. At that juncture, vitiligo affected the total body surface area, resembling albinism, along with near-total poliosis and significant photosensitivity. Pembrolizumab was tried but had to be stopped after three cycles due to the reoccurrence of grade 3 enterocolitis. Progression of malignant melanoma with new brain, lung, liver, subcutaneous, and colonic metastases led to the patient's demise. CONCLUSION: We report a unique case of severe vitiligo and poliosis that involved total body surface area in a Caucasian man with dark complexion, resembling albinism. Further studies are warranted to evaluate the severity of dermatologic side effects with combination immune checkpoint inhibitor therapy.

Novel Application of Conjunctival Anterior Segment Optical Coherence Tomography Angiography to Assess Ocular Redness.

PURPOSE: The aim of this study was to determine anterior segment optical coherence tomography angiography (AS-OCTA) parameters to assess ocular redness severity. METHODS: AS-OCTA analyses of 60 eyes of 40 patients were grouped according to ocular redness stages using the 5-category validated bulbar redness scale in a cross-sectional retrospective study (groups 1-5). A subset of patients with slit-lamp photographs, total 35 eyes of 23 patients, were assessed with 10-category validated bulbar redness scale for comparison. AS-OCTA images of nasal and temporal bulbar conjunctiva were analyzed. Vessel density (VD) represented the blood flow pixels by the total pixels of image (%); vessel diameter index represented the VD by the skeletonized density; fractal dimension, measured with the box-count method, represented the vessel branching complexity. Averaged nasal and temporal parameters for each eye were correlated to validated bulbar redness scales. RESULTS: There was no statistical difference between groups for age ( P = 0.118), sex ( P = 0.501), eye laterality (OD/OS; P = 0.111), or location (nasal/temporal; P = 0.932). In the 5-category scale, VD significantly increased from group 1 to 2 (31.5 ± 1.9% and 33.4 ± 2.2%, P = 0.023), 2 to 3 (36.0 ± 3.5%, P < 0.001), and 4 to 5 (40.2 ± 2.9 and 46.5 ± 2.8, P < 0.001). The correlations were 0.805 ( P < 0.001) and 0.893 ( P < 0.001) for the 5-category and 10-category scales, respectively. Vessel diameter index showed a significant increase from 1 to 2 (2.90 ± 0.17 and 3.00 ± 0.15; P = 0.004) and 4 to 5 (2.92 ± 0.31 and 3.33 ± 0.08; P = 0.001). The correlations were 0.550 ( P < 0.001) and 0.625 ( P < 0.001) for the respective scales. The fractal dimension showed no significant differences between subsequent groups. The correlations were 0.445 ( P < 0.001) and 0.583 ( P < 0.001), respectively. CONCLUSIONS: Conjunctival AS-OCTA VD was the most reliable parameter to assess ocular redness.

Corneal nerve regeneration is affected by scar location in herpes simplex keratitis: A longitudinal in vivo confocal microscopy study.

PURPOSE: To assess the effect of corneal scar location on corneal nerve regeneration in patients with herpes simplex virus (HSV) keratitis in their affected and contralateral eyes over a 1-year period by in vivo confocal microscopy (IVCM), and to correlate these findings to corneal sensation measured by Cochet-Bonnet Esthesiometer. METHODS: Prospective, longitudinal, case-control study. Bilateral corneal nerve density and corneal sensation were analyzed centrally and peripherally in 24 healthy controls and 23 patients with unilateral HSV-related corneal scars using IVCM. RESULTS: In the central scar (CS) group, total nerve density in the central cornea remained significantly lower compared to controls at follow-up (11.05 ± 1.97mm/mm2, p < 0.001), and no significant nerve regeneration was observed (p = 0.090). At follow-up, total nerve density was not significantly different from controls in the central and peripheral cornea of the peripheral scar (PS) group (all p > 0.05), but significant nerve regeneration was observed in central corneas (16.39 ± 2.39mm/mm2, p = 0.007) compared to baseline. In contralateral eyes, no significant corneal nerve regeneration was observed in central or peripheral corneas of patients with central scars or peripheral scars at 1-year follow-up, compared to baseline (p > 0.05). There was a positive correlation between corneal nerve density and sensation in both central (R = 0.53, p < 0.0001) and peripheral corneas (R = 0.27, p = 0.0004). In the CS group, the corneal sensitivity was <4 cm in 4 (30.8%) and 7 (53.8%) patients in the central and peripheral corneas at baseline, and in 5 (38.5%) and 2 subjects (15.4%) at follow-up, whereas in the PS group only 1 patient (10%) showed a corneal sensation < 4 cm in the central cornea at baseline, and only 1 (10.0%), 3 (30.0%) and 1 (10.0%) patients at follow-up in the central, affected and opposite area of the cornea, respectively. CONCLUSION: The location of HSV scarring in the cornea affects the level of corneal nerve regeneration. Eyes with central corneal scar have a diminished capacity to regenerate nerves in central cornea, show a more severe reduction in corneal sensation in the central and peripheral corneas that persist at follow-up, and have a reduced capability to restore the corneal sensitivity above the cut-off of 4 cm. Thus, clinicians should be aware that CS patients would benefit from closer monitoring for potential complications associated with neurotrophic keratopathy, as they have a lower likelihood for nerve regeneration.

