Pterygium is a multifaceted pathology that displays apparent conflicting characteristics: benign (e.g., self-limiting and superficial), bad (e.g., proliferative and potentially recurrent) and ugly (e.g., signs of preneoplastic transformation). The natural successive question is: why are we lacking reports showing that pterygium lesions become life-threatening through metastasis, especially since pterygium has considerable similarities with UV-related malignancies on the molecular level? In this review, we consider how our pathophysiological understanding of the benign pterygium pathology overlaps with ocular surface squamous neoplasia and skin cancer. The three UV-related disorders share the same initial insult (i.e., UV radiation) and responsive repair mechanisms to the ensuing (in)direct DNA damage. Their downstream apoptotic regulators and other cellular adaptations are remarkably alike. However, a complicating factor in understanding the fine line between the self-limiting nature of pterygium and the malignant transformation in other UV-related diseases is the prominent ambiguity in the pathological evaluation of pterygium biopsies. Features of preneoplastic transformation (i.e., dysplasia) are used to define normal cellular reactions (i.e., atypia and metaplasia) and vice versa. A uniform grading system could help in unraveling the true nature of this ancient disease and potentially help in identifying the earliest intervention point possible regarding the cellular switch that drives a cell’s fate towards cancer.

Pterygium is a common eye disease, linked to an increased exposure to UV radiation and dry environments. The associated pathology culminates in visual impairment and, in some rare cases, blindness. However, there remains a lot of uncertainty concerning the pathogenesis of this fibrovascular lesion. As the composition of the tear film provides a reflection into the pathological changes at the ocular surface, tear analysis represents an ideal approach to gain insight in the progression of disease following pterygiectomy. This study enrolled 19 patients and age/gender-matched healthy controls. Tear film levels of interleukin- (IL-) 6, IL-8, and vascular endothelial growth factor (VEGF) were investigated over time, and preoperative concentrations were linked to corneal neovascularization and pterygium size. Diminished tear film levels were found in unilateral patients who show no clinical signs of pterygium recurrence over a period of one year. Hence, our results highlight the potential of using the course of IL-6, IL-8, and VEGF levels in tears as biomarkers for recovery. In addition, when focusing on the affected eyes (i.e., primary and recurrent pterygium), we detected fold changes in preoperative cytokine concentrations to correspond with disease severity. As our proposed biomarkers did not reveal a linear relationship with corneal neovascularization nor the invasive behaviour of pterygium, no exact role in the pterygium pathology could be established. Hence, our data point to these factors being contributors rather than decisive players in the pathological processes.

Neuroglobin is a heme protein of which increased levels provide neuroprotection against amyloid proteinopathy and hemorrhagic damage. These cellular stressors involve the promotion of ferroptosis—an iron-dependent, lipid peroxide-accreting form of cell death. Hence, we questioned whether neuroglobin could oppose ferroptosis initiation. We detected human neuroglobin (hNgb)-EGFP-expressing SH-SY5Y cells to be significantly more resistant to ferroptosis induction, identifying 0.68-fold less cell death. To elucidate the underlying pathways, this study investigated hNgb-protein interactions with a Co-IP-MS/MS approach both under a physiological and a ferroptotic condition. hNgb binds to proteins of the cellular iron metabolism (e.g., RPL15 and PCBP3) in an unstressed condition and shows an elevated binding ratio towards cell death-linked proteins, such as HNRNPA3, FAM120A, and ABRAXAS2, under ferroptotic stress. Our data also reveal a constitutive interaction between hNgb and the longevity-associated heterodimer XRCC5/XRCC6. Disentangling the involvement of hNgb and its binding partners in cellular processes, using Ingenuity Pathway Analysis, resulted in the integration of hNgb in the ubiquitination pathway, mTOR signaling, 14-3-3-mediated signaling, and the glycolysis cascade. We also detected a previously unknown strong link with motor neuropathies. Hence, this study contributes to the elucidation of neuroglobin’s involvement in cellular mechanisms that provide neuroprotection and the upkeep of homeostasis.

