Urinary peptidome analyses for the diagnosis of chronic kidney disease in dogs

Chronic kidney disease (CKD) is clinically important in canine medicine. Current diagnostic tools lack sensitivity for detection of subclinical CKD. The aim of the present study was to evaluate urinary peptidome analysis for diagnosis of CKD in dogs. Capillary electrophoresis coupled to mass spectrometry analysis demonstrated presence of approximately 5400 peptides in dog urine. Comparison of urinary peptide abundance of dogs with and without CKD led to the identification of 133 differentially excreted peptides (adjusted P for each peptide <0.05). Sequence information was obtained for 35 of these peptides. This 35 peptide subset and the total group of 133 peptides were used to construct two predictive models of CKD which were subsequently validated by researchers masked to results in an independent cohort of 20 dogs. Both models diagnosed CKD with an area under the receiver operating characteristic (ROC) curve of 0.88 (95% confidence intervals [CI], 0.72–1.0). Most differentially excreted peptides represented fragments of collagen I, indicating possible association with fibrotic processes in CKD (similar to the equivalent human urinary peptide CKD model, CKD273). This first study of the urinary peptidome in dogs identified peptides that were associated with presence of CKD. Future studies are needed to validate the utility of this model for diagnosis and prediction of progression of canine CKD in a clinical setting

Cardiac troponin I levels in canine pyometra

BACKGROUND: Myocardial injury may contribute to unexpected deaths due to pyometra. To detect myocardial damage, measurement of cardiac troponin I (cTnI) is currently the most sensitive and specific method. The aims of the present study were to evaluate presence of myocardial damage in canine pyometra by analysis of cTnI, to explore whether myocardial injury was associated with systemic inflammatory response syndrome (SIRS) and to evaluate whether other clinical or laboratory parameters were associated with cTnI increase. METHODS: Preoperative plasma levels of cTnI were investigated in 58 female dogs with pyometra and 9 controls. The value of physical examination findings, haematological, serum biochemical and pro-inflammatory (CRP and TNF-α) parameters as possible predictors of increased cTnI levels was also evaluated. RESULTS: Seven dogs with pyometra (12%) and one control dog (11%) had increased levels of cTnI. In the pyometra group, the levels ranged between 0.3–0.9 μg l(-1 )and in the control dog the level was 0.3 μg l(-1). The cTnI levels did not differ significantly between the two groups. No cardiac abnormalities were evident on preoperative physical examinations. Four of the pyometra patients died within two weeks of surgery, of which two were examined post mortem. In one of these cases (later diagnosed with myocarditis and disseminated bacterial infection) the cTnI levels increased from 0.9 μg l(-1 )preoperatively to 180 μg l(-1 )the following day when also heart arrhythmia was also detected. The other patient had cTnI levels of 0.7 μg l(-1 )with no detectable heart pathology post mortem. CTnI increase was not associated with presence of SIRS. There was a trend for the association of cTnI increase with increased mortality. No preoperative physical examination findings and few but unspecific laboratory parameters were associated with increased cTnI levels. CONCLUSION: Increased cTnI levels were observed in 12% of the dogs with pyometra. The proportions of dogs with cTnI increase did not differ significantly in the pyometra group compared with the control group. CTnI increase was not associated with presence of SIRS. A trend for association of cTnI increase and mortality was observed. Preoperative physical examination findings and included laboratory parameters were poor predictors of increased cTnI levels

