Annual Research & Review in Biology

Canine pyometra is a common reproductive disorder of intact diestrual bitches. The disease is associated with inflammation and infection of the uterus leading to generalized illness. The varied reactions of the host to infection, inflammation, or trauma are collectively known as the acute-phase response (APR) and encompass a wide range of pathophysiological responses. One of these systemic responses to disease is an increase in the production by the liver of a number of serum proteins, which are known collectively as the acute-phase proteins (APP). The most important APPs in dogs are C-reactive protein (CRP) and serum amyloid A (SAA). When diagnosed with pyometra, the preferred and most effective treatment is ovariohysterectomy, a procedure that in itself causes systemic inflammation due to the tissue trauma caused by surgery. The purpose of this study was to identify the effect pyometra and ovariohysterectomy have on acute phase proteins and routinely measured biochemistry variables in serum of affected bitches. We achieved this by evaluating concentrations of CRP and SAA, as well as of some biochemistry parameters in serum before and after ovariohysterectomy in bitches with pyometra. Blood samples were collected from the cephalic vein immediately prior to and 24h, 72h and 168 hours after initiation of surgery. The levels of CRP, SAA and some biochemistry parameters - total protein (TP), albumin (ALB), globulin (GLOB), as well as activity of some enzymes: alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (AKLP) and lactate dehydrogenase (LDH) were determined

Aims: The aim of this study was to investigate the impact of pyometra and its surgical treatment, ovariohysterectomy, on acute phase proteins, specifically C-reactive protein (CRP) and serum amyloid A (SAA), as well as on routine serum biochemistry variables in affected bitches. By evaluating the concentrations of CRP and SAA, alongside key biochemistry parameters before and after surgery, this research seeks to elucidate the physiological and biochemical responses to both the disease and its treatment, providing insight into the systemic effects of pyometra and the surgical stress response in dogs.

Study Design:  The study design involved a cohort of twenty-seven female dogs, divided into two groups: seventeen clinically healthy intact female dogs serving as the control group, which were examined for routine health care or vaccinations, and ten female dogs diagnosed with pyometra at the Faculty of Veterinary Medicine, University of Sarajevo. The diagnosis of pyometra was made based on case history, clinical signs, ultrasonography, and confirmed by gross examination of the pus-filled uterus during and after ovariohysterectomy. The surgical procedure was performed using a standard mid-line approach under general inhalation anesthesia, with no post-operative complications reported.

Place and Duration of Study: Faculty of Veterinary Medicine, University of Sarajevo, between June 2012 and September 2012.

Methodology: Blood samples were collected from the distal cephalic vein into serum separating tubes (Vacutainer SST; Becton Dickinson, USA) for CRP and SAA measurement and serum biochemical evaluation just before the ovariohysterectomy (base values) as well as after 24 hours (day one), 72 hours (day three) and 168 hours (day seven). Blood samples from control dogs were collected only once. Serum samples were prepared by centrifugation (1.500 × g for 10 min) and stored in plain micro tubes (Eppendorf, Germany) at −20°C prior to analysis. Serum CRP concentrations in the dogs were measured with ELISA microplate reader (Anthos 2001 ELISA-reader, Anthos Mikrosysteme, Krefeld, Germany) using a commercial canine CRP enzyme-linked immunosorbent assay (ELISA) kit (Tridelta PhaseTM Range CRP Assay, Tridelta Development Limited, County Kildare, Ireland). For SAA, the analyses were performed using also a commercially available ELISA kit (Tridelta PhaseTM Range SAA Assay, Tridelta Development Limited, County Kildare, Ireland). Total protein (TP), albumin (ALB), globulin (GLOB) concentrations and alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALKP) and lactate dehydrogenase (LDH) activities were determined by using standard procedures and slides by Catalyst Dx Chemistry Analyzer, Idexx.

Statistical analyses were performed using the SPSS software package (for Windows, Version 11.5, SPSS Inc, USA). All data were shown as mean ± standard deviation. The results were analyzed using the independent t-test. A P-value less than 0.05 were considered statistically significant.

Results: Our results indicated significantly higher concentrations of CRP and SAA in dogs with pyometra throughout the study period compared to the control group, with CRP concentrations rising up to 15-fold and SAA up to 10-fold. Notably, these increases in CRP and SAA concentrations were mostly observed before surgery, with only limited additional increases post-surgery. This suggests that while ovariohysterectomy induces a proinflammatory response, it does not significantly escalate the levels of these proteins beyond the initial disease impact. The variability observed in CRP and SAA levels points to potential interindividual differences, impacting their diagnostic value. However, on the last postoperative day, the significantly elevated levels of both proteins in dogs with pyometra suggest a persistent, though less severe, inflammatory process, with a subsequent decline as homeostasis was gradually restored.

Conclusion: The role of the acute phase response is to limit inflammation, remove the damaging factor and restore homeostasis. The systemic effects of pyometra are reflected by acute phase proteins and several biochemistry parameters.