The introduction of tissue engineering has allowed scientists to push the boundaries and treat seriously damaged ocular surface epithelia. They have managed to do this through the development of biological substitutes that restore, maintain or improve tissue function. To ensure the generation of a therapeutically safe and effective graft, knowledge on the transcriptional profile of native and cultured ocular surface epithelia is of undeniable value. Gene expression studies are, however, only as reliable as their proper selection of internal reaction controls or reference genes. In this study, we determined the expression stability of a number of reference genes: 18s rRNA, ACTB, ATP5B, CyC1, EIF4A2, GAPDH, RPL13A, SDHA, TOP1, UBC, and YWHAZ in primary isolates as well as in ex vivo cultured ocular surface epithelia explants (day 0 and/or day 14). Expression stability of the reference genes was assessed with both the geNorm and NormFinder software that use a pairwise comparison and a model-based approach, respectively. Our results extend the general recommendation of using multiple reference genes for normalization purposes to our model systems and provide an overview of several references genes that are likely to be stable in similar culture protocols.

The application of in vivo bioluminescent imaging in infectious disease research has significantly increased over the past years. The detection of transgenic parasites expressing wildtype firefly luciferase is however hampered by a relatively low and heterogeneous tissue penetrating capacity of emitted light. Solutions are sought by using codon-optimized red-shifted luciferases that yield higher expression levels and produce relatively more red or near-infrared light, or by using modified bioluminescent substrates with enhanced cell permeability and improved luminogenic or pharmacokinetic properties. In this study, the in vitro and in vivo efficacy of two modified bioluminescent substrates, CycLuc1 and AkaLumine-HCl, were compared with that of D-luciferin as a gold standard. Comparisons were made in experimental and insect-transmitted animal models of leishmaniasis (caused by intracellular Leishmania species) and African trypanosomiasis (caused by extracellular Trypanosoma species), using parasite strains expressing the red-shifted firefly luciferase PpyRE9. Although the luminogenic properties of AkaLumine-HCl and D-luciferin for in vitro parasite detection were comparable at equal substrate concentrations, AkaLumine-HCl proved to be unsuitable for in vivo infection follow-up due to high background signals in the liver. CycLuc1 presented a higher in vitro luminescence compared to the other substrates and proved to be highly efficacious in vivo, even at a 20-fold lower dose than D-luciferin. This efficacy was consistent across infections with the herein included intracellular and extracellular parasitic organisms. It can be concluded that CycLuc1 is an excellent and broadly applicable alternative for D-luciferin, requiring significantly lower doses for in vivo bioluminescent imaging in rodent models of leishmaniasis and African trypanosomiasis.

One key element to the health of the ocular surface encompasses the presence of gel-forming mucins in the pre-ocular tear film. Conjunctival goblet cells are specialized epithelial cells that secrete mucins necessary for tear film stability and general homeostasis. Their dysfunction can be linked to a range of ocular surface inflammation disorders and chronic injuries. To obtain new perspectives and angles to tackle mucin deficiency, the need for an accurate evaluation of their presence and corresponding mucin secretion in ex vivo conjunctival cultures has become a requisite. In vitro, goblet cells show a significant decrease in the production and secretion of gel-forming mucins, accompanied by signs of dedifferentiation or transdifferentiation. Explant cultures on laminin-treated CLP-PEG hydrogels can, however, support the production of gel-forming mucins. Together, we challenge the current paradigm to evaluate the presence of cultured goblet cells solely based on their general mucin (MUC) content through imaging analyses, showing the need for additional techniques to assess the functionality of goblet cells. In addition, we broadened the gel-forming mucin profile of in vivo goblet cells with MUC5B and MUC6, while MUC2 and MUC6 is added to the profile of cultured goblet cells.