Blood lactate levels in 31 female dogs with pyometra

<p>Abstract</p> <p>Background</p> <p>Canine pyometra is a life-threatening disease common in countries where spaying of dogs is not routinely performed. The disease is associated with endotoxemia, sepsis, systemic inflammatory response syndrome (SIRS) and a 3–4% mortality rate. Blood lactate analysis is clinically valuable in predicting prognosis and survival, evaluating tissue perfusion and treatment response in human and veterinary critical care settings. The aims of the present study were to investigate 1) the blood lactate levels of female dogs with pyometra by a hand-held analyser and 2) if these levels are related with the clinical status or other biochemical or hematological disorders.</p> <p>Methods</p> <p>In total 31 female dogs with pyometra admitted for surgical ovariohysterectomy and 16 healthy female control dogs were included in the present study. A complete physical examination including SIRS-status determination was performed. Blood samples for lactate concentrations, hematological and biochemical parameters, acid-base and blood gas analysis and other laboratory parameters were collected and subsequently analysed. The diagnosis pyometra was verified with histopathological examination of the uterus and ovaries. Increased hospitalisation length and presence of SIRS were used as indicators of outcome.</p> <p>Results</p> <p>In the pyometra group the median blood lactate level was 1,6 mmol l^-1(range <0.8–2.7 mmol l^-1). In the control group the median lactate level was 1,2 mmol l^-1(range <0.8–2.1 mmol l^-1). Of the 31 bitches 19 (61%) fulfilled 2 or more criteria for SIRS at inclusion, 10 bitches (32%) fulfilled 3 of the SIRS criteria whereas none accomplished more than 3 criteria. Lactate levels did not differ significantly between the pyometra and control group, or between the SIRS positive and SIRS negative dogs with pyometra. Increased lactate concentration (>2.5 mmol l^-1) was demonstrated in one female dog with pyometra (3%), and was not associated with longer hospitalisation or presence of SIRS. Lactate measurement was not indicative of peritonitis. None of the bitches died during or within two months of the hospital stay. The measurements of temperature, heart rate, respiratory rate, percentage bandforms of neutrophilic granulocytes, α₂-globulins, creatinin, pvCO₂, TCO₂and base excess showed significant differences between the SIRS positive and the SIRS negative pyometra cases.</p> <p>Conclusion</p> <p>Increased blood lactate concentrations were demonstrated in 3% (1/31), and SIRS was present in 61% (19/31) of the female dogs with pyometra. Preoperative lactate levels were not related with presence of SIRS or prolonged hospitalisation. Lactate measurement was not indicative of peritonitis. The value of a single and repeated lactate analysis in more severely affected cases remains to be determined.</p

Advances in structure elucidation of small molecules using mass spectrometry

The structural elucidation of small molecules using mass spectrometry plays an important role in modern life sciences and bioanalytical approaches. This review covers different soft and hard ionization techniques and figures of merit for modern mass spectrometers, such as mass resolving power, mass accuracy, isotopic abundance accuracy, accurate mass multiple-stage MS(n) capability, as well as hybrid mass spectrometric and orthogonal chromatographic approaches. The latter part discusses mass spectral data handling strategies, which includes background and noise subtraction, adduct formation and detection, charge state determination, accurate mass measurements, elemental composition determinations, and complex data-dependent setups with ion maps and ion trees. The importance of mass spectral library search algorithms for tandem mass spectra and multiple-stage MS(n) mass spectra as well as mass spectral tree libraries that combine multiple-stage mass spectra are outlined. The successive chapter discusses mass spectral fragmentation pathways, biotransformation reactions and drug metabolism studies, the mass spectral simulation and generation of in silico mass spectra, expert systems for mass spectral interpretation, and the use of computational chemistry to explain gas-phase phenomena. A single chapter discusses data handling for hyphenated approaches including mass spectral deconvolution for clean mass spectra, cheminformatics approaches and structure retention relationships, and retention index predictions for gas and liquid chromatography. The last section reviews the current state of electronic data sharing of mass spectra and discusses the importance of software development for the advancement of structure elucidation of small molecules

Hyperammonemia and systemic inflammatory response syndrome predicts presence of hepatic encephalopathy in dogs with congenital portosystemic shunts

Hepatic encephalopathy (HE) is an important cause of morbidity and mortality in patients with liver disease. The pathogenesis of he is incompletely understood although ammonia and inflammatory cytokines have been implicated as key mediators. To facilitate further mechanistic understanding of the pathogenesis of HE, a large number of animal models have been developed which often involve the surgical creation of an anastomosis between the hepatic portal vein and the caudal vena cava. One of the most common congenital abnormalities in dogs is a congenital portosystemic shunt (cpss), which closely mimics these surgical experimental models of HE. Dogs with a cPSS often have clinical signs which mimic clinical signs observed in humans with HE. Our hypothesis is that the pathogenesis of HE in dogs with a cPSS is similar to humans with HE. The aim of the study was to measure a range of clinical, haematological and biochemical parameters, which have been linked to the development of HE in humans, in dogs with a cPSS and a known HE grade. One hundred and twenty dogs with a cPSS were included in the study and multiple regression analysis of clinical, haematological and biochemical variables revealed that plasma ammonia concentrations and systemic inflammatory response syndrome scores predicted the presence of HE. Our findings further support the notion that the pathogenesis of canine and human HE share many similarities and indicate that dogs with cPSS may be an informative spontaneous model of human HE. Further investigations on dogs with cPSS may allow studies on HE to be undertaken without creating surgical models of HE thereby allowing the number of large animals used in animal experimentation to be reduced

