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Background: The long-term effects of mild Traumatic Brain Injury (TBI) in children and adolescents are increasingly discussed due to their potential impact on psycho-social development and education. This study aims to evaluate post-hospital care of children and adolescents after mild TBI using a physician survey. Methods: A self-developed, pre-tested questionnaire on diagnostics and treatment of TBI in outpatient care was sent to a representative sample of general practitioners and pediatricians in Germany. Results: Datasets from 699 general practitioners, 334 pediatricians and 24 neuropediatricians were available and included in the analysis. Nearly half of the general practitioners and most pediatricians say they treat at least one acute pediatric TBI per year. However, a substantive proportion of general practitioners are not familiar with scales assessing TBI severity and have difficulties assessing the symptoms correctly. Pediatricians seem to have better knowledge than general practitioners when it comes to treatment and outpatient care of TBI. Conclusions: To increase knowledge about TBI in outpatient physicians, targeted training courses should be offered, especially for general practitioners. Moreover, handing out written information about long-term effects and reintegration after TBI should be encouraged in outpatient practice.

BACKGROUND: Dry eye syndrome (DES) is a prevalent condition, but information on risk or protective factors is lacking. OBJECTIVE: We aimed to determine the association between the dietary intake and ratio of n-3 and n-6 fatty acids (FAs) and DES occurrence. DESIGN: Of the 39876 female health professionals in the Women's Health Study (WHS), 32470 women aged 45-84 y who provided information on diet and DES were cross-sectionally studied. We assessed FA intakes by using a validated food-frequency questionnaire and assessed DES by using self-reports of clinically diagnosed cases. Of the sample, 1546 (4.7%) subjects reported DES. We used logistic regression models to estimate the odds ratios (ORs) and 95% CIs to describe the relation of FA intake with DES. RESULTS: After adjustment for demographic factors, hormone therapy, and total fat intake, the OR for the highest versus the lowest quintile of n-3 FAs was 0.83 (95% CI: 0.70, 0.98; P for trend = 0.05). A higher ratio of n-6 to n-3 FA consumption was associated with a significantly increased risk of DES (OR: 2.51; 95% CI: 1.13, 5.58) for >15:1 versus <4:1 (P for trend = 0.01). In addition, tuna consumption [1 serving was 113 g (4 oz)] was inversely associated with DES (OR: 0.81; 95% CI: 0.66, 0.99 for 2-4 servings/wk; OR: 0.32; 95% CI: 0.13, 0.79 for 5-6 servings/wk versus < or =1 serving/wk; P for trend = 0.005). CONCLUSIONS: These results suggest that a higher dietary intake of n-3 FAs is associated with a decreased incidence of DES in women. These findings are consistent with anecdotal clinical observations and postulated biological mechanisms. Copyright © 2005 American Society for Clinical Nutrition [ABSTRACT FROM AUTHOR]

El incremento de la preinscripción de fármacos analgésicos opioides se ha asociado a un aumento de las tasas de abuso y las consecuencias negativas asociadas a su uso inadecuado. Diversos organismos sanitarios internacionales recomiendan la realización de una evaluación global y multidisciplinar del paciente durante todo el proceso terapéutico con opiáceos, con el fin de identificar un posible abuso. Ante la ausencia de quías específicas en el ámbito sanitario español, el objetivo de este artículo ha sido presentar una propuesta de evaluación psicológica, en base a los principales instrumentos de evaluación, disponibles actualmente para evaluar el abuso de opioides. Se establecen pautas para el proceso de evaluación en función de las variables psicológicas que puedan predecir y mantener dicho abuso, clasificando todo ello en función del momento del proceso terapéutico en el que los pacientes se encuentren, Aunque existen instrumentos con buenas propiedades psicométricas, son necesarias futuras investigaciones para la adaptación, traducción y validación de éstos a población española. Al mismo tiempo son necesarios futuros estudios que profundicen en estrategias de prevención e intervención para reducir la probabilidad de abuso en pacientes tratados con fármacos opioides.

The increase in the prescription of opioid analgesics is related to increased rates of opioid abuse and the negative consequences of medication misuse. Several international health organisations recommend comprehensive and multidisciplinary patient assessment for the duration of the opioid treatment in order to identify and prevent medication abuse. Due to the lack of specific clinical guidelines in the Spanish National Health System, the aim of this paper is to present a proposal for psychological assessment based on the main psychological tools currently available for assessing opioid abuse. The assessment guidelines have been established based on the psychological variables that can predict and prolong the abuse, classifying all of the variables depending on the current stage of the therapeutic process for each patient. Although there are instruments with good psychometric properties, further research is necessary to adapt, translate and validate these instruments for use in the Spanish population. Future studies are also needed to investigate intervention and prevention strategies in depth in order to reduce the likelihood of abuse in patients treated with opioids.

Examines the hypothesis that sex steroid hormones are critical etiological factors in the pathogenesis of evaporative dry eye in Sjorgen's syndrome. Regulation of quality and/or quantity of lipids produced by the tissue; Formation of tear film's lipid layer; Estrogen overproduction.

