Table 1. Demographics and clinical features of uveitis patients, and controls.

Table 2. Clinical and immunological features of uveitis patients.

Table 3. Clinical evolution with the methilprednisolone use and anti TB or anti-toxoplasma drugs.

Figure 1. Specific IgG to retinal S protein in autoimmune and infectious uveitis types. X2; p<0.0001...

Figure 2. Specific IgE to retinal S protein in autoimmune and infectious uveitis: X2; p=0.0002.

Figure 3. A. SLPI total levels uveitis group vs control. F test of variance; p=0.0424.

Figure 3. B. SLPI discriminated levels in uveitis group vs control. F test of variance; p=0.0424.

Figure 4. SLPI in levels in infectious uveitis, expressed in pg/ml , pre y post treatment with CS. F...

Figure 5. SLPI levels in autoimmune uveitis, separated in not responder (n=8), partially responder (...

Figure 6. Meprednisone responses in different kind of uveitis. X2; p=0.0365

Introduction

Human secretory leukocyte inhibitor (SLPI) is an 11.7 kDa cationic protein and a member of innate immunity–associated proteins, with significant antiprotease and anti-inflammatory activities1. It is a nonglycosylated highly basic, acid–stable, cysteine-rich, 107 amino acid, single chain poly – peptide. The SLPI gene, along with the elafin gene, is a member of the trapping gene family1,2. Products of this family are characterized by an N–terminal transglutaminase domain substrate and C–terminal four – disulfide core. These two domains (COOH terminal and NH 2 terminal) share about 35% of homology. Each of these domains has distinct enzyme activities. The tertiary structure of the SLPI molecule resembles a boomerang, with each arm carrying one domain. The four in each domain disulfide bridges formed between the cysteine residues, as well as the two-domain interaction, contribute to the conformation and efficacy of the molecule1-5).

The human SLPI gene is localized in chromosome 20q 12-13.2. The SLPI gene consists of four exons and three introns and spanding approximately 2.6 kb1,2. Up to date no polymorphism of the SLPI gene and no state of SLPI deficiency has been found. SLPI has multiple important roles both in normal homoeostasis and at sites of inflammation, and appear to modulate on multiple pathological conditions1,3,5. SLPI has been shown to exhibit an anti-inflammatory property, including down regulation of TNFα and inhibition of NFκβ (6).

Historically SLPI was first isolated from secretions of patients with COPD, cystic fibrosis and experimental asthma3,4,6-10. It has been found to be a potent inhibitor of human leukocyte elastase, human cathepsine G, and human trypsin. Both the anti elastase and anti trypsin activities are mediated by C-terminal domain. The N-terminal domain has no known function. SLPI is produced by neutrophils, macrophages, β-cells of pancreatic islets, epithelial cells investing the renal tubules, acinar cells of parotid and submaxillary glands, and epithelial cells lining mucous membranes of respiratory and alimentary tracts. SLPI was originally isolated from parotid saliva and has been detected in a variety secretion, as whole saliva, seminal fluid, cervical mucus, synovial fluid, breast milk, tears, and cerebral spinal fluid, as in secretions from the nose and bronchi1-13. The SLPI gene was found to be expressed and correlated with poor prognosis in lung, breast, oropharyngeal, bladder, endometrial, ovarian, and colon rectal carcinomas1. SLPI is found in neurons and astrocytes in the ischemic brain tissue13, and SLPI has also been found to play a pivotal role in apoptosis and wound healing1,14-16. Furthermore, various studies have indicated that the SLPI gene is constitutively expressed in epithelial cells and that this expression can be increased by pro-inflammatory stimuli, such as tumor necrosis factor (TNF) and IL–1b7,11. These findings support the notion that epithelial cells respond to inflammatory stimuli by increasing their anti-protease shield, which may minimize proteolytic damage4,5,11. However, it is largely unknown which molecule is essential in mounting the innate immune response, or have the kinetics of these processes been studied5,12.

SLPI has also been shown to inhibit mast cell chymase, a protease released during mast cell degranulation, and inhibit histamine release from mast cell in vitro and in vivo1,17. SLPI has been described in the aqueous humor of patients with cataract and other eye pathologies18. It is known that the eye contains a large number of mast cells, and the choroid has one of the highest densities of mast cells of any tissue in the body18-22. SLPI may play an important role in neutralizing mast cell-released inflammatory mediators in the eye17,18,22.

