Joanna Wessely-Szponder

Crosstalk between coagulation and inflammation in mastitis and metritis in dairy cows

Coagulation and inflammation are closely related as part of the mechanisms of host defence during a severe infection. The aim of this study was to investigate the relation between thrombin as a factor in both the coagulative and inflammatory processes and neutrophil secretory function on the basis of lactoferrin (LF), elastase and myeloperoxidase release in the course of mastitis and metritis in cows. Thrombin generation was measured on the basis of hydrolysis of SAR-PRO-ARG-pNA and lactoferrin concentration was estimated by an ELISA method. The greatest thrombin generation was observed in the metritis group (1.18 +/- 0.62 IU). The level of LF was the highest in the group of cows with mastitis (0.74 +/- 0.55 mg/ml) in the first phase of the disease. In the second phase of the diseases the level of serum LF in cows with mastitis diminished to the value of 0.41 +/- 0.16 mg/ml, whereas in cows with metritis the level of LF increased to 0.51 +/- 0.17 mg/ml. This study reveals that the excessive production of thrombin not only causes hypercoagulatory disorders but also exaggerates neutrophil function by the release of some enzymes which may play a destructive role during disseminated intravascular coagulation (DIC). These enzymes also inhibit anticoagulative systems, thus potentially worsening the course of the disease.

Different activation of monocyte-derived macrophages by antimicrobial peptides at a titanium tibial implantation in rabbits

The aim of our study was to assess the functional and morphological features of monocyte derived Mfs from rabbits with titanium (Ti) tibial implants, their ability to polarize towards M1 or M2 and their reactivity after stimulation with neutrophil antimicrobial peptides extract (AMP). The study was conducted on six White New Zealand rabbits with tibial implants of pure titanium Grade 2. Blood was taken before implantation and 14days after biomaterial implantation. Then, blood-derived Mfs were cultured and their function was assessed on the basis of morphological changes, generation of nitric oxide (NO), superoxide, and proteases release after treatment with LPS or dexamethasone (Dex). As a result of polarization we specified two subpopulations of Mfs with features characteristic of each subtype. M1 stimulated with LPS exhibited an increased NO and superoxide level, and M2 stimulated with Dex which showed higher arginase and lower free radical generation. Addition of AMP evoked further changes in Mfs morphology and function. After 24h stimulation with AMP an increase of NO was observed in all cultures, whereas after 48h it decreased. Production of superoxide lowered, especially after 48h, when M1 generated 6.00±0.2nM and M1/AMP culture generated 5.2±0.1nM of superoxide. Our study revealed that activated Mfs stimulated with AMP demonstrated both pro- and anti-inflammatory features. Moreover, we did not detect significant differences between the response of Mfs cultured from blood derived monocytes before and after implantation of Ti implants.

Elastase, Myeloperoxidase, and Alkaline Phosphatase Release and Free Radical Generation in Neutrophils Isolated from Blood of Sows at Different Stages of Oestrous Cycle

Abstract The aim of the study was to assess the influence of the oestrous cycle phase on neutrophil secretory activity and to extrapolate it to susceptibility to uterine infections. The obtained results indicate that the highest enzyme release seen in the late follicular phase (elastase release was 42.18 ±3.11% of maximal release, myeloperoxidase was 45.0 ±5.12%, and alkaline phosphatase was 44.75 ±9.0%) was related to the level of 17β-oestradiol in plasma. Similarly, a free radical generation was also the most enhanced during this phase. Significantly lower values were obtained from sows during the luteal phase in regard to both enzyme release (36.62 ±3.58% for elastase, 27.87 ±8.7% for myeloperoxidase, and 22.12 ±2.4% for alkaline phosphatase), and that of free radicals (2.28 1.6 μM/106 cells for nitric oxide and 2.47 0.6 nM/106 cells for superoxide).

The Neutrophil Response to Rabbit Antimicrobial Extract After Implantation of Biomaterial into a Bone/Cartilage Defect

Background/aim: In this study, the neutrophil response to an antimicrobial extract was evaluated as a potential marker of inflammatory process after implantation of alginate and carbon fiber biomaterials into a bone or cartilage defect in rabbits. Materials and Methods: The response to biomaterials used was assessed based on enzyme release, generation of reactive oxygen species (ROS) and survival of neutrophils isolated from the rabbits blood after implantation. Results: The implantation of alginate biomaterial increased elastase and alkaline phosphatase release, whereas carbon fibers did not evoke increased elastase release. The implantation of both biomaterials resulted in a higher myeloperoxidase (MPO) release after 30-min incubation. Stimulation with different fractions of antimicrobial peptide (AMP) extract diminished MPO release and nitric oxide generation, as well as slightly reducing neutrophil survival. Conclusion: Our study permits the assessment of the neutrophil intravital response to an implant without the need for preparation of histological sections. Additionally, AMP extract restricted some manifestations of the pro-inflammatory response and may be considered a regulator of neutrophil activity in the early inflammatory phase, preventing rejection of the implant.

Application of Platelet-rich Plasma and Tricalcium Phosphate in the Treatment of Comminuted Fractures in Animals

Aim: To assess the applicability of β-tri-calcium phosphate (TCP) and platelet-rich plasma (PRP) in the treatment of comminuted fractures in small animals. Material and Methods: The experimental study was carried out on 16 New Zealand White rabbits. After creating the bone defect and performing tibial osteotomy, TCP implants containing activated PRP were introduced into the fracture and the defect. The fracture was stabilised using external fixators or intramedullary nails. After 12 weeks, the animals were euthanised, and radiological, histological, scanning electron microscopy and peripheral quantitative computed tomography examinations were performed. The analysis also covered the results of fracture treatment in 37 small animals (cats and dogs) in which treatment with TCP containing PRP was used as an alternative to cancellous bone implantation. Results: Correct bone union was observed in the experimental groups, TCP remained visible at the site of the fracture after 12 weeks. In the clinical application in small animals, bone union was observed in over 91% of treated animals. Conclusion: β-TCP and activated PRP may be an effective method of bone union enhancement in the treatment of comminuted fractures in small animals.