Although several studies have previously focused on the conjunctival epithelial response to surface dryness, little is known about the effect of a dry environment on corneal epithelium, which is the most clinically significant tissue affected in dry eye. The aim of this study was to quantitatively evaluate the effect of desiccating stress on the number of proliferating corneal epithelial cells and corneal epithelial thickness in mice placed in a controlled-environment chamber (CEC) that induces dry eye. Corneal epithelial cell proliferation and thickness were studied in 8- to 12-week-old female BALB/c mice placed in the CEC (temperature: 22.3+/-0.7 degrees C; relative humidity: 22.5+/-4.5%; airflow: 15 L/min) for 7 days and compared to a control group of mice with no dry eye. Actively proliferating cells were identified by immunofluorescence using a FITC-conjugated antibody against the Ki-67 protein, a cell proliferation marker expressed during active phases of the cell cycle. To detect the spatial distribution of proliferative cells, Ki-67(+) cells were counted in three areas of the epithelium: center, periphery, and limbus. Corneal epithelial thickness was evaluated in the central cornea after staining with hematoxylin-eosin. Results from each experimental group were compared using the Mann-Whitney test. The number of Ki-67(+) cells observed in the corneal epithelium of mice exposed to the CEC was significantly higher in each area (center: 32.1+/-1.1; periphery: 94.2+/-5.3; limbus: 4.0+/-1.5) than in the control group (center: 13.2+/-1.0, p=0.02; periphery: 42.9+/-2.3, p=0.02; limbus: 0.0, p=0.01). In mice subjected to desiccating stress, a significant number of Ki-67(+) positive cells were detected in the basal and suprabasal cell layers (central area 46%; periphery 30.8%: limbus 0%), whereas in the control group the cells were exclusively distributed through the basal cell layer. Ki-67(+) cells were not found in the corneal stroma or endothelium in any group. The corneal epithelium was found to be significantly thicker in dry eye mice (54.94+/-6.09 microm) as compared to the controls (43.9+/-6.23 microm, p<0.0001) by a mean of 25%. These results demonstrate that desiccating stress increases corneal epithelial turnover and thickness, similar to what is observed in other chronic inflammatory states of other epithelialized surfaces. The CEC can facilitate the study of the regulation of epithelial cell function and turnover at the molecular and cellular levels under desiccating stress conditions.

Background: Corneal grafting is by far the most common form of transplantation. Many grafts suffer from immune rejection and current therapies are associated with many side effects, requiring more effective and safe therapies. alpha-Melanocyte stimulating hormone (alpha-MSH) is a neuropeptide that suppresses host inflammatory defense mechanisms. The purpose of this study was to determine the role of local therapy with alpha-MSH on corneal allograft survival, and the mechanisms by which it may influence graft outcome., Methods: Orthotopic corneal transplantation was performed, with recipients receiving subconjunctival alpha-MSH or sham injections twice weekly. Grafts were followed up for 70 days, and graft inflammation/opacification was compared between the two groups in a masked fashion. Graft infiltration and ocular gene expression of select inflammatory cytokines was evaluated at different timepoints. Additionally, allospecific delayed-type hypersensitivity was compared among the groups 3 weeks posttransplantation., Results: Results showed a significant increase in corneal graft survival in alpha-MSH-treated recipients compared with controls. Although 75% of allografts in alpha-MSH-treated hosts survived at 70 days, 43% survived in controls (P=0.04). Graft infiltration studies demonstrated a significant decrease in the number of mononuclear and polymorphonuclear cells in alpha-MSH-treated mice compared with controls at days 7 and 14 after transplantation. Furthermore, allospecific delayed-type hypersensitivity and gene expression of interferon-gamma and interleukin-2 showed a significantly reduced expression in alpha-MSH-treated mice compared with controls., Conclusions: This study provides for the first time, in vivo evidence that treatment with local alpha-MSH may significantly reduce allorejection of orthotopic transplants.

Corneal transplantation is the oldest and the most common form of solid tissue transplantation in humans. Immunologic graft rejection is one of the main causes of short and long-term graft failure. Rejection involves donor tissue recognition and destruction by allo-specific immune cells of the recipient. This review outlines (1) the immunobiology of transplantation, with reference to ocular immune privilege, (2) factors that confer "high-risk" status to a graft and (3) the pathophysiologic mechanisms of corneal transplant rejection.

Objective: To study the efficacy of topical application of alpha-linolenic acid (ALA) and linoleic acid (LA) for dry eye treatment., Methods: Formulations containing ALA, LA, combined ALA and LA, or vehicle alone, were applied to dry eyes induced in mice. Corneal fluorescein staining and the number and maturation of corneal CD11b(+) cells were determined by a masked observer in the different treatment groups. Real-time polymerase chain reaction was used to quantify expression of inflammatory cytokines in the cornea and conjunctiva., Results: Dry eye induction significantly increased corneal fluorescein staining; CD11b(+) cell number and major histocompatibility complex Class II expression; corneal IL-1alpha and tumor necrosis factor alpha (TNF-alpha) expression; and conjunctival IL-1alpha, TNF-alpha, interferon gamma, IL-2, IL-6, and IL-10 expression. Treatment with ALA significantly decreased corneal fluorescein staining compared with both vehicle and untreated controls. Additionally, ALA treatment was associated with a significant decrease in CD11b(+) cell number, expression of corneal IL-1alpha and TNF-alpha, and conjunctival TNF-alpha., Conclusions: Topical ALA treatment led to a significant decrease in dry eye signs and inflammatory changes at both cellular and molecular levels., Clinical Relevance: Topical application of ALA omega-3 fatty acid may be a novel therapy to treat the clinical signs and inflammatory changes accompanying dry eye syndrome.

