Ross W. Lambert

Androgens and Dry Eye in Sjögren's Syndromea

ABSTRACT: Sjögrens syndrome is an extremely complex and currently incurable autoimmune disorder, which occurs primarily in females, and is associated with lacrimal gland inflammation, meibomian gland dysfunction, and severe dry eye. We hypothesize that androgen deficiency, which reportedly occurs in primary and secondary Sjögrens syndrome (e.g., systemic lupus erythematosus, rheumatoid arthritis), is a critical etiologic factor in the pathogenesis of dry eye syndromes. We further hypothesize that androgen treatment to the ocular surface will promote both lacrimal and meibomian gland function and alleviate both “aqueous‐deficient” and “evaporative” dry eye. Our results demonstrate that androgens regulate both lacrimal and meibomian gland function, and suggest that topical androgen administration may serve as a safe and effective therapy for the treatment of dry eye in Sjögrens syndrome.

Influence of gender and the endocrine environment on the distribution of androgen receptors in the lacrimal gland

Androgens are known to regulate both the structure and function of lacrimal tissue in a variety of species. To explore the endocrine basis for this hormone action, the following study was designed to: (1) determine the cellular distribution of androgen receptors in the lacrimal gland; and (2) examine the influence of gender and the endocrine environment on the glandular content of these binding sites. Lacrimal glands were obtained from intact, castrated, hypophysectomized, diabetic or sham-operated male or female adult rats, mice or hamsters, as well as from orchiectomized rats exposed to placebo compounds or physiological levels of testosterone. The cellular location of androgen receptors was evaluated by utilizing an immunoperoxidase protocol, in which a purified rabbit polyclonal antibody to the rat androgen receptor was used as the first antibody. Our findings with lacrimal glands showed that: (1) androgen receptors are located almost exclusively in nuclei of epithelial cells; (2) the cellular distribution or intranuclear density of these binding sites is far more extensive in glands of males, as compared to females; (3) orchiectomy or hypophysectomy, but not sham-surgery or diabetes, lead to a dramatic reduction in the immunocytochemical expression of androgen receptors; and (4) testosterone administration to orchiectomized rats induces a marked increase in androgen receptor content, relative to that in placebo-exposed glands. Our results also reveal that a 10 kb androgen receptor mRNA exists in the rat lacrimal gland. Overall, these findings demonstrate that gender and the endocrine system may significantly influence the distribution of androgen binding sites in rat lacrimal tissue. Moreover, our results show that androgens up-regulate their own lacrimal gland receptors.

Androgen control of secretory component mRNA levels in the rat lacrimal gland.

The purpose of this investigation was to determine whether the known gender-related differences in, and the endocrine control of, the production of secretory component (SC) by the rat lacrimal gland are associated with alterations in SC mRNA content. Levels of SC mRNA were measured in lacrimal tissues of intact, sham-operated, castrated, hypophysectomized, and testosterone-treated male and female adult rats by Northern blot procedures, which utilized a specific, [alpha-32P]-labelled rat SC cDNA probe. For control purposes, SC mRNA amounts were standardized to the beta-actin content in experimental blots. The location of SC mRNA in lacrimal glands was evaluated by in situ hybridization techniques, which involved exposure of tissue sections to sense or anti-sense [35S]-labelled SC RNA probes. Our results demonstrate that: (1) lacrimal glands of male rats contain a significantly greater amount of SC mRNA than those of female rats, and that this difference co-exists with distinct, gender-associated variations in the distribution of SC mRNA in lacrimal tissue; (2) orchiectomy or hypophysectomy, but not ovariectomy or sham surgery, leads to a marked decline in the lacrimal SC mRNA content; and (3) testosterone, but not placebo, administration to castrated male and female rats induces a significant increase in the SC mRNA levels in lacrimal tissue. Overall, these findings show that gender, androgens and the hypothalamic-pituitary axis exert a considerable influence on the SC mRNA content in the rat lacrimal gland.

Effect of sialodacryoadenitis virus exposure on acinar epithelial cells from the rat lacrimal gland

Sialodacryoadenitis virus (SDAV), a RNA coronavirus, induces degenerative, necrotic and atrophic alterations in acinar epithelial cells of the rat lacrimal gland. To begin to explore the underlying mechanism(s) of this viral effect, we sought in the present study to: (1) determine whether SDAV invades and replicates in lacrimal gland acinar cells in vitro and (2) assess whether short-term SDAV challenge interferes with the viability or function of acinar cells in vitro. For comparison we also evaluated the relative infectivity of SDAV in acinar epithelial cells from lacrimal, submandibular and parotid glands, given that salivary tissues are known to be highly susceptible to SDAV infection in vivo. Acinar epithelial cells from lacrimal, submandibular or parotid glands were isolated from male rats, exposed briefly to SDAV or control cell antigen and then cultured for four, eight or twelve days. At experimental termination, SDAV titers in both media and sonicated cell extracts were evaluated by plaque assay titration on mouse L2 cell monolayers. To evaluate functional aspects of lacrimal gland acinar cells, SDAV-infected cells were incubated in the presence or absence of dihydrotestosterone and culture media were analyzed by RIA to measure the extent of the androgen-induced increase in secretory component (SC) production. Our results showed that: (1) SDAV invades and replicates in lacrimal gland acinar cells, Viral challenge resulted in a significant, time-dependent increase in SDAV titers, that were primarily cell-associated and greatly exceeded amounts contained in the original inoculum; (2) SDAV infection did not compromise lacrimal acinar cell viability or prevent the cellular SC response to androgens. Viral presence, though, did often attenuate the magnitude of this hormone action; and (3) SDAV infects salivary acinar cells, but the kinetics and magnitude or viral replication in lacrimal, submandibular and parotid cells showed considerable variations. These findings demonstrate that SDAV invades and replicates in acinar epithelial cells from lacrimal and salivary glands. The resulting release of infectious progeny may play a role in the SDAV-induced pathology of exocrine tissues in vivo.

