Freeze-drying protocols and methods of maintaining the in-vitro biological activity of horse platelet lysate.

Platelet lysate, derived from platelets, are valuable biological products rich in bioactive molecules. Their use promotes tissue healing and modulates inflammation. However, maintaining the stability and bioactivity of platelet lysate is challenging since they degrade rapidly at room temperature. This study focused on the possibility to confer enhanced stability to freeze-dried equine platelet lysate as an alternative to platelet-rich plasma (PRP). Platelet lysate (PL) was derived from PRP and freeze-dried either as such or using various adjuvants. Primary cell cultures of porcine Vascular Wall-Mesenchymal Stem Cells were treated with different PL formulations, and cell viability was assessed using an MTT assay. Overall, the addition of PL significantly improved cell viability as compared to controls without growth factor supplementation or with foetal bovine serum. Notably, the freeze-drying process maintained the effectiveness of the PL for at least a week. Furthermore, the study revealed that varying the horse as the source of PL could yield varying effects on cell viability. Detailed freeze-drying protocols were established, including freezing, primary drying and secondary drying phases, and the type of adjuvant. This study demonstrated the potential of freeze-dried equine PL as a viable alternative to PRP and highlighted the importance of precise freeze-drying protocols and adjuvants for standardization. Equine PL showed promise for medical treatment in horses, offering advantages such as extended shelf life, ease of handling, and reduced transportation costs, with the potential for broadened therapeutic usage.

Bronchoalveolar Lavage Cytology in Severe Equine Asthma: Cytocentrifugated versus Sediment Smear Preparations.

Equine asthma is a common respiratory disease that may affect horses of any age. The diagnosis of severe equine asthma (SEA) (historically referred as recurrent airway obstruction or RAO) is based mainly on the history of the animal and clinical signs, which are further supported by the cytological examination of the bronchoalveolar lavage (BAL). This can also be helpful in monitoring the inflammation of the lower airways in response to environmental management and medication. The cytocentrifugated preparation is usually considered the method of choice for BAL cytological interpretation. The aim of this study was to compare the results in terms of differential cell counts (DCC) in BAL cytology performed on sedimented smears and cytocentrifugated preparations. To carry this out, 48 BAL samples were collected from six horses with SEA that were subjected to a process of exacerbation of the disease by environmental stimuli, which was later followed by the appropriate treatment. Each collected BAL fluid was equally divided into duplicate portions: one processed by cytocentrifugation and one by sediment smear from simple centrifugation. Cytologic examination of all BAL by both methods showed poor concordance in DCC, although it was still able to allow diagnostic recognition of severe lung neutrophilic disorders. These results suggest that sediment smear preparation, although remaining a useful method in general equine practice associated with clinical assessments in the diagnosis of SEA under conditions where there is no possibility of using a cytocentrifuge, cannot be considered a comparable alternative.

Short-Term Survival and Postoperative Complications Rates in Horses Undergoing Colic Surgery: A Multicentre Study.

The occurrence of colic could be influenced by the characteristics of a population, geographical area, and feeding management. The aim of this study was to report the short-term postoperative complications and survival rates and to identify factors that might affect the outcome of horses that underwent colic surgery in three Italian surgical referral centres. Data of horses subjected to colic surgery in three referral centres (2018-2021) were analysed. Comparisons of the outcomes were performed using a Mann-Whitney or a Chi square test. Areas under the receiver operating characteristic (ROC) curve and multivariable logistic regression analysis were used for parameters that were significant in the previous univariate analysis. The goodness-of-fit of the model was assessed using the Akike information criterion (AIC). Significance was defined as p < 0.05, and odds ratios and 95% confidence intervals were calculated as percentages. A total of 451 horses were included. The survival rate was 68.5% of all of the horses that underwent colic surgery and 80% of the horses surviving anaesthesia. Age, BCS, PCV and TPP before and after surgery, amount of reflux, type of disease, type of lesion, duration of surgery, surgeon's experience, and amount of intra- and postoperative fluids administered influenced the probability of short-term survival. The multivariate analysis revealed that PCV at arrival, TPP after surgery, and BCS had the highest predictive power. This is the first multicentre study in Italy. The results of this study may help surgeons to inform owners regarding the prognosis of colic surgery.

