OCULAR IMMUNONOLOGY

1. OCULAR IMMUNONOLOGY Class 12 Dr. Pittler

2. OVERVIEW: • Essential review of molecular & cellular events • The compartmentalization of immunoglobulins • Ocular immune privilege & its functions • The 1st line of defense in the cornea • Progressive ocular surface disease • Intraocular infection and the loss of immune • privilege. • Complement reactions. • Inflammatory processes. • Graves’ disease – autoimmunity.

3. ESSENTIAL REVIEW: The immunoglobulins (Igs) Immunoglobulins are 1st responder molecules for the immune system. By binding to antigens they “identify” the substance as foreign. They initiate immunoreaction mechanisms and they reinforce (= amplify) further immune reactions. Igs that are important on the ocular surface are: IgG IgA (secretory) IgM phagocyte rapid mover limited 1st responder (attraction) (tear film) (complement activation)

4. THIS TABLE SHOWS THE MAJOR Igs AND THER LOCATIONS: NOTE: ● Only IgA (sec); IgG and IgM are found in the tear film. ● That only IgA (sec) and IgG are found in the aqueous ● that no Igs at all are found in the lens (nor in any other part of the eye#) ● these values represent the immunologically quiescent eye (nothing is going on – no infections, inflammation or disease processes). #except within the blood vessels

5. OTHER INNATE IMMUNOLOGICAL MOLECULES IN TEAR FILM: LYSOZYME – lyses peptidoglycans of Gram+ bacteria. LACTOFERRIN – the name comes from the Latin words for milk and iron. Essentially, lactoferrin ties up iron atoms and keeps keeping them from being used by iron-requiring bacteria: including both Gm+ and and Gm- as well as fungi. The protein weighs 80 kDa. LIPOCALIN – a peptide in tears whose role in immuno-chemistry is not understood. IRON BINDING SITES

6. CELLS THAT CONTRIBUTE TO ACQUIRED IMMUNE DEFENSE IN THE EYE (LACRIMAL GLAND) The upper image shows a plasma cell (green) and a Th cell (yellow). The lower image shows a Tc cells (blue) attacking and killing an infected cell. All three cells types are normally present in the lacrimal gland and are released to anterior segment tissues on demand (i. e., an immunological threat). Plasma cells are B cells that are terminally differentiated to manufacture antibodies. Helper T cells have a variety of functions involved in reinforcing the development and activities of plasma cells, mast cells, antigen presenting cells, macrophages and killer cells. Killer cells (cytotoxic killer cells NOT NK cells) will attack infected cells and cause their destruction. HELPER CELL PLASMA CELL KILLER CELL

7. THE CONCEPT OF IMMUNE PRIVILEGE AND “ACAID” In order to understand how cells and molecules work together in the eye’s immune system, it also becomes necessary to understand the immune privilege of the eye. Basically, immune privilege is an adaptation of the body’s immune system to suppress to some degree the normal immune response that occurs in the body. No one completely understands how it works, but it is a system that helps the eye avoid massive reactions (especially in the cornea) that would result in a disastrous loss of ocular tissue. It also makes possible corneal transplants without significant rejection of the tissue. The system allows normally antigenic cells and tissues to be tolerated in the anterior chamber and has been termed anterior chamber-associated immune deviation (ACAID).

8. FLOW OF IMMUNE TRAFFIC WITHOUT IMMUNE PRIVILEGE NEW Abs Afferent pathways ( ) bring antigenic particles to the lymph node ( ) where an APC sensitizes Th cells. These cells move to the spleen ( ) and from there to the ocular MALT (mucosal associated lymphoid tissue) ( ). Here and in the lacrimal gland, plasma cells produce antibodies to respond to antigenic stimuli ( )

9. FLOW OF IMMUNE TRAFFIC WITH IMMUNE PRIVILEGE NEW Abs↓ Cellular and aqueous origin of some immunosuppressants: TGFb, MSH, and VIP.

10. FLOW OF IMMUNE TRAFFIC WITH IMMUNE PRIVILEGE NEW Abs↓ These cytokines migrate to plasma cells and inhibit their production of antobodies.

11. The cytokines, such as TGFb, all operate by the same basic, hormone-like mechanism. They bind to a receptor protein and transform their signal by means of a series of protein bindings (cascade) to a protein that binds to DNA. The “signal” communicates to the specific DNA directed RNA polymerase whether to upregulate or downregulate the rate of protein synthesis (via mRNA). This signal eventually gets to plasma cells concerning the synthesis of antibodies. It demonstrates how immune privilege works in the cornea. TRANSFORMING GROWTH FACTOR b Receptor protein DNA binding protein Cascade proteins Inhibition of Ab protein synthesis

12. WHAT HAPPENS WHEN IMMUNE DEFENSES MUST OVERCOME PRIVILEGE? A point is reached when an invading organism can no longer be tolerated and the ocular surface is threatened. IgA (sec), IgG and IgM are the 1st defense mechanisms to respond. IgA (sec) is always present in sufficient concentrations to begin the response. However, it is IgM that may often be the 1st responder as can be seen by the table below: Here the concentration of IgG1 is increased by ~11% and IgG4 by ~700% in an active herpes infection.

