Evaluation of Accessory Lacrimal Gland in Muller’s Muscle Conjunctival Resection Specimens for Precursor Cell Markers and Biological Markers of Dry Eye Disease

Abstract

Purpose The accessory lacrimal glands (ALG) are an understudied component of the tear functional unit, even though they are important in the development of dry eye syndrome (DES). To advance our understanding of aging changes, regenerative potential and histologic correlates to human characteristics, we investigated human ALG tissue from surgical samples to determine the presence or absence of progenitor cell markers and lacrimal epithelial markers and to correlate marker expression to relevant patient characteristics. Materials and Methods ALG tissues obtained from Muller’s Muscle Conjunctival Resection (MMCR) specimens were created using tissue microarrays (TMAs). Immunofluorescence staining of MMCR sections was performed using primary antibodies specific to cell protein markers. Cell marker localization in TMAs was then assessed by two blinded observers using a standardized scoring system. Patient characteristics including age, race, and status of ocular surface health were then compared against expression of stem cell markers. Results Human ALG expressed a number of epithelial markers, and in particular, histatin-1 was well correlated with the expression of epithelial markers and was present in most acini. In addition, we noted the presence of precursor cell markers nestin, ABCG2 and CD90 in ALG tissue. There was a decrease in precursor cell marker expression with increasing age. Finally, we noted that a negative association was present between histatin-1 expression and DES. Conclusions Thus, we report for the first time that human ALG tissues contain precursor marker positive cells and that this marker expression may decrease with increasing age. Moreover, histatin-1 expression may be decreased in DES. Future studies will be performed to use these cell markers to isolate and culture lacrimal epithelial cells from heterogeneous tissues, determine the relevance of histatin-1 expression to DES and isolate candidate precursor cells from ALG tissue.

Presence of Histatin-1 in Human Tears and Association with Aqueous Deficient Dry Eye Diagnosis: A Preliminary Study

“…The main findings of this study are that H1 is present in human tears and levels of H1 are significantly reduced in patients with ADDE. This study represents the first robust demonstration of H1 in human tears, building upon initial findings amongst multiple groups 14,15,17,22 .…”

“…In this study, patients were considered to have ADDE if their Schirmer I at 5 minutes was less than 10 mm of wetting, vital dye staining of the cornea ≥1 on the National Eye Institute-Corneal Staining Score (NEI-CSS), and an OSDI score greater than 13 20,21 . We focused primarily on H1 as this is one of the most studied in saliva, has been demonstrated in the lacrimal functional unit, and could be an interesting tear component to replace due to its epithelial trophic functions 17,22 .…”

Histatins, wound healing, and cell migration

“…These proteins are synthesized and secreted by the serous acini of the parotid, submandibular, sublingual, and von Ebner glands (Ahmad, Piludu, Oppenheim, Helmerhorst, & Hand, 2004;Sabatini & Azen, 1989;Schenkels, Veerman, & Nieuw Amerongen, 1995;vanderSpek et al, 1989;Takano, Malamud, Bennick, Oppenheim, & Hand, 1993). Intriguingly, recent reports have detected the expression of these peptides in other tissues, for instance, histatin-1 in human lacrimal gland (Ali et al, 2017;Shah et al, 2016), melanoma cell lines, and metastatic lesions of patients diagnosed with melanoma (Yaguchi & Kawakami, 2012), and histatin-5 in human ocular surface (Steele, Jumblatt, Smith, & Pierce, 2002).…”

Human Lacrimal Gland Gene Expression

“…The development of the lacrimal gland and branching organogenesis has been the topic of multiple studies of late [8, 11, 19]. These studies in addition to our own gene expression studies which have shown the presence of stem cell markers in adult lacrimal tissue are important to developing regenerative technologies for lacrimal gland atrophy, disease and hypofunction.…”