VALIDATION OF AN ELECTROPHORETIC METHOD TO DETECT URINARY ALBUMIN IN CATS.

Urine samples were collected from 66 feline hospital admissions by cystocentesis or free catch, and analyzed (UPC, HRE). Within-assay imprecision for high (H), medium (M) and low (L) concentrations, between-assay imprecision, linearity/accuracy and percentage recovery were determined for albumin. Albumin was assessed by urine to creatinine ratio (UAC), percentage (ALB%) and mg/dl (ALB mg/dl). Cases were categorized: a) as chronic kidney disease with (CKDC) and without complications (CKD), feline lower urinary tract disease (FLUTD) or other, and b) as proteinuric (P), borderline proteinuric (BP), or not proteinuric (NP), and c) by type of sediment active (A) or not active (NA). Results: HRE within-assay CV was 2.58%(H), 5.31%(M), 3.42%(L), between-assay CV was 3.52%, recovery percentage was 97%, 93% and 109% for high, medium and low concentration, respectively, and linearity was high (r=0.99). Compared with UPC at different cut-off values, HRE showed higher specificity and PPV but lower sensitivity and NPV than dipstick to measure proteinuria. Dipsticks showed frequent false positives compared with HRE. Based on UPC, measures of albuminuria were higher in proteinurics than non-proteinurics, and ALB% correlated to UPC (r=0.35, R2=0.12, p<0.05) and UAC (r=0.41, R2=0.17, p<0.05). Samples with active sediment had higher ALB% than samples with inactive sediment. ALB% was not different between proteinuria groups (P, BP, NP). UPC, UAC, ALB mg/dl, ALB% were not different across pathologies, even if CKD had higher values. Conclusions: HRE is accurate and precise for measuring urinary albumin in cats. UPC, UAC and ALB% were correlated. Increases in filtered albumin and proteinuria indicate a greater glomerular involvement in renal disease

URINE PROTEOME IN DOGS AFFECTED BY LEISHMANIASIS AND CUSHING\u2019S SYNDROME

Background: Urinary proteome is a topic not deeply investigated in dogs. Leishmaniasis (L) and Cushing\u2019s Syndrome (CS) are frequently associated with persistent proteinuria. Evaluation of urine proteome with sensitive methods is considered a non invasive procedure to monitor the therapy and select novel biomarkers. Objectives: Comparison of urinary electrophoretic profiles between healthy (H) and diseased dogs (L, CS) at the time of diagnosis and during the treatment monitoring. Methods: Urine samples were collected from 49 dogs (12 H, 26 L, 11 CS) and Urine Protein and Albumin to Creatinine ratios (UPC, UAC), HRE and SDS-PAGE were performed. Electrophoretic profiles were analyzed by ImageJ\ua9; proteins were identified by ESI-Q-TOF. Results: UPC and UAC (mean\ub1SD) were significantly higher (p<0.01) in L (3.47\ub14.76 and 1.69\ub13.19) and CS (2.55\ub13.69 and 1.79\ub13.65) compared to H dogs (0.11\ub10.11 and 0.008\ub10.008). HRE was useful with urine protein concentration higher than 40 mg/dl, and revealed different electrophoretic patterns between healthy (mainly albumin band is present) and diseased dogs (also other bands can be present). Urinary albumin concentration determined by HRE correlated well with the data obtained by routine immunoturbidimetric method (r=0.91, p<0.01). SDS-PAGE allowed to visualize an \u201chealthy profile\u201d with 20-25 different bands in urine of H dogs; the two most abundant ones have been identified as uromodulin and albumin. Urine samples from L and CS dogs presented a significantly higher number of protein bands (35-40), a decrease of uromodulin and an increase of albumin band intensities and the appearance of other bands, particularly in the range of 55-14 kDa; among them haptoglobin, superoxide dismutase1 and arginine esterase were identified by ESI-Q-TOF in urine of L dogs. Qualitative changes of protein profiles were evidenced during the treatment in CS and L dogs, indicating in some cases an improvement of renal damage. Conclusion: HRE could represent a rapid screening test for diagnosis of renal damage. SDS-PAGE is a sensitive and promising technique to investigate qualitative proteinuria in healthy and diseased dogs. Urine proteome analysis could help clinicians to better characterize the proteinuria even in absence of renal histopathology

Cadmium accumulation and antioxidant responses in Sparus aurata exposed to waterborne cadmium.

Cadmium (Cd), a nonessential trace element, is rapidly accumulated by most living organisms and subsequently exerts its toxicity at different molecular levels. This study exposed gilthead sea bream (Sparus aurata) to waterborne 0.1 mg/l Cd for 11 days and investigated the Cd accumulation pattern, lipid oxidation, and response of antioxidant defences. At the end of the experiment, mean Cd concentrations in gills and liver, the organs most prone to metal accumulation, were 209.4 and 371.7 ng/g ww, respectively. Muscle did not show any Cd retention during the 11 days of exposure. In liver, the cytosolic fraction of the metal was chelated into the nontoxic form by metallothionein (MT), a specific Cd-inducible protein. Zn and Cu concentrations were not influenced by Cd exposure. Glutathione (GSH) concentrations and the antioxidant enzyme activities of GSH reductase and GSH peroxidase showed an overall decreasing trend. In addition, lipid and aqueous hydroperoxide levels did not show any significant variation. Oxidative stress indirectly generated by Cd seems to be compensated for by the different biochemical systems tailored to decrease cellular damage. In particular, the negative effects of Cd accumulation in tissues were probably counteracted by the induction of MT