13. As a herpes virus infection progresses, IgM and IgA (sec) contribute to the defense of the tissue. IgM, in particular, may begin a complement reaction which is usually limited since IgM is such as large molecule. However, in the tissue erosions caused by the damaging virus, the molecule is able to penetrate to the erosions themselves and bring the complement reaction to completion. On the right are shown the typical dendritic lesions of a herpes infection (arrow). IgG levels can also be considerably more elevated in other infections. For example, in acute hemorrhagic conjunctivitis, the levels of IgG can reach 1300 mg/ mL.

14. OCULAR COMPLEMENT Whether complement is fixed by IgG or IgM is irrelevant to the large molecular weight of the initial C1q complex (410 kD). That also limits its activity in the corneal stroma. Although there is evidence for the presence of complement components in the aqueous, the sequence must still be initiated by an immuno- globulin in the classical pathway. In addition to the destruction of accessable herpes virus, complement fixation can also facilitate the removal of Staphylococcus aureus (in conjunction with lysozyme activity) [Gm+ ];Neisseria gonorrhoea; and Haemophilus influenzae [both Gm- ]. It must be remembered that complement causes two effects in its immune activities: the initiation of inflammation and membrane destruction of antigenic cells. Complement seems to be more involved in pore perforation of antigenic cells in the cornea.

15. OCULAR INFLAMMATION Ocular inflammation is not a desirable process in ocular tissues due to the real possibility of tissue destruction mechanisms that include: phagocytosis and the invasion and growth of blood vessels. That is the reason that the eye is immunoprivileged unless an immune attack overwhelms immune suppression. The picture on the right shows a phago- cyte with pseudopodia reaching out to engulf bacterial rods. The process unfortunately causes the production of highly reactive forms of oxygen (e. g., O2 –[superoxide] and HOCl [hypochlorous acid “chlorox”] that not only destroy offending micro- organisms, but also breakdown essential collagen fibers in the corneal.

16. GRAVES’ DISEASE AND AUTOIMMUNITY – OCULAR EFFECTS Graves’ disease is initially an autoimmune pathology that begins at the thyroid gland. The thyroid gland (shown on the right) controls basal metabolism by its secretion of T3 and T4 hormones. In the disease (generally in the 3rd or 4th decade of life) individuals begin to experience an increase in body temperature; become hyperactive; have increased GI activity as well as weight loss and increased appetite. There are also frequent ocular effects (called “ophthalmopathy”) in which the eyes become pushed forward in their sockets.

17. The reason for the initial effects at the thyroid gland are related to an invasion of lymphocytes that are responding to a perceived “antigenic” presence in the gland. These lymphocytes synthesize a type of antibody known as TSI (thyroid stimulating immunoglobulin). TSI mimics the ability of TSH (thryoid stimulating hormone) to bind to the TSH receptor on thyroid follicular cells. The Result of this is an Increase in: the uptake of iodine Into the cell, an Increase in cell volume and an increase in the release of T3 and T4 into the body.

18. This is a typical Graves’ disease patient with ocular effects that is indicated by the wide-eye stare (upper arrow) due to eyes that have been pushed forward due to the engorgement of surrounding fat and muscle tissue. The conditions include: periorbital and lid edema; lid retraction; restriction of muscle movement; proptosis of the eyes and possible vsual acuity/ field defects. Although some of these characteristics can be attributed to an increase in circulating T3 and T4, some cannot: i. e., the engorgement of fat and muscle tissue.

19. The picture shows the appearance of fat (green arrow) and muscle tissues (red arrow) in normal and Graves’ disease patients’ eyes at autopsy. Note the engorgement of muscles in the periocular tissues in the Graves’ disease patient. The engorgement of muscles is thought to be due to a separate autoimmune reaction at these tissues and not to increased T3 and T4.

20. STUDY GUIDE FOR OCULAR IMMUNOCHEMISTRY • What are the Ig’s that are commonly found in the anterior segment and what are their roles. • What is lactoferrin and how does it work? • How does immune privilege (ACAID) work in the cornea? • What is the basic mechanism by which immunosuppressant cytokines work? • If IgM and C1q are impeded from entering the corneal stroma – how is it possible for complement to attack herpes virus? • What are the destructive processes in corneal inflammation? • What is the mechanism in the body by which Graves’ disease operates? How does the disease start? • What is the possible immune mechanism in the eye by which ophthalmopathy is produced in Graves’ disease?