In uveitis it is important to differentiate between diseases of an autoimmune or presumed autoimmune origin and those clearly due to an infectious process, such as toxoplasmosis, tuberculosis or a common bacterial infection. Although most cases of intraocular inflammatory disease are not produced by active infection, a systemic immunologic abnormality may be present19-21. To the best of our knowledge, scarce information exists in relationship between SLPI and different types of uveitis. The emerging role of SLPI in uveitis, as well as the functional and physical properties, suggests that this molecule may become an important new mechanism in the regulation and control of ocular inflammation.

In the present study, we determined the occurrence of serum SLPI levels in sera from patients with autoimmune and infectious uveitis defined by clinical and immunological features, and we evaluated them as a possible marker for disease prognosis in relationship with corticosteroid use.

Material and methods

Patients

We studied 50 patients with uveitis, 18 males and 32 females (aged 38±18.4, ranged 7–77 years). The diagnosis of uveitis was established by the clinical and immunological criteria as previously described19-21. According to the etiology of uveitis19,20, the patients were classified into the following groups: a) patients with infectious uveitis (n=16; 2 males and 14 females), and b) patients with autoimmune uveitis (n=34; 16 males and 18 females). Infectious uveitis (IU) was separated by the etiology in: 1) common bacterial uveitis (CBU) (n=7), 2) tuberculosis uveitis (TBU) (n=5), 3) toxoplasmic uveitis (Tx U) (n=4). The AIU patients (n=34) were treated for 3 months with methylprednisolone in decreased immunosuppressive doses from 120 mg/day to 10 mg in alternating days. In case of poor response to this treatment, patients were included in tacrolimus protocol from 5 mg/day to 1 mg/day for at least 1 year of treatment. Serum samples were collected from patients with uveitis and from age-matched healthy volunteer individuals (control group) (n=16), and stored in aliquots at –40°C, until required.

The patients with uveitis of different etiologies were controlled on their clinical evolution and divided in subgroups of severe relapsing uveitis patients with poor visual acuity and evolution, and patients with benign evolution and only slight impairment of vision.

The studies were approved by the Institutional Review Board and The Ethical Committee of “Fundación Ver” and comply with the tenets of the Declaration of Helsinki.

Reactives

Preparation of retinal S antigen

Retinal S antigen was prepared from fresh bovine eyes according to the method of Dorey et al.23 This method involved extraction with hypotonic buffer followed by 50% ammonium sulfate precipitation, gel filtration and affinity chromatography.

Human SLPI Immunoassay Quantikine was purchased in R&D System (USA).

Enzyme – Linked Immunosorbent Assay

Quantitation of Specific IgG to Retinal S Antigen: This was assessed in sera of all uveitis patients and normal controls by ELISA20,21,24. Briefly, ELISA plates were coated with retinal S antigen (1 g/100 l) in carbonate-bicarbonate buffer (pH=9.6) overnight at 4°C. The unbound proteins were removed by washing five times with PBS – 0.1% Tween 20. Wells were then blocked for 1 hr with 4% BSA/PBS at 37°C and incubated for 30 min with 1:100 dilutions in 1% BSA / PBS of uveitis and control serum samples. Plates were washed as described previously and incubated with 1:1000 dilution of anti-human IgG peroxidase conjugated (Sigma) for 30 min. Again, the plates were washed as described previously and bound secondary antibody was allowed to react with the substrate, o-phenylenediamine (Sigma) for 30 min. The reaction was stopped by adding 4NSO4H 2 and the optical density was determined in a microplate reader at 492 nm. A sample was considered positive if the optical density was two or more standard deviation above the mean of the normal control subjects. The sample result was obtained by the mean OD of triplicate points; for each sample these should agree to within 10%. The assays were performed three times: positive and negative results appeared in the same – patient serum samples with and intraassay CV of 9%; the interassay CV for the first to the second assay was 3.7 %, and that for the first vs the third assay was 5.5 %.

Quantitation of Specific IgE to Retinal S Antigen. This was assessed in sera of all uveitis patients and normal controls by ELISA (20, 21, 24). Briefly, ELISA plates were coated with retinal S antigen (1 μg/100 μl) in carbonate-bicarbonate buffer (pH=9.6) overnight at 4°C. The unbound proteins were removed by washing five times with PBS – 0.1% Tween 20. Wells were then blocked for 1 hr with 1% BSA/PBS at room temperature and incubated for 2 hr at room temperature with 1:4 dilutions in 1% BSA / PBS of uveitis and normal serum samples. Plates were washed five times with PBS – 0.1% Tween 20 and then incubated at room temperature with 1:1000 dilution of anti-human IgE alkaline phosphatase conjugated (Sigma) for 2 hr. Again, the plates were washed as described previously and bound secondary antibody was allowed to react with 100

l/ well of the substrate, p-nitrophenylphosphate (Sigma) for 30 min. The reaction was stopped by adding 3 M NaOH (100 μl/well) and the optical density was determined in a microplate reader at 405 nm. A sample was considered positive if the optical density was two or more standard deviation above the mean of the normal control subjects. The assays were performed three times: a positive or a negative result appeared in thee same – patient serum samples with an intraassay CV of 5%; the interassay CV for the first vs the second determination was 4.8% and the first vs the third determination was 7.9%.