Objective: To evaluate whether hemangiogenesis, lymphangiogenesis, and concomitant invasion of mononuclear phagocytes occurring after high-risk corneal transplantation in already vascularized high-risk recipient corneal beds increase the risk for subsequent immune rejection., Methods: Three intrastromal sutures were left in place for 6 weeks in the corneas of BALB/c mice, causing neovascularization. Three weeks after suture removal, keratoplasty was performed (donors C57BL/6 mice). The treatment group received a vascular endothelial growth factor A (VEGF-A)-neutralizing cytokine trap at 0, 4, 7, and 14 days postoperatively (Fc protein was used as the control treatment). Morphometry was performed in corneal flat mounts using lymphatic endothelial hyaluronan receptor-1 (a specific lymphatic endothelial marker), CD31 (a panendothelial marker), and F4/80 (a marker for mononuclear phagocytes)., Results: After corneal transplantation, significant additional hemangiogenesis (mean area covered by vessels [SD], 68% [18%] postoperatively vs 40% [18%] preoperatively; P = .03) and lymphangiogenesis (12% [1.3%] postoperatively vs 9% [2.8%] preoperatively; P = .03) were observed. Postoperative neutralization of VEGF-A inhibited operation-induced hemangiogenesis (35% [8%]; P = .007) and lymphangiogenesis (6% [1.6%]; P = .03) and decreased the recruitment of mononuclear phagocytes into the graft (mean [SD], 501 cells/mm(2) [152] in treated mice vs 684 cells/mm(2) [35] in Fc controls; P = .03). After 8 weeks, 23% of the treated corneas were not rejected, whereas all control corneas were rejected after 21 days (P = .007)., Conclusions: Neutralization of VEGF-A after high-risk corneal transplantation effectively inhibits postoperative hemangiogenesis, lymphangiogenesis, and recruitment of antigen-presenting cells and improves corneal graft survival., Clinical Relevance: Blocking of VEGF-A after high-risk corneal transplantation may be a novel approach to improve graft survival.

Purpose: To report a case of a 34-year-old woman presenting with keratoglobus and posterior polymorphous corneal dystrophy (PPMD)., Methods: Observational case report including ophthalmologic examination, topographic findings, and specular microscopy findings., Results: This patient presented with corneal steepening on topography consistent with keratoglobus, as well as large areas of irregular polymorphous changes of the corneal endothelium on specular microscopy consistent with PPMD., Conclusions: We report the first case with clinical features of both keratoglobus and PPMD. This report brings forth the description of keratoglobus findings on Orbscan topography.

Programmed death-ligand (PD-L)1 and PD-L2, newer B7 superfamily members, are implicated in the negative regulation of immune responses and peripheral tolerance. To examine their function in alloimmunity, we used the murine model of orthotopic corneal transplantation. We demonstrate that PD-L1, but not PD-L2, is constitutively expressed at high levels by the corneal epithelial cells, and at low levels by corneal CD45+ cells in the stroma, whereas it is undetectable on stromal fibroblasts and corneal endothelial cells. Inflammation induces PD-L1 up-regulation by corneal epithelial cells, and infiltration of significant numbers of PD-L1+CD45+CD11b+ cells. Blockade with anti-PD-L1 mAb dramatically enhances rejection of C57BL/6 corneal allografts by BALB/c recipients. To examine the selective contribution of donor vs host PD-L1 in modulating allorejection, we used PD-L1-/- mice as hosts or donors of combined MHC and minor H-mismatched corneal grafts. BALB/c grafts placed in PD-L1-/- C57BL/6 hosts resulted in pronounced T cell priming in the draining lymph nodes, and universally underwent rapid rejection. Allografts from PD-L1-/- C57BL/6 donors were also significantly more susceptible to rejection than wild-type C57BL/6 grafts placed into BALB/c hosts, primarily as a result of increased T cell infiltration rather than enhanced priming. Taken together, our results identify differential roles for recipient vs donor PD-L1 in regulating induction vs effector of alloimmunity in corneal grafts, the most common form of tissue transplantation, and highlight the importance of peripheral tissue-derived PD-L1 in down-regulating local immune responses.

Corneal transplants normally enjoy a high percentage of survival, mainly because the eye is an immune-privileged site. When allograft failure occurs, it is most commonly due to rejection, an immune-mediated reaction that targets the corneal endothelium. While the exact mechanism by which the endothelium is targeted is still unknown, we postulate that corneal endothelial cell loss during allograft failure is mediated by apoptosis. Furthermore, because corneal endothelial cells do not normally regenerate, we hypothesize that suppressing apoptosis in the graft endothelium will promote transplant survival. In a murine model of transplantation, TUNEL staining and confocal microscopy showed apoptosis of the graft endothelium occurring in rejecting corneas as early as 2 weeks posttransplantation. We found that bcl-xL protected cultured corneal endothelial cells from apoptosis and that lentiviral delivery of bcl-xL to the corneal endothelium of donor corneas significantly improved the survival of allografts. These studies suggest a novel approach to improve corneal allograft survival by preventing apoptosis of the endothelium.

Dexamethasone, a synthetic corticosteroid, is widely used as a potent anti-inflammatory drug in various diseases including corneal angiogenesis. However, dexamethasone's impact on interleukin (IL)-1beta-dependent inflammatory angiogenesis is unknown. Here, we show that dexamethasone inhibits IL-1beta-induced neovascularization and the expression of the angiogenesis-related factors, vascular endothelial growth factor-A, KC, and prostaglandin E(2) in the mouse cornea 2 days after IL-1beta implantation. IL-1beta caused IkappaB-alpha phosphorylation in corneal stromal cells but not in infiltrated CD11b(+) cells 2 days after IL-1beta implantation. In contrast, both cell types were positive for phosphorylated IkappaB-alpha 4 days after IL-1beta implantation. Dexamethasone significantly inhibited IkappaB-alpha phosphorylation 2 and 4 days after IL-1beta implantation. Furthermore, dexamethasone inhibited IL-1beta-induced expression of vascular endothelial growth factor-A, KC, and prostaglandin E(2), and signaling of nuclear factor (NF)-kappaB in corneal fibroblasts in vitro. A selective NF-kappaB inhibitor attenuated IL-1beta-induced corneal angiogenesis. These findings suggest that NF-kappaB activation in the corneal stromal cells is an important early event during IL-1beta-induced corneal angiogenesis and that dexamethasone inhibits IL-1beta-induced angiogenesis partially via blocking NF-kappaB signaling.

