Chronic kidney disease (CKD) is a major cause of mortality in cats, but sensitive and specific biomarkers for early prediction and monitoring of CKD are currently lacking. The present study aimed to apply proteomic techniques to map the urine proteome of the healthy cat and compare it with the proteome of cats with CKD. Urine samples were collected by cystocentesis from 23 healthy young cats and 17 cats with CKD. One-dimensional sodium-dodecyl-sulfate polyacrylamide gel electrophoresis (1D-SDS-PAGE) was conducted on 4-12% gels. Two-dimensional electrophoresis (2DE) was applied to pooled urine samples from healthy cats (n = 4) and cats with CKD (n = 4), respectively. Sixteen protein bands and 36 spots were cut, trypsin-digested and identified by mass spectrometry. 1D-SDS-PAGE yielded an overall view of the protein profile and the separation of 32 ± 6 protein bands in the urine of healthy cats, while CKD cats showed significantly fewer bands (P < 0.01). 2-DE was essential in fractionation of the complex urine proteome, producing a reference map that included 20 proteins. Cauxin was the most abundant protein in urine of healthy cats. Several protease inhibitors and transport proteins that derive from plasma were also identified, including alpha-2-macroglobulin, albumin, transferrin, haemopexin and haptoglobin. There was differential expression of 27 spots between healthy and CKD samples (P < 0.05) and 13 proteins were unambiguously identified. In particular, increased expression of retinol-binding protein, cystatin M and apolipoprotein-H associated with decreased expression of uromodulin and cauxin confirmed tubular damage in CKD cats suggesting that these proteins are candidate biomarkers.

Background: Hermann's tortoise, Testudo hermanni, is currently on the International Union for Conservation of Nature (IUCN) red list of endangered species. Reptile medicine relies also on laboratory analyses to evaluate health status, but reference ranges for hematology and biochemistry variables and protein electrophoresis in plasma of healthy tortoises are not available. Objective: The purposes of this study were to establish reference ranges for select hematologic and biochemical variables in clinically healthy Hermann's tortoises, and evaluate the impact of sex and season. Methods: Blood samples were collected from 34 healthy tortoises at the end of September and beginning of July. Blood smears, HCT, concentrations of HGB and select plasma biochemical analytes, select enzyme activities, and plasma protein fractions were evaluated. Reference ranges were determined and checked for influence of sex and sampling time point. Results: Typical reptilian RBC and WBC were observed in blood smears. HCT and concentrations of HGB, uric acid and urea, and ALT and AST activities were significantly higher in males than in females. Concentrations of glucose, uric acid, and phosphate, and AST activity were significantly higher at the beginning of July, whereas concentrations of urea and Cl were higher at the end of September prior to hibernation. The electrophoretic protein fractions included albumin, and a, b, and c globulins. Conclusions: The reference ranges defined in the present study are useful for clinical tortoise medicine and conservation. Sex and seasonal sampling were identified as factors significantly affecting hematology and blood chemistry analytes; they should be taken into consideration when assessing tortoise health status.

Urinary proteome and metabolome were studied in healthy and CKD dogs. •Proteomics highlighted a decrease of uromodulin and an increase of albumin.• 1 H NMR evidenced 17 metabolites significantly different between healthy and CKD dogs.•Proteomics and metabolomics successfully suggested putative biomarkers for CKD.

SUMMARYAlkaline phosphatase (AP) catalyses the detachment of phosphate residues from different substrates. Its activity has been demonstrated in seminal plasma and spermatozoa from porcine and other mammalian species; anyway, the role of AP in male reproduction has not been clarified yet and the aim of this study was to determine AP function in boar sperm capacitation and in vitro fertilization (IVF). AP activity was assayed in seminal plasma and in uncapacitated and in vitro capacitated (IVC) spermatozoa; in addition, capacitation was studied in presence of different doses of AP (1.2 and 2.5 IU/mL). The effect of different doses of AP (1.2 and 2.5 IU/mL) on several sperm parameters after IVC (viability, acrosome integrity with FITC-PSA, capacitation status with CTC staining, tyrosine phosphorylation) and on fertilizing ability during IVF were also evaluated. High AP activity was detected in seminal plasma, in particular in sperm-rich fraction; a lower activity was detected in uncapacitated spermatozoa while a significant decrease was evidenced after IVC. Viability was not changed by AP supplementation of the capacitating medium, whereas acrosome integrity and capacitation status were significantly affected by 1.2 and 2.5 doses, with a dose-dependent decrease in acrosome-reacted cells as well as in CTC B pattern displaying cells. As for sperm head protein phosphorylation, a decrease in relative fluorescence was detected in AP 2.5 group, if compared with capacitated one. After IVF, a dose-dependent decrease in penetrated oocytes was recorded, with an increase in monospermic zygote rate. In conclusion, we demonstrated that AP activity decreases under capacitating condition and that addition of AP to spermatozoa during capacitation results in a depression of the capacitating process and IVF. We can infer that AP plays a role in keeping spermatozoa quiescent until they are ejaculated and in modulating the acquisition of the fertilizing ability.

Feline morbillivirus (FeMV) is a newly discovered paramyxovirus infecting domestic cats and its role in the pathogenesis of feline chronic kidney disease (CKD) has been suggested, however not confirmed. The primary aim of the study was to evaluate the renal damage associated with FeMV infection in cats. In this retrospective study, clinical and clinicopathological data were compared among 14 FeMV naturally infected, 21 CKD and 22 healthy cats. FeMV positive cats had serum chemistry analytes and main urine chemistry results similar to the healthy subjects. FeMV positive cats had significantly decreased urine specific gravity (median 1054, range 1022–1065) and urine creatinine (median 227.23 mg/dL, range 83.02–489.75) when compared with healthy cats (median 1067, range 1040–1080, p < 0.001; median 406.50 mg/dL, range 195.32–575.58; p < 0.001, respectively). Urine protein:creatinine ratio (UPC) results of FeMV and CKD were not different (median 0.20, range 0.08–1.03; median 0.23, range 0.10–0.80, respectively), however UPC results were significantly increased in both groups, if compared with healthy cats (median 0.1, range 0.04–0.250, p < 0.01). Based on clinical data, serum creatinine concentration, urine specific gravity and UPC results, CKD was suspected by clinicians in 3/14 FeMV cats. Urine protein sodium-dodecyl-sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in 10/13 (77%) FeMV cats indicated a tubular pattern, with a decrease of uromodulin and an increase in the number and intensity of low molecular weight proteins. FeMV infection can be associated with different grades of renal dysfunction ranging from mild tubular proteinuria with less concentrated urine to azotemia in cats younger than those typically affected by CKD.