SLPI assay - sample preparation

Bring all reagents and samples to room temperature before use. All samples and standards are assayed in duplicate. The determination was performed according to the manufacturer’s instructions.

All serum samples require at least a 20 fold dilution in to calibrator diluent RD5A.

SLPI was measured by an enzyme linked immunosorbent assay (ELISA). Assay plates (96 well) were coated with 0.025 µg/ml recombinant SLPI (R&D Systems) in PBS and 1% 400 mmol/L carbonate buffer; 100 µl were added to each well. Plates were left 60 min at room temperature. Blocking was carried out with 400 μl /well blocking/protecting solution (polyvinilpyrrolidone 2%, BSA 5 mg/ml, preservatives: EDTA 5 mmol/L, Tris 50 mmol/L for 30 min. Plates were washed with wash buffer (150 mmol/L NaCl 100 mmol/L Tris, 0.05% Tween -20; pH: 7-7.5), 150 μl of standard sample and 50 μl of anti-SLPI; 2 μg /ml; diluted in ELISA buffer (150 mmol/L NaCl, 100 mmol/L Tris, 50 mmol/L phenol red solution, 2 mmol/L EDTA, 1 mmol/L 2-methylisothiazolone), 1 mmol/L bromonitrodioxane, 2 mg/ml BSA, 0.05% Tween 20; (pH=7.2) were added to each well. A non-specific binding well (200 µl buffer only) and two B0 wells (150 µl buffer; 50 µl anti-SLPI) we included on each plate. Standards were added in triplicate and their concentration range was from 5000 – 9.8 pg/ml. Incubation was performed in a plate shaker at room temperature for 2 h; and the plates were then washed. One hundred μl/well of peroxidase – labeled anti-sheep/goat immunoglobulin G (IgG), Fab fragment raised in donkey (1:1000 of stock in ELISA buffer) were added. Incubation was on shaker as before and the plates were washed again. Two hundred μl substrate, (0.3 g/L urea – hydrogen peroxide, 0.1 g/L tetramethyl benzidine in 100 mmol/L sodium acetate; pH=6) were added to each well. After 2-10 min, the reaction was stopped by adding 50 μl/well 2 N SO4H2. The optical density was determined in a microplate reader at 450 nm. The minimum detectable dose of SLPI is typically less 25 pg/ml.

Statistical analysis

Statistical significance was performed by ANOVA and Chi square and Fisher’s exact tests, using GrafPad and SPSS software´s, version 10 . We considered p< 0.05 statistically significant.

Results

The clinical and immunological features of uveitis patients are detailed in Tables 1 / 2 / 3.

The autoimmune uveitis group was divided by clinical evolution after treatment with methylprednisolone and tacrolimus in: a) active autoinmune uveitis (AAIU) (n= 22) and inactive autoinmune uveitis (IAIU) (n=12). The infectious uveitis group (n=16) was divided in common bacterial uveitis (CBU) (n=7); tuberculosis uveitis (n=5) and toxoplasmic uveitis (n=4).

Both groups presented severe relapsing uveitis patients with poor evolution (n=16) and b) patients with benign evolution (n=34) (Table 3).

The active autoimmune uveitis presented poor evolution in 12 of 16 cases in group of severe relapsing uveitis patients, and the four remaining cases were two cases from the toxoplasmic subgroup, one case from bacterial uveitis, and one of the TB uveitis, all with poor outcome (Table 3).

The specific IgG to retinal S antigen was negative in all cases of TBU, and positive in 1 of 7 CBU, the Tx U was positive in 4 of 4 cases. The specific IgG for retinal S antigen was positive in all cases of AAIU as well as in one case of IAIU. The normal control group was negative for specific IgG to retinal S protein in all of cases (p< 0.0001) (Figure 1).

The specific IgE to retinal S antigen was negative in TBU and CBU and positive in 3 of 4 cases of Tx U and 18 of 22 patients with AAIU as well as 7 of 12 IAIU. The normal control group was negative for specific IgE to retinal S antigen in all cases (p< 0.0001) (Figure 2).

The specific IgG to retinal S protein overlapped with specific IgE to that antigen in 75 % of autoimmune uveitis patients.