Objective: To determine whether the ocular anterior segment (aqueous humor and cornea) actively inhibits dendritic cell (DC) maturation., Methods: Dendritic cells were injected into syngeneic corneas or conjunctivae, and their surface major histocompatibility complex class II expression in response to the local milieu was assessed using confocal microscopy. Immature DCs were cocultured with corneal supernatant or with aqueous humor to evaluate their regulation of DC phenotypic and functional maturity., Results: In contrast to conjunctivally injected DCs, DCs injected into the cornea resisted up-regulation in expression of surface major histocompatibility complex class II. Corneal supernatant-treated and aqueous humor-treated DCs retained their immaturity, as reflected by high antigen uptake but low costimulatory molecule (CD80 and CD86) expression and poor T-cell stimulation. Anti-transforming growth factor beta(2) treatment of aqueous humor and of corneal supernatant led to complete and partial blockade of their inhibition of DC maturation, respectively. However, alpha-melanocyte-stimulating hormone and calcitonin gene-related peptide had no demonstrable effect on DC maturation., Conclusion: Cornea and aqueous humor, principally through transforming growth factor beta(2,) promote generation of phenotypically and functionally immature DCs. Clinical Relevance Our results indicate that relative immune quiescence in the cornea and in the anterior segment is actively maintained in part by the inhibitory effect of transforming growth factor beta(2) on resident DCs and by their suppression of T-cell-mediated immune and inflammatory responses.

Objective: To investigate the role of very late antigen 1 (VLA-1) (also known as integrin receptor alpha(1)beta(1)) in corneal transplantation inflammation and allograft survival., Methods: Cell infiltration and vasculogenesis (both angiogenesis and lymphangiogenesis) associated with allodisparate corneal transplantation were assessed in VLA-1-deficient conditions and controls by immunofluorescent microscopic studies. Corneal allograft survival was also assessed after anti-VLA-1 antibody treatment and in VLA-1 knockout recipient mice., Results: Anti-VLA-1 antibody treatment leads to a profound reduction in the granulocytic, monocytic, and T-cell infiltration after corneal transplantation. In addition, corneal angiogenesis and lymphangiogenesis were both significantly suppressed in VLA-1 knockout mice. Remarkably, universal graft survival was observed in both anti-VLA-1 antibody treatment and knockout mice., Conclusions: Very late antigen 1 blockade markedly reduces inflammation and inflammation-induced tissue responses, including vasculogenic responses, associated with corneal transplantation and promotes allograft survival., Clinical Relevance: These studies offer insights into important integrin-mediated mechanisms of corneal transplant-related inflammation and provide possible new integrin-based immunotherapies for transplant rejection.

Most current animal models of dry eye have a single causative mechanism and do not take into consideration the influence of environmental conditions on tear secretion and associated ocular surface signs. Since immunity and inflammation have been implicated in dry eye pathogenesis, and different mouse strains are known to have differentially biased immune responses, we conducted the present study to test the hypothesis that strains with specifically polarized T cell responses (T helper-1 [Th1] vs. T helper-2 [Th2]) develop differential signs of dry eye when exposed to a controlled low humidity setting. Eight to 12-week-old BALB/c (Th2 biased) and C57BL/6 (Th1 biased) mice were placed in a controlled environment chamber (CEC) where relative humidity (RH), temperature (T), and air flow (AF) were continuously regulated and monitored. Mice were exposed to specific environmental controlled conditions (RH=15.5+/-3.8%, AF=15 l/min, T=21-23 degrees C) for 3 to 7 days. Aqueous tear production by means of the cotton thread test, corneal fluorescein staining (NEI grading scheme, score 0-15) and goblet cell density in the superior and inferior conjunctivae were measured by a masked observer. No statistically significant differences between the groups were found at baseline. Statistically significant decreases in tear secretion were seen after exposure to the CEC environment. Mean cotton thread wetting was 1.9+/-0.2 (baseline), 1.4+/-0.3 (day 3), and 0.9+/-0.2 mm (day 7) for BALB/c mice, and 1.7+/-0.3 (baseline), 0.9+/-0.3 (day 3), and 0.4+/-0.2 mm (day 7) for C57BL/6 mice. These mice showed reduced tear secretion as compared to BALB/C at each time point tested (P<.005, t-test). Both BALB/c and C57BL/6 mice showed a significant increase in corneal fluorescein staining at both day 3 and day 7 as compared to baseline. With exposure to the CEC goblet cell density significantly decreased in the superior and inferior conjunctivae in BALB/c mice, while it remained unchanged in C57BL/6 mice. This study indicates that exposure of non-pharmacologically modified mice to a low humidity environment in the CEC can lead to significant alterations in tear secretion, goblet cell density, and acquisition of dry eye-related ocular surface signs which are strain-specific.

Purpose: Trafficking of corneal antigen-presenting cells (APC) to draining lymph nodes (LN) is critical in triggering immune responses. However, very little is known about the molecular regulation of this pathway. We investigated the expression and function of the chemokine receptor CCR7 in mediating corneal APC migration in inflammation., Methods: Expression of CCR7 and its ligands, CCL21 and CCL19, in the normal and inflamed corneas was analyzed by RT-PCR and immunofluorescence staining. The phenotype of CCR7-expressing cells was identified by double-staining with different cell surface markers. To trace the trafficking of APC to draining LN, we injected corneal grafts with Alexa488-conjugated ovalbumin (OVA) and transplanted to syngeneic recipients. CCR7 expression on the Alexa488-conjugated OVA+ cells in the ipsilateral draining LN was analyzed by flow cytometry. To determine the functional role of CCR7, we injected anti-CCL21 neutralizing antibody subconjunctivally after corneal transplantation and analyzed changes in numbers of OVA+ cells in the draining LN. Each experiment was repeated at least three times., Results: Both CCR7 and its ligand CCL21 were significantly upregulated in inflamed corneas as measured by RT-PCR and immunofluorescence staining. CCR7+ cells were detected especially in the corneal periphery near LYVE-1+ lymphatic vessels. CCR7+ cells were universally CD11b+CD11c+, and a majority were major histocompatibility complex class II positive, suggesting a monocytic dendritic cell lineage and a relative state of maturation. Forty-eight h after syngeneic transplantation with OVA-loaded grafts, CCR7 expression was detected on the OVA+ cells in both the host corneal beds and the draining LN. Local administration of anti-CCL21 led to a significant suppression in the flow of OVA+CD11c+ cells to the draining LN., Conclusions: These data suggest that in inflammation, APC expressing CCR7 on their cell surface interact with CCL21 to facilitate their migration from the cornea to draining LN via afferent lymphatics.