Neural-Endocrine Control of Secretory Component Synthesis by Lacrimal Gland Acinar Cells: Specificity, Temporal Characteristics and Molecular Basis

The ocular surface appears to be protected from bacterial and viral pathogens by polymeric IgA antibodies.1 These antibodies, which are produced by plasma cells in the lacrimal gland, bind to secretory component (SC) in the basolateral membrane of acinar epithelial cells, and are then transported to the apical membrane and secreted into tears.2 Of interest, the lacrimal secretion of sIgA, as well as free SC, appears to be significantly influenced by gender and hormones from the hypothalamic-pituitary-gonadal axis.2 Thus, almost 10 years ago, it was found that the concentrations of free SC and IgA in the tears of male rats were 2 to 5-fold higher than those in tears of female rats. These gender-associated differences in tear SC and IgA were shown to be caused by androgens.2 For example, castration led to a significant reduction in SC levels in tears of males, while having no impact on SC content in tears of females. In addition, administration of testosterone for 4 days to castrated male or female rats significantly increased tear SC levels. These studies showed that exposure of castrated rats to androgens resulted in alterations of lacrimal SC production.2 Moreover, later experiments demonstrated that this androgen control of SC, as well as IgA, is modulated by factors from the hypothalamus and pituitary.2 However, this research did not address whether androgens act directly on lacrimal tissue to change SC or IgA, or indirectly via an androgen-sensitive site, which in tum acts to modulate lacrimal SC and IgA.

Sialodacryoadenitis virus infection of rat lacrimal gland acinar cells.

The secretory immune system of the eye is designed to protect the ocular surface against microbial challenge and infectious disease.1 This immunological role is mediated primarily through secretory IgA (sIgA) antibodies, which are produced by plasma cells in interstitial areas of the lacrimal gland and are selectively transported to tears by secretory component (SC), the polymeric TgA receptor.1 After delivery to the eye’s anterior surface, sIgA antibodies may act to prevent viral internalization, inhibit bacterial colonization, curtail parasitic infestation and attenuate toxin-related damage.2

Neural, Endocrine and Immune Regulation of Secretory Component Production by Lacrimal Gland Acinar Cells

Our previous research has shown that the endocrine, nervous and immune systems regulate the production of secretory component (SC), the polymeric immunoglobulin receptor, by acinar (epithelial) cells from the rat lacrimal gland.1 Thus, acinar cell exposure in vitro to androgens (e. g., dihydrotestosterone [DHT]), vasoactive intestinal peptide (VIP), the adrenergic agonist, isoproterenol, cyclic AMP analogues (e. g., 8-bromoadenosine 3’:5’- cyclic monophosphate [bcAMP]), cyclic AMP inducers (e. g., cholera toxin, PGE2), phosphodiesterase inhibitors (e. g., 3-isobutyl-l-methylxanthine), IL-lα, IL-lβ, or TNF-α results in a significant increase in SC output.2-4 Conversely, cellular treatment with the cholinergic agent, carbachol, causes a significant suppression of both androgen-induced and basal SC release by acinar cells.3,4

Influence of the Endocrine Environment on Herpes Virus Infection in Rat Lacrimal Gland Acinar Cells

Throughout the world, one of the greatest single causes of keratoconjuctivitis sicca is Sjogren’s syndrome:1 this disease is an exceedingly complex autoimmune disorder, which occurs almost exclusively in females, and involves a progressive, immune-mediated destruction of lacrimal tissue and consequent dry eye.2 The etiology of Sjogren’s sydrome is unclear, although recent research has suggested that viral infection and hormone action may play significant roles in the pathogenesis and/or expression of this autoimmune disease.3,4 Thus, primary infection by herpes viruses (e.g. Epstein-Barr virus, cytomegalovirus, herpes virus-6), or retroviruses, may elicit the lymphocytic accumulation, immune cell activation and associated inflammation evident in affected lacrimal tissues.3,4 Moreover, sex steroids may: [a] influence the onset and/or severity of lacrimal gland immunopathology in Sjogren’s syndrome;4 and [b] potentially alter viral activity in infected lacrimal tissues, given that such hormones are known to modulate herpes virus transcription.5,6

Androgen Regulation of Ocular Mucosal- and Auto-Immunity

At present, it is known that androgens exert a tremendous impact on the development, expression and function of the immune system in both health and disease. In brief, these hormones have been shown to: [a] influence the maturation, proliferation and/or activity of pluripotent stem cells, B cells, T cells and macrophages; [b] modulate the synthesis, secretion and/or action of antibodies, cytokines and growth factors; [c] alter the production of autoantibodies and the formation of immune complexes; [d] affect the immune response to, and clearance of, antigens; [e] control the generation of thymic factors and secretory component (SC), the IgA antibody receptor; and [f] modify the rejection of allografts, the extent of graft vs. host disease, the magnitude of inflammation, and the onset and severity of autoimmune disease.e.g.1–6 The precise nature of these androgen actions is exceedingly dependent upon the specific target cell, the local microenvironment, and the particular immunological process, and may result in enhancement, inhibition, or no effect, on immune expression.3 Nevertheless, it is quite apparent that androgens control multiple aspects of humoral, cell-mediated, mucosal and auto-immunity.