The total SLPI in both uveitis( Infectious and autoimmune uveitis) vs control was in uveitis 372.5±25.97 pg/ml and control group 346±24 pg/ml; with F test to compare variance, p value =0.0424 (Figures 3A and 3B).

The SLPI levels was in Infectious uveitis BIU Pre-Cs treatment 288.6±26.63 pg/ml, and post-CS was 286.3±9.551 pg/ml, p=NS (Figure 4).

The SLPI determination in autoimmune group pre-and post-CS treatment was separated by the responses in AIU no responder group (n=8), SLPI level was in the basal 350.8±±29.5 pg/ml and post-CS 337.8± 29.5 pg ml; partial response group to CS (n=9) basal 393.8± 27.25 pg/ml, post-CS 399.3± 30.7pg/ml; and the group with improvement (n=17) the basal was 450.5±17.8 pg/ml, post=CS 504±18,62 pg/ml, p< 0.0001 (Figures 5 and 6).

Discussion

Uveitis is a generic term that encompasses a variety of intraocular inflammatory responses to infectious or autoimmune origin18-21,25-28.

Despite considerable progress in elucidating the immunopathogenesis of these ocular disorders, there is still scarce information about reliable immunologic markers of disease evolution. In this context, no significant differences in the levels of specific IgG and IgE to retinal S protein were found in our patients when compared with previous studies20,21. In agreement with these studies, the specific IgG and IgE to retinal S antigen was positive in autoimmune uveitis as well as in toxoplasmic uveitis. These specific IgG and IgE to retinal S antigen probably represent CD4+ Th2 lymphocyte activation20,21,28.

Some authors believe that the evidence for autoimmunity to retinal antigens expressed in both autoantibodies above described, are sufficient markers to define autoimmune ocular disease19-21,27.

In the other infectious ocular condition, autoantibodies against retinal S antigen, were found to be negative. In this context, we have previously shown the importance of specific IgG and IgE autoantibodies to retinal S antigen in autoimmune and infectious uveitis20, as well as more recently the presence of circulating anti-galectin antibodies in similar eye conditions21.

The innate immune system is a complex and dynamic system that continuously adapts itself to the changing environment to prevent infections and certainly has gained renewed interest over the past two decades1,10,15,29-36.

The image of innate immunity as a more primitive response has evolved towards a system that interfaces with adaptive immune response. The fact remains that innate inflammatory effectors mechanisms through proteases still have considerable potential to damage host tissues1,14,16,29,35. In response to these enzymes, systemic anti-proteases such as α1-PI, α1-antichimotrypsin, and α2-macroglobulin are synthesized in the liver and reach tissues through the blood stream1,4,11,31,32. On the other hand, alarm anti-proteases like SLPI and elafin, are produced locally in the site of inflammation in response to interleukin-1 and TNF1,5,12,18. The balance between proteases and their inhibitors may play a modulating roll in many biological processes. SLPI as a potent protease inhibitor with anti-inflammatory and antimicrobial activities, has been found to be present normally in the normal human tear fluid, in the cornea12,16,18, and sera13,37, and to be synthesized by the human ciliary epithelium38 and in the aqueous humor in different eye pathologies18. Probably the main function of SLPI is to protect local tissue against the detrimental consequence of inflammation1,34,35.

It is known that the eye contains a large number of mast cells, and the choroid has one of the highest densities of mast cells of any tissue in the body18-20,22. Human mast cells have pro-inflammatory selective mediators located in the granules and released upon degranulation. Histamine has been widely used as a marker of mast cell degranulation and increased level of histamine has been observed in a number of diseases including: chronic urticaria, ischemic heart disease patients ongoing coronaroangiography and allergic diseases such as asthma, rhinitis, otitis media, indicating that mediator is involved in the pathogenesis of these diseases1,3,4,7,9,13,17,21,39,40. Numerous anti-allergic drugs such as di-sodiun cromoglicate or anti-histamines were reported to inhibit in vitro of anti IgE induced histamine release from human mast cells17,22,40. In recent years, inhibition of tryptase and chymase by SLPI was considered as a novel class of natural mast cell stabilizer. SLPI inhibit IgE dependent release from human mast cells and enzymatic activity of chymase, suggesting, that it may play a protective role in autoimmune uveitis18,20,40. The natural chymase inhibitor SLPI probably inhibits the histamine release from various sources of human mast cells, and contributes to local regulatory mechanism of these cell degranulation both physiologically or pathophysiologically17,22,40. On the other hand, a plethora of toxic inflammatory products, i.e., serine proteinase, are released from stimulated leukocytes during the uveitic inflammatory process and subsequent degradation of tissue ensues18.