Purpose: Many corneal grafts undergo immune rejection, and current therapies are associated with many side effects. The purpose of this study was to identify critical chemokine pathways involved in generating the alloimmune response to corneal transplants., Methods: Orthotopic corneal transplantation was performed in fully mismatched strains. Cytokine and chemokine receptor gene expression was determined by the RNase protection assay. Knockout (KO) strains for chemokine-chemokine receptors that are upregulated after transplantation underwent corneal transplantation. Results derived from KO murine hosts were compared with cyclosporine (Cy) therapy. In addition to graft survival, graft infiltration, allospecific delayed-type hypersensitivity (DTH), and cytokine expression were compared among the recipient groups., Results: Initial experiments revealed gene upregulation of the chemokine receptors CCR1, -2, and -5 after corneal allorejection. Although CCR1 KO hosts showed a significant increase in graft survival compared with wild-type (WT) hosts, allografts in CCR5, CCR2/CCL3(MIP-1alpha), CXCR3, CXCL10/IP-10, and CCL3/MIP-1alpha KO mice did not show a significant improvement in graft survival. Further, CCR1 KO hosts showed a significantly higher survival rate than with systemic Cy therapy in WT hosts. Moreover, graft infiltration by leukocytes and gene expression of proinflammatory cytokines were reduced in CCR1 KO mice compared with both Cy treated and untreated WT mice, as was the induction of allospecific DTH., Conclusions: These studies provide, for the first time, evidence that targeting of specific chemokine pathways can significantly promote survival of corneal transplants, and suggest that select deletion or suppression of CCR1 can be a useful therapeutic target in corneal transplant immunity.

Over the past 20 years it has become clear that dry eye syndrome (DES) or keratoconjunctivitis sicca (KCS) is a complex multifactorial disease characterized by an immune and inflammatory process that affects the lacrimal glands and ocular surface. In this paradigm, inflammation is seen as both the cause and consequence of conjunctival and corneal cell damage. In this chapter, we identify the unique characteristics of the lacrimal gland, the role of epithelial cells, regulatory T cells, and cytokines in maintaining ocular surface homeostasis and tear secretion function. We analyze the factors inducing loss of the lacrimal gland homeostasis and its consequences, and in so doing hope to provide a picture of the role of the immune system in the pathophysiology of KCS and useful information to help understand the complexity of DES.

Autoimmune disorders of the ocular surface represent a clinically heterogeneous group of conditions where acute and chronic autoreactive mechanisms can cause significant damage to the eye. When severe and affecting the epithelium and substantia propria of the conjunctiva, cicatrization can ensue, leading to significant mechanical alterations as a result of the fibrosis. These conditions, though generally infrequent, can be the cause of profound pathology and visual disability, and often need systemic immune modulation for therapy.

Corneal antigen-presenting cells (APCs) were thought to reside exclusively in the peripheral cornea. However, recent evidence demonstrates that the central cornea is also endowed with a heterogeneous population of bone marrow-derived cells, including epithelial Langerhans cells (LCs) and anterior stromal dendritic cells (DCs), which under certain conditions can function as APCs. While the corneal periphery contains mature and immature resident bone marrow-derived DCs, the central cornea is endowed exclusively with highly immature/precursor-type DCs. During inflammation, a majority of resident DCs undergo maturation by acquiring high expression of major histocompatibility complex class II antigens and B7 (CD80/CD86) and CD40 costimulatory molecules. Further, macrophages are present in the posterior corneal stroma. In transplantation, donor-derived DCs migrate to host cervical lymph nodes and activate host T cells via the direct pathway when allografts are placed in inflamed, but not normal uninflamed, host beds. Migration of DCs to cervical lymph nodes is, in part, regulated by the vascular endothelial growth factor receptor-3 (VEGFR-3) that is expressed on corneal DCs. Blockade of the VEGFR-3 signaling significantly suppresses corneal DC trafficking to draining lymph nodes and rejection of corneal transplants. Much remains unknown about the function of these cells including their role in innate responses as well as in tolerance. Regardless, these data revise the tenet that the cornea is immune privileged due to a lack of resident lymphoreticular cells per se, but suggest that the cornea is capable of actively participating in the immune response to foreign antigens and autoantigens, rather than being a passive bystander. Additionally, one important aspect of immune privilege is likely the ocular 'imposition' of the immature phenotype on its resident bone marrow-derived cells.

Objective: To determine whether aging is associated with significant alterations in the polar and neutral lipid profiles in human meibomian gland secretions., Methods: Meibomian gland secretions were collected from both eyes of younger and older men and women. Samples were processed for high-performance liquid chromatography or mass spectrometry and for the analysis of associated spectra of fragment ions. Subjects also underwent slitlamp evaluations of the eyelid., Results: Aging is associated with numerous significant alterations in the lipid profiles of human meibomian gland secretions. Analysis of polar and neutral lipid patterns identified ions that were significantly different in secretions of younger vs older men and women, as well as ions that varied significantly only between men and women. Correlation coefficients within, but not between, groups were high. Aging was accompanied by increased opacity of meibomian gland secretions and by eyelid and eyelid margin changes., Conclusion: Aging is associated with significant sex-related alterations in the polar and neutral lipid profiles of human meibomian gland secretions., Clinical Relevance: The observed changes may contribute to the age-related increase in the prevalence of dry eye syndromes.