Lidocaine constant rate infusion in isoflurane anesthetized neonatal foals.

Introduction: In horses, lidocaine infusion is administered intraoperatively for analgesia and for a reduction of inhalant anaesthetic requirement. The objective of the study was to describe the anaesthetic effects of lidocaine infusion in isoflurane anaesthetised foals. Methods: Twelve foals (<3 weeks old) undergoing surgery were included in the study (LIDO group). Foals were premedicated with midazolam and butorphanol IV, anaesthesia was induced with ketamine and propofol IV and maintained with isoflurane. Lidocaine was administered intraoperatively at 0.05 mg/kg/min. Also, the anaesthetic records of 11 foals in which lidocaine was not administered intraoperatively were retrospectively evaluated and they were considered as a historical control group (HC). Heart rate (HR), mean arterial pressure (MAP) and fraction of expired isoflurane were monitored continuously. Time of extubation, time to reach sternal recumbency and standing were recorded. The quality of recovery was assessed. Results: HR decreased in both groups compared with baseline values and intraoperatively the differences were statistically significant (p = 0.01 and p = 0.03 respectively in the LIDO and HC groups). Intraoperatively the HR was significantly lower in the LIDO group (71.2 ± 13.4 bpm) compared with the HC group (87.1 ± 17.7 bpm) (p = 0.0236). The number of foals requiring inotropic support (LIDO n = 7 and HC n = 9) was not statistically associated with the treatment group (p = 0.371). The extubation time, the time to reach the sternal recumbency and the quality of recovery did not differ significantly between the two groups (p = 0.7 and p = 0.6 respectively). Discussion: In conclusion, in anaesthetised foals the addition of lidocaine does not provide a sparing effect on isoflurane requirement, and it does not interfere with the quality of recovery, however it decreases significantly the HR, which is pivotal in foals for the maintenance of cardiac output and peripheral perfusion. Therefore, a continuous patient monitoring is essential.

The Comparison of Latero-Medial versus Dorso-Palmar/Plantar Drilling for Cartilage Removal in the Proximal Interphalangeal Joint.

The aims of the present study were to compare the percentages of articular cartilage removed using a lateral drilling approach of the proximal interphalangeal joint (PIPJ) and a dorsal drilling approach, and to assess the usefulness of digital fluoroscopy when performing a lateral drilling approach. Sixty cadaveric PIPJs were drilled using a surgical drill bit to remove the articular cartilage. The limbs were divided into three groups containing 10 forelimbs and 10 hindlimbs each. One group received the dorsal drilling approach, the second one received the lateral drilling approach and the last one received the lateral drilling approach under digital fluoroscopy guidance. The percentage of articular cartilage removed from each articular surface was assessed using Adobe Photoshop ® software. The percentages of removed cartilage turned out to be significantly higher with lateral approach, especially under fluoroscopic guidance, both in the forelimbs (p = 0.00712) and hindlimbs (p = 0.00962). In conclusion, the lateral drilling approach seems to be a minimally invasive technique with which to perform PIPJ arthrodesis, even more efficient than the previously reported dorsal approach.

Immunohistochemical Expression of Neurokinin-A and Interleukin-8 in the Bronchial Epithelium of Horses with Severe Equine Asthma Syndrome during Asymptomatic, Exacerbation, and Remission Phase.

Severe equine asthma (EA) syndrome is a chronic obstructive disease characterized by exaggerated contraction, inflammation, and structural alteration of the airways in adult horses, when exposed to airborne molds and particulate material. However, little is known about the relationship between the degree and type of inflammation on one hand, and the severity of the disease and the response to treatment on the other. Furthermore, to date, very few studies evaluate the diagnostic value of histology and immunohistochemical features of endoscopic biopsies on subjects with severe equine asthma. To investigate the expression of two inflammatory markers (NKA and IL-8) before, during, and after the exacerbation of severe EA, a histological and immunohistochemical study was carried out on a series of biopsy samples collected by bronchoscopy from six EA-affected horses subjected to process exacerbation through environmental stimuli and then to pharmacological treatment. The application of a histological biopsy scoring system revealed a significant difference between control cases and the EA-affected horses in all experimental phases (asymptomatic, early exacerbation phase, late exacerbation phase, and remission phase). For immunohistochemistry (IHC), only the intensity of NKA positivity increases significantly between control horses and the EA horses at late exacerbation and remission phases. In EA-affected horses, a difference was detected by comparing histology between asymptomatic and remission phase, meanwhile, NKA and IL-8 showed no differences between the experimental phases. Based on these results we can assert that: (1) The endoscopic biopsies generate reliable and homogeneous samples in the entire bronchial tree; (2) the clinical improvement associated with treatment is characterized by a significant worsening of the histological findings; and (3) the NKA immunopositivity seems to increase significantly rather than decrease, as one would have expected, after pharmacological treatment. Further studies are necessary both to implement the number of samples and to use other markers of inflammation to characterize the potential role of cytokines in the diagnosis and therapeutic approach of severe equine asthma.