Acute pancreatitis as an adverse effect of COVID-19 vaccination.

While vaccination against COVID-19 has significantly improved the morbidity and mortality of the disease, with the increase in the administration of COVID-19 vaccines, it is more likely to observe their rare side effects in the clinical settings. Herein, we report a case of an 82-year-old man with history of coronary artery disease, prostate cancer in remission, gastroesophageal reflux disease, and hypothyroidism, who presented with acute pancreatitis few hours after receiving the third dose of Pfizer-BioNTech BNT162b2 mRNA COVID-19 vaccine, without other identified etiology. His symptoms were mild and he was discharged in a stable condition after improvement in his condition with supportive care.

Automated Image Threshold Method Comparison for Conjunctival Vessel Quantification on Optical Coherence Tomography Angiography.

Purpose: To determine the impact of image binarization and the best thresholding method for conjunctival optical coherence tomography angiography (OCTA). Methods: Vessel density (VD) of 14 OCTA conjunctival images (nine nasal and five temporal conjunctivas, and eight right and six left eyes) from normal subjects was analyzed. The binarization of gold-standard images, created by removing pixels that do not represent vessels on ImageJ software, was assessed by three masked graders to determine consistency of VD for images. Various thresholding methods on ImageJ, including manual, 1-, 2- and 3-step processes, were performed on unprocessed images for comparison. Interclass correlation coefficient (ICC) ≥0.750 were classified as good reliability and selected for calculation of the performance of the pixel location in the binarized images of each method. Results: Analysis of the gold-standard threshold method achieved an ICC of 0.816 with excellent agreement (R2 = 0.965, P < 0.001). From a total 28 different methods and variations performed, only nine methods performed with good reliability, including two 1-step thresholds, six 2-step thresholds, and one 3-step threshold method. Overall, 2-step threshold methods were more reliable than 3-step threshold methods. The 2-step method of Bandpass filter + Phansalkar local threshold (LT) showed the best performance with mean pixel accuracy of 86.9% ± 6.8%, area under the curve of 0.826, sensitivity of 79.0%, and specificity 86.1%. Conclusions: Bandpass filter + Phansalkar LT was the best method for VD measurement in conjunctival OCTA. Most commonly reported threshold methods showed unsatisfactory agreement. There is a need in the OCTA field for a standardized method to allow comparison between different studies. Translational Relevance: The proposed threshold method using a widely accessible and commonly used software provides an accurate VD measurement for future OCTA studies.

Acute Pancreatitis in Individuals with COVID-19: A Case Report and Critical Review of Literature.

Involvement of gastrointestinal tract has been reported in individuals diagnosed with COVID-19. Herein, we report a case of 65-year-old woman with type 2 diabetes mellitus, hypertension, and end-stage renal disease undergoing hemodialysis who was initially diagnosed with COVID-19 on a screening test. During the course of the disease, her respiratory symptoms remained mild; however, she developed acute pancreatitis leading to severe hypertension and hyperosmolar hyperglycemic state. During the hospitalization and treatment of acute pancreatitis, hyperglycemia, and hypertension, her condition improved and she was discharged in stable condition.

Development of Monoclonal B-cell Lymphocytosis Shortly After Treatment with Nivolumab and Subsequent Progression to Chronic Lymphocytic Leukemia.

