MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression in a variety of physiological processes. Several studies have documented a diagnostic potential of circulating miRNAs in different neoplastic and inflammatory conditions in both humans and animals. Recently, also urinary miRNAs have been investigated in humans. Urinary miRNAs could potentially be useful non-invasive biomarkers in various urological disease conditions in cats.

The objective of this study was to; 1) investigate if miRNAs can be detected in feline urine; 2) evaluate the performance of different extraction kits for the best miRNA yield and sample purity; 3) assess stability of urinary miRNAs during different storage conditions.

Five cats were characterized as healthy based on clinical examination, hematology and biochemistry, urinalysis. Urine samples were obtained by cystocentesis. Three samples were collected from each cat: one immediately frozen at -80°C, one at -20°C, one refrigerated for 24 hours (+4°C) and subsequently frozen at -20°C. Two methods for RNA isolation were investigated (microRNA purification kit [Norgen Biotek Corp.] and miRNeasy mini kit [Qiagen]). Expression levels of 3 miRNAs (let-7a, miR-16, and miR-23a) were tested by quantitative real-time PCR and normalised to a spiked-in synthetic miRNA. Data was processed and statistically analysed by GenEx 6 Pro program (MultiD). An Anova test was applied to compare storage temperatures.

It was possible to detect the investigated miRNAs in urine from healthy cats. The miRNeasy mini kit performed markedly better with less contaminants following cDNA synthesis. No significant difference in miRNA levels could be detected among storage temperatures (range of fold change for each miRNA at +4°C, -20°C, -80°C: (let-7a: 4.91–5.64; 0–6.25; 5.67–6.51), (miR-16:3.61–6.54; 0–9.32; 4.02–8.99), miR-23a: 3.7–5.21; 0–5.21; 4.78–5.37).

MiRNAs are stably expressed in urine from healthy cats independently of storage temperature. Further studies are needed to assess the value of miRNAs as diagnostic biomarkers in urogenital conditions in cats.

Disclosures

No disclosures to report.