First isolation of Klebsiella variicola from a horse pleural effusion.

BACKGROUND: Respiratory diseases are the second most common cause of illnesses in horses, their etiology can be viral, bacterial, immune-mediated, or mechanical (Racklyeft and Love DN, Aust Vet J 78:549-59, 2000; Austin et al., J Am Vet Med Assoc 207:325-328, 1995; Arroyo et al., J Vet Intern Med 31:894-900, 2017). Klebsiella variicola is a Gram-negative bacterium that was initially identified as an endophyte in soil and plants such as bananas, rice, sugar cane and maize but recent studies have identified this microorganism as an emerging pathogen in humans (Rodríguez-Medina et al., Emerg Microbes Infect 8:973-988, 2019; Fontana et al., J Clin Microbiol 57:e00825-18, 2019; Rosenblueth et al., Syst Appl Microbiol 27:27-35, 2004). This paper describes, for the first time to our knowledge, the isolation of K. variicola from pleural effusion in a male adult horse. CASE PRESENTATION: 17-years Italian Saddle Horse with respiratory distress and fever was admitted to the Veterinary Teaching Hospital of the Department of Veterinary Medical Sciences, University of Bologna. At home, the patient had undergone antibiotic therapy without clinical improvement. Vital signs on admission revealed an increased respiratory rate, tachycardia, pyrexia and weight loss. The animal was submitted for collateral examination including thoracic radiology and ultrasound and thoracoscopy that showed bilateral pleural effusion associated with multifocal pulmonary atelectasis. During the thoracoscopic examination, that confirmed the presence of a seropurulent pleural effusion, a sample of pleural fluid was collected and Gram-negative bacteria were isolated and subjected to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) that allowed the identification of K. variicola. The isolate was sensitive to amikacin, cefazolin, enrofloxacin, marbofloxacin, tetracycline, and trimethoprim-sulfamethoxazole;the horse was treated with Oxytetracycline and amikacin. Despite a general health improvement of the subject, the pleural effusion did not resolve after treatment. CONCLUSIONS: This paper describes, for the first time, the isolation of K. variicola in a horse with respiratory disease. The misidentification between K. variicola and K. pneumoniae has caused unawareness about significant aspects of this bacterial species. In fact, even though in animals the role of this bacterium is not clear, in humans it has been recognized as an emerging pathogen. The use of new methods for bacterial identification will probably lead to the isolation of a greater number of strains which will have to be studied to acquire knowledge that will be useful to clarify the clinical importance and relevance of K. variicola also in animals.

Localisation of cannabinoid and cannabinoid-related receptors in the equine dorsal root ganglia.