INTRODUCTION: Immune checkpoint inhibitors, such as programmed death (PD)-1 inhibitor nivolumab, are currently widely used in treatment of various malignancies. Due to their widespread application, any new potential adverse effects due to these agents necessitate careful assessment. CASE REPORT: We report a case of an 81-year-old man with recurrent high-risk malignant melanoma who underwent a 12-month adjuvant treatment with nivolumab. Shortly after the course of nivolumab, he developed monoclonal B-cell lymphocytosis (MBL) which subsequently progressed to chronic lymphocytic leukemia (CLL). MANAGEMENT AND OUTCOME: The patient is currently doing clinically well in Rai stage 0. Malignant melanoma remains in remission. CONCLUSION: Considering the pathophysiologic plausibility of nivolumab inducing B-cell dysregulation via PD-1 inhibition, we suggest further studies on potential association between nivolumab and B-cell malignancies.

Effect of herpes simplex keratitis scar location on bilateral corneal nerve alterations: an in vivo confocal microscopy study.

AIMS: To evaluate the impact of herpes simplex virus (HSV)-induced scar location on bilateral corneal nerve alterations using laser in vivo confocal microscopy (IVCM). METHODS: Central and peripheral corneal subbasal nerve density (CSND) were assessed bilaterally in 39 patients with unilateral HSV-induced corneal scars (21 central scars (CS), 18 peripheral scars (PS)) using IVCM. Results were compared between patients and 24 age-matched controls. CSND was correlated to corneal sensation for all locations. RESULTS: Overall patients revealed significant decrease of CSND in the central and peripheral cornea (9.13±0.98 and 6.26±0.53 mm/mm2, p<0.001), compared with controls (22.60±0.77 and 9.88±0.49 mm/mm2). CS group showed a decrease in central (8.09±1.30 mm/mm2) and total peripheral nerves (5.15±0.62 mm/mm2) of the affected eyes, whereas PS group demonstrated a decrease in central (10.34±1.48 mm/mm2) and localised peripheral nerves only in the scar area (4.22±0.77 mm/mm2) (all p<0.001). In contralateral eyes, CSND decreased in the central cornea of the CS group (16.88±1.27, p=0.004), and in the peripheral area, mirroring the scar area in the affected eyes of the PS group (7.20±0.87, p=0.032). Corneal sensation significantly decreased in the whole cornea of the affected, but not in contralateral eyes (p<0.001). A positive correlation between CSND and corneal sensation was found in all locations (p<0.001). CONCLUSIONS: Patients with HSV scar demonstrate bilateral CSND decrease as shown by IVCM. CSND and corneal sensation decrease in both central and peripheral cornea in affected eyes, although only in the scar area in PS group. Interestingly, diminishment of CSND was found locally in the contralateral eyes, corresponding and mirroring the scar location in the affected eyes.

The Effects of Helicobacter Pylori Eradication on Liver Function and Metabolic Profile in Non-diabetic Non-alcoholic Steatohepatitis: A 5-year Randomized Clinical Trial.

BACKGROUND: To evaluate the effects of Helicobacter pylori (HP) eradication on liver function tests (LFT) and fat content (LFC) in non-diabetic non-alcoholic steatohepatitis (NASH). METHODS: This randomized clinical trial included dyspeptic HP infected non-diabetic NASH participants. The intervention arm received HP eradication treatment, while the control arm did not get any HP treatment. In the meantime, the standard management of NASH was performed in both trial arms. Mean alterations in LFT were the primary outcome and the secondary outcomes included the mean changes in LFC and serum metabolic profile. The trial follow-up period was 5 years. RESULTS: 40 participants (female: 20), with a mean age of 41.58 (±12.31) years, were enrolled in the study. The HP eradication arm included 20 participants (female: 11) with a mean age of 40.25 (±10.59) years, and the control arm consisted of 20 individuals (female: 9) with a mean age of 42.90 (±13.97) years. The tests of within-subjects effects showed a significant decrease in mean serum alanine aminotransferase (ALT; P=0.007), triglyceride (TG; P=0.04), cholesterol (P=0.004), and fasting blood sugar (FBS; P<0.001), and an increase in high-density lipoprotein (HDL; P=0.04) in both research groups during the study period. The tests of between-subjects effects demonstrated a more significant decrement of FBS in HP eradicated patients than the controls (P=0.02). The reduction in waist circumference, aspartate aminotransferase (AST), ALT, alkaline phosphatase, triglyceride, cholesterol, low-density lipoprotein, insulin, and LFC were more prominent in the intervention group than the controls; however, these differences were not statistically significant. CONCLUSION: Adding HP eradication treatment to standard NASH treatment showed more therapeutic effect thanthe standard NASH treatment protocol alone regarding the decrement of FBS in participants with dyspeptic non-diabetic NASH. Considering the non-statistically significant improvement in other metabolic indices and LFT in this trial, further studies are recommended.