In the present study we evaluated the SLPI serum levels in different uveitis conditions. SLPI was elevated in active autoimmune uveitis, when compared with inactive autoimmune, common bacterial and TB uveitis. Toxoplasmic uveitis responded similarly to active autoimmune uveitis and probably represented the rupture of the blood –retinal barrier accompanying the uveitic inflammation with release of retinal S antigen, as have been previously described19-21.

The SLPI level was increased in patients with active autoimmune uveitis, probably reflecting the local and sys The SLPI level was increased in patients with active autoimmune uveitis, probably reflecting the local and sys temic anti-inflammatory response in this group of patients18-21. Toxoplasmic retinochoroiditis, however, behaves differently from other infectious uveitis presenting similar SLPI levels compared with active autoimmune uveitis. These findings may be explained by retinal tissue disruption and SLPI released by local mast and systemic inflammatory cells. This uveitis subgroup also presented severe impairment of vision.

In the case of infectious uveitis secondary to tuberculosis, the SLPI level was similar to control group and different to active autoimmune uveitis. In the first step the macrophages respond to Mycobacterium tuberculosis by regulating expression of gene products that initiate a host innate response to this micro-organism and elevated SLPI. In the second part, the presence of these particular antigen metabolites of mycobacterium tuberculosis has more relevance, because they stimulate predominantly proliferation of CD4+ Th1 lymphocyte profile. The CD4+ Th1 lymphocyte response produced high levels of IL-12 and INFγ.

The common bacterial endophtalmitis presented normal SLPI serum level. The INFγ, produced inhibition of SLPI expression and low SLPI serum levels and strong delayed hypersensitivity reaction and subsequent granuloma formation20,41.

This feature was comparable to those of the control group, inactive autoimmune uveitis and TB uveitis. Infectious uveitis has been ascribed to Th1 and CD8+ cytotoxic lymphocytes pathways19-21,28. The more important function of SLPI is to protect local tissue against the detrimental consequences of inflammation1,36,37,42. The SLPI has a broad-spectrum antibiotic activity that includes bactericidal and antifungal properties1,30,35,42. The local increase of SLPI in patients found with uveitis18 is different to our feature in serum; this difference probably is explained by locally up-regulation in response to the local infection similar to uterine epithelial cells in pre and post menopausal women. The post menopausal women decreased SLPI bactericidal activity, and in our bacterial endophtalmitis group the majority of patients were localized in post menopausal status1,35.

The common bacterial uveitis presented benign evolution with antibiotic treatment and only slight impairment of vision in most cases, with only one case becoming blind.

The inactive autoimmune uveitis patients presented benign evolution to Methylprednisolone treatment. In this sub-group the SLPI levels was similar to the control group and the clinical activity of uveitis was improved dramatically with the treatment. In the active autoimmune uveitis as well as toxoplasmic uveitis the clinical evolution was poor. These different forms of responses to treatment with corticosteroids probably reflected the activation of glucocorticoid receptor inducing transcription of genes such as SLPI. At lower concentrations glucocorticoid reduce inflammatory gene induced by NFκβ or AP-1 via association between these factors6,8,43,44. SLPI uptake into the inflammatory cells was rapid and widespread. The protein is not simply bound to the cell membrane. It is internalized efficiently as evidenced by high level into the cells cytoplasmic and nuclear fractions. SLPI could bind DNA directly because it does by its cationic charge. Due to SLPI´s ability to inhibit LPS induced NFκβ, and new information that SLPI can localize to the nucleus and bind DNA, Taggart et al. considered the possibility that SLPI might bind to NFκβ sites5,8,39,43-45. SLPI interaction with other consensus transcription factor binding has been also assessed. Although there was no binding of SLPI-GATA, Sp1, CREB consensus oligonucleotides, strong binding of SLPI-AP1 probe has been observed indicating that SLPI may also bind sites other than NFκβ. The SLPI bind to the IL-8 and TNFα promoters; which contains multiple NFκβ. They do not interact with IL-10 promoters, nor contain NFκβ sites. SLPI decreased LPS induced TNF α and IL-8 productions but not IL-106,43-45. The ability of SLPI to enter the nucleus is particularly interesting due to the cationic nature of this protein and its potential to bind negatively charged regions of DNA. Taggart et al. demonstrated that SLPI can bind DNA and NFκβ sites in specific manner. SLPI present degradation on IK βα and β, and subsequently inhibits NFκβ activity6,43. The patients that responded poorly to corticosteroid treatment were included in a tacrolimus protocol. This immunosuppressive drug an immunophilin-binding agent, without important complications, and partially improve in 60% of the active autoimmune uveitis.