BACKGROUND: Growing evidence recognises cannabinoid receptors as potential therapeutic targets for pain. Consequently, there is increasing interest in developing cannabinoid receptor agonists for treating pain. As a general rule, to better understand the actions of a drug, it would be of extreme importance to know the cellular distribution of its specific receptors. The localisation of cannabinoid receptors in the dorsal root ganglia of the horse has not yet been investigated. OBJECTIVES: To localise the cellular distribution of canonical and putative cannabinoid receptors in the equine cervical dorsal root ganglia. STUDY DESIGN: Qualitative and quantitative immunohistochemical study. METHODS: Cervical (C6-C8) dorsal root ganglia were collected from six horses (1.5 years of age) at the slaughterhouse. The tissues were fixed and processed to obtain cryosections which were used to investigate the immunoreactivity of canonical cannabinoid receptors 1 (CB1R) and 2 (CB2R), and for three putative cannabinoid-related receptors: nuclear peroxisome proliferator-activated receptor alpha (PPARα), transient receptor potential ankyrin 1 (TRPA1) and serotonin 5-HT1a receptor (5-HT1aR). RESULTS: The neurons showed immunoreactivity for CB1R (100%), CB2R (80% ± 13%), PPARα (100%), TRPA1 (74% ± 10%) and 5-HT1aR (84% ± 6%). The neuronal satellite glial cells showed immunoreactivity for CB2R, PPARα, TRPA1 and 5-HT1aR. MAIN LIMITATIONS: The low number of horses included in the study. CONCLUSIONS: This study highlighted the expression of cannabinoid receptors in the sensory neurons and glial cells of the dorsal root ganglia. These findings could be of particular relevance for future functional studies assessing the effects of cannabinoids in horses to manage pain.

Expression of Proteinase-Activated Receptor 2 During Colon Volvulus in the Horse.

Large colon volvulus in horses is associated with a poor prognosis, especially when ischemic-reperfusion injury of the affected intestinal tract develops. Proteinase-activated receptor 2 (PAR2) plays an important role in the pathogenesis of inflammation in the gastrointestinal tract. The aim of this study was to evaluate the distribution and expression of PAR2 in colonic pelvic flexure of horses spontaneously affected by large colon volvulus (CVH group). Eight horses admitted for severe abdominal colon volvolus and which underwent surgery were included. Colon samples were collected after enterotomy. Data previously obtained from healthy horses were used as a control group. Histologic evaluation was carried out to grade the severity of the colon lesions. Immunofluorescence, western blot and quantitative polymerase chain reaction (RT-qPCR) were carried out on colon samples to evaluate PAR2 expression. In addition, the transcriptional profile of cytokines and chemokines was evaluated using RT2 Profiler™ PCR Array Horse Cytokines & Chemokines. Three out of the eight patients were euthanised due to clinical deterioration. Immunostaining for PAR2 was observed in the enterocytes, intestinal glands and neurons of the submucosal and myenteric plexi. In the CVH horses, the expression of PAR2 mesenger RNA (mRNA) did not differ significantly from that of the healthy animals; western blots of the mucosa of the colon tracts showed a clear band of the expected molecular weight for PAR2 (~44 kDa) and a band smaller than the expected molecular weight for PAR2 (25kDa), suggesting its activation. The gene expressions for C-X-C motif ligand 1 (CXCL1); interleukin 8 (IL8), macrophage inflammatory protein 2 beta (MIP-2BETA) were upregulated in the colic horses as compared with the colons of the healthy horses. Therefore, in the present study, the expression and activation of PAR2 in the colons of horses in the presence of an inflammatory reaction like that occurring in those with spontaneous colon volvulus was confirmed.

Evaluation of Protein Carbonyl Content in Healthy and Sick Hospitalized Horses.

Literature on the protein carbonyl content (PCC) evaluation in horses is scarce, thus the aims were to evaluate the PCC in healthy and SIRS (Systemic Inflammatory Response Syndrome) horses and to investigate the performances of PCC in terms of sensitivity, specificity, and likelihood ratio in identifying SIRS positive and negative horses. A total of 72 adult horses were included. All the horses underwent to a complete physical examination, blood analysis, and were evaluated for the SIRS status. Blood samples were collected once in healthy horses and at admission time, then 24, 48, 72, and 96 h after admission in sick animals. PCC was evaluated using a method previously described. Data were statistically analyzed to verify differences in PCC between healthy vs. SIRS positive or SIRS negative horses at all sampling time. The receiver operating characteristic (ROC) curve was performed to verify sensitivity and specificity of PCC in the diagnosis of SIRS-positive and SIRS negative horses. The healthy horses were standardbred mares with a median age of 8.5 years. The sick horses were 31/54 females, 16/54 geldings, and 7/54 stallions of different breeds and with a median age of 12 years old. Eight out of 54 sick horses were SIRS negative, while 46/54 were SIRS positive. Statistically significant differences were obtained between healthy and SIRS positive horses, while no differences were observed between healthy and SIRS negative horses at any sampling time. The best cutoff value of PCC to discriminate between SIRS positive, SIRS negative, and healthy horses, the sensitivity and specificity of cutoff point, the area under receiver operating characteristic curve, the 95% confidence intervals, and the likelihood ratio were reported. We found higher PCC values in sick SIRS-positive horses vs. healthy ones with a decrement over time, while no differences at admission, nor during the observational period, were obtained in sick but SIRS-negative horses. The value of 0.049 nmol/ml/mg is reported as a potential cutoff for the diagnosis of SIRS positivity vs. healthy horses with a sensibility of 74.5% and a specificity of 72.2%. In conclusion, PCC seems to be a sensitive and specific marker for SIRS in horses.