Herpes simplex virus-1 KOS-63 strain is virulent and causes titer-dependent corneal nerve damage and keratitis.

To investigate the acute clinical, immunological, and corneal nerve changes following corneal HSV-1 KOS-63 strain inoculation. Corneas of C57BL/6 mice were inoculated with either low dose (Ld) or high dose (Hd) HSV-1 KOS-63 or culture medium. Clinical evaluation was conducted up to 7 days post inoculation (dpi). Viral titers were assessed by standard plaque assay. Excised corneas were stained for CD45 and beta-III tubulin. Corneal flow cytometry was performed to assess changes in leukocyte subpopulations. Corneal sensation was measured using a Cochet-Bonnet esthesiometer. Naïve, sham-infected (post scarification), and McKrae-infected C57BL/6 corneas served as two negative and positive controls, respectively. Compared to Ld infected mice, Hd HSV-1 KOS-63 demonstrated higher incidence of corneal opacity (1.5 ×) and neovascularization (2.6 × ; p < 0.05). At 7 dpi Hd infected mice showed more severe corneal opacity (2.23 vs. 0.87; p = 0.0003), neovascularization (6.00 vs. 0.75; p < 0.0001), and blepharitis (3.11 vs. 2.06; p = 0.001) compared to the Ld group. At 3 dpi epitheliopathy was significantly larger in the Hd group (23.59% vs. 3.44%; p = 0.001). Similarly, corneal opacity was significantly higher in Hd McKrae-infected corneas as compared with Ld McKrae-infected corneas at 3 and 5 dpi. No significant corneal opacity, neovascularization, blepharitis, and epitheliopathy were observed in naïve or sham-infected mice. Higher viral titers were detected in corneas (1 and 3 dpi) and trigeminal ganglia (TG) (3 and 5 dpi) in Hd versus Ld KOS-63 groups (p < 0.05). Leukocyte density showed a gradual increase over time from 1 to 7 dpi in both KOS-63 and McKrae-infected corneas. Corneal flow cytometric analysis (3 dpi) demonstrated a higher percentage of Gr-1 + (71.6 vs. 26.3) and CD11b + (90.6 vs. 41.1) cells in Hd versus Ld KOS-63 groups. Corneal nerve density significantly decreased in both Hd KOS-63 and Hd McKrae infected corneas in comparison with naïve and sham-infected corneas. At 3 dpi corneal nerve density was lower in the Hd versus Ld KOS-63 groups (16.79 vs. 57.41 mm/mm2; p = 0.004). Corneal sensation decreased accordingly at 5 and 7 dpi in both Ld and Hd KOS-63-infected mice. Corneal inoculation with HSV-1 KOS-63 strain shows acute keratitis and nerve degeneration in a dose-dependent fashion, demonstrating virulence of this strain.

Plasmacytoid dendritic cells in the eye.

Plasmacytoid dendritic cells (pDCs) are a unique subpopulation of immune cells, distinct from classical dendritic cells. pDCs are generated in the bone marrow and following development, they typically home to secondary lymphoid tissues. While peripheral tissues are generally devoid of pDCs during steady state, few tissues, including the lung, kidney, vagina, and in particular ocular tissues harbor resident pDCs. pDCs were originally appreciated for their potential to produce large quantities of type I interferons in viral immunity. Subsequent studies have now unraveled their pivotal role in mediating immune responses, in particular in the induction of tolerance. In this review, we summarize our current knowledge on pDCs in ocular tissues in both mice and humans, in particular in the cornea, limbus, conjunctiva, choroid, retina, and lacrimal gland. Further, we will review our current understanding on the significance of pDCs in ameliorating inflammatory responses during herpes simplex virus keratitis, sterile inflammation, and corneal transplantation. Moreover, we describe their novel and pivotal neuroprotective role, their key function in preserving corneal angiogenic privilege, as well as their potential application as a cell-based therapy for ocular diseases.

