Preclinical study of a new matrix to help the ocular surface in dry eye disease.

Dry eye disease (DED), a multifactorial disease of the tears and ocular system, causes loss of tear film homeostasis with damage to the ocular surface. This study aimed to assess whether a peculiar matrix based on sodium hyaluronate (HA), xanthan gum (XNT), glycine (GLY) and betaine (BET) as osmoprotectants, could be involved in biological responses. Wound healing assay on human corneal epithelial (HCE) cells in monolayer showed a synergistic effect of the combination of HA + XNT (**p ≤ 0.01) together with an efficient extracellular matrix remodeling of the formulation in SkinEthic™ HCE 3D-model sought by integrin beta-1 (ITGß1) expression and morphological analysis by hematoxylin and eosin (H&E), compared to a reference marketed product. The synergistic effect of HA + XNT + GLY + BET showed an antioxidant effect on HCE cells (***p ≤ 0.001). Real-time PCR analysis showed that the combination of GLY + BET seemed to ameliorate the effect exhibited by the single osmoprotectants in reducing tumor necrosis factor-alpha (TNFα, #p ≤ 0.05), interleukin-1 beta (IL1ß, ####p ≤ 0.0001) and cyclooxygenases-2 (COX2, ####p ≤ 0.0001) genes in SIRC cells under hyperosmotic stress. Furthermore, pretreatment with XNT, alone and in combination (##p ≤ 0.01), reduced COX2 expression in human non-small cell lung cancer cells (A549). Finally, the formulation was well-tolerated following q.i.d. ocular administration in rabbits during a 28-day study. Due to the synergistic effect of its components, the matrix proved able to repair the ocular surface restoring cell homeostasis and to protect the ocular surface from pro-inflammatory pathways activation and oxidative damage, thus behaving as a reactive oxygen species (ROS) scavenger.

Characterisation of three variants of estrogen receptor beta mRNA in the common sole, Solea solea L. (Teleostei).

In all vertebrates, estrogen action is mediated by cognate nuclear receptors. In this study, we cloned the different transcripts of the estrogen receptor beta (ERbeta) gene of the common sole, Solea solea. 5'-RLM-RACE (RNA Ligase-Mediated 5'-Rapid Amplification of cDNA Ends) and 3'-RACE analyses revealed three isoforms of different length, called Long, Intermediate and Short isoforms, consisting of 2212, 1531 and 1207 b, respectively. The Long isoform is characterised by an open reading frame (ORF) encoding 589aa, with an estimated molecular weight of 65kDa. Phylogenetic analysis established that it belongs to the teleost ERbeta1 or ERbetaa cluster. The Intermediate isoform encodes a 490-aa protein, which lacks the first 99aa of the Long isoform, but still retains a complete DNA-binding domain (DBD). In the Short variant (363aa-long), all the N-terminal region, down to the two zinc fingers included, is missing, thus crippling DBD. ERbeta transcription was analysed by semiquantitative RT-PCR with specific primers, common to the Long and Intermediate isoforms, in various sole tissues, such as brain, gills, muscle, stomach, intestine, spleen, head kidney, kidney, liver and gonads. This analysis revealed that ERbeta displays a widespread or ubiquitous pattern of transcription, with the highest levels being found in the gonads and liver.