Tear Film Proteome of Healthy Domestic Cats

Abstract

The aim of this study was to investigate the proteins found in tear film of healthy domestic cats. Schirmer tear test strips were used to collect tear samples of twelve healthy cats, which were mixed, centrifuged, and placed in a single 1.5 mL microtube that was frozen at -20°C, until analysis by two-dimensional polyacrylamide gel and mass spectrometry associated with high-performance liquid chromatography. The resulting spectra were analyzed and compared with the Swiss-Prot search tool. Forty peptides were detected in the analyzed protein fragments of 90 spots, with 16 proteins identified. Of these, the authors confirmed what has been already found in other studies: lactotransferrin, serum albumin, allergenic lipocalins, and neutrophil gelatinase-associated lipocalin. Others were considered novel in tear film samples of all species: cyclin-dependent protein kinase, serine/arginine repetitive matrix protein, apelin receptor, secretory protein related to C1q/TNF, Wee1, α-1,4 glucan phosphorylase, and WD repeat domain 1. The network was divided into 11 clusters, and a biological function was assigned. Most of the proteins have functions in the defense and maintenance of feline ocular surface homeostasis. Serum albumin is a bottleneck protein, with a high betweenness value. This paper is a pioneer in reporting, in-depth, the tear film proteome of domestic cats.

Figure 1

Two-dimensional polyacrylamide gel (2D-SDS-PAGE).

Figure 2

Graphical representation exposing the distribution of proteins and their abundances (%), which can be considered a signature for the protein composition of tear film of healthy cats. The different colors in the abundance bars refer to the different spots where the same protein was identified, that is, for each protein, there is a bar with the sum of all the abundances found for itself.

Figure 3

Interaction network of Mus musculus proteins homologous to those identified in Felis catus. Betweenness value represented by the color of the nodes, the lightest being the lowest value and the darkest being the highest value. It represents the ability of the node to join several clusters, or groups of nodes. Proteins with an elevated betweenness value have a high interaction with other proteins and are called bottleneck proteins. The width of the border of the nodes represents the value of node degree, so that the greater the width of the border, the greater the value and vice versa. The node degree property represents the number of connections that cross a single node, and the proteins with an above average node degree value are called hubs, which have an important regulatory function in the network. The clusters are subsets of nodes that are mostly connected to each other. In Figure 3, we have 11 clusters, each grouped in numbered circles. For each cluster, a biological process was assigned with the lowest corrected p value, according to the BiNGO tool. Therefore, in this network figure, for each cluster, the biological function, the number of the cluster, and its p value were indicated (the closer to zero, the more reliable). No annotation data were generated for cluster 8 (no data).