Topical Adoptive Transfer of Plasmacytoid Dendritic Cells for Corneal Wound Healing.

Plasmacytoid dendritic cells (pDCs) are crucial for corneal homeostasis through secretion of various anti-angiogenic molecules and growth factors. Due to its avascular nature, only a limited number of adoptively transferred cells home to the cornea, when administered systemically. In addition, local adoptive transfer of cells poses several challenges and the clinical application of commonly used techniques is limited. Herein, we detail a novel approach for local adoptive transfer of pDCs to the cornea for the treatment of corneal wounds. This approach utilizes a commonly used fibrin sealant as a means of transferring previously isolated cells locally on the cornea. The technique is simple, reproducible, and is accompanied with successful transfer and integration of a substantial number of the cells to the cornea. Application of this approach to transfer pDCs promotes corneal wound healing. Furthermore, this technique can be applied for adoptive transfer of any cell of interest to the cornea.

Correlation of corneal immune cell changes with clinical severity in dry eye disease: An in vivo confocal microscopy study.

PURPOSE: To evaluate corneal immune dendritiform cell (DC) changes in dry eye disease (DED) using in vivo confocal microscopy (IVCM) and to correlate IVCM parameters with clinical severity. METHODS: This was a retrospective, cross-sectional study including 300 eyes of 150 DED patients and 49 eyes of 49 age-matched controls. Severity of DED was based on the Dry Eye Workshop (DEWS) classification. IVCM images of subbasal layer of the central cornea were analyzed for DC density and morphology (including number of dendrites per DC, DC size and DC field). RESULTS: DC density was significantly higher in DED compared to controls (93.4 ± 6.3 vs. 25.9 ± 3.9 cells/mm2; P < 0.001). Morphologically, number of dendrites, DC size and field were significantly larger in DED (3.3 ± 0.1, 106.9 ± 4.7 µm2, 403.8 ± 20.1 µm2 than controls (2.3 ± 0.1, 62.5 ± 5.7 µm2, 241.4 ± 24.4 µm2, P < 0.001). Significantly higher DC density compared to controls was observed as early as Level 1 DED severity (87 ± 10 cells/mm2, p < 0.001. Significant morphological changes in DC were detected for Levels 2 to 4 (p=<0.001, and p =< 0.05) for dendrites and DC field, respectively. Similarly, DC size showed significant increase at DED level 3-4. (p < 0.05). Linear regression analysis showed that both conjunctival and corneal staining were independently associated with DC density, while corneal staining was independently associated with DC morphology. CONCLUSION: DC density and morphology correlated with clinical severity of DED. While, DC density is increased in mild DED, morphological changes are seen only in severe cases. IVCM may be a powerful tool to detect early immune changes and may complement clinical examination in DED.

Effect of Dry Eye Disease on the Kinetics of Lacrimal Gland Dendritic Cells as Visualized by Intravital Multi-Photon Microscopy.

The lacrimal gland (LG) is the main source of the tear film aqueous layer and its dysfunction results in dry eye disease (DED), a chronic immune-mediated disorder of the ocular surface. The desiccating stress (DS) murine model that mimics human DED, results in LG dysfunction, immune cell infiltration, and consequently insufficient tear production. To date, the immune cell kinetics in DED are poorly understood. The purpose of this study was to develop a murine model of intravital multi-photon microscopy (IV-MPM) for the LG, and to investigate the migratory kinetics and 3D morphological properties of conventional dendritic cells (cDCs), the professional antigen presenting cells of the ocular surface, in DED. Mice were placed in a controlled environmental chamber with low humidity and increased airflow rate for 2 and 4 weeks to induce DED, while control naïve transgenic mice were housed under standard conditions. DED mice had significantly decreased tear secretion and increased fluorescein staining (p < 0.01) compared to naïve controls. Histological analysis of the LG exhibited infiltrating mononuclear and polymorphonuclear cells (p < 0.05), as well as increased LG swelling (p < 0.001) in DED mice compared to controls. Immunofluorescence staining revealed increased density of cDCs in DED mice (p < 0.001). IV-MPM of the LG demonstrated increased density of cDCs in the LGs of DED mice, compared with controls (p < 0.001). cDCs were more spherical in DED at both time points compared to controls (p < 0.001); however, differences in surface area were found at 2 weeks in DED compared with naïve controls (p < 0.001). Similarly, 3D cell volume was significantly lower at 2 weeks in DED vs. the naïve controls (p < 0.001). 3D instantaneous velocity and mean track speed were significantly higher in DED compared to naïve mice (p < 0.001). Finally, the meandering index, an index for directionality, was significant increased at 4 weeks after DED compared with controls and 2 weeks of DED (p < 0.001). Our IV-MPM study sheds light into the 3D morphological alterations and cDC kinetics in the LG during DED. While in naïve LGs, cDCs exhibit a more dendritic morphology and are less motile, they became more spherical with enhanced motility during DED. This study shows that IV-MPM represents a robust tool to study immune cell trafficking and kinetics in the LG, which might elucidate cellular alterations in immunological diseases, such as DED.