Background: Murine models of high-risk and low-risk corneal transplantation were used to determine the role of keratocyte apoptosis in the failure of orthotopic allogeneic corneal transplants., Materials and Methods: Normal (low-risk, low-rejecting) and inflamed/vascularized (high-risk, high-rejecting) BALB/c recipient beds received fully mismatched C57BL/6 corneal allografts. Apoptosis was detected in the corneal stroma at various time points using an in situ terminal deoxynucleotide tranferase-mediated dUTP nick-end labeling assay, and ex vivo via Western analysis for active caspase-3. Apoptosis was also measured in a (donor-type) C57BL/6 keratocyte cell line after stimulation of Fas or via use of various pro-inflammatory cytokines., Results: Significantly more apoptotic cells were present in the stroma of rapidly rejecting high-risk corneal allografts compared with low-risk grafts. Apoptotic cells were shown to be nearly uniformly CD45 and hence of a non-hematopoetic lineage. Apoptosis, however, was not present in highly inflamed but ungrafted corneas. Apoptosis was induced in keratocytes in vitro by dual stimulation with agonistic Fas mAb and either interleukin-1beta or tumor necrosis factor-alpha., Conclusion: Apoptosis of resident non-bone marrow-derived fibroblastic cells of the corneal stroma is strongly correlated with the failure of corneal allografts, particularly in the highly inflamed microenvironment of the high-risk allograft.

Background/aim: Human tears contain hundreds of proteins that may exert a significant influence on tear film stability, ocular surface integrity, and visual function. The authors hypothesise that many of these proteins originate from the meibomian gland. This study's aim was to begin to develop the proteomic methodology to permit the testing of their hypothesis., Methods: Meibomian gland secretions were collected from the lower eyelids of adult volunteers and placed in a chloroform-methanol mixture. Samples were partitioned in a biphasic system and non-lipid phase materials were reduced, alkylated, and trypsin digested to obtain peptides for protein identification. This peptide mixture was separated by micro-capillary reverse phase chromatography and the effluent examined by nano-electrospray MS and data dependent MS/MS. SEQUEST software was used to identify proteins from the MS/MS spectra., Results: The methodological approach to date has permitted the identification of more than 90 proteins in human meibomian gland secretions. Proteins include the alpha2-macroglobulin receptor, IgA alpha chain, farnesoid X activated receptor, interferon regulatory factor 3, lacritin precursor, lactotransferrin, lipocalin 1, lysozyme C precursor, potential phospholipid transporting ATPase IK, seven transmembrane helix receptor (also termed somatostatin receptor type 4), testes development related NYD-SP21 (also termed high affinity IgE receptor beta subunit), and TrkC tyrosine kinase., Conclusions: These findings indicate that the meibomian gland secretes a number of proteins into the tear film. It is quite possible that these proteins contribute to the dynamics of the tear film in both health and disease.

It is now widely acknowledged that vasculogenesis and immunogenic inflammation are intricately related in many tissues. However, less understood is to what extent blood (heme) angiogenesis (HA) and lymphangiogenesis (LG) are differentially regulated, and in turn contribute potentially to different aspects of adaptive immunity generated to corneal antigens. Herein, we will provide a brief synopsis of relevant work in this field by focusing particularly on the differential regulation of immunity by these two distinct vasculogenic processes in the cornea, illustrating that in spite of significant redundancies in the mechanisms that induce these vascular processes in the cornea, there are also important distinctions in what aspects of immunity are promoted by them.

Purpose: Lymphatic vessel endothelial hyaluronic acid receptor (LYVE-1) is a newly discovered lymphatic-specific marker. To date, there is no report of its expression on conjunctival cells. The purpose of this study was to investigate the expression of LYVE-1 in normal conjunctiva, to phenotype LYVE-1+ cells, and to study changes in their expression levels during corneal inflammation., Methods: Flat-mounted conjunctivae or cross sections of eyeballs were harvested from BALB/c mice (6-8 weeks of age) for immunofluorescent confocal microscopic studies., Results: The data demonstrate, for the first time, that in addition to its expression on lymphatic vessels, LYVE-1 was expressed on CD45+, CD11b+, and CD31- conjunctival cells, indicating a bone-marrow-derived monocytic lineage. Surprisingly, the number of cells that expressed LYVE-1 decreased during corneal inflammation, in conjunction with ingrowth of lymphatics into the cornea., Conclusions: A new population of monocytic cells has been found to express LYVE-1 in normal conjunctiva. These cells that normally express LYVE-1 may act as a reservoir for lymphangiogenesis and cell recruitment when the immune system is challenged.

Purpose: To determine the differential gene expression of chemokine species and leukocyte infiltration of grafts in the early pre-rejection postoperative period after high-risk (HR) compared to normal-risk (NR) corneal transplantation., Methods: Fully mismatched and syngeneic corneal grafts were performed in NR (avascular) and HR (vascularized) recipient beds of BALB/c murine hosts. Gene expression levels of a panel of chemokines were determined by a multiprobe ribonuclease protection assay system. The profiles of infiltrating cells into the corneal grafts at the same times were determined immunohistochemically., Results: Compared to NR transplantation, HR eyes exhibited significantly higher mRNA levels for macrophage inflammatory protein (MIP)-2 and monocyte chemotactic protein-1 (MCP-1) on day 1, and for eotaxin on days 1 and 3 after transplantation. By day 6 after transplantation, still well before graft rejection, significantly higher levels of RANTES, eotaxin, MIP-1alpha, MIP-1beta, MIP-2, and MCP-1 were detected in HR eyes. The overexpression of MIPs in HR eyes correlated with a significant increase in the number of infiltrating macrophages (p<0.01), and neutrophils (p<0.05) in HR compared to NR recipients. Low levels of eosinophil and mast cell infiltration were observed in all grafts, with a modest increase in mast cell infiltration (p<0.05) in HR compared to NR grafts., Conclusions: These results suggest that increased expression of gene products for select chemokines, in particular those that mediate recruitment of innate immune cells, in the early period after HR corneal transplantation is related to the enhanced leukocytic infiltration of grafts observed in HR keratoplasty.