The increased frequency of SLPI in sera of patients with intraocular disorders might be a direct consequence of the release of tissue-associated SLPI during the inflammatory response, from the mast cells of the choroid, as well as from the ciliary epithelium18-22,46.

Concluding remarks:

Furthermore, we have demonstrated in the present study that serum SLPI levels correlated with worse evolution of ocular inflammatory disease in patients with active autoimmune uveitis. The presence of these autoantibodies, as well as the high serum SLPI levels, suggests their potential use for the prognosis of inflammatory ocular diseases. The elevated SLPI in patients with active autoimmune uveitis reflect the presence of a double inflammatory process, expressed locally as well as systemic. Probably, SLPI may be considered as a marker of disease activity in auto-immune as well as in toxoplasmic uveitis.

1. Doumas S; Kolokotronis A; Stefanopoulos P: Anti-Inflammatory and Antimicrobial Roles of Secretory Leukocyte Protease Inhibitor. Infection and Immunity 2005; 1271 – 4.

2. Stetler G; Brewer MT; Thompson RC: Isolation and sequence of a human gene encoding a potent inhibitor of leukocyte proteases. Nucleic Acids Res 1986; 14:7883 – 96.

3. Wright CD, Havill AM, Middleton SC, Kashem MA, Lee PA, Dripps DJ, O´ Riordan, Bevilacqua MP, Abraham W M. Secretory Leukocyte Protease Inhibitor Prevents Allergen-Induced Pulmonary Responses in Animal Models of Asthma. J Pharmacol Exp Ther 1999; 289: 1007 – 14.

4. Vogelhmeier CA; Gillissen CA; Buhl R: Use of Secretary Leukoprotease Inhibitor to Augment Lung Antineutrophil Elastase Activity. Chest 1996; 110:261 – 6.

5. Williams SE; Brown A; Sallenave J-M: SLPI and Elafin: one glove, many fingers. Clin Sci (London) 2006; 110:21 – 35.

6. Taggart CC, Cryan SA, Weldon S, Gibbons A, Greene CM, Kelly E, Low TB, O´ Neill SJO, McElvaney NG. Secretory leucoprotease inhibitor binds to NF κß binding sites in monocytes and inhibits p65 binding. JEM 2005; 202: 1659 – 8.

7. Sallenave, J.M., Shulmann, J., Crossley, J., Jordana, M., Gauldie, J. Regulation of secretory leukocyte inhibitor (SLPI) and elastase-specific inhibitor (ESI/Elafin) in human airway epithelial cells by cytokines and neutrophilic enzymes. Am J Respir Cell Mol Biol 1994; 11:733 – 41.

8. Usmani OS, Ito K, Maneechotesuwan K, Ito M, Johnson M, Barnes PJ, Adcock IM. Glucocorticoid Receptor Nuclear Translocation in Airway Cells after Inhaled Combination Therapy. Am J Repir Crit Care Med 2005; 172: 704 – 12.

9. Hollander C, Nyström M, Janciauskiene S, Westin U. Human mast cells decrease SLPI levels in type II – like alveolar cell model, in vitro. Cancer Cell Inter 2003; 3: 14 – 22.

10. Bingle L, Cross SS, High AS, Wallace WA, Rassl D, Yuan G, Hellstrom I, Campos MA, Bingle CD. WFDC2 (HE4): A potential role in the innate immunity of the oral cavity and respiratory tract and development of adenocarcinomas of the lung. Resp Research 2006; 7 : 61 – 70

11. Van Wetering S, Van Der Linden AC, Van Sterkenburg MAJA, De Boer WI, Kuijpers ALA, Schalkwijk J, Hiemstra PS. Regulation of SLPI and elafin release from bronchial epithelial cells by neutrophils defensins. Am J Physiol Lung Cell Mol Physiol 2000; 278: L 51 – L 58.

12. Sathe S, Sakata M, Beaton AR, Sack RA. Identification, origins and the diurnal role of the principal serine protease inhibitors in human tear fluid. Curr Eye Res 1998; 17:348 – 62.

13. Wang X; Li X; Xu L, Zhan Y, Yaish – Ohad S, Erhardt JA, Barone FC, Feuerstein GZ: Up-Regulation of Secretory Leukocyte Protease Inhibitor (SLPI) in the Brain after Ischemic Stroke: Adenoviral Expression of SLPI Protects Brain from Ischemic Injury. Mol Pharmacol 2003; 64:833 – 40.