Data for: Urinary peptidome analyses for diagnosis of chronic kidney disease in dogs

CE-MS analyses were performed using a Beckman Coulter Proteome Lab PA800 capillary electrophoresis system (Beckman Coulter, Fullerton, USA) on-line coupled to a micrOTOF II MS (Bruker Daltonic, Bremen, Germany). The electro-ionization sprayer (Agilent Technologies, Palo Alto, CA, USA) was grounded, and the ion spray interface potential was set to –4.5 kV. Data acquisition and MS acquisition methods were automatically controlled by the CE via contact-close-relays. Spectra were accumulated every 3 s, over a range of m/z 350 to 3000. In the next step the MosaiquesVisu software package was applied to deconvolute mass spectral ion peaks, because ionization produced ions at different charged states from the original urinary peptides. This deconvolution step groups these differently charged ions into single peptides with unique real mass. Only signals observed in a minimum of three consecutive spectra with a signal-to-noise ratio of at least 4 were considered. Signals with a calculated charge of 1+ were automatically excluded to minimize interference with matrix compounds or drugs. Capillary electrophoresis migration time and MS-detected mass were normalized by the definition of 950 clusters of peptides covering a range of 17.23 to 47.74 minutes in CE migration time and 807 to 16399 kDa in molecular mass. Samples were normalized by peptide abundance (intensity) calibration based on 141 endogenous internal urinary polypeptide standards displaying the highest frequency and stability in all analysed samples, to compensate for differences in hydration status and urine volume between dogs. Each polypeptide present in the list was defined by its normalized migration time [min], molecular mass [kDa], and signal intensity detected. Using a Microsoft Structured Query Language database, all detected polypeptides were deposited, matched, and annotated in order to allow for further comparison between groups. The criteria applied to consider a polypeptide identical was that within different samples, the mass deviation was lower than 50 ppm for masses &lt; 4 kDa, 150 ppm for masses &gt;6 kDa, and between 50-150 ppm for masses between 4 and 6 kDa. Acceptable migration time deviation was between 1 and 2.5 minutes

Data for: Urinary peptidome analyses for diagnosis of chronic kidney disease in dogs

CE-MS analyses were performed using a Beckman Coulter Proteome Lab PA800 capillary electrophoresis system (Beckman Coulter, Fullerton, USA) on-line coupled to a micrOTOF II MS (Bruker Daltonic, Bremen, Germany). The electro-ionization sprayer (Agilent Technologies, Palo Alto, CA, USA) was grounded, and the ion spray interface potential was set to –4.5 kV. Data acquisition and MS acquisition methods were automatically controlled by the CE via contact-close-relays. Spectra were accumulated every 3 s, over a range of m/z 350 to 3000. In the next step the MosaiquesVisu software package was applied to deconvolute mass spectral ion peaks, because ionization produced ions at different charged states from the original urinary peptides. This deconvolution step groups these differently charged ions into single peptides with unique real mass. Only signals observed in a minimum of three consecutive spectra with a signal-to-noise ratio of at least 4 were considered. Signals with a calculated charge of 1+ were automatically excluded to minimize interference with matrix compounds or drugs. Capillary electrophoresis migration time and MS-detected mass were normalized by the definition of 950 clusters of peptides covering a range of 17.23 to 47.74 minutes in CE migration time and 807 to 16399 kDa in molecular mass. Samples were normalized by peptide abundance (intensity) calibration based on 141 endogenous internal urinary polypeptide standards displaying the highest frequency and stability in all analysed samples, to compensate for differences in hydration status and urine volume between dogs. Each polypeptide present in the list was defined by its normalized migration time [min], molecular mass [kDa], and signal intensity detected. Using a Microsoft Structured Query Language database, all detected polypeptides were deposited, matched, and annotated in order to allow for further comparison between groups. The criteria applied to consider a polypeptide identical was that within different samples, the mass deviation was lower than 50 ppm for masses &lt; 4 kDa, 150 ppm for masses &gt;6 kDa, and between 50-150 ppm for masses between 4 and 6 kDa. Acceptable migration time deviation was between 1 and 2.5 minutes.THIS DATASET IS ARCHIVED AT DANS/EASY, BUT NOT ACCESSIBLE HERE. TO VIEW A LIST OF FILES AND ACCESS THE FILES IN THIS DATASET CLICK ON THE DOI-LINK ABOV