Purpose: To develop a controlled-environment chamber (CEC) for mice and verify the effects of a low-humidity setting on ocular surface signs in normal mice., Methods: Eight- to 12-week-old BALB/c mice were used in a controlled-environment chamber (CEC) where relative humidity (RH), temperature (T), and airflow (AF) are regulated and monitored. Mice were placed into the CEC and exposed to specific environmentally controlled conditions (RH = 18.5% +/- 5.1%, AF = 15 L/min, T = 21-23 degrees C) for 3, 7, 14, and 28 days. Control mice were kept in a normal environment (RH = 50%-80%, no AF, T = 21-23 degrees C) for the same duration. Aqueous tear production by means of the cotton thread test, corneal fluorescein staining (score, 0-15), and goblet cell density in the superior and inferior conjunctiva were measured by a masked observer., Results: No statistically significant differences between the groups were found at baseline. Decreased tear secretion and increased corneal fluorescein staining were significantly present on day 3, 7, 14, and 28 in animals kept in the CEC. Goblet cell density was significantly decreased in the superior conjunctiva on day 7, and on day 3, 7, and 14 in the inferior conjunctiva in the CEC-kept mice compared with control animals., Conclusions: This study indicates that exposure of normal mice to a low-humidity environment in a CEC can lead to significant alterations in tear secretion, goblet cell density, and acquisition of dry eye-related ocular surface signs.

Lymphangiogenesis, the formation of new lymphatic vessels, is important for tumor metastasis and induction of immunity to peripheral antigens including organ transplants. We herein describe a novel mouse model of spontaneous, secondary lymphangiogenesis in the normally avascular cornea. corn1 mice, which suffer from a deletion in the gene encoding the cytoskeletal protein destrin, develop hemangiogenesis as well as spontaneous outgrowth of LYVE-1+++/CD31+ lymphatic vessels into the cornea starting at age 4 weeks. Corneal lymphangiogenesis is delayed in onset, is less intense, and regresses earlier compared with hemangiogenesis. Moreover, the lymphangiogenesis is preceded only by a mild recruitment of CD45+ inflammatory cells into the cornea. In contrast to mice with inflammation-induced hem- and lymphangiogenesis, corn1 mice do not develop breakdown of the blood-aqueous barrier. Finally, in this novel mouse model, a blocking anti-VEGFR3 antibody significantly inhibited not only lymph- but also hemangiogenesis. In summary, destrin deletion has differential effects on spontaneous hem- and lymphangiogenesis in the normally avascular cornea and represents a novel mouse model to study the mechanisms of lymphangiogenesis and to test the antihem- and antilymphangiogenic properties of known or new antiangiogenic agents.

Purpose: To report acute unilateral hypopyon uveitis as an initial presenting feature of relapsing acute lymphoblastic leukemia (ALL) in an adult patient., Design: Observational case report., Methods: Anterior chamber paracentesis was performed in a 56-year-old male presenting with treatment-resistant unilateral hypopyon while in the remission phase of ALL., Results: Examination of the aqueous humor aspirate revealed presence of malignant cells compatible with the previous bone marrow biopsy and subsequent spinal tap results., Conclusions: Atypical hypopyon uveitis can be an indication of relapsing ALL, even in adults. Prompt anterior chamber aspiration is required for the correct diagnosis and subsequent treatment.

Purpose: To characterize the chemokines and chemokine receptors that mediate the effect of proinflammatory cytokines, interleukin (IL)-1 and tumor necrosis factor (TNF)-alpha, on the recruitment of MHC class II(+) Langerhans cells (LCs) in the corneal epithelium., Methods: A standard model for corneal LC recruitment, application of cautery to the central corneal surface was used, and the differential gene expression levels of a panel of chemokines and chemokine receptors were determined by RNase protection assay. Chemokine receptor-knockout mice were used to evaluate the recruitment of MHC class II(+) LCs to the corneal epithelium. To determine the sensitivity of selected chemokines to IL-1 and TNF-alpha stimulation, the chemokine gene expression pattern was analyzed after blockade of IL-1 and TNF receptors., Results: CCR1, -2, and -5 were overexpressed in corneas after cauterization. Topical administration of soluble TNF receptor I and IL-1 receptor antagonist, which abrogated corneal LC recruitment, significantly suppressed the gene transcription levels of the ligands of CCR1 and/or -5, regulated on activation normal T-cell expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1alpha, and MIP-1beta. The recruitment of major histocompatibility complex (MHC) class II(+) LC was significantly suppressed in CCR5(-/-) mice and blockade of RANTES and MIP-1beta, but not in CCR1(-/-), CCR2(-/-)/MIP-1alpha(-/-), or MIP-1alpha(-/-) mice. The evaluation of epithelial CD11c(+) LC cells by confocal microscopy revealed coexpression for CCR5 primarily among B7(-) (CD80(-)/CD86(-)) subsets of these LCs but not among the mature B7(+) subsets of CD11c(+) LCs., Conclusions: These data suggest that CCR5 plays a critical role in mediating recruitment and mobilization of MHC class II(+) LCs into the corneal epithelium. Targeting CCR5 and its ligands may be a new strategy for modulating immunity.