14. Ashcroft GS; Lei K; Jin W, Longenecker G, Kulkarni AB, Greenwell-Wild T, Hale-Donze H, McGrady G, Song XY, Wahl SM: Secretory leukocyte protease inhibitor mediates non-redundant functions necessary for normal wound healing. Nat Med 2000; 6: 1147 – 53.

15. Odaka C; Mizuochi T; Yang J; Ding A: Murine Macrophages Produce Secretory Leukocyte Protease Inhibitor During Clearance of Apoptotic Cells: Implications for Resolution of the Inflammatory Response. J Immunol 2003; 171:1507 – 14.

16. Sorensen OE; Cowland JB; Theilgaard-Mönch K; Liu L; Ganz T, Borregaard N: Wound Healing and Expression of Antimicrobial Peptides/Polypeptides in Human Keratinocytes, a Consequence of Common Growth Factors. J Immunol 2003; 170:5583 – 9.

17. HE S-H, Chen P, Chen HQ: Modulation of enzymatic activity of human mast cell tryptase and chymase by protease inhibitors. Acta Pharmacol Sin 2003; 24: 923 – 9.

18. Janciauskiene S, Brandt L, Wallmark A, Westin U; Krakau T: Secreted leukocyte protease inhibitor is present in aqueous humours from cataract and other eye pathologies. Exp Eye Research 2006; 82: 505 – 11.

19. Nussenblatt RB, Palestine AG: Concepts of disease pathogenesis. In Uveitis, Fundamentals and Clinical Practice. 1st ed. Chicago, Year Book Medical 1989, pp 21 – 52.

20. Muiño JC, Juárez CP, Luna JD, Castro CC, Wolf EG, Ferrero M, Romero-Piffiguer MD. The Importance of Specific IgG and IgE Autoantibodies to Retinal S Antigen, Total Serum IgE, and sCD23 Levels in Autoimmune and Infectious Uveitis. J. Clin Immunol 1999; 19: 215 – 22.

21. Romero MD, Muiño JC, Bianco GA, Ferrero M, Juárez CP, Luna JD, Rabinovich GA. Circulating Anti-galectin-1 antibodies Are Associated with the Severity of Ocular Disease in Autoimmune and Infectious uveitis. IOVS 2006; 47: 1550 – 6.

22. Irani AM: Ocular Mast Cells and Mediators: Immunol Allergy Clin North Am 1997; 17: 1 – 18

23. Dorey O, Cozette J, Faure JO: A simple rapid method for isolation of retinal S antigen. Ophtalmol Rev 1982; 14: 249 – 55.

24. Nieto A, Carbonetto CA. Enzimoinmuno-ensayo. In Inmunología e Inmunoquímica, Margni RA (Ed). Buenos Aires, Panamericana, 1989; pp. 571 – 86.

25. Rosenbaum JT, Becker MT, Smith JR: Immunologic Ocular disease. In Clinical Immunology, Principles and Practice. Rich RR, Fleisher TA, Shearer WT, Kotzin BL, Schroeder HW (Eds). Mosby, London 2001, pp 86.1 – 86.12.

26. Forester JV, Menamin PC. Immunopathogenic Mechanisms in Intraocular Inflammation. In Immune Response and the Eye. Streilein JW, (Ed). Chemical Immunology. Basel, Karger, 1999; 73: pp 159 – 85.

27. Nussenblatt RB, Gery I, Weiner HL, Ferris FL, Shiloach J, Remaley N, Perry C, Caspi RR, Haffler DA, Foster C, and Whitcup SM. Treatment of uveitis by oral administration of retinal antigens: Results of a phase I/II randomized masked trial. Am J Ophtalmol 1997; 123: 583 – 92.

28. de Smet MD, Dayan M, Nussenblatt RB. A novel method for the determination of T – cell proliferative responses in patients with uveitis. Ocular Immunol Inflamm 1998; 6: 139 – 207.

29. Abbas AK, Litchman AH. Innate Immunity – The Early Defense against Infections in Basic Immunology. Saunders, Philadelphia 2006; pp 21 – 39.

30. Vos JB, van Sterkenburg MA, Rabe KF, Schalkwijk J, Hiemstra PS, Datson N. Transcriptional response of bronchial epithelial cells to Pseudomonas aeruginosa: identification of early mediators of host defense. Physiol Genomics 2005; 21: 324 – 36.

31. Sánchez MC, Barcelona PF, Luna JD, Ortiz SG, Juárez PC, Riera CM, Chiabrando GA. Low – density lipoprotein receptor – related protein – 1 (LRP – 1) in rat model of oxygen induced retinal neovascularization. Exp Eye Research, 2006 in press. Available on line at www. Science direct. com

32. Piitulainen E, Carlson J, Ohlsson K, Sverger T. α1 – antitrypsin Deficiency in 26 Year – Old Subjects. Lung, Liver and Protease/ Protease Inhibitor Studies. Chest 2005; 128: 2076 – 81.