Characterization of Resident Corneal Plasmacytoid Dendritic Cells and Their Pivotal Role in Herpes Simplex Keratitis.

The presence and potential functions of resident plasmacytoid dendritic cells (pDCs) in peripheral tissues is unclear. We report that pDCs constitutively populate naïve corneas and are increased during sterile injuries or acute herpes simplex virus 1 (HSV-1) keratitis. Their local depletion leads to severe clinical disease, nerve loss, viral dissemination to the trigeminal ganglion and draining lymph nodes, and mortality, while their local adoptive transfer limits disease. pDCs are the main source of HSV-1-induced IFN-α in the corneal stroma through TLR9, and they prevent re-programming of regulatory T cells (Tregs) to effector ex-Tregs. Clinical signs of infection are observed in pDC-depleted corneas, but not in pDC-sufficient corneas, following low-dose HSV-1 inoculation, suggesting their critical role in corneal antiviral immunity. Our findings demonstrate a vital role for corneal pDCs in the control of local viral infections.

A randomized, sham-controlled trial of intraductal meibomian gland probing with or without topical antibiotic/steroid for obstructive meibomian gland dysfunction.

IMPORTANCE: Obstructive meibomian gland dysfunction (MGD) can be refractory to medical therapy. Intraductal meibomian gland (MG) probing may offer a potential therapeutic approach for these patients, but no randomized trials have been conducted to date. OBJECTIVE: To assess clinical changes after intraductal MG probing for patients with refractory obstructive meibomian gland dysfunction. DESIGN: Randomized, double-masked, sham-controlled clinical trial. SETTING: Single-center, tertiary referral center. PARTICIPANTS: 42 patients with refractory obstructive MGD associated with lid tenderness. INTERVENTIONS: Enrolled patients received one of the following treatments: 1) MG probing plus post-procedural topical sulfacetamide/prednisolone ointment (Blephamide®), 2) MG probing plus post-procedural lubricating ointment (GenTeal), or 3) sham probing plus GenTeal ointment. The probing was performed on the upper lids of both eyes. MAIN OUTCOME MEASURES: Primary outcome measures were symptoms as measured by Ocular Surface Disease Index (OSDI) and Symptom Assessment iN Dry Eye (SANDE), as well as tear break-up time (TBUT). Secondary outcome measures were other clinical signs. Safety of the procedure was also evaluated by investigating the treatment-related adverse events. At baseline and 4 weeks after the procedure a masked observer evaluated the following outcome measures: symptom questionnaires, including OSDI and SANDE, upper lid tenderness, lid margin telangiectasia, corneal fluorescein staining, conjunctival lissamine green staining, TBUT, Schirmer's test, and meibomian glands yielding liquid secretion (MGYLS). RESULTS: Compared to baseline, the MG probing/Blephamide® group showed significant improvements in both OSDI and SANDE scores and the MG probing/GenTeal group demonstrated a significant improvement only in SANDE score. In contrast, the Sham/GenTeal group did not show any statistically significant changes in symptoms. There were no statistically significant changes in clinical signs in any group at the 4-week visit, except for improvement of lid tenderness in the sham probing group. CONCLUSIONS: MG probing/Blephamide® results in a significant improvement in symptoms in patients with refractory obstructive MGD without any significant effect on clinical signs. Larger studies are warranted to determine the efficacy of MG probing. TRIAL REGISTRATION: Clinicaltrials.gov(identifier NCT02256969, Filed on 08/13/2014).