Context: Low-level lead exposure may increase the risk for a number of chronic age-related diseases. Several studies have documented the presence of lead in lenses with cataract. The intrusion of lead into the lens may alter lens redox status and cause protein conformational changes that decrease lens transparency., Objective: To determine the relationship of cumulative lead exposure with the development of cataract., Design, Setting, and Participants: Tibial (cortical) and patellar (trabecular) bone lead levels were measured by K x-ray fluorescence between 1991 and 1999 in a subset of participants in the Normative Aging Study (NAS), a Boston-based longitudinal study of aging in men. Among the first 795 NAS participants to have bone lead levels measured, we reviewed eye examination data (collected routinely every 3-5 years) for the period after the bone lead measurements were taken. We limited the population to men aged 60 years and older who had sufficient eye examination information available (n = 642). Blood lead levels were also measured., Main Outcome Measures: Cataract assessment was done while masked to the lead level results. A participant was considered to have cataract if there was documentation for either eye of cataract surgery or a cataract graded clinically as 3+ or higher on a 4-point scale. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated as estimates of the magnitude and significance of the relationship of lead exposure with cataract, in logistic regression models., Results: The mean age of the study participants was 69 years and cataract was identified in 122 men. The age-adjusted OR (95% CI) for cataract for men in the highest vs lowest quintile of tibia lead level was 2.68 (1.31-5.50). Further adjustment for pack-years of cigarette smoking, diabetes, blood lead levels, and intake of vitamin C, vitamin E, and carotenoids resulted in an OR of 3.19 (95% CI, 1.48-6.90). For patella lead level, there was an increased risk of cataract in the highest vs lowest quintile (OR, 1.88; 95% CI, 0.88-4.02), but the trend was not significant (P = .16). Blood lead levels, more indicative of short-term exposure levels, were not significantly associated with cataract (OR, 0.89; 95% CI, 0.46-1.72; P = .73)., Conclusions: These epidemiological data suggest that accumulated lead exposure, such as that commonly experienced by adults in the United States, may be an important unrecognized risk factor for cataract. This research suggests that reduction of lead exposure could help decrease the global burden of cataract.

Many ocular surface tests have been developed for the clinical diagnosis of keratoconjunctivitis sicca, and several have been used to 'validate' animal models of Sjogren's and non-Sjogren's dry eye. However, many of these tests themselves have not been systematically studied or standardized, and yet their use in animal models of dry eye is common. This review provides a rational approach and systematic review of the tear film and ocular surface tests described in the literature that may be applicable to the current animal models of dry eye, with particular emphasis on their limitations, along with some suggestions regarding their standardization and applications in eye research.

Vascular endothelial growth factor-3 (VEGFR-3), also known as Fms-like tyrosine kinase receptor 4 (FLT-4), was thought to be expressed exclusively on the lymphatic endothelium, high endothelial venules, and rarely on vascular endothelium. It plays a critical role in the development of lymphatics and cancer metastasis. Very recently, however, VEGFR-3 expression has been identified on dendritic cells (DCs) in the inflamed cornea, and related to the trafficking of these cells to lymphoid organs. The current study was performed to evaluate the expression of VEGFR-3 in the conjunctiva. The conjunctiva and limbus of normal and inflamed murine eyes were excised and stained for VEGFR-3. Immunofluorescence double staining for CD11b, CD11c, CD31, CD45, GR-1, CD3, CD80, LYVE-1 and class II major histocompatibility complex (MHC) antigen expression, using confocal microscopy, was performed to further phenotype the VEGFR-3+ cells. VEGFR-3 and LYVE-1 expression was observed on lymphatic, but not blood vessel, endothelium. In addition, we also detected expression of VEGFR-3 on non-endothelial CD45+ bone marrow-derived cells in the conjunctiva of normal and, in an increased number, in inflamed eyes. These cells were uniformly CD11b+, CD3-, and Gr-1-, suggesting a monocytic origin, similar to the VEGFR-3+ cells in the cornea. Nearly half of the VEGFR-3+ cells were also positive for MHC class II expression, and none were positive for CD80 (B7-1), indicating their relative immature status. In contrast to the recently described VEGFR3+ corneal cells, however, VEGFR-3+ conjunctival cells did not express the DC marker CD11c. We conclude that in addition to its known role in lymphangiogenesis, VEGFR-3 is also expressed by a conjunctival monocyte/macrophage lineage, implicating a potential relationship between lymphangiogenesis and leukocyte trafficking in the ocular surface.

Corneal grafts were until recently considered entirely devoid of resident APCs, giving rise to the tenet that alloantigen recognition is mediated exclusively by the indirect (host APC-dependent) pathway. The recent discovery of a resident myeloid corneal dendritic cell population that is normally MHC class II(-) but can readily up-regulate class II expression during inflammation led us to hypothesize that under certain conditions the direct pathway of allosensitization becomes operative. To test this, corneal allotransplants were performed in either inflamed (high-risk (HR)) or uninflamed (low-risk (LR)) host beds in mice, and the frequencies of host T cells activated via the direct pathway were determined. We found that directly primed CD4(+) T cells were detected in the HR but not LR setting, and these cells displayed a clear Th1 phenotype by 2 wk after grafting. Moreover, the use of MHC class II knockout donor tissue led to significantly enhanced survival of HR but not LR allografts. Finally, we show that donor corneal APC demonstrate high expression of CD40, CD80, and CD86 costimulatory molecules when derived from HR but not LR grafts. These data are the first to report that a functional donor APC-dependent direct response is elicited in corneal transplant hosts when the graft bed is inflamed and underscore the relevance of the graft microenvironment in dictating the pathway of allosensitization.