Proteinase Activated Receptor 4 in the Jejunum of Healthy Horses and of Horses With Epiploic Hernia.

Proteinase activated receptor 4 (PAR4) in the gastrointestinal tract is involved in the regulation of inflammation and pain pathways. The aim of the present study was to evaluate the distribution and expression of PAR4 in the jejunum of healthy horses and in the pathologic tracts from horses undergoing surgery for herniation of the small intestine through the epiploic foramen. Eight healthy horses (Group H) and eight horses with epiploic hernia (Group EH) were included; the jejunum samples were collected at the slaughter or intraoperatively after enterectomy, respectively. To evaluate PAR4 expression in sections of the jejunum, immunofluorescence, western blot and quantitative polymerase chain reaction (qRT-PCR) were performed. Immunohistochemistry of PAR4 in the jejunum of the healthy horses showed that receptors are predominantly expressed in the immune cell population scattered throughout the lamina propria of the mucosa and in the submucosa. Quantitative PCR data demonstrated that PAR4 mRNA was detectable in all of the samples analyzed without any difference between the H and the EH groups, however the PAR4 protein level was significantly lower in the jejunums of the EH horses. In the Group EH horses, PAR4 immunoreactivity was mainly expressed in the mast cells and was extensively distributed in the sierosa. In the lamina propria of mucosa of Group EH, leukocytes were less abundant than in Group H. In this study, the distribution and expression of PAR4 in the jejunums of the healthy horses and in those with spontaneous occurring epiploic hernia was demonstrated.

Treatment of Septic Arthritis With Acoustic Cavitation and Lavage: A Case Report.

A 2-year-old Quarter Horse filly was admitted to our facility with a two-day history of trauma caused by barbed wire on the front face of the right front limb fetlock joint. A septic arthritis was confirmed with the synovial fluid examination. The cytologic evaluation of synovial fluid showed 101,000 leukocytes/mm3, 90% neutrophils, and a total protein concentration of 4 g/dL. It was, therefore, decided to perform an arthroscopic lavage using an ultrasonic device with the tip inserted inside the synovial cavity to exploit the phenomenon of acoustic cavitation. After the removal of the synovial pannus, the probe was left to vibrate in the area near the visible penetrating wound inside the joint until three liters of saline was finished. Further studies are needed for understanding and applying this technology in different equine fields and with different bacterial species. However, these first results are promising for the efficacy of this innovative procedure.

Paraoxonase-1 activity evaluation as a diagnostic and prognostic marker in horses and foals.

BACKGROUND: In several species, paraoxonase-1 (PON-1) decreases during inflammation, because of the presence of oxidative stress; its measurement recently has been validated in horses, but its role as a clinical biomarker is unknown. OBJECTIVES: To evaluate sensitivity, specificity and likelihood ratio of PON-1 activity to identify systemic inflammatory response syndrome (SIRS)-positive horses or horses with a poor prognosis. ANIMALS: One hundred seventy-two blood samples from 58 sick horses from 3 different veterinary hospitals. METHODS: In a cross-sectional study, PON-1 activity was measured upon admission and at 24-hour intervals until discharge or death, and results were analyzed based on SIRS status and outcome. RESULTS: No statistically significant difference was found in median PON-1 activity between SIRS and non-SIRS cases or between survivors and non-survivors except for mares, in which PON-1 activity was significantly lower in SIRS-positive horses (P = .05). The sensitivity of PON-1 activity in identifying horses with SIRS or negative outcome was low (0.0%-46.2% depending on the examined group) but its specificity was high (87.0%-100.0%). However, when PON-1 is low, the likelihood of death is 2.40-3.89 times higher than the likelihood of survival. Repeated measurement of PON-1 after treatment does not predict outcome. CONCLUSIONS AND CLINICAL IMPORTANCE: Evaluation of PON-1 activity in horses with inflammation might be advisable in the future, but only low activity at admission may be relevant in predicting SIRS or negative outcome.