Visualization of microneuromas by using in vivo confocal microscopy: An objective biomarker for the diagnosis of neuropathic corneal pain?

PURPOSE: The diagnosis of neuropathic corneal pain (NCP) is challenging, as it is often difficult to differentiate it from conventional dry eye disease (DED). In addition to eye pain, NCP can present with similar signs and symptoms of DED. The purpose of this study is to find an objective diagnostic sign to identify patients with NCP, using in vivo confocal microscopy (IVCM). METHODS: This was a comparative, retrospective, case-control study. Patients with clinical diagnosis of NCP (n = 25), DED (n = 30), and age- and sex-matched healthy controls (n = 16), who underwent corneal imaging with IVCM (HRT3/RCM) were included. Central corneal IVCM scans were analyzed by 2 masked observers for nerve density and number, presence of microneuromas (terminal enlargements of subbasal corneal nerve) and/or nerve beading (bead-like formation along the nerves), and dendritiform cell (DC) density. RESULTS: There was a decrease in total nerve density in both NCP (14.14 ± 1.03 mm/mm2) and DED patients (12.86 ± 1.04 mm/mm2), as compared to normal controls (23.90 ± 0.92 mm/mm2; p < 0.001). However, total nerve density was not statistically different between NCP and DED patients (p = 0.63). Presence of nerve beading was not significantly different between patients and normal controls (p = 0.15). Interestingly, microneuromas were observed in all patients with NCP, while they were not present in any of the patients with conventional DED (sensitivity and specificity of 100%). DC density was significantly increased in both NCP (71.89 ± 16.91 cells/mm2) and DED patients (111.5 ± 23.86 cells/mm2), as compared to normal controls (24.81 ± 4.48 cells/mm2 (p < 0.05). However, there was no significant difference in DC density between DED and NCP patients (p = 0.31). CONCLUSION: IVCM may be used as an adjunct diagnostic tool for the diagnosis of NCP in the presence of neuropathic symptoms. Microneuromas may serve as a sensitive and specific biomarker for the diagnosis of NCP.

Intravital Multiphoton Microscopy of the Ocular Surface: Alterations in Conventional Dendritic Cell Morphology and Kinetics in Dry Eye Disease.

Dry eye disease (DED) is a multifactorial disease of the ocular surface, characterized by loss of tear film homeostasis and ocular symptoms, in which neurosensory abnormalities have recently been shown to play an etiological role. Although the role of inflammation has been widely studied in DED, the kinetics of immune cells of the ocular surface in this complex disease are hereto unclear. Herein, we utilized intravital multiphoton imaging on transgenic mice to investigate the 3D morphology and kinetics of conventional dendritic cells (cDCs) and the role of ocular surface sensory nerves in regulating them in both the naïve state and experimental DED. Mice with DED had significantly lower tear secretion (p < 0.01), greater corneal fluorescein staining (p < 0.001), and higher cDC density in the ocular surface (p < 0.05), compared to naïve mice. cDCs in DED mice showed morphological alterations in the limbus, exhibiting smaller surface area (p < 0.001) and volume (p < 0.001) compared to naïve mice. Furthermore, corneal cDCs showed greater sphericity in DED mice compared to naïve mice (p < 0.01). In addition, limbal cDCs displayed significantly increased migratory kinetics in DED, including mean track speed, 3D instantaneous velocity, track length, and displacement, compared to naïve mice (all p < 0.05). In mice with DED, cDCs showed a higher meandering index in the limbus compared to central cornea (p < 0.05). In DED, cDCs were less frequently found in contact with nerves in the limbus, peripheral, and central cornea (p < 0.05). cDCs in contact with nerves demonstrated a larger surface area (p < 0.001) and volume (p < 0.001), however, they exhibited less sphericity (p < 0.05) as compared to cDCs not in contact with nerves in naïve mice. Importantly, cDCs in contact with nerves during DED had a decreased track length, displacement, mean track speed, and 3D instantaneous velocity compared to those not in contact with nerves (all p < 0.05). Taken together, we present in vivo evidence of altered cDC kinetics and 3D morphology in DED. Furthermore, apparent neuronal contact significantly alters cDC kinetics and morphological characteristics, suggesting that ocular surface nerves may play a direct role in mediating immune responses in DED.