The diseases of ocular surface and the cornea

The diseases of ocular surface and the cornea are very common in ophthalmological practice and as a result there is a continuous, need for novel therapeutic options. Here, we demonstrated a corneal distribution of GPR35, a potential molecular target for new drugs (; ). The search for the expression profile of gene performed on the basis of the records deposited in the public GEO repository revealed a medium level of in cornea, which is comparable to the expression level found in the retina, conjunctiva and uvea. A lower expression level was found in neuronal tissues, while high levels were found in gastrointestinal tissues and blood cells; this expression profile stays in line with the literature (). Western blotting and immunohistochemical experiments confirmed our preliminary bioinformatics analysis. The immunoblotting experiment revealed the presence of low and high molecular weight protein species that are immunoreactive to anti-GPR35 antibody; the detected protein bands were ∼34 kDa and ∼68 kDa in size, respectively. This result indicates that GPR35 exists as both monomeric and dimeric receptor in the corneal and scleral tissues. Oligomerization is a common feature of GPCRs and it shapes the signaling and pharmacology of the receptors (; ; ). Recent yeast two-hybrid studies demonstrated that GPR35 can oligomerize with several other GPCRs () which stays in line with our results on the homo-dimerization capacity of GPR35. However, the physiological relevance of this phenomenon requires further investigation. Interestingly the GPR35 dimer seems to be a Evans Blue tetrasodium salt receptor form of the receptor, as it caused stronger staining in the western blot. According to our immunohistochemical results, GPR35 immunoreactivity was found in all the examined corneas including normal corneas, KC and corneas with FECD. Immunohistochemical analysis revealed the presence of GPR35 mainly in corneal epithelium and endothelium. Only a slight staining for GPR35 was detected in stroma. Normal corneas and KC corneas showed a very similar distribution of GPR35. However, a slightly less intensive immunoreactivity or its irregular distribution was found in the endothelium of corneas with FECD. Since the previously published microarray experiments did not show any significant differences in the expression of GPR35 genes between normal and FECD corneas () such an altered pattern of GPR35 immunoreactivity can be attributed to pleomorphism, polymegethism and reduction of endothelial cell density reported in corneas of FECD patients (). FECD is the leading indication for endothelial keratoplasty. There are numerous theories on the etiology of the endothelial destruction in FECD, which are of genetic or environmental origin (; ). A role of oxidative stress and the possible inflammation has been confirmed in the pathophysiology of FECD (; ; ). Until now, no indication of the role of GPR35 in pathological processes leading to FECD exist in the literature. Therefore, our finding may add a new direction for research on the pathophysiology of this disease. The first discovered endogenous ligand for GPR35 is KYNA (). Recently, we have revealed the presence of KYNA in tears (). Furthermore, we demonstrated the presence of enzymes synthetizing KYNA – kynurenine aminotransferases KAT I, II and III in normal human corneas (, ). It is worth emphasizing that corneal endothelium and epithelium showed the strongest reactivity for all three KAT isoforms. There was a slight positive staining of corneal stroma for KAT I and II. KAT III immunoreactivity was found only in the stroma of the limbal region (, ). This pattern of KATs distribution closely imitated the immunoreactivity pattern of GPR35. Therefore, these results prove that the same eye structures contain GPR35 and may produce KYNA, its agonist. The potential role of KYNA in the cornea has been recently substantiated by the finding that KYNA increased the viability and proliferation of corneal epithelium in model of corneal epithelium (). However, the involvement of GPR35 in this effect has not yet been investigated.