The First Case of Botulism in a Donkey.

Botulism, a severe neuroparalytic disease that can affect humans, all warm-blooded animals, and some fishes, is caused by exotoxins produced by ubiquitous, obligate anaerobic, spore-forming bacteria belonging to the genus Clostridium and named botulinum neurotoxin (BoNT)-producing clostridia. This report presents the case of a 3-year-old donkey mare referred for progressive and worsening dysphagia of four days' duration. Her voluntary effort in eating and drinking was conserved, and she was able to slow chew without swallowing. A complete neurological examination was performed, and botulism was strongly suspected. The ability to swallow feed and water returned on the tenth day of hospitalization and improved progressively. The jenny was discharged from the hospital after fifteen days. During the hospitalization, the Italian National Reference Centre for Botulism confirmed the diagnosis: mare's feces were positive for BoNT/B and Clostridium botulinum type B.

Proteinase-activated receptor 2 distribution and expression in equine small intestine tracts following herniation through the epiploic foramen.

Proteinase-activated receptor 2 (PAR2) is a G-protein-coupled receptor for trypsin and mast cell tryptase; it is highly expressed at the intestinal level with multiple functions, such as epithelial permeability and intestinal motility. The aim of the study was to evaluate the distribution and expression of proteinase-activated receptor 2 in the small intestine during herniation through epiploic foramen. In this prospective clinical study, eight horses admitted for colic and which underwent exploratory laparotomy were considered. During surgery, the jejunum or the ileum was sampled by enterectomy. Morphological examination (histology, PAR2 immunohistochemistry) and molecular biology analysis (western blot and quantitative polymerase chain reaction) were carried out on the resected intestinal samples. The Marginal Injured Tracts (MITs) and Central Injury Tracts (CITs) were defined as the oral and caudal marginal segments of the resected bowel tract and as the geometric centre of the intestinal ischaemic lesion length, respectively. The PAR2 immunoreactivity was particularly evident in the epithelial cells, with higher immunoreactivity in the MIT rather than in the CIT. Moreover, a different immune localisation was observed in the MITs at the cell membrane level and in the CITs in the cytoplasm. No statistical difference was observed in PAR2 mRNA and protein (44kDa) expression between the MIT and the CIT. The PAR2 protein content in the intestinal tracts which were removed from horses with herniation was lower when compared with the control animals. This study provided data concerning the PAR2 presence and distribution in horses with intestinal herniation through the epiploic foramen.

Pharmacokinetic profile and partitioning in red blood cells of romifidine after single intravenous administration in the horse.

The aims of this study were to assess the plasma concentrations of romifidine in horses after intravenous injection, to evaluate the red blood cell (RBC) partitioning of the anaesthetic drug, and to improve knowledge regarding its sedative effect in horses describing the pharmacokinetic model. Eight adult Standardbred horses received a single bolus of romifidine at a dosage of 100 µg/kg. Blood samples (5 mL) were collected immediately before romifidine administration (t0), and at 2, 5, 10, 15, 20, 25, 30, 40, 50, 60, 75, 90, 105, 120, 150 and 180 min after injection. A sedation score was recorded at the same time. The romifidine concentrations in plasma and red blood cells were determined by high performance liquid chromatography (HPLC). The plasma and red blood cell concentrations were correlated with the sedation at each time point. Romifidine produced a satisfactory level of sedation in all animals. The sedation was detectable in all horses for up to 105 min. All the animals returned to normal without any behavioural changes at 180 min. The romifidine concentrations in the red blood cells were significantly higher (P < 0.01) at all time points than those in the plasma. The T1/2ß was 148.67 ± 61.59 min and body clearance was 22.55 ± 6.67 mL/kg per min. The results showed that after a single bolus administration of romifidine, a partitioning in the RBCs was detected.

