We compared the expression of mRNAs and miRNAs in 20 AAK patients and 20 healthy subjects and identified novel potential biomarkers of this rare and challenging disease. We focused not only on genes that may associate with AAK but also on those related to clinical features of the disease. We hope our study can provide a new method to help evaluate and treat patients.
We applied WGCNA to the analysis of non-tumor disease, identified DGEs with the intersection of multiple approaches, and presented their affiliation with miRNAs, trying to identify the most qualified genes to be biomarkers. The regulation of differential miRNAs and DEGs was shown to elucidate their expression level change and the regulatory relationship between them in AAK. Although there were studies showing that AAK deteriorated with age [11], we didn’t find a specific age with a high incidence of the disease. Based on function analysis and pathway enrichment, we predicted the small molecule active drugs most likely to be helpful and demonstrated their possible effectiveness.
A few highly conserved signaling pathways are vitally important for eye development. Known as a vital tumor suppressor in carcinogenesis, CHL1 works by repressing the PI3K/AKT signaling pathway [12]. As a neurotransmitter, it is involved in cognitive activities [13] and some neurological diseases. It guides neuronal survival and growth in the nervous system and can regulate the regeneration of axons [14]. It is significantly lower in female than in male patients and may become a potential biomarker for the diagnosis of female patients. We found that the regulator of CHL1, miR-30a-5p, was decreased in AAK patients, further demonstrating that CHL1 may play a critical role in the process. SOCS family proteins are mainly in charge of excessive cytokine signaling restraint, and because of the ability to regulate certain allergic autoimmune diseases, SOSC3 gets special attention [15]. Experimental results indicate that SOSC3 is a key regulator of physiology, also participating in immune homeostasis. Dysregulation of SOSC3 can cause a variety of diseases, including cancer, autoimmune diseases, and neurodegenerative disorders. A study showed that SOCS3 induced regrowth of retinal axons and the formation of functional synapses in the superior colliculus after optic nerve injury [16]. Although the upregulation of miR-30a-5p was able to decrease the SOCS3 protein level, the promotion of miR-19a-3p[17] or miR-185[3] was also associated with reduced SOCS3 expression. It was shown that miR-204-5p can inhibit cell proliferation, migration, invasion, and metastasis [18]. Our study found it was downregulated in AAK, which may be related to the process of corneal conjunctivation, in which the conjunctival epithelium invades the cornea and causes the formation of new blood vessels. We found that miR-224-5p, upregulated in AAK, may contribute to understanding the disease mechanism, while other researchers found that it was increased in the pathogenesis of hepatocellular carcinoma [19]. In total, these factors may be optimal diagnostic biomarkers and inform the development of a therapeutic strategy for this disease.
Balancing anti- and pro-angiogenic factors helps maintain the avascular environment of the corneal limbus in AAK. Some researchers believe that the conjunctiva can invade the cornea and strengthen conjunctival cell signaling, which leads to clinical symptoms of neovascularization [20]. The classic symptoms always appear after childhood, including corneal pannus and corneal conjunctivation [21]. Existing clinical treatments depend on the severity of the ocular surface signs; artificial tears can relieve slight symptoms, while severe cases need amniotic membrane transplantation, keratoplasty, or even limbo-keratoplasty [22]. By screening for genes related to AAK, our study may help with early diagnosis and differential diagnosis to prevent serious consequences and extend the treatment window, working as an accessory examination.
Unfortunately, there were some certain limitations in this study. The sample size was insufficient, the clinical information was inadequate, and there was a lack of datasets for validation. The expression validation of the five miRNAs was consistent with the present bioinformatics analysis. Additional in vitro and in vivo experiments, such as cell culture and establishment of an animal model, are required to further investigate the potential mechanisms underlying AAK. Furthermore, applying only conjunctival cells was not an overwhelming support, so using these results as diagnostic methods requires further examination. The present study may provide a research basis for the diagnosis and treatment of AAK.