33. Muiño JC, Castro CC, Gagliardi JR, Wolff EG, Gómez RM, Gugliemone H, Alé AM, Ferrero M, Romero MD. Regulación de la respuesta inflamatoria de la aspergilosis alérgica broncopulmonar por antígenos Clase III del complejo mayor de histocompatibilidad (C4, C2, FB) CD 23 soluble y fallas de la Fagocitosis. Rev Med Córdoba 1997; 85: 2 – 12.

34. Jana NK, Gray LR, Shugars DC. Human Immunodeficiency Virus Type 1 Stimulates the Expression and Production of Secretory Leukocyte Protease Inhibitor (SLPI) in Oral Epithelial Cell: role for SLPI in Innate Mucosal Immunity. J Virol 2005; 79: 6432 – 40.

35. King AE, Critchley HOD, Kelly RW. Presence of secretory leukocyte protease inhibitor in human endometrium and first trimester decidua suggest an antibacterial protective role. Mol Hum Rerprod 2000; 6: 191 – 6.

36. Greene CM, McElvaney NG, O´ Neill SJ, Taggart CC. Secretory Leucoprotease Inhibitor Impairs Toll-Like Receptor 2 and 4 Mediated Responses in Monocytic Cells. Infection and Immunity 2004; 72: 3684 – 7.

37. Grobmyer SR, Barie PS, Nathan CF, Fuortes M, Lin E, Lowry SF, Wright CD, Weyant MJ, Hydo L, Reeves F, Shiloh Mu, Ding A: Secretory leukocyte protease inhibitor, an inhibitor of neutrophil activation, is elevated in serum in human sepsis and experimental endotoxemia. Crit Care Med 2000; 28: 1276 – 82.

38. Ortego J, Escribano J, Coca-Prados M. Gene expression of proteases and protease inhibitors in the human ciliary epithelium and ODM-2 cells. Exp. Eye Res. 1997; 65:289 – 99.

39. Henrisksen PA, Hitt M, Xing Z, Wang J, Haslett C, Riesmersma RA, Webb DJ, Kotelevtsev YV, Sallenave JM. Adenoviral Gene Delivery of Elafin and Secretory Leukocyte Protease Inhibitor Attenuates NF-κß – Dependent Inflammatory Responses of Human Endothelial Cells and Macrophages to Atherogenic Stimuli. J Immunol 2004; 172: 4535 – 44.

40. He S, Xie H, Zhang X-J, Wang S-J : Inhibition of histamine release from human mast cells by natural chymase inhibitors. Acta Pharmacol Sin 2004; 25: 822 – 6.

41. Ding A, Yang J, Shi S, Ehrt S. Inductions of macrophage-derived SLPI by Mycobacterium tuberculosis depends on TLR 2 but not MyD88. Immunology 2005; 116 – 381 – 9.

42. da Silva MCA, Zahm JM, Gras D, Bajolet O, Abely M, Hinnrasky J, Milliot M, de Assis MC, Hologne C, Bonnet N, Merten M, Plotkowski MC, Puchele E. Dynamic interaction between airway epithelial cells and Staphylococcus aureus. Am J Physiol Lung Cell Mol Physiol 2004; 287: L 542 – L 551.

43. Butler MW, Robertson I, Greene CM, O´ Neill SJO, Taggart CC, McElvaney NG. Elafin prevents lipopolysaccharide-induced APQ and NF-κß activation via and effect on the ubiquitin-proteasome pathway. JBC Papers in press. Published on September 15, 2006. Http//www.jbc.org/cgi/doi/10.1074/jbc/M604844200

44. Ito K, Yamamura S, Essilfie-Quaye S, Cosio B, Ito M, Barnes PJ, Adcock IM, Histone deacetylase 2 – mediated deacetylation of the glucocorticoid receptor enables NF-κß suppression. JEM 2005; 203: 7 – 13.

45. Taggart CC, Greene CM, Smith SG, Levine RL, McCray PB, O´ Neill SO, Mc Elvaney NG. Inactivation of Human ß-Defensins 2 and 3 by Elastolytic Cathepsins. J Immunol 2003; 170 931 – 7

46. T Novak, F Fortune, L Bergmeier, I Khan, E Hagi-Pavli. Neutrophil elastase and endogenous inhibitors in Behçet’s disease saliva. Clin Exp Immunol 2020 Oct;202(1):93-105. doi: 0.1111/cei.13483. Epub 2020 Jul 29

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