The influence of environmental variables on platelet concentration in horse platelet-rich plasma.

Platelet-rich plasma (PRP) commonly refers to blood products which contain a higher platelet (PLT) concentration as compared to normal plasma. Autologous PRP has been shown to be safe and effective in promoting the natural processes of soft tissue healing or reconstruction in humans and horses. Variability in PLT concentration has been observed in practice between PRP preparations from different patients or from the same individual under different conditions. A change in PLT concentration could modify PRP efficacy in routine applications. The aim of this study was to test the influence of environmental, individual and agonistic variables on the PLT concentration of PRP in horses. Six healthy Standardbred mares were exposed to six different variables with a one-week washout period between variables, and PRP was subsequently obtained from each horse. The variables were time of withdrawal during the day (morning/evening), hydration status (overhydration/dehydration) treatment with anti-inflammatory drugs and training periods on a treadmill. The platelet concentration was significantly higher in horses treated with a non-steroidal anti-inflammatory drug (P = 0.03). The leukocyte concentration increased 2-9 fold with respect to whole blood in the PRP which was obtained after exposure to all the variable considered. Environmental variation in platelet concentration should be taken into consideration during PRP preparation.

Serum amyloid A, haptoglobin, and ferritin in horses with colic: Association with common clinicopathological variables and short-term outcome.

Equine colic may be associated with an acute phase response (APR). Measurement of acute phase proteins (APPs) allows the detection of an APR and may help clinicians in monitoring the disease; however, the role of APPs in colic is unclear. This study aimed to evaluate the clinical usefulness of serum amyloid A (SAA), haptoglobin and ferritin in combination with an extended clinicopathological profile in equine colic. The medical records of 54 horses were retrospectively selected. Horses were grouped based on outcome (survivors vs. non-survivors), diagnosis (ischaemic/strangulating vs. non-ischaemic/non-strangulating), and treatment (medical treatment vs. surgery). Laboratory data were compared, and a logistic regression analysis was performed for outcome prediction upon admission. A high percentage of horses had abnormal SAA (29/54), haptoglobin (20/54), and ferritin (31/54) concentrations. In particular, haptoglobin was below the reference interval in 13/54 horses. Non-survivors had significantly decreased haptoglobin and increased ferritin concentrations compared with survivors. The ischaemic/strangulating group had significantly increased creatinine and ferritin and decreased haptoglobin concentrations compared with the non-ischaemic/non-strangulating group. Creatinine was the only significant predictor of mortality in the regression analysis. In conclusion, APPs including SAA, haptoglobin, and ferritin combined with clinicopathological variables may help clinicians to understand the pathogenesis of APR and underline potential complications of equine colic. The reduction in haptoglobin concentration may suggest haemolysis or muscle fibre damage; ferritin may indicate alteration in iron metabolism and tissue damage. Further prospective studies are needed to assess diagnostic and prognostic values of APPs in colic horses.

Proteinase-activated receptor 2 expression in the intestinal tract of the horse.

Proteinase-activated receptor 2 (PAR2) is a G-protein-coupled receptor for trypsin and mast cell tryptase; it is highly expressed at the intestinal level with multiple functions, such as epithelial permeability and intestinal motility. Many proteases activate PAR2 during tissue damage, suggesting a role of the inflammatory response receptors. The aim of the study was to evaluate the distribution and expression of PAR2 in the jejunum, the ileum and the pelvic flexure, using samples collected from healthy adult horses after slaughter. Proteinase-activated receptor 2 immunoreactivity (PAR2-IR) was observed in the enterocytes, intestinal glands, the smooth muscle of the muscularis mucosae, and the longitudinal and circular muscle layers; there were no differences in the distribution of PAR2-IR in the different sections of the intestinal tract. The protein expression level showed that the relative amount of the PAR2 content in the mucosa of the intestinal tract decreased from the small to the large intestine while the PAR2 mRNA analysed showed similar values. This study provides relevant findings concerning the distribution of the PAR2 in the intestines of healthy horses and represents the starting point for evaluating the role of the PAR2 during strangulative intestinal disease and consequent systemic intestinal reperfusion/injury complications in horses in order to identify and employ antagonist PAR2 molecules.